Journal articles on the topic 'Other chemical sciences not elsewhere classified'

BackgroundThe diet of the koala (Phascolarctos cinereus) is comprised almost exclusively of foliage from the genusEucalyptus(family Myrtaceae).Eucalyptusproduces a wide variety of potentially toxic plant secondary metabolites which have evolved as chemical defences against herbivory. The koala is classified as an obligate dietary specialist, and although dietary specialisation is rare in mammalian herbivores, it has been found elsewhere to promote a highly-conserved but low-diversity gut microbiome. The gut microbes of dietary specialists have been found sometimes to enhance tolerance of dietary PSMs, facilitating competition-free access to food. Although the koala and its gut microbes have evolved together to utilise a low nutrient, potentially toxic diet, their gut microbiome has not previously been assessed in conjunction with diet quality. Thus, linking the two may provide new insights in to the ability of the koala to extract nutrients and detoxify their potentially toxic diet.MethodThe 16S rRNA gene was used to characterise the composition and diversity of faecal bacterial communities from a wild koala population (n = 32) comprising individuals that predominately eat either one of two different food species, one the strongly preferred and relatively nutritious speciesEucalyptus viminalis, the other comprising the less preferred and less digestible speciesEucalyptus obliqua.ResultsAlpha diversity indices indicated consistently and significantly lower diversity and richness in koalas eatingE. viminalis. Assessment of beta diversity using both weighted and unweighted UniFrac matrices indicated that diet was a strong driver of both microbial community structure, and of microbial presence/absence across the combined koala population and when assessed independently. Further, principal coordinates analysis based on both the weighted and unweighted UniFrac matrices for the combined and separated populations, also revealed a separation linked to diet. During our analysis of the OTU tables we also detected a strong association between microbial community composition and host diet. We found that the phyla Bacteroidetes and Firmicutes were co-dominant in all faecal microbiomes, with Cyanobacteria also co-dominant in some individuals; however, theE. viminalisdiet produced communities dominated by the generaParabacteroidesand/orBacteroides, whereas theE. obliqua-associated diets were dominated by unidentified genera from the family Ruminococcaceae.DiscussionWe show that diet differences, even those caused by differential consumption of the foliage of two species from the same plant genus, can profoundly affect the gut microbiome of a specialist folivorous mammal, even amongst individuals in the same population. We identify key microbiota associated with each diet type and predict functions within the microbial community based on 80 previously identifiedParabacteroidesand Ruminococcaceae genomes.

This article offers an approach through administrative and cultural history to the problems associated with gathering and processing data for the Spanish national census of 1920, and by implication for earlier Spanish censuses. It focuses on the Basque province of Guipúzcoa, making use of correspondence between the central statistical office in Madrid, the provincial jefe de estadística and the localities, and of reports on three problematic towns within the province. The issues that emerge regarding ‘undercounting’, the definition of administrative boundaries and the classification of demographic characteristics are set in the wider context of census-taking practices and problems elsewhere in Spain and in other cultures.

AbstractObjectivesTo estimate the dietary intakes of heterocyclic amines (HCAs), to examine the intakes in relation to socio-economics, lifestyle and other dietary factors and to compare the classification of subjects by intake of HCA versus intake of meat and fish.DesignCross-sectional analysis within the Malmö Diet and Cancer (MDC) cohort. Data were obtained from a modified diet history, a structured questionnaire on socio-economics and lifestyle, anthropometric measurements and chemical analysis of HCAs. HCA intake was cross-classified against meat and fish intake. The likelihood of being a high consumer of HCAs was estimated by logistic regression analysis. Dietary intakes were examined across quintiles of HCA intake using analysis of variance.SettingBaseline examinations conducted in 1991–1994 in Malmö, Sweden.SubjectsA sub-sample of 8599 women and 6575 men of the MDC cohort.ResultsThe mean daily HCA intake was 583 ng for women and 821 ng for men. Subjects were ranked differently with respect to HCA intake compared with intake of fried and baked meat and fish (κ = 0.13). High HCA intake was significantly associated with lower age, overweight, sedentary lifestyle and smoking. Intakes of dietary fibre, fruits and fermented milk products were negatively associated with HCA intake, while intakes of selenium, vegetables, potatoes, alcohol (among men) and non-milk-based margarines (among women) were positively associated with HCA intake.ConclusionsThe estimated daily HCA intake of 690 ng is similar to values obtained elsewhere. The present study suggests that lifestyle factors (e.g. smoking, physical activity, fruit and vegetable intakes, and types of milk products and margarines) may confound associations between HCA intake and disease. The poor correlation between HCA intake and intakes of fried meat and fish facilitates an isolation of the health effects of HCAs.

This study was carried out to ascertain the characterization and classification of soils along river Donga in Donga local government area of Taraba State. Three profile pits were dug on the flat terrain for the physio-chemical properties of the soils. The pits were described and samples were collected for routine laboratory analysis for the selected physio-chemicals properties. Data collected were analyzed statistically using analysis of variance (ANOVA) The result of the laboratory analysis of the physical and chemical properties indicated that particle size distribution of sand particle dominated other fraction of fine earth materials. This can be attributed to the parent materials that gave rise to the soils.The pH (H2O) was slightly acidic in reaction in all the pedons studied (6.44, 5.82 and 5.67). The organic carbon content ranges from very low to low compared to the maximum level (5%) for tropical soils. Available phosphorus obtained exceeded the critical limits of 8.0 to >18mg/kg, which indicates that the available phosphorus were high. The exchangeable bases (Ca, Mg, K & Na) as indicated in the table 3 that Ca, Mg, K & Na were low in all the pedons studied. The results obtained from analyzed profile samples showed that percentage (%) base saturation values for all the pedons were high. The Correlation was done to determine non-significant and significant values among soil physio-chemical properties studied at 5% and 1% probability level. In addition, after the profile description and examination of the soil properties of the study area, the soils were classified using USDA soil taxonomy (2014). Pedons 1 and 2 are classified into the order of Alfisols and sub-group of Typical Haplustalf while pedon 3 is also classified into Alfisols but sub-group of Arenickandiustals (Hypereuticlixisels).

Although well over 1000 journal articles, conference papers, books, technical notes and theses have been written on the subject of information technology (IT) evaluation, only a relatively small subset of this literature has been concerned with the core issues of what precisely is meant by the term ‘value’ and with the process of making (specifically) IT investment decisions. All too often, the problem and highly complex issue of value is either simplified, ignored or assumed away. Instead the focus of much of the research to date has been on evaluation methodologies and, within this literature, there are different strands of thought which can be classified as partisan, composite and meta approaches to evaluation. Research shows that a small number of partisan techniques are used by most decision makers with a minority using a single technique and a majority using a mixture of such techniques of whom a substantial minority use a formal composite approach. It is argued that, in mapping the set of evaluation methodologies on to what is termed the investment opportunity space, that there is a limit to what can be achieved by formal rational evaluation methods. This limit becomes evident when decision makers fall back on ‘gut feel’ and other non-formal/rigorous ways of making decisions. It is suggested that an understanding of these more complex processes and decision making, in IT as elsewhere, needs tools drawn from philosophy and psychology.

Winter wheat is one of the major cereal crops in China. The spatial distribution of winter wheat planting areas is closely related to food security; however, mapping winter wheat with time-series finer spatial resolution satellite images across large areas is challenging. This paper explores the potential of combining temporally aggregated Landsat-8 OLI and Sentinel-2 MSI data available via the Google Earth Engine (GEE) platform for mapping winter wheat in Shandong Province, China. First, six phenological median composites of Landsat-8 OLI and Sentinel-2 MSI reflectance measures were generated by a temporal aggregation technique according to the winter wheat phenological calendar, which covered seedling, tillering, over-wintering, reviving, jointing-heading and maturing phases, respectively. Then, Random Forest (RF) classifier was used to classify multi-temporal composites but also mono-temporal winter wheat development phases and mono-sensor data. The results showed that winter wheat could be classified with an overall accuracy of 93.4% and F1 measure (the harmonic mean of producer’s and user’s accuracy) of 0.97 with temporally aggregated Landsat-8 and Sentinel-2 data were combined. As our results also revealed, it was always good to classify multi-temporal images compared to mono-temporal imagery (the overall accuracy dropped from 93.4% to as low as 76.4%). It was also good to classify Landsat-8 OLI and Sentinel-2 MSI imagery combined instead of classifying them individually. The analysis showed among the mono-temporal winter wheat development phases that the maturing phase’s and reviving phase’s data were more important than the data for other mono-temporal winter wheat development phases. In sum, this study confirmed the importance of using temporally aggregated Landsat-8 OLI and Sentinel-2 MSI data combined and identified key winter wheat development phases for accurate winter wheat classification. These results can be useful to benefit on freely available optical satellite data (Landsat-8 OLI and Sentinel-2 MSI) and prioritize key winter wheat development phases for accurate mapping winter wheat planting areas across China and elsewhere.

Nickel laterite deposits developed on ultramafic rocks have traditionally been a significant source of Ni and Co and recently of Sc. Although the Loma de Hierro deposit (Venezuela) has been in operation for more than 50 years, it lacks detailed studies on the mineralogical and geochemical composition of the lateritic profile. In this study, we present a geochemical and mineral description of the main carrier phases of Ni and Co in a complete profile of the deposit. The selected weathering profile has been developed from a partially serpentinized harzburgitic protolith and has a well-developed saprolitic horizon covered by a thin limonitic horizon. The geochemical signature of the profile is characterized by a significant Mg and Si decrease towards the top of the saprolite, with a clearly visible Mg discontinuity. The main Ni-bearing minerals are secondary serpentine (1–4 wt.% NiO) and kerolite-pimelite-dominated garnierite mixtures with serpentine (18–22 wt.% NiO). Limonite is rich in goethite (0–1.85 wt. % NiO), gibbsite, and Mn-oxy-hydroxides. The latter have intermediate compositions between lithiophorite and asbolane (2–13 wt.% CoO). The highest Sc grades (40–68 ppm) were observed in the limonite with amounts positively correlated with Fe content. Rare earth elements are mainly concentrated in the upper part of the saprolite horizon (60–80 ppm), while they have a lower content in the limonite (7–45 ppm). In this horizon, rare earth elements are clearly associated with Fe, indicating adsorption and/or coprecipitation. This association was not observed in the saprolite, suggesting that other minerals (e.g., clay minerals) are controlling their concentration; more information is needed to identify the rare earth element-bearing minerals. The lateritic profile of Loma de Hierro can be classified as representative of hydrated Mg silicate deposits, and was formed in a context of continuous tectonic uplift and a low water table conditions favoring the development of a thick saprolitic horizon and the precipitation of kerolite-pimelite-dominated garnierites.

Signet-ring cell carcinoma of the prostate gland (SRCCP) an uncommon and aggressive malignant tumour of the prostate gland which is characterized by histopathology examination features of compression of the nucleus into the form of a crescent by a large cytoplasmic vacuole. SRCCPs that have so far been reported have been either (a) primary tumours, metastatic tumours with the primary tumour elsewhere with the gastro-intestinal tract being the site of the primary tumour but the primary tumour could originate elsewhere, and additionally some reported SRCCPs have been classified as carcinoma of unknown primary. SRCCP could be a pure tumour or a tumour that is contemporaneously associated with other types of tumour including various variants of adenocarcinoma. SRCCP can manifest in various ways including: Incidental finding following prostatectomy that has been undertaken for a presumed benign prostatic hyperplasia, lower urinary tract symptoms, visible and non-visible haematuria, raised levels of serum PSA but some SRCCPs have been diagnosed with normal / low levels of serum PSA, there may be a history of dyspepsia in cases of metastatic signet-ring cell carcinoma in association with contemporaneous primary signet-ring cell carcinoma of the stomach or there may be a past history of surgical treatment for signet-ring cell carcinoma of the gastrointestinal tract, or bleeding from the gastrointestinal tract in cases of upper gastrointestinal tract and rectal bleeding as well as change in bowel habit for primary tumours of the anorectal region, retention of urine, and rarely a rectal mass in the case of SRCCP with an anorectal primary tumour. In order to exclude a primary signet ring cell carcinoma elsewhere, a detailed past medical history is required as well as radiology imaging including contrast – enhanced computed tomography (CECT) scan and contrast-enhanced magnetic resonance imaging (CEMRI) scan as well as upper gastrointestinal endoscopy and colonoscopy to exclude a primary lesion within the gastrointestinal tract. Diagnosis of SRCCP requires utilization of the histopathology and immunohistochemistry examination features of prostate biopsy, prostatic chips obtained from trans-urethral resection of prostate specimen or radical prostatectomy specimen. SRCCPs upon immunohistochemistry staining studies tend to show tumour that tend to exhibit positive staining for the following tumour markers as follows: PSA – positive staining for PSA has been variable in some studies, AE1/AE3, CAM 5.2, Ki-67 with a mean of 8%, PAS-diastase, Mucicarmine (50%), Alcian blue (60%), Alpha-methyl-acyl coenzyme A racemase (P504S), and Cytokeratin 5/6. SRCCPs also tend to exhibit negative staining for: Bcl2 (rare positive), and CEA (80%). Traditionally the treatment of Primary Signet-Ring Cell Carcinoma of the Prostate Gland has tended to be similar to the treatment of the traditional adenocarcinoma of the prostate gland which does include: hormonal treatment, radiotherapy, and surgery. Nevertheless, considering that primary SRCCPs and metastatic SRCCPs that have been reported in the literature have generally tended to be associated with an aggressive biological behaviour, even though there is no consensus opinion on the treatment of the disease it would be strongly recommended that these tumours that tend to be associated with rapid progress of the disease and poor survival there is an urgent need to treat all these tumours with aggressive surgery including radical prostatectomy plus adjuvant therapies including: radical radiotherapy, combination chemotherapy, selective prostatic angiography and super-selective embolization of the artery feeding the tumour including intra-arterial infusion of chemotherapy agents directly to the tumour, radiofrequency ablation of the tumour as well as irreversible electroporation of the tumour which should form part of a global multicentre study of various treatment options. With regard to metastatic signet-ring cell carcinomas of the prostate gland with a contemporaneous primary tumour elsewhere the primary tumour should also be treated by radical and complete excision of the primary tumour plus radical surgery and aggressive adjuvant therapy. Considering that SRCCPs have tendered not to respond well to available chemotherapy agents, there is need for urologists, oncologists, and pharmacotherapy research workers to identify new chemotherapy medicaments that would more effectively and safely destroy signet-ring cell tumours in order to improve upon the prognosis.

Lamprophyres are minor intrusions with atypical sources and crystallisation sequences. Among lamprophyres, the calc-alkaline (CAL) type on which this study focuses has the least distinctive chemistry and petrology. CAL correspond to small-volume mafic intrusions characterised by the early fractionation of amphibole and (or) biotite. In the Archean Superior Province (Canada), CAL are temporally and spatially related to several gold deposits and may thus be relevant to mineral exploration. This study focuses on several altered and metamorphosed intrusions of the Abitibi and La Grande subprovinces, which were designated lamprophyres based on field observations. Several criteria established from thin sections, whole-rock chemical analyses, and SEM data are applied to the studied rocks to distinguish CAL from other types of magma. As a result, only one of the studied dykes has the morphology, chemistry, and petrology typical of CAL, while the other intrusions are either too altered to be classified or may correspond to metamorphosed and metasomatized gabbro and diorite. This study shows that thin sections and whole-rock chemical analyses are not always sufficient to unequivocally classify an altered and metamorphosed intrusion as a CAL. Also, intrusions as challenging to recognise as CAL should not be used by exploration geologists to prospect for orogenic gold deposits. Much remains to be done to document the distribution and volume represented by lamprophyres in Archean greenstone belts and to confirm their spatial dependence with gold deposits.

Melagranites (colour index > 20, with biotite > garnet > cordierite) constitute ∼0.1% of the area of the 7300 km2 peraluminous South Mountain Batholith (SMB), Nova Scotia. The melagranites occur as small bodies showing sharp to gradational contacts against the Meguma Supergroup country rocks, and coeval mingling contacts against other facies of the batholith. They also occur as elliptical or blocky metre-scale enclaves elsewhere in the SMB. Characteristic petrological features of the melagranites include high modal abundances of sulphide minerals, strongly reacted metasedimentary xenoliths, mafic mineral-rich clots, apparent porphyritic textures with highly variable proportions of alkali feldspar megacrysts, and allotriomorphic-granular textures. Chemically and isotopically, melagranite rocks have wide compositional variations. In most major-element, trace-element, and isotopic variation diagrams, the melagranites lie on mixing lines between the more abundant granodioritic and monzogranitic phases of the SMB and the metasedimentary rocks of the Meguma Supergroup. Textural evidence, supported by published experimental evidence, suggests that the garnet, cordierite, and K-feldspar are peritectic phases resulting from incongruent melting of the pelitic fraction of the Meguma metasedimentary country rocks. The field relations, mineral assemblages, textural features, and chemical compositions of the melagranites all point to the melagranites as highly concentrated contamination zones in the SMB, representing small portions of the batholith that have failed either to complete the assimilation process or to disperse their contaminants widely in the batholith. As such, these rarely preserved melagranites provide petrogenetic information disproportionate in importance to their abundance in the batholith, especially about the significant role of contamination and assimilation in determining the physical and chemical composition of the SMB. Without preservation of melagranites in the SMB, and by extension all granite bodies, the petrogenetic importance of contamination is difficult to assess, even with trace-element and isotopic data. The present study shows that high quality field observations are as important in deciphering petrogenesis as chemical data.

After the 1991 Gulf War, veterans of the conflict from the United States, United Kingdom, Canada, Australia and other nations described chronic idiopathic symptoms that became popularly known as ‘Gulf War Syndrome’. Nearly 15 years later, some 250 million dollars in United States medical research has failed to confirm a novel war-related syndrome and controversy over the existence and causes of idiopathic physical symptoms has persisted. Wartime exposures implicated as possible causes of subsequent symptoms include oil well fire smoke, infectious diseases, vaccines, chemical and biological warfare agents, depleted uranium munitions and post-traumatic stress disorder. Recent historical analyses have identified controversial idiopathic symptom syndromes associated with nearly every modern war, suggesting that war typically sets into motion interrelated physical, emotional and fiscal consequences for veterans and for society. We anticipate future controversial war syndromes and maintain that a population-based approach to care can mitigate their impact. This paper delineates essential features of the model, describes its public health and scientific underpinnings and details how several countries are trying to implement it. With troops returning from combat in Afghanistan, Iraq and elsewhere, the model is already getting put to the test.

AbstractThe Dawson Cut Forest Bed lies in the lower part of thick, late Cenozoic loess deposits in the Fairbanks area. It is associated with several distal tephra beds that provide age control and offer the opportunity of its recognition elsewhere in central Alaska. EC tephra (named herein) occurs in the uppermost part of the Dawson Cut Forest Bed and its petrographic and chemical properties point to a co-magmatic relationship with PA tephra, which has not been found in direct association with the forest bed. Both tephra beds are pink and have unusually high Cl in their glass shards, which readily separates them from all other tephra beds in the Fairbanks area. They were produced by discrete eruptions, closely spaced in time. PA tephra has a glass-fission-track age of 2.02 ± 0.14 myr, indicating that the Dawson Cut Forest Bed must be about 2 million years old. The Palisades tephra (named herein) has very similar properties to these two tephra beds, suggesting that the buried forest bed just above it at the Palisades site on the Yukon River, about 250 km west of Fairbanks, correlates with the Dawson Cut Forest Bed.

ABSTRACT The natural occurrence of asbestos (NOA) in rocks and soil has been known for many years in several areas of the world, differently from the natural presence of asbestiform minerals. In Italy, the mapping of NOA is mandatory according to the 2001 and 2003 regulations. An investigation, not yet concluded, has revealed that in Italy, NOA is represented by chrysotile and tremolite asbestos with minor amounts of actinolite asbestos and anthophyllite asbestos. A field survey conducted in the Italian Western Alps (IWA), dealing with the natural occurrence of asbestiform minerals non-asbestos classified and not regulated, started many years ago and is still ongoing. It revealed that the following kinds of asbestiform silicates are present (in decreasing order of frequency): asbestiform polygonal serpentine and asbestiform antigorite, asbestiform diopside, asbestiform carlosturanite, asbestiform forsterite, asbestiform sepiolite, asbestiform balangeroite, and asbestiform talc. The asbestiform non-silicates brugnatellite and brucite have been rarely detected. Outside the IWA, asbestiform zeolite (erionite and offretite), asbestiform sodium amphibole (fluoro-edenite), and a few other asbestiform silicates have been also detected. For some asbestiform minerals, the identification is problematic and needs the use of transmission electron microscopy combining imaging at high magnification and electron diffraction and chemical data. This investigation is particularly important to distinguish four kinds of asbestiform minerals (antigorite, polygonal serpentine, carlosturanite, and balangeroite) from chrysotile since only the last one is regulated. The issue is much more complicated by the intergrowth of different fibrous species on the submicrometer scale.

Aegla papudo is a freshwater crab endemic to Chile. Although known to inhabit turbid watercourses with gravel, sand, filamentous algae, leaf litter, and no tree cover; however, is scarce the information on the proportions of these components, physical-chemical aspects of the water, population abundance, and reproductive season. Its presence in the Cordillera El Melon led us to describe the habitat. Between 2018 and 2019, the species was searched for in seven streams. Its habitat was described, the density calculated, the water’s physical-chemical parameters recorded. The variable that explained the abundance was analyzed with a model of Poisson. The species was recorded in the Los Coiles and El Gallo streams (62 and 24 individuals respectively) with a mean density of 1 individual/m2. The abundance was explained by the volume of the mud with gravel and pebble. This investigation is the first to determinethe abundance of adults and juveniles, as well as the variable that explained it. Environments with transparent water and tree cover are new aspects of the habitat. Fecal coliforms varied remarkably. Juvenile individuals present in August suggest that reproduction takes place between summer and fall. The International Union for Conservation of Nature does not register the species; however, we agree with other researchers who propose that the species be classified as Endangered due to known threats.

Ten groundwater samples from hand dug wells were collected at Irun Akoko within the basement complex of part of southwestern Nigeria. The aim of the study was to determine the groundwater suitability for both drinking and irrigation purposes. The groundwater samples were analysed for some physical and chemical constituents including pH, Total Dissolved Solids (TDS), turbidity, Na, Ca, Mg, K, Cl, SO4, PO4 and NO3. The pH (7.0 – 7.9) and TDS (12.26 – 19.35 mg/l) of the groundwater indicate that the water is neutral to slightly alkaline and fresh respectively. The order of relative abundance for concentration of cations in the groundwater is K>Ca>Na>Mg, while that of anions is Cl>SO4>NO3>PO4. The concentrations of potassium in about 90% and chloride in about 40 % of the samples were above the World Health Organization (WHO) standard for drinking water. Other hydro-geochemical parameters Ca, Mg, Na, SO4, PO4 and NO3 falls within the WHO standards. Values of Sodium Absorption Ratio (SAR) classified the water as excellent for irrigation purposes. The Soluble Sodium Percentage (SSP) values classified eighty percent of the water to be of good quality and the remaining samples as fair for irrigation purposes. However, using the Magnesium Adsorption Ratio (55.0 – 68.9 %) the samples may have hazardous effects on the soil. Generally, groundwater in the study area is suitable for both domestic and irrigation uses.

The invertebrate species play different roles above and below ground in a soil ecosystem. They are also responsible for a complex interaction, which is manifested in soils as self-organized systems of different sizes and functions. These invertebrates are sensitive to any changes in land management activities and soil physio-chemical properties. Therefore, the current study aims to examine the ecosystem functions of soil invertebrates from different agriculture fields and establish the relationships of soil physio-chemical properties. Soil samples were collected from four different fields: three are from the main campus, namely Universiti Putra Malaysia (Center of Environmental and Forensic Studies (CEFS), papaya and organic farm), and one is from Nilai, Negeri Sembilan. The soil physical properties, such as moisture, temperature, particle size, bulk density, and soil porosity, were recorded. Soil chemical properties (EC, pH, total C&N, C/N ratio, organic matter, extractable P, Ca, Mg, K, and Na, CEC) were also determined. Pitfall traps and Berlese funnel were used as sampling methods. The invertebrates were stored in 70% ethanol and identified using soil invertebrate morphological classification, which was classified based on body size and biological functional groups. Macrofauna composition in the papaya farm was abundant from other sites at 80.44%, and ecosystem engineers comprised 39.56%. By contrast, mesofauna was abundant with 43.22% in organic farms, and litter transformers were abundant in CEFS at 54.05%. Meanwhile, microfauna was high in organic farms at 2.85%. Predatory populations were also high in Nilai farm at 49.29%. The CCA analysis showed that the physical and chemical properties of soil influence soil fauna density and diversity. The present finding concludes that the activities of ecosystem functions of soil invertebrates were considerably affected by agricultural and management activities

The surface sediment samples of Kongsfjorden were analyzed for the Spheroidal Carbonaceous Particles (SCP) in an attempt to document the SCP in the environment other than previously studied inland lakes and snow and also to understand the probable source of industrial atmospheric pollution. The SCP are derived from the combustion of fossil fuels at high temperatures and are not produced naturally. They are chemically inert in both sediment and water and thus provide the indestructible record of atmospherically deposited pollutants in remote areas and anthropogenic impact on pristine environments. The SCP were recovered from different locations of the fjord and were classified according to their size range (small particles, 5-10 µm – large particles, 20-50 µm). The characterization of the spherical carbonaceous particles (shape, size, morphology, color etc.) was done under the light microscope. The detailed morphological features and chemical composition of SCP were studied using Scanning Electron Microscope equipped with Energy Dispersive X-Ray (SEM-EDX). The result shows that in term of a source of pollution in the area, long-range transportation is the major source of pollution but local sources cannot be ignored. This is a first attempt to study the SCP from the Kongsfjorden.

Hydrochemical characteristics and aquifer properties present a better understanding of the mitigation of groundwater pollution, which has become one of the leading environmental concerns and threats to the sustainable ecosystem. Seventy-seven groundwater samples were collected from Sargodha District (Pakistan) and characterized for their physical and chemical properties. The analytical data were processed for the evaluation of the processes that control the groundwater chemistry using various drinking and agricultural indices with statistical and hydrochemical modeling. The predominant hydrochemical type was found to be Ca-HCO3 type, followed by Na-HCO3 and Mg-Ca-Cl types. The present study showed that the main factors controlling the groundwater chemistry were the prevalent rock dominance alongside the weathering of silicates, solubilization of carbonates, and cation exchange processes. Entropy water quality index (EWQI) revealed that 6.51% represented “poor water,” while 7.79% were considered “extremely poor” for drinking purposes. However, USSL classification, Wilcox diagram, and other agricultural indices (RCS, SAR, %Na, MH, PI, and PS) showed that the majority of the samples were classified as suitable for irrigation purpose. However, 16% of the samples for %Na and 24% of the samples for MH were not suitable for agricultural purposes. Overall, the groundwater quality was affected by the anthropogenic stress in the study area.

I compared the effects of potassium sulfate and potassium chloride fertiliser, at rates up to 1280 kg/ ha K, on the concentrations in petioles of the youngest fully expanded leaves and in harvested tubers of potassium, calcium (tubers only), magnesium, phosphorus, chloride and sulfur (petioles only) and on potassium: magnesium ratios in petioles. For the sites classified as deficient and marginal according to soil test potassium concentrations, increased application rates of potassium sulfate increased the potassium and sulfur concentrations and potassium: magnesium ratios in petioles and decreased magnesium, phosphorus and chloride concentrations. In tubers both potassium and magnesium concentrations increased as potassium sulfate rates increased, while calcium and phosphorus concentrations showed negative trends. Compared with potassium sulfate, potassium chloride resulted in higher potassium, calcium and chloride concentrations and lower sulfur concentrations. Chloride concentrations in petioles from many sites were much higher than those reported in other studies and were related to high chloride concentrations in irrigation waters. Significant correlations were obtained between bicarbonate- extractable potassium concentrations in the surface (0-1 5-cm) soils and potassium concentrations in petioles (r2 = 0.75***) and tubers (r2 = 0.84***). As bicarbonate-extractable potassium concentrations in the surface soils increased from 40 to 600 mg/kg, the potassium concentrations in tubers increased from 1.69 to 2.69% compared with an increase from 5.72 to 1376% in petioles. Significant correlations were obtained between relative yields ((mean treatment yields/maximum mean treatment yie1d)x 100) and potassium concentrations (dry weight basis) in petioles and tubers and potassium:magnesium ratios in petioles. The percentage of variance in relative yield accounted for by each regression was 67.9, 31.0 and 56.9% respectively. The potassium concentration in petioles was the most effective plant index to predict yield response to potassium. For petioles and tubers prognostic critical potassium concentrations of 11.30 k 0.26% and 2.19 � 0.11% respectively were established. Alternatively, a prognostic critical potassium:magnesium ratio of 14.8 � 0.8% was determined. The choice of the mathematical model used influenced the critical values obtained.

Contaminants, in freezing ground or elsewhere in the world, are of concern not simply because of their presence but because of their potential for detrimental effects on human health, the biota, or other valued aspects of the environment. Understanding these effects is central to any attempt to manage or remediate contaminated land. The polar regions are different from other parts of the world, and it would be naïve to assume that the mass of information developed in temperate regions can be applied without modification to the polar regions. Despite their obvious environmental similarities, there are important differences between the Arctic and Antarctic. The landmass of the Arctic is much warmer than that of the Antarctic and as a result has a much greater diversity and abundance of flora. Because of its proximity to industrial areas in the Northern Hemisphere, the Arctic also experiences a higher input of contaminants via long-range aerial transport. In addition, the Arctic, with its indigenous population and generally undisputed territorial claims, has long been the subject of resource utilisation, including harvesting of living resources, mineral extraction, and the construction of military infrastructure. The history of human activity in Antarctica is relatively brief, but in this time there has been a series of quite distinct phases, culminating in the Antarctic now holding a unique position in the world. Activities in the Antarctic are governed by the Antarctic Treaty, which contains provisions dealing with environmental matters. The differences between the polar regions and the rest of the world, and between the Arctic and the Antarctic, significantly affect scientific and engineering approaches to the remediation of contamination in polar regions. This paper compares and contrasts the Arctic and Antarctic with respect to geography, configuration, habitation, logistics, environmental guidelines, regulations, and remediation protocols. Chemical contamination is also discussed in terms of its origin and major concerns and interests, particularly with reference to current remediation activities and site-restoration methodology.

Ferromanganese-oxide deposits dredged from four seamounts (Welker, Miller, Murray, and Patton) in the Gulf of Alaska Seamount Province include poorly crystallized microlaminated crusts on basalt substrate, well-crystallized Mn-oxide veins in epiclastic sedimentary rocks, and crystalline Mn-oxide layers and micronodules in phosphorite. The principal rock types dredged are alkali-basalt pillow fragments and tuffaceous conglomerate and sandstone. The glassy rims of pillow fragments, the glassy goundmass of large volcanic clasts, and the tuffaceous component of the sediment are altered to palagonite. Other low-temperature alteration products include phillipsite, smectite, and carbonate-apatite.Thick (10–50 mm) Fe–Mn crusts consist mainly of δ-MnO2; straight and cuspate growth laminae indicate variable growth rates and periods of nondeposition. A larger number of detrital particles toward the top of thick crusts record the increasing influence of active volcanoes of the Aleutian arc during northwestward movement of the Pacific plate. Thick crusts on basalt substrate have higher Mn/Fe ratios and lower Co content than Fe–Mn crusts from low-latitude seamounts of the central Pacific region. Thin (< 10 mm) crusts on volcaniclastic substrate contain todorokite and birnessite and have higher Mn/Fe ratios, Ni, and Cu and lower Fe and Co than thick Gulf of Alaska crusts.Veins of todorokite and cryptomelane with complex internal structure occur in altered tuffaceous sandstone and conglomerate from Miller Seamount. Fibrous todorokite has a composition similar to those of other marine examples but may contain up to 7% Mn2+ in M2 sites. Microprobe analysis of the marine cryptomelane indicates a composition that is approximately (K,Ba)1–2(Mn4+,Co)7–8O16∙x(H2O).A third type of Fe–Mn deposit in phosphorite is an intimate mixture of todorokite (and minor δ-MnO2)-bearing layers and micronodules, carbonate-apatite, and phillipsite that encloses grains of altered volcanic glass and lithic fragments.The microlaminated structure, mineralogy (predominantly δ-MnO2), and composition (Mn/Fe ratio and transition metal, rare earth element, U, and Th contents) of the thick crusts are characteristic of hydrogenetic Fe–Mn crusts elsewhere in the Pacific. Conversely, the crystallinity and chemical composition of the Mn-oxide veins and thin crusts indicate formation during diagenesis of the volcanogenic sediment substrate. Mn and other transition metals are mobilized during low-temperature oxidative alteration (palagonitization) of basaltic volcanic glass; the oxidation of Fe2+ to Fe3+ during palagonitization and the dissolution of the dilute biogenic fraction of the sediment combine to lower the Eh of ambient pore fluid and enhance the mobility of Mn2+. Diagenesis in the phosphatic sandstone from Patton Seamount involves organic-rich sediment and pore waters elevated in phosphorus owing to upwelling above a large volcanic edifice.

Secondary production of Chasmagnathus granulatus was calculated at the Refugio de Vida Silvestre Bahía Samborombón, Argentina (36° 16' S and 57° 06' W). Sampling was conducted on nine occasions between March 2001 and February 2003, crabs were collected by hand, physico-chemical variables, granulometry and organic matter contents of the sediments were registered. Crabs were classified as male, female and undifferentiated, measured (total carapace width: CW) and weighed (wet and dry weight: DW at 60 °C, during 48 hours). A correlation analysis between CW and DW was made. Morphometric growth of C. granulatus was by the application of the power function (y = a x b), where the carapace width (CW) was used as an independent variable. Males, females and undifferentiated individuals were analysed separately as well as all together as a group. The data were fitted indicating a positive allometry (constant of allometry b > 3), the males showing the greatest allometric value. The individuals (n = 957 juveniles and adults) were separated in cohorts by the polymodal width-frequency distribution converted into normal curves. Three cohorts were found during the whole study period, and two cohorts coexisting in each sampling date. Ovigerous females were caught on December 2001, 2002 and February 2003. The size-frequency method was used to estimate the annual production. The major contribution to production was carried out by the mature individuals, in particular those with size between 25 and 30 mm, but on the other hand, only few individuals measuring from 10 to 20 mm were collected. The annual production of C. granulatus was estimated in 7.76 g.m-2. The biomass (expressed as total dry weight) varied between 0.55 and 1.85 g.m-2, with the greater values being registered during autumn and spring, and the lower values during summer.

A comprehensive study was carried out to assess the microbiological, nutritional, biochemical and essential oil characteristics of three Egyptian traditional cultivars seeds, namely, cumin (Cuminum cyminum) and coriander (Coriandum sativum) spices as well as basil whole herb (Ocimum basilicum) collected from different Egyptian export centers as being ready for export. The found values for humidity in dry seeds of cumin (7.4%) and coriander (6.4%), as well as total ash and ash insoluble in acid (in cumin 7.7% and 0.74%, but in coriander 5.3% and 0.55%, respectively), were lower than the maximum limits indicated by the Egyptian Specification Standards (ES) and by International Standards Organization (ISO) for cumin and coriander seeds. Analysis of essential minerals in seed spices and herbs indicated that they were rich in K, Ca, Na, Fe and Zn. The total bacterial count was low content in seeds of cumin and coriander as well as fresh whole basil herb. The microbiological load in all tested seed spices and herbs was found lower than those indicated by the ES and ISO for cumin and coriander seeds. Yields in hydro-distilled essential oils (EOs) were the highest in cumin seeds (3.762%), while both coriander and basil herbs had lower amounts (0.285% and 0.686%, respectively). EOs contents were found higher than the maximum limits for cumin (1.5% - 2.5% on a dry weight basis), but within the limits for coriander (0.1% - 0.5% on a dry weight basis) as indicated by the ES and ISO for cumin and coriander seed oils. Gas chromatography of extracted EOs from seeds of cumin and coriander as well as basil herbs indicated the presence of 41, 35 and 47 compounds, respectively, where cumin aldehyde was the major component in cumin volatiles but was linalool in volatiles of both coriander seeds and basil herbs. EOs of basil herbs grown in Egypt were of the high linalool chemotype which was characterized by high contents of linalool and relatively lower amounts of eugenol. However, the major compounds in the three tested EOs from seeds or herbs grown in Egypt are in accordance with literature reports from different parts of the world. Volatile oil components in EOs of the three tested Egyptian spices and herbs were classified into groups, based on the relative area (%). The proportion of the major and the other main components in EOs from seeds of cumin and coriander cultivars were within the ranges indicated by both the ES and ISO for cumin seed oils (cumin aldehyde between 15% - 46%) and for coriander seed oils (linalool between 65% - 78%). The aim of the present work was to Assessment study certain commonly used Egyptian spices and herbal products for characterizing their physical, biochemical and microbiological properties.

The swell-shrink behavior of expansive clay soil cause problems in the construction such as the occurrence of bumpy road, cracking and various other problems, therefore stabilization of the soil is needed to improve the behavior of the soil.Red brick is a building material that contains chemical compounds that can increase the compressive strength of a building. Red brick waste which is rarely used is then reused as a cement mixture.This study aims to determine the type and classification of soil in Cibingbin village, a village located in Cibaliung sub-district, Pandeglang district-Banten province, based on the USCS soil classification system, and to determine the value of unconfined compression test (qu) with the addition of red brick powder with variations the percentage additions were 0%, 5%, 10%, and 15% with curing times of 0, 7, 14, and 28 days.From the results of the study, it was found that the soil classification in Cibingbin village was classified as high plasticity organic clay (OH) according to the USCS classification system. The lowest qu value was obtained at 0 days of curing with the percentage of added ingredients 0%, 5%, 10%, 15% increasing the qu value was 1,316 kg/cm2, 1,655 kg/cm2, 2,242 kg/cm2; 2,607 kg/cm2, and the largest qu value was obtained at 14 days of curing with the percentage of added material 0%, 5%. , 10%, 15% obtained 1,639 kg/cm2, 3,189 kg/cm2, 3,440 kg/cm2, dan 3,603 kg/cm2. Keywords: Stabilization, Unconfined Compression Strength, Waste Red Brick

Low-resolution Geological Survey of Pakistan (GSP) maps surrounding the region of interest show oolitic and fossiliferous limestone occurrences correspondingly in Samanasuk, Lockhart, and Margalla hill formations in the Hazara division, Pakistan. Machine-learning algorithms (MLAs) have been rarely applied to multispectral remote sensing data for differentiating between limestone formations formed due to different depositional environments, such as oolitic or fossiliferous. Unlike the previous studies that mostly report lithological classification of rock types having different chemical compositions by the MLAs, this paper aimed to investigate MLAs’ potential for mapping subclasses within the same lithology, i.e., limestone. Additionally, selecting appropriate data labels, training algorithms, hyperparameters, and remote sensing data sources were also investigated while applying these MLAs. In this paper, first, oolitic (Samanasuk), fossiliferous (Lockhart and Margalla) limestone-bearing formations along with the adjoining Hazara formation were mapped using random forest (RF), support vector machine (SVM), classification and regression tree (CART), and naïve Bayes (NB) MLAs. The RF algorithm reported the best accuracy of 83.28% and a Kappa coefficient of 0.78. To further improve the targeted allochemical limestone formation map, annotation labels were generated by the fusion of maps obtained from principal component analysis (PCA), decorrelation stretching (DS), X-means clustering applied to ASTER-L1T, Landsat-8, and Sentinel-2 datasets. These labels were used to train and validate SVM, CART, NB, and RF MLAs to obtain a binary classification map of limestone occurrences in the Hazara division, Pakistan using the Google Earth Engine (GEE) platform. The classification of Landsat-8 data by CART reported 99.63% accuracy, with a Kappa coefficient of 0.99, and was in good agreement with the field validation. This binary limestone map was further classified into oolitic (Samanasuk) and fossiliferous (Lockhart and Margalla) formations by all the four MLAs; in this case, RF surpassed all the other algorithms with an improved accuracy of 96.36%. This improvement can be attributed to better annotation, resulting in a binary limestone classification map, which formed a mask for improved classification of oolitic and fossiliferous limestone in the area.

In numerous species, egg chemoattractants play a critical role in guiding sperm towards unfertilized eggs (sperm chemotaxis). Until now, the known functions of sperm chemotaxis include increasing the effective target size of eggs, thereby promoting sperm–egg encounters, and facilitating species recognition. Here, we report that in the broadcast spawning mussel, Mytilus galloprovincialis , egg chemoattractants may play an unforeseen role in sexual selection by enabling sperm to effectively ‘choose’ between the eggs of different conspecific females. In an initial experiment, we confirmed that sperm chemotaxis occurs in M. galloprovincialis by showing that sperm are attracted towards unfertilized eggs when given the choice of eggs or no eggs in a dichotomous chamber. We then conducted two cross-classified mating experiments, each comprising the same individual males and females crossed in identical male × female combinations, but under experimental conditions that offered sperm ‘no-choice’ (each fertilization trial took place in a Petri dish and involved a single male and female) or a ‘choice’ of a female's eggs (sperm were placed in the centre of a dichotomous choice chamber and allowed to choose eggs from different females). We show that male-by-female interactions characterized fertilization rates in both experiments, and that there was remarkable consistency between patterns of sperm migration in the egg-choice experiment and fertilization rates in the no-choice experiment. Thus, sperm appear to exploit chemical cues to preferentially swim towards eggs with which they are most compatible during direct sperm-to-egg encounters. These results reveal that sperm differentially select eggs on the basis of chemical cues, thus exposing the potential for egg chemoattractants to mediate mate choice for genetically compatible partners. Given the prevalence of sperm chemotaxis across diverse taxa, our findings may have broad implications for sexual selection in other mating systems.

Background The argus snakehead (Ophiocephalus argus Cantor) is a highly nutritious, freshwater, cultured bony fish with a high economic value. The health of the fish is closely related to its blood cells, which are critical for oxygen transport, natural defense, and immunity. We investigated the morphometry, microstructure, and cytochemical characteristics of the peripheral blood cells of O. argus. Our results may provide the basic reference values needed to monitor the health of this fish for large-scale cultivation. Methods The number of blood cells in O. argus were counted on a hemocytometer and their size was measured using a micrometer under light microscope. The morphology and classification of the blood cells were studied using Wright’s staining and the cytochemical characteristics were studied using seven chemical stains including peroxidase (POX), Sudan black B (SBB), periodic acid-Schiff (PAS), acid phosphatase (ACP), alkaline phosphatase (ALP), chloroacetic acid AS-D naphthol esterase (AS-D), and α-naphthol acetate esterase (α-NAE). Results The peripheral blood cells in O. argus can be classified as erythrocytes, leukocytes, and thrombocytes; of which, females had 2.9597 million/mm3, 88,400/mm3, and 43,600/mm3, respectively, and males had 3.0105 million/mm3, 105,500/mm3, and 34,000/mm3, respectively. Leukocytes consisted of neutrophils, monocytes, large lymphocytes, and small lymphocytes. Eosinophils and basophils were not found. Monocytes were the most numerous leukocytes identified, followed by neutrophils and small lymphocytes, while large lymphocytes were the least frequently identified. Cytochemical staining showed that erythrocytes were only positive for PAS staining. Neutrophils were strongly positive for POX, SBB, and ACP, and positive for all the other cytochemical stains. Monocytes were positive for PAS and α-NAE and were weakly positive for ACP and AS-D staining. Large lymphocytes were positive for PAS and were weakly positive for ALP, AS-D, and α-NAE staining. Small lymphocytes were positive for PAS and weakly positive for AS-D and α-NAE staining. Thrombocytes were positive for PAS and were weakly positive for ACP and AS-D, but negative for the remaining cytochemical stains. The morphology of peripheral blood cells in O. argus was generally similar to that of other fish species, while the cytochemical staining patterns showed clear species specificity.

Food image recognition system has various applications now a day. In this paper, we have used a machine learning supervised approach and Support Vector Machine to classify different food images. SVM has been classified to detect and recognize food images with the least modification. By applying various filters like a texture filter, a segmentation method, clustering, and a SVM approach we have achieved more accuracy than other machine learning approaches with manually extracting features. Sustenance is an indivisible piece of people groups lives. we tend to apply a convolution neural network (CNN) to the undertakings of analyst work and perceiving sustenance pictures. Clarification for the wide decent variety of styles of nourishment, and picture acknowledgment of sustenance things are typically unpleasant difficulties. Nevertheless, profound learning has been demonstrated starting late to be a genuinely extreme picture acknowledgment framework, and CNN could be a dynamic approach to managing profound learning. CNN showed on a very basic level higher precision than did old-fashioned help vector-machine-based courses with carefully assembled decisions. For sustenance picture disclosure, CNN likewise demonstrated fundamentally count higher precision than a standard technique. Generally higher precision than standard techniques.

Abstract Introduction Multiple Myeloma (MM) is considered a malignancy of post germinal center long-lived plasma cells. Nevertheless T-cell independent antigen stimulation before the exposure of the B-cell to the germinal center can happen and results to IgM secreting short lived plasma cells and lymphoplasmacytes representing thus a potential alternative normal counterpart for IgM plasma cell dyscrasias. IgM myeloma is an infrequent subtytpe of MM with an estimated prevalence of 0.5%. Due to its rarity little is known about its characteristics and prognosis in comparison with Waldestrom’s macroglobulinemia (WM) and the other MM subtypes. Purpose To identify the characteristics and the prognosis of IgM MM, and compare it predominantly with WM and subsequently with the rest of the MM subtypes. Methods We interogatted our Multiple Myeloma Data Base for cases of IgM MM and their respective Overall Survival (OS), Progression Free Survival (PFS), bone disease as defined by x-Rays, PET-CT and MRI, Gene Expression Profile (GEP), and common disease characteristics (anemia,calcium, creatinine) and compare it to the prognosis of WM and non-IgM MM. Diagnosis was based on the morphological and immunophenotypical findings of pathologically examined biopsy specimens along with the presence or not of typical clinical characteristics of MM (lytic bone lesions, hypercalcemia, renal failure) or typical clinical characteristics of WM (organomegaly, lymphadenopathy). Results There were 22 confirmed IgM MM cases. 14 of them presented at MIRT at initial diagnosis while 8 had previously been treated elsewhere. Osteolytic bone lesions and/or pathological fractures by x-ray and CT examination were evident in 16 cases. For the remaining 6 cases active bone focal lesions by either MRI or PET were identified in three. There was no organomegaly evident in cases with an available PET/CT at baseline, while only one had evidence of hilar and mediastinal lymphadenopathy along with calcified lung nodules. Elevated creatinine levels (>2.0 mg/dl) were evident in 4 cases at initial diagnosis. Their disease characteristics are depicted in the table 1. Median OS for IgM MM was 4.9 years while PFS could not be accurately estimated due to lack of data on patients treated elsewhere. Median OS for a historical control of 158 WM cases in MIRT was 9.2 years (Clin Lymphoma Myeloma Leuk. 11(1):139-42). Median OS of the WM group remained largely unaffected, even when the subgroup of the WM cases requiring treatment was analyzed (9.0 years).To further clarify if the IgM MM differs in terms of OS from the other isotypes of MM, we compared the IgM group to a group of 61 non-IgM MM cases which were matched by important prognostic clinical factors (age, creatinine> 2mg/dl, LDH>190u/L, b-2M >5.5mg/dl and Albumin<3.5gr/dl). No statistical difference was found for OS (p=0.846). Out of 22 cases, 14 of them had available GEP data on initial diagnosis. In 6 of these cases the cyclin D1 gene expression was high enough to be consistent with a t(11;14) translocation at FISH analysis, one case was consistent with a t(14;16) translocation, one with a t(4;14) translocation and two more were classified as belonging to the hyperdiploid subgroup. A comparative genomic analysis was performed on the IgM MM, the non-IgM MM and WM cases with available GEP data at initial diagnosis (14, 61 and 42 cases respectively). 1155 probesets that had expression level significantly different between WM and non IgM MM (FDR<3E-06) were identified. Then, the expression values of these 1155 probesets in all GEP samples, including WM, non IgM MM, and IgM MM, were used to build a clustering tree. We found that IgM MM mainly clustered with non IgM MM, supporting the findings of the clinical data. Conclusion IgM MM is a discrete clinical entity that should be distinguished from WM. Bone disease is evident in the majority of the cases, especially when specialized radiological techniques are incorporated at the initial work up. It holds a distinct prognosis from WM, while when balanced for prognostic factors that hold importance in MM it does not differ from the other MM isotypes. Finally analysis of the genetic data further supports the resemblance between IgM MM and the non IgM MM, and the difference with WM. Disclosures: Zhang: University of Arkansas for Medical Sciences: Co-inventor of the DNA probes for FISH of IGHC/IGHV (14q32), MMSET/FGFR3 (4p16), CCND3 (6p21), CCND1 (11q13), MAF (16q23), and MAFB (20q12) loci, sub. to the US Patent & Trademark Office as Prov. App# 61/726,327: Methods of Detecting 14q32 Translocations, Co-inventor of the DNA probes for FISH of IGHC/IGHV (14q32), MMSET/FGFR3 (4p16), CCND3 (6p21), CCND1 (11q13), MAF (16q23), and MAFB (20q12) loci, sub. to the US Patent & Trademark Office as Prov. App# 61/726,327: Methods of Detecting 14q32 Translocations Patents & Royalties.

The following seven papers were presented on May 16, 1984, at the Geological Association of Canada and Mineralogical Association of Canada joint annual meeting. The special session, organized by R. W. Macqueen and J. A. Coope, contained 10 papers and was sponsored by the Mineral Deposits Division of the Geological Association of Canada.Our objective in organizing the special session was to examine organically based processes and relationships that may be of major importance to the origin of ore deposits. As noted by Fyfe (1984), the concept of the geochemical cycle focuses attention on pathways of chemical elements and isotopes of the Earth's system during geologic history. It is clear from the chemistry of carbon-rich materials that a wide range of elements is concentrated directly or indirectly by biological processes operating as part of the geochemical cycle. Two of the papers of the special session examine some of these concentration processes, although definitive links to actual ore deposits cannot be made yet. Beveridge and Fyfe document the remarkable ability of the anionic cell walls of certain bacteria to concentrate metals and to provide sites for nucleation and growth of minerals. In a related paper, Mann and Fyfe show that several species of simple freshwater green algae readily concentrate large amounts of uranium under both experimental and natural conditions (Elliot Lake and Thames River, Ontario).Two papers deal with aspects of sulphate reduction. Birnbaum and Wireman describe controlled experiments that suggest that sulphate-reducing bacteria may be involved in the selective replacement of sulphate-evaporite minerals by silica and in the precipitation of silica in association with sulphide mineral phases in banded iron formations. Their work focuses directly on the effect that bacterial sulphate reduction has on silica solubility. Trudinger et al. examine the question of mechanisms of sulphate reduction at temperatures less than 200 °C and the bearing this has on origin of sulphide for low-temperature sulphide ore deposits. Although there is empirical evidence favouring abiological sulphate reduction at temperatures in the vicinity of 100 °C, Trudinger et al. have not been able to demonstrate abiological reduction of sulphate under controlled laboratory conditions and at temperatures under about 200 °C. Perhaps catalysts, as yet undiscovered, are involved in this process in nature.Impressive progress has been made in understanding the diagenetic evolution of organic matter in response to heat and pressure in geological environments: excellent reviews are found in Barnes et al. (1984) and Bustin et al. (1985). Simoneit's paper examines and reviews the genesis of petroleum in a most unusual setting, that of the active ocean ridge spreading centre of Guaymas Basin, Gulf of California. There, in the vicinity of black smokers and associated metallic sulphide deposits, petroleum originates instantaneously geologically as a result of hydrothermal activity. The question of genetic involvement of organic matter in the origin of the metallic sulphides (e.g., reduction of sulphate to H2S) cannot be answered yet for this setting with the available data.The final two special session papers included here are concerned with organic matter associated with mineralization in Canadian Shield Precambrian settings. Willingham et al. demonstrate that Elliot Lake – Blind River Early Proterozoic uranium deposits with minor amounts of associated gold also contain kerogen-like organic matter. Some of this organic matter has anomalously rich amounts of gold and uranium and appears to have originated as mats of cyanobacteria, possibly with the ability to concentrate these metals. For a number of settings in the Archean-aged Abitibi greenstone belt of Ontario and Quebec, Springer demonstrates that carbon, at least partly of organic origin, is closely associated with some gold deposits. Her interpretation is that carbon activated by shear-zone-associated hydrothermal fluids has provided sites for fixing some of the gold.Three of the papers given at the special session are not included here. H. T. Shacklette reviewed metal uptake by young conifer trees, demonstrating that nursery-grown seedlings of several species readily concentrated a variety of metals, including lead, zinc, tin, and gold, over a 7 year period. This work is of interest to those involved in geochemical prospecting and is now published elsewhere (King et al. 1984). R. W. Macqueen presented quantitative data on the genesis of sulphide by abiological bitumen–sulphate reactions at the Pine Point lead–zinc property, Northwest Territories, Canada (Macqueen and Powell 1983; Powell and Macqueen 1984). Although Trudinger et al. have not been able to demonstrate abiological reduction of sulphate at temperatures approximating those of Pine Point [Formula: see text], the data presented by Macqueen (Powell and Macqueen 1984) are consistent with the amounts, alteration, and composition of bitumens at Pine Point, as well as with the presence of native sulphur and the sulphur isotope compositions of the various Pine Point sulphur species. This work is continuing, and a more extensive account is in preparation. J. R. Watterson examined relationships between freezing climates and the local chemical behaviour of gold in the weathering cycle, concluding that ice-induced accumulation of organic acids, bacteria, and other organic matter at mineral surfaces may increase rates of chemical attack, leading to dissolution of normally insoluble metals such as gold (Watterson 1986).Interest in organic aspects of the geochemical cycle, including ore deposition, is growing dramatically (e.g., Fyfe 1984). Although the following papers address a limited range of topics within the field, they do indicate some of the diversity and variety of active processes and associations between metallic elements and organic components. Perhaps, in the not too distant future, we will be able to identify or even discover whole classes of ore deposits that owe their origin directly to organic influences operating within the geochemical cycle.

AbstractThe question of the origin of life interested people for centuries. All existing views on this subject can be classified into different areas of human knowledge about the world: natural sciences, philosophy, and theology (religion). It is interesting to look at them closer and to classify all the typical groups of the theories about the origins of life. We can then see links existing between them and relationships that indicate their own nature. Nowadays, driving forces of prebiological chemical evolution and the mode of explanation of the transition ‘non-life into life’ give a great variety of solutions. The differences between the theories, however, as well as the current controversies in the scientific community (what ‘in the beginning’ was?; where and when the prebiotic evolution may took place? etc.) will be shown as of secondary importance for the theories’ systematization in comparison with several much more profound philosophical assumptions underlying the origin-of-life-studies. My proposal to organize and classify different types of the theories of genesis of life allows for extracting conceptions of different kind: metaphysical and scientific, and also for comparing them with each other. Some of them answer the question of the emergence of life in general and others on the question of the origin of life on the Earth only. In the perspective of contemporary scientific research on the origin of life it seems interesting that two main ideas concerning the problem of the origin of life, spontaneous generation and panspermia, are still present as presuppositions of certain theories but have been modified. Thus a ‘philosophical key’ seems to be the most appropriate to systematize all kinds of theories on the origin of life. This paper is an attempt to justify the position adopted. Most important conclusion is that the philosophical basis or implications, which are irreducibly present and possible to be found within the scientific theories of the origin of life, indicates that this problem is not just the strictly scientific one; it is the philosophical problem, too; thus it cannot be fully solved merely through referring to the empirical aspect of biogenesis.

Environments containing significant concentration of NaCl such as salt lakes harbor extremophiles microorganisms which have a great biotechnology interest. To explore the diversity of Bacteria in Chott Tinsilt (Algeria), an isolation program was performed. Water samples were collected from the saltern during the pre-salt harvesting phase. This Chott is high in salt (22.47% (w/v). Seven halophiles Bacteria were selected for further characterization. The isolated strains were able to grow optimally in media with 10–25% (w/v) total salts. Molecular identification of the isolates was performed by sequencing the 16S rRNA gene. It showed that these cultured isolates included members belonging to the Halomonas, Staphylococcus, Salinivibrio, Planococcus and Halobacillus genera with less than 98% of similarity with their closest phylogenetic relative. The halophilic bacterial isolates were also characterized for the production of biosurfactant and industrially important enzymes. Most isolates produced hydrolases and biosurfactants at high salt concentration. In fact, this is the first report on bacterial strains (A4 and B4) which were a good biosurfactant and coagulase producer at 20% and 25% ((w/v)) NaCl. In addition, the biosurfactant produced by the strain B4 at high salinity (25%) was also stable at high temperature (30-100°C) and high alkalinity (pH 11).Key word: Salt Lake, Bacteria, biosurfactant, Chott, halophiles, hydrolases, 16S rRNAINTRODUCTIONSaline lakes cover approximately 10% of the Earth’s surface area. The microbial populations of many hypersaline environments have already been studied in different geographical regions such as Great Salt Lake (USA), Dead Sea (Israel), Wadi Natrun Lake (Egypt), Lake Magadi (Kenya), Soda Lake (Antarctica) and Big Soda Lake and Mono Lake (California). Hypersaline regions differ from each other in terms of geographical location, salt concentration and chemical composition, which determine the nature of inhabitant microorganisms (Gupta et al., 2015). Then low taxonomic diversity is common to all these saline environments (Oren et al., 1993). Halophiles are found in nearly all major microbial clades, including prokaryotic (Bacteria and Archaea) and eukaryotic forms (DasSarma and Arora, 2001). They are classified as slight halophiles when they grow optimally at 0.2–0.85 M (2–5%) NaCl, as moderate halophiles when they grow at 0.85–3.4 M (5–20%) NaCl, and as extreme halophiles when they grow at 3.4–5.1 M (20–30%) NaCl. Hyper saline environments are inhabited by extremely halophilic and halotolerant microorganisms such as Halobacillus sp, Halobacterium sp., Haloarcula sp., Salinibacter ruber , Haloferax sp and Bacillus spp. (Solomon and Viswalingam, 2013). There is a tremendous demand for halophilic bacteria due to their biotechnological importance as sources of halophilic enzymes. Enzymes derived from halophiles are endowed with unique structural features and catalytic power to sustain the metabolic and physiological processes under high salt conditions. Some of these enzymes have been reported to be active and stable under more than one extreme condition (Karan and Khare, 2010). Applications are being considered in a range of industries such as food processing, washing, biosynthetic processes and environmental bioremediation. Halophilic proteases are widely used in the detergent and food industries (DasSarma and Arora, 2001). However, esterases and lipases have also been useful in laundry detergents for the removal of oil stains and are widely used as biocatalysts because of their ability to produce pure compounds. Likewise, amylases are used industrially in the first step of the production of high fructose corn syrup (hydrolysis of corn starch). They are also used in the textile industry in the de-sizing process and added to laundry detergents. Furthermore, for the environmental applications, the use of halophiles for bioremediation and biodegradation of various materials from industrial effluents to soil contaminants and accidental spills are being widely explored. In addition to enzymes, halophilic / halotolerants microorganisms living in saline environments, offer another potential applications in various fields of biotechnology like the production of biosurfactant. Biosurfactants are amphiphilic compounds synthesized from plants and microorganisms. They reduce surface tension and interfacial tension between individual molecules at the surface and interface respectively (Akbari et al., 2018). Comparing to the chemical surfactant, biosurfactant are promising alternative molecules due to their low toxicity, high biodegradability, environmental capability, mild production conditions, lower critical micelle concentration, higher selectivity, availability of resources and ability to function in wide ranges of pH, temperature and salinity (Rocha et al., 1992). They are used in various industries which include pharmaceuticals, petroleum, food, detergents, cosmetics, paints, paper products and water treatment (Akbari et al., 2018). The search for biosurfactants in extremophiles is particularly promising since these biomolecules can adapt and be stable in the harsh environments in which they are to be applied in biotechnology.OBJECTIVESEastern Algeria features numerous ecosystems including hypersaline environments, which are an important source of salt for food. The microbial diversity in Chott Tinsilt, a shallow Salt Lake with more than 200g/L salt concentration and a superficies of 2.154 Ha, has never yet been studied. The purpose of this research was to chemically analyse water samples collected from the Chott, isolate novel extremely or moderate halophilic Bacteria, and examine their phenotypic and phylogenetic characteristics with a view to screening for biosurfactants and enzymes of industrial interest.MATERIALS AND METHODSStudy area: The area is at 5 km of the Commune of Souk-Naâmane and 17 km in the South of the town of Aïn-Melila. This area skirts the trunk road 3 serving Constantine and Batna and the railway Constantine-Biskra. It is part the administrative jurisdiction of the Wilaya of Oum El Bouaghi. The Chott belongs to the wetlands of the High Plains of Constantine with a depth varying rather regularly without never exceeding 0.5 meter. Its length extends on 4 km with a width of 2.5 km (figure 1).Water samples and physico-chemical analysis: In February 2013, water samples were collected from various places at the Chott Tinsilt using Global Positioning System (GPS) coordinates of 35°53’14” N lat. and 06°28’44”E long. Samples were collected randomly in sterile polythene bags and transported immediately to the laboratory for isolation of halophilic microorganisms. All samples were treated within 24 h after collection. Temperature, pH and salinity were measured in situ using a multi-parameter probe (Hanna Instruments, Smithfield, RI, USA). The analytical methods used in this study to measure ions concentration (Ca2+, Mg2+, Fe2+, Na+, K+, Cl−, HCO3−, SO42−) were based on 4500-S-2 F standard methods described elsewhere (Association et al., 1920).Isolation of halophilic bacteria from water sample: The media (M1) used in the present study contain (g/L): 2.0 g of KCl, 100.0/200.0 g of NaCl, 1.0 g of MgSO4.7HO2, 3.0 g of Sodium Citrate, 0.36 g of MnCl2, 10.0 g of yeast extract and 15.0 g agar. The pH was adjusted to 8.0. Different dilutions of water samples were added to the above medium and incubated at 30°C during 2–7 days or more depending on growth. Appearance and growth of halophilic bacteria were monitored regularly. The growth was diluted 10 times and plated on complete medium agar (g/L): glucose 10.0; peptone 5.0; yeast extract 5.0; KH2PO4 5.0; agar 30.0; and NaCl 100.0/200.0. Resultant colonies were purified by repeated streaking on complete media agar. The pure cultures were preserved in 20% glycerol vials and stored at −80°C for long-term preservation.Biochemical characterisation of halophilic bacterial isolates: Bacterial isolates were studied for Gram’s reaction, cell morphology and pigmentation. Enzymatic assays (catalase, oxidase, nitrate reductase and urease), and assays for fermentation of lactose and mannitol were done as described by Smibert (1994).Optimization of growth conditions: Temperature, pH, and salt concentration were optimized for the growth of halophilic bacterial isolates. These growth parameters were studied quantitatively by growing the bacterial isolates in M1 medium with shaking at 200 rpm and measuring the cell density at 600 nm after 8 days of incubation. To study the effect of NaCl on the growth, bacterial isolates were inoculated on M1 medium supplemented with different concentration of NaCl: 1%-35% (w/v). The effect of pH on the growth of halophilic bacterial strains was studied by inoculating isolates on above described growth media containing NaCl and adjusted to acidic pH of 5 and 6 by using 1N HCl and alkaline pH of 8, 9, 10, 11 and 12 using 5N NaOH. The effect of temperature was studied by culturing the bacterial isolates in M1 medium at different temperatures of incubation (4°C–55°C).Screening of halophilic bacteria for hydrolytic enzymes: Hydrolase producing bacteria among the isolates were screened by plate assay on starch, tributyrin, gelatin and DNA agar plates respectively for amylase, lipase, protease and DNAse activities. Amylolytic activity of the cultures was screened on starch nutrient agar plates containing g/L: starch 10.0; peptone 5.0; yeast extract 3.0; agar 30.0; NaCl 100.0/250.0. The pH was 7.0. After incubation at 30 ºC for 7 days, the zone of clearance was determined by flooding the plates with iodine solution. The potential amylase producers were selected based on ratio of zone of clearance diameter to colony diameter. Lipase activity of the cultures was screened on tributyrin nutrient agar plates containing 1% (v/v) of tributyrin. Isolates that showed clear zones of tributyrin hydrolysis were identified as lipase producing bacteria. Proteolytic activity of the isolates was similarly screened on gelatin nutrient agar plates containing 10.0 g/L of gelatin. The isolates showing zones of gelatin clearance upon treatment with acidic mercuric chloride were selected and designated as protease producing bacteria. The presence of DNAse activity on plates was determined on DNAse test agar (BBL) containing 10%-25% (w/v) total salt. After incubation for 7days, the plates were flooded with 1N HCl solution. Clear halos around the colonies indicated DNAse activity (Jeffries et al., 1957).Milk clotting activity (coagulase activity) of the isolates was also determined following the procedure described (Berridge, 1952). Skim milk powder was reconstituted in 10 mM aqueous CaCl2 (pH 6.5) to a final concentration of 0.12 kg/L. Enzyme extracts were added at a rate of 0.1 mL per mL of milk. The coagulation point was determined by manual rotating of the test tube periodically, at short time intervals, and checking for visible clot formation.Screening of halophilic bacteria for biosurfactant production. Oil spread Assay: The Petridis base was filled with 50 mL of distilled water. On the water surface, 20μL of diesel and 10μl of culture were added respectively. The culture was introduced at different spots on the diesel, which is coated on the water surface. The occurrence of a clear zone was an indicator of positive result (Morikawa et al., 2000). The diameter of the oil expelling circles was measured by slide caliber (with a degree of accuracy of 0.02 mm).Surface tension and emulsification index (E24): Isolates were cultivated at 30 °C for 7 days on the enrichment medium containing 10-25% NaCl and diesel oil as the sole carbon source. The medium was centrifuged (7000 rpm for 20 min) and the surface tension of the cell-free culture broth was measured with a TS90000 surface tensiometer (Nima, Coventry, England) as a qualitative indicator of biosurfactant production. The culture broth was collected with a Pasteur pipette to remove the non-emulsified hydrocarbons. The emulsifying capacity was evaluated by an emulsification index (E24). The E24 of culture samples was determined by adding 2 mL of diesel oil to the same amount of culture, mixed for 2 min with a vortex, and allowed to stand for 24 h. E24 index is defined as the percentage of height of emulsified layer (mm) divided by the total height of the liquid column (mm).Biosurfactant stability studies : After growth on diesel oil as sole source of carbone, cultures supernatant obtained after centrifugation at 6,000 rpm for 15 min were considered as the source of crude biosurfactant. Its stability was determined by subjecting the culture supernatant to various temperature ranges (30, 40, 50, 60, 70, 80 and 100 °C) for 30 min then cooled to room temperature. Similarly, the effect of different pH (2–11) on the activity of the biosurfactant was tested. The activity of the biosurfactant was investigated by measuring the emulsification index (El-Sersy, 2012).Molecular identification of potential strains. DNA extraction and PCR amplification of 16S rDNA: Total cellular DNA was extracted from strains and purified as described by Sambrook et al. (1989). DNA was purified using Geneclean® Turbo (Q-BIO gene, Carlsbad, CA, USA) before use as a template in polymerase chain reaction (PCR) amplification. For the 16S rDNA gene sequence, the purified DNA was amplified using a universal primer set, forward primer (27f; 5′-AGA GTT TGA TCM TGG CTC AG) and a reverse primer (1492r; 5′-TAC GGY TAC CTT GTT ACG ACT T) (Lane, 1991). Agarose gel electrophoresis confirmed the amplification product as a 1400-bp DNA fragment.16S rDNA sequencing and Phylogenic analysis: Amplicons generated using primer pair 27f-1492r was sequenced using an automatic sequencer system at Macrogene Company (Seoul, Korea). The sequences were compared with those of the NCBI BLAST GenBank nucleotide sequence databases. Phylogenetic trees were constructed by the neighbor-joining method using MEGA version 5.05 software (Tamura et al., 2011). Bootstrap resembling analysis for 1,000 replicates was performed to estimate the confidence of tree topologies.Nucleotide sequence accession numbers: The nucleotide sequences reported in this work have been deposited in the EMBL Nucleotide Sequence Database. The accession numbers are represented in table 5.Statistics: All experiments were conducted in triplicates. Results were evaluated for statistical significance using ANOVA.RESULTSPhysico-chemical parameters of the collected water samples: The physicochemical properties of the collected water samples are reported in table 1. At the time of sampling, the temperature was 10.6°C and pH 7.89. The salinity of the sample, as determined in situ, was 224.70 g/L (22,47% (w/v)). Chemical analysis of water sample indicated that Na +and Cl- were the most abundant ions (table 1). SO4-2 and Mg+2 was present in much smaller amounts compared to Na +and Cl- concentration. Low levels of calcium, potassium and bicarbonate were also detected, often at less than 1 g/L.Characterization of isolates. Morphological and biochemical characteristic feature of halophilic bacterial isolates: Among 52 strains isolated from water of Chott Tinsilt, seven distinct bacteria (A1, A2, A3, A4, B1, B4 and B5) were chosen for further characterization (table 2). The colour of the isolates varied from beige, pale yellow, yellowish and orange. The bacterial isolates A1, A2, A4, B1 and B5 were rod shaped and gram negative (except B5), whereas A3 and B4 were cocci and gram positive. All strains were oxidase and catalase positive except for B1. Nitrate reductase and urease activities were observed in all the bacterial isolates, except B4. All the bacterial isolates were negative for H2S formation. B5 was the only strain positive for mannitol fermentation (table 2).We isolated halophilic bacteria on growth medium with NaCl supplementation at pH 7 and temperature of 30°C. We studied the effect of NaCl, temperature and pH on the growth of bacterial isolates. All the isolates exhibited growth only in the presence of NaCl indicating that these strains are halophilic. The optimum growth of isolates A3 and B1 was observed in the presence of 10% NaCl, whereas it was 15% NaCl for A1, A2 and B5. A4 and B4 showed optimum growth in the presence of 20% and 25% NaCl respectively. A4, B4 and B5 strains can tolerate up to 35% NaCl.The isolate B1 showed growth in medium supplemented with 10% NaCl and pH range of 7–10. The optimum pH for the growth B1 was 9 and they did not show any detectable growth at or below pH 6 (table 2), which indicates the alkaliphilic nature of B1 isolate. The bacterial isolates A1, A2 and A4 exhibited growth in the range of pH 6–10, while A3 and B4 did not show any growth at pH greater than 8. The optimum pH for growth of all strains (except B1) was pH 7.0 (table 2). These results indicate that A1, A2, A3, A4, B4 and B5 are neutrophilic in nature. All the bacterial isolates exhibited optimal growth at 30°C and no detectable growth at 55°C. Also, detectable growth of isolates A1, A2 and A4 was observed at 4°C. However, none of the bacterial strains could grow below 4°C and above 50°C (table 2).Screening of the halophilic enzymes: To characterize the diversity of halophiles able to produce hydrolytic enzymes among the population of microorganisms inhabiting the hypersaline habitats of East Algeria (Chott Tinsilt), a screening was performed. As described in Materials and Methods, samples were plated on solid media containing 10%-25% (w/v) of total salts and different substrates for the detection of amylase, protease, lipase and DNAse activities. However, coagulase activity was determined in liquid medium using milk as substrate (figure 3). Distributions of hydrolytic activity among the isolates are summarized in table 4.From the seven bacterial isolates, four strains A1, A2, A4 and B5 showed combined hydrolytic activities. They were positive for gelatinase, lipase and coagulase. A3 strain showed gelatinase and lipase activities. DNAse activities were detected with A1, A4, B1 and B5 isolates. B4 presented lipase and coagulase activity. Surprisingly, no amylase activity was detected among all the isolates.Screening for biosurfactant producing isolates: Oil spread assay: The results showed that all the strains could produce notable (>4 cm diameter) oil expelling circles (ranging from 4.11 cm to 4.67 cm). The average diameter for strain B5 was 4.67 cm, significantly (P < 0.05) higher than for the other strains.Surface tension and emulsification index (E24): The assimilation of hydrocarbons as the sole sources of carbon by the isolate strains led to the production of biosurfactants indicated by the emulsification index and the lowering of the surface tension of cell-free supernatant. Based on rapid growth on media containing diesel oil as sole carbon source, the seven isolates were tested for biosurfactant production and emulsification activity. The obtained values of the surface tension measurements as well as the emulsification index (E24) are shown in table 3. The highest reduction of surface tension was achieved with B5 and A3 isolates with values of 25.3 mN m−1 and 28.1 mN m−1 respectively. The emulsifying capacity evaluated by the E24 emulsification index was highest in the culture of isolate B4 (78%), B5 (77%) and A3 (76%) as shown in table 3 and figure 2. These emulsions were stable even after 4 months. The bacteria with emulsification indices higher than 50 % and/or reduction in the surface tension (under 30 mN/m) have been defined as potential biosurfactant producers. Based on surface tension and the E24 index results, isolates B5, B4, A3 and A4 are the best candidates for biosurfactant production. It is important to note that, strains B4 and A4 produce biosurfactant in medium containing respectively 25% and 20% (w/v) NaCl.Stability of biosurfactant activities: The applicability of biosurfactants in several biotechnological fields depends on their stability at different environmental conditions (temperatures, pH and NaCl). For this study, the strain B4 appear very interesting (It can produce biosurfactant at 25 % NaCl) and was choosen for futher analysis for biosurfactant stability. The effects of temperature and pH on the biosurfactant production by the strain B4 are shown in figure 4.biosurfactant in medium containing respectively 25% and 20% (w/v) NaCl.Stability of biosurfactant activities: The applicability of biosurfactants in several biotechnological fields depends on their stability at different environmental conditions (temperatures, pH and NaCl). For this study, the strain B4 appear very interesting (It can produce biosurfactant at 25 % NaCl) and was chosen for further analysis for biosurfactant stability. The effects of temperature and pH on the biosurfactant production by the strain B4 are shown in figure 4. The biosurfactant produced by this strain was shown to be thermostable giving an E-24 Index value greater than 78% (figure 4A). Heating of the biosurfactant to 100 °C caused no significant effect on the biosurfactant performance. Therefore, the surface activity of the crude biosurfactant supernatant remained relatively stable to pH changes between pH 6 and 11. At pH 11, the value of E24 showed almost 76% activity, whereas below pH 6 the activity was decreased up to 40% (figure 4A). The decreases of the emulsification activity by decreasing the pH value from basic to an acidic region; may be due to partial precipitation of the biosurfactant. This result indicated that biosurfactant produced by strain B4 show higher stability at alkaline than in acidic conditions.Molecular identification and phylogenies of potential isolates: To identify halophilic bacterial isolates, the 16S rDNA gene was amplified using gene-specific primers. A PCR product of ≈ 1.3 kb was detected in all the seven isolates. The 16S rDNA amplicons of each bacterial isolate was sequenced on both strands using 27F and 1492R primers. The complete nucleotide sequence of 1336,1374, 1377,1313, 1305,1308 and 1273 bp sequences were obtained from A1, A2, A3, A4, B1, B4 and B5 isolates respectively, and subjected to BLAST analysis. The 16S rDNA sequence analysis showed that the isolated strains belong to the genera Halomonas, Staphylococcus, Salinivibrio, Planococcus and Halobacillus as shown in table 5. The halophilic isolates A2 and A4 showed 97% similarity with the Halomonas variabilis strain GSP3 (accession no. AY505527) and the Halomonas sp. M59 (accession no. AM229319), respectively. As for A1, it showed 96% similarity with the Halomonas venusta strain GSP24 (accession no. AY553074). B1 and B4 showed for their part 96% similarity with the Salinivibrio costicola subsp. alcaliphilus strain 18AG DSM4743 (accession no. NR_042255) and the Planococcus citreus (accession no. JX122551), respectively. The bacterial isolate B5 showed 98% sequence similarity with the Halobacillus trueperi (accession no. HG931926), As for A3, it showed only 95% similarity with the Staphylococcus arlettae (accession no. KR047785). The 16S rDNA nucleotide sequences of all the seven halophilic bacterial strains have been submitted to the NCBI GenBank database under the accession number presented in table 5. The phylogenetic association of the isolates is shown in figure 5.DICUSSIONThe physicochemical properties of the collected water samples indicated that this water was relatively neutral (pH 7.89) similar to the Dead Sea and the Great Salt Lake (USA) and in contrast to the more basic lakes such as Lake Wadi Natrun (Egypt) (pH 11) and El Golea Salt Lake (Algeria) (pH 9). The salinity of the sample was 224.70 g/L (22,47% (w/v). This range of salinity (20-30%) for Chott Tinsilt is comparable to a number of well characterized hypersaline ecosystems including both natural and man-made habitats, such as the Great Salt Lake (USA) and solar salterns of Puerto Rico. Thus, Chott Tinsilt is a hypersaline environment, i.e. environments with salt concentrations well above that of seawater. Chemical analysis of water sample indicated that Na +and Cl- were the most abundant ions, as in most hypersaline ecosystems (with some exceptions such as the Dead Sea). These chemical water characteristics were consistent with the previously reported data in other hypersaline ecosystems (DasSarma and Arora, 2001; Oren, 2002; Hacěne et al., 2004). Among 52 strains isolated from this Chott, seven distinct bacteria (A1, A2, A3, A4, B1, B4 and B5) were chosen for phenotypique, genotypique and phylogenetique characterization.The 16S rDNA sequence analysis showed that the isolated strains belong to the genera Halomonas, Staphylococcus, Salinivibrio, Planococcus and Halobacillus. Genera obtained in the present study are commonly occurring in various saline habitats across the globe. Staphylococci have the ability to grow in a wide range of salt concentrations (Graham and Wilkinson, 1992; Morikawa et al., 2009; Roohi et al., 2014). For example, in Pakistan, Staphylococcus strains were isolated from various salt samples during the study conducted by Roohi et al. (2014) and these results agreed with previous reports. Halomonas, halophilic and/or halotolerant Gram-negative bacteria are typically found in saline environments (Kim et al., 2013). The presence of Planococcus and Halobacillus has been reported in studies about hypersaline lakes; like La Sal del Rey (USA) (Phillips et al., 2012) and Great Salt Lake (Spring et al., 1996), respectively. The Salinivibrio costicola was a representative model for studies on osmoregulatory and other physiological mechanisms of moderately halophilic bacteria (Oren, 2006).However, it is interesting to note that all strains shared less than 98.7% identity (the usual species cut-off proposed by Yarza et al. (2014) with their closest phylogenetic relative, suggesting that they could be considered as new species. Phenotypic, genetic and phylogenetic analyses have been suggested for the complete identification of these strains. Theses bacterial strains were tested for the production of industrially important enzymes (Amylase, protease, lipase, DNAse and coagulase). These isolates are good candidates as sources of novel enzymes with biotechnological potential as they can be used in different industrial processes at high salt concentration (up to 25% NaCl for B4). Prominent amylase, lipase, protease and DNAase activities have been reported from different hypersaline environments across the globe; e.g., Spain (Sánchez‐Porro et al., 2003), Iran (Rohban et al., 2009), Tunisia (Baati et al., 2010) and India (Gupta et al., 2016). However, to the best of our knowledge, the coagulase activity has never been detected in extreme halophilic bacteria. Isolation and characterization of crude enzymes (especially coagulase) to investigate their properties and stability are in progress.The finding of novel enzymes with optimal activities at various ranges of salt concentrations is of great importance. Besides being intrinsically stable and active at high salt concentrations, halophilic and halotolerant enzymes offer great opportunities in biotechnological applications, such as environmental bioremediation (marine, oilfiel) and food processing. The bacterial isolates were also characterized for production of biosurfactants by oil-spread assay, measurement of surface tension and emulsification index (E24). There are few reports on biosurfactant producers in hypersaline environments and in recent years, there has been a greater increase in interest and importance in halophilic bacteria for biomolecules (Donio et al., 2013; Sarafin et al., 2014). Halophiles, which have a unique lipid composition, may have an important role to play as surface-active agents. The archae bacterial ether-linked phytanyl membrane lipid of the extremely halophilic bacteria has been shown to have surfactant properties (Post and Collins, 1982). Yakimov et al. (1995) reported the production of biosurfactant by a halotolerant Bacillus licheniformis strain BAS 50 which was able to produce a lipopeptide surfactant when cultured at salinities up to 13% NaCl. From solar salt, Halomonas sp. BS4 and Kocuria marina BS-15 were found to be able to produce biosurfactant when cultured at salinities of 8% and 10% NaCl respectively (Donio et al., 2013; Sarafin et al., 2014). In the present work, strains B4 and A4 produce biosurfactant in medium containing respectively 25% and 20% NaCl. To our knowledge, this is the first report on biosurfactant production by bacteria under such salt concentration. Biosurfactants have a wide variety of industrial and environmental applications (Akbari et al., 2018) but their applicability depends on their stability at different environmental conditions. The strain B4 which can produce biosurfactant at 25% NaCl showed good stability in alkaline pH and at a temperature range of 30°C-100°C. Due to the enormous utilization of biosurfactant in detergent manufacture the choice of alkaline biosurfactant is researched (Elazzazy et al., 2015). On the other hand, the interesting finding was the thermostability of the produced biosurfactant even after heat treatment (100°C for 30 min) which suggests the use of this biosurfactant in industries where heating is of a paramount importance (Khopade et al., 2012). To date, more attention has been focused on biosurfactant producing bacteria under extreme conditions for industrial and commercial usefulness. In fact, the biosurfactant produce by strain B4 have promising usefulness in pharmaceutical, cosmetics and food industries and for bioremediation in marine environment and Microbial enhanced oil recovery (MEOR) where the salinity, temperature and pH are high.CONCLUSIONThis is the first study on the culturable halophilic bacteria community inhabiting Chott Tinsilt in Eastern Algeria. Different genera of halotolerant bacteria with different phylogeneticaly characteristics have been isolated from this Chott. Culturing of bacteria and their molecular analysis provides an opportunity to have a wide range of cultured microorganisms from extreme habitats like hypersaline environments. Enzymes produced by halophilic bacteria show interesting properties like their ability to remain functional in extreme conditions, such as high temperatures, wide range of pH, and high salt concentrations. These enzymes have great economical potential in industrial, agricultural, chemical, pharmaceutical, and biotechnological applications. Thus, the halophiles isolated from Chott Tinsilt offer an important potential for application in microbial and enzyme biotechnology. In addition, these halo bacterial biosurfactants producers isolated from this Chott will help to develop more valuable eco-friendly products to the pharmacological and food industries and will be usefulness for bioremediation in marine environment and petroleum industry.ACKNOWLEDGMENTSOur thanks to Professor Abdelhamid Zoubir for proofreading the English composition of the present paper.CONFLICT OF INTERESTThe authors declare that they have no conflict of interest.Akbari, S., N. H. Abdurahman, R. M. Yunus, F. Fayaz and O. R. Alara, 2018. Biosurfactants—a new frontier for social and environmental safety: A mini review. Biotechnology research innovation, 2(1): 81-90.Association, A. P. H., A. W. W. Association, W. P. C. Federation and W. E. Federation, 1920. Standard methods for the examination of water and wastewater. American Public Health Association.Baati, H., R. Amdouni, N. Gharsallah, A. Sghir and E. Ammar, 2010. Isolation and characterization of moderately halophilic bacteria from tunisian solar saltern. Current microbiology, 60(3): 157-161.Berridge, N., 1952. Some observations on the determination of the activity of rennet. Analyst, 77(911): 57b-62.DasSarma, S. and P. Arora, 2001. Halophiles. Encyclopedia of life sciences. Nature publishishing group: 1-9.Donio, M. B. S., F. A. Ronica, V. T. Viji, S. Velmurugan, J. S. C. A. Jenifer, M. Michaelbabu, P. Dhar and T. Citarasu, 2013. Halomonas sp. Bs4, a biosurfactant producing halophilic bacterium isolated from solar salt works in India and their biomedical importance. SpringerPlus, 2(1): 149.El-Sersy, N. A., 2012. Plackett-burman design to optimize biosurfactant production by marine Bacillus subtilis n10. Roman biotechnol lett, 17(2): 7049-7064.Elazzazy, A. M., T. Abdelmoneim and O. Almaghrabi, 2015. Isolation and characterization of biosurfactant production under extreme environmental conditions by alkali-halo-thermophilic bacteria from Saudi Arabia. Saudi journal of biological Sciences, 22(4): 466-475.Graham, J. E. and B. Wilkinson, 1992. Staphylococcus aureus osmoregulation: Roles for choline, glycine betaine, proline, and taurine. Journal of bacteriology, 174(8): 2711-2716.Gupta, S., P. Sharma, K. Dev and A. Sourirajan, 2016. Halophilic bacteria of lunsu produce an array of industrially important enzymes with salt tolerant activity. Biochemistry research international, 1: 1-10.Gupta, S., P. Sharma, K. Dev, M. Srivastava and A. Sourirajan, 2015. A diverse group of halophilic bacteria exist in lunsu, a natural salt water body of Himachal Pradesh, India. SpringerPlus 4(1): 274.Hacěne, H., F. Rafa, N. Chebhouni, S. Boutaiba, T. Bhatnagar, J. C. Baratti and B. Ollivier, 2004. Biodiversity of prokaryotic microflora in el golea salt lake, Algerian Sahara. Journal of arid environments, 58(3): 273-284.Jeffries, C. D., D. F. Holtman and D. G. Guse, 1957. Rapid method for determining the activity of microorgan-isms on nucleic acids. Journal of bacteriology, 73(4): 590.Karan, R. and S. Khare, 2010. Purification and characterization of a solvent‐stable protease from Geomicrobium sp. Emb2. Environmental technology, 31(10): 1061-1072.Khopade, A., R. Biao, X. Liu, K. Mahadik, L. Zhang and C. Kokare, 2012. Production and stability studies of the biosurfactant isolated from marine Nocardiopsis sp. B4. Desalination, 3: 198-204.Kim, K. K., J.-S. Lee and D. A. Stevens, 2013. Microbiology and epidemiology of Halomonas species. Future microbiology, 8(12): 1559-1573.Lane, D., 1991. 16s/23s rRNA sequencing in nucleic acid techniques in bacterial systematics. Stackebrandt e., editor;, and goodfellow m., editor. Chichester, UK: John Wiley & Sons.Morikawa, K., R. L. Ohniwa, T. Ohta, Y. Tanaka, K. Takeyasu and T. Msadek, 2009. Adaptation beyond the stress response: Cell structure dynamics and population heterogeneity in Staphylococcus aureus. Microbes environments, 25: 75-82.Morikawa, M., Y. Hirata and T. J. B. e. B. A.-M. Imanaka, 2000. A study on the structure–function relationship of lipopeptide biosurfactants. Biochimica et biophysica acta, 1488(3): 211-218.Oren, A., 2002. Diversity of halophilic microorganisms: Environments, phylogeny, physiology, and applications. Journal of industrial microbiology biotechnology, 28(1): 56-63.Oren, A., 2006. Halophilic microorganisms and their environments. Springer science & business media.Oren, A., R. Vreeland and L. Hochstein, 1993. Ecology of extremely halophilic microorganisms. The biology of halophilic bacteria, 2(1): 1-8.Phillips, K., F. Zaidan, O. R. Elizondo and K. L. Lowe, 2012. Phenotypic characterization and 16s rDNA identification of culturable non-obligate halophilic bacterial communities from a hypersaline lake, la sal del rey, in extreme south texas (USA). Aquatic biosystems, 8(1): 1-5.Post, F. and N. Collins, 1982. A preliminary investigation of the membrane lipid of Halobacterium halobium as a food additive 1. Journal of food biochemistry, 6(1): 25-38.Rocha, C., F. San-Blas, G. San-Blas and L. Vierma, 1992. Biosurfactant production by two isolates of Pseudomonas aeruginosa. World Journal of microbiology biotechnology, 8(2): 125-128.Rohban, R., M. A. Amoozegar and A. Ventosa, 2009. Screening and isolation of halophilic bacteria producing extracellular hydrolyses from howz soltan lake, Iran. Journal of industrial microbiology biotechnology, 36(3): 333-340.Roohi, A., I. Ahmed, N. Khalid, M. Iqbal and M. Jamil, 2014. Isolation and phylogenetic identification of halotolerant/halophilic bacteria from the salt mines of Karak, Pakistan. International journal of agricultural and biology, 16: 564-570.Sambrook, J., E. F. Fritsch and T. Maniatis, 1989. Molecular cloning: A laboratory manual, 2nd edn. Cold spring harbor laboratory, cold spring harbor, New York.Sánchez‐Porro, C., S. Martin, E. Mellado and A. Ventosa, 2003. Diversity of moderately halophilic bacteria producing extracellular hydrolytic enzymes. Journal of applied microbiology, 94(2): 295-300.Sarafin, Y., M. B. S. Donio, S. Velmurugan, M. Michaelbabu and T. Citarasu, 2014. Kocuria marina bs-15 a biosurfactant producing halophilic bacteria isolated from solar salt works in India. Saudi journal of biological sciences, 21(6): 511-519.Smibert, R., 1994. Phenotypic characterization. In methods for general and molecular bacteriology. American society for microbiology: 611-651.Solomon, E. and K. J. I. Viswalingam, 2013. Isolation, characterization of halotolerant bacteria and its biotechnological potentials. International journal scientific research paper publication sites, 4: 1-7.Spring, S., W. Ludwig, M. Marquez, A. Ventosa and K.-H. Schleifer, 1996. Halobacillus gen. Nov., with descriptions of Halobacillus litoralis sp. Nov. and Halobacillus trueperi sp. Nov., and transfer of Sporosarcina halophila to Halobacillus halophilus comb. Nov. International journal of systematic evolutionary microbiology, 46(2): 492-496.Tamura, K., D. Peterson, N. Peterson, G. Stecher, M. Nei and S. Kumar, 2011. Mega5: Molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Molecular biology evolution, 28(10): 2731-2739.Yakimov, M. M., K. N. Timmis, V. Wray and H. L. Fredrickson, 1995. Characterization of a new lipopeptide surfactant produced by thermotolerant and halotolerant subsurface Bacillus licheniformis bas50. Applied and environmental microbiology, 61(5): 1706-1713.Yarza, P., P. Yilmaz, E. Pruesse, F. O. Glöckner, W. Ludwig, K.-H. Schleifer, W. B. Whitman, J. Euzéby, R. Amann and R. Rosselló-Móra, 2014. Uniting the classification of cultured and uncultured bacteria and archaea using 16s rRNA gene sequences. Nature reviews microbiology, 12(9): 635-645

At Jeongseon Maedun Cave Site, the Bronze Age human skeletons are almost completely restored and preserved in good condition. It is of great importance, as the Site shows the burial cave customs of a type of tomb that is little investigated in Korean peninsula. The burial cave comes under a mass tomb in which several individuals are buried, including at least three persons, adults and children. It may be a joint tomb (family tomb) judging from the aspect of excavation and the sex and age of the buried. In particular, human bones in Tomb No. 1 with the most remaining area were buried by burying aside above an ash layer that was intentionally built. The stone chamber is assumed to be a stone coffin, and burial goods include arrowhead and red burnished pottery. Based on the result of an analysis of sex and age, No. 1 is of a woman in her mid-30s to 50s; No. 2, of a late-middle-aged to elderly man; and No. 3, of a young child aged 3-6. Measurement of bones in No. 1 reveals that the height is about 151-156㎝ and the cranium type is classified to be high and long. High cranium type is unique characteristic of the inhabitants of the Korean peninsula like other ancient human skeletons; However, the narrow and long cranium type(hyperdolichocrany, the cranial index 66.9) can be differentiated from that. This morphometric characteristic is considered as an unique and inherent feature that the residents around Maedun Cave shared. It has high correlation in the cranium type and archaeological aspects with the Bronze Age human skeletons excavated in Hwangseok-ri, Jecheon. As for pathological features, porotic hyperostosis and cribra orbitalia were found in the cranium; oral cavity and periodontal diseases, in the upper jaw and teeth; and osteoarthritis in the spine and ribs. Lifestyles including diseases, health, and nutritional status could be inferred from such features. Furthermore, the stable isotope analysis, which is of a chemical analytic method of diet, reveals that the proportion of total animal protein consumed in the diet was low, whereas the intake of farm crops(C4 plant), such as millet, foxtail millet and broomcorn was high. This dietary characteristic corresponds with the results of Hwangseok-ri, Jecheon and Joongdo, Chuncheon, enabling the restoration of dietary life of central region residents of the Korean peninsula in the Bronze Age.

Signet-ring cell carcinoma of the urinary bladder is an uncommon histopathology variant of carcinoma of urinary bladder which has been stated to account for 0.5% and 2% of primary malignant tumours of the urinary bladder. Signet-ring cell carcinoma of the urinary bladder is stated to either arise from the wall of the urinary bladder or from remnants of the urachus, or signet-ring cell carcinoma of the urinary bladder could also develop as a metastatic tumour that has ensued a primary signet-ring cell carcinoma that had arisen from a number sites of the body some of which include: the stomach, colon, or breast, the appendix and other organs. It has been iterated that the least common type of signet-ring cell carcinoma is primary signet-ring cell carcinoma and that up to 2013 less than 100 cases had been reported. Signet-ring cell carcinoma of the urinary bladder can affect males as well as females, young individuals or adults. Signet-ring cell carcinoma of the urinary bladder could be diagnosed incidentally or it may present with non-specific symptoms that simulate the symptoms of other urinary bladder tumours including: lower urinary tract symptoms, haematuria, abdominal pain / discomfort or loin pain, retention of urine, feeling unwell, or weight loss. Microscopy examination of the tumour whether it was obtained by means of trans-urethral resection or by cystectomy would tend to demonstrate a tumour that is comprised of signet-ring cells that contain peripherally pushed hyperchromatic nuclei, intra-cytoplasmic mucin, as well as lakes of extracellular mucin. The tumour cells could be arranged in lobules, and separated by fibrovascular septae. There tends to be visualization of mitosis as well as evidence of necrosis. The tumour tends to be seen within the underlying stroma and quite often within the detrusor muscle and up to the extra-vesical fat quite often. Immunohistochemistry staining studies of signet-ring cell carcinoma of the urinary bladder would tend to show tumour cells that exhibit positive staining for: Cytokeratin including cytokeratin 7, CAM 5.2, AE1/3, and 34ßE12; Vimentin; Peanut lectin agglutinin; Ulex europaeus agglutinin. In signet ring cell carcinoma of urinary bladder immunohistochemistry staining of the tumour may also show tumour cells that exhibit positive staining for the following tumour markers: CK, CK7, CK20; CDX2; Villin - There could be a small amount of positive staining for Villin. In signet-ring cell carcinoma of the urinary bladder, immunohistochemistry studies of the tumour may demonstrate tumour cells that do exhibit negative staining for the ensuing tumour markers: Vimentin, (this does show therefore that some tumours would stain positive and others would stain negative); GATA3, and P53. To confirm whether a signet-ring cell carcinoma of the urinary bladder is a pure primary tumour or metastatic tumour does require detailed history taking with evidence of previously treated signet-ring cell carcinoma elsewhere and comparing the pathology features of the tumours, the undertaking of radiology imaging including ultrasound scan, computed tomography scan or magnetic resonance imaging scan of abdomen and pelvis as well as upper gastrointestinal endoscopy and lower gastrointestinal endoscopy to ascertain if there are any lesions within the gastrointestinal tract and taking biopsies of any suspicious lesion found for pathology examination and comparing the features of the lesions with the urinary tract tumours. There is no consensus opinion of the treatment of signet-ring cell carcinomas of the urinary bladder even though it has been realised that primary signet ring cell carcinomas have tended to be more invasive and higher staged as well as associated with very poor prognosis in comparison with the traditional urothelial carcinoma. Treatment options that have been utilized have included: trans-urethral resection of tumour, radical cystectomy alone or radical cystectomy plus adjuvant therapy and despite utilization of radical cystectomy and adjuvant therapy majority of patients tend to die. There are sporadic reports of isolated cases of good short-term, medium-term, and long-term survival usually if the tumour is diagnosed at an early stage. Early diagnosis, aggressive complete surgical excision of primary and metastatic signet-ring cell carcinomas and utilization appropriate combination adjuvant therapies would provide the best treatment of curative intent. Additionally, there is an anecdotal report of an effective treatment of an advanced metastatic primary signet-ring cell carcinoma of the urinary bladder with utilization of docetaxel which resulted in destruction of the tumour cells without an operation which would indicate that some chemotherapy agents could be good enough for the successful treatment of signet-ring cell carcinomas of the bladder including primary and metastatic tumours. Therefore, it is possible that novel treatment options of treatment of signet-ring cell carcinoma of the urinary bladder including an appropriate chemotherapy plus additional non-operative treatments including cryotherapy, radiotherapy, radiofrequency ablation, irreversible electroporation, selective angiography and chemical infusion of chemotherapy agents into the tumour plus immunotherapy could be explored as treatment options. There is a global need for urologists, oncologists, and pharmacotherapy research workers to identify new chemotherapy medicaments that would safely and effectively destroy primary and metastatic signet-ring cell tumours in order to improve upon the outcome of the disease. A global multi-centre trial of various aggressive treatment options should be commenced quickly.

Polychlorinated dibezno-p-dioxins (PCDD) and dibenzofurans (PCDF) are compounds with similar structure, physicochemical and toxicological properties. They are not used commercially, they are formed as by-products during certain industrial processes, combustion, failures, etc. LD50 values (0.002-300 mg / kg) depend on the species of animal tested and the chemical structure of the compound. On chronic toxicity, the information available mainly relates to 2,3,7,8-TCDD and 2,3,4,7,8-PeCDF. Potential routes of human exposure are: the digestive system, lungs and skin. These compounds are accumulated mainly in the liver and adipose tissue. Their polar metabolites may undergo conjugation with glucuronic acid and glutathione. The main routes of excretion are bile and feces. In mammals, PCDD / F are also eliminated in breast milk. The results of mutagenicity and genotoxicity tests of PCDD (mainly 2,3,7,8-TCDD) and PCDF and their effects on fertility and reproduction are inconsistent. Among PCDD and PCDF, the compound that most strongly affects fertility, reproduction and fetal development is 2,3,7,8-TCDD. Epidemiological studies are the basis for assessing the carcinogenic potential of dioxins (including 2,3,7,8-TCDD) and furans in humans. Cohorts include those professionally exposed to chlorophenols, phenoxyacetic herbicides and a mixture of polychlorinated dibenzodioxins and furans. PCDD / F have a common mechanism of toxic action associated with the Ah receptor. PCDD / F are considered to be inducers of several enzymes (e.g. CYP1A) and modulators of hormones and growth factors. CYP1A1 activity is one of the most sensitive indicators of exposure to 2,3,7,8-TCDD. Adenocarcinomas and hepatocellular carcinomas as well as bile ducts have been found in rats and mice after 2,3,7,8-TCDD. Tumor changes have also been observed in other organs. NTP studies also showed carcinogenic effects 2,3,4,7,8-PeCDF. According to IARC, sufficient evidence of a carcinogenic effect on humans exists only for 2,3,7,8-TCDD (CAS: 1746-01-6) and 2,3,4,7,8 PeCDF (CAS: 57117-31-4). Other PCDD / F cannot be classified as carcinogenic to humans. The basis for determining the MAC value for the mixture of PCDD and PCDF was the results of the assessment of the risk of developing additional liver cancer in people exposed in the work environment for 2,3,7,8-TCDD in 2017. This risk was estimated at 1 · 10-4 for 40 years of exposure to the compound at a concentration of 18 pg / m3. In the case of combined exposure, the content of polychlorinated dibenzo-p-dioxins and furans in the tested samples, as well as their maximum acceptable levels are expressed in the form of the so-called toxicity equivalent (TEQ). For the PCDD and PCDF mixture, we propose the value of 18 pg WHO2006-TEQ / m3. The result expressed as pg WHO-TEQ / m3 is not a de facto concentration, but a determination of the toxicity of the mixture of dioxin and furan congeners contained in the sample in relation to TCDD. This article discusses the problems of occupational safety and health, which are covered by health sciences and environmental engineering.

Aim: To make a comparison between functional results in intra-articular displaced calcaneus fractures which are treated conservatively and operative treatment Study design: A randomized controlled trial Place and Duration:This study was conducted at ward 17 JPMC Karachi from January 20221 to January 2022. Methodology: This study includes 32 individuals who had displaced intra-articular calcaneal fractures. These individuals were classified into two groups, each group with 16 individuals. They were randomly assigned to either operative or conservative treatment. Clinical results, as well as radiological results (Modified Rowe’s score), were assessed at a 1-year follow-up. Results: All results were measured using Modified Rowe’s score. With a 1-year follow-up of the operative group, according to Modified Rowe’s score, 6 individuals had excellent results representing 37.5%, 8 individuals had good results representing 50%, and only 2 individuals had satisfactory results representing 12.5%. On the other hand, with a 1-year follow-up of the conservative group, only 2 individuals had excellent results representing 12.5%, 7 individuals had good results representing 43.75%, 5 individuals had satisfactory results representing 31.25%, and 2 individuals had poor results representing 12.5%. A few challenges were seen in each group. In the operative group, delayed wound healing was a complication seen in 2 cases while in the conservative group, peroneal tendonitis, subtalar arthritis and malunion were seen as a complication. Conclusion: In conclusion, the operative group was seen to be more effective to cure displaced intra-articular calcaneus fractures rather than the comparative group. Only a few complications, better functional recovery, early rehabilitation, and early weight bearing were seen in the operative treatment. Keywords: Calcaneus fractures, functional outcome, rehabilitation, conservative treatment

Os igarapés urbanos são afetados pela supressão da floresta ripária o que ocasiona degradação ambiental. Devido às atividades antrópicas, esses ambientes são alterados e descaracterizados, sofrendo profundas mudanças em suas estruturas aquática e terrestre. Essas alterações se refletem na qualidade das águas, e usualmente são estimadas através da análise quantitativa de parâmetros hidrológicos e físico-químicos. O Índice de Integridade do Habitat (IIH) é um método qualitativo de avaliação das condições ambientais dos igarapés. Este estudo busca avaliar se a variação da integridade do habitat reflete a variação hidrológica e físico-química decorrente das alterações antrópicas nos ambientes aquáticos. Foram medidas as variáveis ambientais da zona ripária (Índice de Integridade do Habitat - IIH e cobertura vegetal média); do canal (vazão, largura e profundidade médias) e da água (pH, condutividade, oxigênio dissolvido e turbidez médias, temperatura média) em de seis igarapés urbanos de Santarém- PA. Dos seis igarapés analisados, cinco encontram-se em estado degradado e com a área de proteção permanente em desacordo com o Código Florestal Brasileiro. Foi avaliada a correlação do IIH com as variáveis ambientais e análise de variância para verificar se houve diferença significativa entre os valores das variáveis ambientais nos pontos amostrados. Verificou-se que o Índice de Integridade do Habitat - IIH apresentou uma correlação significativa (p<0,05) com as variáveis oxigênio dissolvido e condutividade, sendo capaz de captar a influência destas variáveis na qualidade da água. Em termos de variação espacial dos parâmetros físico-químicos, o Ponto 6 se destacou com o maior número de diferenças significativas em relação aos demais pontos amostrados e foi o único classificado pelo IIH como íntegro, demonstrando que o IIH foi capaz de detectar a variação da qualidade da água e que pode ser uma ferramenta para um rápido e efetivo diagnóstico das condições ambientais de corpos d’água. Diante dos resultados, medidas de recuperação devem ser tomadas para minimizar e reverter as fontes de impacto que promovem a degradação dos igarapés urbanos na cidade de Santarém- PA.Palavras-chave: Gestão de microbacias urbanas, qualidade da água, mata ciliar. Effects of the environmental integrity of the riparian zone and its influence on the water quality of urban streams in Santarém-Pa, Amazonia, Brazil A B S T R A C TUrban streams are affected by the suppression of riparian forest, which causes environmental degradation. Due to human activities, these environments are altered and uncharacterized, undergoing profound changes in their aquatic and terrestrial structures. These changes are reflected in the water quality and are usually estimated through the quantitative analysis of hydrological and physical-chemical parameters. The Habitat Integrity Index (IIH) is a qualitative method for assessing the environmental conditions of streams. This study aims to assess whether the variation in habitat integrity reflects the hydrological and physical-chemical variation resulting from anthropic changes in aquatic environments. The environmental variables from the riparian zone (Habitat Integrity Index - IIH and average vegetation cover), from the channel (mean flow, width, and depth) and from the water (pH, average conductivity, dissolved oxygen and turbidity, average temperature) were measured; in six urban streams in Santarém-PA. Among the six streams analyzed, five are in a degraded state and have a permanent protection area that does not comply with the Brazilian Forest Code. The correlation of the IIH with the environmental variables and an analysis of variance was performed in order to verify if there was a significant difference between the environmental variables’ values at the sampled points. It was found that the Habitat Integrity Index - IIH showed a significant correlation (p <0.05) with the variables dissolved oxygen and conductivity, being able to capture the influence of these variables on water quality. In terms of spatial variation of the physical-chemical parameters, the Point 6 stood out with the largest number of significant differences in relation to the other sampled points and was the only one classified by the IIH as intact, which demonstrates that the IIH could detect the variation of the water quality and that it can be a tool for a quick and effective diagnosis of the environmental conditions of water bodies. In view of the results, recovery measures must be taken to minimize and reverse the sources of impact that promote the degradation of urban streams in the city of Santarém-PA.Keywords Management of urban micro basins, water quality, riparian forest.

The question of the origin of life interested people for centuries. All existing views on this subject can be classified into different areas of our knowledge of the world: natural sciences, philosophy, and theology. Some theories (perhaps the majority) contain more or less explicit elements from all of these areas. Thus, it is helpful to take a closer look at them and to classify all the typical groups of theories about the origins of life. We can in this way stress their mutual connections and clarify their own nature. Nowadays, driving forces of pre-biological chemical evolution and the explanation of the transition from “non-life into life” present a great variety of solutions. The differences between the theories, however, as well as the current controversies in the scientific community (e.g., what was “in the beginning”?; where did prebiotic evolution take place? etc.), will be shown to be of secondary importance in comparison with several much more profound philosophical assumptions underlying the origin-of-life-studies. The attempt to organize and classify different types of theories on the genesis of life allows to take into account different kinds of perspectives (theistic, philosophical and scientific), and to compare them to each other. The most general division between theories is based on a distinction between metaphysical conceptions and scientific ones. Some theories answer the question of the emergence of life in general, whereas others tackle the question of the origin of life on Earth only. Interestingly, two traditional ideas concerning the problem of the origin of life (i.e., spontaneous generation and panspermia) are still at play in contemporary scientific research, albeit in a modified form. In the perspective of contemporary scientific research on the origin of life it seems interesting that two main ideas concerning the problem of the origin of life, spontaneous generation and panspermia, are still present as presuppositions of certain theories but have been modified. Moreover, it is evident that the theistic view of the origin of life (creation) does not have to fall into conflict with contemporary scientific theories. Rather, they are complementary. This article is an extension, explanation and refinement of the proposed scheme of the main types of theories on the origin of life. An attempt to classify various biogenesis theories is also proposed. One of the most important questions that will be addressed concerns the philosophical presumptions of biogenetics still informing current research as well as scientific explanations of the origin of life.

Abstract Background: Adult T-cell leukemia-lymphoma (ATL), which is a hematological malignancy related to human T-lymphotropic virus type I (HTLV-1), is known as a malignant lymphoid disease with poor prognosis. Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is considered a treatment modality to contribute prolonging the survival in some population of patients (Katsuya H et al. 2015 Blood). A small study indicated that ATL cells frequently have chromosomal abnormalities including various numerical aberrations and complex structural abnormalities (Kamada N et al. Cancer Res. 1992). However, there has been no large study to examine the relationship between chromosomal abnormalities and survival. Here we report the impact of chromosomal abnormalities on survival in ATL patients with conventional chemotherapy. Patients and Methods: In this study, we extracted a population of patients from the database of ATL-PI project, which is a nation-wide survey of ATL patients newly diagnosed between January 2000 and May 2009, and the overall results were reported elsewhere (Katsuya H et al. JCO 2012, Blood 2015). Fifteen hundred and twelve patients were registered in this project, and we excluded the patients who received allo-HSCT, and who were diagnosed with smoldering and chronic type ATL. Moreover, patients whose chromosomal analysis with G-banding stain showed normal, multiploid or no mitosis, or no information on chromosomal analysis and on the factors to determine the ATL-PI were also excluded. As a consequence, 210 patients with acute (n=157) and lymphoma (n=53) types were selected for this analysis. Abnormal karyotypes were analyzed from the aspect of numerical and structural break points. To identify the specific abnormalities and to avoid the secondary changes, the stem-line karyotype, which meant the simplest abnormal clone, was used as the representative karyotype for this study. A statistical analysis was performed with 'EZR' (Kanda T. BMT 2013). We examined an overall survival (OS) in relation to chromosomal abnormalities using cox proportional hazard regression model, and p-value of less than 0.05 was considered as statistically significant. Results: Among the 210 patients, 119 and 91 patients were male and female, respectively. The median age of patients was 66 years (40-89). With respect to other factors of ATL-PI, the median level of soluble interleukin-2 receptor (sIL-2R), serum albumin, were 30800 U/mL (620-1,219,000), 3.6 g/dL (1.6-5.3), respectively. Almost all patients were diagnosed with stage II-IV of clinical stage (n=200), and almost half of patients certified 2-4 of ECOG-PS (n=113). Sixty-nine, 106, and 35 patients were classified as high, intermediate and low risk by ATL-PI, respectively. Specimens used for chromosomal analysis were taken from lymph nodes (n=77), bone marrow (n=72), peripheral blood (n=50), pleural fluids (n=4), ascites (n=4) and 1 each of stomach, sinusoidal tumor, and tonsillar lesions. The median number of numerical abnormalities and marker chromosomes were 2 (0-20) and 1 (0-20), and the median number of structural break points was 6 (0-21). The numerical abnormalities were loss of sex chromosomes (31%) followed in decreasing order by -14 (24%), -13 (19%), -9 (12%) and -17 (12%) while the structural break points were located at 1p (30%), 3q (30%), 14q (30%), 6q (26%) and 1q (22%). The median OS was 286 days (1-2565). ECOG-PS, serum albumin level, sIL-2R, and structural break points at 9q or 19p, and the number of structural break points over 4 or 5 points were shown to be the significant factors affecting the survival by univariate analysis. Then, the multivariate analysis indicated that 2-4 of ECOG-PS (HR: 2.021, 95%CI: 1.436-2.791, p<0.001), serum albumin level < 3.5 g/dl (HR: 1.631, 95%CI: 1.282-2.074, P<0.001), and chromosomally structural break points at 9q (HR: 1.584, 95%CI: 1.096-2.289, p<0.001) and 19p (HR: 1.765, 95%CI: 1.038-2.999, p<0.001) were factors that negatively contributed to OS. Discussions and Conclusion: This is the first large study showing the impact of chromosomal abnormality in acute and lymphoma types ATL on OS. It demonstrated that structural break points at 9q and 19p were the independent risk factors for OS in addition to those of ATL-PI. Disclosures Katsuya: The Uehara Memorial Foundation: Research Funding. Utsunomiya:Daiichi Sankyo Co., Ltd.: Speakers Bureau. Tsukasaki:Daiichi Sankyo Co., Ltd.: Consultancy; Takeda: Research Funding. Suzumiya:Takeda: Honoraria; Astellas: Research Funding; Kyowa Hakko kirin: Research Funding; Chugai: Honoraria, Research Funding; Toyama Chemical: Research Funding; Eisai: Honoraria, Research Funding.

Nickel (Ni) laterites are regolith materials derived from ultramafic rocks and play an important role in the world's Ni production. Ni-laterite deposits are the supergene enrichment of Ni formed from the intense chemical and mechanical weathering of ultramafic parental rocks. In Vietnam, the weathering profile containing Ni laterite was first discovered in the Ha Tri massive (Cao Bang). This profile develops on the Ha Tri serpentinized peridotite rocks classified to the Cao Bang mafic-ultramafic complex (North Vietnam) and exhibits thick weathered zone (10 - 15m). This work carried out a detailed study of the weathering profile at the center of Ha Tri massive. Samples from different horizons of the profile were collected and analyzed in detail by XRF, XRD and SEM-EDX methods to establish the relationship between the Ni-rich supergene products and the parental peridotites (lherzolite) rocks in Ha Tri massive. The results show that the saprolite horizon is most Ni-rich in the weathering profile in Ha Tri. In this horizon, Ni-silicate minerals of garnierite group such as pimelite, nepouite and other Mg-Ni silicates have been found. The appearance of minerals of garnierite group is due to the exchange of Mg by Ni during weathering of peridotite minerals, especially olivine, which leads to the enrichment of the supergene Ni. The occurrence of Ni silicates suggests the existence of the supergene Ni ore in the weathering profile of the Ha Tri massive.References Bosio N.J., Hurst J.V., Smith R.L., 1975. Nickelliferousnontronite, a 15 Å garnierite, at Niquelandia, Goias Brazil. Clays Clay Miner., 23, 400-403. Brand N.W., Butt C.R.M., Elias M., 1998. Nickel Laterites: Classification and features. AGSO Journal of Australian Geology & Geophysics, 17(4), 81-88. Bricker O.P., Nesbitt H.W. and Gunter W.D., 1973. The stability of talc. American Mineralogist, 58, 64-72. Brindley G.W. and Hang P.T., 1973. The nature of garnierites. Structures, chemical composition and color characteristics. Clay and Clay Minerals, 21, 27-40. Brindley G.W. and Maksimovic Z., 1974. The nature and nomenclature of hydrous nickel-containing silicates. Clay Minerals, 10, 271-277. Brindley G.W. and Wan H.M., 1975. Composition structures and thermal behavior of nickel containing minerals in thelizardite-ne´pouite series. American Mineralogist, 60, 863-871. Brindley G.W., Bish D.L. and Wan H.M., 1979. Compositions, structures and properties of nickel containing minerals in the kerolite-pimelite series. American Mineralogist, 64, 615-625. Cluzel D. and Vigier B., 2008. Syntectonic mobility of supergene nickel ores from New Caledonia (Southwest Pacific). Evidence from faulted regolith and garnierite veins. Resource Geology, 58, 161-170. Colin F., Nahon D., Trescases J.J., Melfi A.J., 1990. Lateritic weathering of pyroxenites at Niquelandia, Goais, Brazil: The supergene behavior ofnickel: Economic Geology, 85, 1010-1023. Das S.K., Sahoo R.K., Muralidhar J., Nayak B.K., 1999. Mineralogy and geochemistry of profilesthrough lateritic nickel deposits at Kansa,Sukinda, Orissa. Joural of Geoogical. SocietyIndia, 53, 649-668. Decarreau A., Colin F., Herbillon A., Manceau A., Nahon D., Paquet H., Trauth-Badaud D.,Trescases J.J., 1987. Domain segregation in NiFe-Mg-Smectites. Clay Minerals, 35, 1-10. Freyssinet P., Butt C.R.M. and Morris R.C., 2005. Oreforming processes related to lateritic weathering. Economic Geology, 100th aniversary volume, 681-722.Garnier J., Quantin C., Martins E.S., Becquer T., 2006. Solid speciation and availability of chromium in ultramafic soils from Niquelandia, Brazil. Journal of Geochemical Exploration, 88, 206-209. Garnier J., Quantin C., Guimarães E., Becquer T., 2008. Can chromite weathering be a source of Cr in soils? Mineralogy Magazine, 72, 49-53. Gleeson S.A., Butt C.R. and Elias M., 2003. Nickel laterites: A review. SEG Newsletter, 54, 11-18. Gleeson S.A., Butt C.R., Wlias M., 2003. Nickellaterites: a review. SEG Newsletter, Society of Economic Geology, 54. Available from www.segweb.org. Golightly J.P., 1981. Nickeliferous laterite deposits. Economic Geology, 75th Anniversary volume, 710-735. Golightly J.P., 2010. Progress in understanding the evolution of nickel laterite. Society of Economic Geology, In Special Publication, 15, 451-485. Manceau A. and Calas G., 1985. Heterogeneous distribution of nickel in hydrous silicates from New Caledonia ore deposits. American Mineralogist, 70, 549-558. Nguyen Van Pho, 2013. Tropic weathering in Vietnam (in Vietnamese). Pubisher Science and Technology, 365p.Ngo Xuan Thanh, Tran Thanh Hai, Nguyen Hoang, Vu Quang Lan, S. Kwon, Tetsumaru Itaya, M. Santosh, 2014. Backarc mafic-ultramafic magmatism in Northeastern Vietnam and its regional tectonic significance. Journal of Asian Earth Sciences, 90, 45-60.Pelletier B., 1983. Localisation du nickel dans les minerais ‘‘garnieritiques’’ de Nouvelle-Caledonie. Sciences Ge´ologique: Me´moires, 73, 173-183.Pelletier B., 1996. Serpentines in nickel silicate ores from New Caledonia. In Grimsey E.J., and Neuss I. (eds): Nickel ’96, Australasian Institute of Miningand Metallurgy, Melbourne, Publication Series 6(9), 197-205. Proenza J.A., Lewis J.F., Galı´ S., Tauler E., Labrador M., Melgarejo J.C., Longo F. and Bloise G., 2008. Garnierite mineralization from Falcondo Ni-laterite deposit (Dominican Republic). Macla, 9, 197-198. Soler J.M., Cama J., Galı´ S., Mele´ndez W., Ramı´rez, A., andEstanga, J., 2008. Composition and dissolution kinetics ofgarnierite from the Loma de Hierro Ni-laterite deposit,Venezuela. Chemical Geology, 249, 191-202. Springer G., 1974. Compositional and structural variations ingarnierites. The Canadian Mineralogist, 12, 381-388. Springer G., 1976. Falcondoite, nickel analogue of sepiolite. The Canadian Mineralogist, 14, 407-409.Svetlitskaya T.V., Tolstykh N.D., Izokh A.E., Phuong Ngo Thi, 2015. PGE geochemical constraints on the origin of the Ni-Cu-PGE sulfide mineralization in the Suoi Cun intrusion, Cao Bang province, Northeastern Vietnam. Miner Petrol, 109, 161-180.Tran Trong Hoa, Izokh A.E., Polyakov G.V., Borisenko A.S., Tran Tuan Anh, Balykin P.A., Ngo Thi Phuong, Rudnev S.N., Vu Van Van, Bui An Nien, 2008. Permo-Triassic magmatism and metallogeny of Northern Vietnam in relation to the Emeishan plume. Russ. Geol. Geophys., 49, 480-491.Trescases J.J., 1975. L'évolution supergene des roches ultrabasiques en zone tropicale: Formation de gisements nikelifères de Nouvelle Caledonie. Editions ORSTOM, Paris, 259p.Tri T.V., Khuc V. (eds), 2011. Geology and Earth Resources of Vietnam. Publishing House for Science and Technology, 645p (in English). Villanova-de-Benavent C., Proenza J.A., GalíS., Tauler E., Lewis J.F. and Longo F., 2011. Talc- and serpentine-like ‘‘garnierites’’ in the Falcondo Ni-laterite deposit, Dominican Republic. ‘Let’s talk ore deposits’, 11th Biennial Meeting SGA 2011, Antofagasta, Chile, 3p.Wells M.A., 2003. Goronickel laterite deposit. New Caledonia. CRC LEME, p.3.

Supplemental Instruction (SI) has been a successful implementation into institutions worldwide. It serves as a means of reducing attrition and increasing the overall learning of course material. The City College of New York (CCNY) has recently implemented SI to General Chemistry I courses to examine whether or not SI would help students succeed in the course and understand and grasp the course material better. SI was made available several times a week during flexible times to students who are struggling in the course. Our method of data collection is a Likert-type and open-ended questionnaire that was distributed at the end of each of the semesters to SI participants in an anonymous fashion. Furthermore, we compared the grades and performance of students participating in SI with those who did not. The collected data enabled us to examine the impact of implementing Supplemental Instruction (SI) in General Chemistry I at CCNY. Our data show that SI was beneficial, contributed to improving students’ understanding of course material, and increased their success rate. About 80 percent of students who failed the first exam and participated in SI obtained a passing grade compared to 11 percent of those who did not.Keywords: supplemental instruction, chemistry education research, student learningINTRODUCTION Supplemental instruction is derived from the term developmental education and stems from a branch of Learning Assistance Centers or LACS [1]. To better understand the origins of Supplemental Instruction, we must first understand the framework that set up this type of education. Developmental education originates in the 1600s and focuses mainly on the assumption that each student has multifaceted talents that can be developed academically, opposed to focusing on the deficits of a student [1]. The main goal was to naturally adapt the students’ population to higher education through means of social and academic domains allowing the students to grow in multiple dimensions in their academic skills [1].Developmental education later paved the way for academic programs such as Learning Assistance Centers (LACs). It was viewed as an extension of the classroom and did not discriminate between faculty and students, and within the student population did not discriminate between students who performed to standard and those who performed below par. LACs were for everyone who wanted to be academically enriched [1]. As a new program, LACs had six main objectives: “higher course grades for participating students; central location for students to receive tutorial assistance; a referral source to other helping agencies; a comprehensive library of basic study aids; a training agency for paraprofessionals, peer counselors, and tutors; and a center for faculty development.” [1]. These goals made it very clear that this service was not to be mislabeled as remedial, which was a main topic of interest when it comes to implementing new services, like this one, in schools. LACs are essentially a tool used to attain higher education, not provide a remedial course. Another important reason as to why LACs were adopted into other higher education institutions is to increase student retention [1]. LACs jump-started improved learning across campuses not only in the United States but across the world.In the 1990’s developmental or remedial courses were not allowed to be offered at public four-year universities, thus making supplemental instruction a very attractive candidate to solving the issue with students’ academic integrity and attrition [1]. College faculty members were drawn to SI due to its minute fiscal commitment and that it strongly promoted individual self-learning for the students, this meant less time spent teaching for instructors [1].SI or supplemental instruction is as it states - supplemental instruction offered to the students enrolled in a course. SI gives the students the opportunity to work in a cooperative setting on problems and is supported by peer instructors [2]. SI works to increase academic performance and retention but utilizing both collaborative studying in addition to independent studying and analysis of study strategies. SI sessions are offered for traditionally difficult high risk course; ‘historically difficult’ courses are also identified by analyzing the grade distribution of courses throughout each academic division, “courses in which 30% or more of the students receive D or F grades or withdraw, become targets” [3]. One of the main factors that attribute to the individuality of SI programs is that it focuses on historically difficult courses [4] and large classes where students have little opportunity for interaction with the professor or other students [5]. Labeling a course as historically difficult allows you to categorize the class as challenging without placing blame on the professor or the students.SI sessions are scheduled on a weekly basis and all students enrolled in the course are encouraged to join. SI sessions act as a continuation of the lesson learned during in-class lecture with the instructor. Being that SI sessions are not tied down to class time, students can attend whenever and however many times they wish without the restriction of a required course preventing them. Constant feedback will be given based on the student’s grasp of the material taught in class, thus giving them ample time and motivation to alter their study methods to adapt, if need be. Research on SI suggests that problem-solving skills and study strategies learned in SI courses are transferable to other courses which leads to improved performance [6]. Supplemental instruction sessions are typically led by an SI leader, who would be a former student that succeeded in the same course, however it is not uncommon for a professor to join an SI session for a course. SI may improve students’ retention and graduation because it offers the students the opportunity to network socially which speeds up the acclimatization to the college experience [7].SI provides a way to combine “how do I teach myself this” with “what do I need to know”. While SI helps students gain a more thorough understanding of the material, it also helps students to condition themselves and alters their thinking to accommodate for the type of information they are processing. SI differs from a remedial tutoring course because it is created based on the course versus the students [1]. This potentially makes attending SI sessions more attractive to students who are afraid or shy to ask for help.SI is not a remedial course, it is considered a mediator connecting the gap between the new information presented to students and the students current knowledge and practices, “SI bridges the gap between the current knowledge base and the acquisition of new knowledge by focusing on the refinement of the learning skills indispensable for acquiring new knowledge” [8]. Successfully implemented SI programs were able to track those students who attended and the impact it made towards their grade, for data that was collected institutionally [9]. SI can be effectively used to develop study skills, increase motivation and improve performance in participants [10].According to Wolfe, “…there appear to be benefits of SI in courses where students are being newly introduced to chemical concepts and methods, but these benefits seem to drop off when students are more experienced with the material.” [5]. The results further go on to conclude that SI is most beneficial to entry level science courses such as, General Chemistry I and Organic Chemistry I. It seems as though SI is a great addition into institutions to overall help attrition rates and more generally help students improve within their courses. More research has been done to give us a wider range of knowledge on SI within Chemistry courses [9]. SI can be used to address the continuous decline of fundamental chemical knowledge noticed in recent college graduates [11].In large lecture classes, there is a disconnect between the lecturer and the student. Supplemental Instruction (SI) is often offered in a more intimate setting where the SI instructors can have one-on-one time with students to individually hone in on their specific problems, whether it is conceptual or problem-solving. As for learning environment, SI is usually held in a smaller more accommodating room where it is a less intimidating more inviting space. This setting provides a social interaction component in which usual Chemistry lectures don’t provide. In addition to the more personal tutoring time available, students are also encouraged to interact in a common social setting. This allows for an opportunity of collaborative learning and may help foster a sense of community [12].The attraction to SI in a college setting is due to its short and long-term effects. The short-term effects of SI sessions are to instill a deeper understanding of the coursework within students. Whereas the long-term goals are two-fold, the academic goal is to change the way students learn, study, and understand information so that the knowledge stays with them. Students who participate in SI earn higher grades not only the course but in subsequent semesters, which is due to improved study habits and is considered a long-term positive outcome of SI [13]. The second long-term goal is to develop better social interaction skills within these students.Many public institutions have implemented SI and thus have uncovered several significant findings. Findings that were previously mentioned pertaining to the outcomes of SI includes achieving higher percentages of passing grades [1, 14]. Some studies also found higher graduation rates among participants [1]. Despite their better performance, SI takers often score lower on academic success predictors such as the SAT and ACT exams [14]; increases in performance, therefore, appear to not occur because stronger students use SI. SI helps promote problem-solving skills because it involves the learner in the construction of knowledge and information processing based on their prior experiences which could contribute to cognitive development [15-16]. Furthermore, SI has been shown to improve motivation and enhance study habits and understanding difficult concepts [17]. Participants in SI have been found to have better control on their achievement, self-esteem, and confidence [18]. SI has also been found to higher academic self-efficacy and grades [19]. Studies have reported that students who participated in SI in introductory courses in Biology or Chemistry courses tended to continue onto upper level courses in the field; SI has a positive effect on students’ enrollment in upper level courses [20-21]. We should note that Rabitoy and co-authors reported that SI enhanced students’ achievement in STEM fields and this was greater for females and students of color [22]. METHODS While researching the topic of SI as a whole, we were able to uncover many studies that were in favor of this new implementation in institutions. Our research study is unique because it targets only at risk students and provides an added value to the curriculum. SI was implemented into The City College of New York (CCNY) as a test run in the Department of Chemistry and Biochemistry. The CCNY is an urban minority serving institute that offers a diverse selection of classes, which vary from the arts to the sciences and everything in between. We wanted to try implementing SI into the sciences to see if at risk science students can benefit from it. Grasping the S in stem, our main goal is to use the successful methods of SI to see what benefits it can bring to general science courses.Beginning during the Spring semester of 2018, SI was implemented at The CCNY’s General Chemistry I course which is composed of lecture, lab, and Peer-Led Team Learning (PLTL) workshop. PLTL is implemented as an integrated part of the course [23]. In the General Chemistry course, after exam one was distributed and grades were given back, an announcement was made introducing SI. The lecture professor would encourage students who received a failing grade, classified as a grade below a 65 out of 100 on exam one, to attend SI as means of improving in the course. This makes our study unique since we are only targeting students who are struggling the course and are not likely to earn a passing grade. It is noteworthy that PLTL offers a collaborative learning experience for all students. SI instructors targeted weaknesses in study habits, socialization, conceptual-understanding, and problem-solving skills.SI was made available for five days a week and on those days, SI was scheduled around classes times which makes it easier for students with busy schedules involving work and class to attend. During SI sessions there are instructors that have been hand-picked by instructors from a cohort of recitation leaders and who have successfully completed the course and have received a grade of A in the course. SI attendance was not mandatory.Throughout the first session available after the Supplemental Instruction announcement was made to the lecture class, those who chose to attend, because SI was not mandatory and it was strongly recommended for struggling students. Students brought their first exam with them to be discussed. This initial discussion is a one-on-one between the student and instructor, which serves as a more social component to learning whereas that isn’t available during class time. The first exam was then discussed so that the instructor can now sift through the student’s strengths and weaknesses to later target them in the weeks to come. The overall composition of our Supplementary Instruction sessions was influenced by the previous studies done and researched, hand-picking methods that seemed to work and leaving off ones that were less successful. SI sessions included cooperative learning activities, and addressed students’ misconceptions and difficulties, explored difficult concepts in depth, and enhanced problem-solving skills [24].Every week during Supplementary Instruction sessions students were encouraged to come back to further work through their weaknesses. During this trial specifically, it was noticed by the SI instructors that a lot of students actually struggle with basic problem-solving skills and reasoning. To help combat this, worksheets were made each week based on the lecture topics covered in class, based on problem-solving to help build their skills. Aside from doing the worksheets, students were encouraged to bring in topics or questions they have on the material so the SI instructors can further clarify and explain. Furthermore, students came to SI with questions that they were struggling with. Students provided questions for the SI instructor to explain which created a mutually beneficial relationship [11, 25].Our research question is: How does Supplemental Instruction in General Chemistry I impact students’ learning, conceptual understanding, retention, and attitudes?In order to properly assess how beneficial SI is to the way students learn Chemistry, data were collected at the end of the Spring and Fall of 2018 semesters, as well as Spring 2019. We used a combination Likert-type and open-ended questionnaire, as well as, grade comparisons. The Likert-type section included a five-point scale, where (1) Strongly Disagree, (2) Disagree, (3) Neutral, (4) Agree, (5) Strongly Agree. For each question, the average was taken which helps make sense of the data. For the open ended questions, questions 1 through 3, we created and used a rubric to score the questions on a scale from 1 to 5 similar to what was mentioned above. For question 4, we compiled the answers and created pie charts based on the type and number of responses.As previously stated, at the end of the semester surveys were printed and distributed to each SI session. Students that attended SI were all encouraged to participate in this survey. A total of 44 out of 60 students were available and willing to participate in the optional survey.RESULT AND DISCUSSION Graphical depictions of each Likert-type question along with average responses are shown in Figure 1. FIGURE 1. Averages for the answers for each of the Likert-type questionnaire. Our data clearly shows that students overall had a positive learning experience with SI. The students agree that SI improved their basic problem-solving skills, understanding of concepts covered, and attaining a better grade in the course. Furthermore, SI participation encouraged students to practice problems and tackle their weaknesses in the course. SI participants believe that the SI instructors targeted their weak areas and helped them improve. Finally, students felt that SI was worth the time and effort they put into it and that it was beneficial and contributed to improvements in grades.FIGURE 2. Averages of the short-answer responses were numerically scaled using a rubric.Figure 2 shows that students overall had a positive learning experience with SI and that it helped them better understand the concepts. Furthermore, the participants would enthusiastically would recommend SI to other Chemistry students.FIGURE 3. The pie chart above shows the beneficial components of SI to studentsFigure 3 shows the parts of SI that students found beneficial. Overall, there were many useful features of the SI program that facilitated students’ learning and succeeding in the course. The students appreciated the time spent on problem-solving, the flexibility of the offerings – refers to the times and days, the individualized learning experience, and the one-on-one support, guidance, and tutoring. From the Likert-type questions given in part one of the survey, responses were broken down for each given question in order to visualize the impact of SI for the students/participants. The average of the responses can be seen above and was about 4.8. The corresponding results can be viewed above as shown in Figure 1. To turn our attention to Figure 1 ‘SI has helped me to improve my basic problem solving skills’, upon meeting with the SI instructor team at The CCNY we were made aware that students have trouble with basic problem-solving which leads to the disconnect in understanding how to build upon this knowledge. When students lack this basic skill, it is difficult for them to move on and build upon this skill as the course progresses, which ultimately leads to their failure to meet the level of understanding for each lecture exam. From the responses, we can see that majority of the students feel as though their problem solving skills have developed as a result of participating in SI. Questions 6-8 deal with an overall belief about the experience of SI participation. Based on Figure 1, SI helped students better understand the course material, and had an overall positive response as 100% of students felt like they did have a better understanding of the course material after attending SI. Students overwhelmingly agree that SI is very beneficial to the way they learn Chemistry. Our data show that SI is well received by the participants which is consistent with other researchers [11]. Short answer questions 1-3 were made in order to have a more in-depth understanding of the student experience with SI this past semester. These questions were formed as short answer so students were more inclined to share their opinions of SI. The collected data shows that the students strongly agree that SI has been very beneficial for ways unique to each student. Some students enjoyed the SI instructors and their way of teaching and helping the students understand the material. While others enjoyed how personal the experience was. It wasn’t a ‘one size fits all’ experience but an experience that was personalized to each student’s needs, strengths, and weaknesses. Short answers for question 4 also had very personalized responses. Question 4 in particular – ‘What features of SI did you find to be most beneficial to you?’ was very helpful in understanding what students appreciate most about SI, and served as a standard to which should be upheld. Responses ranged from the flexible times SI was offered to the small class setting which is extremely different than the lecture, and more comfortable. Students also enjoyed and benefited from the one-on-one attention they received in SI session. For that reason, students would make time to attend the SI section of their ‘favorite instructor’. Furthermore, the individualized learning experience was appealing to SI participants. The overall goal is to get each student to understand complex concepts and if one instructor can better help someone to reach this goal; SI is a success. The overall responses show that SI is very beneficial and contributed a great deal to the way that students learn Chemistry. One main issue we have encountered during this initial phase of implementation is low number of students who self-select to participate in SI. Only about 10 percent of students who failed the first exam, participated in SI. One reason could be that students might not buy into the philosophy of SI. A second reason might be that students do not think that SI would help them better understand the content and succeed on the course. A third reason could be due to lack of motivation in these students. Another reason could be attributed to the lack of maturity in students who are taking General Chemistry I. We use the term ‘lack of maturity’ loosely in this sense to mean the lack of knowledge of how to conduct oneself in a college setting [26]. Most commonly, freshman students come straight from high school where they have not yet acquired good study techniques or methods which work best for them. These reasons combined with the hardship required to understand such complex concepts that they’re learning for the very first time can deter students. Our data show that students who participated in SI after failing the first exam achieved a success rate of about 80%. Students who failed the first exam and did not participate in SI had an 11% chance of passing the course. This data is consistent with several research in the field indicating that students who participate in SI have a higher chance of successfully completing the course with a passing grade [14, 27]. We feel these data make powerful statement for the added value of SI in General Chemistry courses and its impact on students learning and attitudes.CONCLUSION In conclusion, our research study on SI provided valuable data into students’ learning and conceptual understanding of content in General Chemistry. Furthermore, our findings provide insights into students’ attitudes about SI implementation and its benefits to the participants. Our research data supports the introduction of SI into General Chemistry courses and provides students with learning skills, socialization competencies, problem-solving skills, and the knowledge required to successfully complete the course. We believe that our data supports the notion that SI had a significantly affected the participants grades in a positive manner and it should be noted that that majority of SI participants earned a passing grade in the course. Our results and data on the implementation of SI in General Chemistry courses improved students’ attitudes towards the subject matter, learning, socialization, and study habits. We are confident we can reach a larger population and improve our recruitment so more students can experience SI and increase their chances of successful completion of the course. As an implication for curriculum, we would recommend supplemental instruction for all introductory courses STEM fields. ACKNOWLEDGMENT We also would like to thank the Department of Psychology at the CCNY and CUNY Coordinated Undergraduate Education (CUE) for funding our Supplemental Instruction program.

Introduction Methods Resuts Discussion Conclusions Acknowledgments Authors' contributions Competing interests Ethical approval References Effect of Methotrexate and Omega-3 Combination on Cytogenetic Changes of Bone Marrow and Some Enzymatic Antioxidants: An Experimental Study Inaam N. Ali1, Muthana M. Awad2, Alaa S. Mahmood2,* 1 Water and Environment Directorate, Ministry of Sciences and Technology, Baghdad, Iraq 2 Department of Biology, College of Science, University of Anbar, Anbar, Iraq * Corresponding author: A. S. Mahmood (alaashm91@gmail.com) Abstract: Objective: To assess the effect of methotrexate and omega-3 combination on cytogenetic changes of bone marrow and activities of some enzymatic antioxidants. Methods: Fifty-six mature male Wistar rats were divided into two experimental groups and a control group. The first experimental group was sub-divided into three sub-groups depending on the concentration of methotrexate (MTX): X1 (0.05 mg/kg MTX), X2 (0.125 mg/kg MTX) and X3 (0.250 mg/kg MTX), which were given intraperitoneally on a weekly basis for eight weeks. The second experimental group (MTX and omega-3 group) was also sub-divided into three sub-groups (Y1, Y2 and Y3), which were injected intraperitoneally with 0.05, 0.125 and 0.25 mg/kg MTX, respectively, weekly for eight weeks accompanied by the oral administration of 300 mg/kg omega-3. The rats of the control group were given distilled water. The enzymatic activity of catalase (CAT), superoxide dismutase (SOD) and glutathione reductase (GR) were measured in the sera of rats. In addition, the mitotic index (MI) and chromosomal aberrations of bone marrow were also studied. Results: MTX resulted in a significant decrease in the activities of CAT, SOD and GR compared to the controls. It also increased the MI and chromosomal aberrations of rat bone marrows. On the other hand, omega-3 significantly increased the activities of the investigated enzymatic antioxidants and reduced the MI and chromosomal aberrations in treated mice when given in combination with MTX. Conclusions: MTX has a genotoxic effect on the bone marrow by increasing the MI and all types of chromosomal aberrations and decreasing the enzymatic activity of CAT, SOD and GR. The addition of omega-3 can lead to a protective effect by reducing the toxic and mutagenic effects of MTX. Keywords: Methotrexate, Omega-3, Antioxidant, Wistar rat, Chromosomal aberration, Mitotic index 1. Introduction Methotrexate (MTX) is a folic acid antagonist because of their chemical similarity [1]. Vezmar et al. [2] showed that MTX affects the synthesis of nucleic acids deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) by interfering with the biosynthesis of thymine and purines. It also directly affects the rapidly dividing and intact cells, especially those in the mucous membranes of the mouth, intestine and bone marrow [3]. Omega-3 is a type of unsaturated fats, which are classified as essential fatty acids that cannot be manufactured by the body and should be taken with food [4]. Sources of omega-3 include fish oils, such as salmon, sardines and tuna, as well as soybeans, walnuts, raisins and linseed, almonds and olive oils [5]. Omega-3 is used in the prevention of a number of diseases such as rheumatoid arthritis, ulcerative colitis, asthma, atherosclerosis, cancer, and cardiovascular diseases [6]. A large amount of evidence indicates that omega-3 fatty acids have significant health benefits, including anti-inflammatory and antioxidant properties besides their effect on blood cholesterol levels [7]. Antioxidants retard the oxidation process by different mechanisms such as the removal of free radicals [8]. Enzymatic antioxidants include catalase (CAT), which is the first line of defense in the cell that removes hydrogen peroxide formed during biological processes by converting it into an aldehyde, and superoxide dismutase (SOD). There are three major families of SOD enzymes: manganese SOD (Mn-SOD) in the mitochondria and peroxisomes, iron SOD (Fe-SOD) in prokaryote cells and copper/zinc SOD (Cu-Zn SOD) in the cytoplasm of eukaryote cells [9]. Therefore, changes in the metal co-factors (manganese, iron, copper and zinc) can alter the effectiveness of SOD and may lead to diseases as a result of oxidative stress [10]. Glutathione reductase (GR) is also an enzymatic antioxidant that converts the oxidized glutathione to the reduced glutathione in the presence of NADPH, which is oxidized to NADP [11]. Therefore, the aim of the present study was to assess the effects of MTX and omega-3 on the cytogenetic changes of bone marrow as well as the activities of CAT, SOD and GR enzymatic antioxidants in male rats. 2. Method 2.1. Laboratory animals and experimental design Fifty-six mature male Wistar rats (Rattus norvegicus), aged 10–12 weeks old and weighing 250–300 gm, were used in the present study. The rats were kept in separate cages, with natural 13- hour light and 11-hour dark periods in a contamination-free environment with a controlled temperature (28.0 ± 1.0°C). In addition, rats were maintained on a standard diet and tap water ad libitum. The rats were randomly allocated to two experimental groups and a control group. The first experimental group (MTX group) included 24 rats injected intraperitoneally with different MTX dilutions with distilled water [12]. It was sub-divided into three sub-groups (eight rats per sub-group) according to MTX concentration as follows: X1 (0.05 mg/kg MTX), X2 (0.125mg/kg MTX) and X3 (0.25 mg/kg MTX). All rats were given a single dose of the specified MTX concentration weekly for eight weeks. The second experimental group (MTX and omega-3 group) included 24 rats allocated to three sub-groups (Y1, Y2 and Y3), which were injected intraperitoneally with 0.05, 0.125 and 0.25 mg/kg MTX, respectively, weekly for eight weeks accompanied by the oral administration of 300 mg/kg omega-3. The control group included eight rats that were intraperitoneally injected with distilled water and given a single dose of distilled water orally weekly for eight weeks. 2.2. Blood collection and processing After the end of the dosing period, 5 ml of blood were withdrawn from the heart (by cardiac puncture) using a 5 cc disposable syringe. The collected blood was immediately poured into a clean sterile screw-capped tube (plain tube) and left for coagulation in a water bath at 37°C for 15 minutes. After coagulation of blood, the plain tube was centrifuged for 5 minutes at 1500 rpm. Then the samples were stored at -20°C for subsequent analysis. 2.3. Measurement of the activity of antioxidant enzymes The antioxidant activities of CAT, SOD and GR were measured using enzyme-linked immunosorbent assay kits purchased from Kamiya Biomedical Company (Seattle, WA, US), according to the manufacturer's instructions. 2.4. Cytogenetic study of bone marrow Rats were killed by cervical dislocation, and their hip bones were cleaned from surrounding muscles and then dissected by cutting both ends of the bone. Five milliliters of physiological buffered saline were injected inside the bone to withdraw bone marrow into a test tube. Tubes were centrifuged at 2000 rpm/10 minutes. The supernatant was then removed, and 10 ml of KCL solution (0.075 M) were added to the sediment. The mixture was then incubated at 37 °C in a water bath for 30 minutes, with shaking from time to time. The tubes were then centrifuged at 2000rpm/10 minutes to remove the supernatant. However, 5 ml of a freshly prepared fixative solution (methanol: glacial acetic acid 1:3) were added gradually in the form of droplets into the inner wall of the tube with constant mixing. After that, the tubes were placed at 4 °C for half an hour to fix the cells. This process was repeated for three times, and the cells were then suspended in 2 ml of the fixative solution. The tubes were centrifuged at 2000 rpm for 5 minutes, and the supernatant was then removed while the cells were re-suspended in 1-2 ml of cold fixative solution. After shaking the tubes, 4–5 drops were then taken from each tube onto a clean slide from a height of about three feet to provide an opportunity for the cells and nuclei to spread well. The slides were stained with acridine orange solution (0.01%) for 4–5 minutes, incubated in Sorensen’s buffer (0.06M, pH 6.5) for a minute. and then examined using a fluorescence microscope Olympus BX 51 America at a wavelength of 450–500 nm [13, 14]. A total of 1000 cells were examined, and both dividing and non-dividing cells were calculated [13]. Mitotic index (MI) was calculated according to the following formula [13]: MI= No. of dividing cells / 1000 × 100 2.5. Analysis of chromosomal aberrations of bone marrow cells A total of 1000 dividing cells were examined on the stained slides under a fluorescence microscope at a wavelength of 45–500 nm. The examined cells were at the first metaphase of the mitotic division, where chromosomal aberrations are clear and can be easily seen [13]. 2.6. Statistical analysis Data were analyzed using the Statistical Analysis System (SAS®) software, version 9.1 (Cary, NC, USA) [15]. Effects were expressed as mean ± standard error (SE) and statistically compared using a completely randomized design analysis of variance and least significant differences. Differences at P values <5 were considered statistically significant. 3. Results 3.1. Effects of MTX and MTX-omega-3 combination on antioxidant enzymatic activities Table (1) shows significantly lower SOD activities among rats treated with MTX or MTX-omega-3 compared to controls. Moreover, sera of rats receiving relatively high doses of MTX (sub-groups X2 and X3) showed the lowest enzymatic activities of 4.29 ± 0.01 IU and 3.93 ± 0.11 IU, respectively. On the other hand, CAT activity differed significantly between treated and control rats as well as among treated rats themselves, In this respect, the controls showed the highest activity of 39.38 ±0.02 IU, while those receiving the highest MTX concentration, either alone or in combination with omega-3 (sub-groups X3 and Y3), showed the lowest activities of 30.97 ± 0.03 IU and 32.12± 0.06 IU, respectively. Regarding GR activity, control rats showed a higher activity of 53.09± 0.05 IU compared to treated ones; however, the differences in GR activities in rats given low doses of MTX, either alone or in combination with omega-3 (sub-groups X1 and Y1), were not statistically significant. On the other hand, rats in sub-groups X3 and Y3 showed the lowest GR activities of 34.59 ± 0.63 IU and 37.15 ±0.01, respectively, with statistically significant differences from other sub-groups. 3.2. Effects of MTX and MTX-omega-3 combination on mitotic index of bone marrow cells Figure (1) shows a significant decrease in the MI in all treated groups compared to control. In addition, there was a reverse association between MTX concentration and MI, where rats treated with the highest dose of MTX (sub-group X3) showed a significant decrease in MI compared to all other treated rat sub-groups. In addition, rats in sub-groups treated with MTX and omega-3 (sub-groups Y1, Y2 and Y3) showed a significant increase in MI compared to their counterpart rats receiving MTX only. Table 1. Activity of antioxidant enzymes in rats treated with MTX and MTX-omega-3 Group Enzymatic activity (mean± SE) SOD (IU) CAT (IU) GR (µmol) Control 6.41±0.02 a 39.38±0.02 a 53.09±0.05 a X1 (0.05 mg MTX/ kg) 5.33±0.01 b 37.81±0.01 c 51.12±0.06 a Y1 (0.05 mg MTX + 300 mg omega-3/ kg) 6.08±0.04 a 38.40±0.02 b 51.97±0.03 a X2 (0.125 mg MTX/ kg) 4.29±0.01 cd 33.13±0.01 e 42.34±0.03 b Y2 (0.125 mg MTX + 300 mg omega-3/ kg) 4.99±0.40 b 36.68±0.02 d 43.02±3.04 b X3 (0.25 mg MTX/ kg) 3.93±0.11 d 30.97±0.03 g 34.59±0.63 c Y3 (0.25 mg MTX + 300 mg omega-3/ kg) 4.47±0.02 c 32.12±0.06 f 37.15±0.01 c SE, Standard error; IU, international unit; SOD, superoxide dismutase; CAT, catalase; GR, glutathione reductase; *statistically significant at P < 0.05; **statistically significant at P < 0.01. Means with different letters within the same column showed a statistically significant difference. 3.3. Effects of MTX and MTX-omega-3 combination on chromosomal aberrations of bone marrow cells Rats receiving higher concentrations of MTX (sub-group X3) showed a significant increase in all types of chromosomal aberrations, i.e., chromatid gaps, chromosome gaps, chromatid breaks, chromosome breaks, deletions and simple fragments (Figure 2 and Table 2) than those of the control group or other treated sub-groups. All rats treated with MTX-omega-3 combination showed a significant decrease in almost all types of chromosomal aberrations compared to their counterpart rats receiving MTX alone (Table 2). Figure 1. Effect of MTX and MTX-omega-3 on the MI of bone marrow cells of treated rats compared to the controls. The groups X1 (0.05 MTX), X2 (0.125 MTX) and X3 (0.250 MTX) were compared to the control group, while the groups Y1 (0.05 MTX+ omega-3), Y2 (0.125 MTX+ omega-3) and Y3 (0.25 MTX+ omega-3) were compared to X1, X2 and X3, respectively. Figure 2. Effect of MTX and MTX-omega-3 on chromosomal aberration as seen under fluorescence microscope after staining with acridine orange: (1) a simple fragment; (2) a chromatid gap; (3) a chromosomal gap (A) and a chromosomal break (B). 4. Discussion The present experiment reveals that the addition of omega-3 to MTX alleviates its effects on the activities of the antioxidant enzymes CAT, SOD and GR, and decreases the MI as well as all types of chromosomal aberrations in the bone marrow cells. Daham et al. [16] showed that the decline in antioxidants associated with chemotherapy is attributed to the increase in lipid peroxidation caused by these kinds of drugs, which increase the level of free radicals. In addition, Weijl et al. [17] showed that some chemotherapeutic drugs have a negative effect on the antioxidant levels such as GR, whose activity decreases as a result of its involvement in many cellular processes such as cell defenses against the toxicity of some compounds. Al-Dalawy et al. [18] found that the decrease in the level of SOD is an evidence of its increased activity due to the increased release of free radicals. MTX causes an increase in the release of free radicals, including the OH radical that causes direct damage to DNA [16]. Al-Helaly [19] showed that the amount of food taken has an effect on antioxidants, where nutritional deficiency decreases the antioxidant levels, thus increasing free radicals that cause damage to DNA. Table 2. Chromosomal aberrations of bone marrow cells in rats treated with MTX and MTX-omega-3 Group Type of chromosomal aberration(mean ± SE) Chromatid gap Chromosome Gap Chromatid breaks Chromosome breaks Deletion Simple Fragments Chromosomal aberration (%) Control 1.33±0.33 e 0.00±0.00 e 1.67±0.33 c 0.33±0.15 c 0.00±0.00 0.67±0.33 cd 0.04±0.005 f X1 2.75±0.47 cd 1.50±0.28 cd 2.50±0.64 bc 1.00±0.41 bc 0.50±0.28 bc 0.75±0.25 bcd 0.09±0.02 de Y1 1.75±0.47 de 0.75±0.25 de 1.50±0.28 c 1.00±0.00 bc 0.75±0.25 abc 0.75±0.25 abc 0.065±0.005 ef X2 4.67±0.33 b 2.67±0.33 ab 2.67±0.33 bc 1.67±0.33 ab 0.67±0.33 abc 1.67±0.33 ab 0.14±0.006 bc Y2 3.00±0.00 c 2.00±0.00 bc 3.00±0.057 bc 1.33±0.33 b 0.67±0.33 abc 0.33±0.15 d 0.106±0.003 cd X3 6.80±0.37 a 3.00±0.31 a 4.60±0.74 a 2.40±0.24 a 1.40±0.24 a 1.80±0.37 a 0.20±0.017 a Y3 5.60±0.40 ab 2.40±0.24 ab 3.60±0.24 ab 1.80±0.20 ab 1.20±0.20 ab 1.40±0.24 abc 0.16±0.003 b LSD 1.231** 0.814** 0.602** 0.841** 0.774* 0.941** 3.499* SE, Standard error; * statistically significant at P < 0.05; ** statistically significant at P < 0.01. Means with different letters within the same column showed a statistically significant difference. X1 (0.05 mg MTX/ kg); X2 (0.125 mg MTX/ kg); X3 (0.25 mg MTX/ kg); Y1 (0.05 mg MTX + 300 mg omega-3/ kg); Y2 (0.125 mg MTX + 300 mg omega-3/ kg); Y3 (0.25 mg MTX + 300 mg omega-3/ kg). In the present study, the intraperitoneal administration of MTX to rats also caused a decrease in the MI of bone marrow and a significant increase in the rate of abnormal chromosomal aberration compared to the control rats. This finding is consistent with those reported previously [20], [21]. The effect of MTX can be attributed to its ability to interfere with the genetic material, leading to the appearance of toxic and mutagenic consequences. Rushworth et al. [22] reported that MTX leads to a lack of dihydrofolate reductase, which is the key to the growth and cell division processes. This, in turn, leads to a reduction of the nucleotides involved in the building of DNA and, therefore, to a stop or obstruction of the repair mechanisms of the damaged DNA. In addition, Wong and Choi [23] concluded that MTX inhibits the action of enzymes controlling the purine metabolism, which leads to the accumulation of adenosine in addition to the damage of the molecule itself and to the occurrence of chromosomal aberrations. Jafer et al. [24] reported the ability of MTX to induce chromosomal aberration in humans or animals by preventing the repair of DNA and affecting the proteins found in chromosomes. These findings were also confirmed by Hussain et al. [25], who found that MTX causes an increase in chromosomal aberrations. In the present study, the MI showed a significant increase in rat sub-groups treated with MTX-omega-3 combination, but there was a decrease in the rate of chromosomal aberration, which confirms the role of omega-3 unsaturated fatty acids in protecting the cell from the impact of free radicals [26], [27]. Attia and Nasr [28] reported the antioxidant effect of omega-3, which was attributed to the reduction in lipid peroxidation and the increase in SOD and CAT or the stimulation of GR. It is noteworthy that GR leads to the synthesis of reduced glutathione, which is important in the defense of the cell against toxic substances and the prevention of the occurrence of mutations [29]. 5. Conclusions MTX significantly decreases the activity of enzymatic antioxidants, reduce the MI and increase the chromosomal aberrations of all types in bone marrow. This gives further evidence on the genotoxic effects of MTX on the bone marrow. On the other hand, omega-3 shows a protective effect by reducing the toxic and mutagenic effects of MTX. Acknowledgments The authors thank the staff of the Water and Environment Directorate, Ministry of Science and Technology, Baghdad, Iraq for their cooperation. They also thank Dr. Jasim Al-Niami for his technical and scientific guidance. Authors' contributions INA, MMA and ASM contributed to the study design and analyzed data. All authors contributed to the manuscript drafting and revising and approved the final submission. Competing interests The authors declare that they have no competing interests associated with this article. Ethical approval The ethical clearance of this study was obtained from the Ethics Committee of the College of Science, University of Anbar (Reference No. A. D. 51 in 30/8/2015). References Yuen CW, Winter ME. Methotrexate (MTX). In: Basic clinical pharmacokinetics, Winter ME, editor. Philadelphia, USA: Lippincott Williams & Wilkins; 2010. p.p. 304–25. Google Scholar Vezmar S, Becker A, Bode U, Jaehde U. Biochemical and clinical aspects of methotrexate neurotoxicity. Chemotherapy 2003; 49: 92–104. DOI PubMed - Google Scholar Tian H, Cronstein BN. Understanding the mechanisms of action of methotrexate implications for the treatment of rheumatoid arthritis. Bull NYU Hosp Jt Dis 2007; 65: 168–73. PubMed - Google Scholar El-Khayat Z, Rasheed WI, Elias T, Hussein J, Oraby F, Badawi M, et al. Protective effect of either dietary or pharmaceutical n-3 fatty acids on bone loss in ovariectomized rats. Maced J Med Sci 2010; 3: 9–16. DOI - Google Scholar Kris-Etherton PM, Harris WS, Appel LJ; Nutrition Committee. Fish consumption, fish oil, omega-3 fatty acids and cardiovascular disease. Arterioscler Thromb Vasc Biol 2003; 23: e20–30. DOI - PubMed - Google Scholar Calder PC. Polyunsaturated fatty acids and inflammation. Prostaglandins Leukot Essent Fatty Acids 2006; 75: 197–202. DOI - PubMed - Google Scholar Begin ME, Ells G, Das UN, Horrobin DF. Differential killing of human carcinoma cells supplemented with n-3 and n-6 polyunsaturated fatty acids. J Natl Cancer Inst 1986; 77: 1053–62. DOI - PubMed - Google Scholar Shan B, Cai YZ, Sun M, Corke H. Antioxidant capacity of 26 spice extracts and characterization of their phenolic constituents. J Agric Food Chem. 2005; 53: 7749–59. DOI - PubMed - Google Scholar Matiax J, Quiles JL, Huertas JR, Battino M. Tissue specific interactions of exercise, dietary fatty acids, and vitamin E in lipid peroxidation. Free Radic Biol Med 1998; 24 : 511–21. DOI - PubMed - Google Scholar Dean RT, Fu S, Stocker R, Davies MJ. Biochemistry and pathology of radical-mediated protein oxidation. Biochem J 1997; 324: 1–8. PubMed - Google Scholar Bashir A, Perham RN, Scrutton NS, Berry A. Altering Kinetic mechanism and enzyme stability by mutagenesis of the dimmer interface of glutathione reductase. Biochem J 1995; 312: 527–33. PubMed - Google Scholar Perret-Gentil MI. Rat Biomethodology. Laboratory Animal Resources Center. The University of Texas at San Antonio. [Cited 1 Feb. 2015]. Available from: https://www.utdallas.edu/research/docs/rat_biomethodology/ Allen JW, Shuler CF, Menders RW, Olatt SA. A simplified technique for in vivo analysis of sister chromatid exchange using 50 bromodeoxyuridine tablets. Cytogenet Cell Genet 1977; 18: 231–7. DOI PubMed - Google Scholar Forsum U, Hallén A. Acridine orange staining of urethral and cervical smears for the diagnosis of gonorrhea. Acta Derm Venereol 1979; 59: 281–2. PubMed - Google Scholar Statistical Analysis System user's guide. Version 9.1. Cary, NC, USA: SAS Institute Inc.; 2012. Daham HH, Rahim SM, Al-Hmesh MJ. The effect of radiotherapy and chemotherapy in several physiological and biochemical parameters in cancer patients. Tikrit J Pure Sci 2012; 17: 83–91. Weijl N, Elseendoorm TJ, Lentjes EG, Hopman CD, Wipkink-Bakker A, Zwinderman AH, et al. Supplementation with antioxidant micronutrients and chemotherapy-induced toxicity in cancer patients treated with cisplatin-based chemotherapy: a randomised, double-blind placebo-controlled study. Eur J Cancer 2004; 40: 1713–23. DOI - PubMed - Google Scholar Al-Dalawy SS, Al-Salehy FK, Al-Sanafi AI. Efficient enzymatic antioxidants for oxidative stress syndrome in patients with hypertension. J Dhi Qar Sci 2008; 2: 32–3. Al-Helaly LA. Some antioxidant enzymes in workers exposed to pollutants. Raf J Sci 2011; 22: 29–38. Google Scholar Othman GO. Protective effects of linseed oil against methotrexate induced genotoxicity in bone marrow cells of albino mice Mus musculus. ZJPAS. 2016; 28: 49–53. Google Scholar Ashoka CH, Vijayalaxmi KK. Cytogenetic effects of methotrexate in bone marrow cells of Swiss albino mice. Int J Sci Res Edu 2016; 4: 4828–34. DOI - Google Scholar Rushworth D, Mathews A, Alpert A, Cooper Dihydrofolate reductase and thymidylate synthase transgenes resistant to methotrexate interact to permit novel transgene regulation. J Biol Chem 2015; 290: 22970–9. DOI - PubMed - Google Scholar Wong PT, Choi SK. Mechanisms and implications of dual-acting methotrexate in folate-targeted nanotherapeutic delivery. Int J Mol Sci 2015; 16: 1772–90. DOI - PubMed - Google Scholar Jafer ZMT, Shubber EK, Amash HS. Cytogenetic analysis of Chinese hamster lung fibroblasts spontaneously resistant to methotrexate. Nucleus 2001; 44: 28–35. Google Scholar Hussain ZK, AL-Mhdawi F, AL-Bakri N. Effect of methotrexate drug on some parameters of kidney in newborn mice. Iraqi J Sci 2014; 55: 968–73. Google Scholar Ghazi-Khansari M, Mohammadi-Bardbori A. Captopril ameliorates toxicity induced by paraquat in mitochondria isolated from the rat liver. Toxicol in Vitro 2007; 21: 403–7. DOI - PubMed - Google Scholar Dinic-olivira RJ, Sousa C, Remiao F, Durte JA, Navarro SA, Bastos L, et al. Full survival of paraquat-exposed rats after treatment with sodium salicylate. Free Radic Biol Med 2007; 42: 1017–28. DOI - PubMed - Google Scholar Attia AM, Nasr HM. Dimethoate-induced changes in biochemical parameters of experimental rat serum and its neutralization by black seed (Nigella sativa) oil. Slovak J Anim Sci 2009; 42: 87–94. Google Scholar Al-Rubaie AH.M. Effect of natural honey and mitomycin C on the effectiveness of the enzyme glutathione reductase in mice Mus musculus. Babylon Uni J 2008; 15: 1385–91.

Introduction Methods Resuts Discussion Conclusions Acknowledgments Authors' contributions Competing interests Ethical approval References Effect of Methotrexate and Omega-3 Combination on Cytogenetic Changes of Bone Marrow and Some Enzymatic Antioxidants: An Experimental Study Inaam N. Ali1, Muthana M. Awad2, Alaa S. Mahmood2,* 1 Water and Environment Directorate, Ministry of Sciences and Technology, Baghdad, Iraq 2 Department of Biology, College of Science, University of Anbar, Anbar, Iraq * Corresponding author: A. S. Mahmood (alaashm91@gmail.com) Abstract: Objective: To assess the effect of methotrexate and omega-3 combination on cytogenetic changes of bone marrow and activities of some enzymatic antioxidants. Methods: Fifty-six mature male Wistar rats were divided into two experimental groups and a control group. The first experimental group was sub-divided into three sub-groups depending on the concentration of methotrexate (MTX): X1 (0.05 mg/kg MTX), X2 (0.125 mg/kg MTX) and X3 (0.250 mg/kg MTX), which were given intraperitoneally on a weekly basis for eight weeks. The second experimental group (MTX and omega-3 group) was also sub-divided into three sub-groups (Y1, Y2 and Y3), which were injected intraperitoneally with 0.05, 0.125 and 0.25 mg/kg MTX, respectively, weekly for eight weeks accompanied by the oral administration of 300 mg/kg omega-3. The rats of the control group were given distilled water. The enzymatic activity of catalase (CAT), superoxide dismutase (SOD) and glutathione reductase (GR) were measured in the sera of rats. In addition, the mitotic index (MI) and chromosomal aberrations of bone marrow were also studied. Results: MTX resulted in a significant decrease in the activities of CAT, SOD and GR compared to the controls. It also increased the MI and chromosomal aberrations of rat bone marrows. On the other hand, omega-3 significantly increased the activities of the investigated enzymatic antioxidants and reduced the MI and chromosomal aberrations in treated mice when given in combination with MTX. Conclusions: MTX has a genotoxic effect on the bone marrow by increasing the MI and all types of chromosomal aberrations and decreasing the enzymatic activity of CAT, SOD and GR. The addition of omega-3 can lead to a protective effect by reducing the toxic and mutagenic effects of MTX. Keywords: Methotrexate, Omega-3, Antioxidant, Wistar rat, Chromosomal aberration, Mitotic index 1. Introduction Methotrexate (MTX) is a folic acid antagonist because of their chemical similarity [1]. Vezmar et al. [2] showed that MTX affects the synthesis of nucleic acids deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) by interfering with the biosynthesis of thymine and purines. It also directly affects the rapidly dividing and intact cells, especially those in the mucous membranes of the mouth, intestine and bone marrow [3]. Omega-3 is a type of unsaturated fats, which are classified as essential fatty acids that cannot be manufactured by the body and should be taken with food [4]. Sources of omega-3 include fish oils, such as salmon, sardines and tuna, as well as soybeans, walnuts, raisins and linseed, almonds and olive oils [5]. Omega-3 is used in the prevention of a number of diseases such as rheumatoid arthritis, ulcerative colitis, asthma, atherosclerosis, cancer, and cardiovascular diseases [6]. A large amount of evidence indicates that omega-3 fatty acids have significant health benefits, including anti-inflammatory and antioxidant properties besides their effect on blood cholesterol levels [7]. Antioxidants retard the oxidation process by different mechanisms such as the removal of free radicals [8]. Enzymatic antioxidants include catalase (CAT), which is the first line of defense in the cell that removes hydrogen peroxide formed during biological processes by converting it into an aldehyde, and superoxide dismutase (SOD). There are three major families of SOD enzymes: manganese SOD (Mn-SOD) in the mitochondria and peroxisomes, iron SOD (Fe-SOD) in prokaryote cells and copper/zinc SOD (Cu-Zn SOD) in the cytoplasm of eukaryote cells [9]. Therefore, changes in the metal co-factors (manganese, iron, copper and zinc) can alter the effectiveness of SOD and may lead to diseases as a result of oxidative stress [10]. Glutathione reductase (GR) is also an enzymatic antioxidant that converts the oxidized glutathione to the reduced glutathione in the presence of NADPH, which is oxidized to NADP [11]. Therefore, the aim of the present study was to assess the effects of MTX and omega-3 on the cytogenetic changes of bone marrow as well as the activities of CAT, SOD and GR enzymatic antioxidants in male rats. 2. Method 2.1. Laboratory animals and experimental design Fifty-six mature male Wistar rats (Rattus norvegicus), aged 10–12 weeks old and weighing 250–300 gm, were used in the present study. The rats were kept in separate cages, with natural 13- hour light and 11-hour dark periods in a contamination-free environment with a controlled temperature (28.0 ± 1.0°C). In addition, rats were maintained on a standard diet and tap water ad libitum. The rats were randomly allocated to two experimental groups and a control group. The first experimental group (MTX group) included 24 rats injected intraperitoneally with different MTX dilutions with distilled water [12]. It was sub-divided into three sub-groups (eight rats per sub-group) according to MTX concentration as follows: X1 (0.05 mg/kg MTX), X2 (0.125mg/kg MTX) and X3 (0.25 mg/kg MTX). All rats were given a single dose of the specified MTX concentration weekly for eight weeks. The second experimental group (MTX and omega-3 group) included 24 rats allocated to three sub-groups (Y1, Y2 and Y3), which were injected intraperitoneally with 0.05, 0.125 and 0.25 mg/kg MTX, respectively, weekly for eight weeks accompanied by the oral administration of 300 mg/kg omega-3. The control group included eight rats that were intraperitoneally injected with distilled water and given a single dose of distilled water orally weekly for eight weeks. 2.2. Blood collection and processing After the end of the dosing period, 5 ml of blood were withdrawn from the heart (by cardiac puncture) using a 5 cc disposable syringe. The collected blood was immediately poured into a clean sterile screw-capped tube (plain tube) and left for coagulation in a water bath at 37°C for 15 minutes. After coagulation of blood, the plain tube was centrifuged for 5 minutes at 1500 rpm. Then the samples were stored at -20°C for subsequent analysis. 2.3. Measurement of the activity of antioxidant enzymes The antioxidant activities of CAT, SOD and GR were measured using enzyme-linked immunosorbent assay kits purchased from Kamiya Biomedical Company (Seattle, WA, US), according to the manufacturer's instructions. 2.4. Cytogenetic study of bone marrow Rats were killed by cervical dislocation, and their hip bones were cleaned from surrounding muscles and then dissected by cutting both ends of the bone. Five milliliters of physiological buffered saline were injected inside the bone to withdraw bone marrow into a test tube. Tubes were centrifuged at 2000 rpm/10 minutes. The supernatant was then removed, and 10 ml of KCL solution (0.075 M) were added to the sediment. The mixture was then incubated at 37 °C in a water bath for 30 minutes, with shaking from time to time. The tubes were then centrifuged at 2000rpm/10 minutes to remove the supernatant. However, 5 ml of a freshly prepared fixative solution (methanol: glacial acetic acid 1:3) were added gradually in the form of droplets into the inner wall of the tube with constant mixing. After that, the tubes were placed at 4 °C for half an hour to fix the cells. This process was repeated for three times, and the cells were then suspended in 2 ml of the fixative solution. The tubes were centrifuged at 2000 rpm for 5 minutes, and the supernatant was then removed while the cells were re-suspended in 1-2 ml of cold fixative solution. After shaking the tubes, 4–5 drops were then taken from each tube onto a clean slide from a height of about three feet to provide an opportunity for the cells and nuclei to spread well. The slides were stained with acridine orange solution (0.01%) for 4–5 minutes, incubated in Sorensen’s buffer (0.06M, pH 6.5) for a minute. and then examined using a fluorescence microscope Olympus BX 51 America at a wavelength of 450–500 nm [13, 14]. A total of 1000 cells were examined, and both dividing and non-dividing cells were calculated [13]. Mitotic index (MI) was calculated according to the following formula [13]: MI= No. of dividing cells / 1000 × 100 2.5. Analysis of chromosomal aberrations of bone marrow cells A total of 1000 dividing cells were examined on the stained slides under a fluorescence microscope at a wavelength of 45–500 nm. The examined cells were at the first metaphase of the mitotic division, where chromosomal aberrations are clear and can be easily seen [13]. 2.6. Statistical analysis Data were analyzed using the Statistical Analysis System (SAS®) software, version 9.1 (Cary, NC, USA) [15]. Effects were expressed as mean ± standard error (SE) and statistically compared using a completely randomized design analysis of variance and least significant differences. Differences at P values <5 were considered statistically significant. 3. Results 3.1. Effects of MTX and MTX-omega-3 combination on antioxidant enzymatic activities Table (1) shows significantly lower SOD activities among rats treated with MTX or MTX-omega-3 compared to controls. Moreover, sera of rats receiving relatively high doses of MTX (sub-groups X2 and X3) showed the lowest enzymatic activities of 4.29 ± 0.01 IU and 3.93 ± 0.11 IU, respectively. On the other hand, CAT activity differed significantly between treated and control rats as well as among treated rats themselves, In this respect, the controls showed the highest activity of 39.38 ±0.02 IU, while those receiving the highest MTX concentration, either alone or in combination with omega-3 (sub-groups X3 and Y3), showed the lowest activities of 30.97 ± 0.03 IU and 32.12± 0.06 IU, respectively. Regarding GR activity, control rats showed a higher activity of 53.09± 0.05 IU compared to treated ones; however, the differences in GR activities in rats given low doses of MTX, either alone or in combination with omega-3 (sub-groups X1 and Y1), were not statistically significant. On the other hand, rats in sub-groups X3 and Y3 showed the lowest GR activities of 34.59 ± 0.63 IU and 37.15 ±0.01, respectively, with statistically significant differences from other sub-groups. 3.2. Effects of MTX and MTX-omega-3 combination on mitotic index of bone marrow cells Figure (1) shows a significant decrease in the MI in all treated groups compared to control. In addition, there was a reverse association between MTX concentration and MI, where rats treated with the highest dose of MTX (sub-group X3) showed a significant decrease in MI compared to all other treated rat sub-groups. In addition, rats in sub-groups treated with MTX and omega-3 (sub-groups Y1, Y2 and Y3) showed a significant increase in MI compared to their counterpart rats receiving MTX only. Table 1. Activity of antioxidant enzymes in rats treated with MTX and MTX-omega-3 Group Enzymatic activity (mean± SE) SOD (IU) CAT (IU) GR (µmol) Control 6.41±0.02 a 39.38±0.02 a 53.09±0.05 a X1 (0.05 mg MTX/ kg) 5.33±0.01 b 37.81±0.01 c 51.12±0.06 a Y1 (0.05 mg MTX + 300 mg omega-3/ kg) 6.08±0.04 a 38.40±0.02 b 51.97±0.03 a X2 (0.125 mg MTX/ kg) 4.29±0.01 cd 33.13±0.01 e 42.34±0.03 b Y2 (0.125 mg MTX + 300 mg omega-3/ kg) 4.99±0.40 b 36.68±0.02 d 43.02±3.04 b X3 (0.25 mg MTX/ kg) 3.93±0.11 d 30.97±0.03 g 34.59±0.63 c Y3 (0.25 mg MTX + 300 mg omega-3/ kg) 4.47±0.02 c 32.12±0.06 f 37.15±0.01 c SE, Standard error; IU, international unit; SOD, superoxide dismutase; CAT, catalase; GR, glutathione reductase; *statistically significant at P < 0.05; **statistically significant at P < 0.01. Means with different letters within the same column showed a statistically significant difference. 3.3. Effects of MTX and MTX-omega-3 combination on chromosomal aberrations of bone marrow cells Rats receiving higher concentrations of MTX (sub-group X3) showed a significant increase in all types of chromosomal aberrations, i.e., chromatid gaps, chromosome gaps, chromatid breaks, chromosome breaks, deletions and simple fragments (Figure 2 and Table 2) than those of the control group or other treated sub-groups. All rats treated with MTX-omega-3 combination showed a significant decrease in almost all types of chromosomal aberrations compared to their counterpart rats receiving MTX alone (Table 2). Figure 1. Effect of MTX and MTX-omega-3 on the MI of bone marrow cells of treated rats compared to the controls. The groups X1 (0.05 MTX), X2 (0.125 MTX) and X3 (0.250 MTX) were compared to the control group, while the groups Y1 (0.05 MTX+ omega-3), Y2 (0.125 MTX+ omega-3) and Y3 (0.25 MTX+ omega-3) were compared to X1, X2 and X3, respectively. Figure 2. Effect of MTX and MTX-omega-3 on chromosomal aberration as seen under fluorescence microscope after staining with acridine orange: (1) a simple fragment; (2) a chromatid gap; (3) a chromosomal gap (A) and a chromosomal break (B). 4. Discussion The present experiment reveals that the addition of omega-3 to MTX alleviates its effects on the activities of the antioxidant enzymes CAT, SOD and GR, and decreases the MI as well as all types of chromosomal aberrations in the bone marrow cells. Daham et al. [16] showed that the decline in antioxidants associated with chemotherapy is attributed to the increase in lipid peroxidation caused by these kinds of drugs, which increase the level of free radicals. In addition, Weijl et al. [17] showed that some chemotherapeutic drugs have a negative effect on the antioxidant levels such as GR, whose activity decreases as a result of its involvement in many cellular processes such as cell defenses against the toxicity of some compounds. Al-Dalawy et al. [18] found that the decrease in the level of SOD is an evidence of its increased activity due to the increased release of free radicals. MTX causes an increase in the release of free radicals, including the OH radical that causes direct damage to DNA [16]. Al-Helaly [19] showed that the amount of food taken has an effect on antioxidants, where nutritional deficiency decreases the antioxidant levels, thus increasing free radicals that cause damage to DNA. Table 2. Chromosomal aberrations of bone marrow cells in rats treated with MTX and MTX-omega-3 Group Type of chromosomal aberration(mean ± SE) Chromatid gap Chromosome Gap Chromatid breaks Chromosome breaks Deletion Simple Fragments Chromosomal aberration (%) Control 1.33±0.33 e 0.00±0.00 e 1.67±0.33 c 0.33±0.15 c 0.00±0.00 0.67±0.33 cd 0.04±0.005 f X1 2.75±0.47 cd 1.50±0.28 cd 2.50±0.64 bc 1.00±0.41 bc 0.50±0.28 bc 0.75±0.25 bcd 0.09±0.02 de Y1 1.75±0.47 de 0.75±0.25 de 1.50±0.28 c 1.00±0.00 bc 0.75±0.25 abc 0.75±0.25 abc 0.065±0.005 ef X2 4.67±0.33 b 2.67±0.33 ab 2.67±0.33 bc 1.67±0.33 ab 0.67±0.33 abc 1.67±0.33 ab 0.14±0.006 bc Y2 3.00±0.00 c 2.00±0.00 bc 3.00±0.057 bc 1.33±0.33 b 0.67±0.33 abc 0.33±0.15 d 0.106±0.003 cd X3 6.80±0.37 a 3.00±0.31 a 4.60±0.74 a 2.40±0.24 a 1.40±0.24 a 1.80±0.37 a 0.20±0.017 a Y3 5.60±0.40 ab 2.40±0.24 ab 3.60±0.24 ab 1.80±0.20 ab 1.20±0.20 ab 1.40±0.24 abc 0.16±0.003 b LSD 1.231** 0.814** 0.602** 0.841** 0.774* 0.941** 3.499* SE, Standard error; * statistically significant at P < 0.05; ** statistically significant at P < 0.01. Means with different letters within the same column showed a statistically significant difference. X1 (0.05 mg MTX/ kg); X2 (0.125 mg MTX/ kg); X3 (0.25 mg MTX/ kg); Y1 (0.05 mg MTX + 300 mg omega-3/ kg); Y2 (0.125 mg MTX + 300 mg omega-3/ kg); Y3 (0.25 mg MTX + 300 mg omega-3/ kg). In the present study, the intraperitoneal administration of MTX to rats also caused a decrease in the MI of bone marrow and a significant increase in the rate of abnormal chromosomal aberration compared to the control rats. This finding is consistent with those reported previously [20], [21]. The effect of MTX can be attributed to its ability to interfere with the genetic material, leading to the appearance of toxic and mutagenic consequences. Rushworth et al. [22] reported that MTX leads to a lack of dihydrofolate reductase, which is the key to the growth and cell division processes. This, in turn, leads to a reduction of the nucleotides involved in the building of DNA and, therefore, to a stop or obstruction of the repair mechanisms of the damaged DNA. In addition, Wong and Choi [23] concluded that MTX inhibits the action of enzymes controlling the purine metabolism, which leads to the accumulation of adenosine in addition to the damage of the molecule itself and to the occurrence of chromosomal aberrations. Jafer et al. [24] reported the ability of MTX to induce chromosomal aberration in humans or animals by preventing the repair of DNA and affecting the proteins found in chromosomes. These findings were also confirmed by Hussain et al. [25], who found that MTX causes an increase in chromosomal aberrations. In the present study, the MI showed a significant increase in rat sub-groups treated with MTX-omega-3 combination, but there was a decrease in the rate of chromosomal aberration, which confirms the role of omega-3 unsaturated fatty acids in protecting the cell from the impact of free radicals [26], [27]. Attia and Nasr [28] reported the antioxidant effect of omega-3, which was attributed to the reduction in lipid peroxidation and the increase in SOD and CAT or the stimulation of GR. It is noteworthy that GR leads to the synthesis of reduced glutathione, which is important in the defense of the cell against toxic substances and the prevention of the occurrence of mutations [29]. 5. Conclusions MTX significantly decreases the activity of enzymatic antioxidants, reduce the MI and increase the chromosomal aberrations of all types in bone marrow. This gives further evidence on the genotoxic effects of MTX on the bone marrow. On the other hand, omega-3 shows a protective effect by reducing the toxic and mutagenic effects of MTX. Acknowledgments The authors thank the staff of the Water and Environment Directorate, Ministry of Science and Technology, Baghdad, Iraq for their cooperation. They also thank Dr. Jasim Al-Niami for his technical and scientific guidance. Authors' contributions INA, MMA and ASM contributed to the study design and analyzed data. All authors contributed to the manuscript drafting and revising and approved the final submission. Competing interests The authors declare that they have no competing interests associated with this article. Ethical approval The ethical clearance of this study was obtained from the Ethics Committee of the College of Science, University of Anbar (Reference No. A. D. 51 in 30/8/2015). References Yuen CW, Winter ME. Methotrexate (MTX). In: Basic clinical pharmacokinetics, Winter ME, editor. Philadelphia, USA: Lippincott Williams & Wilkins; 2010. p.p. 304–25. Google Scholar Vezmar S, Becker A, Bode U, Jaehde U. Biochemical and clinical aspects of methotrexate neurotoxicity. Chemotherapy 2003; 49: 92–104. DOI PubMed - Google Scholar Tian H, Cronstein BN. Understanding the mechanisms of action of methotrexate implications for the treatment of rheumatoid arthritis. Bull NYU Hosp Jt Dis 2007; 65: 168–73. PubMed - Google Scholar El-Khayat Z, Rasheed WI, Elias T, Hussein J, Oraby F, Badawi M, et al. Protective effect of either dietary or pharmaceutical n-3 fatty acids on bone loss in ovariectomized rats. Maced J Med Sci 2010; 3: 9–16. DOI - Google Scholar Kris-Etherton PM, Harris WS, Appel LJ; Nutrition Committee. Fish consumption, fish oil, omega-3 fatty acids and cardiovascular disease. Arterioscler Thromb Vasc Biol 2003; 23: e20–30. DOI - PubMed - Google Scholar Calder PC. Polyunsaturated fatty acids and inflammation. Prostaglandins Leukot Essent Fatty Acids 2006; 75: 197–202. DOI - PubMed - Google Scholar Begin ME, Ells G, Das UN, Horrobin DF. Differential killing of human carcinoma cells supplemented with n-3 and n-6 polyunsaturated fatty acids. J Natl Cancer Inst 1986; 77: 1053–62. DOI - PubMed - Google Scholar Shan B, Cai YZ, Sun M, Corke H. Antioxidant capacity of 26 spice extracts and characterization of their phenolic constituents. J Agric Food Chem. 2005; 53: 7749–59. DOI - PubMed - Google Scholar Matiax J, Quiles JL, Huertas JR, Battino M. Tissue specific interactions of exercise, dietary fatty acids, and vitamin E in lipid peroxidation. Free Radic Biol Med 1998; 24 : 511–21. DOI - PubMed - Google Scholar Dean RT, Fu S, Stocker R, Davies MJ. Biochemistry and pathology of radical-mediated protein oxidation. Biochem J 1997; 324: 1–8. PubMed - Google Scholar Bashir A, Perham RN, Scrutton NS, Berry A. Altering Kinetic mechanism and enzyme stability by mutagenesis of the dimmer interface of glutathione reductase. Biochem J 1995; 312: 527–33. PubMed - Google Scholar Perret-Gentil MI. Rat Biomethodology. Laboratory Animal Resources Center. The University of Texas at San Antonio. [Cited 1 Feb. 2015]. Available from: https://www.utdallas.edu/research/docs/rat_biomethodology/ Allen JW, Shuler CF, Menders RW, Olatt SA. A simplified technique for in vivo analysis of sister chromatid exchange using 50 bromodeoxyuridine tablets. Cytogenet Cell Genet 1977; 18: 231–7. DOI PubMed - Google Scholar Forsum U, Hallén A. Acridine orange staining of urethral and cervical smears for the diagnosis of gonorrhea. Acta Derm Venereol 1979; 59: 281–2. PubMed - Google Scholar Statistical Analysis System user's guide. Version 9.1. Cary, NC, USA: SAS Institute Inc.; 2012. Daham HH, Rahim SM, Al-Hmesh MJ. The effect of radiotherapy and chemotherapy in several physiological and biochemical parameters in cancer patients. Tikrit J Pure Sci 2012; 17: 83–91. Weijl N, Elseendoorm TJ, Lentjes EG, Hopman CD, Wipkink-Bakker A, Zwinderman AH, et al. Supplementation with antioxidant micronutrients and chemotherapy-induced toxicity in cancer patients treated with cisplatin-based chemotherapy: a randomised, double-blind placebo-controlled study. Eur J Cancer 2004; 40: 1713–23. DOI - PubMed - Google Scholar Al-Dalawy SS, Al-Salehy FK, Al-Sanafi AI. Efficient enzymatic antioxidants for oxidative stress syndrome in patients with hypertension. J Dhi Qar Sci 2008; 2: 32–3. Al-Helaly LA. Some antioxidant enzymes in workers exposed to pollutants. Raf J Sci 2011; 22: 29–38. Google Scholar Othman GO. Protective effects of linseed oil against methotrexate induced genotoxicity in bone marrow cells of albino mice Mus musculus. ZJPAS. 2016; 28: 49–53. Google Scholar Ashoka CH, Vijayalaxmi KK. Cytogenetic effects of methotrexate in bone marrow cells of Swiss albino mice. Int J Sci Res Edu 2016; 4: 4828–34. DOI - Google Scholar Rushworth D, Mathews A, Alpert A, Cooper Dihydrofolate reductase and thymidylate synthase transgenes resistant to methotrexate interact to permit novel transgene regulation. J Biol Chem 2015; 290: 22970–9. DOI - PubMed - Google Scholar Wong PT, Choi SK. Mechanisms and implications of dual-acting methotrexate in folate-targeted nanotherapeutic delivery. Int J Mol Sci 2015; 16: 1772–90. DOI - PubMed - Google Scholar Jafer ZMT, Shubber EK, Amash HS. Cytogenetic analysis of Chinese hamster lung fibroblasts spontaneously resistant to methotrexate. Nucleus 2001; 44: 28–35. Google Scholar Hussain ZK, AL-Mhdawi F, AL-Bakri N. Effect of methotrexate drug on some parameters of kidney in newborn mice. Iraqi J Sci 2014; 55: 968–73. Google Scholar Ghazi-Khansari M, Mohammadi-Bardbori A. Captopril ameliorates toxicity induced by paraquat in mitochondria isolated from the rat liver. Toxicol in Vitro 2007; 21: 403–7. DOI - PubMed - Google Scholar Dinic-olivira RJ, Sousa C, Remiao F, Durte JA, Navarro SA, Bastos L, et al. Full survival of paraquat-exposed rats after treatment with sodium salicylate. Free Radic Biol Med 2007; 42: 1017–28. DOI - PubMed - Google Scholar Attia AM, Nasr HM. Dimethoate-induced changes in biochemical parameters of experimental rat serum and its neutralization by black seed (Nigella sativa) oil. Slovak J Anim Sci 2009; 42: 87–94. Google Scholar Al-Rubaie AH.M. Effect of natural honey and mitomycin C on the effectiveness of the enzyme glutathione reductase in mice Mus musculus. Babylon Uni J 2008; 15: 1385–91.

The jalapeño is derived from the Capsicum genus of the family Solanaceae. Jalapeños are members of a diverse group, which also include ancho poblano, cayenne, serrano, Anaheim, banana, Asian, habanero, and Hungarian wax peppers. Hot peppers are classified by their heat and shape. The heat of the pepper comes from the chemical compound capsaicin, which is measured by the Scoville scale. This 8-page fact sheet is a guide of jalapeño and other hot pepper varieties used in Florida was written by Monica Ozores-Hampton and Gene McAvoy, and published by the UF Department of Horticultural Sciences, October 2014.

AbstractThe fungal pathogens of spruce are well known in Europe and elsewhere. Therefore, it was surprising to discover a new fungal species and genus in Central Europe that attacks the pollen cones of three spruce species. The new ascomycete forms apothecia on stromatized pollen cones of Norway spruce (Picea abies) and Serbian spruce (Picea omorika) in mountain areas and on West Himalayan spruce (Picea smithiana) planted in urban lowland regions of Switzerland, Germany, and Italy. It was also detected in France, based on metabarcode sequences deposited in the GlobalFungi database. Its sudden appearance and the different origins of the host trees in Europe and Asia leave the origin of the fungus unclear. The new fungus might be a neomycete for Europe. A phylogenetic analysis using SSU, LSU, ITS, RPB2, and TEF1 sequences classified the fungus as a member of Sclerotiniaceae (Helotiales, Leotiomycetes). However, it differs morphologically from the other genera of this family in having an ascus without apical apparatus containing four mainly citriform spores with 16 nuclei each. Furthermore, it is the only known cup fungus that parasitizes pollen cones of conifers by stromatizing their tissue and infecting pollen grains. The fungus does not seem to cause major damage to the spruce populations, as only a few pollen cones per tree are affected. All this leads us to describe the newly discovered fungus as the new species and new genus Microstrobilinia castrans, the fungus that castrates pollen cones of spruce.

Two lander-based devices, the Bubble-Box and GasQuant-II, were used to investigate the spatial and temporal variability and total gas flow rates of a seep area offshore Oregon, United States. The Bubble-Box is a stereo camera–equipped lander that records bubbles inside a rising corridor with 80 Hz, allowing for automated image analyses of bubble size distributions and rising speeds. GasQuant is a hydroacoustic lander using a horizontally oriented multibeam swath to record the backscatter intensity of bubble streams passing the swath plain. The experimental set up at the Astoria Canyon site at a water depth of about 500 m aimed at calibrating the hydroacoustic GasQuant data with the visual Bubble-Box data for a spatial and temporal flow rate quantification of the site. For about 90 h in total, both systems were deployed simultaneously and pressure and temperature data were recorded using a CTD as well. Detailed image analyses show a Gaussian-like bubble size distribution of bubbles with a radius of 0.6–6 mm (mean 2.5 mm, std. dev. 0.25 mm); this is very similar to other measurements reported in the literature. Rising speeds ranged from 15 to 37 cm/s between 1- and 5-mm bubble sizes and are thus, in parts, slightly faster than reported elsewhere. Bubble sizes and calculated flow rates are rather constant over time at the two monitored bubble streams. Flow rates of these individual bubble streams are in the range of 544–1,278 mm3/s. One Bubble-Box data set was used to calibrate the acoustic backscatter response of the GasQuant data, enabling us to calculate a flow rate of the ensonified seep area (∼1,700 m2) that ranged from 4.98 to 8.33 L/min (5.38 × 106 to 9.01 × 106 CH4 mol/year). Such flow rates are common for seep areas of similar size, and as such, this location is classified as a normally active seep area. For deriving these acoustically based flow rates, the detailed data pre-processing considered echogram gridding methods of the swath data and bubble responses at the respective water depth. The described method uses the inverse gas flow quantification approach and gives an in-depth example of the benefits of using acoustic and optical methods in tandem.

Thermophysical properties of rock and soil are important parameters that affect the efficiency of shallow geothermal energy utilization. This paper analyzes the thermophysical parameters (specific heat capacity and thermal conductivity) and their influencing factors based on 6467 sample data in 24 provincial capitals, China. The statistical distributions of thermophysical properties are analyzed based on histogram plots. Linear regression analysis is conducted to investigate the correlation between thermophysical properties and lithology (classified as metamorphic rocks, igneous rocks, chemical sedimentary rocks, clastic sedimentary rocks, and loose sediments in this study)), density, and water content. The results show that the thermophysical properties are influenced predominantly by lithology. The measured values of thermophysical properties generally show normal distribution characteristics. The specific heat capacity of loose sediments is relatively high, the thermal conductivity of other four types are relatively high. The specific heat capacity is negatively correlated with density, positively correlated with water content, whereas thermal conductivity is positively correlated with density, and negatively correlated with water content. The findings obtained in this study provide evidence and guidelines for the investigation, evaluation, and development of shallow geothermal resources.

Abstract @@@In recent times, repellents and synthetic drugs have been identified as having negative toxicity effects on humans and the environment. Apart from the unfavourable effects on man and livestock caused by these chemicals-based (synthetic) repellents, they are also expensive, non-biodegradable, and no more effective because mosquitoes are getting adapted. With these drawbacks, an eco-friendly plant-based insecticide as a substitute is needed urgently. This paper reviews the extraction and use of essential oil from the leaves of Mentha spicata, Ocimum gratissimum, and Moringa oleifera as mosquito repellent. Carvone, Eugenol, and 9-Octadecenoic acid were discovered to be the most active components in the M. spicata, O. gratissimum, and M. oleifera extracts, respectively, using gas chromatography-mass spectrometry (GC-MS). Highlights In recent times, repellents and synthetic drugs have been identified as having negative toxicity effects on humans and the environment. Apart from the unfavorable effects on man and livestock caused by these chemical-based (synthetic) repellents, they are also expensive, non-biodegradable, and no more effective because mosquitoes are getting adapted. An eco-friendly plant-based insecticide as a substitute is needed urgently. Diseases transmitted by mosquitoes are still a significant reason for the global mortality rate, with over 700 million individuals experiencing such diseases every year. With the proper formulation of other repellent forms using their oils, they can replace non-degradable synthetic mosquito repellents since they are eco-friendly. In general, the mosquitocidal activity and percentage protection of plant extract increase with increasing concentration of the extracts in different formulations. This paper is our original work. We certify that this manuscript has not been published in part or whole elsewhere in any language, and it has not been submitted to any other journal for reviews.

In the present study the occurrence of the Giant African snail, Achatina fulica reported from Jalgaon, Dapoli District Ratnagiri (M.S), India in various field during rainy season but their occurrence is throughout the year. Due to hibernation, these are not much active as compared to rainy season. According to native people of the area, severe plant and vegetable loss was not only due to infestation but also due to the stinking smell of the mucus layer in wetland dry conditions released by the Giant African snail, Achatina fulica different plants. The pest, also classified as a large-scale invader, has been known to be difficult to exterminate without harming nearby organisms as well as ecosystems. Hence, there should be a proper method of control measure of this pest. The following article describes possible methods of control measures researched since ages chemical as well as non-chemical. Even though a lot were proven to be unsuccessful, some methods do play an important role in the extermination of this species with respect to collective efforts, destruction scale, costings etc.Mechanical extermination is a lot more effective than the chemicals introduced which have a possibility of getting washed away due to rains or other possible circumstances. This article mainly focuses on discussing previously studied control methods and defining the criteria needed to develop a suitable pesticide.

Electronic waste (e-waste) recycling presents an opportunity to reclaim materials from a secondary resource and to create jobs and other economic opportunities. E-waste consists of various materials such as metals, plastics, glass, and other chemical substances. Some of these materials are hazardous if processed or disposed of improperly. Therefore, e-waste is classified as hazardous in South African law up until the hazardous components are removed. With the appropriate infrastructure and technology, a large portion of materials contained in e-waste can be reclaimed, and any adverse impacts of irresponsible management prevented. The private sector has played a proactive role in shaping the South African waste economy, and the government is taking strides to draw up enabling regulatory frameworks. Through a literature review and stakeholder engagements, this paper unpacks the organisation of the South African e-waste recycling industry. We consider whether the legal environment drives a common vision for a circular e-waste economy and probe the barriers to e-waste recycling across the value chain. The findings indicate that the development of the e-waste recycling sector in South Africa is dependent on a robust collection network and the enabling of local end-processing, refining, and manufacturing capacity. The availability and quality of input material and the development of local refining and manufacturing capacity are co-dependent and should be addressed simultaneously.