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1
Zilliges, Yvonne. "Molekulare Funktionsanalyse von Microcystin in Microcystis aeruginosa PCC 7806." Doctoral thesis, Humboldt-Universität zu Berlin, Mathematisch-Naturwissenschaftliche Fakultät I, 2008. http://dx.doi.org/10.18452/15782.
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Microcystine sind die wohl bekanntesten cyanobakteriellen Toxine. Sie werden im Wesentlichen durch die im Süßwasser weltweit verbreitete, koloniebildende Gattung Microcystis synthetisiert. Die biologische Funktion dieser Peptide ist jedoch ungeklärt. In dieser Studie wurde die Fragestellung erstmals über einen globalen Ansatz auf molekularer Ebene analysiert. Die proteomischen Analysen zwischen M. aeruginosa PCC 7806/ Wildtyp und einigen Microcystin-freien Mutanten deuten auf eine physiologische Rolle der Microcystine. Microcystine beeinflussen die Abundanz zahlreicher Proteine. Prominentester Vertreter ist RubisCO – Schlüsselenzym des Calvin Zyklus. RubisCO und andere im 2D selektierte Proteine konnten außerdem als mögliche zelluläre Bindepartner des Microcystins identifiziert werden. Möglicherweise bindet MC an bestimmte Cysteinreste dieser Proteine. Mit dem Knockout der mcy-Gene geht außerdem eine Überexpression eines Morphotyp-spezifischen Proteins einher, das MrpC genannt wurde. Dieses Protein vermittelt möglicherweise Zell-Zell-Interaktionen in Microcystis.<br>Microcystins are the most common cyanobacterial toxins found in freshwater lakes and reservoirs throughout the world. They are frequently produced by the unicellular, colonial cyanobacterium Microcystis; however, the role of the peptide for the producing organismen is poorly understood. In this study we describe the first global approach to investigate this topic on a molecular level. Proteomic studies with M. aeruginosa PCC 7806 wild-type and several microcystin-deficient mutants indicated a physiological function for microcystin. Microcystin was shown to influence the abundance of several proteins which have an intra- or extracellular function. A prominent candidate is RubisCO, the key enzyme of the calvin cycle. RubisCO and other proteins, initially selected by 2D analysis, are putative cellular binding partners of microcystin. A potentially interaction mechanismen is the kovalent binding of microcystin to cysteine residues of the protein. Moreover, several knockouts of microcystin biosynthesis genes result in an overexpression of a putative morpho-type specific factor, named MrpC. This protein possibly mediates cell-cell interactions in Microcystis.
2
Phelan, Richard Reginald. "Microcystin enhances the fitness of microcystin producing cyanobacteria at high light intensities by either preventing or retarding photoinhibition." Thesis, Nelson Mandela Metropolitan University, 2013. http://hdl.handle.net/10948/d1020957.
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Several genera of cyanobacteria produce microcystin, a monocyclic peptide, with a unique chemical structure. To date, there have been over a 100 different structural variants of microcystin which have been identified. Microcystin production is affected by numerous environmental factors. However, the primary modulating factor for intracellular microcystin quota is the intracellular N:C ratio. No clearly defined biological role has been described for microcystin. Proposed roles for microcystin include defence against plankton grazers, metal chelation, an infochemical and a protectant against oxidative stress. There is sufficient evidence to support a biological role for microcystin in photosynthesis: microcystin is predominantly located in the thylakoid membranes, the microcystin gene cluster is differentially expressed as a function of light and a growth advantage for the microcystin producer in saturating light intensities. The purpose of this study is to investigate a possible biological role for microcystin in preventing photoinhibition and thus explaining the growth advantage observed in toxin-producers over non-toxin-producers. The uptake of exogenous microcystin was observed in Synechocystis PCC 6803 which was internalized and located in the thylakoid membranes and caused the inhibition of photosynthesis. Microcystin variants and increasing concentrations of microcystin-LR had no effect on the fluidity of the thylakoid membranes. The exposure of thylakoid membranes from Synechocystis PCC 6803 to physiologically relevant concentrations of different microcystin variants resulted in the inhibition of photosystem II activity but not photosystem I activity. The inhibition of photosystem II was variant dependent and concentration dependent for microcystin-LR and microcystin-RR. Chlorophyll a fluorescence data showed that photosystem II inhibition was caused by the inhibition of the oxygen evolving complex. Furthermore, a completion study revealed that the microcystin-producing Microcystis PCC 7806 had a competitive advantage over the non-microcystin producing ΔmcyA mutant of Microcystis PCC 7806 at high light intensities. The data indicates that microcystin protects the toxin-producer by either retarding or preventing photoinhibition and thus identifying the first data supported function for microcystin in cyanobacteria.
3
Phelan, Richard Reginald. "A potential biological role for microcystin in photosynthesis in Microcystis Aeruginosa." Thesis, Nelson Mandela Metropolitan University, 2009. http://hdl.handle.net/10948/1285.
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Neither the ecological role nor the metabolic function of microcystin is known. Cellular microcystin concentrations correlate to cellular nitrogen status for a given environmental phosphorous concentration and specific growth rate. Microcystin production is enhanced when the rate of nitrogen accumulation exceeds the relative specific growth rate and/or when cellular N:C ratios exceed the Redfield ratio as a function of reduced carbon fixation, suggesting enhanced production of microcystin under carbon stress. Additionally, a strong correlation between medium phosphate and carbon fixation, and the negative correlation between medium phosphate and microcystin combined with the cellular localization of microcystin in thylakoids supports a possible role for microcystin in enhancement of photosynthesis. Batch cultures of both Microcystis aeruginosa PCC7806 and a mcyA- knockout mutant of PCC7806 were therefore cultured at different light intensities and media treatments, so as to vary cellular N:C ratios and concentrations, and sampled for analysis of microcystin concentration, cell numbers and residual medium nitrates. Inter-strain differences in photosynthetic electron transfer rates and levels were monitored using a Hansatech PEA fluorometer and compared to cellular microcystin concentrations. An enhanced survival was observed at high light, where the toxic strain survived while the nontoxic strain became chlorotic. A strong correlation (r2 = 0.907, p< 0.001, N=22) between microcystin concentration and growth rate was observed at high light conditions. No such advantage was observed at optimal or low-light conditions and media composition had no significant effect on the relationship between toxicity and survival at high light. PCC7806 showed elevated PI(abs) values compared to the mcyA knockout strain, which indicates an increased stability of PSII. A strong correlation between PI(abs) and microcystin (r = 0.88, p< 0.005, N=15) was observed for cultures grown in modified BG11 containing 25 mM under continuous illumination of 37 μmol of photons m-2.s-1. No correlation was observed between PI(abs) and microcystin for the other treatments. The toxin producer had significantly higher values for density of active reaction centers and ii quantum efficiency compared to the mutant. A decrease in F0 in the mutant suggests degradation of the phycobiliproteins, whereas PCC7806 didn’t show a significant decrease in F0 Data indicate that microcystins play a role in photosynthesis by preventing chlorosis in saturating light conditions either by enhancing the redox stability of the phycobiliproteins or PS II, thus preventing photooxidation.
4
Downing, T. G. "The role of nitrogen in the regulation of microcystin content in Microcystis aeruginosa." Thesis, Stellenbosch : Stellenbosch University, 2005. http://hdl.handle.net/10019.1/50523.
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Thesis (PhD)--University of Stellenbosch, 2005.<br>ENGLISH ABSTRACT: Several genera of cyanobacteria produce a range of toxins. The increased
rate of eutrophication of surface fresh waters due to anthropogenic inputs has
resulted in more frequent and severe cyanobacterial bloom events. Such
bloom events make impoundments unsuitable for recreational use and
increase the cost of production of potable water due to the necessity for
removal of toxins released from cells during the purification process.
Microcystis aeruginosa is the major freshwater bloom-forming toxic
cyanobacterium. Concentrations of the hepatotoxin, microcystin, are highly
variable in blooms. Published literature on environmental conditions leading to
increased microcystin production was often contradictory and in many cases
did not consider all relevant parameters. However, environmental nitrogen
and phosphorus, temperature and light, and growth rate were implicated in
regulation of toxin content. The purpose of this work was therefore to
investigate environmental factors (specifically nitrogen and phosphorus) and
cellular activities (specifically carbon fixation and nitrogen uptake rates and
growth rate) involved in the modulation of microcystin production in M.
aeruginosa in order to clarify the role of these parameters, and in an attempt
to identify regulatory mechanisms for microcystin production. Environmental
nitrogen, phosphorus and growth rate were shown to co-modulate microcystin
production in M. aeruginosa. Adequate phosphorus is required for
photosynthetic carbon fixation. Phosphorus uptake by M. aeruginosa is
strongly correlated with carbon fixation rate. Although microcystin content
increased with increasing nitrogen:phosphorus ratios in culture medium,
under phosphorus limitation microcystin content was lower irrespective of
nitrogen concentrations. This observation and the requirements for fixed
carbon for nitrogen assimilation therefore prompted investigation of the effects
of cellular carbon fixation and nitrogen uptake in the modulation of microcystin
production. Microcystin production was found to be enhanced when nitrogen
uptake rate relative to carbon fixation rate was higher than that required for
balanced growth. The cellular nitrogen:carbon ratio above which microcystin
concentrations increased substantially, corresponded to the Redfield ratio for balanced growth. Investigation of potential regulatory mechanisms involving
the cyanobacterial nitrogen regulator, NtcA, yielded putative NtcA binding
sites indicative of repression in the microcystin synthetase gene cluster. In
culture, the polypeptide synthetase module gene, mcyA, and ntcA were
inversely expressed as a function of carbon-fixation:nitrogen-uptake potential.
However, no increase or decrease in microcystin production could be linked to
either glutamine, glutamate or a-ketoglutarate, metabolites that are involved in
regulation of ntcA. The role of NtcA in regulation of microcystin production
could therefore not be confirmed. In conclusion, these data suggest that
microcystin production is metabolically regulated by cellular C:N balance and
specific growth rate. The primary importance of nitrogen and carbon was
demonstrated by a simple model where only nitrogen uptake, carbon fixation
and growth rate were used to predict microcystin levels. The model also
explains results previously described in literature. Similarly, an artificial neural
network model was used to show that the carbon fixation dependence on
phosphorus allows accurate prediction of microcystin levels based on growth
rate and environmental nitrogen and phosphorus.<br>AFRIKAANSE OPSOMMING: Verskeie genera van sianobakterieë produseer 'n verskeidenheid van
toksiene. Die toename in die tempo van eutrofikasie van varswater
oppervlaktes as gevolg van antropogeniese insette veroorsaak al hoe meer
en al hoe erger sianobakteriële infestasies. Dit veroorsaak probleme vir
ontspanninggebruik van hierdie waters en verhoog die koste van produksie
van drinkbare water as gevolg van die noodsaak om die toksiene wat deur die
selle gedurende die suiweringsproses vrygelaat word te verwyder. Microcystis
aeruginosa is die belangrikste varswater bloeisel-vormende toksiese
sianobakterium. Die konsentrasie van die hepatotoksien mikrosistien is hoogs
varieerbaar in sulke bloeisels. Gepubliseerde literatuur oor die
omgewingskondisies wat lei na verhoogde mikrosistienproduksie is dikwels
weersprekend en neem in vele gevalle nie al die relevante parameters in ag
nie. Desnieteenstaande word omgewingstikstof, fosfor, temperatuur en lig,
asook groeisnelheid, geïmpliseer in die regulering van toksieninhoud. Die doel
van hierdie navorsing was dus om omgewingsfaktore (spesifiek stikstof en
fosfor) en sellulêre aktiwiteite (spesifiek koolstoffiskering en die snelheid van
stikstofopname en van groei) betrokke by die modulering van
mikrosistienproduksie in M. aeruginosa te ondersoek in 'n poging om die rol
van hierdie parameters te verstaan en om regulatoriese meganismes vir
mikrosistienproduksie te identifiseer. In hierdie studie is aangetoon dat
omgewingstikstof en fosfor sowel as groeisnelheid mikrosistienproduksie in M.
aeruginosa ko-moduleer. Genoegsame fosfor word benodig vir fotosintetiese
koolstoffiksering. Fosforopname deur M. aeruginosa korreleer sterk met die
snelheid van koolstoffiksering. Alhoewel mikrosistieninhoud toegeneem het
met 'n toename in die stikstof:fosfor verhouding in die kultuurmedium, was die
mikrosistieninhoud onder kondisies van fosforlimitering laer ongeag die
stikstofkonsentrasie. Hierdie waarneming, tesame met die noodsaak van
gefikseerde koolstof vir stikstofassimilering, het gelei na 'n studie van die
effekte van sellulêre koolstoffiksering and stikstofopname op die modulering
van mikrosistienproduksie. Dit is gevind dat mikrosistienproduksie verhoog
was wanneer die snelheid van stikstofopname relatief tot die snelheid van koolstoffiksering hoër was as die waarde wat benodig word vir gebalanseerde
groei. Die sellulêre stikstof:koolstof verhouding waarbo
mikrosistienkonsentrasies beduidend verhoog is stem ooreen met die
Redfield verhouding vir gebalanseerde groei. 'n Ondersoek na potensiële
reguleringsmeganismes waarby die sianobakteriële stikstofreguleerder NtcA
betrokke is het gelei na die ontdekking van moontlike NtcA bindingseteis; dit
kan dui op die repressie van die mikrosistiensintetase geengroepering. Onder
kultuurkondisies is gevind dat die geen vir die polipeptiedsintetase module,
mcyA, en ntcA omgekeerd uitgedruk word as 'n funksie van
koolstofopname:stikstofopname potensiale. Geen toename of afname in
mikrosistienproduksie kon egter gekoppel word aan óf glutamien, óf
glutamaat, óf a-ketoglutaraat nie, metaboliete wat betrokke is by die
regulering van ntcA. Die rol van NtcA in die regulering van
mikrosistienproduksie kon dus nie bevestig word nie. Die gevolgtrekking is
dus gemaak dat mikrosistienproduksie metabolies gereguleer word deur die
C:N balans en die spesifieke groeisnelheid. Die primêre belang van stikstof en
koolstof is gedemonstreer deur 'n eenvoudige model waarin slegs
stikstofopname, koolstoffiksering en groeisnelheid gebruik word om
mikrosistienvlakke te voorspel. Die model verklaar ook resultate wat tevore in
die literatuur beskryf is. Soortgelyk is 'n artifisiële neurale netwerkmodel
gebruik om te toon dat die afhanklikheid van koolstoffiksering van fosfor
akkurate voorspelling van mikrosistienvlakke gebaseer of groeisnelheid en
omgewingstikstof en fosfor moontlik maak.
5
Sember, Craig Stewart. "The effect of nutrient levels and ratios on the growth of Microcystis aeruginosa and microcystin production." Thesis, University of Port Elizabeth, 2002. http://hdl.handle.net/10948/287.
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This study reports the findings on the effect of nitrates and phosphates on the biomass and toxin production of various strains of the unicellular non-nitrogen fixing cyanobacterium, Microcystis aeruginosa. The occurrence of blooms of Microcystis aeruginosa and microcystin in freshwater impoundments across the globe has been on the increase lately due to increased levels of eutrophication, resulting in human and animal deaths and illness, as well as drinking and recreational water foulment. A range of environmental factors have been shown to effect growth and microcystin production. Existing literature however is somewhat contradictory as to the effects of these physical and chemical factors on toxin production. Therefore Microcystis aeruginosa strains were cultured under batch and continuous conditions to determine the effect of nitrate and phosphate concentrations and ratios on biomass and toxin production. Cultures were analysed with regards to internal nutrient stores, biomass production, nutrient depletion, photosynthetic efficiency and microcystin production. Results showed that microcystin production correlated to growth rate, photosynthetic efficiency and internal nitrogen stores and that an optimal N:P ratio was associated with microcystin levels, growth rate and photosynthetic efficiency. Results therefore led to the conclusion that the nitrogen, carbon, and phosphate balance within the cell is closely associated with microcystin production. Whether or not microcystin is produced to maintain this balance or produced as a function of this balance remains to be determined.
6
Delaney, James M. "The biological activity of microcystin-LR, isolated from the cyanobacterium Microcystis aeruginosa against insects." Thesis, University of Newcastle Upon Tyne, 1996. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.308015.
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7
Zemskov, Ivan [Verfasser]. "Total Synthesis of Microcystin-LR, Microcystin-LF, and Unnatural Derivatives thereof / Ivan Zemskov." Konstanz : Bibliothek der Universität Konstanz, 2016. http://d-nb.info/1169046711/34.
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8
Meissner, Sven Verfasser], and Elke [Akademischer Betreuer] [Dittmann. "Implications of Microcystin Production in Microcystis aeruginosa PCC 7806 / Sven Meissner ; Betreuer: Elke Dittmann-Thünemann." Potsdam : Universität Potsdam, 2015. http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-75199.
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9
Meissner, Sven [Verfasser], and Elke [Akademischer Betreuer] Dittmann-Thünemann. "Implications of Microcystin Production in Microcystis aeruginosa PCC 7806 / Sven Meissner ; Betreuer: Elke Dittmann-Thünemann." Potsdam : Universität Potsdam, 2015. http://d-nb.info/1218398965/34.
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10
Velkme, Erik. "A novel method for antisense oligonucleotide gene expression manipulation in toxigenic cyanobacterial species, Microcystis aeruginosa." OpenSIUC, 2020. https://opensiuc.lib.siu.edu/theses/2781.
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Algal blooms caused by toxigenic cyanobacterial species are an increasing economic burden globally, as high anthropogenic inputs of nitrogen and phosphorous, coupled with rising levels of atmospheric CO2, promote eutrophication and enhance bloom proliferation. Of the freshwater bloom forming species, Microcystis aeruginosa has garnered the most attention due to the production of toxic secondary metabolites known as microcystins. These cyclic peptides are potent eukaryotic protein phosphatase 1 and 2A inhibitors, and can induce hepatic damage if concentration levels reach above the World Health Organization level of 1 µg/L. Current mitigation strategies of water column disruption or by use of broad acting chemicals, are limited in their range and may cause unwanted off target effects to the surrounding biota. Antisense oligonucleotides are short single-stranded DNA polymers that hybridize with transcribed mRNA, and suppress translation of protein products through steric hindrance of ribosomes, or by RNAse H degradation of the DNA/RNA bound complex. While antisense oligonucleotide applications have proven successful in the pharmaceutical industry, their potential remains largely unexplored in environmental contexts. For this reason, we investigated the knockdown of microcystin synthetase gene cluster mcyE in M. aeruginosa. We found that ionic charge neutralization coupled with heat shock were effective chemical competence based methods for delivery, mcyE transcript abundance in cells treated with phosphodiester linked antisense oligonucleotides significantly decreased in RT-qPCR analysis, and production of intracellular microcystin significantly decreased over a 24 hour period (-1.9 fg/cell). This work demonstrates a novel proof of concept for the potential use of exogenous antisense oligonucleotides to target M. aeruginosa in harmful algal bloom occurrences.
11
Woller-Skar, M. Megan. "Zebra Mussel (Dreissena Polymorpha) Promotion of Cyanobacteria in Low-Nutrient Lakes and the Subsequent Production and Fate of Microcystin." Bowling Green State University / OhioLINK, 2009. http://rave.ohiolink.edu/etdc/view?acc_num=bgsu1256928121.
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12
Ha, Joo Hyun. "A study on characteristics of microcystin-producing cyanobacterial bloom and microcystin production using real-time PCR." 京都大学 (Kyoto University), 2009. http://hdl.handle.net/2433/124526.
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Drever, Matthew. "Generating microcystin antibodies by phage display." Thesis, University of Aberdeen, 2007. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.445137.
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A naïve human antibody fragment phage display library was screened against conjugated microcystin-LR (MCLR). Phage antibodies were eluted with free MCLR to promote the isolation of free antigen binders. Isolated antibody fragments were cloned into a soluble expression vector and single-chain antibody (scAb) expressed in a bacterial host. All scAbs bound free antigen in an indirect competition ELISA and levels of sensitivity were determined. The most responsive clone 3A8 had an 800-fold increase in sensitivity to MCLR than previously documented scAbs and was able to detect levels below guidelines for MCLR in drinking water set by the World Health Organisation (WHO) at 1 μg/L. Despite its sensitivity to MCLR, full characterisation revealed low levels of cross-reactivity with other microcystin variants. <i>In vitro</i> affinity maturation was employed to increase cross-reactivity to other microcystin variants in scab 3A8. A chain-shuffled library was constructed where a naïve V<sub>L</sub> repertoire was shuffled with clone 3A8 V<sub>H</sub> chain. The library was screened and isolated phage antibodies cloned and expressed as scAbs. Chain-shuffled clone CSB-D9 displayed a 50-fold increase in cross-reactivity for a single microcystin variant and was capable of detecting all variants available below the guideline levels of 1 μg/L. The highly cross-reactive scab was used for the determination of total microcystin content in cyanobacterial extracts and results showed good correlation with HPLC quantification. Immunoaffinity chromatography utilising scab CSB-D9 was used for the concentration of trace amount of microcystin from large volumes of water, out-performing columns generated with anti-microcystin whole antibody molecules. The ability of scab CSB-D9 to protect cultured hepatocytes against microcystin-induced apoptosis was also determined. Results indicated a possible therapeutic application for human antibody fragments isolated from phage display libraries.
14
Qiao, Qin. "Reprotoxic effects of microcystins and secondary metabolites produced by cyanobacteria Microcystis in adult medaka fish." Thesis, Paris, Muséum national d'histoire naturelle, 2016. http://www.theses.fr/2016MNHN0022/document.
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Les efflorescences de cyanobactéries sont susceptibles d’avoir des effets néfastes sur les organismes des écosystèmes aquatiques, ainsi que sur les populations environnantes, notamment à travers la production de nombreuses molécules potentiellement toxiques (appelées cyanotoxines). Jusqu'à présent, une des cyanotoxines les plus étudiées est la microcystine (MC). Cette thèse a pour objectif d’évaluer la toxicité potentielle sur la reproduction de la MC-LR et de l'extrait d'une souche de Microcystis productrice de MCs en étudiant leurs effets toxiques sur le foie et les gonades de poissons medaka adultes exposés de manière aiguë ou chronique.Une étude complète du foie des poissons médaka deux sexes a été menés par ailleurs, attestant d'un fort dimorphisme sexuel aussi bien au niveau cellulaire que moléculaire et souligne les importantes spécificités métaboliques du foie entre les deux sexes, notamment pour le maintien de la compétence de reproduction chez les poissons medaka adultes femelles.Dans l'étude des effets induits par une exposition aiguë, les poissons medaka adultes ont été exposés par gavage à 10 μg.g-1 bw de MC-LR pure pendant 1 heure. L'examen histologique et l'immunolocalisation des MCs du foie de poisson traité par la MC-LR ont révélé des lésions hépatiques sévères ainsi qu'une distribution intense de la MC-LR dans le foie, localisée particulièrement dans le cytoplasme et dans le noyau des hépatocytes. Dans la gonade des poissons traités, la MC-LR a été détectée dans les tissus conjonctifs de l'ovaire et des testicules. De plus, l’observation par microscopie électronique couplé à la technique d’immunogold a révélé, pour la première fois, que la MC-LR était également détectable dans le chorion, le cytoplasme et le vitellus des ovocytes matures.Au cours des études des effets induits par l’exposition chronique, les poissons medaka adultes ont été exposés durant 28 jours par balnéation à 1 et 5 μg.L-1 de MC-LR et à un extrait de la souche de Microcystis aeruginosa (PCC 7820) productrice de microcystines (5 μg.L-1 équivalent MC-LR). Ces résultats ont révélé que la MC-LR et l'extrait de Microcystis induisent des effets délétères sur différents paramètres de reproduction, tels la fécondité et le taux d’éclosion des embryons. La cause principale de ces perturbations de la reproduction semblent principalement résulté d’un dysfonction hépatique globale induite par les traitements aux MCs (hépatotoxiques, notoires), plutôt qu’à des effets directs sur les gonades. Dans l'ensemble, les résultats de cette thèse démontrent que même si les microcystines pourraient avoir un impact direct, mais modéré, sur la fonction gonadique en induisant une cytotoxicité dans les cellules somatiques gonadiques et les cellules reproductrices, elle semble avoir principalement avoir un impact indirect sur la fonction reproductrice en perturbant la fonction hépatique générale. Ces données améliorent notre compréhension des processus liés à la toxicité potentielle des cyanotoxines pour la reproduction chez un poisson modèle, et fait d’une manière générale progresser questionnement quant à la protection des populations exposées à ces cyanotoxines<br>Cyanobacterial blooms threaten human health as well as other living organisms of the aquatic environment, particularly due to the production of natural toxic components (called cyanotoxins). So far, one of the most studied cyanotoxins is the microcystin (MC). This thesis evaluated the potential reproductive toxicity of MC-LR and the extract of one Microcystis strain (MC-producing) by investigating their toxic effects on the liver and gonad of adult medaka fish with one acute and one chronic study.An investigation of the metabolic specificities of the liver in two genders of medaka fish was performed prior to the MC-containing exposure, which attests to a strong sexual dimorphism of medaka liver, and highlights the importance of metabolic adjustments of the liver for maintaining the reproductive competency in adult medaka fish.In the acute study, adult medaka fish were administered with 10 μg.g-1 bw of pure MC-LR for 1 hour by gavage. The histological examination and immunolocalization of the MC-treated fish liver revealed a severe liver lesion along with an intense distribution of MC-LR in the liver, being particularly localized in the cytoplasm and nucleus of hepatocytes. In the gonad of MC-treated fish, MC-LR was shown to be present in the connective tissue of ovary and testis. Additionally, immunogold electron microscopy, for the first time, revealed that MC-LR was also localized in the chorion, cytoplasm and yolk vesicles of oocytes.Overall, the results of this thesis demonstrates that MC might directly impact gonadal function by inducing cytotoxicity in gonadal somatic cells and reproductive cells, and it could also impact the reproductive function indirectly by disturbing the general liver function. This improves our understanding of the potential reproductive toxicity of cyanotoxins in model fish, and advances our current knowledge on the protection of aquatic organism populations as well as human health from cyanotoxin issues
15
Kehr, Jan-Christoph. "Characterisation of the lectin microvirin from Microcystis aeruginosa PCC 7806 and new insights into the role of microcystin." Doctoral thesis, Humboldt-Universität zu Berlin, Mathematisch-Naturwissenschaftliche Fakultät I, 2009. http://dx.doi.org/10.18452/15994.
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Sowohl in Süßwasserseen als auch in marinen Gewässern kommt es immer wieder zu Massenentwicklungen von Cyanobakterien, sogenannten “Blüten”. In Seen werden diese oftmals von Cyanobakterien der Gattung Microcystis dominiert, deren Arten häufig Toxine bilden. Die verbreitesten dieser Toxine sind die leberschädigen Microcystine, die eine Klasse nichtribosomal synthetisierter Peptide darstellen. Nachdem die toxische Wirkung der Microcystine bisher als deren Hauptfunktion angesehen wurde, deuten neuere Forschungsergebnisse darauf hin, dass Microcystine eine andere Primärfunktion für die Produzenten besitzen. Im Rahmen dieser Studie wurde Microvirin (Mvn), ein putatives Lektin aus Microcystis aeruginosa PCC 7806, von dem angnommen wurde, dass es funktional mit Microcystin assoziiert ist, charakterisiert. Zunächst konnte gezeigt werden, dass Mvn tatsächlich zuckerbindende Aktivität besitzt und spezifisch Mannan, ein Oligosaccharid aus Mannoseuntereinheiten, erkennt. Bindestudien zeigten, dass Zucker dieses Typs auf der Zelloberfläche von M. aeruginosa PCC 7806 lokalisiert sind und eine Bindestelle für das sekretierte Mvn darstellen. Mit Hilfe fluoreszenzmikroskopiebasierender Methoden wurde gezeigt, dass sowohl Mvn als auch das korrespondierende Mannanoligosaccharid stammspezifisch sind. Weiterhin konnte durch PCR gezeigt werden, dass das mvn-Gen in allen getesteten Microcystis-Stämmen vorkommt, die auch Gene für die Microcystinbiosynthese besitzen. Eine direkte Interaktion von Microcystin und Mvn konnte in vitro bestätigt werden. Microcystin bindet dabei über seinen N-Methyl-Dehydroalaninrest kovalent an die reduzierten Cysteinreste des Proteins. Ein Einfluss auf die Oligomerisierung des Proteins wurde festgestellt. Microcystin bindet an Cysteinreste von Proteinen, und es konnte gezeigt werden, dass dies besonders unter oxidativen Stressbedingungen geschieht. Die Daten liefern somit weitere Indizien für eine Rolle von Microcystin in der Stressadaptation.<br>Cyanobacteria frequently appear as so-called “water-blooms” during summer months. Cyanobacteria of the genus Microcystis, whose species often dominate freshwater lakes, produce toxins that represent a potential threat for humans and animals. The most prominent toxins are the non-ribosomally synthesised hepatotoxic microcystins. Toxicity has been considered the main function of these peptides, but recent studies propose different primary functions of microcystins for their producers. The involvement of microcystins in the response to oxidative stress was proposed recently. Within this study the putative lectin microvirin (Mvn), which was suggested to be functionally related to microcystin, was characterised. Initially it was shown that Mvn does indeed possess a carbohydrate binding activity, and specificity for mannan, an oligosaccharide made of mannose subunits, was proven. Binding studies using fluorescence-labelled Mvn and antibodies identified carbohydrates of this type at the cell surface of M. aeruginosa being a binding site for the secreted Mvn. Fluorescence microscopy techniques were employed to show that Mvn as well as the corresponding mannan oligosaccharide are strain-specific. Additionally it was shown by PCR that the mvn gene is present in all tested Microcystis strains possessing microcystin biosynthesis genes. A direct interaction of microcystin and Mvn was confirmed in vitro. Microcystin covalently binds to the reduced cysteine residues of the protein via its N-methyl-dehydroalanine moiety. An impact on the oligomerisation state of Mvn was observed. Microcystin seems to bind cysteine residues in an unspecific manner in vivo, and it was shown that this occurs especially under conditions of oxidative stress such as iron depletion and exposition to high light. Hence, the data provide further evidence for an involvement of microcystins in stress adaptation.
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Racine, Marianne. "Metabolic Variation in the Toxigenic Cyanobacterium Microcystis Aeruginosa." Thesis, Université d'Ottawa / University of Ottawa, 2018. http://hdl.handle.net/10393/37718.
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Cyanobacteria are notorious for their potential to produce toxins with human health effects, particularly the hepatotoxic microcystins (MCs), but cyanobacteria also produce other bioactive compounds. A wide variety of oligopeptides including aeruginosins, cyanopeptolins and cyanobactins may be as toxic as MCs. To investigate the production of these compounds, an UPLC QTOF-MS/MS method was developed to compare the metabolomic profiles of various strains of a common bloom-forming and toxigenic species, Microcystis aeruginosa, as well as those obtained from lakes with mixed cyanobacterial assemblages. Although many compounds could not be confirmed, MCs were rarely the dominant secondary metabolite in any sample. Since the biological role of MCs remains unknown, I tested the hypothesis that MCs provide protection against oxidative stress as induced through exposure to the herbicide atrazine and UV radiation in pure cultures of toxic vs non-toxic strains. Results were inconclusive and varied between strains suggesting other mechanisms exist to counter oxidative stress.
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Humpage, Andrew Raymond. "Tumour promotion by the cyanobacterial toxin microcystin /." Title page, contents and abstract only, 1997. http://web4.library.adelaide.edu.au/theses/09PH/09phh9258.pdf.
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Waack, Julia. "Uptake and depuration of cyanotoxins in the common blue mussel Mytilus edulis." Thesis, Robert Gordon University, 2017. http://hdl.handle.net/10059/2447.
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Cyanobacteria produce a variety of secondary metabolites which possess amongst others antifungal, antibacterial, and antiviral properties. Being primary producers they are also a vital component within the food web. However, certain strains also produce toxic metabolites such as the hepatotoxins microcystin (MC) and nodularin (NOD). Their toxicity in combination with the increasing global occurrence has resulted in a drinking water guideline limit of 1 μg L-1 being issued by the World Health Organisation (WHO). However, these toxins are not only present in water, but can be accumulated by fish and shellfish. Currently, no regulations regarding cyanotoxin contaminated seafood has been established despite similar toxicity to routinely monitored marine toxins such as domoic acid (DA). To facilitate regular monitoring, a high performance liquid chromatography photo diode array (HPLC-PDA) analysis method for the detection of DA was optimised to enable the simultaneous detection of DA and nine cyanotoxins. This method was then utilised to determine cyanotoxin concentration in laboratory cyanobacteria strains. To assess the accumulation and depuration of cyanotoxins in the common blue mussel Mytilus edulis, three feeding trials were performed. During these, mussels were exposed to two cyanobacteria strains, Nodularia spumigena KAC66, Microcystis aeruginosa PCC 7813, both individually and simultaneously. A rapid dose dependent accumulation of cyanotoxins was observed with maximum concentration of 3.4 -17 μg g-1 ww accumulated by M. edulis, which was followed by a much slower depuration observed. During the final feeding trial, with N. spumigena KAC 66 and M. aeruginosa PCC7813, cyanotoxins were still detectable following 27 days of depuration. Mortality in all studies was 7% or less indicating that most mussels were unaffected by the maximum dose of 480 μg L-1 NOD (feeding study 1), 390 μg L-1 MC (feeding study 2), or 130 μg L-1 total cyanotoxins (feeding trial 3), respectively. Mortality in negative control tanks was lower throughout all three feeding trials ( < 1 - 2.6%). Consumption of a typical portion size (20 mussels) would result in ingestion of cyanotoxins at levels significantly higher than the WHO recommended tolerable daily intake (TDI) of 2.4 μg NOD and/or MCs for a 60 kg adult. This value was exceeded not only during the exposure period (maximum levels 270 - 1370 μg cyanotoxins per 20 mussels), but also at the end of the depuration period 39-600 μg cyanotoxins per 20 mussels. These results illustrated that cyanotoxin monitoring of seafood should be considered not only during, but also following bloom events. In an attempt to investigate the cyanotoxin budget of the experimental system, not only mussels, but cyanobacteria cultures, the tank water, and the mussel faeces were also analysed for their cyanotoxin content. Results showed that large quantities of MCs and NOD were unaccounted for during all exposure trials. The combined effect of cyanotoxin metabolism in M. edulis, biotic and/or abiotic degradation, protein binding, and losses during the extraction and analysis were thought to have contributed to the unaccounted cyanotoxin fraction. Mussel flesh was analysed for the presence of glutathione or cysteine conjugates, however, there was no evidence of their occurrence in the samples tested. Due to these discrepancies in the toxin budget of the system, the introduction of correction factors for the analysis of cyanotoxins in M. edulis was suggested in order to protect the general public.
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Marshall, Randall S. "Environmental Isolations, Community Nutrient Ratio Effects, and Allelopathy of Microcystis from Grand Lake Saint Mary's." University of Cincinnati / OhioLINK, 2014. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1397735879.
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Davenport, Emily J. "Diel regulation of metabolic functions of a western Lake Erie Microcystis bloom informed by metatranscriptomic analysis." Bowling Green State University / OhioLINK, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=bgsu1479477147782031.
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Arif, Abdul Rahman Thaslim. "An investigation on the effects of cyanopeptides on the growth and secondary metabolite production of Microcystis aeruginosa PCC7806." Thesis, Robert Gordon University, 2016. http://hdl.handle.net/10059/1582.
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Cyanobacteria are one of the oldest forms of photosynthetic life and may have contributed significantly to the evolution of oxygen into the then anoxic environment. Cyanobacteria are also one of the best sources of natural secondary metabolites (cyanopeptides) some of which have harmful effects on the ecosystem, while others may be beneficial. It is known that these secondary metabolites are continuously produced during growth, however, it is not known whether the producing cyanobacteria actually benefit from these metabolites. The overarching aim of this study was to answer the question ‘Why do cyanobacteria produce secondary metabolites?’. With this aim in mind, preliminary work focused on understanding the growth and secondary metabolite production characteristics of Microcystis aeruginosa PCC7806. The technique of labelling secondary metabolites with 15N was successfully employed in differentiation and quantification of ex-novo and de-novo metabolites. The effect of exogenous cyanopeptides such as microcystins, aerucyclamides, anabaenopeptins, aeruginosamide, cyanopeptolin and aeruginosin on M. aeruginosa PCC7806 was evaluated using a rapid bioassay approach along with an automated cell enumeration technique. The results indicate that at least some cyanopeptides (microcystins-LR, microcystin-LF, aeruginosamide, anabaenopeptin B and aerucyclamide A) induce significant changes to cell division and metabolite production rate. In an ecological scenario, the release of such secondary metabolites by lysing cells (such as when blooms collapse), may be perceived as an alarm signal by surrounding live cells, which may in turn slow cell division and prepare for re-invasion. This may be a strategy for species survival and dominance. While the results from this study do not confirm a role for cyanopeptides, it is thought that the results are clearly indicative of the role played by cyanopeptides for the producing organism. In order to confirm a role, it is recommended that monitoring ribosomally synthesised metabolites (e.g. aerucyclamides) along with chlorophyll-a gene expression, with sophisticated techniques such as qPCR are used.
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Borges, Renata Maria Cortez. "Identificação e quantificação de microcistinas por HPLC em reservatórios de água." Universidade de São Paulo, 2008. http://www.teses.usp.br/teses/disponiveis/97/97131/tde-24102012-125936/.
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A oferta de água vem se tornando cada vez mais diminuta à medida que a população, a indústria e a agricultura se expandem. A contaminação dos mananciais gerada pelo descarte de efluentes domésticos e industriais leva a eutrofização, processo pela qual grande aporte de nutrientes, particularmente fosfatos, leva ao crescimento excessivo de algas. As cianobactérias são microrganismos procariontes, que vivem nos ambientes mais adversos. A floração dessas algas quando presente em mananciais destinados ao consumo humano gera sérios problemas à saúde humana, pois algumas dessas algas podem gerar toxinas, conhecidas como hepatotoxinas, neurotoxinas e dermatotoxinas, de acordo com sua ação farmacológica. Dentre as hepatotoxinas encontramos a microcistina, um heptapeptídeo cíclico que pode levar à morte em horas ou dias. O objetivo desse estudo foi viabilizar a técnica HPLC, já proposta por outros autores, para quantificar microcistinas-LR em reservatórios de água, em escala empresarial para ser implementada em laboratórios de análise de águas. Para o desenvolvimento da técnica, foram utilizadas amostras de uma lagoa com floração de Microcystis. Para determinar a eficiência da técnica cromatográfica, foram realizados estudos com outro método, através do kit ELISA. Nessa etapa do trabalho, verificou-se que a técnica HPLC é mais sensível e viável para a quantificação das microcistinas. Nos experimentos realizados com a cromatografia líquida, observou-se que a coluna C-18 LiChrosorb (25 cm) 7 Sm utilizada no método, e o solvente metanol apresentaram grande influência nos resultados. À medida que os experimentos foram executados, verificou-se o decréscimo da sensibilidade da coluna. Os resultados foram satisfatórios apenas após a limpeza realizada na coluna, onde os padrões apresentaram uma curva de correlação igual a 0,92. Este fato leva a concluir que as colunas precisam ser renovadas para análises mais sensíveis, como no caso das microcistinas. As amostras extraídas com metanol apresentaram resultados relevantes, isto é, quanto maior a concentração de metanol utilizado na extração, maior a concentração de microcistina-LR obtida nos resultados, concluindo o metanol ser o solvente apropriado para a extração. Por fim, conclui-se que o método desenvolvido é viável, apresentando algumas dificuldades para sua implantação em escala empresarial.<br>The water supply has been decreasing more and more as the population, industry and agriculture expand. The contamination of the water sources generated by the domestic and industrial effluents discharges leads to the eutrophization, process where the large presence of nutrients, particularly phosphates, causes excessive increase of algae. The cyanobacteria are procaryote microorganisms which live in the most diverse environments. The florescence of the algae when present in sources directed to human consumption generates serious problems to the human health, for some of them may produce toxins known as hepatotoxins, neurotoxins and dermotoxins, according to their pharmacological action. Among the hepatotoxins, the microcystin, a cyclic heptapeptide that can lead to death in hours or days, is found. The objective of this study was to make feasible the use of the HPLC technique, already proposed by other authors, to quantify microcystins-LR in water reservoirs, in enterprise scale to be implemented in water analysis laboratories. In order to develop the technique, samples of water from a pond with Microcystis florescence were utilized. To evaluate the efficiency of the chromatographic technique, studies were performed with another method, through the ELISA kit. In this phase of the work, it was verified that the HPLC technique is the most sensible and viable for the quantification of the microcystins. It was observed, in the experiments performed with the liquid chromatography, that the column C-18 LiChrosorb (25 cm) 7 Sm utilized in the method and the methanol solvent presented great influence in the results. As the experiments were realized, the decrease of the sensibility of the column was verified. The results were satisfactory only after the column being cleaned, when the patterns presented a curve of correlation equal to 0.92. This fact leads to the conclusion that the columns need to be renewed for more sensible analysis, like in the case of the microcystins. The samples extracted with methanol presented relevant results, that is, the greater the concentration of the methanol utilized, the higher the concentration of microcystin-LR obtained in the results, leading to the conclusion that methanol was the solvent adequate to the extraction. Finally, it was concluded that the method developed is feasible, presenting some difficulties concerning its implantation in enterprise scale.
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Root, Hannah Patricia Biotechnology & Biomolecular Sciences Faculty of Science UNSW. "Transcription regulation of hepatotoxins microcystin and nodularin from cyanobacteria." Publisher:University of New South Wales. Biotechnology & Biomolecular Sciences, 2008. http://handle.unsw.edu.au/1959.4/43351.
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The role and function of hepatotoxins microcystin and nodularin produced by M.aeruginosa PCC 7806 and N. spumigena NSORlO respectively have yet to be elucidated. The mode of transcriptional regulation of these toxins, incorporating DNA binding proteins, was investigated, as an attempt to further understand the key control mechanisms acting on the toxins. The DNA binding proteins that control nitrogen and iron responsive transcription, NtcA and Fur, were identified from M. aeruginosa PCC7806 and N. spumigena NSOR10. Cloning and over-expression in E. coli was followed by mobility shift assays to determine binding characteristics of NtcA and Fur to the promoters, mcyA/D and ndaA/C, those regions that control the toxin encoding gene clusters in M. aeruginosa PCC 7806 and N. spumigena NSOR10, respectively. The results from these studies suggested a role for iron and nitrogen in the transcriptional control of microcystin and nodularin. biosynthesis. As NtcA and Fur classically act to regulate nitrogen and iron dependent genes, a link may be made to the putative function and control of microcystin and nodularin. By identifying the transcription factors NtcA and Fur in these genera, a greater understanding of the link between nutrient levels in the environment and hepatotoxin production in cyanobacteria may be possible.
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Cross, David Michael. "Analytical methods for cyanobacterial toxins." Thesis, University of Bath, 1997. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.390310.
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Reitz, Laura A. "Quantification of Microcystin Production and Loss Rates for the Spatiotemporal Distribution of Microcystis aeruginosa Blooms in Lake Erie." Bowling Green State University / OhioLINK, 2020. http://rave.ohiolink.edu/etdc/view?acc_num=bgsu1594907425606966.
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Campbell, Dennis Leslie. "Laboratory and field investigations into the cyanobacterial hepatotoxin, microcystin-LR." Thesis, University of Dundee, 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.388382.
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Huang, Yuzhou. "Evaluating the Removal of Microcystin Variants with Powdered Activated Carbon." The Ohio State University, 2020. http://rave.ohiolink.edu/etdc/view?acc_num=osu1593536125422238.
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Graham, Jennifer L. "Environmental factors influencing microcystin distribution and concentration in Midwestern lakes /." free to MU campus, to others for purchase, 2004. http://wwwlib.umi.com/cr/mo/fullcit?p3137703.
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Downs, Kerry. "The effect of selenium in the detoxification of the microcystin hepatotoxins." Thesis, University of Port Elizabeth, 2002. http://hdl.handle.net/10948/284.
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Blooms of cyanobacteria have been known to cause illness in humans and death in wild and domestic animals. One of the toxins produced by cyanobacteria is microcystin, which is a potent hepatotoxin. Microcystin is taken up by bile acid transporters in the intestine and transported into the liver. After exposure to acute doses of microcystin, severe haemorrhage has been observed along with apoptotic and necrotic hepatocytes. The cytoskeletal structure of the hepatocytes is disrupted and oxidative stress is induced. Selenium, a known anti-oxidant, has been shown to induce increased activity of glutathione peroxidase. Glutathione peroxidase removes peroxides from cells protecting them from oxidative stress. This study set out to determine if selenium could play a role in preventing the damage to mice livers due to microcystin toxin. The protective role of selenium was explored in three main studies: in the first study, the ability of selenium to increase the survival time of mice exposed to a lethal dose of toxin was determined. In the second study the mice were exposed to sublethal chronic doses of toxin over 30 days. The ability of selenium to minimise liver damage under these conditions was determined. The final study investigated the mechanism of the protective effect of selenium. The results of the first study suggested that selenium could extend survival time. In the second study the selenium supplemented mice showed a reduction in the extent of the increase in liver weight and a decrease in the amount of lipid peroxidation induced compared to the mice that received only toxin. The histology of the selenium supplemented mice also showed a decrease in the severity and amount of morphological changes in the liver. The third study indicated that the protection shown by selenium might be mediated by an increase in the glutathione peroxidase (GPX) activity in selenium supplemented mice. This increase in GPX activity would increase the removal of the lipid hydroperoxides and prevent the damage they would cause in the cell. A further result indicated an increase in glutathione S-transferase in only the toxin control mice when compared to the selenium supplemented and control mice. ii In conclusion selenium offers protection against microcystin but further studies need to be done to provide statistically valid results to clarify the level of protection.
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Zastepa, Arthur. "Fate and Persistence of Microcystin Congeners in Lakes and Lake Sediments." Thesis, Université d'Ottawa / University of Ottawa, 2014. http://hdl.handle.net/10393/30453.
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Cyanobacterial blooms and their toxins are a major water quality and potential health risk around the world. This thesis developed an analytical method for microcystin congeners in sediments in order to examine their fate in lakes and establish the history of toxin-producing cyanobacteria in relation to environmental change using lake sediments. A novel method for both intra- and extracellular microcystins in lake sediments was developed, consisting of accelerated solvent extraction, hydrophilic-lipophilic balance solid phase extraction and multiple reaction monitoring-based HPLC-MS/MS quantitation. The method achieved comparable recoveries of intra- and extracellular cyanotoxins based on nine microcystins and nodularin (marine analogue). The analytical method was validated using surficial and deeper sediments from seven lakes of diverse geography and trophic state. To study the fate of microcystins, a multi-year, whole lake study of Microcystis blooms was conducted to obtain both in situ and in vitro half-life estimates of microcystin-LA (MC-LA), an understudied, but increasingly reported microcystin. MC-LA appeared to undergo slower rates of decomposition and persist longer than the more frequently studied MC-LR. Experimentally, high light intensity increased in vitro decomposition of dissolved MC-LA while high temperature enhanced decomposition in the particulate phase. Sediment deposition measurements and estimates of sediment-pore water distribution coefficients, sediment accumulation rates, and diffusive fluxes indicated that microcystin congeners differ in their fate. Notably, MC-LA preferentially distributed into pore water and remobilized (by diffusion) from sediments and into overlying water while MC-RR adsorbed more strongly to sediment particles. Finally, the sediment record of an eutrophic lake of major recreational importance was examined to identify possible drivers of toxigenic cyanobacteria and determine if the perceived increase in toxigenic cyanobacteria could be corroborated. Microcystins were detected to the bottom of the core (early 1800s), indicating that toxigenic cyanobacteria were present prior to the first permanent settlements. Microcystins were significantly correlated with changes in diatom-inferred nutrients (DI-TP and DI-TKN) within the sediment core as well as with specific algal pigments. Sediment microcystins in the upper layers also significantly correlated with a 20-year monitoring record for water column microcystins suggesting that sediment microcystins can be used as a proxy for past surface water conditions.
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Su, Robin. "Microcystin-LR (MC-LR) Toxicity In The Gut-Liver Signaling Axis." University of Toledo Health Science Campus / OhioLINK, 2020. http://rave.ohiolink.edu/etdc/view?acc_num=mco1588673649090272.
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Villars, Kathryn E. Villars. "Removal of Microcystin-LR from Drinking Water Using Granular Activated Carbon." The Ohio State University, 2018. http://rave.ohiolink.edu/etdc/view?acc_num=osu1532007603377473.
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Cornish, Benjamin J. P. A. "The destruction of the cyanobacterial toxin microcystin-LR by semiconductor photocatalysis." Thesis, Robert Gordon University, 2000. http://hdl.handle.net/10059/3094.
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In fresh waters where cyanobacteria (blue-green algae) flourish, dense growths known as blooms occur. Such blooms present a threat to human and animal health as many of these cyanobacteria produce toxins. One such group of toxins are the microcystins which are hepatotoxic resulting in haemoraging and tumour promotion in the liver. There have been several reports of human poisonings resulting from the presence of cyanotoxins in potable waters, some of which have resulted in fatalities. The most frequently cited cyanotoxin in these poisonings has been microcystin-LR, which has prompted the World Health Organisation (WHO) to set a guideline for the recommended safe level of this toxin in drinking water of 1 mgl-1. Removal of microcystin-LR from potable waters has proven to be inefficient using conventional water treatment techniques such as coagualtion, filtration and chemical oxidation using chlorine. While activated carbon adsorption and membrane filtration have been shown to physically remove microcystin-LR from water the toxin is not destroyed. Recently the use of photocatalysis was shown to rapidly degrade microcystin-LR even at high concentrations. The process involves the illumination of a titanium dioxide catalyst with ultraviolet (UV) light to produce highly oxidising hydroxyl radicals in solution. While several researchers have demonstrated the process's effectiveness in degrading the toxin none have determined the fate of the compound, or if the toxicity related to microcystin-LR has been removed. This study was carried out to determine if photocatalytic oxidation of microcystin-LR was suitable as a treatment method for potable water supplies. Analysis of treated toxin samples by high performance liquid chromatography (HPLC) with photo-diode array detection (PDA) and mass spectroscopy established that the toxin was not completely degraded during photocatalysis. A simple toxicity assessment however indicated that by-products were non-toxic. Using the data from this work a proposed pathway for toxin destruction was produced giving the speculative identity of some of the by-products. The use of hydrogen peroxide to enhance UV mediated destruction of microcystin-LR has been previously reported. There have also been reports of the enhancement of photocatalytiC reaction in the presence of this oxidant. The work carried out in this study demonstrated that the destruction of microcystin-LR by photocatalysis was both more rapid and more efficient when hydrogen peroxide was present in the system. The use of a fixed film flow reactor was also investigated for microcystin-LR destruction. While degradation of the toxin occurred it was demonstrated that batch reactors were more efficient as a treatment method. The effectiveness of the photocatalytic process on microcystin-RR, -LW and -LF was also investigated. While destruction of a" the variants occurred during photocatalytic treatment each microcystin demonstrated different rates and efficiencies of photooxidation. It was concluded from this study that photocatalysis is a promising treatment method for the removal of microcystin-LR and other variants from potable waters. Further research however is required to assess if the tumour promoting effects of microcystin-LR are rendered inactive and to determine the behaviour of the toxins degradation in natural water supplies. The study also allowed for speculation as to how the degradation of the toxin occurred during the photocatalytic process.
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Jagani, Neelam V. "Evaluating Home Point-of-Use Reverse Osmosis Membrane Systems for Removal of Cyanotoxins." University of Toledo / OhioLINK, 2018. http://rave.ohiolink.edu/etdc/view?acc_num=toledo1524844338053604.
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Zheng, Bingxue. "Quantitative Analysis and Determination of Microcystin in water by Capillary Electrophoresis Mass Spectrometry." FIU Digital Commons, 2014. http://digitalcommons.fiu.edu/etd/1538.
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The presence of harmful algal blooms (HAB) is a growing concern in aquatic environments. Among HAB organisms, cyanobacteria are of special concern because they have been reported worldwide to cause environmental and human health problem through contamination of drinking water. Although several analytical approaches have been applied to monitoring cyanobacteria toxins, conventional methods are costly and time-consuming so that analyses take weeks for field sampling and subsequent lab analysis. Capillary electrophoresis (CE) becomes a particularly suitable analytical separation method that can couple very small samples and rapid separations to a wide range of selective and sensitive detection techniques. This paper demonstrates a method for rapid separation and identification of four microcystin variants commonly found in aquatic environments. CE coupled to UV and electrospray ionization time-of-flight mass spectrometry (ESI-TOF) procedures were developed. All four analytes were separated within 6 minutes. The ESI-TOF experiment provides accurate molecular information, which further identifies analytes.
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Lee, Jung Ju. "Removal of Microcystin-LR from Drinking Water Using Adsorption and Membrane Processes." The Ohio State University, 2009. http://rave.ohiolink.edu/etdc/view?acc_num=osu1229026536.
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DeMarco, Jonathan R. "Cyanobacterial Blooms in Chautauqua Lake, NY: Nutrient Sources and Toxin Analyses." Bowling Green State University / OhioLINK, 2021. http://rave.ohiolink.edu/etdc/view?acc_num=bgsu1625052848648708.
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Ross, Catherine M. "The Feasibility of Applying an Industrial Hygiene Sampling Method to Measure Airborne Microcystin." University of Toledo / OhioLINK, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=toledo1513365627637454.
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Haase, Maxwell D. "Effects of the Algal Toxin Microcystin on Fishes in the James River, Virginia." VCU Scholars Compass, 2015. http://scholarscompass.vcu.edu/etd/3941.
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With the global rise in frequency of harmful algal blooms in estuarine environments comes an increase in prevalence of toxic metabolites, such as microcystin (MC), that some of the cyanobacteria involved will produce. At high concentrations, MC may accumulate in consumer tissues and have deleterious effects on organisms; however impacts of the toxin on aquatic living resources at ecologically relevant concentrations have not been widely documented. We analyzed the effects of MC on juveniles of five fish species from the James River, Virginia to determine if MC has the potential to impede growth. Using three separate experimental approaches, it was shown that exposure to concentrations of the toxin currently observed in the James River estuary do not appear to significantly impact the growth or survivorship of tested fish species. Extraneous factors in parts of the study led to an inability to draw clear conclusions on mortality or growth impacts; however it is evident from the experiments that at least some of the fish species have biological mechanisms in place that allow them to effectively eliminate the toxin from their systems. An ability to extricate the toxin suggests the possibility for fishes to withstand MC exposures and sustain few negative health impacts at low MC concentrations.
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Alambo, Katherine I. "Cyanobacteria North of 60°: Environmental DNA Approaches." Thesis, Université d'Ottawa / University of Ottawa, 2017. http://hdl.handle.net/10393/35665.
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Cyanobacterial blooms, such as those recently reported in Great Slave Lake (GSL, NWT), have sparked concern over the occurrence of toxic blooms in the North. This study investigated past and present incidences of cyanobacteria in lakes above latitude 60° N. The abundance of the toxin (microcystin) gene mcyE, as well as genes common to all cyanobacteria (16S rRNA) and bacteria (glnA) were quantified from lake sediment cores using ddPCR. Individual colony isolates from a surface bloom in Yellowknife Bay (GSL) in August 2015 were amplified and identified as non-toxigenic Dolichospermum lemmermannii. Very low levels of microcystin genes were detected through the sediment archives (over ~100-150 yr) of GSL and other lakes, as well as in the plankton of GSL. While recent increases in mcyE were not observed, an increase in the cyanobacterial 16S rRNA and glnA genes was seen through time. In the high Arctic Meretta Lake, gene abundance profiles reflected the effects of past eutrophication and recovery.
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Bury, Nicolas R. "The effects of cyanobacteria on fish." Thesis, University of Dundee, 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.367494.
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Ji, Bo. "Effects of environmental factors on the growth and microcystins production of Microcystis aeruginosa." HKBU Institutional Repository, 2006. http://repository.hkbu.edu.hk/etd_ra/670.
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Halvorson, Rebecca Ann. "Raman Spectroscopy for Monitoring of Microcystins in Water." Thesis, Virginia Tech, 2010. http://hdl.handle.net/10919/76924.
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Cyanobacterial blooms are of great concern to the drinking water treatment industry due to their capacity to produce microcystins and other cyanotoxins that are deadly to humans, livestock, pets, and aquatic life at low doses. Unfortunately, the strategies currently employed for cyanotoxin detection involve laborious analyses requiring significant expertise or bioassay kits that are subject to numerous false positives and negatives. These methods are incapable of providing rapid, inexpensive, and robust information to differentiate between the >80 cyanotoxin variants potentially present in an aqueous sample.
The use of Raman spectroscopy for identification and quantification of the ubiquitous cyanotoxin microcystin-LR (MC-LR) was examined. Raman spectra readily reflect minute changes in molecular structure, spectra can be collected through water or glass, portable Raman spectrometers are increasingly available, and through surface enhanced Raman spectroscopy (SERS) it is possible to achieve femto or picomolar detection limits for a variety of target species. Drop coating deposition Raman (DCDR) was successfully implemented for quantitation of 2-100 ng of MC-LR deposited in 2 ?L of aqueous sample, even without the use of a specifically designed DCDR substrate or Raman signal enhancements. Reproducible MC-LR Raman spectra were observed for both fresh and aged DCDR samples, and the MC-LR Raman spectrum remained identifiable through a matrix of >80% DOM by mass. DCDR methods show tremendous potential for the rapid, simple, and economical detection of cyanotoxins in environmental matricies at environmentally relevant concentrations.<br>Master of Science
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Júlio, Maria de Fátima de Jesus Leal. "Carbon key-properties for microcystin adsorption in drinking water treatment: structure or surface chemistry?" Master's thesis, Faculdade de Ciências e Tecnologia, 2011. http://hdl.handle.net/10362/7066.
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Dissertação para Obtenção de Grau de Mestre em Engenharia Química e Bioquímica<br>The carbon key-properties (structure and surface chemistry) for microcystin-LR (MC-LR) adsorption onto activated carbon were investigated. Waters with an inorganic background matrix approaching that of the soft natural water (2.5 mM ionic strength) were used. Also, model waters with controlled ionic make-up and NOM surrogate with similar size of MC-LR (tannic acid - TA) with MC-LR extracts were tested with activated carbon NORIT 0.8 SUPRA. For this AC, two particle sizes, 125-180 μm and 63-90 μm were tested. The surface chemistry of NOR 125-180 μm was modified by thermal treatment and was also preloaded with TA. The integrated analysis of carbon’s chemical and textural characterization and of kinetic and isotherm modeling using non-linear models allowed concluding that: i) the heating method is an efficient and simple process for reducing a relatively hydrophilic activated carbon and thereby enhancing its MC-LR adsorption capacity; ii) from a combination of the modification of the carbon surface chemistry and the carbon structure, it is demonstrates that both properties play an important role in the adsorption process, although carbon surface chemistry seems to be more important than its porous structure – MC-LR adsorption correlated with meso and macroporous volume and particularly well with carbon hydrophobicity (inverse of oxygen content); iii) the smaller the particle size, the more important is external mass transfer over intraparticle diffusion; iv) similar sized NOM strongly competes with MC-LR for the same AC sites; v) direct competition governs the simultaneous MC-LR and NOM adsorption; vi) the preloading phenomena reduces significantly the performance of activated carbon adsorption.
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Thees, Alison. "Identification and Characterization of Lake Erie Bacteria that Degrade the Microcystin Toxin MC-LR." University of Toledo Health Science Campus / OhioLINK, 2018. http://rave.ohiolink.edu/etdc/view?acc_num=mco15254429474901.
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Ihle, Tilo. "Die raum-zeitliche Variation von Microcystis spp. (Cyanophyceae) und Microcystinen in der Talsperre Quitzdorf (Sachsen)." Doctoral thesis, Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2008. http://nbn-resolving.de/urn:nbn:de:bsz:14-ds-1214480543195-41264.
Full textAbstract:
Cyanobakterien bilden zahlreiche bioaktive Substanzen mit zum Teil humantoxischer Relevanz. Nicht selten spielen dabei zyklische Peptide, zu denen unter anderem die Microcystine (MCYST) gehören, eine Schlüsselrolle. MCYST werden u.a. von Microcystis KÜTZING EX LEMMERMANN 1907 gebildet. Erkenntnisse zur ökophysiologischen Funktion der MCYST, die zweifelsfrei bei den Produzenten selbst zu suchen ist, liegen bisher kaum vor. Mit Hilfe von Freilanduntersuchungen sollten im Rahmen der vorliegenden Arbeit Kenntnisse zu einer möglichen ökologischen Funktion der MCYST erweitert und vertieft werden. Grundlage stellte dabei die Phänologie von Microcystis als einer der bedeutendsten limnischen MCYST-Produzenten dar. Microcystis zeigt im Freiland einen charakteristischen annuellen Lebenszyklus mit benthisch-pelagischer Kopplung. Ziel der vorliegenden Arbeit war es, die phänologischen Phasen des Lebenszyklus von Microcystis im Freiland zu differenzieren sowie die Dynamik der MCYST während dieser Phasen kompartimentübergreifend gesamtheitlich zu erfassen. Über eine MCYST-Massenbilanzierung sollen anschließend die dem annuellen Zyklus zugrundeliegenden Teilprozesse quantifiziert und zusammengeführt werden. Vordergründiges Anliegen war es, Phasen einzugrenzen, bei denen MCYST möglicherweise eine ökophysiologische Funktion haben könnte. Der annuelle Lebenszyklus von Microcystis wurde anhand von Biomasseänderungen am Sediment und im Pelagial der TS Quitzdorf in die phänologischen Phasen Überwinterung, Reinvasion, pelagisches Wachstum und Sedimentation unterteilt: Intakte, im Herbst aus dem Freiwasser aussedimentierte, Microcystis-Kolonien überwintern am Sediment und steigen im Frühjahr und Frühsommer zurück ins Freiwasser auf. Dort erfolgt der Wachstumsprozess, dem sich im darauffolgenden Herbst erneut ein Zusammenbruch und die Sedimentation der Freiwassergemeinschaft anschließt. Die benthisch-pelagische Kopplung wirkt dabei als interannuelles Bindeglied. Zwischen dem annuellen Lebenszyklus von Microcystis und der MCYST-Dynamik wurde eine enge Bindung nachgewiesen: Änderungen der absoluten MCYST-Konzentrationen während der Übergangsphasen Aufstieg (Frühjahr) und Sedimentation (Herbst) zeigen, dass MCYST mit den aufsteigenden bzw. aussedimentierenden Microcystis-Kolonien aus dem bzw. in das Sediment ‚transportiert’ werden. Ausschließlich während der pelagischen Phase, die sich dem Reinvasionsprozess anschließt, kommt es in Abhängigkeit vom Wachstum der Produzenten und deren Sukzession zur Neubildung von MCYST. Während den Wintermonaten wurden MCYST am Sediment intrazellulär ‚konserviert’. Der Verlauf der pelagischen MCYST-Konzentration wurde mit Hilfe eines Wachstumsmodells nachgebildet. In dieses Modell wurde die genetische Variabilität der MCYST-Produzenten sowie eine mögliche physiologische Steuerung der MCYST-Synthese über die Verfügbarkeit des anorganischen Kohlenstoffs integriert. Der prinzipielle Verlauf zeigte dabei weitestgehend Koinzidenz zwischen den real gemessenen und den simulierten MCYST-Konzentrationswerten. Abweichungen zwischen beiden konnten mit Hilfe des gesamtheitlich kompartimentübergreifenden MCYST-Bilanzierungsansatzes – in erster Linie über benthisch-pelagische Kopplungsprozesse – plausibel erklärt werden. Der Habitatwechsel ist für Microcystis prinzipiell mit Verlusten (Seneszenz/Lyse oder möglicherweise Apoptose) verbunden, sowohl für MCYST-Produzenten und Nichtproduzenten. Die auffallende Stabilität der benthischen MCYST-Zellquote während der Überwinterung gibt Grund zur Annahme, dass eine Funktion von MCYST am/im Sediment eher unwahrscheinlich ist. Da MCYST über derart lange Zeiträume am Sediment intrazellulär ‚konserviert’ werden, ist eine Bedeutung der MCYST während der Reinvasionsphase und in der frühen pelagischen Phase nicht auszuschließen. Im Speziellen wurde eine mögliche ökologische Funktion von MCYST in Zusammenhang mit der Variation der Koloniegröße bzw. dem epiphytischen Bewuchs von Microcystis-Kolonien mit Pseudanabaena mucicola geprüft: Aus dem Zusammenhang zwischen extra-/intrazellulärer MCYST-Konzentration und der Microcystis-Koloniegrößenverteilung waren keine konsistenten Schlussfolgerungen abzuleiten, welche auf eine Steuerung der Koloniebildung durch MCYST deuten. Vor dem Hintergrund, dass MCYST keinen nachweislich allelopathischen Effekt auf den Epibionten Pseudanabaena mucicola ausüben, wurde postuliert, dass zwischen dem beobachteten epiphytischen Besiedlungs-/Verteilungsmuster und der MCYST-Produktion ein indirekter Zusammenhang besteht, welcher die zeitweise Einnischung von Pseudanabaena mucicola auf Microcystis-Kolonien ermöglicht. Die Ergebnisse der vorliegenden Untersuchung lassen weder unmittelbar noch mittelbar eine Variabilität der ökophysiologischen Bedeutung von MCYST, die im Zusammenhang mit der raum-zeitlichen Verteilung potentieller Produzenten steht, erkennen. Eine divergierende Funktion der MCYST auf intra- bzw. extrazellulärer Ebene kann nicht zwingend ausgeschlossen werden. Die Mehrzahl der aus der MCYST-Phänologie und MCYST-Bilanzierung abzuleitenden Schlussfolgerungen deutet allerdings eher auf eine Funktion auf (intra-)zellulärer Ebene hin, wie etwa die Effizienzsteigerung des Kohlenstoffmetabolismus (d.h. der intrazellulä-ren Akkumulation anorganischen Kohlenstoffs) während der pelagischen (Wachstums-)Phase der Produzenten.
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Bortoli, Stella de. "Investigação da biossíntese de toxinas produzidas por cepas de cianobactérias." Universidade de São Paulo, 2011. http://www.teses.usp.br/teses/disponiveis/9/9141/tde-29092011-164054/.
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A demanda crescente de água doce de boa qualidade são problemas atuais e mundiais, além do descaso com os dejetos lançados nos ambientes aquáticos que comprometem a qualidade dos recursos hídricos. Um dos parâmetros que atesta a potabilidade da água é a presença de cianobactérias e cianotoxinas. Cianobactérias são microrganismos procariontes aeróbicos fotoautróficos que sintetizam as cianotoxinas. Estes compostos podem ser classificados de acordo com seus mecanismos de ação em hepatotóxicos, neurotóxicos e dermatotóxicos. Por sua diversidade, representam diferentes riscos não só ao ecossistema e a outros organismos dos ambientes aquáticos, como também aos seres humanos. Esse projeto visou o isolamento e cultivo de cepas de cianobactérias produtoras de toxinas para a investigação da biossíntese desses compostos. Com este intuito, foram realizadas coletas de água em três reservatórios no estado de São Paulo e um no Paraná. Cepas de cianobactérais foram isoladas, identificadas e analisadas quanto à produção de toxinas. Uma cepa de Microcystis aeruginosa (LTPNA 02) produtora de microcistinas (MC-LR, MC-RR, MC-YR, MC-LF, MC-LW e desm-MC-LR e desm- MC-RR) foi escolhida para ser estudada frente diferentes condições de cultivo e ter o seu crescimento, produção de toxinas e expressão gênica estudados. Foram utilizados os meios de cultura já referidos na literatura: ASM-1 (N:P=1, 10 e 20), MLA (N:P=10), Bold 3N (N:P=16) e BG-11 (N:P=10 e 100). Para acompanhar o crescimento, dois métodos foram utilizados: contagem de células e espectrofotometria. As toxinas foram quantificadas por LC-MS - QTrap. A análise da expressão gênica foi realizada por reação de PCR em tempo real pelo método de quantificação relativa ΔΔCt. Foi observada diferença no crescimento da cepa estudada nos diferentes meios de cultivo empregados. A contagem das células permitiu a identificação das fases logarítmica e total de crescimento. Durante a fase logarítmica, três experimentos demonstraram diferenças estatísticas quando comparadas ao controle (p<0,05). Ao se avaliar o crescimento total, quatro experimentos foram menores (p<0,01). As leituras das absorvâncias e a contagem de células demonstraram alta correlação Para ambas as leituras em 680 nm e 750 nm o coeficiente de correlação (r) esteve entre 0,93 e 0,99. A quantificação das microcistinas (MC) foi realizada por LC-MS - QTrap. Foram quantificadas as variantes MC-LR, MR-RR e MC-YR. Apesar da relação toxina/célula ser distinto para cada experimento, não representou grande variação naqueles realizados com meio ASM-1 (N:P 1; 10 e 20), meio MLA (N:P=10) e BG11(N:P=10). O experimento realizado em Bold3N (N:P=16,6) apresentou menor concentração de toxina/célula e as variantes MC-LR e MC-YR não foram detectadas. Por outro lado, o experimento realizado em BG-11 (N:P=100) apresentou a maior relação toxina por célula. Estes resultados sugerem que o excesso de nitrato seja um fator estressante para o desenvolvimento e crescimento da cepa de M. aeruginosa avaliada e ao mesmo tempo um fator estimulante para a produção das toxinas analisadas. Os experimentos que avaliaram a expressão dos genes 16S e mcyB em relação ao gene da ficocianina (controle endógeno) foram realizados em meio ASM-1 (N:P=10 e 100) e BG 11 (N:P= 10 e 100). Os parâmetros anteriores, como crescimento e produção de toxinas também foram avaliados. Novamente foram encontradas diferenças entre as fases de crescimento e produção de toxina, porém a expressão dos genes avaliados não demonstrou variação significativa entre os experimentos. Porém ambos os genes avaliados demonstraram menor expressão nos experimentos condizidos em (N:P=100).<br>There is a great concern these days about potable and good quality water due to the increase of the population needs and also to the arising problems with contamination caused by anthropogenic sources. The presence of cyanobacteria and cyanotoxins are some parameters that attest water potability. Cyanobacteria are prokaryotic aerobic photoautotrophic microorganisms that may synthesize cyanotoxins. These compounds can be classified as hepatotoxic, neurotoxic and dermatotoxic according to their action mechanisms. Because of their diversity, they may represent different risks, not only to their ecosystem and other aquatic living organisms, but also to human beings. The aim of this project was the isolation and cultivation of cyanotoxin-producing cyanobacteria for further investigation on the biosynthesis of these compounds. Water samples from three different reservoirs in São Paulo state and one in Paraná state were collected in order to isolate cyanobacteria strains and accomplish their identification and to evaluate the toxin production. The Microcystis aeruginosa (LTPNA 02) microcystin producer strain (MCLR, MC-RR, MC-YR, MC-LF, MC-LW, desm-MC-LR and desm-MC-RR) was chosen to be grown in different cultivation conditions and later analyzed for its growth rate, toxin production and gene expression. All culture media used in this research were chosen according to the literature: ASM-1 (N:P=1, 10 and 20), MLA (N:P=10), Bold 3N (N:P=16) and BG-11 (N:P=10 and 100). To evaluate growth rate, two techniques were used: cell counting and absorbance determination in two different wavelengths (680 nm and 750 nm). Toxins were quantified by LC-MS in a hybrid triple-quadrupole instrument (Qtrap). Gene expression was assessed by real time PCR, using the ΔΔCt relative quantification method. Cell counting allowed total growth and logarithmic phase identification. During the last, three experiments showed statistical difference from control group (p<0,05). Four experiments resulted in a lower total growth rate (p<0,05). A high correlation between cell counting and absorbance levels was found for both wavelengths tested. Correlation coefficients (r) were from 0,93 to 0,99. Three microcystin variants (MC-LR, MR-RR e MC-YR) were quantified by LC-MS. The toxin content per cell was calculated and showed no statistc variation among those experiments performed on ASM-1 (N:P 1; 10 and 20), MLA (N:P=10) and BG-11 (N:P=10). The lowest toxin/cell concentration was found for Bold3N (N:P=16,6) medium, where MC-LR and MC-YR production was not detected. On the other hand, the experiment with BG-11 (N:P=100) medium showed the highest toxin/cell content. These results suggest that high levels of nitrate in the culture medium may be a stressing factor for the development and growth of the M. aeruginosa tested strain, as well as a disturbing factor for microcystin production. Gene expression experiments regarding 16S and mycB genes using the phycocyanin gene as endogen control were performed on ASM-1 (N:P=10 and 100) and BG 11 (N:P= 10 and 100) media, along with the evaluation of growth rate and toxin production. Differences between growth rates and toxin production were once more observed, however gene expression did not show a significant variation among experiments.
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Krishnan, Anjali. "ASSESSMENT OF BIODEGRADATION AND TOXICOLOGICAL EFFECTS OF MICROCYSTINS." Kent State University / OhioLINK, 2019. http://rave.ohiolink.edu/etdc/view?acc_num=kent1560600592119155.
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Stanic, Dina. "Characterization of Oscillatoria spp. and their Role in Black Band Disease of Coral." FIU Digital Commons, 2010. http://digitalcommons.fiu.edu/etd/243.
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Black band disease (BBD) is a cyanobacterial dominated pathogenic consortium that affects corals worldwide. Recently two cyanobacteria (Oscillatoria strains 101-1 and 100-1) were isolated into culture from BBD. The aim of this study was to characterize the strains and assess their role in BBD pathogenesis. Light, scanning electron and transmission electron microscopy, coupled with 16S rRNA gene sequencing, were used for taxonomic characterization. Cyanotoxin production was assessed by enzyme-linked immunosorbent assay. Toxin identification was performed by high performance liquid chromatography. The ability of the strains to initiate BBD was tested on host coral fragments of Siderastrea siderea and Diploria strigosa under controlled laboratory conditions. Results showed that both Oscillatoria sp. strains caused infection that resulted in complete lysis of coral tissue. Both strains produced a cyanotoxin, identified as microcystin-LR, with production affected by different environmental factors. This study provides evidence that BBD Oscillatoria is a key component in BBD pathogenicity.
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Chianella, Iva. "Development of affinity sensors for Microcystin-LR based on a computationally designed molecularly imprinted polymer." Thesis, Cranfield University, 2003. http://dspace.lib.cranfield.ac.uk/handle/1826/10744.
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In this work the development of affinity sensors for the detection of microcystin-LR
based on a computationally designed artificial receptor is presented. Microcystin-LR is
a cyclic heptapeptide hepatotoxin produced by Cyanobacteria (aquatic organisms also
known as blue-green algae), which during blooms period can release toxins in water.
Clinical signs of hepatotoxicosis have been observed in domestic animals and livestock
and recently also in humans. At present, analysis of these toxins is achieved largely
using conventional, time consuming and expensive techniques such as chromatographic
methods (HPLC, TLC) and immunoassay. Therefore, the necessity of an easy and
inexpensive method of analysis such a biosensor is becoming urgent. In this work an
artificial receptor for microcystin-LR was synthesised using a combined approach of
molecular imprinting and computer modelling. A computer-aided rational design was
applied to study microcystin-LRlmonomers interactions in order to find an optimal
composition for the synthesis of the receptor. The optimised composition, suggested by
computer modelling, consisted in 1 mol of2-acrylamido-2-methyl-propanesulfonic acid
and 6 mol ofurocanic acid ethyl ester for 1 mol of template. This monomer composition
was then used to synthesise a molecularly imprinted polymer (MIP) and an enzyme-
linked competitive assay was developed to characterise the computational receptor. In
the assay, computational MIP was able both to detect 0.1 ~g rl of microcystin-LR and
to distinguish the analyte among analogues such as microcystin-YR, microcystin-RR
and nodularin.
The computationally designed receptor was then used as a sensing element for the
construction of sensor devices. A MIP-based piezoelectric sensor, capable of detecting
35 ~g rl of toxin in water, was developed. In order to improve the system sensitivity,
the computational polymer was also used as a material in solid-phase extraction (SPE)
for samples pre-concentration. The receptor was able to pre-concentrate up to 1,000 fold
tap water samples spiked with only 1 J.1g rl of toxin. By combining MIP-based SPE and
piezoelectric sensor an improved system with a minimum detectable concentration of
toxin of 0.35 ~g rl was achieved.
Encouraging preliminary results were also obtained in developing a MIP-based
electrochemical sensor.