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Soll, David R. "The Ins and Outs of DNA Fingerprinting the Infectious Fungi". Clinical Microbiology Reviews 13, n.º 2 (1 de abril de 2000): 332–70. http://dx.doi.org/10.1128/cmr.13.2.332.
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SUMMARY DNA fingerprinting methods have evolved as major tools in fungal epidemiology. However, no single method has emerged as the method of choice, and some methods perform better than others at different levels of resolution. In this review, requirements for an effective DNA fingerprinting method are proposed and procedures are described for testing the efficacy of a method. In light of the proposed requirements, the most common methods now being used to DNA fingerprint the infectious fungi are described and assessed. These methods include restriction fragment length polymorphisms (RFLP), RFLP with hybridization probes, randomly amplified polymorphic DNA and other PCR-based methods, electrophoretic karyotyping, and sequencing-based methods. Procedures for computing similarity coefficients, generating phylogenetic trees, and testing the stability of clusters are then described. To facilitate the analysis of DNA fingerprinting data, computer-assisted methods are described. Finally, the problems inherent in the collection of test and control isolates are considered, and DNA fingerprinting studies of strain maintenance during persistent or recurrent infections, microevolution in infecting strains, and the origin of nosocomial infections are assessed in light of the preceding discussion of the ins and outs of DNA fingerprinting. The intent of this review is to generate an awareness of the need to verify the efficacy of each DNA fingerprinting method for the level of genetic relatedness necessary to answer the epidemiological question posed, to use quantitative methods to analyze DNA fingerprint data, to use computer-assisted DNA fingerprint analysis systems to analyze data, and to file data in a form that can be used in the future for retrospective and comparative studies.
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DeScenzo, R. A. "Use of (CAT)5as a DNA Fingerprinting Probe for Fungi". Phytopathology 84, n.º 5 (1994): 534. http://dx.doi.org/10.1094/phyto-84-534.
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Meyer, Wieland, Anke Koch, Claudia Niemann, Birgit Beyermann, J�rg T. Epplen e Thomas B�rner. "Differentiation of species and strains among filamentous fungi by DNA fingerprinting". Current Genetics 19, n.º 3 (março de 1991): 239–42. http://dx.doi.org/10.1007/bf00336493.
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Lockhart, S. R., C. Pujol, S. Joly e D. R. Soll. "Development and use of complex probes for DNA fingerprinting the infectious fungi". Medical Mycology 39, n.º 1 (janeiro de 2001): 1–8. http://dx.doi.org/10.1080/mmy.39.1.1.8.
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WATANABE, M., K. LEE, K. GOTO, S. KUMAGAI, Y. SUGITA-KONISHI e Y. HARA-KUDO. "Rapid and Effective DNA Extraction Method with Bead Grinding for a Large Amount of Fungal DNA". Journal of Food Protection 73, n.º 6 (1 de junho de 2010): 1077–84. http://dx.doi.org/10.4315/0362-028x-73.6.1077.
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To identify a rapid method for extracting a large amount of DNA from fungi associated with food hygiene, extraction methods were compared using fungal pellets formed rapidly in liquid media. Combinations of physical and chemical methods or commercial kits were evaluated with 3 species of yeast, 10 species of ascomycetous molds, and 4 species of zygomycetous molds. Bead grinding was the physical method, followed by chemical methods involving sodium dodecyl sulfate (SDS), cetyl trimethyl ammonium bromide (CTAB), and benzyl chloride and two commercial kits. Quantity was calculated by UV absorbance at 260 nm, quality was determined by the ratio of UV absorbance at 260 and 280 nm, and gene amplifications and electrophoresis profiles of whole genomes were analyzed. Bead grinding with the SDS method was the most effective for DNA extraction for yeasts and ascomycetous molds, and bead grinding with the CTAB method was most effective with zygomycetous molds. For both groups of molds, bead grinding with the CTAB method was the best approach for DNA extraction. Because this combination also is relatively effective for yeasts, it can be used to extract a large amount of DNA from a wide range of fungi. The DNA extraction methods are useful for developing gene indexes to identify fungi with molecular techniques, such as DNA fingerprinting.
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Chiang, Yu-Chung, Chang-Hung Chou, Shong Huang e Tzen-Yuh Chiang. "Possible consequences of fungal contamination on the RAPD fingerprinting in Miscanthus (Poaceae)". Australian Journal of Botany 51, n.º 2 (2003): 197. http://dx.doi.org/10.1071/bt02021.
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Fungal contamination has been frequently reported in higher plants. In Miscanthus species, a wide range of fungal flora has also been recorded previously, including an investigation based on nrITS amplification. In order to understand the effects of the fungal genomes on the random amplified polymorphic DNA (RAPD) fingerprinting, callus specimens were obtained from the tissue culture of shoot apices of Miscanthus. RAPD fingerprinting with 60 oligoprimers was conducted with genomic DNA extracted from leaf tissue collected in the field and from the greenhouse, as well as callus derived from the same individuals. Extra bands were detected in the RAPD fingerprints amplified with 44 primers (84.6%) from the genomic DNA of both the field and greenhouse leaf tissue of most Miscanthus taxa examined, except for M. sinensis var. condensatus. Positive PCR amplification of organelle DNA non-coding spacers with both leaf and callus DNA ruled out the possibility that such DNA fingerprinting discrepancies were due to loss of organelles in the callus after consecutive subcultures. Among the 44 primers, one yielded no amplified fragments from the callus DNA, indicating that the amplified DNA fragments from leaf-tissue DNA were likely to be derived from fungi. The contaminating fungal DNA not only caused the overestimation of genetic diversity in the host plants, but also interfered with the phylogenetic inference. Systematic inconsistency occurred between the UPGMA dendrograms of leaf and callus DNA fingerprints. The detection of contaminating fungal DNA suggested that precautions are required for PCR-based fingerprinting when field materials are used for DNA resources. A method for quick screening of the contaminating fungal DNA with universal primers for the nrITS (internal transcribed spacer) region is suggested.
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Becerra-LopezLavalle, L. Augusto, Jennifer A. Saleeba e Bruce R. Lyon. "Molecular identification of fungi isolated from stem tissue of Upland cotton (Gossypium hirsutum)". Australian Journal of Botany 53, n.º 6 (2005): 571. http://dx.doi.org/10.1071/bt04092.
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Molecular techniques such as restriction fragment length polymorphism (RFLP) analysis, random amplification of polymorphic DNA (RAPD) fingerprinting, and DNA sequencing and database comparison, were employed to identify fungi isolated from field-grown cotton plants (Gossypium hirsutum L.). DNA fragments of between 510 and 590 bp, representing the two rDNA (rDNA) internal transcribed spacers (ITS1 and ITS2) and the intervening 5.8S rRNA gene, were amplified from the fungi with eukaryotic consensus primers. Subsequent digestion with the restriction endonucleases AluI, CfoI, HaeIII, HinfI and HpaII enabled the allotment of all 57 isolates to 13 different groups. Restriction analysis was supported by RAPD–PCR analysis of multiple isolates and rDNA sequencing of representative fungi from each group. Sequence alignment and comparison with rDNA sequences of other fungi available in GenBank allowed for putative identification of three different taxa of Fusarium, two taxa each of Cladosporium, Diaporthe and Nectria, and one taxon each of Alternaria, Ampelomyces, Bartalinia, Phaeosphaeria and Rhizoctonia. Many of the stem-colonising fungi identified in this study are either pathogenic on cotton or have elsewhere been found to act as biocontrol agents.
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Oh, S., D. P. Kamdem, D. E. Keathley e K. H. Han. "Detection and Species Identification of Wood-Decaying Fungi by Hybridization of Immobilized Sequence-Specific Oligonucleotide Probes with PCR-Amplified Fungal Ribosomal DNA Internal Transcribed Spacers". Holzforschung 57, n.º 4 (26 de junho de 2003): 346–52. http://dx.doi.org/10.1515/hf.2003.052.
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SummaryWe developed an effective detection method for wood-decaying fungi by hybridization of immobilized Sequence-Specific Oligonucleotide Probes with florescent-labeled PCR-amplified fungal rDNA internal transcribed spacer sequences. This method takes advantage of both the sequence specificity of Southern blot hybridization and the sensitivity of the previously reported PCR-based fungal species identification methods. Bothin vitrocultured fungal strains and naturally decaying wood samples were used to demonstrate that this method is robust and practical for detection of incipient wood-decaying fungi. It can be a useful tool for microbial ecology, plant pathology, protection of wood products in service, preservation efforts for high-value furniture and wood-based art and DNA fingerprinting for tracking the source of contamination of wood decay fungi.
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Gadkar, Vijay, Alok Adholeya e T. Satyanarayana. "Randomly amplified polymorphic DNA using the M13 core sequence of the vesicular–arbuscular mycorrhizal fungi Gigaspora margarita and Gigaspora gigantea". Canadian Journal of Microbiology 43, n.º 8 (1 de agosto de 1997): 795–98. http://dx.doi.org/10.1139/m97-115.
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Vesicular–arbuscular mycorrhizal (VAM) fungi are obligate symbionts, and a primary benefit provided to the host is the alleviation of stress. The recalcitrance of these fungi to grow in pure culture has spurred researchers to develop an alternative form of cultivation, namely the root organ culture (ROC). This synthetic form of production is new and efforts were made to use randomly amplified polymorphic DNA with the M13 minisatellite sequence as the polymerase chain reaction primer to look into polymorphism, if any, in the spores of Gigaspora margarita produced both in vitro and in situ (soil). The fingerprint patterns obtained from in vitro and in situ spores were similar. Extramatrical structures, such as auxiliary cells, were also examined by DNA fingerprinting. Their amplification pattern did not vary from the mother or daughter spores. A few interesting observations were made. For instance, the mother spore, which seemed hollow and inactive after germination, nevertheless contained nuclei after 4 months under in vitro conditions and generated a fingerprint pattern. The fingerprint pattern for Gigaspora margarita was different from that of Gigaspora gigantea, indicating that the minisatellite sequence could be exploited for identifying VAM fungi. ROC appears to be a truly representative system, in the sense that it mimics the essential features of the complex rhizosphere, allowing the fungi to complete their life cycle without any induced genetic changes per se. Key words : root organ culture, arbuscular mycorrhiza, M13 minisatellite sequence, randomly amplified polymorphic DNA.
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Witfeld, Frederick, Dominik Begerow e Marco Alexandre Guerreiro. "Improved strategies to efficiently isolate thermophilic, thermotolerant, and heat-resistant fungi from compost and soil". Mycological Progress 20, n.º 3 (março de 2021): 325–39. http://dx.doi.org/10.1007/s11557-021-01674-z.
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AbstractThermophilic, thermotolerant and heat-resistant fungi developed different physiological traits, enabling them to sustain or even flourish under elevated temperatures, which are life-hostile for most other eukaryotes. With the growing demand of heat-stable molecules in biotechnology and industry, the awareness of heat-adapted fungi as a promising source of respective enzymes and biomolecules is still increasing. The aim of this study was to test two different strategies for the efficient isolation and identification of distinctly heat-adapted fungi from easily accessible substrates and locations. Eight compost piles and ten soil sites were sampled in combination with different culture-dependent approaches to describe suitable strategies for the isolation and selection of thermophilous fungi. Additionally, an approach with a heat-shock treatment, but without elevated temperature incubation led to the isolation of heat-resistant mesophilic species. The cultures were identified based on morphology, DNA barcodes, and microsatellite fingerprinting. In total, 191 obtained isolates were assigned to 31 fungal species, from which half are truly thermophilic or thermotolerant, while the other half are heat-resistant fungi. A numerous amount of heat-adapted fungi was isolated from both compost and soil samples, indicating the suitability of the used approaches and that the richness and availability of those organisms in such environments are substantially high.
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Gonçalves, Micael F. M., Ana C. Esteves e Artur Alves. "Revealing the hidden diversity of marine fungi in Portugal with the description of two novel species, Neoascochyta fuci sp. nov. and Paraconiothyrium salinum sp. nov." International Journal of Systematic and Evolutionary Microbiology 70, n.º 10 (1 de outubro de 2020): 5337–54. http://dx.doi.org/10.1099/ijsem.0.004410.
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Fungi are ubiquitous organisms with a wide distribution in almost all ecosystems, including marine environments. Coastal and estuarine ecosystems remain poorly unexplored as fungal habitats, potentially harbouring a hidden diversity with important ecological roles. During an extensive survey of marine fungi in coastal and estuarine Portuguese environments, a collection of 612 isolates was obtained from water, algae, sponges and driftwood. From these, 282 representative isolates were selected through microsatellite-primed PCR (MSP-PCR) fingerprinting analysis, which were identified based on DNA sequence data. The collection yielded 117 taxa from 38 distinct genera, which were identified using DNA sequence analysis. Overall, fungal community composition varied with host/substrate, but the most abundant taxa in the collection were Cladosporium cladosporioides, Penicillium terrigenum, Penicillium brevicompactum and Fusarium equiseti/incarnatum complex. The occurrence of a high fungal diversity harbouring novel species was disclosed. Through a multilocus phylogeny based on ITS, tub2 and tef1-α sequences, in conjunction with morphological and physiological data, we propose Neoascochyta fuci sp. nov. and Paraconiothyrium salinum sp. nov.
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Kunová, Simona, Eva Ivanišová, Jana Žiarovská, Lucia Zamiešková, Soňa Felšöciová, Anka Trajkovska Petkoska, Daniela Nikolovska Nedelkoska e Miroslava Kačániová. "Differences between microbiota, phytochemical, antioxidant profile and dna fingerprinting of cabernet sauvignon grape from Slovakia and Macedonia". Potravinarstvo Slovak Journal of Food Sciences 14 (28 de outubro de 2020): 945–53. http://dx.doi.org/10.5219/1353.
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This study aimed to evaluate the microbiota, phytochemical, antioxidant profile and DNA fingerprinting of Cabernet Sauvignon grapes from Slovakia and R. North Macedonia. There were used two samples of grape berries (one sample from Slovakia and one from Macedonia). Each sample was analyzed in triplicate. The bacteria were cultivated on Plate count agar (PCA), microscopic filamentous fungi were cultivated on Malt extract agar (MEA). MALDI-TOF MS Biotyper mass spectrometry was used for the identification of microorganisms (bacteria and yeasts) and microscopic filamentous fungi with manuals. DPPH method was used to determine of antioxidant activity of grape berries. Phytochemical and antioxidant profiles were evaluated in grape berries samples. Total genomic DNA was extracted from mature grapes by GeneJET Plant Genomic DNA Purification Kit. The number of bacteria was higher in the sample of Macedonian grape (4.13 log CFU.g-1) in comparison to the grape from Slovakia as well as the number of yeasts was also higher in the Macedonian sample (2.57 log CFU.g-1). Antioxidant activity of Slovak grape berries was 0.55 mg TEAC.g-1 and of Macedonian grape, berries was 0.51 mg TEAC.g-1. Total polyphenol content was higher in grape from Slovakia (0.81 mg GAE.g-1) than in grape from Macedonia (0.77 mg GAE.g-1), while total flavonoid content was 0.57 and 0.17 mg QE.g-1 in Slovak grape and Macedonian grape, respectively. Total phenolic acid content was higher in the sample from Macedonia (0.40 mg CAE.g-1) compared to the grape from Slovakia (0.24 mg CAE.g-1). Total anthocyanin content was also higher in Macedonian grape (0.46 mg.g-1) compared to the Slovak sample (0.05 mg.g-1). The total polymorphism for all of the used primers of 87.5% was obtained for the Macedonian sample of Cabernet Sauvignon and 89.4% for the Slovak sample.
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White, John, Jeppe Nielsen e Anne Madsen. "Potential Respiratory Deposition and Species Composition of Airborne Culturable, Viable, and Non-Viable Fungi during Occupancy in a Pig Farm". Atmosphere 11, n.º 6 (16 de junho de 2020): 639. http://dx.doi.org/10.3390/atmos11060639.
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Fungal species composition and site of deposition within the airways affects whether diseases develop and where they may arise. The aim of this study is to obtain knowledge regarding the potential deposition of airborne culturable, viable, and non-viable fungi in the airways of pig farm workers, and how this composition changes over multiple sampling days. Airborne fungi were sampled using impactors and subsequently analyzed using amplicon sequencing and matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) fingerprinting. The geometric mean aerodynamic diameter (Dg) of airborne particles with culturable airborne fungi were not affected by sampling days and ranged in size between 3.7 and 4.6 µm. Amplicon sequencing of the internal transcribed spacer region of the rRNA gene operon, in combination with DNA interchelating agents, revealed a large presence of non-viable fungi, but several pathogenic and toxic fungal species were detected in the viable portion. The diversity was found to be significantly associated with the sampling day but did not change significantly over multiple sampling rounds during the same day. The non-viable fraction contained genera typically associated with the pig gastrointestinal tract, such as Kazachstania and Vishniacozyma. In conclusion, the Dg of culturable fungi was between 3.7 and 4.6 µm, and the Dg of the viable and total fungi was 1.5 and 2.1 µm, respectively. The species composition changed over the multiple sampling days.
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Kageyama, Stacie A., Nancy Ritchie Posavatz, Kirk E. Waterstripe, Sarah J. Jones, Peter J. Bottomley, Kermit Cromack e David D. Myrold. "Fungal and bacterial communities across meadow–forest ecotones in the western Cascades of Oregon". Canadian Journal of Forest Research 38, n.º 5 (maio de 2008): 1053–60. http://dx.doi.org/10.1139/x07-221.
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Meadows are natural dynamic features of forested mountain landscapes of the Pacific Northwest. Proportions of meadows and forests change with environmental conditions and disturbance history. We investigated the belowground microbial communities associated with these two vegetation types and how they change across the meadow–forest transition at two sites in Oregon. Soils were sampled along replicate transects extending from meadow into forest. We quantified total bacterial and fungal biomass using direct microscopy and described the composition of bacterial and fungal communities using a DNA-based fingerprinting technique. Bacterial biomass was similar in meadow and forest soils, but fungal biomass was significantly higher in forest soil. Meadow and forest soils had distinct communities of bacteria and fungi. Bacterial communities near the meadow–forest boundary reflected current vegetation, but fungal communities under meadow vegetation near the forest edge were intermediate in composition between those found in meadow and forest soils. The more gradual transition observed with fungal communities may reflect the influence of tree roots and their associated ectomycorrhizal fungi or possibly colonization by saprotrophic fungi associated with tree litter accumulating near the forest edge. Invasion of forest-associated fungi into the meadow soils may presage subsequent expansion of forest vegetation into meadows.
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Okuda, Toru, Mieko Yanagisawa, Fumihiro Fujimori, Yuri Nishizuka, Yuki Takehana e Masato Sugiyama. "New isolation methods and polymerase chain reaction strain discrimination techniques for natural products screening programs". Canadian Journal of Botany 73, S1 (31 de dezembro de 1995): 946–54. http://dx.doi.org/10.1139/b95-343.
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A new isolation method of fungi was established by using the Percoll density gradient centrifugation. We found that the difference in the specific gravity allowed us to separate mixtures of fungal spores of Aspergillus, Fusarium, Listeromyces, Penicillium, and Trichoderma. When this technique was applied to soil samples, a wider variety of fungi were isolated compared with those obtained from an ordinary serial dilution method. For screening purposes, it is necessary to eliminate redundant strains from the strains initially isolated. For the efficient discrimination of strains, the random amplified polymorphic DNA (RAPD) method was applied. The establishment of a standard method for RAPD enabled us to minimize the deviation of Rf values between gels to less than 0.03. Conversion of the Rf values to binary matrices provided a matrix data that could be analyzed statistically. Based on the RAPD, it was possible to select genetically distinct fungal strains within a species. Key words: fingerprinting, isolation method, microbial screening, PCR, Percoll density gradient, RAPD.
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Prat, Chantal, Olaya Ruiz-Rueda, Rosalia Trias, Enriqueta Anticó, Dimitra Capone, Mark Sefton e Lluís Bañeras. "Molecular Fingerprinting by PCR-Denaturing Gradient Gel Electrophoresis Reveals Differences in the Levels of Microbial Diversity for Musty-Earthy Tainted Corks". Applied and Environmental Microbiology 75, n.º 7 (5 de fevereiro de 2009): 1922–31. http://dx.doi.org/10.1128/aem.02758-08.
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ABSTRACT The microbial community structure of cork with marked musty-earthy aromas was analyzed using denaturing gradient gel electrophoresis of amplified ribosomal DNA. Cork stoppers and discs were used for DNA extraction and were analyzed by using selective primers for bacteria and fungi. Stoppers clearly differed from discs harboring a different fungal community. Moreover, musty-earthy samples of both types were shown to have a specific microbiota. The fungi Penicillium glabrum and Neurospora spp. were present in all samples and were assumed to make only a small contribution to off-odor development. In contrast, Penicillium islandicum and Penicillium variabile were found almost exclusively in 2,4,6-trichloroanisole (TCA) tainted discs. Conversely, Rhodotorula minuta and Rhodotorula sloofiae were most common in cork stoppers, where only small amounts of TCA were detected. Alpha- and gammaproteobacteria were the most commonly found bacteria in either control or tainted cork stoppers. Specific Pseudomonas and Actinobacteria were detected in stoppers with low amounts of TCA and 2-methoxy-3,5-dimethylpyrazine. These results are discussed in terms of biological degradation of taint compounds by specific microorganisms. Reliable and straightforward microbial identification methods based on a molecular approach provided useful data to determine and evaluate the risk of taint formation in cork.
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McDonald, B. A., R. E. Pettway, R. S. Chen, J. M. Boeger e J. P. Martinez. "The population genetics of Septoria tritici (teleomorph Mycosphaerella graminicola)". Canadian Journal of Botany 73, S1 (31 de dezembro de 1995): 292–301. http://dx.doi.org/10.1139/b95-259.
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The DNA-based markers of molecular genetics were combined with the analytical tools of population genetics to learn about the population biology of the wheat pathogen Mycosphaerella graminicola. DNA-based genetic markers, including restriction fragment length polymorphisms in nuclear and mitochondrial DNA, DNA fingerprints, and electrophoretic karyotypes were used in combination to show that the amount and distribution of genetic variation within and among field populations of M. graminicola is similar around the world. Measures of gametic disequilibrium suggested that the sexual stage of reproduction has a more significant impact on the genetic structure of M. graminicola populations than asexual reproduction. A field experiment conducted over a 3-year period showed that populations had a high degree of genetic stability over time. The potential effects of selection were quantified in a cultivar mixture experiment with four wheat cultivars that varied in resistance to M. graminicola. In combination, these experiments demonstrated the utility of selectively neutral genetic markers to elucidate the population genetics of fungi. Key words: genetic diversity, wheat, gene flow, RFLPs, DNA fingerprinting.
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Verbruggen, Erik, Eiko E. Kuramae, Remy Hillekens, Mattias de Hollander, E. Toby Kiers, Wilfred F. M. Röling, George A. Kowalchuk e Marcel G. A. van der Heijden. "Testing Potential Effects of Maize Expressing the Bacillus thuringiensis Cry1Ab Endotoxin (Bt Maize) on Mycorrhizal Fungal Communities via DNA- and RNA-Based Pyrosequencing and Molecular Fingerprinting". Applied and Environmental Microbiology 78, n.º 20 (10 de agosto de 2012): 7384–92. http://dx.doi.org/10.1128/aem.01372-12.
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ABSTRACTThe cultivation of genetically modified (GM) crops has increased significantly over the last decades. However, concerns have been raised that some GM traits may negatively affect beneficial soil biota, such as arbuscular mycorrhizal fungi (AMF), potentially leading to alterations in soil functioning. Here, we test two maize varieties expressing theBacillus thuringiensisCry1Ab endotoxin (Bt maize) for their effects on soil AM fungal communities. We target both fungal DNA and RNA, which is new for AM fungi, and we use two strategies as an inclusive and robust way of detecting community differences: (i) 454 pyrosequencing using general fungal rRNA gene-directed primers and (ii) terminal restriction fragment length polymorphism (T-RFLP) profiling using AM fungus-specific markers. Potential GM-induced effects were compared to the normal natural variation of AM fungal communities across 15 different agricultural fields. AM fungi were found to be abundant in the experiment, accounting for 8% and 21% of total recovered DNA- and RNA-derived fungal sequences, respectively, after 104 days of plant growth. RNA- and DNA-based sequence analyses yielded most of the same AM fungal lineages. Our research yielded three major conclusions. First, no consistent differences were detected between AM fungal communities associated with GM plants and non-GM plants. Second, temporal variation in AMF community composition (between two measured time points) was bigger than GM trait-induced variation. Third, natural variation of AMF communities across 15 agricultural fields in The Netherlands, as well as within-field temporal variation, was much higher than GM-induced variation. In conclusion, we found no indication that Bt maize cultivation poses a risk for AMF.
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Trigiano, R. N., G. Caetano-Anollés, B. J. Bassam e M. T. Windham. "309 GENOMIC ANALYSIS OF TWO FUNGI CAUSING ANTHRACNOSE OF DOGWOOD (CORNUS SPECIES) IN THE EASTERN UNITED STATES". HortScience 29, n.º 5 (maio de 1994): 474e—474. http://dx.doi.org/10.21273/hortsci.29.5.474e.
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DNA Amplification Fingerprinting (DAF) was used to characterize ten isolates of Discula destructiva Redlin and three isolates of an undescribed species of Discula, the causal organisms of dogwood (Cornus species) anthracnose. Isolates were obtained throughout the disease range in the eastern United States and DAF profiles generated with ten arbitrary oligonucleotide primers. Very few polymorphic loci (27/298) were detected between isolates of D. destructiva; whereas, a greater number were observed between and among the isolates of Discula species. Relationships among and between the two fungal groups were analyzed using PAUP and UPGMA and indicate that the genome of D. destructiva is highly conserved throughout the distribution. In contrast, isolates of Discula species exhibited greater variability. This suggests that D. destructive was recently introduced to the eastern United States.
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Valinsky, Lea, Gianluca Della Vedova, Tao Jiang e James Borneman. "Oligonucleotide Fingerprinting of rRNA Genes for Analysis of Fungal Community Composition". Applied and Environmental Microbiology 68, n.º 12 (dezembro de 2002): 5999–6004. http://dx.doi.org/10.1128/aem.68.12.5999-6004.2002.
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ABSTRACT Thorough assessments of fungal diversity are currently hindered by technological limitations. Here we describe a new method for identifying fungi, oligonucleotide fingerprinting of rRNA genes (OFRG). ORFG sorts arrayed rRNA gene (ribosomal DNA [rDNA]) clones into taxonomic clusters through a series of hybridization experiments, each using a single oligonucleotide probe. A simulated annealing algorithm was used to design an OFRG probe set for fungal rDNA. Analysis of 1,536 fungal rDNA clones derived from soil generated 455 clusters. A pairwise sequence analysis showed that clones with average sequence identities of 99.2% were grouped into the same cluster. To examine the accuracy of the taxonomic identities produced by this OFRG experiment, we determined the nucleotide sequences for 117 clones distributed throughout the tree. For all but two of these clones, the taxonomic identities generated by this OFRG experiment were consistent with those generated by a nucleotide sequence analysis. Eighty-eight percent of the clones were affiliated with Ascomycota, while 12% belonged to Basidiomycota. A large fraction of the clones were affiliated with the genera Fusarium (404 clones) and Raciborskiomyces (176 clones). Smaller assemblages of clones had high sequence identities to the Alternaria, Ascobolus, Chaetomium, Cryptococcus, and Rhizoctonia clades.
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Gryta, Agata, e Magdalena Frąc. "Methodological Aspects of Multiplex Terminal Restriction Fragment Length Polymorphism-Technique to Describe the Genetic Diversity of Soil Bacteria, Archaea and Fungi". Sensors 20, n.º 11 (9 de junho de 2020): 3292. http://dx.doi.org/10.3390/s20113292.
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The molecular fingerprinting methods used to evaluate soil microbial diversity could also be used as effective biosensors for the purposes of monitoring ecological soil status. The biodiversity of microorganisms is a relevant index of soil activity and there is a necessity to develop tools to generate reliable results for an emerging approach in the field of environmental control using microbial diversity biosensors. This work reports a method under development for determining soil microbial diversity using high efficiency Multiplex PCR-Terminal Restriction Fragment Length Polymorphism (M-T-RFLP) for the simultaneous detection of bacteria, archaea and fungi. Three different primer sets were used in the reaction and the analytical conditions were optimized. Optimal analytical conditions were achieved using 0.5 µM of primer for bacteria and 1 µM for archaea and fungi, 4 ng of soil DNA template, and HaeIII restriction enzyme. Comparative tests using the proposed analytical approach and a single analysis of each microorganism group were carried out to indicate that both genetic profiles were similar. The Jaccard similarity coefficient between single and multiplexing approach ranged from 0.773 to 0.850 for bacteria and fungi, and 0.208 to 0.905 for archaea. In conclusion, the multiplexing and pooling approaches significantly reduced the costs and time required to perform the analyses, while maintaining a proper effectiveness.
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Campbell, Michael, Michael Campbell, Rachael Adams, Emily Dobry, Kara Dobson, Kara Dobson, Veronica Stefanick e Jessica Till. "The Sprout Regulating Compound 1,4-Dimethylnaphthalene Exhibits Fungistatic Activity". Journal of Agronomy Research 1, n.º 3 (10 de janeiro de 2019): 27–34. http://dx.doi.org/10.14302/issn.2639-3166.jar-18-2502.
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The compound 1,4-dimethylnaphthalene, originally isolated from dormant potatoes, is currently in use as a commercial sprout inhibitor. Growers and processors report a reduction in fungal infections in potatoes treated with DMN resulting in increased yields. To assess the effects of DMN on fungal growth a culture of Fusarium oxysporum was isolated from potato tubers and identified via DNA fingerprinting using the 18ITS ribosomal region. Growth of F. oxysporum was inhibited by 31% after four days of exposure to DMN but overall rate of spore germination was not affected by DMN treatment. The growth of additional fungi, including Alternaria alternata, Aspergillus niger, Epicoccum nigrum, Gnomoniopsis smithogilvyi, Phoma medicaginis, and Pythium ultimum was inhibited by DMN as was suppression of sporulation in A. niger. These results suggest that DMN is fungistatic at the application levels examined.
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Moon, Christina D., Brian A. Tapper e Barry Scott. "Identification of Epichloë Endophytes In Planta by a Microsatellite-Based PCR Fingerprinting Assay with Automated Analysis". Applied and Environmental Microbiology 65, n.º 3 (1 de março de 1999): 1268–79. http://dx.doi.org/10.1128/aem.65.3.1268-1279.1999.
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ABSTRACT Epichloë endophytes are a group of filamentous fungi that include both sexual (Epichloë) and asexual (Neotyphodium) species. As a group they are genetically diverse and form both antagonistic and mutualistic associations with temperate grasses. We report here on the development of a microsatellite-based PCR system for fingerprinting this group of fungi with template isolated from either culture or infected plant material. M13mp19 partial genomic libraries were constructed for size-fractionated genomic DNA from two endophyte strains. These libraries were screened with a mixture of DIG-labeled dinucleotide and trinucleotide repeat probes. Positive clones were sequenced, and nine unique microsatellite loci were identified. An additional microsatellite was serendipitously identified in the 3′ untranscribed region of the 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase gene from N. lolii Lp19. Primers were designed for each locus and a panel of endophytes, from different taxonomic groupings, was screened to determine the degree of polymorphism. On the basis of these results a multiplex assay was developed for strain identification with fluorescently labeled primers for five of these loci. Using this system the size of the products amplified can be precisely determined by automated analysis, and an allele profile for each strain can be readily generated. The assay was shown to resolve endophyte groupings to the level of known isozyme phenotype groupings. In a blind test the assay was used successfully to identify a set of endophytes in planta. A reference database of allele sizes has been established for the panel of endophytes examined, and this will be expanded as new strains are analyzed.
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Sudini, H., C. R. Arias, M. R. Liles, K. L. Bowen e R. N. Huettel. "Comparison of Soil Fungal Community Structure in Different Peanut Rotation Sequences Using Ribosomal Intergenic Spacer Analysis in Relation to Aflatoxin-Producing Fungi". Phytopathology® 101, n.º 1 (janeiro de 2011): 52–57. http://dx.doi.org/10.1094/phyto-03-10-0072.
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The present study focuses on determining soil fungal community structure in different peanut-cropping sequences by using a high-resolution DNA fingerprinting technique: ribosomal intergenic spacer analysis (RISA). This study was initiated to determine fungal community profiles in four peanut-cropping sequences (continuous peanut, 4 years of continuous bahiagrass followed by peanut, peanut-corn-cotton, and peanut-cotton rotations), with a special focus to evaluate whether the profiles under investigation may have also indicated microbial differences that could affect Aspergillus flavus populations. Results indicated 75% similarities among fungal communities from the same cropping sequences as well as with similar times of sampling. Polymerase chain reaction (PCR)-based detection of A. flavus directly from these soils was carried out using A. flavus-specific primers (FLA1 and FLA2) and also through quantitative estimation on A. flavus and A. parasiticus agar medium. Population levels of A. flavus in soil samples ranged from zero to 1.2 × 103 CFU g–1 of soil (based on culturable methods); however, the fungus was not detected with A. flavus-specific primers. The minimum threshold limit at which these aflatoxin-producing fungi could be detected from the total soil genomic DNA was determined through artificial inoculation of samples with 10-fold increases in concentrations. The results indicated that a minimum population density of 2.6 × 106 CFU g–1 of soil is required for PCR detection in our conditions. These results are useful in further determining the relative population levels of these fungi in peanut soils with other soil fungi. This is a new approach to understanding soil fungal communities and how they might change over time and under different rotation systems.
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Meyer, Wieland, e Thomas G. Mitchell. "Polymerase chain reaction fingerprinting in fungi using single primers specific to minisatellites and simple repetitive DNA sequences: Strain variation inCryptococcus neoformans". Electrophoresis 16, n.º 1 (1995): 1648–56. http://dx.doi.org/10.1002/elps.11501601273.
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Ahmad, Rana Zaheer, Fuad Ameen, Rida Khalid, Mousa A. Alghuthaymi, Reem Alsalmi e Chunjie Li. "A Brief History of Endophyte Detection Techniques in Grasses". Sustainable Agriculture Research 8, n.º 3 (27 de julho de 2019): 66. http://dx.doi.org/10.5539/sar.v8n3p66.
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Endophytes are the plant mutualists that live asymptomatically inside plant tissue and are found in nearly whole plant kingdom. Endophytic fungi receive shelter and nutrition from host plants and in return provide great advantages to the host. Grasses are a useful forage species and are of great agricultural and socio-economic value. The presence of endophytes in these grasses provide protection, persistence and improved yield against herbivores, insects, pathogens, drought and several other biotic and abiotic stresses. This review summarizes traditional and modern molecular techniques to identify endophytes from turf and forage grasses. Traditional approaches include direct observation, staining, laser micro dissection and pressure catapulting and cultivation-dependent methods that provide a morphological identification of endophytic mycobiota in grass tissues. Earlier studies on endophytes using these methods resulted in several technical implications which molecular approaches are able to solve now-a-days. Molecular approaches include DNA extraction, PCR based DNA Fingerprinting techniques, Denaturing Gradient Gel Electrophoresis, Sanger sequencing, Pyrosequencing, Immunoblot assay, Biosensors, DNA Barcoding and Molecular Phylogenetics etc. A comparison of these detection techniques will facilitate other researchers as well to develop new ways for the detection of endophytes that will contribute to the improvement of grassland in future.
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Rajala, Tiina, Mikko Peltoniemi, Taina Pennanen e Raisa Mäkipää. "Relationship between wood-inhabiting fungi determined by molecular analysis (denaturing gradient gel electrophoresis) and quality of decaying logs". Canadian Journal of Forest Research 40, n.º 12 (dezembro de 2010): 2384–97. http://dx.doi.org/10.1139/x10-176.
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We investigated the fungal communities inhabiting decaying logs in a seminatural boreal forest stand in relation to host tree species, stage of decay, density, diameter, moisture, C to N ratio, Klason lignin content, and water- and ethanol-soluble extractives. Communities were profiled using denaturing gradient gel electrophoresis fingerprinting of the rDNA ITS1 region coupled with sequencing of fungal DNA extracted directly from the wood. In addition, polypore fruit bodies were inventoried. Logs from different tree species had different fungal communities and different physicochemical properties (e.g., C to N ratio, density, ethanol extractives, and diameter). Ascomycetes comprised a larger portion of communities inhabiting deciduous birch ( Betula spp.) and European aspen ( Populus tremula L.) logs compared with those living on coniferous Norway spruce ( Picea abies (L.) Karst.) and Scots pine ( Pinus sylvestris L.). A relationship between mycelial community structure and density of decaying spruce logs suggested a succession of fungi with mass loss of wood. The fruit body inventory underestimated fungal diversity in comparison with the culture-free denaturing gradient gel electrophoresis analysis that also detected inconspicuous but important species inhabiting decaying wood.
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MacNeil, L., T. Kauri e W. Robertson. "Molecular techniques and their potential application in monitoring the microbiological quality of indoor air". Canadian Journal of Microbiology 41, n.º 8 (1 de agosto de 1995): 657–65. http://dx.doi.org/10.1139/m95-091.
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Health effects associated with poor indoor air quality have created a need for accurate, reproducible methods of monitoring the microbiological content of indoor air. Improved methods of detection may allow researchers to clarify the effect of individual species present in the indoor environment on human health. This review discusses the shortcomings of current methods of identification and detection and focuses on the potential for molecular techniques in this emerging field. Probe techniques, restriction endonuclease analysis, karyotyping, and DNA and polymerase chain reaction fingerprinting methods available to detect and identify bacteria and fungi significant in the indoor air environment are discussed. Problems that may be encountered using these techniques are also considered. The authors have included a brief discussion on current air sampling techniques as well as adapting these techniques for use with molecular detection methods.Key words: indoor air microbiology, microbiological air sampling, molecular detection methods.
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Pagano, M. C. "Rhizobia associated with neotropical tree Centrolobium tomentosum used in riparian restoration". Plant, Soil and Environment 54, No. 11 (2 de dezembro de 2008): 498–508. http://dx.doi.org/10.17221/436-pse.
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<I>Centrolobium tomentosum</I> is a tropical legume tree indicated for functional and structural restoration of riparian areas. This study was conducted to characterize the rhizobia isolated from nodules of <I>C. tomentosum in situ</I> and to determine their capacity of renodulation, in an experimental area of land rehabilitation in the Rio Doce valley. Nodulation potential to inoculation with 2 selected fast-growing <I>Rhizobium</I> strains separately and a mixed inoculum of arbuscular mycorrhizal fungi was evaluated by the use of antibiotics resistance. Flood disturbance were observed not to affect renodulation by fast-growing strains. DNA fingerprinting RAPD (random amplified polymorphic DNA) and lipopolysaccharides (LPS) profiles were used to examine molecular relationships among field isolates, inoculants and reference strains. Maximal renodulation was exhibited by strain BHCBAb1 after 24 months after transplantation. <I>Centrolobium tomentosum</I> forms symbiosis with fast- and slow-growing <I>Rhizobium</I> strains, and it is suggested that their nursery culture could be improved by inoculation of selected strain under low nitrogen-input conditions.
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Wang, Xiaojing, Xinxin Wang e Gu Feng. "Optimised nitrogen fertiliser management achieved higher diversity of arbuscular mycorrhiza fungi and high-yielding maize (Zea mays L.)". Crop and Pasture Science 66, n.º 7 (2015): 706. http://dx.doi.org/10.1071/cp14160.
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The integrated soil–crop system management (ISSM) approach can potentially mitigate the loss of biodiversity in agricultural landscapes, ensuring crop yield with lower nitrogen (N) fertiliser input and minimised environmental pollution. The aim of this study was to test the hypotheses that overuse of N fertiliser could reduce the biodiversity of arbuscular mycorrhizal fungi (AMF) and that ISSM could help to maintain higher AMF biodiversity than the conventionally managed system in maize (Zea mays L.). The AMF community composition under three different treatments (conventionally managed, N-optimised and non-N-fertilised fields) was assessed by using both spore-based morphological taxonomy and DNA-based T-RFLP fingerprinting approaches. Maize roots in intensively managed fields formed functioning mycorrhizal symbioses even when a high rate of N fertiliser was applied. AMF diversity was higher under optimised N input, whereas AMF richness decreased when more N fertiliser was used. The N-optimised farms had AMF communities similar to those in the conventionally managed fields. The ISSM approach is recommended for sustaining crop yields without incurring continuing environmental costs and for maintaining AMF communities in intensively managed agro-ecosystems, especially in rapidly developing countries.
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Andrade, Orlando, Gastón Muñoz, Rafael Galdames, Paola Durán e Rodrigo Honorato. "Characterization, In Vitro Culture, and Molecular Analysis of Thecaphora solani, the Causal Agent of Potato Smut". Phytopathology® 94, n.º 8 (agosto de 2004): 875–82. http://dx.doi.org/10.1094/phyto.2004.94.8.875.
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The fungus Thecaphora solani (syn.: Angiosorus solani), the causal agent of potato smut, was cultivated in vitro for the first time. Teliospores obtained from galls of infected potato plants were used to inoculate commonly used solid and liquid media. The teliospores produced two kinds of vegetative tissue depending on the nutrient status of the media. A very slow radial-growing, hyaline, and septate mycelium, as usually seen in most of the in vitro-cultivated filamentous fungi, was obtained in wateragar medium after 30 to 40 days. On the other hand, a white, sponge-like mycelial mass was obtained in HCM + 1% activated charcoal, and on common potato dextrose agar or malt-yeast-peptone solid or liquid media, after 40 to 50 days under lab conditions. The identity among teliospores and the sponge-like mycelial mass was corroborated by DNA fingerprinting and partial sequencing of the large subunit (LSU) rDNA region. The sexual cycle of the pathogen was completed under lab conditions based on the development of teliospores on the sponge-like mycelial mass. The first attempt to reproduce the disease under controlled conditions was successful, inducing a gall in a cv. Desirée potato explant cultivated in vitro inoculated with radial-growing mycelia. Phylogenetic analysis of LSU rDNA data of the genus Thecaphora and other smut fungi confirmed the initial classification of the pathogen as T. solani.
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Goodwin, Stephen B., Jessica R. Cavaletto, Cees Waalwijk e Gert H. J. Kema. "DNA Fingerprint Probe from Mycosphaerella graminicola Identifies an Active Transposable Element". Phytopathology® 91, n.º 12 (dezembro de 2001): 1181–88. http://dx.doi.org/10.1094/phyto.2001.91.12.1181.
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DNA fingerprinting has been used extensively to characterize populations of Mycosphaerella graminicola, the Septoria tritici blotch pathogen of wheat. The highly polymorphic DNA fingerprints of Mycosphaerella graminicola were assumed to reflect the action of transposable elements. However, there was no direct evidence to support that conclusion. To test the transposable element hypothesis, the DNA fingerprint probe pSTL70 was sequenced, along with three other clones from a subgenomic library that hybridized with pSTL70. Analysis of these sequences revealed that pSTL70 contains the 3′ end of a reverse transcriptase sequence plus 29- and 79-bp direct repeats. These are characteristics of transposable elements identified in other organisms. Southern analyses indicated that either the direct-repeat or reverse-transcriptase sequences by themselves essentially duplicated the original DNA fingerprint pattern, but other portions of pSTL70 contained single-copy DNA. Analysis of 60 progeny from a sexual cross between two Dutch isolates of Mycosphaerella graminicola identified several new bands that were not present in the parents. Thus, the putative transposable element probably is active during meiosis. Tests of single-spore isolates revealed gains or losses of one or more DNA fingerprint bands in 4 out of 10 asexual lines derived from isolate IPO94269. Therefore, DNA fingerprint patterns produced by the putative transposable element were capable of changes during asexual reproduction of this isolate. Probe pSTL70 did not hybridize at high stringency to genomic DNAs from other fungi related to Septoria and Mycosphaerella. These results indicate that the DNA fingerprint probe pSTL70 most likely identifies a transposable element in Mycosphaerella graminicola that may have been acquired recently, and appears to be active during both sexual and asexual reproduction.
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Pancher, Michael, Marco Ceol, Paola Elisa Corneo, Claudia Maria Oliveira Longa, Sohail Yousaf, Ilaria Pertot e Andrea Campisano. "Fungal Endophytic Communities in Grapevines (Vitis vinifera L.) Respond to Crop Management". Applied and Environmental Microbiology 78, n.º 12 (6 de abril de 2012): 4308–17. http://dx.doi.org/10.1128/aem.07655-11.
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ABSTRACTWe studied the distribution of fungal endophytes of grapevine (Vitis viniferaL.) plants in a subalpine area of northern Italy, where viticulture is of high economic relevance. We adopted both cultivation-based and cultivation-independent approaches to address how various anthropic and nonanthropic factors shape microbial communities. Grapevine stems were harvested from several locations considering organic and integrated pest management (IPM) and from the cultivars Merlot and Chardonnay. Cultivable fungi were isolated and identified by internal-transcribed-spacer sequence analysis, using a novel colony-PCR method, to amplify DNA from fungal specimens. The composition of fungal communities was assessed using a cultivation-independent approach, automated ribosomal intergenic spacer analysis (ARISA). Multivariate statistical analysis of both culture-dependent and culture-independent data sets was convergent and indicated that fungal endophytic communities in grapevines from organically managed farms were different from those from farms utilizing IPM. Fungal communities in plants of cv. Merlot and cv. Chardonnay overlapped when analyzed using culture-dependent approaches but could be partially resolved using ARISA fingerprinting.
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Himmelreich, Uwe, Ray L. Somorjai, Brion Dolenko, Ok Cha Lee, Heide-Marie Daniel, Ronan Murray, Carolyn E. Mountford e Tania C. Sorrell. "Rapid Identification of Candida Species by Using Nuclear Magnetic Resonance Spectroscopy and a Statistical Classification Strategy". Applied and Environmental Microbiology 69, n.º 8 (agosto de 2003): 4566–74. http://dx.doi.org/10.1128/aem.69.8.4566-4574.2003.
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ABSTRACT Nuclear magnetic resonance (NMR) spectra were acquired from suspensions of clinically important yeast species of the genus Candida to characterize the relationship between metabolite profiles and species identification. Major metabolites were identified by using two-dimensional correlation NMR spectroscopy. One-dimensional proton NMR spectra were analyzed by using a staged statistical classification strategy. Analysis of NMR spectra from 442 isolates of Candida albicans, C. glabrata, C. krusei, C. parapsilosis, and C. tropicalis resulted in rapid, accurate identification when compared with conventional and DNA-based identification. Spectral regions used for the classification of the five yeast species revealed species-specific differences in relative amounts of lipids, trehalose, polyols, and other metabolites. Isolates of C. parapsilosis and C. glabrata with unusual PCR fingerprinting patterns also generated atypical NMR spectra, suggesting the possibility of intraspecies discontinuity. We conclude that NMR spectroscopy combined with a statistical classification strategy is a rapid, nondestructive, and potentially valuable method for identification and chemotaxonomic characterization that may be broadly applicable to fungi and other microorganisms.
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Iquebal, Mir Asif, Sarika Jaiswal, Vineet Kumar Mishra, Rahul Singh Jasrotia, Ulavappa B. Angadi, Bhim Pratap Singh, Ajit Kumar Passari et al. "Fungal Genomic Resources for Strain Identification and Diversity Analysis of 1900 Fungal Species". Journal of Fungi 7, n.º 4 (12 de abril de 2021): 288. http://dx.doi.org/10.3390/jof7040288.
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Identification and diversity analysis of fungi is greatly challenging. Though internal transcribed spacer (ITS), region-based DNA fingerprinting works as a “gold standard” for most of the fungal species group, it cannot differentiate between all the groups and cryptic species. Therefore, it is of paramount importance to find an alternative approach for strain differentiation. Availability of whole genome sequence data of nearly 2000 fungal species are a promising solution to such requirement. We present whole genome sequence-based world’s largest microsatellite database, FungSatDB having >19M loci obtained from >1900 fungal species/strains using >4000 assemblies across globe. Genotyping efficacy of FungSatDB has been evaluated by both in-silico and in-vitro PCR. By in silico PCR, 66 strains of 8 countries representing four continents were successfully differentiated. Genotyping efficacy was also evaluated by in vitro PCR in four fungal species. This approach overcomes limitation of ITS in species, strain signature, and diversity analysis. It can accelerate fungal genomic research endeavors in agriculture, industrial, and environmental management.
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Buchan, Alison, Steven Y. Newell, Melissa Butler, Erin J. Biers, James T. Hollibaugh e Mary Ann Moran. "Dynamics of Bacterial and Fungal Communities on Decaying Salt Marsh Grass". Applied and Environmental Microbiology 69, n.º 11 (novembro de 2003): 6676–87. http://dx.doi.org/10.1128/aem.69.11.6676-6687.2003.
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ABSTRACT Both bacteria and fungi play critical roles in decomposition processes in many natural environments, yet only rarely have they been studied as an integrated microbial community. Here we describe the bacterial and fungal assemblages associated with two decomposition stages of Spartina alterniflora detritus in a productive southeastern U.S. salt marsh. 16S rRNA genes and 18S-to-28S internal transcribed spacer (ITS) regions were used to target the bacterial and ascomycete fungal communities, respectively, based on DNA sequence analysis of isolates and environmental clones and by using community fingerprinting based on terminal restriction fragment length polymorphism (T-RFLP) analysis. Seven major bacterial taxa (six affiliated with the α-Proteobacteria and one with the Cytophagales) and four major fungal taxa were identified over five sample dates spanning 13 months. Fungal terminal restriction fragments (T-RFs) were informative at the species level; however, bacterial T-RFs frequently comprised a number of related genera. Amplicon abundances indicated that the salt marsh saprophyte communities have little-to-moderate variability spatially or with decomposition stage, but considerable variability temporally. However, the temporal variability could not be readily explained by either successional shifts or simple relationships with environmental factors. Significant correlations in abundance (both positive and negative) were found among dominant fungal and bacterial taxa that possibly indicate ecological interactions between decomposer organisms. Most associations involved one of four microbial taxa: two groups of bacteria affiliated with the α-Proteobacteria and two ascomycete fungi (Phaeosphaeria spartinicola and environmental isolate “4clt”).
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Chou, Chang-Hung, Yu-Chung Chiang e Tzen-Yuh Chiang. "Genetic variability and phytogeography of Miscanthus sinensis var. condensatus, an apomictic grass, based on RAPD fingerprints". Canadian Journal of Botany 78, n.º 10 (1 de outubro de 2000): 1262–68. http://dx.doi.org/10.1139/b00-102.
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DNA fingerprinting using random amplified polymorphic DNA (RAPD) markers was employed to investigate the genetic variation within and among populations of Miscanthus Anderss. sinensis var. condensatus (Hack.) Makino, an apomictic grass distributed along the coasts of Taiwan and Ryukyu Islands. A total of 250 plants from three Taiwanese populations (Southeast Coast, Orchid Islet, and Green Islet) and two populations from Ryukyu (Ishigaki and Amami-O-Shima Islets) were sampled. The amplified products of 40 random primers showed monomorphic banding patterns within all populations as well as among the three populations from Taiwan. Low genetic variation (with only two polymorphic loci), but significant differentiation, was detected between populations from Taiwan and Ryukyu (ΦCT = 0.864) and between populations (ΦST = 1.0) from Ishigaki and Amami-O-Shima Islets. In contrast, a high level of variation was exhibited in the outcrossing Miscanthus sinensis var. glaber (Nakai) Li. In addition to apomictic reproduction, low genetic variation across populations of M. sinensis var. condensatus may be a result of high salinity acting as a selective agent. With the cost of reduced genetic heterogeneity, apomixis may have provided a mechanism for avoiding the transmission of endophytic fungi. The phytogeographic pattern of M. sinensis var. condensatus, as reflected by the RAPD data, likely represents isolation between Taiwan and Ryukyu since the mid-Pleistocene.Key words: apomixis, Miscanthus sinensis var. condensatus, phytogeography, population differentiation, RAPD, system of mating.
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Turzhanova, Ainur, Oxana N. Khapilina, Asem Tumenbayeva, Vladislav Shevtsov, Olesya Raiser e Ruslan Kalendar. "Genetic diversity of Alternaria species associated with black point in wheat grains". PeerJ 8 (5 de maio de 2020): e9097. http://dx.doi.org/10.7717/peerj.9097.
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The genus Alternaria is a widely distributed major plant pathogen that can act as a saprophyte in plant debris. Fungi of this genus frequently infect cereal crops and cause such diseases as black point and wheat leaf blight, which decrease the yield and quality of cereal products. A total of 25 Alternaria sp. isolates were collected from germ grains of various wheat cultivars from different geographic regions in Kazakhstan. We investigated the genetic relationships of the main Alternaria species related to black point disease of wheat in Kazakhstan, using the inter-primer binding site (iPBS) DNA profiling technique. We used 25 retrotransposon-based iPBS primers to identify the differences among and within Alternaria species populations, and analyzed the variation using clustering (UPGMA) and statistical approaches (AMOVA). Isolates of Alternaria species clustered into two main genetic groups, with species of A.alternata and A.tennuissima forming one cluster, and isolates of A. infectoria forming another. The genetic diversity found using retrotransposon profiles was strongly correlated with geographic data. Overall, the iPBS fingerprinting technique is highly informative and useful for the evaluation of genetic diversity and relationships of Alternaria species.
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Prince, James P., e Steven D. Tanksley. "Restriction fragment length polymorphisms in plant breeding and genetics". Proceedings of the Royal Society of Edinburgh. Section B. Biological Sciences 99, n.º 3-4 (1992): 23–29. http://dx.doi.org/10.1017/s0269727000005479.
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SynopsisThe usefulness of restriction fragment length polymorphisms (RFLPs) in plant breeding and genetics is discussed, with particular emphasis on tagging genes, map-based cloning, the assessment of genetic variability and distances, and comparative genome mapping.The Department of Plant Breeding and Biometry has currently established tight linkages between RFLPs and more than 20 genes of economic importance. Approximately half of these genes confer resistance to major pathogens including nematodes, bacteria, fungi, and viruses. Other genes tagged are involved in various aspects of crop quality.Locating genes with respect to DNA markers on an RFLP map provides a starting point for cloning the genes by chromosome walking. This strategy is currently being pursued for three disease-resistance genes that have been placed on the tomato RFLP map; Pto (resistance to Pseudomonas syringae), Mi (resistance to root-knot nemotodes) and Tm2a (resistance to tobacco mosaic virus). Further discussion will include the construction of a yeast artificial chromosome library and the collection of additional DNA markers in the regions of interest through RAPD analysis of nearly isogenic lines.The assessment of genetic variability and fingerprinting varieties based on RFLP data will be briefly discussed.Comparative genome mapping in the family Solanaceae has allowed the relationships among tomato, potato, and pepper to be unravelled. Tomato and potato share a near perfect conservation of gene order throughout their genomes. In contrast, while pepper shares most of its single copy DNA with tomato and potato, the order of these markers is highly rearranged compared with the other two species.
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Somai, Benesh M., Ralph A. Dean, Mark W. Farnham, Thomas A. Zitter e Anthony P. Keinath. "Internal Transcribed Spacer Regions 1 and 2 and Random Amplified Polymorphic DNA Analysis of Didymella bryoniae and Related Phoma Species Isolated from Cucurbits". Phytopathology® 92, n.º 9 (setembro de 2002): 997–1004. http://dx.doi.org/10.1094/phyto.2002.92.9.997.
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Didymella bryoniae (anamorph Phoma cucurbitacearum) is the causal agent of gummy stem blight, although other Phoma species are often isolated from cucurbit plants exhibiting symptoms of the disease. The molecular and phylogenetic relationships between D. bryoniae and these Phoma species are unknown. Isolates of D. bryoniae and Phoma obtained from cucurbits grown at various geographical locations in the United States were subjected to random amplified polymorphic DNA (RAPD) analysis and internal transcribed spacer (ITS) sequence analysis (ITS-1 and ITS-2) to determine the molecular and phylogenetic relationships within and between these fungi. Using RAPD fingerprinting, 59 isolates were placed into four phylogenetic groups, designated RAPD group (RG) I, RG II, RG III, and RG IV. D. bryoniae isolates clustered in either RG I (33 isolates), RG II (12 isolates), or RG IV (one isolate), whereas all 13 Phoma isolates clustered to RG III. There was greater than 99% sequence identity in the ITS-1 and ITS-2 regions between isolates in RG I and RG II, whereas isolates in RG III, P. medicaginis ATCC 64481, and P. exigua ATCC 14728 clustered separately. On muskmelon seedlings, a subset of RG I isolates were highly virulent (mean disease severity was 71%), RG II and RG IV isolates were slightly virulent (mean disease severity was 4%), and RG III isolates were nonpathogenic (disease severity was 0% for all isolates). The ITS sequences indicate that RG I and RG II are both D. bryoniae, but RAPD fingerprints and pathogenicity indicate that they represent two different molecular and virulence subgroups.
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Trung, Trịnh Thành, Đinh Thị Tuyết Vân, Nguyễn Phương Liên, Đào Thị Lương e Dương Văn Hợp Dương Văn Hợp Dương Văn Hợp. "Potential application on preparation for bio-fertilizer using Bacillus velezensis strains isolated from various regions in Vietnam". Vietnam Journal of Biotechnology 15, n.º 1 (20 de abril de 2018): 169–79. http://dx.doi.org/10.15625/1811-4989/15/1/12332.
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Bacillus velezensis is a species belonging to the bacterial group of B. subtilis species complex. Due to various beneficial traits to plants, the bacterium has been received considerable attention for application in disease control and crop productivity. In this study, we isolated 15 B. velezensis strains from various soils collected in regions of Hoang Lien, Cuc Phuong, Bach Ma, Chu Yang Sin and Con Dao. All of the strains demonstrated antagonistic activity against phythopathogenic fungi of Fusarium oxysporum, Sclerotium hydrophilum, Rhizoctonia solani and Phytophthora capsici and rice blight bacterium Xanthomonas oryzae. Those strains were capable of solubilizing Ca3(PO4)2 and producing plant growth hormone of indole acetic acid (IAA). Moreover, those strains produced a series of extracellular hydrolytic enzymes such as lipase, protease, amylase, CMCase, xylanase and chitinase. Analysis of genetic diversity based on rep - PCR fingerprinting technique showed that primer ERIC had greater discrimination than primer GTG5. Number of DNA banding patterns generated by primer ERIC and GTG5 were 8 and 13, respectively. Combination of DNA banding patterns of both primers showed 14 different genotypes among 15 B. velezensis strains. Using multilocus sequencing technique, genetic diversity was confirmed by a clear discrimination of strain CP 1604 and SP 1901 in two different clusters on phylogenetic tree. Due to the possession of many beneficial traits to plant growth, pesticide protection and production of many hydrolytic enzymes, the B. velezensis strains in this study have potential application on preparation of bio-fertilizer. Further investigation is needed in order to select an appropriate strain for this application.
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SENGUPTA, RAJIB, DHUNDY R. BASTOLA e HESHAM H. ALI. "CLASSIFICATION AND IDENTIFICATION OF FUNGAL SEQUENCES USING CHARACTERISTIC RESTRICTION ENDONUCLEASE CUT ORDER". Journal of Bioinformatics and Computational Biology 08, n.º 02 (abril de 2010): 181–98. http://dx.doi.org/10.1142/s0219720010004616.
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Restriction Fragment Length Polymorphism (RFLP) is a powerful molecular tool that is extensively used in the molecular fingerprinting and epidemiological studies of microorganisms. In a wet-lab setting, the DNA is cut with one or more restriction enzymes and subjected to gel electrophoresis to obtain signature fragment patterns, which is utilized in the classification and identification of organisms. This wet-lab approach may not be practical when the experimental data set includes a large number of genetic sequences and a wide pool of restriction enzymes to choose from. In this study, we introduce a novel concept of Enzyme Cut Order — a biological property-based characteristic of DNA sequences which can be defined and analyzed computationally without any alignment algorithm. In this alignment-free approach, a similarity matrix is developed based on the pairwise Longest Common Subsequences (LCS) of the Enzyme Cut Orders. The choice of an ideal set of restriction enzymes used for analysis is augmented by using genetic algorithms. The results obtained from this approach using internal transcribed spacer regions of rDNA from fungi as the target sequence show that the phylogenetically-related organisms form a single cluster and successful grouping of phylogenetically close or distant organisms is dependent on the choice of restriction enzymes used in the analysis. Additionally, comparison of trees obtained with this alignment-free and the legacy method revealed highly similar tree topologies. This novel alignment-free method, which utilizes the Enzyme Cut Order and restriction enzyme profile, is a reliable alternative to local or global alignment-based classification and identification of organisms.
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Köhl, J., B. H. Groenenboom-de Haas, P. Kastelein, V. Rossi e C. Waalwijk. "Quantitative Detection of Pear-Pathogenic Stemphylium vesicarium in Orchards". Phytopathology® 99, n.º 12 (dezembro de 2009): 1377–86. http://dx.doi.org/10.1094/phyto-99-12-1377.
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Isolates of Stemphylium vesicarium causing brown spot of pear can be distinguished from nonpathogenic isolates of S. vesicarium from pear or from other hosts on the basis of distinctive amplified fragment length polymorphism fingerprinting profiles. DNA fragments specific for isolates pathogenic to pear were identified and a quantitative polymerase chain reaction (PCR) was developed on the sequence from one of these specific DNA loci. This TaqMan PCR has a high sensitivity with a dynamic range for reliable quantification between 1 ng and 100 fg of DNA. The method detected pear-pathogenic isolates of S. vesicarium originating from four different European countries and various regions within those countries. No cross-reaction was found with either the nonpathogenic isolates of S. vesicarium tested or isolates belonging to other Stemphylium spp. or related fungi. The pathogen was detected on leaves with brown-spot symptoms originating from six different locations in The Netherlands, Italy, and Spain. Pear-pathogenic S. vesicarium populations were monitored on crop residues in two Dutch orchards between October 2007 and October 2008. Brown spot had been observed at both orchards at the end of the growing season of 2007. In one location, pear-pathogenic S. vesicarium was detected only sporadically on crop residues and no brown-spot symptoms were observed on fruit in 2008. At the other location, a pathogenic population was found on fallen pear leaves and on other crop residues but this population decreased during winter. From the beginning of the growing season in 2008 onward, the pathogen population could not be detected and the disease incidence was only 0.6%. The TaqMan PCR will allow more detailed studies on epidemiology of brown spot and on the effect of disease control measures.
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Rovná, Katarína, Eva Ivanišová, Jana Žiarovská, Peter Ferus, Margarita Terentjeva, Przemysław Łukasz Kowalczewski e Miroslava Kačániová. "Characterization of Rosa canina Fruits Collected in Urban Areas of Slovakia. Genome Size, iPBS Profiles and Antioxidant and Antimicrobial Activities". Molecules 25, n.º 8 (19 de abril de 2020): 1888. http://dx.doi.org/10.3390/molecules25081888.
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The studies of plant bacterial endophytes, colonizing the plant tissues without any signs of diseases, are essential for understanding of ecological interactions. The aim of our study is to detect microbiological contamination and to assess the antimicrobial, antioxidant activity, total phenolic, carotenoid content, genome size, and ploidy of non-cultivated Rosa canina sampled from urban areas. Samples of Rosa canina fruits were collected in three locations in Slovakia. The highest total viable count and the Enterobacteriaceae count in fruits were 4.32 log CFU/g and 4.29 log CFU/g, respectively. Counts of the mesophilic anaerobic sporulating bacteria, Pseudomonas spp., and of the microscopic fungi and yeasts were 3.00, 2.15 log CFU/g, 3.65 log CFU/g, and 2.76 log CFU/g, respectively. Regarding the antimicrobial activity, Escherichia coli and Klebsiela oxytoca were the most sensitive species among the assayed microorganisms to the treatment with the ethanolic extracts of Rosa canina fruits. The fruits were rich in bioactive compounds, polyphenols, and carotenoids, that could be related to their antioxidant activity. Genome sizes of analyzed samples ranged from 2.3 to 2.96. DNA-based fingerprinting obtained by iPBS markers of the Rosa canina var. lapidicola Heinr. Braun., was characterized by some distinctive inserted loci. An interdisciplinary study was performed for the dog roses from different parts of Slovakia that resulted in deeper characterization of this species.
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Knapp, B. A., J. Seeber, S. M. Podmirseg, E. Meyer e H. Insam. "Application of denaturing gradient gel electrophoresis for analysing the gut microflora ofLumbricus rubellusHoffmeister under different feeding conditions". Bulletin of Entomological Research 98, n.º 3 (28 de abril de 2008): 271–79. http://dx.doi.org/10.1017/s0007485308006056.
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AbstractThe earthworm,Lumbricus rubellus, plays an essential role in soil ecosystems as it affects organic matter decomposition and nutrient cycling. By ingesting a mixture of organic and mineral material, a variety of bacteria and fungi are carried to the intestinal tract of the earthworm. To get a better understanding of the interactions betweenL. rubellusand the microorganisms ingested, this study tried to reveal if the diet affects the composition of the gut microflora ofL. rubellusor if its intestinal tract hosts an indigenous, species-specific microbiota. A feeding experiment withL. rubelluswas set up; individuals were collected in the field, transferred to a climate chamber and fed with food sources of different quality (dwarf shrub litter, grass litter or horse dung) for six weeks. DNA was extracted from the guts of the earthworms, as well as from the food sources and the surrounding soil, and further analysed by a molecular fingerprinting method, PCR-DGGE (Polymerase Chain Reaction – Denaturing Gradient Gel Electrophoresis). We were able to demonstrate that the gut microbiota was strongly influenced by the food source ingested and was considerably different to that of the surrounding soil. Sequencing of dominant bands of the bacterial DGGE fingerprints revealed a strong occurrence of y-Proteobacteria in all gut samples, independent of the food source. A specific microflora in the intestinal tract ofL. rubellus, robust against diet changes, could not be found.
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Brown, Eric, e Emma Allen-Vercoe. "Analysis of the fungal, archaeal and bacteriophage diversity in the human distal gut". SURG Journal 4, n.º 2 (11 de março de 2011): 75–82. http://dx.doi.org/10.21083/surg.v4i2.1331.
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The composition and role of bacteria in the human gut has been studied intensely and is a burgeoning field of scientific research. However, there is a relative lack of research on other microorganisms which compose our gut flora such as bacteriophage, archaea and fungi. The aim of our study was to begin to fill this gap. The archaeal, fungal and bacteriophage diversity in the gut was analyzed using a PCR-DGGE fingerprinting method on fecal samples from 3 healthy donors. These samples were inoculated into chemostats and the microbes were grown in continuous culture to model the interactions of our flora in vitro. Norepinephrine was also added to the chemostats to test the microbial community’s reaction to stress. Here we report that, relative to bacteria, fungal and archaeal diversity in the gut is low. The archaeal populations seemed stable over time varied depending on the individual. Fungal populations were more variable over time and changes in the community structure were observed after the addition of norepinephrine. DNA sequence analysis confirmed the presence of fungal species that are not yet cultured, yet are residents of the gut. Species of Podophage can also be detected as residents of the gut based on sequence analysis. It is clear that there is a core set of archaeal and fungal species living as residents in the gut. Bacteriophage are also present but their ecological role and effect on the microbial community in the gut is unknown.
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du PLESSIS, ERIKA M., FRANCOIS DUVENAGE e LISE KORSTEN. "Determining the Potential Link between Irrigation Water Quality and the Microbiological Quality of Onions by Phenotypic and Genotypic Characterization of Escherichia coli Isolates". Journal of Food Protection 78, n.º 4 (1 de abril de 2015): 643–51. http://dx.doi.org/10.4315/0362-028x.jfp-14-486.
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The potential transfer of human pathogenic bacteria present in irrigation water onto fresh produce was investigated, because surface water sources used for irrigation purposes in South Africa have increasingly been reported to be contaminated with enteric bacterial pathogens. A microbiological analysis was performed of a selected river in Limpopo Province, South Africa, that is often contaminated with raw sewage from municipal sewage works and overhead irrigated onions produced on a commercial farm. Counts of Escherichia coli, coliforms, aerobic bacteria, fungi, and yeasts and the prevalence of E. coli O157:H7, Salmonella, and Listeria monocytogenes were determined. Identities of bacterial isolates from irrigation water and onions were confirmed using matrix-assisted laser desorption ionization–time of flight mass spectrometry, PCR, and biochemical tests. To establish a potential link between the microbiological quality of the irrigation source and the onions, the E. coli isolates from both were subjected to antibiotic resistance, virulence gene, and enterobacterial repetitive intergenic consensus PCR analyses. River water E. coli counts exceeded South African Department of Water Affairs and World Health Organization irrigation water guidelines. Counts of aerobic bacteria, coliforms, fungi, and yeasts of onions from the market were acceptable according to Department of Health Directorate, Food Control, South Africa, microbiological guidelines for ready-to-eat fresh fruits and vegetables. E. coli O157:H7, Salmonella, and L. monocytogenes were not detected in onions, whereas only Salmonella was detected in 22% of water samples. Matrix-assisted laser desorption ionization–time of flight mass spectrometry and PCR identification of E. coli isolates from water and onions correlated. Of the 45 E. coli isolates from water and onions, 42.2% were resistant to multiple antibiotics. Virulence genes eae, stx1, and stx2 were detected in 2.2, 6.6, and 2.2% of the E. coli isolates, respectively. Phenotypic (antimicrobial) and genotypic (virulence gene prevalence, DNA fingerprinting) analyses showed a link between river, dam, irrigation pivot point, and onion E. coli isolates.
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Vohník, Martin. "Bioerosion and fungal colonization of the invasive foraminiferan <i>Amphistegina lobifera</i> in a Mediterranean seagrass meadow". Biogeosciences 18, n.º 8 (30 de abril de 2021): 2777–90. http://dx.doi.org/10.5194/bg-18-2777-2021.
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Abstract. Foraminiferans are diverse micro- to macroscopic protists abundant especially in (sub)tropical seas, often forming characteristic benthic communities known as “living sands”. Numerous species have migrated through the Suez Canal to the Mediterranean and one of them, i.e., Amphistegina lobifera, turned invasive, gradually outcompeting the indigenous species. At some places, A. lobifera creates thick seabed sediments, thus becoming an important environmental engineer. However, little is known about the turnover of its shells in the invaded ecosystems. Using vital staining, stereomicroscopy, scanning electron microscopy, and cultivation and DNA fingerprinting, I investigated the vital status, destruction/decomposition and mycobiota of A. lobifera in the rhizosphere of the dominant Mediterranean seagrass Posidonia oceanica in an underwater Maltese meadow (average 284 shells g−1, representing 28.5 % of dry substrate weight), in comparison with epiphytic specimens and P. oceanica roots. While 78 % of the epiphytes were alive, nearly all substrate specimens were dead. On average, 80 % of the epiphytes were intact compared to 21 % of the substrate specimens. Abiotic dissolution and mechanical damage played only a minor role, but some bioerosion was detected in 18 % and >70 % of the epiphytic and substrate specimens, respectively. Few bioerosion traces could be attributed to fungi, and the majority probably belonged to photoautotrophs. The seagrass roots displayed fungal colonization typical for this species and yielded 81 identified isolates, while the surface-sterilized substrate specimens surprisingly yielded no cultivable fungi compared to 16 other identified isolates obtained from the epiphytes. While the epiphytes' mycobiota was dominated by ascomycetous generalists also known from terrestrial ecosystems (alongside with, for example, a relative of the “rock-eating” extremophiles), the roots were dominated by the seagrass-specific dark septate endophyte Posidoniomyces atricolor and additionally contained a previously unreported lulworthioid mycobiont. In conclusion, at the investigated locality, dead A. lobifera shells seem to be regularly bioeroded by endolithic non-fungal organisms, which may counterbalance their accumulation in the seabed substrate.
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ben Omar, Nabil, e Fr�d�ric Ampe. "Microbial Community Dynamics during Production of the Mexican Fermented Maize Dough Pozol". Applied and Environmental Microbiology 66, n.º 9 (1 de setembro de 2000): 3664–73. http://dx.doi.org/10.1128/aem.66.9.3664-3673.2000.
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ABSTRACT The dynamics of the microbial community responsible for the traditional fermentation of maize in the production of Mexican pozol was investigated by using a polyphasic approach combining (i) microbial enumerations with culture media, (ii) denaturing gradient gel electrophoresis (DGGE) fingerprinting of total community DNA with bacterial and eukaryotic primers and sequencing of partial 16S ribosomal DNA (rDNA) genes, (iii) quantification of rRNAs from dominant microbial taxa by using phylogenetic oligonucleotide probes, and (iv) analysis of sugars and fermentation products. AStreptococcus species dominated the fermentation and accounted for between 25 and 75% of the total flora throughout the process. Results also showed that the initial epiphytic aerobic microflora was replaced in the first 2 days by heterofermentative lactic acid bacteria (LAB), including a close relative ofLactobacillus fermentum, producing lactic acid and ethanol; this heterolactic flora was then progressively replaced by homofermentative LAB (mainly close relatives of L. plantarum, L. casei, and L. delbrueckii) which continued acidification of the maize dough. At the same time, a very diverse community of yeasts and fungi developed, mainly at the periphery of the dough. The analysis of the DGGE patterns obtained with bacterial and eukaryotic primers targeting the 16S and 18S rDNA genes clearly demonstrated that there was a major shift in the community structure after 24 h and that high biodiversity—according to the Shannon-Weaver index—was maintained throughout the process. These results proved that a relatively high number of species, at least six to eight, are needed to perform this traditional lactic acid fermentation. The presence ofBifidobacterium, Enterococcus, and enterobacteria suggests a fecal origin of some important pozol microorganisms. Overall, the results obtained with different culture-dependent or -independent techniques clearly confirmed the importance of developing a polyphasic approach to study the ecology of fermented foods.
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de Boer, Wietse, Johan H. J. Leveau, George A. Kowalchuk, Paulien J. A. Klein Gunnewiek, Edwin C. A. Abeln, Marian J. Figge, Klaas Sjollema, Jaap D. Janse e Johannes A. van Veen. "Collimonas fungivorans gen. nov., sp. nov., a chitinolytic soil bacterium with the ability to grow on living fungal hyphae". International Journal of Systematic and Evolutionary Microbiology 54, n.º 3 (1 de maio de 2004): 857–64. http://dx.doi.org/10.1099/ijs.0.02920-0.
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A polyphasic approach was used to describe the phylogenetic position of 22 chitinolytic bacterial isolates that were able to grow at the expense of intact, living hyphae of several soil fungi. These isolates, which were found in slightly acidic dune soils in the Netherlands, were strictly aerobic, Gram-negative rods. Cells grown in liquid cultures were flagellated and possessed pili. A wide range of sugars, alcohols, organic acids and amino acids could be metabolized, whereas several di- and trisaccharides could not be used as substrates. The major cellular fatty acids were C16 : 0, C16 : 1 ω7c and C18 : 1 ω7c. DNA G+C contents were 57–62 mol%. Analysis of nearly full-length 16S rDNA sequences showed that the isolates were related closely to each other (>98·6 % sequence similarity) and could be assigned to the β-Proteobacteria, family ‘Oxalobacteraceae’, order ‘Burkholderiales’. The most closely related species belonged to the genera Herbaspirillum and Janthinobacterium, exhibiting 95·9–96·7 % (Herbaspirillum species) and 94·3–95·6 % (Janthinobacterium species) 16S rDNA sequence similarity to the isolates. Several physiological and biochemical properties indicated that the isolates could be distinguished clearly from both of these genera. Therefore, it is proposed that the isolates described in this study are representatives of a novel genus, Collimonas gen. nov. Genomic fingerprinting (BOX-PCR), detailed analysis of 16S rDNA patterns and physiological characterization (Biolog) of the isolates revealed the existence of four subclusters. The name Collimonas fungivorans gen. nov., sp. nov. has been given to one subcluster (four isolates) that appears to be in the centre of the novel genus; isolates in the other subclusters have been tentatively named Collimonas sp. The type strain of Collimonas fungivorans gen. nov., sp. nov. is Ter6T (=NCCB 100033T=LMG 21973T).