Phylogenetic characterization and in situ detection of a Cytophaga-Flexibacter-Bacteroides phylogroup bacterium in Tuber borchii vittad. Ectomycorrhizal mycelium.
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Mycorrhizal ascomycetous fungi are obligate ectosymbionts that colonize the roots of gymnosperms and angiosperms. In this paper we describe a straightforward approach in which a combination of morphological and molecular methods was used to survey the presence of potentially endo- and epiphytic bacteria associated with the ascomycetous ectomycorrhizal fungus Tuber borchii Vittad. Universal eubacterial primers specific for the 5' and 3' ends of the 16S rRNA gene (16S rDNA) were used for PCR amplification, direct sequencing, and phylogenetic analyses. The 16S rDNA was amplified directly from four pure cultures of T. borchii Vittad. mycelium. A nearly full-length sequence of the gene coding for the prokaryotic small-subunit rRNA was obtained from each T. borchii mycelium studied. The 16S rDNA sequences were almost identical (98 to 99% similarity), and phylogenetic analysis placed them in a single unique rRNA branch belonging to the Cytophaga-Flexibacter-Bacteroides (CFB) phylogroup which had not been described previously. In situ detection of the CFB bacterium in the hyphal tissue of the fungus T. borchii was carried out by using 16S rRNA-targeted oligonucleotide probes for the eubacterial domain and the Cytophaga-Flexibacter phylum, as well as a probe specifically designed for the detection of this mycelium-associated bacterium. Fluorescent in situ hybridization showed that all three of the probes used bound to the mycelium tissue. This study provides the first direct visual evidence of a not-yet-cultured CFB bacterium associated with a mycorrhizal fungus of the genus Tuber. (+info)
Comparative 16S rRNA analysis of lake bacterioplankton reveals globally distributed phylogenetic clusters including an abundant group of actinobacteria.
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In a search for cosmopolitan phylogenetic clusters of freshwater bacteria, we recovered a total of 190 full and partial 16S ribosomal DNA (rDNA) sequences from three different lakes (Lake Gossenkollesee, Austria; Lake Fuchskuhle, Germany; and Lake Baikal, Russia). The phylogenetic comparison with the currently available rDNA data set showed that our sequences fall into 16 clusters, which otherwise include bacterial rDNA sequences of primarily freshwater and soil, but not marine, origin. Six of the clusters were affiliated with the alpha, four were affiliated with the beta, and one was affiliated with the gamma subclass of the Proteobacteria; four were affiliated with the Cytophaga-Flavobacterium-Bacteroides group; and one was affiliated with the class Actinobacteria (formerly known as the high-G+C gram-positive bacteria). The latter cluster (hgcI) is monophyletic and so far includes only sequences directly retrieved from aquatic environments. Fluorescence in situ hybridization (FISH) with probes specific for the hgcI cluster showed abundances of up to 1.7 x 10(5) cells ml(-1) in Lake Gossenkollesee, with strong seasonal fluctuations, and high abundances in the two other lakes investigated. Cell size measurements revealed that Actinobacteria in Lake Gossenkollesee can account for up to 63% of the bacterioplankton biomass. A combination of phylogenetic analysis and FISH was used to reveal 16 globally distributed sequence clusters and to confirm the broad distribution, abundance, and high biomass of members of the class Actinobacteria in freshwater ecosystems. (+info)
Identification and characterization of bacteria in a selenium-contaminated hypersaline evaporation pond.
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Solar evaporation ponds are commonly used to reduce the volume of seleniferous agricultural drainage water in the San Joaquin Valley, Calif. These hypersaline ponds pose an environmental health hazard because they are heavily contaminated with selenium (Se), mainly in the form of selenate. Se in the ponds may be removed by microbial Se volatilization, a bioremediation process whereby toxic, bioavailable selenate is converted to relatively nontoxic dimethylselenide gas. In order to identify microbes that may be used for Se bioremediation, a 16S ribosomal DNA phylogenetic analysis of an aerobic hypersaline pond in the San Joaquin Valley showed that a previously unaffiliated group of uncultured bacteria (belonging to the order Cytophagales) was dominant, followed by a group of cultured gamma-Proteobacteria which was closely related to Halomonas species. Se K-edge X-ray absorption spectroscopy of selenate-treated bacterial isolates showed that they accumulated a mixture of predominantly selenate and a selenomethionine-like species, consistent with the idea that selenate was assimilated via the S assimilation pathway. One of these bacterial isolates (Halomonas-like strain MPD-51) was the best candidate for the bioremediation of hypersaline evaporation ponds contaminated with high Se concentrations because it tolerated 2 M selenate and 32.5% NaCl, grew rapidly in media containing selenate, and accumulated and volatilized Se at high rates (1.65 microg of Se g of protein(-1) x h(-1)), compared to other cultured bacterial isolates. (+info)
New degenerate Cytophaga-Flexibacter-Bacteroides-specific 16S ribosomal DNA-targeted oligonucleotide probes reveal high bacterial diversity in River Taff epilithon.
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River microbial communities play an important role in global nutrient cycles, and aggregated bacteria such as those in epilithic biofilms may be major contributors. In this study the bacterial diversity of River Taff epilithon in South Wales was investigated. A 16S ribosomal DNA (rDNA) clone library was constructed and analyzed by partial sequencing of 76 of 347 clones and hybridization with taxon-specific probes. The epilithon was found to be very diverse, with an estimated 59.6% of the bacterial populations not accounted for by these clones. Members of the Cytophaga-Flexibacter-Bacteroides division (CFBs) were most abundant in the library, representing 25% of clones, followed by members of the alpha subdivision of the division Proteobacteria (alpha-Proteobacteria), gamma-Proteobacteria, gram-positive bacteria, Cyanobacteria, beta-Proteobacteria, delta-Proteobacteria, and the Prosthecobacter group. This study concentrated on the epilithic CFB populations, and a new set of degenerate 16S rDNA probes was developed to enhance their detection, namely, CFB560, CFB562, and CFB376. The commonly used probe CF319a/b may frequently lead to the underestimation of CFB populations in environmental studies, because it does not fully detect members of the division. CFB560 had exact matches to 95.6% of CFBs listed in the Ribosomal Database Project (release 8.0) small-subunit phylogenetic trees, compared to 60% for CF319a/b. The CFB probes detected 66 of 347 epilithon TAF clones, and 60 of these were partially sequenced. They affiliated with the RDP-designated groups Cytophaga, Sphingobacterium, Lewinella, and Cytophaga aurantiaca. CFB560 and CF319a/b detected 94% (62 of 66) and 48.5% (32 of 66) of clones, respectively, and therefore CFB560 is recommended for future use. Probe design in this study illustrated that multiple degenerate positions can greatly increase target range without adversely effecting specificity or experimental performance. (+info)
Pedobacter cryoconitis sp. nov., a facultative psychrophile from alpine glacier cryoconite.
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On the basis of phenotypic and genotypic characteristics, a novel species belonging to the genus Pedobacter is described. A facultatively psychrophilic, Gram-negative, aerobic, rod-shaped strain, A37(T), was isolated from alpine glacier cryoconite. The non-flagellated and non-spore-forming isolate grew over a temperature range of 1-25 degrees C, showed activities of oxidase, catalase, DNase, protease (gelatin, casein), amylase, beta-glucosidase, beta-galactosidase and beta-lactamase and degraded oil hydrocarbons. A distinct optimum temperature of 15 degrees C was observed for both protease production and oil hydrocarbon biodegradation. Analysis of 16S rDNA revealed that strain A37(T) represents a distinct taxon within PEDOBACTER: DNA from strain A37(T) showed only 19.7 % genetic relatedness to the DNA of Pedobacter piscium. The DNA G+C content was 43.4 mol%. Dominant fatty acids (51 %) were iso-15 : 0 2-OH and 16 : 1omega7c. The strain is assigned to a novel Pedobacter species, for which the name Pedobacter cryoconitis sp. nov. is proposed, with A37(T) (=DSM 14825(T)=LMG 21415(T)) as the type strain. (+info)
Algoriphagus ratkowskyi gen. nov., sp. nov., Brumimicrobium glaciale gen. nov., sp. nov., Cryomorpha ignava gen. nov., sp. nov. and Crocinitomix catalasitica gen. nov., sp. nov., novel flavobacteria isolated from various polar habitats.
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Several cold-adapted strains isolated from a variety of algal-rich Antarctic and Southern Ocean samples formed three distinct groups within the class Flavobacteria, phylogenetically distant from other cultivated species. The first taxon, designated Algoriphagus ratkowskyi gen. nov., sp. nov., was isolated from sea ice and from saline lake cyanobacterial mats and includes non-motile, strictly aerobic, saccharolytic rod-like or serpentine strains that were most closely related to the genus Cyclobacterium according to 16S rDNA sequence analysis (sequence similarity 0.85). The second taxon, designated Brumimicrobium glaciale gen. nov., sp. nov., isolated from sea ice and from continental shelf sediment, formed gliding, rod-like cells that were facultatively anaerobic with a fermentative metabolism. The third taxon, designated Cryomorpha ignava gen. nov., sp. nov., isolated from Southern Ocean particulates and from quartz stone subliths, included strictly aerobic, pleomorphic rod-like cells. Brumimicrobium glaciale and Cryomorpha ignava were most closely allied with 'Microscilla aggregans var. catalatica', which, on the basis of its distinctive taxonomic traits, is also proposed as a new genus and species, Crocinitomix catalasitica gen. nov., sp. nov. It is proposed that the three genera Brumimicrobium, Cryomorpha and Crocinitomix belong to a new family, Cryomorphaceae fam. nov. (type genus Cryomorpha), as they possess generally similar morphological and ecophysiological characteristics and form a common and distinct clade within class FLAVOBACTERIA: (+info)
Dynamics of bacterial and fungal communities on decaying salt marsh grass.
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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 alpha-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 alpha-Proteobacteria and two ascomycete fungi (Phaeosphaeria spartinicola and environmental isolate "4clt"). (+info)
Flexibacter ovolyticus sp. nov., a pathogen of eggs and larvae of Atlantic halibut, Hippoglossus hippoglossus L.
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A psychrotrophic Flexibacter sp., Flexibacter ovolyticus sp. nov., was isolated from the adherent bacterial epiflora of Atlantic halibut (Hippoglossus hippoglossus L.) eggs and was shown to be an opportunistic pathogen for halibut eggs and larvae. The strains which we isolated had the enzymatic capacity to dissolve both the chorion and the zona radiata of the egg shells. A total of 35 isolates were characterized by using morphological and biochemical tests. These strains were rod shaped, gram negative, Kovacs oxidase positive, and pale yellow and exhibited gliding motility. They did not produce acid from any of the wide range of carbohydrates tested. Our isolates had the ability to degrade gelatin, tyrosine, DNA, and Tween 80. Starch, cellulose, and chitin were not degraded. The strains were catalase and nitrate reductase positive, did not produce H2S, and did not grow under anaerobic conditions. F. ovolyticus resembles Flexibacter maritimus, but differs from the latter species in several biochemical and physiological characteristics. DNAs from F. ovolyticus strains had guanine-plus-cytosine contents which ranged from 30.3 to 32.0 mol% (strains EKC001, EKD002T [T = type strain], and VKB004), and DNA-DNA hybridization studies revealed levels of relatedness between F. ovolyticus EKD002T and F. maritimus NCMB 2154T and NCMB 2153 of 42.7 and 30.0%, respectively. Compared with previously described Cytophaga and Flexibacter spp. with low guanine-plus-cytosine contents, F. ovolyticus constitutes a new species. Strain EKD002 (= NCIMB 13127) is the type strain of the new species. (+info)