Phylogenetic diversity of microbial communities of the Posolsk Bank bottom sediments,Lake Baikal

Massive parallel sequencing (the Roche 454 platform) of the 16S rRNA gene fragments was used to investigate microbial diversity in the sediments of the Posolsk Bank cold methane seep. Bacterial communities from all sediment horizons were found to contain members of the phyla Actinobacteria, Bacteroidetes, Deinococcus-Thermus, Firmicutes, Nitrospirae, Chloroflexi, Proteobacteria, and the candidate phyla Aminicenantes (OP8) and Atribacteria (OP9). Among Bacteria, members of the Chloroflexi and Proteobacteria were the most numerous (42 and 46%, respectively). Among archaea, the Thaumarchaeota predominated in the upper sediment layer (40.1%), while Bathyarchaeota (54.2%) and Euryarchaeota (95%) were predominant at 70 and 140 cm, respectively. Specific migration pathways of fluid flows circulating in the zone of gas hydrate stability (400 m) may be responsible for considerable numbers of the sequences of Chloroflexi, Acidobacteria, and the candidate phyla Aminicenantes and Atribacteria in the upper sediment layers and of the Deinococcus-Thermus phylum in deep bottom sediments.     

Phylogenetic analysis of the bacterial communities in marine sediments.

For the phylogenetic analysis of microbial communities present in environmental samples microbial DNA can be extracted from the sample, 16S rDNA can be amplified with suitable primers and the PCR, and clonal libraries can be constructed. We report a protocol that can be used for efficient cell lysis and recovery of DNA from marine sediments. Key steps in this procedure include the use of a bead mill homogenizer for matrix disruption and uniform cell lysis and then purification of the released DNA by agarose gel electrophoresis. For sediments collected from two sites in Puget Sound, over 96% of the cells present were lysed. Our method yields high-molecular-weight DNA that is suitable for molecular studies, including amplification of 16S rRNA genes. The DNA yield was 47 micrograms per g (dry weight) for sediments collected from creosote-contaminated Eagle Harbor, Wash. Primers were selected for the PCR amplification of (eu)bacterial 16S rDNA that contained linkers with unique 8-base restriction sites for directional cloning. Examination of 22 16S rDNA clones showed that the surficial sediments in Eagle Harbor contained a phylogenetically diverse population of organisms from the Bacteria domain (G. J. Olsen, C. R. Woese, and R. Overbeek, J. Bacteriol. 176:1-6, 1994) with members of six major lineages represented: alpha, delta, and gamma Proteobacteria; the gram-positive high G+C content subdivision; clostridia and related organisms; and planctomyces and related organisms. None of the clones were identical to any representatives in the Ribosomal Database Project small subunit RNA database. The analysis of clonal representives in the first report using molecular techniques to determine the phylogenetic composition of the (eu)bacterial community present in coastal marine sediments.     

北极太平洋扇区海洋沉积物细菌多样性的系统发育分析

对北极太平洋扇区3个不同深度的海洋沉积物样品,采用PCR结合变性梯度凝胶电泳(DGGE)技术进行细菌16S rRNA基因V3区序列的系统发育分析。结果表明,同一个沉积物样品不同层次的DGGE电泳图谱不完全相同。从3个沉积物样品中共获得50条序列,大部分序列与从海洋环境尤其海洋沉积物获得的细菌16S rDNA序列相似性较高(88%~100%),归属于变形细菌(Proteobacteria)的gamma亚群、alpha亚群、beta亚群、epsilon亚群、delta亚群,Cytophaga_Flavobacterium_Bacteroides(CFB)群细菌和高G C含量的革兰氏阳性细菌等系统分类群,其中变形细菌(Proteobacteria)的gamma亚群为沉积物中的优势细菌类群。     

Exploring the diversity of bacterial communities in sediments of urban mangrove forests

Municipal sewage, urban runoff and accidental oil spills are common sources of pollutants in urban mangrove forests and may have drastic effects on the microbial communities inhabiting the sediment. However, studies on microbial communities in the sediment of urban mangroves are largely lacking. In this study, we explored the diversity of bacterial communities in the sediment of three urban mangroves located in Guanabara Bay (Rio de Janeiro, Brazil). Analysis of sediment samples by means of denaturing gradient gel electrophoresis (DGGE) of 16S rRNA gene fragments suggested that the overall bacterial diversity was not significantly affected by the different levels of hydrocarbon pollution at each sampling site. However, DGGE and sequence analyses provided evidences that each mangrove sediment displayed a specific structure bacterial community. Although primer sets for Pseudomonas, alphaproteobacterial and actinobacterial groups also amplified ribotypes belonging to taxa not intended to be enriched, sequence analyses of dominant DGGE bands revealed ribotypes related to Alteromonadales, Burkholderiales, Pseudomonadales, Rhodobacterales and Rhodocyclales. Members of these groups were often shown to be involved in aerobic or anaerobic degradation of hydrocarbon pollutants. Many of these sequences were only detected in the sampling sites with high levels of anthropogenic inputs of hydrocarbons. Many dominant DGGE ribotypes showed low levels of sequence identity to known sequences, indicating a large untapped bacterial diversity in mangrove ecosystems.     

Phylogenetic diversity of bacterial and archaeal communities in the anoxic zone of the Cariaco Basin

Microbial community samples were collected from the anoxic zone of the Cariaco Basin at depths of 320, 500, and 1,310 m on a November 1996 cruise and were used to construct 16S ribosomal DNA libraries. Of 60 nonchimeric sequences in the 320-m library, 56 belonged to the epsilon subdivision of the Proteobacteria (epsilon-Proteobacteria) and 53 were closely related to ectosymbionts of Rimicaris exoculata and Alvinella pompejana, which are referred to here as epsilon symbiont relatives (ESR). The 500-m library contained sequences affiliated with the fibrobacteria, the Flexibacter-Cytophaga-Bacteroides division, the division Verrucomicrobia, the division Proteobacteria, and the OP3 candidate division. The Proteobacteria included members of the gamma, delta, epsilon and new candidate subdivisions, and gamma-proteobacterial sequences were dominant (25.6%) among the proteobacterial sequences. As in the 320-m library, the majority of the epsilon-proteobacteria belonged to the ESR group. The genus Fibrobacter and its relatives were the second largest group in the library (23.6%), followed by the delta-proteobacteria and the epsilon-proteobacteria. The 1,310-m library had the greatest diversity; 59 nonchimeric clones in the library contained 30 unique sequences belonging to the planctomycetes, the fibrobacteria, the Flexibacter-Cytophaga-Bacteroides division, the Proteobacteria, and the OP3 and OP8 candidate divisions. The proteobacteria included members of new candidate subdivisions and the beta, gamma, delta, and epsilon-subdivisions. ESR sequences were still present in the 1,310-m library but in a much lower proportion (8.5%). One archaeal sequence was present in the 500-m library (2% of all microorganisms in the library), and eight archaeal sequences were present in the 1,310-m library (13.6%). All archaeal sequences fell into two groups; two clones in the 1,310-m library belonged to the kingdom Crenarchaeota and the remaining sequences in both libraries belonged to the kingdom Euryarchaeota. The latter group appears to be related to the Eel-TA1f2 sequence, which belongs to an archaeon suggested to be able to oxidize methane anaerobically. Based on phylogenetic inferences and measurements of dark CO(2) fixation, we hypothesized that (i) the ESR are autotrophic anaerobic sulfide oxidizers, (ii) sulfate reduction and fermentative metabolism may be carried out by a large number of bacteria in the 500- and 1,310-m libraries, and (iii) members of the Euryarchaeota found in relatively large numbers in the 1,310-m library may be involved in anaerobic methane oxidation. Overall, the composition of microbial communities from the Cariaco Basin resembles the compositions of communities from several anaerobic sediments, supporting the hypothesis that the Cariaco Basin water column is similar to anaerobic sediments.     

Palynological characteristics of the bottom sediments of the southern Barents sea

The distribution of spores and pollen grains was studied in nine sections of bottom sediments up to 50 m thick, in the southern Barents Sea. The Early Cretaceous, Pleistocene, and Holocene ages of the enclosing sediments are determined by palynological analysis. Three Holocene provinces were recognized and traced along the beds, which were formed in the Early, Middle, and Late Holocene according to the scheme of Neishtadt (1957).     

Insights into the diversity and metabolic function of bacterial communities in sediments from Chilean salmon aquaculture sites

Aquaculture is an extremely valuable and rapidly expanding sector worldwide, but concerns exist related to environmental sustainability. The sediment below aquaculture farms receives inputs of antimicrobials, metal-containing products, and organic matter from uneaten food and fecal material. These inputs impact the surrounding marine microbial communities in complex ways; however, functional diversity shifts related to taxonomic composition remain poorly understood. Here, we investigated the effect of pollution from marine fish farms on sediment bacterial communities. We compared the bacterial communities and functional bacterial diversity in surface sediments at salmon aquaculture and reference sites in Chiloé, southern Chile, using Roche 454 pyrosequencing of the 16S ribosomal RNA (rRNA) gene and the predictive metagenomics approach (Phylogenetic Investigation of Communities by Reconstruction of Unobserved States, PICRUSt). Bacterial diversity, measured as the inverse Simpson index, was significantly lower at aquaculture than at reference sites, while species richness, based on Chao’s estimator, was not significantly different. Nevertheless, community composition differed significantly between reference and aquaculture sites. We found that Gammaproteobacteria and several taxa involved in remediating metal contamination and known to have antimicrobial resistances were enriched at aquaculture sites. However, PICRUSt predicted functions indicated a degree of functional redundancy between sites, whereas taxonomic-functional relationships indicated differences in the functional traits of specific taxa at aquaculture sites. This study provides a first step in understanding the bacterial community structure and functional changes due to Chilean salmon aquaculture and has direct implications for using bacterial shifts as indicators of aquaculture perturbations.     

Phylogenetic diversity of bacterial communities in bovine rumen as affected by diets and microenvironments

Phylogenetic analysis was conducted to examine ruminal bacteria in two ruminal fractions (adherent fraction vs. liquid fraction) collected from cattle fed with two different diets: forage alone vs. forage plus concentrate. One hundred forty-four 16S rRNA gene (rrs) sequences were obtained from clone libraries constructed from the four samples. These rrs sequences were assigned to 116 different operational taxonomic units (OTUs) defined at 0.03 phylogenetic distance. Most of these OTUs could not be assigned to any known genus. The phylum Firmicutes was represented by approximately 70% of all the sequences. By comparing to the OTUs already documented in the rumen, 52 new OTUs were identified. UniFrac, SONS, and denaturing gradient gel electrophoresis analyses revealed difference in diversity between the two fractions and between the two diets. This study showed that rrs sequences recovered from small clone libraries can still help identify novel species-level OTUs.     

Phylogenetic diversity of bacterial communities associated with bioremediation of crude oil in microcosms

Bioremediation, mainly by indigenous bacteria, has been regarded as an effective way to clean up oil pollution after an oil spill. In order to obtain a systematic understanding of the succession of bacterial communities associated with oil bioremediation, sediments collected from the Penglai 19-3 oil platform were co-incubated with crude oil. Oil biodegradation was assessed on the basis of changes in oil composition monitored by GC–MS. Changes in the bacterial community structure were detected by two 16S rRNA gene based culture-independent methods, denaturing gradient gel electrophoresis (DGGE) and clone library. The results suggested that crude oil was rapidly degraded during the 30-day bioremediation period. Bacteria affiliated with the genus Pseudomonas dominated all three clone libraries. But dramatic changes were also detected in the process of biodegradation of crude oil. The “professional hydrocarbonocastic bacteria” (e.g., Alcanivorax) became abundant in the two samples during the bioremediation period. Meanwhile, δ-proteobacteria was only detected in the two samples. Information on the bacterial community revealed in this study will be useful in developing strategies for bioremediation of crude oil dispersed in the marine ecosystem.     

东太湖表层沉积物放线菌群落结构多样性及其空间异质性

采用基于16SrRNA基因变性梯度凝胶电泳(DGGE)方法,考察东太湖表层沉积物放线菌群落结构多样性及其空间分布特征.结果显示:东太湖表层沉积物中放线菌群落结构多样性较高,DGGE指纹图谱样品的平均条带数为21.5±2个,群落结构多样性(如Shannon-Wiener指数)在空间尺度上差异不明显;聚类分析表明放线菌群落结构空间分布特征表现为相邻采样位点群落结构较为相似,这可能与相邻位点沉积物理化性质类似有关.典型对应分析结果表明,东太湖表层沉积物中放线菌群落结构变化的主要影响因子为沉积物pH.     

Seasonal and spatial diversity of microbial communities in marine sediments of the South China Sea

This study was conducted to characterize the diversity of microbial communities in marine sediments of the South China Sea by means of 16S rRNA gene clone libraries. The results revealed that the sediment samples collected in summer harboured a more diverse microbial community than that collected in winter, Deltaproteobacteria dominated 16S rRNA gene clone libraries from both seasons, followed by Gammaproteobacteria, Acidobacteria, Nitrospirae, Planctomycetes, Firmicutes. Archaea phylotypes were also found. The majority of clone sequences shared greatest similarity to uncultured organisms, mainly from hydrothermal sediments and cold seep sediments. In addition, the sedimentary microbial communities in the coastal sea appears to be much more diverse than that of the open sea. A spatial pattern in the sediment samples was observed that the sediment samples collected from the coastal sea and the open sea clustered separately, a novel microbial community dominated the open sea. The data indicate that changes in environmental conditions are accompanied by significant variations in diversity of microbial communities at the South China Sea.     

Phylogenetic diversity of gram-positive bacteria cultured from marine sediments

Major advances in our understanding of marine bacterial diversity have been gained through studies of bacterioplankton, the vast majority of which appear to be gram negative. Less effort has been devoted to studies of bacteria inhabiting marine sediments, yet there is evidence to suggest that gram-positive bacteria comprise a relatively large proportion of these communities. To further expand our understanding of the aerobic gram-positive bacteria present in tropical marine sediments, a culture-dependent approach was applied to sediments collected in the Republic of Palau from the intertidal zone to depths of 500 m. This investigation resulted in the isolation of 1,624 diverse gram-positive bacteria spanning 22 families, including many that appear to represent new taxa. Phylogenetic analysis of 189 representative isolates, based on 16S rRNA gene sequence data, indicated that 124 (65.6%) belonged to the class Actinobacteria while the remaining 65 (34.4%) were members of the class Bacilli. Using a sequence identity value of >/=98%, the 189 isolates grouped into 78 operational taxonomic units, of which 29 (37.2%) are likely to represent new taxa. The high degree of phylogenetic novelty observed during this study highlights the fact that a great deal remains to be learned about the diversity of gram-positive bacteria in marine sediments.     

Phylogenetic comparisons of bacterial communities from serpentine and nonserpentine soils

I present the results of a culture-independent survey of soil bacterial communities from serpentine soils and adjacent nonserpentine comparator soils using a variety of newly developed phylogenetically based statistical tools. The study design included site-based replication of the serpentine-to-nonserpentine community comparison over a regional scale ( approximately 100 km) in Northern California and Southern Oregon by producing 16S rRNA clone libraries from pairs of samples taken on either side of the serepentine-nonserpentine edaphic boundary at three geographical sites. At the division level, the serpentine and nonserpentine communities were similar to each other and to previous data from forest soils. Comparisons of both richness and Shannon diversity produced no significant differences between any of the libraries, but the vast majority of phylogenetically based tests were significant, even with only 50 sequences per library. These results suggest that most samples were distinct, consisting of a collection of lineages generally not found in other samples. The pattern of results showed that serpentine communities tended to be more similar to each other than they were to nonserpentine communities, and these differences were at a lower taxonomic scale. Comparisons of two nonserpentine communities generally showed differences, and some results suggest that the geographical site may control community composition as well. These results show the power of phylogenetic tests to discern differences between 16S rRNA libraries compared to tests that discard DNA data to bin sequences into operational taxonomic units, and they stress the importance of replication at larger scales for inferences regarding microbial biogeography.     

Phylogenetic diversity of bacterial and archaeal communities inhabiting the saline Lake Red located in Sovata, Romania

Lake Red is one of the saline lakes which were formed as a consequence of salt massif dissolution at the foot of the Gurghiu Mountains (Central Romania) at the end of the nineteenth century. The lake water had approximately 15?% w/v salt content. Phylogenetic diversity of prokaryotes inhabiting the water and sediment of the lake was studied using cultivation and cultivation-independent methods following a sampling in spring 2009. According to the results of 16S rRNA gene-based denaturing gradient gel electrophoresis (DGGE), the richness of Bacteria was higher than Archaea on the basis of the number and position of dominant bands in the gel. Sequences from DGGE bands were affiliated with Gammaproteobacteria (Halomonas and Alkalilimnicola) and Bacteroidetes (Psychroflexus) as well as Euryarchaeota. Cultivation from five different saline media resulted in 101 bacterial strains of which Gammaproteobacteria (Halomonas, Marinobacter and Salinivibrio) were the most abundant. Firmicutes (Bacillus) and Alphaproteobacteria (Aurantimonas and Roseovarius) were also identified among the isolated strains. The 16S rRNA genes from 82 bacterial and 95 archaeal clones were also phylogenetically analyzed. Bacterial clones were related to various genera of Gammaproteobacteria (Alkalilimnicola, Alkalispirillum, Arhodomonas, Halomonas, Saccharospirillum), Bacteroidetes (Gracilimonas, Psychroflexus) and Alphaproteobacteria (Oceanicola, Roseinatronobacter, Roseovarius). All of the archaeal clones sequenced corresponded to a homologous cluster affiliated with Halopelagius.     

Phylogenetic diversity of culturable bacteria from Antarctic sandy intertidal sediments

The diversity of culturable bacteria associated with sandy intertidal sediments from the coastal regions of the Chinese Antarctic Zhongshan Station on the Larsemann Hills (Princess Elizabeth Land, East Antarctica) was investigated. A total of 65 aerobic heterotrophic bacterial strains were isolated at 4°C. Microscopy and 16S rRNA gene sequence analysis indicated that the isolates were dominated by Gram-negative bacteria, while only 16 Gram-positive strains were isolated. The bacterial isolates fell in five phylogenetic groups: Alpha- and Gammaproteobacteria, Bacteroidetes, Actinobacteria and Firmicutes. Based on phylogenetic trees, all the 65 isolates were sorted into 29 main clusters, corresponding to at least 29 different genera. Based on sequence analysis (<98% sequence similarity), the Antarctic isolates belonged to at least 37 different bacterial species, and 14 of the 37 bacterial species (37.8%) represented potentially novel taxa. These results indicated a high culturable diversity within the bacterial community of the Antarctic sandy intertidal sediments.     

Dynamics of benthic communities in the Central Depression of the Barents Sea

Based on the data collected in five marine expeditions of the Murmansk Marine Biological Institute from 2002 through 2007, the spatial and temporary variability of benthic communities in the Central Depression of the Barents Sea (licensed plot of the Shtokmanovskoe condensed gas deposit) has been analyzed. The range of quantitative characteristics and the variability of species composition of deep-water zoobenthos have been determined. The influence of an insignificant change in the collecting method on the obtained results has been examined.     

Aerobic methanotrophic communities in the bottom sediments of Lake Baikal

The results of the first methodical investigation into the aerobic methanotrophic communities inhabiting the bottom sediments of Lake Baikal are reported. Use of the radioisotopic method revealed methane consumption in 12 10- to 50-cm-long sediment cores. The maximum methane consumption rates (495-737 microl/(dm3 day) were recorded in sediments in the regions of hydrothermal vents and oil and gas occurrence. Methane consumption was most active in the surface layers of the sediments (0-4 cm); it decreased with the sediment depth and became negligible or absent at depths below 20 cm. The number of methanotrophic bacteria usually ranged from 100 to 1000 cells/cm3 of sediment and reached 1 million cells/cm3 in the regions of oil and gas occurrence. The 17 enrichment cultures obtained were represented mainly by morphotype II methanotrophs. Phylogenetic analysis of the enrichment cultures in terms of the amino acid sequence of the alpha subunit of the membrane-bound methane monooxygenase revealed the predominance of methanotrophs of the genus Methylocystis. The results obtained suggest the presence of an active aerobic methanotrophic community in Lake Baikal.     

Phylogenetic diversity of Synechococcus strains isolated from the East China Sea and the East Sea

Phylogenetic relationships among 33 Synechococcus strains isolated from the East China Sea (ECS) and the East Sea (ES) were studied based on 16S rRNA gene sequences and 16S–23S rRNA gene internal transcribed spacer (ITS) sequences. Pigment patterns of the culture strains were also examined. Based on 16S rRNA gene and ITS sequence phylogenies, the Synechococcus isolates were clustered into 10 clades, among which eight were previously identified and two were novel. Half of the culture strains belonged to clade V or VI. All strains that clustered into novel clades exhibited both phycoerythrobilin and phycourobilin. Interestingly, the pigment compositions of isolates belonging to clades V and VI differed from those reported for other oceanic regions. None of the isolates in clade V showed phycourobilin, whereas strains in clade VI exhibited both phycourobilin and phycoerythrobilin, which is in contrast to previous studies. The presence of novel lineages and the different pigment patterns in the ECS and the ES suggests the possibility that some Synechococcus lineages are distributed only in geographically restricted areas and have evolved in these regions. Therefore, further elucidation of the physiological, ecological, and genetic characteristics of the diverse Synechococcus strains is required to understand their spatial and geographical distribution.     

天山一号冰川底部沉积层产蛋白酶耐低温菌株的筛选及其系统发育

[目的]通过对天山一号冰川底部沉积层耐低温菌的分离和其中产蛋白酶菌株的筛选,了解冰川微生物生理多样性和系统发育多样性,为高效低温蛋白酶生物技术的研发奠定基础.[方法]采用稀浓度的R2A、TSB平板涂布分离可培养细菌,通过脱脂乳选择性培养基筛选产蛋白酶的耐低温菌株.对分离菌株表型特征、生理生化特性、最适生长温度、耐盐性、产酶性能进行了比较,结合16S rRNA基因序列同源性分析确定产蛋白酶菌株的多样性和系统进化地位,通过BOX-PCR指纹技术分析16S rRNA基因序列高相似度的近缘菌株的遗传差异.[结果]从125株分离物中筛选到27株产蛋白酶的耐低温菌株,其中21株为适冷菌,仅6株菌为专性嗜冷菌,革兰氏阴性菌居多,假单胞菌属(Pseudomonas)菌株占40.7％.产酶菌株隶属于5个系统发育类群、9个属,其中γ-Proteobacteria、Actinobacteria、CFB(Cytophaga-Flexibacter-Bacteroides)为优势类群.[结论]天山1号冰川底部沉积层冻土中产蛋白酶的耐低温细菌多样性较丰富,本研究筛选得到的同属近缘种群较多,其产酶性状存在差异,适合开展微生物种群的生物地理学研究.