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PubMed2007

Microbial diversity and dynamics in multi- and single-compartment anaerobic bioreactors processing sulfate-rich waste streams.

We investigated bacterial and archaeal community structures and population dynamics in two anaerobic bioreactors processing a carbohydrate- and sulfate-rich synthetic wastewater. A five-compartment anaerobic migrating blanket reactor (AMBR) was designed to promote biomass and substrate staging, which partially separates the processes of methanogenesis and sulfidogenesis in the middle and outer compartment(s) respectively. The second reactor was a conventional, single-compartment upflow anaerobic sludge blanket (UASB) reactor. Both reactors, which were seeded with the same inoculum, performed well when the influent chemical oxygen demand (COD)/SO(4) (2-) mass ratio was 24.4. The AMBR performed worse than the UASB reactor when the influent COD/SO(4) (2-) mass ratio was decreased to 5.0 by raising the sulfate load. Terminal restriction fragment length polymorphism analyses of bacterial 16S rRNA genes showed that the increase in sulfate load had a greater impact on bacterial diversity and community structure for the five AMBR compartments than for the UASB reactor. Moreover, bacterial community profiles across AMBR compartments became more similar through time, indicating a converging, rather than a staged community. While similar populations were abundant in both reactors at the beginning of the experiment, fermenting bacteria (clostridia, streptococci), and sulfate-reducing bacteria became more abundant in the AMBR, after shifting to a higher sulfate load, while a novel Thermotogales-like population eventually became predominant in the UASB reactor. A similar shift in the community structure of the hydrogenotrophic methanogens in the AMBR occurred: representatives of the Methanobacteriaceae out-competed the Methanospirillaceae after increasing the sulfate load in the AMBR, while the archaeal community structure was maintained in the UASB.

Archaea↗
PubMed2001

The microbial diversity in picoplankton enrichment cultures: a molecular screening of marine isolates.

Picoplankton bacteria from a North Sea water sample were cultured under a variety of different conditions (nutrients, temperature, light, agitation, adhesion). Fluorescent in situ hybridization (FISH) analysis of the enrichments showed complex communities which were dominated by gamma-Proteobacteria or beta-Proteobacteria, followed by alpha-Proteobacteria and bacteria from the Cytophaga/Flavobacterium/Bacteroides (CFB) cluster. Among 410 isolates, a high degree of diversity was found, both with respect to colony color and morphology and with respect to genetic diversity. Isolated bacteria were classified into the main taxa by a special PCR approach, termed signature PCR (SIG-PCR). It was based on an oligo primer mixture targeting 16S rDNA which yielded PCR products of taxon-specific lengths. Again, gamma-Proteobacteria dominated (48%), followed by alpha-Proteobacteria (20%). beta-Proteobacteria were rarely isolated (eight strains of 410). The CFB cluster comprised the second largest phylum (14%), and 7.5% of all isolates belonged to the high-GC Gram-positives. Thus, isolated bacteria were representative of enrichment communities with the exception of the beta-Proteobacteria, which were detected in high abundance in certain enrichments by FISH but not isolated, and the high-GC Gram-positives, which were cultivated but not detected by FISH. A genomic fingerprinting technique, randomly amplified polymorphic DNA, showed that among 58 CFB isolates only 18 identical genotypes were found, and among the 84 alpha-Proteobacteria only eight identical genotypes were present. The data show the enormous diversity of cultivated bacteria from picoplankton enrichment cultures of one North Sea water sample, which is only a small fraction of the total picoplankton community.

Journal Article↗
PubMed2006

Impact of microbial diversity on rapid detection of enterohemorrhagic Escherichia coli in surface waters.

Enterohemorrhagic Escherichia coli (EHEC) are a physiologically, immunologically and genetically diverse collection of strains that pose a serious water-borne threat to human health. Consequently, immunological and PCR assays have been developed for the rapid, sensitive detection of presumptive EHEC. However, the ability of these assays to consistently detect presumptive EHEC while excluding closely related non-EHEC strains has not been documented. We conducted a 30-month monitoring study of a major metropolitan watershed. Surface water samples were analyzed using an immunological assay for E. coli O157 (the predominant strain worldwide) and a multiplex PCR assay for the virulence genes stx(1), stx(2) and eae. The mean frequency of water samples positive for the presence of E. coli O157, stx(1) or stx(2) genes, or the eae gene was 50%, 26% and 96%, respectively. Quantitative analysis of selected enriched water samples indicated that even in samples positive for E. coli O157 cells, stx(1)/stx(2) genes, and the eae gene, the concentrations were rarely comparable. Seventeen E. coli O157 strains were isolated, however, none were EHEC. These data indicate the presence of multiple strains similar to EHEC but less pathogenic. These findings have important ramifications for the rapid detection of presumptive EHEC; namely, that current immunological or PCR assays cannot reliably identify water-borne EHEC strains.

Adhesins, Bacterial↗
PubMed2006

Microbial diversity in Maras salterns, a hypersaline environment in the Peruvian Andes.

Maras salterns are located 3,380 m above sea level in the Peruvian Andes. These salterns consist of more than 3,000 little ponds which are not interconnected and act as crystallizers where salt precipitates. These ponds are fed by hypersaline spring water rich in sodium and chloride. The microbiota inhabiting these salterns was examined by fluorescence in situ hybridization (FISH), 16S rRNA gene clone library analysis, and cultivation techniques. The total counts per milliliter in the ponds were around 2 x 10(6) to 3 x 10(6) cells/ml, while the spring water contained less than 100 cells/ml and did not yield any detectable FISH signal. The microbiota inhabiting the ponds was dominated (80 to 86% of the total counts) by Archaea, while Bacteria accounted for 10 to 13% of the 4',6'-diamidino-2-phenylindole (DAPI) counts. A total of 239 16S rRNA gene clones were analyzed (132 Archaea clones and 107 Bacteria clones). According to the clone libraries, the archaeal assemblage was dominated by microorganisms related to the cosmopolitan square archaeon "Haloquadra walsbyi," although a substantial number of the sequences in the libraries (31% of the 16S rRNA gene archaeal clones) were related to Halobacterium sp., which is not normally found in clone libraries from solar salterns. All the bacterial clones were closely related to each other and to the gamma-proteobacterium "Pseudomonas halophila" DSM 3050. FISH analysis with a probe specific for this bacterial assemblage revealed that it accounted for 69 to 76% of the total bacterial counts detected with a Bacteria-specific probe. When pond water was used to inoculate solid media containing 25% total salts, both extremely halophilic Archaea and Bacteria were isolated. Archaeal isolates were not related to the isolates in clone libraries, although several bacterial isolates were very closely related to the "P. halophila" cluster found in the libraries. As observed for other hypersaline environments, extremely halophilic bacteria that had ecological relevance seemed to be easier to culture than their archaeal counterparts.

Altitude↗
PubMed1997

Molecular microbial diversity of an anaerobic digestor as determined by small-subunit rDNA sequence analysis.

The bacterial community structure of a fluidized-bed reactor fed by vinasses (wine distillation waste) was analyzed. After PCR amplification, four small-subunit (SSU) rDNA clone libraries of Bacteria, Archaea, Procarya, and Eucarya populations were established. The community structure was determined by operational taxonomic unit (OTU) phylogenetic analyses of 579 partial rDNA sequences (about 500 bp long). A total of 146 OTUs were found, comprising 133, 6, and 7 from the Bacteria, Archaea, and Eucarya domains, respectively. A total of 117 bacterial OTU were affiliated with major phyla: low-G+C gram-positive bacteria, Cytophaga-Flexibacter-Bacteroides, Proteobacteria, high-G+C gram-positive bacteria, and Spirochaetes, where the clone distribution was 34, 26, 17, 6, and 4%, respectively. The other 16 bacterial OTUs represent 13% of the clones. They were either affiliated with narrow phyla such as Planctomyces-Chlamydia, green nonsulfur bacteria, or Synergistes, or deeply branched on the phylogenetic tree. A large number of bacterial OTUs are not closely related to any other hitherto determined sequences. The most frequent bacterial OTUs represents less than 5% of the total bacterial SSU rDNA sequences. However, the 20 more frequent bacterial OTUs describe at least 50% of these sequences. Three of the six Archaea OTUs correspond to 95% of the Archaea population and are very similar to already known methanogenic species: Methanosarcina barkeri, Methanosarcina frisius, and Methanobacterium formicicum. In contrast, the three other Archaea OTUs are unusual and are related to thermophilic microorganisms such as Crenarchaea or Thermoplasma spp. Five percent of the sequences analyzed were chimeras and were removed from the analysis.

Bacteria, Anaerobic↗
PubMed1999

Molecular analysis of the microbial diversity present in the colonic wall, colonic lumen, and cecal lumen of a pig.

Random clones of 16S ribosomal DNA gene sequences were isolated after PCR amplification with eubacterial primers from total genomic DNA recovered from samples of the colonic lumen, colonic wall, and cecal lumen from a pig. Sequences were also obtained for cultures isolated anaerobically from the same colonic-wall sample. Phylogenetic analysis showed that many sequences were related to those of Lactobacillus or Streptococcus spp. or fell into clusters IX, XIVa, and XI of gram-positive bacteria. In addition, 59% of randomly cloned sequences showed less than 95% similarity to database entries or sequences from cultivated organisms. Cultivation bias is also suggested by the fact that the majority of isolates (54%) recovered from the colon wall by culturing were related to Lactobacillus and Streptococcus, whereas this group accounted for only one-third of the sequence variation for the same sample from random cloning. The remaining cultured isolates were mainly Selenomonas related. A higher proportion of Lactobacillus reuteri-related sequences than of Lactobacillus acidophilus- and Lactobacillus amylovorus-related sequences were present in the colonic-wall sample. Since the majority of bacterial ribosomal sequences recovered from the colon wall are less than 95% related to known organisms, the roles of many of the predominant wall-associated bacteria remain to be defined.

Anaerobiosis↗
PubMed2001

DNA extraction from soils: old bias for new microbial diversity analysis methods.

The impact of three different soil DNA extraction methods on bacterial diversity was evaluated using PCR-based 16S ribosomal DNA analysis. DNA extracted directly from three soils showing contrasting physicochemical properties was subjected to amplified ribosomal DNA restriction analysis and ribosomal intergenic spacer analysis (RISA). The obtained RISA patterns revealed clearly that both the phylotype abundance and the composition of the indigenous bacterial community are dependent on the DNA recovery method used. In addition, this effect was also shown in the context of an experimental study aiming to estimate the impact on soil biodiversity of the application of farmyard manure or sewage sludge onto a monoculture of maize for 15 years.

Bacteria↗
PubMed2001

Microbial diversity of the brine-seawater interface of the Kebrit Deep, Red Sea, studied via 16S rRNA gene sequences and cultivation methods.

The brine-seawater interface of the Kebrit Deep, northern Red Sea, was investigated for the presence of microorganisms using phylogenetic analysis combined with cultivation methods. Under strictly anaerobic culture conditions, novel halophiles were isolated. The new rod-shaped isolates belong to the halophilic genus Halanaerobium and are the first representatives of the genus obtained from deep-sea, anaerobic brine pools. Within the genus Halanaerobium, they represent new species which grow chemoorganotrophically at NaCl concentrations ranging from 5 to 34%. The cellular fatty acid compositions are consistent with those of other Halanaerobium representatives, showing unusually large amounts of Delta7 and Delta11 16:1 fatty acids. Phylogenetic analysis of the brine-seawater interface sample revealed the presence of various bacterial 16S rRNA gene sequences dominated by cultivated members of the bacterial domain, with the majority affiliated with the genus Halanaerobium. The new Halanaerobium 16S rRNA clone sequences showed the highest similarity (99.9%) to the sequence of isolate KT-8-13 from the Kebrit Deep brine. In this initial survey, our polyphasic approach demonstrates that novel halophiles thrive in the anaerobic, deep-sea brine pool of the Kebrit Deep, Red Sea. They may contribute significantly to the anaerobic degradation of organic matter enriched at the brine-seawater interface.

Bacteria, Anaerobic↗
PubMed2005

Longitudinal study of microbial diversity and seasonality in the Mexico City metropolitan area water supply system.

In the Mexico City metropolitan area (MCMA), 70% of the water for 18 million inhabitants is derived from the Basin of Mexico regional aquifer. To provide an overview of the quality of the groundwater, a longitudinal study was conducted, in which 30 sites were randomly selected from 1,575 registered extraction wells. Samples were taken before and after chlorine disinfection during both the rainy and dry seasons (2000-2001). Microbiological parameters (total coliforms, fecal coliforms, streptococci, and Vibrio spp.), the presence of Helicobacter pylori, and physicochemical parameters, including the amount of trihalomethanes (THMs), were determined. Although microorganisms and inorganic and organic compounds were evident, they did not exceed current permissible limits. Chlorine levels were low, and the bacterial counts were not affected by chlorine disinfection. Eighty-four bacterial species from nine genera normally associated with fecal contamination were identified in water samples. H. pylori was detected in at least 10% of the studied samples. About 40% of the samples surpassed the THM concentration allowed by Mexican and U.S. regulations, with levels of chloroform being high. The quality of the water distributed to the MCMA varied between the rainy and dry seasons, with higher levels of pH, nitrates, chloroform, bromodichloromethane, total organic carbon, and fecal streptococci during the dry season. This study showed that the groundwater distribution system is susceptible to contamination and that there is a need for a strict, year-round disinfection strategy to ensure adequate drinking-water quality. This situation in one of the world's megacities may reflect what is happening in large urban centers in developing countries which rely on a groundwater supply.

Bacteria↗
PubMed2000

[Microbial diversity of deep-sea extremophiles--Piezophiles, Hyperthermophiles, and subsurface microorganisms].

Knowledge of our Planet's biosphere has increased tremendously during the last 10 to 20 years. In the field of Microbiology in particular, scientists have discovered novel "extremophiles", microorganisms capable of living in extreme environments such as highly acidic or alkaline conditions, at high salt concentration, with no oxygen, extreme temperatures (as low as -20 degrees C and as high as 300 degrees C), at high concentrations of heavy metals and in high pressure environments such as the deep-sea. It is apparent that microorganisms can exist in any extreme environment of the Earth, yet already scientists have started to look for life on other planets; the so-called "Exobiology" project. But as yet we have little knowledge of the deep-sea and subsurface biosphere of our own planet. We believe that we should elucidate the Biodiversity of Earth more thoroughly before exploring life on other planets, and these attempts would provide deeper insight into clarifying the existence of extraterrestrial life. We focused on two deep-sea extremophiles in this article; one is "Piezophiles", and another is "Hyperthermophiles". Piezophiles are typical microorganisms adapted to high-pressure and cold temperature environments, and located in deep-sea bottom. Otherwise, hyperthermophiles are living in high temperature environment, and located at around the hydrothermal vent systems in deep-sea. They are not typical deep-sea microorganisms, but they can grow well at high-pressure condition, just like piezophiles. Deming and Baross mentioned that most of the hyperthermophilic archaea isolated from deep-sea hydrothermal vents are able to grow under conditions of high temperature and pressure, and in most cases their optimal pressure for growth was greater than the environmental pressure they were isolated from. It is possible that originally their native environment may have been deeper than the sea floor and that there had to be a deeper biosphere. This implication suggests that the deep-sea hydrothermal vents are the windows to a deep subsurface biosphere. A vast array of chemoautotrophic deep-sea animal communities have been found to exist in cold seep environments, and most of these animals are common with those found in hydrothermal vent environments. Thus, it is possible to consider that the cold seeps are also one of slit windows to a deep subsurface biosphere. We conclude that the deep-sea extremophiles are very closely related into the unseen majority in subsurface biosphere, and the subsurface biosphere probably concerns to consider the "exobiology".

Archaea↗
PubMedDate not supplied

[Microbial diversity studies at the Winogradsky Institute of Microbiology].

Due to a chain of circumstances, some of them caused by unfavorable events, the Institute of Microbiology (INMI), Russian Academy of Sciences, continued the tradition of the "golden age" of microbiology of Winogradsky's time. The interest was focused on functional diversity of microorganisms as catalysts of natural chemical processes. The concept of a morphophysiological genus continues to play the key role in general microbiology, serving as an operational entity of the science. Researchers affiliated with INMI pioneered in describing 60 genera, constituting more than 4.9% of the 1228 validated genus designations as of January 1, 2004, and more than 6% of the nonpathogenic cultivable genera deposited at DSMZ. In addition to the formal biodiversity, other aspects of physiological and ecological diversity described by different classification schemes are considered. More recent findings were mostly in the field of alkaliphiles, acidophiles, thermophiles, and halophilic anaerobes, proliferating in extreme environments. Attempts to work out classifications of microorganisms with prognostic potential are presented.

Academies and Institutes↗
PubMed2005

[Bacterial conversion of sulfur-and phosphorous-compounds and microbial diversity in sediments from a near-shore marine-cultural region].

The H2S-producing bacteria and the phosphorous-cycling bacteria in sediments from near-shore marine-cultural region were investigated. Results indicate that the bacterial H2S production in aerobic condition is the dominating process to produce H2S in the sediment of cultural pond. The total counts of H2S-producing bacteria utilizing cysteine and Na2S2O3 were 1.6 x 10(6) and 4.35 x 10(3) cells g(-1) respectively. The counts of sulfate-reducing bacteria in the sediments were very little, only 2.5 x 10(1) g(-1). Further results show that the bacterial counts of decomposing lecithin and secreting phosphatase were 2.17 x 10(5) and 1.21 x 10(6) g(-1) respectively, bacterial counts of dissolving Ca3PO4 were 6.96 x 10(3) g(-1). Traditional taxonomy and partial 16S rRNA gene sequencing on the H2S-producing and phosphate-cycling bacteria indicate that most isolates could be classified as members of the following Genera: Bacillus, Halobacillus, Microbacterium, etc.

Animals↗
PubMed2003

A(r)Ray of hope in analysis of the function and diversity of microbial communities.

The vast majority of microorganisms in the environment remain uncultured, and their existence is known only from sequences retrieved by PCR. As a consequence, our understanding of the ecological function of dominant microbial populations in the environment is limited. We will review microbial diversity studies and show that these may have moved from an extreme underestimation to a potentially severe overestimation of diversity. The latter results from a simple PCR-generated artifact: the cloning of heteroduplex molecules followed by Escherichia coli mismatch repair, which may generate an exponential increase in observed sequence diversity. However, simple modifications to current PCR amplification protocols minimize such artifactual sequences and may bring within our reach estimation of bacterial diversity in environmental samples. Such estimates may spur new culture-independent approaches based on genomic and microarray technology, allowing correlation of phylogenetic identity with the ecological function of unculturable organisms. In particular, we are developing a DNA microarray that enables identification of individual populations active in utilization of specific organic substrates. The array consists of 16S and 23S rDNA-targeted oligonucleotides and is hybridized to RNA extracted from samples incubated with (14)C-labeled organic substrates. Populations that metabolize the substrate can be identified by the radiolabel incorporated in their rRNA after only one to two cell doublings, ensuring realistic preservation of community structure. Thus, the microarray approach may provide a powerful means to link microbial community structure with in situ function of individual populations.

Bacterial Physiological Phenomena↗