Use of real-time PCR for group-specific quantification of aceticlastic methanogens in anaerobic processes: population dynamics and community structures

The TaqMan quantitative PCR (QPCR) method was used to detect and quantify the 16S rRNA genes of aceticlastic methanogens at different taxonomic levels. Three different sets of primers coupled with a TaqMan probe for QPCR assays to detect the 16S rRNA genes of the order Methanosarcinales, as well as the families Methanosarcinaceae and Methanosaetaceae, were separately used. Using these primer and probe sets, the 16S rRNA genes of aceticlastic methanogens in samples from various anaerobic processes (i.e., nine pure cultures, batch experiment, and three different continuous processes including a full-scale digester), were monitored and quantified by QPCR assays. A batch experiment cultivating a mixture of aceticlastic methanogens, was conducted to monitor their population dynamics. Using this group-specific quantification method, the dynamics of a competition between two aceticlastic populations, as modulated by the acetate concentration, could well be described. The target 16S rRNA genes in environmental samples, collected from three different anaerobic processes treating sludge, cheese whey, and synthetic wastewaters, were additionally quantified. The quantified 16S rRNA gene concentrations for all samples successfully represented the community structures of the target methanogens, which were correlated accurately with the operational parameters of the anaerobic processes. It was also successful to demonstrate probe nesting of aceticlastic methanogens at the levels of order and family.     

Sulfate-reducing Bacteria and Mercury Methylation in the Water Column of the Lake 658 of the Experimental Lake Area

Sulfate-reducing bacteria (SRB) appear to be the main mediators of mercury methylation in sediments, which are deemed to be major sites of methylmercury (MMHg) production. However, recent studies have also found significant MMHg formation in the water column of lakes across North America. To investigate the potential involvement of SRB in mercury methylation in the water column of a stratified oligotrophic lake, two of the main families of SRB (Desulfobacteraceae and Desulfovibrionaceae) were quantified by Real-Time Polymerase Chain Reaction of the 16S rRNA gene. MMHg production was measured applying a stable isotope technique using ¹⁹⁸HgCl. Methylation assays were conducted at different water depths and under stimulation with lactate, acetate or propionate and inhibition with molybdate. Desulfobacteraceae and Desulfovibrionaceae16S rRNA gene copies in control samples accounted for 0.05% to 33% and <0.01% to 1.12% of the total bacterial 16S rRNA, respectively. MMHg formation was as high as 0.3 ng L^?1 day^?1 and largest in lactate amended samples. Strain isolation was only achieved in lactate amended media with all isolated strains being SRB belonging to the Desulfovibrio genus according to their 16S rRNA gene sequence. Isolated strains methylated between 0.06 and 0.2% of ¹⁹⁸HgCl per day. Acetate and propionate did not stimulate mercury methylation as much as lactate. Two strains were identified as Desulfovibrio sp. 12ML1 (FJ865472) and Desulfovibrio sp. 12ML3 (FJ865473), based on partial sequences of their 16S rRNA and DSR gene. Methylation assays and bacteria characterization suggest that Desulfovibrionaceae is an important mercury methylators in Lake 658. Supplemental materials are available for this article. Go to the publisher's online edition of Geomicrobiology Journal to view the free supplemental file.     

Changes in microbial diversity in industrial wastewater evaporation ponds following artificial salination

The salinity of industrial wastewater evaporation ponds was artificially increased from 3-7% to 12-16% (w/v), in an attempt to reduce the activity of sulfate-reducing bacteria (SRB) and subsequent emission of H2S. To investigate the changes in bacterial diversity in general, and SRB in particular, following this salination, two sets of universal primers targeting the 16S rRNA gene and the functional apsA [adenosine-5'-phosphosulfate (APS) reductase alpha-subunit] gene of SRB were used. Phylogenetic analysis indicated that Proteobacteria was the most dominant phylum both before and after salination (with 52% and 68%, respectively), whereas Firmicutes was the second most dominant phylum before (39%) and after (19%) salination. Sequences belonging to Bacteroidetes, Spirochaetes and Actinobacteria were also found. Several groups of SRB from Proteobacteria and Firmicutes were also found to inhabit this saline environment. Comparison of bacterial diversity before and after salination of the ponds revealed both a shift in community composition and an increase in microbial diversity following salination. The share of SRB in the 16S rRNA gene was reduced following salination, consistent with the reduction of H2S emissions. However, the community composition, as shown by apsA gene analysis, was not markedly affected.     

Comparison of five PCR assays for detecting Chlamydophila pneumoniae DNA

We compared five different polymerase chain reaction (PCR) assays for the detection of Chlamydophila pneumoniae DNA using highly purified elementary bodies (EBs) and peripheral blood mononuclear cells (PBMCs) from healthy blood donors. The primers were as follows; two targeting the 16S rRNA gene, one targeting the ompA gene, one targeting the Pst-I gene, and one targeting the 53 kDa outer membrane protein gene. The 16S rRNA touchdown enzyme time release (TETR) PCR, the ompA nested PCR and the 53 kDa nested PCR were the most sensitive assays and could detect one or more EB per assay. These three PCRs also had the same reproducibility, but the minimal amount of C. pneumoniae that could be reproducibly detected (10 of 10 testing positive) was 20 EBs. In a sample of specimens from healthy blood donors, we found 5 of 77 (6.5%) PBMCs specimens to have C. pneumoniae DNA according to the nested ompA PCR. Specimens with the 16S rRNA TETR and 53 kDa nested assays were found to have C. pneumoniae DNA 7 of 77 (9.1%) and 18 of 77 (23.4%) specimens, respectively. The other two assays failed to detect even a single positive. However, the detection rate decreased with repeated testing of the same samples. Our newly designed 53 kDa nested PCR may be as useful as the other four recommended PCR assays and may be a more useful assay for the detection of C. pneumoniae DNA from PBMCs.     

Speciating Campylobacter jejuni and Campylobacter coli isolates from poultry and humans using six PCR-based assays

Six previously published polymerase chain reaction (PCR) assays each targeting different genes were used to speciate 116 isolates previously identified as Campylobacter jejuni using routine microbiological techniques. Of the 116 isolates, 84 were of poultry origin and 32 of human origin. The six PCR assays confirmed the species identities of 31 of 32 (97%) human isolates and 56 of 84 (67%) poultry isolates as C. jejuni. Twenty eight of 84 (33%) poultry isolates were identified as Campylobacter coli and the remaining human isolate was tentatively identified as Campylobacter upsaliensis based on the degree of similarity of 16S rRNA gene sequences. Four of six published PCR assays showed 100% concordance in their ability to speciate 113 of the 116 (97.4%) isolates; two assays failed to generate a PCR product with four to 10 isolates. A C. coli-specific PCR identified all 28 hippuricase gene (hipO)-negative poultry isolates as C. coli although three isolates confirmed to be C. jejuni by the remaining five assays were also positive in this assay. A PCR-restriction fragment length polymorphism assay based on the 16S rRNA gene was developed, which contrary to the results of the six PCR-based assays, identified 28 of 29 hipO-negative isolates as C. jejuni. DNA sequence analysis of 16S rRNA genes from four hipO-negative poultry isolates showed they were almost identical to the C. jejuni type strain 16S rRNA sequences ATCC43431 and ATCC33560 indicating that assays reliant on 16S rRNA sequence may not be suitable for the differentiation of these two species.     

Primer and probe sets for group-specific quantification of the genera Nitrosomonas and Nitrosospira using real-time PCR

Use of quantitative real-time PCR (QPCR) with TaqMan probes is increasingly popular in various environmental works to detect and quantify a specific microorganism or a group of target microorganism. Although many aspects of conducting a QPCR assay have become very easy to perform, a proper design of oligonucleotide sequences comprising primers and a probe is still considered as one of the most important aspects of a QPCR application. This work was conducted to design group specific primer and probe sets for the detection of ammonia oxidizing bacteria (AOB) using a real-time PCR with a TaqMan system. The genera Nitrosomonas and Nitrosospira were grouped into five clusters based on similarity of their 16S rRNA gene sequences. Five group-specific AOB primer and probe sets were designed. These sets separately detect four subgroups of Nitrosomonas (Nitrosomonas europaea-, Nitrosococcus mobilis-, Nitrosomonas nitrosa-, and Nitrosomonas cryotolerans-clusters) along with the genus Nitrosospira. Target-group specificity of each primer and probe set was initially investigated by analyzing potential false results in silico, followed by a series of experimental tests for QPCR efficiency and detection limit. In general, each primer and probe set was very specific to the target group and sensitive to detect target DNA as low as two 16S rRNA gene copies per reaction mixture. QPCR efficiency, higher than 93.5%, could be achieved for all primer and probe sets. The primer and probe sets designed in this study can be used to detect and quantify the beta-proteobacterial AOB in biological nitrification processes and various environments.     

Identification and differentiation of Legionella pneumophila and Legionella spp. with real-time PCR targeting the 16S rRNA gene and species identification by mip sequencing

Fluorescent resonance energy transfer probes targeting the 16S rRNA gene were constructed for a sensitive and specific real-time PCR for identification and differentiation of Legionella pneumophila from other Legionella spp. For identification of non-L. pneumophila spp. by direct amplicon sequencing, two conventional PCR assays targeting the mip gene were established.     

Sulfate-reducing bacteria in tubes constructed by the marine infaunal polychaete Diopatra cuprea

Marine infaunal burrows and tubes greatly enhance solute transport between sediments and the overlying water column and are sites of elevated microbial activity. Biotic and abiotic controls of the compositions and activities of burrow and tube microbial communities are poorly understood. The microbial communities in tubes of the marine infaunal polychaete Diopatria cuprea collected from two different sediment habitats were examined. The bacterial communities in the tubes from a sandy sediment differed from those in the tubes from a muddy sediment. The difference in community structure also extended to the sulfate-reducing bacterial (SRB) assemblage, although it was not as pronounced for this functional group of species. PCR-amplified 16S rRNA gene sequences recovered from Diopatra tube SRB by clonal library construction and screening were all related to the family Desulfobacteriaceae. This finding was supported by phospholipid fatty acid analysis and by hybridization of 16S rRNA probes specific for members of the genera Desulfosarcina, Desulfobacter, Desulfobacterium, Desulfobotulus, Desulfococcus, and Desulfovibrio and some members of the genera Desulfomonas, Desulfuromonas, and Desulfomicrobium with 16S rRNA gene sequences resolved by denaturing gradient gel electrophoresis. Two of six SRB clones from the clone library were not detected in tubes from the sandy sediment. The habitat in which the D. cuprea tubes were constructed had a strong influence on the tube bacterial community as a whole, as well as on the SRB assemblage.     

Changes in sulfate-reducing bacterial populations during the onset of black band disease

Factors that facilitate the onset of black band disease (BBD) of corals remain elusive, though anoxic conditions under the complex microbial mat and production of sulfide are implicated in necrosis of underlying coral tissues. This study investigated the diversity and quantitative shifts of sulfate-reducing bacterial (SRB) populations during the onset of BBD using real-time PCR (RT-PCR) and cloning approaches targeting the dissimilatory (bi)sulfite reductase (dsrA) gene. A quantitativePCR (qPCR) assay targeting the 16S rRNA gene also provided an estimate of total bacteria, and allowed the relative percentage of SRB within the lesions to be determined. Three Montipora sp. coral colonies identified with lesions previously termed cyanobacterial patches (CPs) (comprising microbial communities unlike those of BBD lesions), were tagged and followed through time as CP developed into BBD. The dsrA-targeted qPCR detected few copies of the gene in the CP samples (<65 per ng DNA), though copy numbers increased in BBD lesions (>2500 per ng DNA). SRB in CP samples were less than 1% of the bacterial population, though represented up to 7.5% of the BBD population. Clone libraries also demonstrated a shift in the dominant dsrA sequences as lesions shifted from CP into BBD. Results from this study confirm that SRB increase during the onset of BBD, likely increasing sulfide concentrations at the base of the microbial mat and facilitating the pathogenesis of BBD.     

重度肥胖症患者膳食干预前后肠道内硫酸盐还原菌的定量检测

目的检测重度肥胖症患者膳食干预过程中,肠道内硫酸盐还原菌（SRB）的数量变化,为进一步研究SRB与肥胖的关系提供参考。方法以SRB基因组中的重要功能基因一腺苷酰硫酸（APS）还原酶基因作为指示基因,通过实时定量PCR的方法检测了12名重度肥胖症患者在进行膳食干预过程中随着体重的降低,肠道内SRB的数量变化,同时以16SrRNA基因对肠道内总菌进行定量来计算SRB占总菌的相对比例。结果膳食干预过程中,随着患者体重的显著降低,其肠道内SRB占总菌的比例也显著下降。在维持期,患者体重仍和干预前有显著差异,但较干预期有所回升;其体内SRB含量也显著低于干预前,但相比干预期,有所上升。结论研究结果提示肠道菌群中SRB细菌与肥胖的发展有密切关系,为后续研究SRB细菌的功能和作用机理奠定了基础。     

Successional development of sulfate-reducing bacterial populations and their activities in a wastewater biofilm growing under microaerophilic conditions

A combination of fluorescence in situ hybridization, microprofiles, denaturing gradient gel electrophoresis of PCR-amplified 16S ribosomal DNA fragments, and 16S rRNA gene cloning analysis was applied to investigate successional development of sulfate-reducing bacteria (SRB) community structure and in situ sulfide production activity within a biofilm growing under microaerophilic conditions (dissolved oxygen concentration in the bulk liquid was in the range of 0 to 100 microM) and in the presence of nitrate. Microelectrode measurements showed that oxygen penetrated 200 microm from the surface during all stages of biofilm development. The first sulfide production of 0.32 micromol of H(2)S m(-2) s(-1) was detected below ca. 500 microm in the 3rd week and then gradually increased to 0.70 micromol H(2)S m(-2) s(-1) in the 8th week. The most active sulfide production zone moved upward to the oxic-anoxic interface and intensified with time. This result coincided with an increase in SRB populations in the surface layer of the biofilm. The numbers of the probe SRB385- and 660-hybridized SRB populations significantly increased to 7.9 x 10(9) cells cm(-3) and 3.6 x 10(9) cells cm(-3), respectively, in the surface 400 microm during an 8-week cultivation, while those populations were relatively unchanged in the deeper part of the biofilm, probably due to substrate transport limitation. Based on 16S rRNA gene cloning analysis data, clone sequences that related to Desulfomicrobium hypogeium (99% sequence similarity) and Desulfobulbus elongatus (95% sequence similarity) were most frequently found. Different molecular analyses confirmed that Desulfobulbus, Desulfovibrio, and Desulfomicrobium were found to be the numerically important members of SRB in this wastewater biofilm.     

Mapping of picoeucaryotes in marine ecosystems with quantitative PCR of the 18S rRNA gene

A quantitative PCR (QPCR) assay based on the use of SYBR Green I was developed to assess the abundance of specific groups of picoeukaryotes in marine waters. Six primer sets were designed targeting four different taxonomic levels: domain (Eukaryota), division (Chlorophyta), order (Mamiellales) and genus (Bathycoccus, Micromonas, and Ostreococcus). Reaction conditions were optimized for each primer set which was validated in silico, on agarose gels, and by QPCR against a variety of target and non-target cultures. The approach was tested by estimating gene copy numbers for Micromonas, Bathycoccus, and Ostreococcus in seawater samples to which cultured cells were added in various concentrations. QPCR was then used to determine that rRNA gene (rDNA) copy number varied from one to more than 12,000 in 18 strains of phytoplankton. Finally, QPCR was applied to environmental samples from a Mediterranean Sea coastal site and the results were compared to those obtained by Fluorescent in situ hybridization (FISH). The data obtained demonstrate that Chlorophyta and more specifically Mamiellales were important in these waters, especially during the winter picoplankton bloom. The timing of major abundance peaks of the targeted species was similar by QPCR and FISH. When used in conjunction with other techniques such as FISH or gene clone libraries, QPCR appears as very promising to quickly obtain data on the ecological distribution of important phytoplankton groups. Data interpretation must take into account primer specificity and the varying rRNA gene copy number among eukaryotes.     

Characterization of bacterial community associated to biofilms of corroded oil pipelines from the southeast of Mexico

Microbial communities associated to biofilms promote corrosion of oil pipelines. The community structure of bacteria in the biofilm formed in oil pipelines is the basic knowledge to understand the complexity and mechanisms of metal corrosion. To assess bacterial diversity, biofilm samples were obtained from X52 steel coupons corroded after 40 days of exposure to normal operation and flow conditions. The biofilm samples were directly used to extract metagenomic DNA, which was used as template to amplify 16S ribosomal gene by PCR. The PCR products of 16S ribosomal gene were also employed as template for sulfate-reducing bacteria (SRB) specific nested-PCR and both PCR products were utilized for the construction of gene libraries. The V3 region of the 16S rRNA gene was also amplified to analyse the bacterial diversity by analysis of denaturing gradient gel electrophoresis (DGGE). Ribosomal library and DGGE profiles exhibited limited bacterial diversity, basically including Citrobacter spp., Enterobacter spp. and Halanaerobium spp. while Desulfovibrio alaskensis and a novel clade within the genus Desulfonatronovibrio were detected from the nested PCR library. The biofilm samples were also taken for the isolation of SRB. Desulfovibrio alaskensis and Desulfovibrio capillatus, as well as some strains related to Citrobacter were isolated. SRB consists in a very small proportion of the community and Desulfovibrio spp. were the relatively abundant groups among the SRB. This is the first study directly exploring bacterial diversity in corrosive biofilms associated to steel pipelines subjected to normal operation conditions.     

Sulfate-Reducing Bacteria in Tubes Constructed by the Marine Infaunal Polychaete Diopatra cuprea

Marine infaunal burrows and tubes greatly enhance solute transport between sediments and the overlying water column and are sites of elevated microbial activity. Biotic and abiotic controls of the compositions and activities of burrow and tube microbial communities are poorly understood. The microbial communities in tubes of the marine infaunal polychaete Diopatria cuprea collected from two different sediment habitats were examined. The bacterial communities in the tubes from a sandy sediment differed from those in the tubes from a muddy sediment. The difference in community structure also extended to the sulfate-reducing bacterial (SRB) assemblage, although it was not as pronounced for this functional group of species. PCR-amplified 16S rRNA gene sequences recovered from Diopatra tube SRB by clonal library construction and screening were all related to the family Desulfobacteriaceae. This finding was supported by phospholipid fatty acid analysis and by hybridization of 16S rRNA probes specific for members of the genera Desulfosarcina, Desulfobacter, Desulfobacterium, Desulfobotulus, Desulfococcus, and Desulfovibrio and some members of the genera Desulfomonas, Desulfuromonas, and Desulfomicrobium with 16S rRNA gene sequences resolved by denaturing gradient gel electrophoresis. Two of six SRB clones from the clone library were not detected in tubes from the sandy sediment. The habitat in which the D. cuprea tubes were constructed had a strong influence on the tube bacterial community as a whole, as well as on the SRB assemblage.     

Identification and Differentiation of Legionella pneumophila and Legionella spp. with Real-Time PCR Targeting the 16S rRNA Gene and Species Identification by mip Sequencing

Fluorescent resonance energy transfer probes targeting the 16S rRNA gene were constructed for a sensitive and specific real-time PCR for identification and differentiation of Legionella pneumophila from other Legionella spp. For identification of non-L. pneumophila spp. by direct amplicon sequencing, two conventional PCR assays targeting the mip gene were established.     

Culture-based and non-growth-dependent detection of the Burkholderia cepacia complex in soil environments

Burkholderia cepacia complex (Bcc) bacteria reside in soil, plant rhizospheres, and water, but their prevalence and distribution in outdoor environments is not clear. We sampled a variety of soil and rhizosphere environments with which people may have contact: playgrounds, athletic fields, parks, hiking trails, residential yards, and gardens. A total of 91 sites was sampled in three large U.S. cities. In the first phase of the study, putative Bcc isolates were recovered on Burkholderia cepacia selective agar and trypan blue tetracycline medium and subsequently examined for biochemical reactivity and growth at 32 and 22 degrees C. Isolates were further examined by PCR assays targeting Bcc-specific ribosomal DNA and recA gene sequences. Among the 1,013 bacterial isolates examined, 68 were identified as Bcc; 14 (15%) of 91 sampled sites yielded Bcc isolates. In the second phase, DNA was extracted directly from soil samples and examined with PCR assays targeting Bcc 16S rRNA gene sequences. Either 82 or 93% of the soil samples were positive for at least one Bcc genomovar, depending on the PCR assay system used. Cloning and sequencing were performed to check the specificity of the PCR assays. Sequence analysis of the 463-bp 16S rRNA inserts from eight clones indicated that all were from members of the Bcc. The four soil samples from which these clones were generated did not yield isolates identified as Bcc. Based on PCR detection, Bcc appears to be prevalent in soil from urban and suburban environments. Culture-based recovery of Bcc may underestimate environmental populations.     

高脂饲料诱导的肥胖大鼠肠道内硫酸盐还原菌的定量检测

目的 检测高脂饲料诱导大鼠肥胖过程中,大鼠肠道内硫酸盐还原菌( sulfate-reducing bacteria,SRB)的数量变化,为研究SRB与肥胖的关系提供参考.方法 20只Wistar大鼠随机分为2组(每组10只),一组饲喂高脂饲料(HFD组)18周,另一组饲喂正常饲料(NCD组,即对照组)18周.以编码腺苷酰硫酸还原酶α亚基的基因(aprA)作为分子标记,通过荧光定量PCR的方法检测两组大鼠在0、8和18周,肠道内SRB的数量变化;同时,以16S rRNA基因作为标记基因定量大鼠肠道内总菌的数量,以计算肠道内SRB在总菌中的比例变化.结果 分组饲喂8周后,高脂饲料饲喂组大鼠的体重与正常饲料组相比显著升高.对SRB的定量结果显示,饲喂8周和18周,高脂饲料组大鼠肠道内SRB的数量和含量与正常饲料组相比显著升高.结论 大鼠肠道中的硫酸盐还原菌与饮食诱导的肥胖密切相关,为进一步研究SRB在肥胖及其相关代谢疾病的发生发展中的作用提供了依据.     

Phylogenetic diversity and molecular detection of bacteria in gull feces

In spite of increasing public health concerns about the potential risks associated with swimming in waters contaminated with waterfowl feces, little is known about the composition of the gut microbial community of aquatic birds. To address this, a gull 16S rRNA gene clone library was developed and analyzed to determine the identities of fecal bacteria. Analysis of 282 16S rRNA gene clones demonstrated that the gull gut bacterial community is mostly composed of populations closely related to Bacilli (37%), Clostridia (17%), Gammaproteobacteria (11%), and Bacteriodetes (1%). Interestingly, a considerable number of sequences (i.e., 26%) were closely related to Catellicoccus marimammalium, a gram-positive, catalase-negative bacterium. To determine the occurrence of C. marimammalium in waterfowl, species-specific 16S rRNA gene PCR and real-time assays were developed and used to test fecal DNA extracts from different bird (n = 13) and mammal (n = 26) species. The results showed that both assays were specific to gull fecal DNA and that C. marimammalium was present in gull fecal samples collected from the five locations in North America (California, Georgia, Ohio, Wisconsin, and Toronto, Canada) tested. Additionally, 48 DNA extracts from waters collected from six sites in southern California, Great Lakes in Michigan, Lake Erie in Ohio, and Lake Ontario in Canada presumed to be impacted with gull feces were positive by the C. marimammalium assay. Due to the widespread presence of this species in gulls and environmental waters contaminated with gull feces, targeting this bacterial species might be useful for detecting gull fecal contamination in waterfowl-impacted waters.     

Analysis of diversity and activity of sulfate-reducing bacterial communities in sulfidogenic bioreactors using 16S rRNA and dsrB genes as molecular markers

Here we describe the diversity and activity of sulfate-reducing bacteria (SRB) in sulfidogenic bioreactors by using the simultaneous analysis of PCR products obtained from DNA and RNA of the 16S rRNA and dissimilatory sulfite reductase (dsrAB) genes. We subsequently analyzed the amplified gene fragments by using denaturing gradient gel electrophoresis (DGGE). We observed fewer bands in the RNA-based DGGE profiles than in the DNA-based profiles, indicating marked differences in the populations present and in those that were metabolically active at the time of sampling. Comparative sequence analyses of the bands obtained from rRNA and dsrB DGGE profiles were congruent, revealing the same SRB populations. Bioreactors that received either ethanol or isopropanol as an energy source showed the presence of SRB affiliated with Desulfobulbus rhabdoformis and/or Desulfovibrio sulfodismutans, as well as SRB related to the acetate-oxidizing Desulfobacca acetoxidans. The reactor that received wastewater containing a diverse mixture of organic compounds showed the presence of nutritionally versatile SRB affiliated with Desulfosarcina variabilis and another acetate-oxidizing SRB, affiliated with Desulfoarculus baarsii. In addition to DGGE analysis, we performed whole-cell hybridization with fluorescently labeled oligonucleotide probes to estimate the relative abundances of the dominant sulfate-reducing bacterial populations. Desulfobacca acetoxidans-like populations were most dominant (50 to 60%) relative to the total SRB communities, followed by Desulfovibrio-like populations (30 to 40%), and Desulfobulbus-like populations (15 to 20%). This study is the first to identify metabolically active SRB in sulfidogenic bioreactors by using the functional gene dsrAB as a molecular marker. The same approach can also be used to infer the ecological role of coexisting SRB in other habitats.