Ribosomal Tag Pyrosequencing of DNA and RNA Reveals “Rare” Taxa with High Protein Synthesis Potential in the Sediment of a Hypersaline Lake in Western Australia

Little is known about the potential activity of microbial communities in hypersaline sediment ecosystems. Ribosomal tag libraries of DNA and RNA extracted from the sediment of Lake Strawbridge (Western Australia) revealed bacterial and archaeal operational taxonomic units (OTUs) with high RNA/DNA ratios providing evidence for the presence of ‘rare’ but potentially “active” taxa. Among the ‘rare’ bacterial taxa Halomonas, Salinivibrio and Idiomarina showed the highest protein synthesis potential. Rare but ‘active’ archaeal OTUs were related to the KTK 4A cluster and the Marine-Benthic-Groups B and D. We present the first molecular analysis of the microbial diversity and protein synthesis potential of rare microbial taxa in a hypersaline sediment ecosystem.     

Amplicon-based profiling of bacteria in raw and secondary treated wastewater from treatment plants across Australia

In this study, we investigated the use of Illumina high-throughput sequencing of 16S ribosomal RNA (rRNA) amplicons to explore microbial diversity and community structure in raw and secondary treated wastewater (WW) samples from four municipal wastewater treatment plants (WWTPs A–D) across Australia. Sequence reads were analyzed to determine the abundance and diversity of bacterial communities in raw and secondary treated WW samples across the four WWTPs. In addition, sequence reads were also characterized to phenotypic features and to estimate the abundance of potential pathogenic bacterial genera and antibiotic-resistant genes in total bacterial communities. The mean coverage, Shannon diversity index, observed richness (S _obs), and abundance-based coverage estimate (ACE) of richness for raw and secondary treated WW samples did not differ significantly (P > 0.05) among the four WWTPs examined. Generally, raw and secondary treated WW samples were dominated by members of the genera Pseudomonas, Arcobacter, and Bacteroides. Evaluation of source contributions to secondary treated WW, done using SourceTracker, revealed that 8.80–61.4% of the bacterial communities in secondary treated WW samples were attributed to raw WW. Twenty-five bacterial genera were classified as containing potential bacterial pathogens. The abundance of potentially pathogenic genera in raw WW samples was higher than that found in secondary treated WW samples. Among the pathogenic genera identified, Pseudomonas and Arcobacter had the greatest percentage of the sequence reads. The abundances of antibiotic resistance genes were generally low (<0.5%), except for genes encoding ABC transporters, which accounted for approximately 3% of inferred genes. These findings provided a comprehensive profile of bacterial communities, including potential bacterial pathogens and antibiotic-resistant genes, in raw and secondary treated WW samples from four WWTPs across Australia and demonstrated that Illumina high-throughput sequencing can be an alternative approach for monitoring WW quality in order to protect environmental and human health.

     

Transgenic tobacco revealing altered bacterial diversity in the rhizosphere during early plant development

The rhizosphere constitutes a complex niche that may be exploited by a wide variety of bacteria. Bacterium–plant interactions in this niche can be influenced by factors such as the expression of heterologous genes in the plant. The objective of this work was to describe the bacterial communities associated with the rhizosphere and rhizoplane regions of tobacco plants, and to compare communities from transgenic tobacco lines (CAB1, CAB2 and TRP) with those found in wild-type (WT) plants. Samples were collected at two stages of plant development, the vegetative and flowering stages (1 and 3 months after germination). The diversity of the culturable microbial community was assessed by isolation and further characterization of isolates by amplified ribosomal RNA gene restriction analysis (ARDRA) and 16S rRNA sequencing. These analyses revealed the presence of fairly common rhizosphere organisms with the main groups Alphaproteobacteria, Betaproteobacteria, Actinobacteria and Bacilli. Analysis of the total bacterial communities using PCR-DGGE (denaturing gradient gel electrophoresis) revealed that shifts in bacterial communities occurred during early plant development, but the reestablishment of original community structure was observed over time. The effects were smaller in rhizosphere than in rhizoplane samples, where selection of specific bacterial groups by the different plant lines was demonstrated. Clustering patterns and principal components analysis (PCA) were used to distinguish the plant lines according to the fingerprint of their associated bacterial communities. Bands differentially detected in plant lines were found to be affiliated with the genera Pantoea, Bacillus and Burkholderia in WT, CAB and TRP plants, respectively. The data revealed that, although rhizosphere/rhizoplane microbial communities can be affected by the cultivation of transgenic plants, soil resilience may be able to restore the original bacterial diversity after one cycle of plant cultivation.     

Turkey fecal microbial community structure and functional gene diversity revealed by 16S rRNA gene and metagenomic sequences

The primary goal of this study was to better understand the microbial composition and functional genetic diversity associated with turkey fecal communities. To achieve this, 16S rRNA gene and metagenomic clone libraries were sequenced from turkey fecal samples. The analysis of 382 16S rRNA gene sequences showed that the most abundant bacteria were closely related to Lactobacillales (47%), Bacillales (31%), and Clostridiales (11%). Actinomycetales, Enterobacteriales, and Bacteroidales sequences were also identified, but represented a smaller part of the community. The analysis of 379 metagenomic sequences showed that most clones were similar to bacterial protein sequences (58%). Bacteriophage (10%) and avian viruses (3%) sequences were also represented. Of all metagenomic clones potentially encoding for bacterial proteins, most were similar to low G+C Gram-positive bacterial proteins, particularly from Lactobacillales (50%), Bacillales (11%), and Clostridiales (8%). Bioinformatic analyses suggested the presence of genes encoding for membrane proteins, lipoproteins, hydrolases, and functional genes associated with the metabolism of nitrogen and sulfur containing compounds. The results from this study further confirmed the predominance of Firmicutes in the avian gut and highlight the value of coupling 16S rRNA gene and metagenomic sequencing data analysis to study the microbial composition of avian fecal microbial communities.     

Thermal stress responses in the bacterial biosphere of the Great Barrier Reef sponge,Rhopaloeides odorabile

Marine sponges are diverse, abundant and provide a crucial coupling point between benthic and pelagic habitats due to their high filtration rates. They also harbour extensive microbial communities, with many microbial phylotypes found exclusively in sponge hosts and not in the seawater or surrounding environment, i.e. so‐called sponge‐specific clusters (SCs) or sponge‐ and coral‐specific clusters (SCCs). We employed DNA (16S rRNA gene) and RNA (16S rRNA)‐based amplicon pyrosequencing to investigate the effects of sublethal thermal stress on the bacterial biosphere of the Great Barrier Reef sponge Rhopaloeides odorabile. A total of 8381 operational taxonomic units (OTUs) (97% sequence similarity) were identified, affiliated with 32 bacterial phyla from seawater samples, 23 bacterial phyla from sponge DNA extracts and 18 bacterial phyla from sponge RNA extracts. Sublethal thermal stress (31°C) had no effect on the present and/or active portions of the R. odorabile bacterial community but a shift in the bacterial assemblage was observed in necrotic sponges. Over two‐thirds of DNA and RNA sequences could be assigned to previously defined SCs/SCCs in healthy sponges whereas only 12% of reads from necrotic sponges could be assigned to SCs/SCCs. A rapid decline in host health over a 1°C temperature increment suggests that sponges such as R. odorabile may be highly vulnerable to the effects of global climate change.     

Microbial diversity in degraded and non-degraded petroleum samples and comparison across oil reservoirs at local and global scales

Microorganisms have shown their ability to colonize extreme environments including deep subsurface petroleum reservoirs. Physicochemical parameters may vary greatly among petroleum reservoirs worldwide and so do the microbial communities inhabiting these different environments. The present work aimed at the characterization of the microbiota in biodegraded and non-degraded petroleum samples from three Brazilian reservoirs and the comparison of microbial community diversity across oil reservoirs at local and global scales using 16S rRNA clone libraries. The analysis of 620 16S rRNA bacterial and archaeal sequences obtained from Brazilian oil samples revealed 42 bacterial OTUs and 21 archaeal OTUs. The bacterial community from the degraded oil was more diverse than the non-degraded samples. Non-degraded oil samples were overwhelmingly dominated by gammaproteobacterial sequences with a predominance of the genera Marinobacter and Marinobacterium. Comparisons of microbial diversity among oil reservoirs worldwide suggested an apparent correlation of prokaryotic communities with reservoir temperature and depth and no influence of geographic distance among reservoirs. The detailed analysis of the phylogenetic diversity across reservoirs allowed us to define a core microbiome encompassing three bacterial classes (Gammaproteobacteria, Clostridia, and Bacteroidia) and one archaeal class (Methanomicrobia) ubiquitous in petroleum reservoirs and presumably owning the abilities to sustain life in these environments.

     

Simultaneous Amplification of Two Bacterial Genes: More Reliable Method of <Emphasis Type="Italic">Helicobacter pylori</Emphasis> Detection in Microbial Rich Dental Plaque Samples

Polymerase Chain reaction (PCR) assay is considered superior to other methods for detection of Helicobacter pylori (H. pylori) in oral cavity; however, it also has limitations when sample under study is microbial rich dental plaque. The type of gene targeted and number of primers used for bacterial detection in dental plaque samples can have a significant effect on the results obtained as there are a number of closely related bacterial species residing in plaque biofilm. Also due to high recombination rate of H. pylori some of the genes might be down regulated or absent. The present study was conducted to determine the frequency of H. pylori colonization of dental plaque by simultaneously amplifying two genes of the bacterium. One hundred dental plaque specimens were collected from dyspeptic patients before their upper gastrointestinal endoscopy and presence of H. pylori was determined through PCR assay using primers targeting two different genes of the bacterium. Eighty-nine of the 100 samples were included in final analysis. With simultaneous amplification of two bacterial genes 51.6% of the dental plaque samples were positive for H. pylori while this prevalence increased to 73% when only one gene amplification was used for bacterial identification. Detection of H. pylori in dental plaque samples is more reliable when two genes of the bacterium are simultaneously amplified as compared to one gene amplification only.     

Population ecology of nitrifying Archaea and Bacteria in the Southern California Bight

Marine Crenarchaeota are among the most abundant microbial groups in the ocean, and although relatively little is currently known about their biogeochemical roles in marine ecosystems, recognition that Crenarchaeota posses ammonia monooxygenase (amoA) genes and may act as ammonia‐oxidizing archaea (AOA) offers another means of probing the ecology of these microorganisms. Here we use a time series approach combining quantification of archaeal and bacterial ammonia oxidizers with bacterial community fingerprints and biogeochemistry, to explore the population and community ecology of nitrification. At multiple depths (150, 500 and 890 m) in the Southern California Bight sampled monthly from 2003 to 2006, AOA were enumerated via quantitative PCR of archaeal amoA and marine group 1 Crenarchaeota 16S rRNA genes. Based on amoA genes, AOA were highly variable in time – a consistent feature of marine Crenarchaeota– however, average values were similar at different depths and ranged from 2.20 to 2.76 × 10⁴amoA copies ml^?1. Archaeal amoA genes were correlated with Crenarchaeota 16S rRNA genes (r² = 0.79) and the slope of this relationship was 1.02, demonstrating that the majority of marine group 1 Crenarchaeota present over the dates and depths sampled possessed amoA. Two AOA clades were specifically quantified and compared with betaproteobacterial ammonia‐oxidizing bacteria (β‐AOB) amoA genes at 150 m; these AOA groups were found to strongly co‐vary in time (r² = 0.70, P < 0.001) whereas AOA : β‐AOB ratios ranged from 13 to 5630. Increases in the AOA : β‐AOB ratio correlated with the accumulation of nitrite (r² = 0.87, P < 0.001), and may be indicative of differences in substrate affinities and activities leading to periodic decoupling between ammonia and nitrite oxidation. These data capture a dynamic nitrogen cycle in which multiple microbial groups appear to be active participants.     

干湿交替水分胁迫对水稻土细菌群落影响的研究

[目的]连续3次风干-湿润循环培养水稻土,在DNA和RNA水平下,探究细菌对干湿交替胁迫的响应机制,明确风干水稻土能否代替新鲜土壤进行细菌群落组成分析.[方法]针对我国江苏省常熟市水稻土,开展新鲜土壤的3次风干-湿润循环连续培养处理(每次循环中风干、湿润状态各维持7 d),在DNA和RNA水平应用16S rRNA基因高...     

Spatial and Temporal Variations in Bacterial Macromolecule Labeling with [methyl-3H]Thymidine in a Hypertrophic Lake

The incorporation of [methyl-³H]thymidine into three macromolecular fractions, designated as DNA, RNA, and protein, by bacteria from Hartbeespoort Dam, South Africa, was measured over 1 year by acid-base hydrolysis procedures. Samples were collected at 10 m, which was at least 5 m beneath the euphotic zone. On four occasions, samples were concurrently collected at the surface. Approximately 80% of the label was incorporated into bacterial DNA in surface samples. At 10 m, total incorporation of label into bacterial macromolecules was correlated to bacterial utilization of glucose (r = 0.913, n = 13, P < 0.001). The labeling of DNA, which ranged between 0 and 78% of total macromolecule incorporation, was inversely related to glucose uptake (r = -0.823), total thymidine incorporation (r = -0.737), and euphotic zone algal production (r = -0.732, n = 13, P < 0.005). With decreased DNA labeling, increasing proportions of label were found in the RNA fraction and proteins. Enzymatic digestion followed by chromatographic separation of macromolecule fragments indicated that DNA and proteins were labeled while RNA was not. The RNA fraction may represent labeled lipids or other macromolecules or both. The data demonstrated a close coupling between phytoplankton production and heterotrophic bacterial activity in this hypertrophic lake but also confirmed the need for the routine extraction and purification of DNA during [methyl-³H]thymidine studies of aquatic bacterial production.     

Metagenomic analysis of microbial communities across a transect from low to highly hydrocarbon-contaminated soils in King George Island,Maritime Antarctica

Soil samples from a transect from low to highly hydrocarbon-contaminated soils were collected around the Brazilian Antarctic Station Comandante Ferraz (EACF), located at King George Island, Antarctica. Quantitative PCR (qPCR) analysis of bacterial 16S rRNA genes, 16S rRNA gene (iTag), and shotgun metagenomic sequencing were used to characterize microbial community structure and the potential for petroleum degradation by indigenous microbes. Hydrocarbon contamination did not affect bacterial abundance in EACF soils (bacterial 16S rRNA gene qPCR). However, analysis of 16S rRNA gene sequences revealed a successive change in the microbial community along the pollution gradient. Microbial richness and diversity decreased with the increase of hydrocarbon concentration in EACF soils. The abundance of Cytophaga, Methyloversatilis, Polaromonas, and Williamsia was positively correlated (p-value = <.05) with the concentration of total petroleum hydrocarbons (TPH) and/or polycyclic aromatic hydrocarbons (PAH). Annotation of metagenomic data revealed that the most abundant hydrocarbon degradation pathway in EACF soils was related to alkyl derivative-PAH degradation (mainly methylnaphthalenes) via the CYP450 enzyme family. The abundance of genes related to nitrogen fixation increased in EACF soils as the concentration of hydrocarbons increased. The results obtained here are valuable for the future of bioremediation of petroleum hydrocarbon-contaminated soils in polar environments.     

Soil microbiome responses to the short‐term effects of Amazonian deforestation

Slash‐and‐burn clearing of forest typically results in increase in soil nutrient availability. However, the impact of these nutrients on the soil microbiome is not known. Using next generation sequencing of 16S rRNA gene and shotgun metagenomic DNA, we compared the structure and the potential functions of bacterial community in forest soils to deforested soils in the Amazon region and related the differences to soil chemical factors. Deforestation decreased soil organic matter content and factors linked to soil acidity and raised soil pH, base saturation and exchangeable bases. Concomitant to expected changes in soil chemical factors, we observed an increase in the alpha diversity of the bacterial microbiota and relative abundances of putative copiotrophic bacteria such as Actinomycetales and a decrease in the relative abundances of bacterial taxa such as Chlamydiae, Planctomycetes and Verrucomicrobia in the deforested soils. We did not observe an increase in genes related to microbial nutrient metabolism in deforested soils. However, we did observe changes in community functions such as increases in DNA repair, protein processing, modification, degradation and folding functions, and these functions might reflect adaptation to changes in soil characteristics due to forest clear‐cutting and burning. In addition, there were changes in the composition of the bacterial groups associated with metabolism‐related functions. Co‐occurrence microbial network analysis identified distinct phylogenetic patterns for forest and deforested soils and suggested relationships between Planctomycetes and aluminium content, and Actinobacteria and nitrogen sources in Amazon soils. The results support taxonomic and functional adaptations in the soil bacterial community following deforestation. We hypothesize that these microbial adaptations may serve as a buffer to drastic changes in soil fertility after slash‐and‐burning deforestation in the Amazon region.     

Regulation of ribonucleic acid synthesis in Escherichia coli B-r: an analysis of a shift-up. 1. Ribosomal RNA chain growth rates

The chain growth rate for ribosomal RNA was determined for Escherichia coli$\text{B}\text{r}$ growing in succinate (μ = 0.69 doublings/h), glucose (μ = 1.36) and glucose/ amino acids (μ = 2.10) medium. With increasing bacterial growth rate the chain growth rate increases from 4400 to 6300 nucleotides/min. These values are almost twofold higher than the chain growth rate reported for messenger RNA; this implies that, following a nutritional shift-up, the transfer of a relatively small number of RNA polymerase molecules from unstable to stable RNA genes along with the increase in the stable RNA chain growth rate is sufficient to account for the abrupt increase in the net rate of RNA synthesis. Furthermore, our calculations indicate that the linear density of polymerase molecules on the ribosomal DNA template increases with the bacterial growth rate, such that in rapidly growing bacteria all ribosomal RNA genes (48 copies at μ = 3) are nearly saturated with RNA polymerase.     

Microbiome of Pukzing Cave in India shows high antimicrobial activity against plant and animal pathogens

Pukzing cave, the largest cave of Mizoram, India was explored for bacterial diversity. Culture dependent method revealed 235 bacterial isolates using three different treatments. Identity of the microbial species was confirmed by 16S rDNA sequencing. The highest bacterial population was recovered from heat treatment (n = 97;41.2%) followed by normal (n = 79;33.6%) and cold treatment (n = 59;25.1%) indicating dominance of moderate thermophiles. Antimicrobial potential of isolates showed 20.4% isolates having antimicrobial ability against tested pathogens. Amplicon sequencing of PKSI, PKSII and NRP specific genes revealed presence of AMP genes in the microbial population. Six microbial pathogens were selected for screening as they are well known for different disease cause organism in various fields such as agriculture and human health. Cave environment harbors unique microbial flora and hypervariable region V4 is more informative. Higher activity of AMP assay against these microbes indicates that cave microbial communities could be potential source of future genomic resources.     

Bacterial endosymbiont infections in ‘living fossils’: a case study of North American vaejovid scorpions

Bacterial endosymbionts are common among arthropods, and maternally inherited forms can affect the reproductive and behavioural traits of their arthropod hosts. The prevalence of bacterial endosymbionts and their role in scorpion evolution have rarely been investigated. In this study, 61 samples from 40 species of scorpion in the family Vaejovidae were screened for the presence of the bacterial endosymbionts Cardinium, Rickettsia, Spiroplasma and Wolbachia. No samples were infected by these bacteria. However, one primer pair specifically designed to amplify Rickettsia amplified nontarget genes of other taxa. Similar off‐target amplification using another endosymbiont‐specific primer was also found during preliminary screenings. Results caution against the overreliance on previously published screening primers to detect bacterial endosymbionts in host taxa and suggest that primer specificity may be higher in primers targeting nuclear rather than mitochondrial genes.     

Analysis of Bacterial and Fungal Communities in Japanese- and Chinese-Fermented Soybean Pastes Using Nested PCR–DGGE

The microbial diversity of Japanese- and Chinese-fermented soybean pastes was investigated using nested PCR–denaturing gradient gel electrophoresis (DGGE). Five Japanese-fermented soybean paste samples and three Chinese-fermented soybean paste samples were analyzed for bacteria and fungi. Extracted DNA was used as a template for PCR to amplify 16S rRNA and 18S rRNA genes. The nearly complete 16S rRNA and 18S rRNA genes were amplified using universal primers, and the resulting products were subsequently used as a template in a nested PCR to obtain suitable fragments for DGGE. Tetragenococcus halophilus and Staphylococcus gallinarum were found to dominate the bacterial microbiota in Japanese samples, whereas Bacillus sp. was detected as the predominant species in Chinese samples. DGGE analysis of fungi in soybean pastes determined the presence of Aspergillus oryzae and Zygosaccharomyces rouxii in most of the Chinese and Japanese samples. Some differences were observed in the bacterial diversity of Japanese- and Chinese-fermented soybean pastes.     

Microbiota of the <Emphasis Type="Italic">Planalto de Bolona</Emphasis>: an artisanal cheese produced in uncommon environmental conditions in the Cape Verde Islands

The present study aimed to evaluate the dominant microbial community naturally present in the Planalto de Bolona cheese, produced in the Cape Verde Islands. Samples of milk, curd and cheese from two different producers were examined through culture-dependent and independent-methods. Traditional plating and genetic identification of lactic acid bacteria (LAB) and yeast isolates were carried out. Moreover, DNA and RNA extracted directly from samples were subjected to Polymerase Chain Reaction-Denaturing Gradient Gel Electrophoresis (PCR-DGGE). Concerning the LAB population, a total of 278 isolates were identified: Lactococcus lactis subsp. lactis and Enterococcus faecium represented the most isolated species. Regarding yeasts, the analysis of isolates throughout the manufacturing period showed a consistent presence of the genus Candida. Divergences in species detection between culture-dependent and culture-independent methods were observed, as well as between DNA and RNA analysis. PCR-DGGE underlined high heterogeneity among bacterial species while yeast microbiota was dominated by Aureobasidium pullulans at DNA level. The obtained results represent a first approach in the understanding of the Planalto de Bolona cheese microbial ecology and consequently may constitute a first step towards the full comprehension of the microbiota of this artisanal cheese produced in unusual environmental conditions in the Cape Verde Islands.     

Microbial community analysis at crude oil‐contaminated soils targeting the 16S ribosomal RNA,xylM, C23O,and bcr genes

Aims: The analyses targeting multiple functional genes were performed on the samples of crude oil‐contaminated soil, to investigate community structures of organisms involved in monoaromatic hydrocarbon degradation. Methods and Results: Environmental samples were obtained from two sites that were contaminated with different components of crude oil. The analysis on 16S rRNA gene revealed that bacterial community structures were clearly different between the two sites. The cloning analyses were performed by using primers specific for the catabolic genes involved in the aerobic or anaerobic degradation of monoaromatic hydrocarbons, i.e. xylene monooxygenase (xylM), catechol 2,3‐dioxygenase (C23O), and benzoyl‐CoA reductase (bcr) genes. From the result of xylM gene, it was suggested that there are lineages specific to the respective sites, reflecting the differences of sampling sites. In the analysis of the C23O gene, the results obtained with two primer sets were distinct from each other. A comparison of these suggested that catabolic types of major bacteria carrying this gene were different between the two sites. As for the bcr gene, no amplicon was obtained from one sample. Phylogenetic analysis revealed that the sequences obtained from the other sample were distinct from the known sequences. Conclusions: The differences between the two sites were demonstrated in the analyses of all tested genes. As for aerobic cleavage of the aromatic ring, it was also suggested that analysis using two primer sets provide more detailed information about microbial communities in the contaminated site. Significance and Impact of the Study: The present study demonstrated that analysis targeting multiple functional genes as molecular markers is practical to examine microbial community in crude oil‐contaminated environments.     

Microbial diversity of Loki's Castle black smokers at the Arctic Mid‐Ocean Ridge

Hydrothermal vent systems harbor rich microbial communities ranging from aerobic mesophiles to anaerobic hyperthermophiles. Among these, members of the archaeal domain are prevalent in microbial communities in the most extreme environments, partly because of their temperature‐resistant and robust membrane lipids. In this study, we use geochemical and molecular microbiological methods to investigate the microbial diversity in black smoker chimneys from the newly discovered Loki's Castle hydrothermal vent field on the Arctic Mid‐Ocean Ridge (AMOR) with vent fluid temperatures of 310–320 °C and pH of 5.5. Archaeal glycerol dialkyl glycerol tetraether lipids (GDGTs) and H‐shaped GDGTs with 0–4 cyclopentane moieties were dominant in all sulfide samples and are most likely derived from both (hyper)thermophilic Euryarchaeota and Crenarchaeota. Crenarchaeol has been detected in low abundances in samples derived from the chimney exterior indicating the presence of Thaumarchaeota at lower ambient temperatures. Aquificales and members of the Epsilonproteobacteria were the dominant bacterial groups detected. Our observations based on the analysis of 16S rRNA genes and biomarker lipid analysis provide insight into microbial communities thriving within the porous sulfide structures of active and inactive deep‐sea hydrothermal vents. Microbial cycling of sulfur, hydrogen, and methane by archaea in the chimney interior and bacteria in the chimney exterior may be the prevailing biogeochemical processes in this system.