High Prevalence of Methanobrevibacter smithii and Methanosphaera stadtmanae Detected in the Human Gut Using an Improved DNA Detection Protocol

Dridi, Bédis; Henry, Mireille; Khechine, Amel El; Raoult, Didier; Drancourt, Michel

doi:10.1371/journal.pone.0007063

Cited by 331 publications

(319 citation statements)

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“…Methanobrevibacter smithii and Methanosphaera stadtmanae (Methanobacteriales) are considered to be the prevalent methanogens in the human gut (59). In our study, sequences belonging to both species were identified in DGGE fingerprints from inlet wastewater samples and in pyrotag libraries from the first weeks of biofilm development, suggesting that archaeal colonizers at early stages of biofilm development derive from human fecal material in wastewater.…”

Section: Discussionmentioning

confidence: 56%

Changes in Microbial Biofilm Communities during Colonization of Sewer Systems

Auguet

Pijuan

Batista

et al. 2015

Appl Environ Microbiol

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bThe coexistence of sulfate-reducing bacteria (SRB) and methanogenic archaea (MA) in anaerobic biofilms developed in sewer inner pipe surfaces favors the accumulation of sulfide (H 2 S) and methane (CH 4 ) as metabolic end products, causing severe impacts on sewerage systems. In this study, we investigated the time course of H 2 S and CH 4 production and emission rates during different stages of biofilm development in relation to changes in the composition of microbial biofilm communities. The study was carried out in a laboratory sewer pilot plant that mimics a full-scale anaerobic rising sewer using a combination of process data and ). This contrasting trend coincided with a stable SRB community and an archaeal community composed solely of methanogens derived from the human gut (i.e., Methanobrevibacter and Methanosphaera). In turn, CH 4 emissions increased after 1 year of biofilm growth (327.6 ؎ 16.6 mg COD-CH 4 liter ؊1 day ؊1 ), coinciding with the replacement of methanogenic colonizers by species more adapted to sewer conditions (i.e., Methanosaeta spp.). Our study provides data that confirm the capacity of our laboratory experimental system to mimic the functioning of full-scale sewers both microbiologically and operationally in terms of sulfide and methane production, gaining insight into the complex dynamics of key microbial groups during biofilm development. W astewater collection systems, or sewers, consist of an underground network of physical structures-installations composed of pipelines, pumping stations, manholes, and channels that convey wastewaters from their source to the discharge point, usually a wastewater treatment plant (WWTP). Sewer systems thus prevent the direct contact of urban populations to fecal material and potential microbial pathogens, greatly reducing the spread of infectious diseases. Sewers have traditionally been considered only hydraulic transport systems for sewage, although they are in fact "reactors" where complex physicochemical and microbial processes take place. Wastewater microorganisms are diverse and abundant, and they are exposed to a wide range of both inorganic and organic substrates as well as changing conditions along their transport through sewers (1). In this regard, wastewater transport through the pipes facilitates the formation of microbial biofilms that grow attached to the inner surface of sewer pipes (2). Different factors, such as large surface area, low flow velocity near pipe walls, and nutrient availability, may favor microbial colonization of pipe surfaces and biofilm growth. Formation of fully functional biofilms occurs in different steps, from surface conditioning, adhesion of microbial "colonizers," initial growth, and glycocalyx formation, followed by secondary colonization and growth (3).Anaerobic conditions in sewer pipes favor the accumulation of both sulfide (H 2 S) and methane (CH 4 ) as end products of different microbial metabolisms, i.e., anaerobic respiration of organic matter by sulfate-reducing bacteria (SRB) and methanogenic archae...

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Section: Discussionmentioning

confidence: 56%

Changes in Microbial Biofilm Communities during Colonization of Sewer Systems

Auguet

Pijuan

Batista

et al. 2015

Appl Environ Microbiol

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“…The relative abundance of the mcrA gene also appeared to be stable and stayed at low levels (log10-4.82 ± 0.82 at day 1, log10-5.2 ± 0.26 at day 2, log10-5.15 ± 0.76 at day 4, log10-3.82±1.18 at day 8 and log10-4.93 ± 0.64 at day16) over the entire duration of the conventionalisation, suggesting that the methanogen population was among the early colonisers and had already reached its final population size early in the experiment. Although the relatively low absolute abundance of the mcrA gene may be an underestimation due to DNA isolation procedures that were not optimised for lysis of the methanogens (Dridi et al, 2009), the stability of the gene's abundance in the microbiota is most probably an accurate reflection of the evolution of this microbial group over time. In contrast, the dsr gene appeared to be B100-fold more abundant in the ecosystem during later stages of the conventionalisation (Figure 2a), increasing from Blog10-2.54 ± 0.2 during days 1-2 to log10-4.4±0.86 at days 4, 8 and 16 post-conventionalisation.…”

Section: Dynamic Establishment Of Microbial Communitiesmentioning

confidence: 99%

Gut bacteria–host metabolic interplay during conventionalisation of the mouse germfree colon

et al. 2012

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The interplay between dietary nutrients, gut microbiota and mammalian host tissues of the gastrointestinal tract is recognised as highly relevant for host health. Combined transcriptome, metabonome and microbial profiling tools were employed to analyse the dynamic responses of germfree mouse colonic mucosa to colonisation by normal mouse microbiota (conventionalisation) at different time-points during 16 days. The colonising microbiota showed a shift from early (days 1 and 2) to later colonisers (days 8 and 16). The dynamic changes in the microbial community were rapidly reflected by the urine metabolic profiles (day 1) and at later stages (day 4 onward) by the colon mucosa transcriptome and metabolic profiles. Correlations of host transcriptomes, metabolite patterns and microbiota composition revealed associations between Bacilli and Proteobacteria, and differential expression of host genes involved in energy and anabolic metabolism. Differential gene expression correlated with scyllo-and myo-inositol, glutamine, glycine and alanine levels in colonic tissues during the time span of conventionalisation. Our combined time-resolved analyses may help to expand the understanding of host-microbe molecular interactions during the microbial establishment.

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“…Results were expressed as the number of M. oralis cell/ml of specimen (Table 1). A quantification synthetic plasmid was used as an internal control to monitor PCR inhibition, and the total bacterial load was measured as previously described (11). The PCR program was 95°C for 5 min, followed by 40 cycles of 95°C for 1 s, 60°C for 35 s, and 45°C for 30 s. A calibration curve was done by measuring the cycle threshold (C T ) value of a serial dilution of M. oralis (10Eϩ01 to 10Eϩ09).…”

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confidence: 99%

Real-Time PCR Quantification of Methanobrevibacter oralis in Periodontitis

et al. 2013

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bA real-time PCR assay developed to quantify Methanobrevibacter oralis indicated that its inoculum significantly correlated with periodontitis severity (P ‫؍‬ 0.003), despite a nonsignificant difference in prevalence between controls (3/10) and patients (12/22) (P ‫؍‬ 0.2, Fisher test). The M. oralis load can be used as a biomarker for periodontitis. Periodontitis is an anaerobic infection possibly leading to loss of teeth (1-3). It likely results from infection with microbial complexes comprising methanogens (4, 5), mostly Methanobrevibacter oralis (6)(7)(8). In this study, we correlated the M. oralis load as measured by real-time PCR in subgingival plaque with the severity of periodontitis.All patients diagnosed with periodontitis from October 2011 to June 2012 were included. The control individuals with generally healthy gingiva were volunteers. All patients underwent interviews for medical and dental history, intraoral examination to determine bleeding on probing (BOP), probing depth (PD), and plaque index (PI), and radiographs (9); periodontal status was also scored as described previously (10) ( Table 1). All individuals signed an informed-consent form, and the ethics committee of the IFR 48, University of Aix-Marseille, approved the protocol.Subgingival plaque specimens (50 l) collected from 3-to 12-mm periodontal pockets in both patients and controls were suspended in 1 ml Tris-HCl (0.05 M, pH 7.5) buffer. After homogenization, a 250-l aliquot was shaken with 0.3 g of acid-washed beads (Յ106 mm; Sigma, Saint-Quentin-Fallavier, France) in a FastPrep-24 instrument (MP Biomedical Europe, Illkirch, France) at a speed of 6.5 m/s (full speed) for 90 s. The supernatant was incubated overnight at 56°C with 180 ml of lysis buffer and 25 l proteinase K (20 mg/ml) in the Qiagen EZ1 DNA tissue kit (Qiagen, Courtaboeuf, France). After a second cycle of mechanical lysis, the supernatant was incubated for 10 min at 100°C, and total DNA was then extracted using the same kit. Negative controls consisting of sterile DNA-free water were introduced in all the manipulations. The specificity of the M. oralis-cnp602P probe (6-carboxyfluorescein [FAM]-5=AGCAGTGCACCTGCTGATA TGGAAGG-3=) (Applied Biosystems, Courtaboeuf, France) and the primer pair M. oralis-cnp602F (5=-GCTGGTGTAATCGAAC CTAAACG-3=) and M. oralis-cnp602R (5=-CACCCATACCCGG ATCCATA-3=) (Eurogentec, Seraing, Belgium) was verified in silico using the BLAST program at NCBI (http://www.ncbi.nlm .nih.gov/BLAST) and further experimentally ensured by incorporating DNA extracted from Methanobrevibacter smithii ATCC 35061 T , Methanosphaera stadtmanae ATCC 43021 T , Methanomassiliicoccus luminyensis CSURP135 T , Escherichia coli, Salmonella enterica, Staphylococcus aureus, and Treponema denticola ATCC 35405T . Real-time PCR assays were performed with a CFX96 Touch real-time PCR detection system (Bio-Rad, Marnes-laCoquette, France) using the Mastermix PCR kit (Eurogentec) with 5 pmol of each primer and probe and 5 l of about 2 g of DNA into a 20-l final volume. M. oralis DSMZ ...

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High Prevalence of Methanobrevibacter smithii and Methanosphaera stadtmanae Detected in the Human Gut Using an Improved DNA Detection Protocol

Cited by 331 publications

References 28 publications

Changes in Microbial Biofilm Communities during Colonization of Sewer Systems

Changes in Microbial Biofilm Communities during Colonization of Sewer Systems

Gut bacteria–host metabolic interplay during conventionalisation of the mouse germfree colon

Real-Time PCR Quantification of Methanobrevibacter oralis in Periodontitis

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