Identification of bacterial populations in drinking water using 16S rRNA-based sequence analyses

Revetta, Randy P.; Pemberton, Adin; Lamendella, Regina; Iker, Brandon C.; Domingo, Jorge W. Santo

doi:10.1016/j.watres.2009.11.008

Cited by 100 publications

(78 citation statements)

References 40 publications

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“…Across all gated events, 232 total 16S sequences were within the domain Bacteria. Some of the most highly detected genera such as Acinetobacter and Sphingomonas are consistent with those found in microbial communities associated with drinking water distribution systems (17,38,39). From the 32 wells with 100 low-fluorescent events in each, 6 wells produced a 16S sequence.…”

Section: Resultssupporting

confidence: 66%

Candidate phylum TM6 genome recovered from a hospital sink biofilm provides genomic insights into this uncultivated phylum

McLean

Lombardo

Badger

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

169

166

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The "dark matter of life" describes microbes and even entire divisions of bacterial phyla that have evaded cultivation and have yet to be sequenced. We present a genome from the globally distributed but elusive candidate phylum TM6 and uncover its metabolic potential. TM6 was detected in a biofilm from a sink drain within a hospital restroom by analyzing cells using a highly automated single-cell genomics platform. We developed an approach for increasing throughput and effectively improving the likelihood of sampling rare events based on forming small random pools of single-flow-sorted cells, amplifying their DNA by multiple displacement amplification and sequencing all cells in the pool, creating a "mini-metagenome." A recently developed single-cell assembler, SPAdes, in combination with contig binning methods, allowed the reconstruction of genomes from these mini-metagenomes. A total of 1.07 Mb was recovered in seven contigs for this member of TM6 (JCVI TM6SC1), estimated to represent 90% of its genome. High nucleotide identity between a total of three TM6 genome drafts generated from pools that were independently captured, amplified, and assembled provided strong confirmation of a correct genomic sequence. TM6 is likely a Gram-negative organism and possibly a symbiont of an unknown host (nonfree living) in part based on its small genome, low-GC content, and lack of biosynthesis pathways for most amino acids and vitamins. Phylogenomic analysis of conserved single-copy genes confirms that TM6SC1 is a deeply branching phylum. (1), which is accomplished using multiple displacement amplification (MDA) (2-5) of genomic DNA to obtain sufficient template. We applied a high-throughput strategy to capture and sequence genomes of bacteria from a biofilm in a hospital sink including pathogens, such as the oral periodontal pathogen (Porphyromonas gingivalis) (6) and uncultivated members (this study). Despite the fact that a typical person spends ∼90% of their time indoors (7), our knowledge of the microbial diversity of the indoor environment has only recently begun to be explored using culture-independent methods (8, 9). Biofilms within water distribution systems in particular are thought to be diverse microbial communities and potential reservoirs of disease-causing organisms in the indoor environment. Several pathogens including Escherichia coli, Legionella pneumophila (10-13), Vibrio cholerae (14), and Helicobacter pylori (15, 16) have been detected in biofilms within water distribution systems. A recent 16S rRNA gene (abbreviated henceforth as 16S unless otherwise stated) molecular survey also revealed significant loads of Mycobacterium avium in showerhead biofilms (17). Based on these findings, indoor environments can clearly serve as significant reservoirs of pathogenic bacteria, and therefore there is great interest in investigating the rare and abundant bacterial species within biofilms in these environments.One approach to capture uncultivated bacteria is to isolate single bacterial cells by fluorescent activat...

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Section: Resultssupporting

confidence: 66%

Candidate phylum TM6 genome recovered from a hospital sink biofilm provides genomic insights into this uncultivated phylum

McLean

Lombardo

Badger

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

169

166

View full text Add to dashboard Cite

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“…Because DNA-based approaches could not differentiate between dead and live cells in the environments, it is difficult to get a true profile of indigenous eukaryotes under the interference of exogenous 18S rRNA gene. To resolve this issue, RNA targeting assays may be promising, which have been used to study active bacterial communities in bulk water and drinking water biofilm (Keinanen-Toivola and Revetta, 2006;Revetta et al, 2010).…”

Section: Eukaryotic Communitiesmentioning

confidence: 99%

Pyrosequencing analysis of eukaryotic and bacterial communities in faucet biofilms

Liu

Guo

et al. 2012

Science of The Total Environment

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“…Although these techniques provide for important screening tools for recent fecal contamination, the majority of microorganisms present in the environment are not culturable and methods such as high throughput sequencing are required to characterize unculturable bacteria in water supplies (Eichler et al, 2006;Revetta et al, 2010;Lin et al, 2014;Gomez-Alvarez et al, 2016).…”

Section: Introductionmentioning

confidence: 99%

Bacteria in drinking water sources of a First Nation reserve in Canada

Farenhorst

Jahan

et al. 2017

Science of The Total Environment

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Approximately 20% of the 600 First Nations reserves across Canada are under a drinking water advisory, often due to unacceptable levels of bacteria. In this study, we detected fecal bacteria at an alarmingly high frequency in drinking water sources in a fly-in First Nations community, most notably in buckets/drums of homes without running water where Escherichia coli levels ranged from 20 to 62,000 CFU/100mL. The water leaving the water treatment plant was free of E. coli and its free residual chlorine concentration (0.67 mg/L) was within the range typically observed for treated water in Canada. Water samples from taps in homes served by cisterns, and those sampled from the water truck and community standpipe, always showed unacceptable levels of E. coli (1 to 2,100 CFU/100mL) and free residual chlorine concentrations below the 0.2 mg/L required to prevent bacterial regrowth. Samples from taps in homes served by piped water had lower levels of E. coli (0 to 2 CFU/100mL). DNA-and RNA-based 16S rRNA Illumina sequencing demonstrated that piped and cisterns water distribution systems showed an abundance of viable cells of Alphaproteobacteria indicative of biofilm formation in pipes and cisterns. The alpha diversity, based on observed OTUs and three other indices, was lowest in water truck samples that supplied water to the cistern and the low free residual chlorine concentration (0.07 mg/L) and predominance of Betaproteobacteria (63% of viable cells) that were immediately detected after the truck had filled up at the water treatment plant was indicative of contamination by particulate matter. Given these findings, First Nation residents living without running water and relying on inadequate water distribution systems are at higher risk of contracting water-born illnesses. We urge all governments in Canada to expand their investments in supporting and sustaining water as a human right in Canada's First Nations communities. 3 GRAPHICAL ABSTRACT

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Identification of bacterial populations in drinking water using 16S rRNA-based sequence analyses

Cited by 100 publications

References 40 publications

Candidate phylum TM6 genome recovered from a hospital sink biofilm provides genomic insights into this uncultivated phylum

Candidate phylum TM6 genome recovered from a hospital sink biofilm provides genomic insights into this uncultivated phylum

Pyrosequencing analysis of eukaryotic and bacterial communities in faucet biofilms

Bacteria in drinking water sources of a First Nation reserve in Canada

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