Combining flow cytometry and 16S rRNA gene pyrosequencing: A promising approach for drinking water monitoring and characterization

Prest, Emmanuelle I.; El-Chakhtoura, J.; Hammes, Frederik; Saikaly, Pascal E.; Loosdrecht, M.C.M. van; Vrouwenvelder, Johannes S.

doi:10.1016/j.watres.2014.06.020

Cited by 114 publications

(98 citation statements)

References 43 publications

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“…Similar observations were reported in the literature [23]. However, Proteobacteria are among the most abundant phyla of tap water microbiome in many studies [21][22][23][24].…”

Section: Biodiversity Of Bacteria Forming Biofilm In Cold-and Hot-tapsupporting

confidence: 89%

Application of NGS for studying the antibiotic resistant microbiome in Wrocław tap water

Siedlecka

Piekarska

2018

E3S Web Conf.

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Abstract. The NGS method was used to investigate the biodiversity of ARB dwelling in tap water, able to form biofilm on bullion media supplemented with antibiotics: β-lactam, fluoroquinolone, 3 rd generation cephalosporin and tetracycline. Biofilms formed by cold-and hot-tap water microorganisms were also examined. The presence of 10 ARGs in plasmid DNA of cultivated biomass was tested using a standard PCR. The analyses revealed that the most predominant families were Sphingomonadaceae, Paenibacillaceae, Enterobacteriaceae, Bacillaceae and Sphingomonadaceae in FQ, C3G, T, C-W and H-W samples, respectively (explanations of abbreviations in the text). The ARGs did not correlate with resistance phenotypes.

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“…Similar observations were reported in the literature [23]. However, Proteobacteria are among the most abundant phyla of tap water microbiome in many studies [21][22][23][24].…”

Section: Biodiversity Of Bacteria Forming Biofilm In Cold-and Hot-tapsupporting

confidence: 89%

Application of NGS for studying the antibiotic resistant microbiome in Wrocław tap water

Siedlecka

Piekarska

2018

E3S Web Conf.

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“…FCM has become the method of choice for quantifying microbial cells in aquatic samples, combining high sample throughput with speed and accuracy Van Nevel et al, 2017). Moreover, FCM has proved to be a useful tool not only for assessing cell abundance and viability but also for microbial community profiling Prest et al, 2014Prest et al, , 2013Ramseier et al, 2011).…”

Section: Introductionmentioning

confidence: 99%

Flow-cytometric quantification of microbial cells on sand from water biofilters

et al. 2018

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a b s t r a c tRapid quantification of absolute microbial cell abundances is important for a comprehensive interpretation of microbiome surveys and crucial to support theoretical modelling and the design of engineered systems. In this paper, we propose a protocol specifically optimised for the quantification of microbial abundances in water biofilters using flow cytometry (FCM). We optimised cell detachment from sand biofilter particles for FCM quantification through the evaluation of five chemical dispersants (NaCl, Triton-X100, CaCl 2 , sodium pyrophosphate (PP), Tween 80 combined with PP), different mechanical pretreatments (low and high energy sonication and shaking) and two fixation methods (glutaraldehyde and ethanol). The developed protocol was cross-compared using other established and commonly employed methods for biomass quantification in water filter samples (adenosine triphosphate (ATP) quantification, real-time quantitative PCR (qPCR) and volatile solids (VS)). The highest microbial count was obtained by detaching the biofilm from biofilter grains and dispersing clusters into singles cells using Tween 80 and sodium pyrophosphate combined with four steps of high energy sonication (27W, for 80 s each step); glutaraldehyde was shown to be the best fixative solution. The developed protocol was reliable and highly reproducible and produced results that are comparable to data from alternative quantification methods. Indeed, high correlations were found with trends obtained through ATP and qPCR (r ¼ 0.98 and r ¼ 0.91) measurements. The VS content was confirmed as an inaccurate method to express biomass in sand samples since it correlated poorly with all the other three methods (r ¼ 0.005 with FCM, 0.002 with ATP and 0.177 with qPCR). FCM and ATP showed the strongest agreement between absolute counts with a slope of the correlation equal to 0.7, while qPCR seemed to overestimate cell counts by a factor of ten. The rapidity and reproducibility of the method developed make its application ideal for routine quantification of microbial cell abundances on sand from water biofilters and thus useful in revealing the ecological patterns and quantifying the metabolic kinetics involved in such systems.

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“…The complex interactions between environmental factors from inside and outside the cave must be taken into account to better understand the biological processes. Finally, as reported by several authors (Krstulovic et al 2013;Prest et al 2014), combining flow cytometry with molecular analyses and nextgeneration sequencing (NGS) is an appealing approach in identifying heterotrophic and phototrophic communities proliferating in different environments.…”

Section: Resultsmentioning

confidence: 93%

“…FCM is a well-known method for the analysis of microbial communities in both freshwater and marine ecosystems (Lebaron et al 2002;Estrada et al 2004;Fernández et al 2008;Zhao et al 2012;Rekik et al 2013;Prest et al 2014;Dhib et al 2016) but is hardly ever observed in the case of microorganisms colonizing terrestrial environments, especially subterranean ones such as caves. The aim of this study was to provide relevant information on microbial communities in caves, specifically provided by the use of FCM, taking into account relationships with environmental factors as well as tourism management practices in this particular habitat.…”

Section: Introductionmentioning

confidence: 99%

Microbial composition and ecological features of phototrophic biofilms proliferating in the Moidons Caves (France): investigation at the single-cell level

Borderie

Denis

Barani

et al. 2016

Environ Sci Pollut Res

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The authors investigated the microbial composition of phototrophic biofilms proliferating in a show cave using flow cytometry for the first time in such a context. Results are based on several biofilms sampled in the Moidons Caves (France) and concern both heterotrophic prokaryotes and autotrophic microorganisms. Heterotrophic microorganisms with low nucleic acid content were dominant in biofilms, as can be expected from the oligotrophic conditions prevailing within the cave. Analysis of the biofilm autotrophic components revealed the presence of several taxa, particularly the unicellular green algae Chlorella minutissima, specifically well adapted to this cave. Relationships between flow cytometry results and environmental variables determined in the cave were established and discussed so as to better understand biofilm proliferation processes in caves.

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Combining flow cytometry and 16S rRNA gene pyrosequencing: A promising approach for drinking water monitoring and characterization

Cited by 114 publications

References 43 publications

Application of NGS for studying the antibiotic resistant microbiome in Wrocław tap water

Application of NGS for studying the antibiotic resistant microbiome in Wrocław tap water

Flow-cytometric quantification of microbial cells on sand from water biofilters

Microbial composition and ecological features of phototrophic biofilms proliferating in the Moidons Caves (France): investigation at the single-cell level

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