Rapid monitoring of Escherichia coli and Enterococcus spp. in bathing water using Reverse Transcription-quantitative PCR

Bergeron, Peggy; Oujati, H.; Cuenca, Vicente Catalán; Mestre, J.M. Huguet; Courtois, Sophie

doi:10.1016/j.ijheh.2011.07.013

Cited by 19 publications

(12 citation statements)

References 31 publications

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“…Studies of qPCR methods for water testing have advanced from laboratory analysis of frozen filters [2, 11, 14, 26–28] and data analyses using correlations or linear regression methods. Some of those studies [11, 14, 29–32] as well as newer investigations [13, 16, 17] evaluated associations between the exceedance of threshold values for culture and qPCR methods. However, because these studies generally did not include a time series of daily measurements, comparisons were limited to Day0 culture threshold exceedance and Day0 qPCR threshold exceedance.…”

Section: Discussionmentioning

confidence: 99%

Monitoring urban beaches with qPCR vs. culture measures of fecal indicator bacteria: Implications for public notification

et al. 2017

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BackgroundThe United States Environmental Protection Agency has established methods for testing beach water using the rapid quantitative polymerase chain reaction (qPCR) method, as well as “beach action values” so that the results of such testing can be used to make same-day beach management decisions. Despite its numerous advantages over culture-based monitoring approaches, qPCR monitoring has yet to become widely used in the US or elsewhere. Considering qPCR results obtained on a given day as the best available measure of that day’s water quality, we evaluated the frequency of correct vs. incorrect beach management decisions that are driven by culture testing.MethodsBeaches in Chicago, USA, were monitored using E. coli culture and enterococci qPCR methods over 894 beach-days in the summers of 2015 and 2016. Agreement in beach management using the two methods, after taking into account agreement due to chance, was summarized using Cohen’s kappa statistic.ResultsNo meaningful agreement (beyond that expected by chance) was observed between beach management actions driven by the two pieces of information available to beach managers on a given day: enterococci qPCR results ofsamples collected that morning and E. coli culture results of samples collected the previous day. The E. coli culture beach action value was exceeded 3.4 times more frequently than the enterococci qPCR beach action value (22.6 vs. 6.6% of beach-days).ConclusionsThe largest evaluation of qPCR-based beach monitoring to date provides little scientific rationale for continued E. coli culture testing of beach water in our setting. The observation that the E. coli culture beach action value was exceeded three times as frequently as the enterococci qPCR beach action value suggests that, although the beach action values for bacteria using different measurement methods are thought to provide comparable information about health risk, this does not appear to be the case in all settings.

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Section: Discussionmentioning

confidence: 99%

Monitoring urban beaches with qPCR vs. culture measures of fecal indicator bacteria: Implications for public notification

et al. 2017

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“…The protocol we employed in our study differs from most studies that have used rRNA-based RT-qPCR to detect fecal bacteria. For example, some studies only used rRNA and therefore did not compare the detection frequency of fecal bacteria using rDNA and rRNA. , Second, in many studies RNA and DNA extractions have been performed using different subsamples. ,, Simultaneous RNA and DNA extraction enables more accurate comparison as it eliminates the intrinsic variability between subsamples. The use of the same sample for both RNA and DNA extraction reduces costs, saves storage space and shortens the extraction time (i.e., one lysis step per sample).…”

Section: Discussionmentioning

confidence: 99%

Detection of Fecal Bacteria and Source Tracking Identifiers in Environmental Waters Using rRNA-Based RT-qPCR and rDNA-Based qPCR Assays

Pitkänen

Ryu

Elk

et al. 2013

Environ. Sci. Technol.

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In this study, we evaluated the use of RT-qPCR assays targeting rRNA gene sequences for the detection of fecal bacteria in water samples. We challenged the RT-qPCR assays against RNA extracted from sewage effluent (n = 14), surface water (n = 30), and treated source water (n = 15) samples. Additionally, we applied the same assays using DNA as the qPCR template. The targeted fecal bacteria were present in most of the samples tested, although in several cases, the detection frequency increased when RNA was used as the template. For example, the majority of samples that tested positive for E. coli and Campylobacter spp. in surface waters, and for human-specific Bacteroidales, E. coli, and Enterococcus spp. in treated source waters were only detected when rRNA was used as the original template. The difference in detection frequency using rRNA or rDNA (rRNA gene) was sample- and assay-dependent, suggesting that the abundance of active and nonactive populations differed between samples. Statistical analyses for each population exhibiting multiple quantifiable results showed that the rRNA copy numbers were significantly higher than the rDNA counterparts (p < 0.05). Moreover, the detection frequency of rRNA-based assays were in better agreement with the culture-based results of E. coli, intestinal enterococci, and thermotolerant Campylobacter spp. in surface waters than that of rDNA-based assays, suggesting that rRNA signals were associated to active bacterial populations. Our data show that using rRNA-based approaches significantly increases detection sensitivity for common fecal bacteria in environmental waters. These findings have important implications for microbial water quality monitoring and public health risk assessments.

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“…Several studies have addressed the dynamics and abundance of enterococci in seawater and sediment using different approaches, including MF and molecular assays based on qPCR [29], [35] and RT-qPCR [8], [36]. However, little is known of the impact of fish farms on the origin and spread of antibiotic-resistant strains and related AR genes [4], [23].…”

Section: Discussionmentioning

confidence: 99%

Aquaculture Can Promote the Presence and Spread of Antibiotic-Resistant Enterococci in Marine Sediments

et al. 2013

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Aquaculture is an expanding activity worldwide. However its rapid growth can affect the aquatic environment through release of large amounts of chemicals, including antibiotics. Moreover, the presence of organic matter and bacteria of different origin can favor gene transfer and recombination. Whereas the consequences of such activities on environmental microbiota are well explored, little is known of their effects on allochthonous and potentially pathogenic bacteria, such as enterococci. Sediments from three sampling stations (two inside and one outside) collected in a fish farm in the Adriatic Sea were examined for enterococcal abundance and antibiotic resistance traits using the membrane filter technique and an improved quantitative PCR. Strains were tested for susceptibility to tetracycline, erythromycin, ampicillin and gentamicin; samples were directly screened for selected tetracycline [tet(M), tet(L), tet(O)] and macrolide [erm(A), erm(B) and mef] resistance genes by newly-developed multiplex PCRs. The abundance of benthic enterococci was higher inside than outside the farm. All isolates were susceptible to the four antimicrobials tested, although direct PCR evidenced tet(M) and tet(L) in sediment samples from all stations. Direct multiplex PCR of sediment samples cultured in rich broth supplemented with antibiotic (tetracycline, erythromycin, ampicillin or gentamicin) highlighted changes in resistance gene profiles, with amplification of previously undetected tet(O), erm(B) and mef genes and an increase in benthic enterococcal abundance after incubation in the presence of ampicillin and gentamicin. Despite being limited to a single farm, these data indicate that aquaculture may influence the abundance and spread of benthic enterococci and that farm sediments can be reservoirs of dormant antibiotic-resistant bacteria, including enterococci, which can rapidly revive in presence of new inputs of organic matter. This reservoir may constitute an underestimated health risk and deserves further investigation.

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Rapid monitoring of Escherichia coli and Enterococcus spp. in bathing water using Reverse Transcription-quantitative PCR

Cited by 19 publications

References 31 publications

Monitoring urban beaches with qPCR vs. culture measures of fecal indicator bacteria: Implications for public notification

Monitoring urban beaches with qPCR vs. culture measures of fecal indicator bacteria: Implications for public notification

Detection of Fecal Bacteria and Source Tracking Identifiers in Environmental Waters Using rRNA-Based RT-qPCR and rDNA-Based qPCR Assays

Aquaculture Can Promote the Presence and Spread of Antibiotic-Resistant Enterococci in Marine Sediments

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