Same-day Enterococcus qPCR results of recreational water quality at two Toronto beaches provide added public health protection and reduced beach days lost.

OBJECTIVES: We evaluated the potential impacts from using a rapid same-day quantitative polymerase chain reaction (qPCR) monitoring method for beach posting outcomes at two Toronto beaches. METHODS: In total, 228 water samples were collected at Marie Curtis Park East and Sunnyside Beaches over the 2021 summer season. Water samples were processed using the USEPA 1609.1 Enterococcus qPCR-based method. Escherichia coli (E. coli) culture data and daily beach posting decisions were obtained from Toronto Public Health. RESULTS: No significant correlation was observed between previous-day and same-day (retrospective) E. coli enumeration results at any Sunnyside Beach transect, and only relatively low (R = 0.41-0.56) or no significant correlation was observed at sampling transects for Marie Curtis Park East Beach. Comparing our same-day Enterococcus qPCR data to Toronto's 2-day E. coli geometric mean beach posting decisions, we noted the need for additional postings for 1 (2%) and 3 (8%) missed health-risk days at Sunnyside and Marie Curtis Park East Beaches, respectively. The qPCR data also pointed to incorrect postings for 12 (31%) and 6 (16%) lost beach days at Sunnyside and Marie Curtis Park East Beaches, respectively. CONCLUSION: Application of a rapid Enterococcus qPCR method at two Toronto beaches revealed 5% of beach posting decisions were false negatives that missed health-risk days, while 23% of decisions were false positives resulting in lost beach days. Deployment of the rapid same-day qPCR method offers the potential to reduce both health risks and unnecessary beach postings.

RéSUMé: OBJECTIFS: Nous avons évalué, à deux plages de Toronto, l'effet possible de l'utilisation d'une méthode de surveillance rapide par PCR quantitative (qPCR) le même jour sur les avis de fermeture ou d'ouverture des plages. MéTHODE: En tout, 228 échantillons d'eau ont été prélevés aux plages Marie Curtis Park East et Sunnyside au cours de la saison estivale 2021. La présence d'Enterococcus dans les échantillons a été détectée par la méthode USEPA 1609.1, utilisant la qPCR. Les données sur les cultures d'Escherichia coli (E. coli) et les avis quotidiens de fermeture ou d'ouverture des plages ont été obtenus auprès du Bureau de santé de Toronto. RéSULTATS: Aucune corrélation significative n'a été observée entre les résultats (rétrospectifs) du dénombrement de E. coli obtenus la veille et le même jour dans les transects de la plage Sunnyside, et une corrélation significative faible (R = 0,41­0,56) ou nulle a été observée dans les transects d'échantillonnage de la plage Marie Curtis Park East. En comparant nos données sur Enterococcus obtenues le même jour par qPCR à la moyenne géométrique des avis de fermeture ou d'ouverture des plages sur deux jours liés à E. coli émis par le Bureau de santé de Toronto, nous avons remarqué qu'il aurait fallu émettre des avis de fermeture pour 1 jour de risques pour la santé manqué (2 %) à la plage Sunnyside et pour 3 jours de risques pour la santé manqués (8 %) à la plage Marie Curtis Park East. Les données de la qPCR ont aussi fait état d'avis de fermeture incorrects ayant entraîné la perte de 12 jours de plage (31 %) à Sunnyside et de 6 jours de plage (16 %) à Marie Curtis Park East. CONCLUSION: L'application d'une méthode de surveillance rapide d'Enterococcus par qPCR à deux plages de Toronto a montré que 5 % des avis étaient des faux négatifs qui n'ont pas détecté des jours de risques pour la santé, et que 23 % étaient des faux positifs qui ont entraîné des jours de plage perdus. Le déploiement de la méthode rapide par qPCR le même jour offre la possibilité de réduire à la fois les risques pour la santé et les avis de fermeture de plages inutiles.

Norovirus risk at a golf course irrigated with reclaimed water: Should QMRA doses be adjusted for infectiousness?

About 25 golf courses in Ontario, Canada have environmental compliance approvals to use reclaimed water for irrigation, where disinfection is confirmed through E. coli limits. A previous study at five Ontario municipal wastewater treatment plants (WWTPs) confirmed that enteric viruses are less susceptible to disinfection than E. coli, when plants provided conventional (secondary or tertiary) treatment and routine (chlorine or UV) disinfection. Here we query whether these four treatment-disinfection scenarios plus 60-day lagoon storage of disinfected effluent would be sufficient to reduce norovirus genogroups I and II (NoV GI and GII) risk of infection to tolerable levels for a golfer who incidentally ingests NoV after handling wet golf balls at a golf course irrigated with reclaimed water. We used our RT-qPCR NoV enumeration datasets from the four treatment-disinfection scenarios above and modeled detected and non-detected NoV by Bayesian inference in 'R'. Monte Carlo simulation included pre-disinfection NoV GI and GII gene copy densities; Ontario WWTP-derived chlorine and UV log10 reductions; literature-derived effluent storage decay parameters and golfer ingested volumes, followed by six different NoV dose-response (DR) models. Quantitative Microbial Risk Assessment (QMRA) results suggest that there is an unacceptable NoV infection risk when using the conservative assumption that all detected NoV particles (RT-qPCR gene copies) are infectious, in both aggregated or disaggregated form. However, after adjusting for PCR target sequences and for infectiousness using data from recently published studies on cultivation of human NoV in human intestinal enteroids, we noted a significant reduction of infection risk. However, this less conservative (i.e., less protective) assumption for water reuse applications such as golf course irrigation may not be corroborated until human NoV are efficiently and routinely grown in cell cultures. In addition, further studies on drivers of NoV risk estimation by DR models are needed, e.g., the extent of NoV particle aggregation resulting from wastewater treatment, as well as the role of immunity. Meantime, regulatory agencies could consider more stringent treatment-disinfection requirements that target enteric viruses rather than E. coli and testing of actual reclaimed irrigation waters.

Enteric viruses in municipal wastewater effluent before and after disinfection with chlorine and ultraviolet light.

In Ontario, Canada, information is lacking on chlorine and ultraviolet (UV) light disinfection performance against enteric viruses in wastewater. We enumerated enteroviruses and noroviruses, coliphages, and Escherichia coli per USEPA methods 1615, 1602, and membrane filtration, respectively, in pre- and post-disinfection effluent at five wastewater treatment plants (WWTPs), with full-year monthly sampling, and calculated log10 reductions (LRs) while WWTPs complied with their monthly geometric mean limit of 200 E. coli/100 mL. Modeling of densities by left-censored estimation and Bayesian inference gave very similar results. Polymerase chain reaction (PCR)-detected enteroviruses and noroviruses were abundant in post-disinfection effluent (mean concentrations of 2.1 × 10+4-7.2 × 10+5 and 2.7 × 10+4-3.6 × 10+5 gene copies (GC)/L, respectively). Chlorine or UV disinfection produced modest LRs for culture- (0.3-0.9) and PCR-detected enteroviruses (0.3-1.3), as well as noroviruses GI + GII (0.5-0.8). Coliphages and E. coli were more susceptible, with LRs of 0.8-3.0 and 2.5, respectively. Sand-filtered effluent produced significantly higher enteric virus LRs (except cultured enteroviruses). Coliphage and human enteric virus densities gave significantly positive correlations using Kendall's Tau test. Enteric viruses are abundant in wastewater effluent following routine chlorine or UV disinfection processes that target E. coli. Coliphages appear to be good indicators for evaluating wastewater disinfection of enteric viruses.

Pathogen Characterization of Fresh and Stored Mesophilic Anaerobically Digested Biosolids.

Culturable bacterial pathogens (Campylobacter, Salmonella, Listeria, Yersinia) and indicators (E. coli, enterococci, Clostridium perfringens) were quantified at six water resource recovery facilities that land apply anaerobically digested biosolids in Ontario, Canada. Cryptosporidium parvum and Giardia lamblia were also quantified by polymerase chain reaction (PCR). Salmonella and Listeria were frequently detected in sludge and liquid biosolids (70-100% of samples) but less often in fresh dewatered cake biosolids (50-60%); with low levels in fresh cake (<100 cells/g dw). Yersinia were in 20 to 30% of samples, typically at very low levels (<10 cell/g dw). Giardia and Cryptosporidium were detected in 80 and 20% of cake biosolids at geometric means of 270 cysts/g dw and 70 oocysts/g dw, respectively. E. coli reduction was typically >2-log10 while pathogen reduction was variable. "Sudden increase" of pathogens was not observed, however, Salmonella and E. coli showed regrowth (at 1 to 3 orders of magnitude) after 2- to 3-day storage at 30 °C.

Helicobacter pylori DNA in dental plaques, gastroscopy, and dental devices.

The role of dental plaque in the transmission of Helicobacter pylori (Hp) is unclear due to variability in the detection rates and techniques used. We used nested PCR to estimate the incidence of Hp in dental plaques of 24 dental hygienists. We found an unexpectedly high incidence (50%) of Hp DNA in dental plaques using sterilized dental probes. Additional treatment of sonication and SDS wash prior to sterilization of dental probes reduced the incidence to 13%. We used the treated probes to assess Hp presence in plaque samples of 47 patients visiting the dental clinic for teeth cleaning. Hp DNA was detected in 24% of cases. Since these data may reflect instrument contamination, we tested dental probes, endoscopes, and endoscopy forceps and found that 12.5-37.5% of them were contaminated. Consequently, dental plaques may be a candidate reservoir for Hp, medical equipment may contribute to Hp transmission, and sample collection techniques can bias the true prevalence of Hp in a population.