Monitoring Waterborne Pathogens in Surface and Drinking Waters. Are Water Treatment Plants (WTPs) Simultaneously Efficient in the Elimination of Enteric Viruses and Fecal Indicator Bacteria (FIB)?

Abstract

Monitoring the quality of water is a requisite to prevent outbreaks related to waterborne diseases, predominantly caused by pathogens like enteric viruses, usually transmitted via the fecal-oral route. This study aimed to survey a group of enteric viruses (Enterovirus, Norovirus genogroups I and II, and hepatitis A virus) in two surface water sources of drinking water, also intending to evaluate the extent of their elimination in the two water treatment plants (WTPs) involved in drinking water production. Correlations between these viruses and fecal indicator bacteria (FIB) were also evaluated. Positive samples for viral RNA were recurrently found by reverse transcription quantitative PCR (RT-qPCR) and quantified, in genomic copies per liter (gc/L) of sampled water. Viral RNAs were detected in 14 out of 27 samples of surface water, and 21 out of 36 samples of drinking water, NoV II having been the most frequently detected in both (0–78.6 gc/L and 0–12.5 gc/L, respectively). Both WTPs showed variable efficacies in the elimination of viral RNA. Only one correlation was found with FIB, between NoV II and intestinal enterococci. These results recommend the monitoring of enteric viruses over time and their inclusion in the mandatory analysis of water quality.

Mengovirus as Process Control Virus in the Monitoring of Genomic RNA and Infectivity of Enteric Viruses in Water Matrices

“…Although a process control virus is highly recommended for the validation and interpretation of water monitoring processes [10,18,30], the low RNA recovery rates obtained for surface water in the present study recommend considering the addition of mengovirus only at the stages following the primary concentration step, where it will still be very useful, namely as control of RNA extraction (as initially proposed in the PCT/EP2007/055407 patent [17]). Furthermore, knowing that mengovirus induces cytopathic effects on Vero E6 cells (this study) and HeLa cells [24,28], its addition at the beginning of the process can make it difficult to interpret assays aiming the evaluation of the infectivity potential (for other virus) of the samples under survey, since the appearance of CPE may be exclusively due to the presence of mengovirus.…”

“…The significant differences between the recovery rates in the two water matrices, found in the present study (confirmed by the application of the Student's t-test), may be due to their very diverse characteristics. The surface water, having not undergone any treatment, contains a higher number of suspended particles and other agents that will be concentrated in the filter, while the drinking water was previously treated by processes that allowed the removal of suspended particles [10]. Consequently, the conditions for mengovirus adhesion to the filter will be different in the two situations, less efficient when surface water is collected.…”

“…A subsample of 20 mL reserved for RNA extraction undergone a tertiary concentration in Vivaspin ® concentrators (Sartorius; Goettingen, Germany) by centrifugation at 8000× g and 4 • C for 6 h or until a volume of about 1 mL was achieved. RNA was then extracted from this concentrated [10,23].…”

“…RNA extractions from the concentrated water samples (Section 2.1) were performed with the QIAamp viral RNA Mini kit (Qiagen; Hilden, Germany), following the manufacturer's instructions. RT-qPCR reactions targeting the detection and quantification of mengovirus genomic RNA were performed on a StepOnePlus thermocycler (Applied Biosystems; Foster City, CA, USA), according to the conditions described in Salvador et al [10,23]. In short, Mengovirus Extraction Control kit (bioMérieux; Marcyl'Etoile, France), was used in 25 µL reaction mixtures consisting of 20 µL of MasterMix and 5 µL of extracted RNA.…”

“…The methodologies for the monitoring of these viruses are complex, in most cases with low recovery rates [9][10][11]. Two of the most commonly applied methodologies are the Method 1615 of the United States Environmental Protection Agency (EPA) [12] and the European methodology ISO 15216-1: 2017 [13].…”