0000000000319647
AUTHOR
Enric Cuevas-ferrando
Bias of library preparation for virome characterization in untreated and treated wastewaters
The use of metagenomics for virome characterization and its implementation for wastewater analyses, including wastewater-based epidemiology, has increased in the last years. However, the lack of standardized methods can led to highly different results. The aim of this work was to analyze virome profiles in upstream and downstream wastewater samples collected from four wastewater treatment plants (WWTPs) using two different library preparation kits. Viral particles were enriched from wastewater concentrates using a filtration and nuclease digestion procedure prior to total nucleic acid (NA) extraction. Sequencing was performed using the ScriptSeq v2 RNA-Seq (LS) and the NEBNext Ultra II RNA …
HEV Occurrence in Waste and Drinking Water Treatment Plants
Hepatitis E virus (HEV), particularly zoonotic genotype 3, is present in environmental waters worldwide, especially in industrialized countries. Thus, monitoring the presence of HEV in wastewater treatment plants (WWTPs) is an emerging topic due to the importance of reusing water on a global level. Given the limited data, this study aimed to monitor the occurrence of HEV in influent and effluent water in waste- and drinking-water treatment plants (WWTPs and DWTPs). To this end, different procedures to concentrate HEV in influent and effluent water from WWTPs and DWTPs were initially evaluated. The evaluated procedures resulted in average HEV recoveries of 15.2, 19.9, and 16.9% in influent, …
Detection Of Genomic Variants Of SARS-CoV-2 Circulating In Wastewater By High-Throughput Sequencing
The use of SARS-CoV-2 metagenomics in wastewater can allow the detection of variants circulating at community level. After comparing with clinical databases, we identified three novel variants in the spike gene, and six new variants in the spike detected for the first time in Spain. We finally support the hypothesis that this approach allows the identification of unknown SARS-CoV-2 variants or detected at only low frequencies in clinical genomes.
SARS-CoV-2 RNA titers in wastewater anticipated COVID-19 occurrence in a low prevalence area
Postprint disponible en http://hdl.handle.net/10261/211707
Evaluation of two different concentration methods for surveillance of human viruses in sewage and their effects on SARS-CoV-2 sequencing
8 páginas, 5 figuras, 1 tabla
Metropolitan wastewater analysis for COVID-19 epidemiological surveillance
The COVID-19 disease, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is a rapidly emerging pandemic which has enforced extreme containment measures worldwide. In the absence of a vaccine or efficient treatment, cost-effective epidemiological surveillance strategies are urgently needed. Here, we have used RT-qPCR for SARS-CoV-2 detection in a series of longitudinal metropolitan wastewaters samples collected from February to April 2020, during the earliest stages of the epidemic in the Region of Valencia, Spain. We were able to consistently detect SARS-CoV-2 RNA in samples taken in late February, when communicated cases in that region were only incipient. We also find…
SARS-CoV-2 RNA in wastewater anticipated COVID-19 occurrence in a low prevalence area
Preprint disponible en http://hdl.handle.net/10261/209125
Monitoring Human Viral Pathogens Reveals Potential Hazard for Treated Wastewater Discharge or Reuse
Wastewater discharge to the environment or its reuse after sanitization poses a concern for public health given the risk of transmission of human viral diseases. However, estimating the viral infectivity along the wastewater cycle presents technical challenges and still remains underexplored. Recently, human-associated crAssphage has been investigated to serve as viral pathogen indicator to monitor fecal impacted water bodies, even though its assessment as biomarker for infectious enteric viruses has not been explored yet. To this end, the occurrence of potentially infectious norovirus genogroup I (GI), norovirus GII, hepatitis A virus (HAV), rotavirus A (RV), and human astrovirus (HAstV) a…
Comparing analytical methods to detect SARS-CoV-2 in wastewater
Wastewater based epidemiology (WBE) has emerged as a reliable strategy to assess the coronavirus disease 2019 (COVID-19) pandemic. Recent publications suggest that SARS-CoV-2 detection in wastewater is technically feasible; however, many different protocols are available and most of the methods applied have not been properly validated. To this end, different procedures to concentrate and extract inactivated SARS-CoV-2 and surrogates were initially evaluated. Urban wastewater seeded with gamma-irradiated SARS-CoV-2, porcine epidemic diarrhea virus (PEDV), and mengovirus (MgV) was used to test the concentration efficiency of an aluminum-based adsorption-precipitation method and a polyethylene…
Platinum chloride-based viability RT-qPCR for SARS-CoV-2 detection in complex samples.
Isolation, contact tracing and restrictions on social movement are being globally implemented to prevent and control onward spread of SARS-CoV-2, even though the infection risk modelled on RNA detection by RT-qPCR remains biased as viral shedding and infectivity are not discerned. Thus, we aimed to develop a rapid viability RT-qPCR procedure to infer SARS-CoV-2 infectivity in clinical specimens and environmental samples. We screened monoazide dyes and platinum compounds as viability molecular markers on five SARS-CoV-2 RNA targets. A platinum chloride-based viability RT-qPCR was then optimized using genomic RNA, and inactivated SARS-CoV-2 particles inoculated in buffer, stool, and urine. Ou…
Viability RT-PCR for SARS-CoV-2: a step forward to solve the infectivity quandary
Background Isolation, contact tracing and restrictions on social movement are being globally implemented to prevent and control onward spread of SARS-CoV-2, even though the infection risk modelled on RNA detection by RT-qPCR remains biased as viral shedding and infectivity are not discerned. Thus, we aimed to develop a rapid viability RT-qPCR procedure to infer SARS-CoV-2 infectivity in clinical specimens and environmental samples. Methods We screened monoazide dyes and platinum compounds as viability molecular markers on five SARS-CoV-2 RNA targets. A platinum chloride-based viability RT-qPCR was then optimized using genomic RNA, and inactivated SARS-CoV-2 particles inoculated in buffer, s…
Spatial and temporal distribution of SARS-CoV-2 diversity circulating in wastewater
Wastewater-based epidemiology (WBE) has proven to be an effective tool for epidemiological surveillance of SARS-CoV-2 during the current COVID-19 pandemic. Furthermore, combining WBE together with high-throughput sequencing techniques can be useful for the analysis of SARS-CoV-2 viral diversity present in a given sample. The present study focuses on the genomic analysis of SARS-CoV-2 in 76 sewage samples collected during the three epidemiological waves that occurred in Spain from 14 wastewater treatment plants distributed throughout the country. The results obtained demonstrate that the metagenomic analysis of SARS-CoV-2 in wastewater allows the detection of mutations that define the B.1.1.…
Assessment of ISO Method 15216 to Quantify Hepatitis E Virus in Bottled Water
Hepatitis E virus (HEV) is one of the causative agents of water-borne human viral hepatitis and considered in Europe an emerging zoonotic pathogen. Analysis of bottled water through a standard method validated for HEV can contribute towards the risk management of this hazard. Putting some recent reports by the European Food Safety Authority in place, this study aimed to assess the performance of the concentration and extraction procedures described in ISO 15216-1:2017 for norovirus and hepatitis A virus on HEV detection. Following the ISO recommendation, the bottled water samples were spiked using serially diluted HEV fecal suspensions together with mengovirus as process control and concent…
Recovering coronavirus from large volumes of water
The need for monitoring tools to better control the ongoing coronavirus disease (COVID-19) pandemic is extremely urgent and the contamination of water resources by excreted viral particles poses alarming questions to be answered. As a first step to overcome technical limitations in monitoring SARS-CoV-2 along the water cycle, we assessed the analytical performance of a dead end hollow fiber ultrafiltration coupled to different options for secondary concentrations to concentrate viral particles from large volume of spiked tap water, seawater and surface water together with two quantitative RT-qPCR detection kits. Spiking the porcine epidemic diarrhea virus (PEDV), an enveloped virus surrogat…
Discrimination of non-infectious SARS-CoV-2 particles from fomites by viability RT-qPCR.
The ongoing coronavirus 2019 (COVID-19) pandemic constitutes a concerning global threat to public health and economy. In the midst of this pandemic scenario, the role of environment-to-human COVID-19 spread is still a matter of debate because mixed results have been reported concerning SARS-CoV-2 stability on high-touch surfaces in real-life scenarios. Up to now, no alternative and accessible procedures for cell culture have been applied to evaluate SARS-CoV-2 infectivity on fomites. Several strategies based on viral capsid integrity have latterly been developed using viability markers to selectively remove false-positive qPCR signals resulting from free nucleic acids and damaged viruses. T…