This study was conducted to evaluate the applicability of crAssphage, pepper mild mottle virus (PMMoV), and tobacco mosaic virus (TMV) as indicators of the reduction of human enteric viruses during ...wastewater treatment. Thirty-nine samples were collected from three steps at a wastewater treatment plant (raw sewage, secondary-treated sewage, and final effluent) monthly for a 13-month period. In addition to the three indicator viruses, eight human enteric viruses human adenoviruses, JC and BK polyomaviruses, Aichi virus 1 (AiV-1), enteroviruses, and noroviruses of genogroups I, II, and IV were tested by quantitative PCR. Indicator viruses were consistently detected in the tested samples, except for a few final effluents for crAssphage and TMV. The mean concentrations of crAssphage were significantly higher than those of most tested viruses. The concentrations of crAssphage in raw sewage were positively correlated with the concentrations of all tested human enteric viruses (p <0.05), suggesting the applicability of crAssphage as a suitable indicator to estimate the concentrations of human enteric viruses in raw sewage. The reduction ratios of AiV-1 (1.8 ± 0.7 log
) were the lowest among the tested viruses, followed by TMV (2.0 ± 0.3 log
) and PMMoV (2.0 ± 0.4 log
). Our findings suggested that the use of not only AiV-1 and PMMoV but also TMV as indicators of reductions in viral levels can be applicable during wastewater treatment.
We investigated the presence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA in wastewater samples in southern Louisiana, USA. Untreated and treated wastewater samples were ...collected on five occasions over a four-month period from January to April 2020. The wastewater samples were concentrated via ultrafiltration (Method A), and an adsorption–elution method using electronegative membranes (Method B). SARS-CoV-2 RNA was detected in 2 out of 15 wastewater samples using two reverse transcription-quantitative polymerase chain reaction (RT-qPCR) assays (CDC N1 and N2). None of the secondary treated and final effluent samples tested positive for SARS-CoV-2 RNA. To our knowledge, this is the first study reporting the detection of SARS-CoV-2 RNA in wastewater in North America, including the USA. However, concentration methods and RT-qPCR assays need to be refined and validated to increase the sensitivity of SARS-CoV-2 RNA detection in wastewater.
Display omitted
•First study in Louisiana, USA reporting the detection of SARS-CoV-2 RNA in wastewater using ultrafiltration.•Two out of seven untreated wastewater samples tested positive for SARS-CoV-2 RNA.•None of the secondary treated and final effluent samples tested positive.•Concentration methods and RT-qPCR assays applied for SARS-CoV-2 RNA detection need further refinement.
The applicability of wastewater-based epidemiology (WBE) has been extensively studied throughout the world with remarkable findings. This study reports the presence and reduction of severe acute ...respiratory syndrome coronavirus 2 (SARS-CoV-2) at two wastewater treatment plants (WWTPs) of Nepal, along with river water, hospital wastewater (HWW), and wastewater from sewer lines collected between July 2020 and February 2021. SARS-CoV-2 RNA was detected in 50%, 54%, 100%, and 100% of water samples from WWTPs, river hospitals, and sewer lines, respectively, by at least one of four quantitative PCR assays tested (CDC-N1, CDC-N2, NIID_2019-nCOV_N, and N_Sarbeco). The CDC-N2 assay detected SARS-CoV-2 RNA in the highest number of raw influent samples of both WWTPs. The highest concentration was observed for an influent sample of WWTP A (5.5 ± 1.0 log10 genome copies/L) by the N_Sarbeco assay. SARS-CoV-2 was detected in 47% (16/34) of the total treated effluents of WWTPs, indicating that biological treatments installed at the tested WWTPs are not enough to eliminate SARS-CoV-2 RNA. One influent sample was positive for N501Y mutation using the mutation-specific qPCR, highlighting a need for further typing of water samples to detect Variants of Concern. Furthermore, crAssphage-normalized SARS-CoV-2 RNA concentrations in raw wastewater did not show any significant association with the number of new coronavirus disease 2019 (COVID-19) cases in the whole district where the WWTPs were located, suggesting a need for further studies focusing on suitability of viral as well as biochemical markers as a population normalizing factor. Detection of SARS-CoV-2 RNA before, after, and during the peaking in number of COVID-19 cases suggests that WBE is a useful tool for COVID-19 case estimation in developing countries.
Display omitted
•First case of detection of SARS-CoV-2 RNA in wastewater and river water in Nepal•SARS-CoV-2 RNA was detected using qPCR in 60% (50/84) of water samples tested.•CDC-N2 assay showed the highest positive ratios among four qPCR assays tested.•Reduction ratios of SARS-CoV-2 RNA at WWTP were higher than those of crAssphage.•N501Y mutation was detected in an influent of WWTP collected in February 2021.
The composition of the gut microbiome in industrialized populations differs from those living traditional lifestyles. However, it has been difficult to separate the contributions of human genetic and ...geographic factors from lifestyle. Whether shifts away from the foraging lifestyle that characterize much of humanity's past influence the gut microbiome, and to what degree, remains unclear. Here, we characterize the stool bacterial composition of four Himalayan populations to investigate how the gut community changes in response to shifts in traditional human lifestyles. These groups led seminomadic hunting-gathering lifestyles until transitioning to varying levels of agricultural dependence upon farming. The Tharu began farming 250-300 years ago, the Raute and Raji transitioned 30-40 years ago, and the Chepang retain many aspects of a foraging lifestyle. We assess the contributions of dietary and environmental factors on their gut-associated microbes and find that differences in the lifestyles of Himalayan foragers and farmers are strongly correlated with microbial community variation. Furthermore, the gut microbiomes of all four traditional Himalayan populations are distinct from that of the Americans, indicating that industrialization may further exacerbate differences in the gut community. The Chepang foragers harbor an elevated abundance of taxa associated with foragers around the world. Conversely, the gut microbiomes of the populations that have transitioned to farming are more similar to those of Americans, with agricultural dependence and several associated lifestyle and environmental factors correlating with the extent of microbiome divergence from the foraging population. The gut microbiomes of Raute and Raji reveal an intermediate state between the Chepang and Tharu, indicating that divergence from a stereotypical foraging microbiome can occur within a single generation. Our results also show that environmental factors such as drinking water source and solid cooking fuel are significantly associated with the gut microbiome. Despite the pronounced differences in gut bacterial composition across populations, we found little differences in alpha diversity across lifestyles. These findings in genetically similar populations living in the same geographical region establish the key role of lifestyle in determining human gut microbiome composition and point to the next challenging steps of determining how large-scale gut microbiome reconfiguration impacts human biology.
Enteric viruses in the aquatic environment are a concern due to the potential for waterborne disease transmission to humans. In Nepal, the Bagmati River serves as a source of drinking and irrigation ...water; therefore, the detection of waterborne enteric pathogens is integral to maintaining human health. The objective of this study was to quantify the crAssphage marker in surface water samples from the Bagmati River between November 2015 and September 2016. Concentrations of crAssphage were then compared to those of other enteric viruses and indicator organisms found in the samples in order to examine the potential of crAssphage as a marker for fecal contamination. CrAssphage was detected in 17% (1/6) of samples from Sundarijal, 100% (6/6) of samples from Thapathali, and 100% (6/6) samples from Chovar, with the highest average concentrations recorded in May 2016 and the lowest average concentrations recorded in September 2016. Overall, crAssphage was present in 72% (13/18) of samples and was strongly correlated with the presence of fecal indicator bacteria
Escherichia coli
(
r
= 0.89) and Enterococcus (
r
= 0.92) and several enteric viruses. The strongest viral correlations were to salivirus (
r
= 0.84), pepper mild mottle virus (
r
= 0.77), Aichi virus 1 (
r
= 0.75), enteroviruses (
r
= 0.76), and tobacco mosaic virus (
r
= 0.71). These results provide evidence for the potential use of crAssphage as a marker for human fecal contamination in river water.
Shiga toxin-producing
Escherichia coli
(STEC) is one of the most commonly heard sources of foodborne outbreaks but presently lacks studies of in the Kathmandu Valley. This study explored the presence ...of STEC in river water (
n
= 17), groundwater (
n
= 83), feces (
n
= 50), and manure (
n
= 20) in the Kathmandu Valley, Nepal. Samples that were confirmed to be
E. coli
–positive using the Colilert assay in previous studies were selected to examine the presence of the
sfmD
,
stx1
, and
stx2
genes using quantitative polymerase chain reaction. Based on Colilert/
sfmD
gene ratios, the percentage of viable cells of
E. coli
in the fecal (2.0% ± 2.5%), manure (0.1% ± 0.1%), river water (3.4% ± 3.1%), and groundwater samples (3.9% ± 5.8%) were determined. The presence of the
stx1
gene was observed in the fecal (6%), river water (53%), and groundwater (2%) samples, whereas the
stx2
gene was detected in the fecal (8%), manure (5%), and river water (71%) samples. Interestingly, the
stx
/
sfmD
gene ratios in the groundwater samples were greater than 100%, suggesting the presence of other
stx1/stx2
-harboring microorganisms. These findings indicated the importance of continuing investigation into waterborne sources of STEC contamination within the Kathmandu Valley.
The use of recycled or reclaimed water for direct or indirect potable uses is increasing because of water scarcity. Wastewater contains pathogens that are capable of causing infections in humans. ...Among the microbial pathogens, viruses pose significant human health risks because of their high environmental persistence and low infectious doses. Routine monitoring of pathogenic viruses is challenging, time-consuming, and expensive. Moreover, the concentrations of most pathogenic viruses are not stable in wastewater throughout the year. As an alternative, viral indicators can be used to predict the presence, behavior, and removal of viral pathogens through wastewater treatment processes. This article highlights the characteristics of various viral indicators, and their usefulness as a proxy for the log reduction of pathogenic viruses for recycled water scheme. Concentrations of indicator viruses, such as pepper mild mottle virus and emerging crAssphage, are 2–3 orders of magnitude greater in untreated sewage than those of bacteriophages and other viral indicators. However, more research is needed for their comparative log reduction with pathogenic viruses. Developing effective methods to determine virus concentrations, the ability to culture indicator viruses, and new detection tools will enhance the application of indicator viruses in water recycling.
This study assessed wastewater quality through the quantification of four human enteric viruses and the applicability of pepper mild mottle virus (PMMoV) and tobacco mosaic virus (TMV) as indicators ...of viral reduction during wastewater treatment. Thirty-three samples were collected from three steps of a wastewater treatment plant in Southern Louisiana, USA for a year between March 2017 and February 2018. Noroviruses of genogroup I were the most prevalent human enteric viruses in influent samples. The concentrations of PMMoV in influent samples (5.9 ± 0.7 log
10
copies/L) and biologically treated effluent samples (5.9 ± 0.5 log
10
copies/L) were significantly higher than those of TMV (
P
< 0.05), and the reduction ratio of PMMoV (1.0 ± 0.8 log
10
) was found comparable to those of TMV and Aichi virus 1. Because of the high prevalence, high correlations with human enteric viruses, and lower reduction ratios, PMMoV was deemed an appropriate indicator of human enteric viral reduction during wastewater treatment process.
Monitoring for SARS-CoV-2 RNA in wastewater through the process of wastewater-based epidemiology provides an additional surveillance tool, contributing to community-level screening and prevention ...efforts as these measurements have preceded disease cases in some instances. Numerous detections of SARS-CoV-2 RNA have been reported globally using various methods, demonstrating the technical feasibility of routine monitoring. However, to reliably interpret data produced from these efforts for informing public health interventions, additional quality control information and standardization in sampling design, sample processing, and data interpretation and reporting are needed. This review summarizes published studies of SARS-CoV-2 RNA detection in wastewater as well as available information regarding sample concentration, extraction, and detection methods. The review highlights areas for potential standardization including considerations related to sampling time and frequency relative to peak fecal loading times; inclusion of appropriate information on sample collection points; sample volume collected; transport and storage conditions; sample concentration procedures; RNA extraction process and performance; effective sample volumes; recovery efficiency testing; PCR inhibition; process controls throughout sample collection and processing; and PCR standard curve performance. Researchers are advised to follow the Minimum Information for Publication of Quantitative Real-Time PCR guidelines. Adhering to these recommendations will enable robust interpretation of wastewater monitoring results and improved inferences regarding the relationship between monitoring results and disease cases.
Monthly sampling was conducted at a drinking water treatment plant (DWTP) in Southern Louisiana, USA from March 2017 to February 2018 to determine the prevalence and reduction efficiency of ...pathogenic and indicator viruses. Water samples were collected from the DWTP at three different treatment stages (raw, secondary-treated, and chlorinated drinking water) and subjected to quantification of seven pathogenic viruses and three indicator viruses pepper mild mottle virus (PMMoV), tobacco mosaic virus (TMV), and crAssphage based on quantitative polymerase chain reaction. Among the seven pathogenic viruses tested, only Aichi virus 1 (AiV-1) (7/12, 58%) and noroviruses of genogroup II (NoVs-GII) (2/12, 17%) were detected in the raw water samples. CrAssphage had the highest positive ratio at 78% (28/36), and its concentrations were significantly higher than those of the other indicator viruses for all three water types (
P
< 0.05). The reduction ratios of AiV-1 (0.7 ± 0.5 log
10
;
n
= 7) during the whole treatment process were the lowest among the tested viruses, followed by crAssphage (1.1 ± 1.9 log
10
;
n
= 9), TMV (1.3 ± 0.9 log
10
;
n
= 8), PMMoV (1.7 ± 0.8 log
10
;
n
= 12), and NoVs-GII (3.1 ± 0.1 log
10
;
n
= 2). Considering the high abundance and relatively low reduction, crAssphage was judged to be an appropriate process indicator during drinking water treatment. To the best of our knowledge, this is the first study to assess the reduction of crAssphage and TMV during drinking water treatment.
PDF
