Pepper Mild Mottle Virus as Indicator of Pollution: Assessment of Prevalence and Concentration in Different Water Environments in Italy

Pepper mild mottle virus (PMMoV), a plant pathogenic virus belonging to the family Virgoviridae, has been proposed as a potential viral indicator for human faecal pollution in aquatic environments. The present study investigated the occurrence, amount and diversity of PMMoV in water environments in Italy. A total of 254 water samples, collected between 2017 and 2019 from different types of water, were analysed. In detail, 92 raw sewage, 32 treated sewage, 16 river samples, 9 estuarine waters, 20 bathing waters, 67 groundwater samples and 18 drinking waters were tested. PMMoV was detected in 79% and 75% of untreated and treated sewage samples, respectively, 75% of river samples, 67% and 25% of estuarine and bathing waters and 13% of groundwater samples. No positive was detected in drinking water. The geometric mean of viral concentrations (genome copies/L) was ranked as follows: raw sewage (2.2 × 10⁶) > treated sewage (2.9 × 10⁵) > river waters (6.1 × 10²) > estuarine waters (4.8 × 10²) > bathing waters (8.5 × 10¹) > groundwater (5.9 × 10¹). A statistically significant variation of viral loads could be observed between raw and treated sewage and between these and all the other water matrices. PMMoV occurrence and viral loads did not display seasonal variation in raw sewage nor correlation with faecal indicator bacteria in marine waters and groundwater. This study represents the first report on the occurrence and quantification PMMoV in different water environments in Italy. Further studies are required to evaluate the suitability of PMMoV as a viral indicator for human faecal pollution and for viral pathogens in waters.

     

Infectious Pepper Mild Mottle Virus and Human Adenoviruses as Viral Indices in Sewage and Water Samples

Food and Environmental Virology - The objective of this study was to compare human adenoviruses (HAdVs) genome and infectivity, polyomaviruses (JC and BK) genome (JCPyVs) and (BKPyVs), Pepper Mild...     

Prevalence and Genetic Characterization of Aichivirus in Environmental Waters in Thailand

Aichivirus 1 (AiV-1) is an enteric virus that has been documented to be the causative agent of diarrhea in humans. It is transmitted by fecal–oral route, through person-to-person contact, consumption of contaminated food or water, or recreation of contaminated water. AiV-1 is highly prevalent in water samples and has been proposed as a potential indicator of fecal contamination in water reservoirs. This study aimed to investigate the prevalence and genetic diversity of AiV-1 in environmental water samples in Thailand. A total of 126 samples were collected monthly from November 2016 to July 2018 from various sources of environmental water including irrigation water, reservoir, river, and wastewater. The presence of AiV-1 was detected by RT-nested PCR of the 3CD region and further analyzed by phylogenetic analysis. The AiV-1 was detected in 28 out of 126 (22.2%) of tested samples. A high frequency of AiV-1 detection was in wastewater (52.4%). All 28 AiV-1 strains detected in this study belonged to the genotype B and were closely related to AiV strains detected previously in environmental waters and in humans worldwide. This study demonstrated, for the first time, the contamination of AiV-1 in various sources of water samples in Thailand and provided a better insight into the prevalence of AiV-1 in environmental waters and its potential risk of human health.

     

Occurrence of <Emphasis Type="Italic">Pepper Mild Mottle Virus</Emphasis> (PMMoV) in Groundwater from a Karst Aquifer System in the Yucatan Peninsula,Mexico

The Yucatan Peninsula of Mexico hosts a karst aquifer system that is the only source of freshwater for the area; however, it is vulnerable to human-mediated contamination. Pepper mild mottle virus (PMMoV) is one of the most abundant RNA viruses associated with human feces, making it a viable indicator for tracking fecal pollution in aquatic environments, including groundwater. In this study, groundwater samples collected from a karst aquifer from fresh and brackish water locations were analyzed for fecal indicator bacteria, somatic and male F+ specific coliphages, and PMMoV during the rainy and dry seasons. Total coliform bacteria were detected at all sites, whereas Escherichia coli were found at relatively low levels <40 MPN/100 ml. The highest average concentrations of somatic and male F+ specific coliphages were 920 and 330 plaque forming units per 100 ml, respectively, detected in freshwater during the rainy season. PMMoV RNA was detected in 85% of the samples with gene sequences sharing 99–100% of nucleotide identity with PMMoV sequences available in GenBank. Quantification of PMMoV genome copies (GC) by quantitative real-time PCR indicated concentrations ranging from 1.7 × 10¹ to 1.0 × 10⁴ GC/L, with the highest number of GC detected during the rainy season. No significant correlation was observed between PMMoV occurrence by season or water type (p > 0.05). Physicochemical and indicator bacteria were not correlated with PMMoV concentrations. The abundance and prevalence of PMMoV in the karst aquifer may reflect its environmental persistence and its potential as a fecal indicator in this karst aquifer system.     

Comprehensive Study on Enteric Viruses and Indicators in Surface Water in Kyoto,Japan, During 2014–2015 Season

Certain enteric viruses that are present in the water environment are potential risk factors of waterborne infections. To better understand the impact of viruses in water, both enteric viruses and their potential indicators should be comparatively investigated. In this study, occurrences of GI- and GII-noroviruses (NoVs), sapovirus (SaV), rotavirus (RoV), Aichi virus 1 (AiV-1), enterovirus (EV), and pepper mild mottle virus (PMMoV) were quantitatively determined in surface water samples in Japan. Additionally, the genotype distribution of GI- and GII-NoVs was determined using a next-generation amplicon sequencing. PMMoV was the most abundant virus regardless of season and location, indicating its usefulness as an indicator for the viral contamination of water. Other potential indicators, AiV and EV, were less abundant than GII-NoV. Viruses other than PMMoV showed seasonality, i.e., EV and other viruses (NoVs, SaV, RoV, and AiV-1) became prevalent during summer and winter, respectively. SaV showed a relatively high abundance at a location that was affected by untreated wastewater. Regarding NoV genotypes, GI.1, GI.2, GI.4, GI.5, GI.6, GII.3, GII.4, GII.6, and GII.17 were found from the surface water samples. GII.4 and GII.17 seemed to have contributed to the high abundance of GII-NoV in the samples. Interestingly, GII.17 strains became prevalent in the water samples before becoming prevalent among gastroenteritis patients in Japan. These findings provide further insights into the properties of viruses as contaminants in the water environment.     

Environmental Surveillance of SARS-CoV-2 RNA in Wastewater and Groundwater in Quintana Roo,Mexico

The presence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA in wastewater has been reported as a result of fecal shedding of infected individuals. In this study, the occurrence of SARS-CoV-2 RNA was explored in primary-treated wastewater from two municipal wastewater treatment plants in Quintana Roo, Mexico, along with groundwater from sinkholes, a household well, and submarine groundwater discharges. Physicochemical variables were obtained in situ, and coliphage densities were determined. Three virus concentration methods based on adsorption-elution and sequential filtration were used followed by RNA isolation. Quantification of SARS-CoV-2 was done by RT-qPCR using the CDC 2020 assay, 2019-nCoV_N1 and 2019-nCoV_N2. The Pepper mild mottle virus, one of the most abundant RNA viruses in wastewater was quantified by RT-qPCR and compared to SARS-CoV-2 concentrations. The use of three combined virus concentration methods together with two qPCR assays allowed the detection of SARS-CoV-2 RNA in 58% of the wastewater samples analyzed, whereas none of the groundwater samples were positive for SARS-CoV-2 RNA. Concentrations of SARS-CoV-2 in wastewater were from 1.8 × 10³ to 7.5 × 10³ genome copies per liter (GC l⁻¹), using the N1 RT-qPCR assay, and from 2.4 × 10² to 5.9 × 10³ GC l⁻¹ using the N2 RT-qPCR assay. Based on PMMoV prevalence detected in all wastewater and groundwater samples tested, the three viral concentration methods used could be successfully applied for SARS-CoV-2 RNA detection in further studies. This study represents the first detection of SARS-CoV-2 RNA in wastewater in southeast Mexico and provides a baseline for developing a wastewater-based epidemiology approach in the area.

     

Evaluation of Virus Reduction by Ultrafiltration with Coagulation–Sedimentation in Water Reclamation

The evaluation of virus reduction in water reclamation processes is essential for proper assessment and management of the risk of infection by enteric viruses. Ultrafiltration (UF) with coagulation–sedimentation (CS) is potentially effective for efficient virus removal. However, its performance at removing indigenous viruses has not been evaluated. In this study, we evaluated the reduction of indigenous viruses by UF with and without CS in a pilot-scale water reclamation plant in Okinawa, Japan, by measuring the concentration of viruses using the real-time polymerase chain reaction (qPCR). Aichi virus (AiV) and pepper mild mottle virus (PMMoV) were targeted in addition to the main enteric viruses of concern for risk management, namely, norovirus (NoV) genogroups I and II (GI and GII) and rotavirus (RoV). PMMoV, which is a plant pathogenic virus and is present at high concentrations in water contaminated by human feces, has been suggested as a useful viral indicator. We also investigated the reduction of a spiked model virus (F-specific RNA bacteriophage MS2) to measure the effect of viral inactivation by both qPCR and plaque assay. Efficiencies of removal of NoV GI, NoV GII, RoV, and AiV by UF with and without CS were >0.5 to 3.7 log₁₀, although concentrations were below the detection limit in permeate water. PMMoV was the most prevalent virus in both feed and permeate water following UF, but CS pretreatment could not significantly improve its removal efficiency (mean removal efficiency: UF, 3.1 log₁₀; CS + UF, 3.4 log₁₀; t test, P > 0.05). CS increased the mean removal efficiency of spiked MS2 by only 0.3 log₁₀ by qPCR (t-test, P > 0.05), but by 2.8 log₁₀ by plaque assay (t-test, P < 0.01). This difference indicates that the virus was inactivated during CS + UF. Our results suggest that PMMoV could be used as an indicator of removal efficiency in water reclamation processes, but cultural assay is essential to understanding viral fate.     

Microbial Source Tracking Analysis Using Viral Indicators in Santa Lucía and Uruguay Rivers,Uruguay

The aim of this study was to determine the origin (human, bovine or porcine) and the concentration of the fecal sources of contamination in waters from Santa Lucía basin and Uruguay River in Uruguay by using host-specific viral markers (adenoviruses and polyomaviruses) as microbial source tracking (MST). Between June 2015 and May 2016, monthly collections of surface water samples were performed in six sites in Santa Lucía basin and four sites in Uruguay River (n = 120 samples). Viral concentration was carried out using an absorption-elution method. Detection and quantification of human and porcine adenovirus (HAdV and PAdV, respectively) and human and bovine polyomavirus (HPyV and BoPyV, respectively) were performed by quantitative PCR (qPCR). To evaluate the infectivity of circulating HAdV, an integrated cell culture-qPCR (ICC-qPCR) was used. A logistic regression analysis was carried out to estimate the influence of environmental variables on the virus presence in surface waters. Overall, HAdV was the prevalent (18%; 21/120) followed by BoPyV (11%; 13/120) and HPyV (3%; 3/120), whereas PAdV was not detected in this study. The mean concentration ranged from 1.5 × 10⁴ genomic copies/L (gc/L) for HAdV to 1.8 × 10² gc/L for HPyV. Infective HAdVs were observed in two out of ten analyzed samples. A significant effect of environmental temperature (p = 0.001) and river (p = 0.012) on the presence of human viruses was found. These results suggest that fecal contamination could affect the water quality of these rivers, showing deficiencies in the procedure of sewage discharge from regional cities, livestock and dairy farms.

     

Detection of Potential Infectious Enteric Viruses in Fresh Produce by (RT)-qPCR Preceded by Nuclease Treatment

Foodborne illnesses associated with contaminated fresh produce are a common public health problem and there is an upward trend of outbreaks caused by enteric viruses, especially human noroviruses (HNoVs) and hepatitis A virus (HAV). This study aimed to assess the use of DNase and RNase coupled to qPCR and RT-qPCR, respectively, to detect intact particles of human adenoviruses (HAdVs), HNoV GI and GII and HAV in fresh produce. Different concentrations of DNase and RNase were tested to optimize the degradation of free DNA and RNA from inactivated HAdV and murine norovirus (MNV), respectively. Results indicated that 10 µg/ml of RNase was able to degrade more than 4 log₁₀ (99.99%) of free RNA, and 1 U of DNase degraded the range of 0.84–2.5 log₁₀ of free DNA depending on the fresh produce analysed. The treatment with nucleases coupled to (RT)-qPCR was applied to detect potential infectious virus in organic lettuce, green onions and strawberries collected in different seasons. As a result, no intact particles of HNoV GI and GII were detected in the 36 samples analysed, HAdV was found in one sample and HAV was present in 33.3% of the samples, without any reasonable distribution pattern among seasons. In conclusion, RT-qPCR preceded by RNase treatment of eluted samples from fresh produce is a good alternative to detect undamaged RNA viruses and therefore, potential infectious viruses. Moreover, this study provides data about the prevalence of enteric viruses in organic fresh produce from Brazil.     

Coxsackievirus B4 as a Causative Agent of Diabetes Mellitus Type 1: Is There a Role of Inefficiently Treated Drinking Water and Sewage in Virus Spreading?

This study proposed to detect the enterovirus (EV) infection in children with type 1 diabetes mellitus (T1D) and to assess the role of insufficiently treated water and sewage as sources of viral spreading. Three hundred and eighty-two serum specimens of children with T1D, one hundred serum specimens of children who did not suffer from T1D as control, and forty-eight water and sewage samples were screened for EV RNA using nested RT-PCR. The number of genome copies and infectious units of EVs in raw and treated sewage and water samples were investigated using real-time (RT)-PCR and plaque assay, respectively. T1D markers [Fasting blood glucose (FBG), HbA1c, and C-peptide], in addition to anti-Coxsackie A & B viruses (CVs A & B) IgG, were measured in control, T1D-negative EV (T1D–EV^?), and T1D-positive EV (T1D–EV⁺) children specimens. The prevalence of EV genome was significantly higher in diabetic children (26.2%, 100 out of 382) than the control children (0%, 0 out of 100). FBG and HbA1c in T1D–EV^? and T1D–EV⁺ children specimens were significantly higher than those in the control group, while c-peptide in T1D–EV^? and T1D–EV⁺ children specimens was significantly lower than that in the control (n = 100; p < 0.001). Positivity of anti-CVs A & B IgG was 70.7, 6.7, and 22.9% in T1D–EV⁺, T1D–EV^?, and control children specimens, respectively. The prevalence of EV genome in drinking water and treated sewage samples was 25 and 33.3%, respectively. The prevalence of EV infectious units in drinking water and treated sewage samples was 8.5 and 25%, respectively. Quantification assays were performed to assess the capabilities of both wastewater treatment plants (WWTPs) and water treatment plants (WTPs) to remove EV. The reduction of EV genome in Zenin WWTP ranged from 2 to 4 log₁₀, while the reduction of EV infectious units ranged from 1 to 4 log₁₀. The reduction of EV genome in El-Giza WTP ranged from 1 to 3 log₁₀, while the reduction of EV infectious units ranged from 1 to 2 log₁₀. This capability of reduction did not prevent the appearance of infectious EV in treated sewage and drinking water. Plaque purification was performed for isolation of separate EV isolates from treated and untreated water and sewage samples. Characterization of the EV amplicons by RT-PCR followed by sequencing of these isolates revealed high homology (97%) with human coxsackievirus B4 (CV B4) in 60% of the isolates, while the rest of the isolates belonged to poliovirus type 1 and type 2 vaccine strains. On the other hand, characterization of the EV amplicons by RT-PCR followed by sequencing for T1D–EV⁺ children specimens indicated that all samples contained CV B4 with the same sequence characterized in the environmental samples. CV B4-contaminated drinking water or treated sewage may play a role as a causative agent of T1D in children.     

Surveillance of Enteric Viruses and Thermotolerant Coliforms in Surface Water and Bivalves from a Mangrove Estuary in Southeastern Brazil

This study was conducted to evaluate the microbiological quality of a mangrove estuary in the Vitória Bay region, Espírito Santo, Brazil. We analyzed the presence and concentration of enteric viruses and thermotolerant coliforms in water, mussels (Mytella charruana and Mytella guyanensis), and oysters (Crassostrea rhizophorae), collected over a 13-month period. Human adenovirus, rotavirus A (RVA), and norovirus genogroup II were analyzed by quantitative PCR. The highest viral load was found in RVA-positive samples with a concentration of 3.0 × 10⁴ genome copies (GC) L⁻¹ in water samples and 1.3 × 10⁵ GC g⁻¹ in bivalves. RVA was the most prevalent virus in all matrices. Thermotolerant coliforms were quantified as colony-forming units (CFU) by the membrane filtration method. The concentration of these bacteria in water was in accordance with the Brazilian standard for recreational waters (< 250 CFU 100 mL⁻¹) during most of the monitoring period (12 out of 13 months). However, thermotolerant coliform concentrations of 3.0, 3.1, and 2.6 log CFU 100 g⁻¹ were detected in M. charruana, M. guyanensis, and C. rhizophorae, respectively. The presence of human-specific viruses in water and bivalves reflects the strong anthropogenic impact on the mangrove and serves as an early warning of waterborne and foodborne disease outbreaks resulting from the consumption of shellfish and the practice of water recreational activities in the region.

     

特异性拟杆菌引物在珠江三角洲的适应性研究

在珠江三角洲地区采集人、猪、牛、狗以及家禽等33个粪便样品,高效提取基因组DNA,选取14种特异性拟杆菌引物进行检验分析,并进一步采用已知污染源类型的水样对其进行验证.结果表明:采用粪便样品DNA专属提取试剂盒和水体样品DNA专属提取试剂盒提取的基因组模版纯度和提取率均符合后续实验要求.拟杆菌通用引物2#(Bac32F/Bac708R)检出率为85%(28/33),特别是对哺乳类动物和鸡的粪便具有更高的检出率.人的拟杆菌特异性引物3#(HF134F/ Bac708R)和4#(HF183F/Bac708R)、反刍动物的拟杆菌特异性引物8#(CF128F/Bac708R)以及猪的拟杆菌特异性引物10#(PF163F/Bac708R)在珠江三角洲地区同时具有较高的灵敏度和较强的特异性.水体样品验证实验与实际污染类型相符,说明拟杆菌通用引物2#、人的特异性引物3# 和4# 以及猪的特异性引物10# 在珠江三角洲地区具有很好的适用性.     

Quantitative Detection and Characterization of Human Adenoviruses in the Buffalo River in the Eastern Cape Province of South Africa

Buffalo River is an important water resource in the Eastern Cape Province of South Africa. Over a 1-year period (August 2010?CJuly 2011), we assessed the prevalence of human adenoviruses (HAdVs) at a total of 6 sites on the river and three dams along its course. HAdVs were detected by real-time quantitative PCR in about 35?% of the samples with concentrations ranging from 1.2?×?10¹ genome copies (GC)/l to 4.71?×?10³ GC/l. HAdVs were detected at 5 of the 6 sampling sites with the detection rate ranging from 8.3?% at Rooikrantz Dam to 92?% at Parkside. The HAdV concentrations across the sampling sites were as follows: Parkside (3.25?×?10²?C4.71?×?10³?GC/); King William??s Town (1.02?×?10²?C4.56?×?10³?GC/l); and Eluxolzweni (1.17?×?10²?C3.97?×?10² GC/l). Significantly (P?<?0.05) higher concentrations were detected at the non-dam sites compared to the dam sites. A very low mean concentration of 1.86?×?10¹ HAdV GC/l was observed at Bridle Drift Dam. While HAdVs were detected only once at Rooikrantz Dam (1.74?×?10¹?GC/l), no HAdV was detected at Maden Dam. Epidemiologically important serotypes, Ad40/41, constituted 83.3?%, while Ad21 made up 16.7?% of the all HAdVs detected and were characterized by qualitative PCR. The Buffalo River presents a public health risk heightened by the presence of Ad 40/41 and Ad21. Our results make imperative the need for assessing water sources for viral contamination in the interest of public health. This work is a significant contribution to the molecular epidemiology of adenoviruses and to the best of our knowledge this is the first report on detection of enteric virus from surface waters in the Eastern Cape.     

Detection and Molecular Characterization of Aichivirus 1 in Wastewater Samples from Uruguay

Aichivirus 1 (AiV-1) is an enteric virus with 30 nm in diameter, belonging to the genus Kobuvirus in the Picornaviridae family being a causative agent of gastroenteritis in humans. The transmission is via the fecal-oral route, through person to person contact, recreation in contaminated waters, or through the consumption of contaminated food or water. The aim of this study was to determine the frequency and the molecular characterization of AiV-1 in wastewater from Uruguay. Biweekly collections from March 2011 to February 2012 were performed in the cities of Bella Unión, Salto, Paysandú, and Fray Bentos, northwestern region of Uruguay. A total of 96 samples were collected; viruses were concentrated by ultracentrifugation, and AiV-1 was detected by using a nested PCR with primers directed to a conserved region (3CD junction) of the viral genome. A high frequency of AiV-1 (n = 54) was observed at all the cities analyzed mainly in the colder months of the year. AiV-1 was not evidenced as an appropriate viral fecal indicator since when compared with other previously detected enteric viruses, no correlation was observed. All 13 characterized AiV-1 belonged to the genotype B after the phylogenetic analysis performed with the sequences obtained from the first round PCR amplicon. This study demonstrates that AiV-1 is a frequently detected enteric viruses present in wastewater and excreted by infected persons in the northwestern region of Uruguay.     

Synergistic Effects of Combined Chlorine and Vitamin B1 on the Reduction of Murine Norovirus-1 on the Oyster (Crassostrea gigas) Surface

This study investigated the synergistic effects of combined chlorine (200, 500, 700, and 1000 ppm) and vitamin B₁ (1000, 2000, and 3000 ppm) on the murine norovirus-1 (MNV-1), a human norovirus (NoV) surrogate, on oyster surface. Vitamin B₁ slightly reduced MNV-1 (0.04–0.3 log-reduction), whereas chlorine significantly reduced MNV-1 (0.4–1.0 log-reduction). The combined chlorine and vitamin B₁ resulted in a 0.52–1.97 log-reduction of MNV-1. The synergistic reduction in the MNV titer was not dependent on the concentrations of chlorine and vitamin B₁, and it ranged between 0.08 and 1.03 log₁₀ PFU/mL. The largest synergistic reduction observed was for the combined 700 ppm chlorine and 1000 ppm vitamin B₁. The pH and mechanical texture of the oysters were not significantly changed by the combined 0–1000 ppm chlorine and 3000 ppm vitamin B₁. The overall sensory acceptability were significantly (P < 0.05) reduced in oysters treated with 1000 ppm chlorine and 3000 ppm vitamin B₁ than in those treated with 0–700 ppm chlorine and 3000 ppm vitamin B₁. This study suggests that the combined 700 ppm chlorine and 3000 ppm vitamin B₁ could potentially be used to reduce NoV on oyster surface without causing concomitant changes in the mechanical texture, pH, or sensory qualities of the oysters.

     

Verification of Bacteroidales 16S rRNA markers as a complementary tool for detecting swine fecal pollution in the Yangtze Delta

To correctly assess and properly manage the public health risks associated with exposure to contaminated water, it is necessary to identify the source of fecal pollution in a watershed. In this study, we evaluated the efficacy of our two previously developed real time-quantitative PCR (qPCR) assays for the detection of swine-associated Bacteroidales genetic markers (gene 1–38, gene 3–53) in the Yangtze Delta watershed of southeastern China. The results indicated that the gene 1–38 and 3-53 markers exhibited high accuracy (92.5%, 91.7% conditional probability, respectively) in detecting Bacteroidales spp. in water samples. According to binary logistic regression (BLR), these two swine-associated markers were well correlated (P < 0.05) with fecal indicators (Escherichia coli and Enterococci spp.) and zoonotic pathogens (E. coli O157: H7, Salmonella spp. and Campylobacter spp.) in water samples. In contrast, concentrations of conventional fecal indicator bacteria (FIB) were not correlated with zoonotic pathogens, suggesting that they are noneffective at detecting fecal pollution events. Collectively, the results obtained in this study demonstrated that a swine-targeted qPCR assay based on two Bacteroidales genes markers (gene 1–38, gene 3–53) could be a useful tool in determining the swine-associated impacts of fecal contamination in a watershed.     

Assessment and Evaluation of an Integrated Hybrid Anaerobic–Aerobic Sewage Treatment System for the Removal of Enteric Viruses

The capability of a cost-effective and a small size decentralized pilot wastewater treatment plant (WWTP) to remove enteric viruses such as rotavirus, norovirus genogroup I (GGI), norovirus genogroup II (GGII), Hepatitis E virus (HEV), and adenovirus was studied. This pilot plant is an integrated hybrid anaerobic/aerobic setup which consisted of anaerobic sludge blanket (UASB), biological aerated filter (BAF), and inclined plate settler (IPS). Both the UASB and BAF are packed with a non-woven polyester fabric (NWPF). Results indicated that the overall log₁₀ reductions of enteric viruses’ genome copies through the whole system were 3.1 ± 1, 3.3 ± 0.5, and 2.6 ± 0.9 log₁₀ for rotavirus, norovirus GGI, and adenovirus, respectively. Reduction efficiency for both norovirus GGII and HEV after the different treatment steps could not be calculated because there were no significant numbers of positive samples for both viruses. The overall reduction of rotavirus infectious units through the whole system was 2.2 ± 0.8 log₁₀ reduction which is very close to the overall log₁₀ reduction of adenovirus infectious units through the whole system which was 2.1 ± 0.8 log₁₀ reduction. There was no considerable difference in the removal efficiency for different rotavirus G and P types. Adenovirus 41 was the only type detected in the all positive samples. Although the pilot WWTP investigated is cost effective, has a small footprint, does not need a long distance network pipes, and easy to operate, its efficiency to remove enteric viruses is comparable with the conventional centralized WWTPs.     

重庆市河水中粪源微生物污染特征及源解析

以重庆主要河流水体作为研究对象,使用传统培养技术评估了细菌总数、粪链球菌、肠球菌、脆弱拟杆菌、总大肠菌群及粪大肠菌群的污染水平;同时以拟杆菌作为特异性指示菌,选取人源性粪便专一指示菌引物（HF183）和猪源性粪便专一指示菌引物（Pig-2-Bac）进行源追踪.微生物培养结果表明：重庆市主要河流在春季有15.4%的研究断面未达到Ⅲ类水质,秋季有61.5%的研究断面未达到Ⅲ类水质.在春季,主城区河流主要受人类粪便污染,区县河流主要受动物粪便污染;在秋季,主城区和区县河流都主要受人类粪便污染.指示微生物指标间Pearson相关性分析结果表明粪链球菌、粪大肠菌群、肠球菌两两之间有显著相关性,肠球菌与脆弱拟杆菌有显著相关性.猪源性拟杆菌特异性生物标记Pig-2-Bac和人源拟杆菌特异性生物标记HF183对人和动物粪便污染区分成功率达100%;用这两种特异性引物对春季水样DNA进行扩增,发现13个采样点均未受到猪源粪便污染,唐家沱、朝天门、鸡冠石、合川受到人源粪便污染.     

Persistent Norovirus Contamination of Groundwater Supplies in Two Waterborne Outbreaks

Microbiological contamination of groundwater supplies causes waterborne outbreaks worldwide. In this study, two waterborne outbreaks related to microbiological contamination of groundwater supplies are described. Analyses of pathogenic human enteric viruses (noroviruses and adenoviruses), fecal bacteria (Campylobacter spp. and Salmonella spp.), and indicator microbes (E. coli, coliform bacteria, intestinal enterococci, Clostridium perfringens, heterotrophic plate count, somatic and F-specific coliphages) were conducted in order to reveal the cause of the outbreaks and to examine the effectiveness of the implemented management measures. Moreover, the long-term persistence of noro- and adenovirus genomes was investigated. Noroviruses were detected in water samples from both outbreaks after the intrusion of wastewater into the drinking water sources. In the outbreak I, the removal efficiency of norovirus genome (3.0 log₁₀ removal) in the sand filter of onsite wastewater treatment system (OWTS) and during the transport through the soil into the groundwater well was lower than the removal efficiencies of E. coli, coliform bacteria, intestinal enterococci, and spores of C. perfringens (6.2, 6.0, > 5.9, and > 4.8 log₁₀ removals, respectively). In the outbreak II, cleaning of massively contaminated groundwater well and drinking water distribution network proved challenging, and noro- and adenovirus genomes were detected up to 3 months (108 days). The long-term persistence study showed that noro- and adenovirus genomes can remain detectable in the contaminated water samples up to 1277 and 1343 days, respectively. This study highlights the transport and survival properties of enteric viruses in the environment explaining their potency to cause waterborne outbreaks.