Virucidal Efficacy of a Hydrogen Peroxide Nebulization Against Murine Norovirus and Feline Calicivirus,Two Surrogates of Human Norovirus

Human noroviruses (HuNoV) are amongst the leading causes of acute non-bacterial gastroenteritis in humans and can be transmitted via person-to-person contact, via contact with contaminated surfaces or by consumption of contaminated food. Contaminated surfaces in healthcare settings contribute to the transmission of viruses. No-touch automated room disinfection systems might prevent such a spread of contamination and thus their virucidal effect needs to be evaluated. The aim of this study was to assess the efficacy of a nebulization system spraying hydrogen peroxide on two main surrogates of HuNoV, namely murine norovirus (MNV) and feline calicivirus (FCV). The viruses were dried on cover glasses and on stainless steel discs and exposed to nebulization. The number of infectious viral particles and genomic copies before and after the nebulization was compared. The efficacy in reducing infectivity of both surrogates was demonstrated. For the infectious viral titre of MNV and FCV, a log₁₀ reduction factor ≥4.84 and 4.85 was observed after nebulization, respectively, for tests on cover glasses and ≥3.90 and 5.30, respectively, for tests on stainless steel discs. Only low reductions in genomic copy numbers were observed for both surrogates. The nebulization of hydrogen peroxide showed a clear virucidal effect on both HuNoV surrogates, MNV and FCV, on two different carriers and the use of nebulization should be promoted in complementarity with conventional disinfection methods in healthcare settings and food processing facilities to reduce viral load and spread of contamination.     

Use of ATP Readings to Predict a Successful Hygiene Intervention in the Workplace to Reduce the Spread of Viruses on Fomites

The purpose of this study was to validate the use of adenosine triphosphate (ATP) for evaluating hygiene intervention effectiveness in reducing viral dissemination in an office environment. The bacterial virus MS-2 was used to evaluate two scenarios, one where the hand of an individual was contaminated and another where a fomite was contaminated. MS-2 was selected as a model because its shape and size are similar to many human pathogenic viruses. Two separate experiments were conducted, one in which the entrance door push plate was inoculated and the other in which the hand of one selected employee was inoculated. In both scenarios, 54 selected surfaces in the office were tested to assess the dissemination of the virus within the office. Associated surface contamination was also measured employing an ATP meter. More than half of the tested hands and surfaces in the office were contaminated with MS-2 within 4 h. Next, an intervention was conducted, and each scenario was repeated. Half of the participating employees were provided hand sanitizer, facial tissues, and disinfecting wipes, and were instructed in their use. A significant (p < 0.05) reduction was observed in the number of surfaces contaminated with virus. This reduction in viral spread was evident from the results of both viral culture and the surface ATP measurements, although there was no direct correlation between ATP measurements with respect to viral concentration. Although ATP does not measure viruses, these results demonstrate that ATP measurements could be useful for evaluating the effectiveness of hygiene interventions aimed at preventing viral spread in the workplace.     

Comparative Virucidal Efficacy of Seven Disinfectants Against Murine Norovirus and Feline Calicivirus,Surrogates of Human Norovirus

Human noroviruses (HuNoV) are the leading cause of acute non-bacterial gastroenteritis in humans and can be transmitted either by person-to-person contact or by consumption of contaminated food. A knowledge of an efficient disinfection for both hands and food-contact surfaces is helpful for the food sector and provides precious information for public health. The aim of this study was to evaluate the effect of seven disinfectants belonging to different groups of biocides (alcohol, halogen, oxidizing agents, quaternary ammonium compounds, aldehyde and biguanide) on infectious viral titre and on genomic copy number. Due to the absence of a cell culture system for HuNoV, two HuNoV surrogates, such as murine norovirus and feline calicivirus, were used and the tests were performed in suspension, on gloves and on stainless steel discs. When, as criteria of efficacy, a log reduction >3 of the infectious viral titre on both surrogates and in the three tests is used, the most efficacious disinfectants in this study appear to be biocidal products B, C and D, representing the halogens, the oxidizing agents group and a mix of QAC, alcohol and aldehyde, respectively. In addition, these three disinfectants also elicited a significant effect on genomic copy number for both surrogate viruses and in all three tests. The results of this study demonstrate that a halogen compound, oxidizing agents and a mix of QAC, alcohol and aldehyde are advisable for HuNoV disinfection of either potentially contaminated surfaces or materials in contact with foodstuffs.     

Prevalence of Human Parainfluenza Viruses and Noroviruses Genomes on Office Fomites

The aim of this study was to evaluate the potential role of office fomites in respiratory (human parainfluenza virus 1—HPIV1, human parainfluenza virus 3—HPIV3) and enteric (norovirus GI—NoV GI, norovirus GII—NoV GII) viruses transmission by assessing the occurrence of these viruses on surfaces in office buildings. Between 2016 and 2017, a total of 130 surfaces from open-space and non-open-space rooms in office buildings located in one city were evaluated for HPIV1, HPIV3, NoV GI, and NoV GII viral RNA presence. Detection of viruses was performed by RT-qPCR method. Study revealed 27 positive samples, among them 59.3% were HPIV3-positive, 25.9% HPIV1-positive, and 14.8% NoV GII-positive. All tested surfaces were NoV GI-negative. Statistical analysis of obtained data showed that the surfaces of office equipment including computer keyboards and mice, telephones, and desktops were significantly more contaminated with respiratory viruses than the surfaces of building equipment elements such as door handles, light switches, or ventilation tracts (χ ² p = 0.006; Fisher’s Exact p = 0.004). All examined surfaces were significantly more contaminated with HPIVs than NoVs (χ ² p = 0.002; Fisher’s Exact p = 0.003). Office fomites in open-space rooms were more often contaminated with HPIVs than with NoVs (χ ² p = 0.016; Fisher’s Exact p = 0.013). The highest average concentration of HPIVs RNA copies was observed on telephones (1.66 × 10² copies/100 cm²), while NoVs on the light switches (1.40 × 10² copies/100 cm²). However, the Kruskal–Wallis test did not show statistically significant differences in concentration levels of viral RNA copies on surfaces between the all tested samples. This study unequivocally showed that individuals in office environment may have contact with both respiratory and enteric viral particles present on frequently touched surfaces.     

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.     

Identification of Enteric Viruses in Foods from Mexico City

Foodborne viruses are a common and, probably, the most under-recognized cause of outbreaks of gastroenteritis. Among the main foods involved in the transmission of human enteric viruses are mollusks, and fruits and vegetables irrigated with wastewater and/or washed with non-potable water or contaminated by contact with surfaces or hands of the infected personnel during its preparation. In this study, 134 food samples were analyzed for the detection of Norovirus, Rotavirus, and Hepatitis A virus (HAV) by amplification of conserved regions of these viruses. From the 134 analyzed samples, 14 were positive for HAV, 6 for Norovirus, and 11 for Rotavirus. This is the first report in Mexico where emphasis is given to the presence of HAV and Norovirus on perishable foods and food from fisheries, as well as Rotavirus on frozen vegetables, confirming the role of vegetables and bivalve mollusks as transmitting vehicles of enteric viruses.     

Environmental Conditions Leading to Shellfish Contamination and Related Outbreaks

Human fecal wastes contain a large variety of viruses that can enter the environment through discharge of waste materials from infected individuals. Despite the high diversity of viruses that are introduced into the environment by human fecal pollution, only a few have been recognized to cause disease in association with consumption of contaminated shellfish. To explain bivalve mollusks contamination, several factors including human epidemiology, virus persistence through sewage treatment plant, and shellfish uptake may be suggested. Considering different outbreaks described in the literature, the most common route for transmission is accidental contamination after heavy rainfall, when extra loads cause an overflow, and release of untreated sewage into the aquatic environment. Outbreak analysis also demonstrates the impact on shellfish consumption of some viral strain transmission and thus their impact on molecular epidemiology, especially for norovirus. To limit shellfish contamination and thus to protect the consumer, the most desirable and effective option is to reduce the viral input.     

Application of a Swab Sampling Method for the Detection of Norovirus and Rotavirus on Artificially Contaminated Food and Environmental Surfaces

Noroviruses and rotaviruses are the leading causes of non-bacterial gastroenteritis in humans worldwide. Virus-contaminated food and surfaces represent an important risk to public health. However, established detection methods for the viruses in food products are laborious and time-consuming. Here, we describe a detailed swabbing protocol combined with real-time RT-PCR for norovirus and rotavirus detection on artificially contaminated food and environmental surfaces. Recovery rates between 2 and 78% for norovirus and between 8 and 42% for rotavirus were determined for contaminated food surfaces of apple, pepper, cooked ham and salami. From contaminated environmental surfaces (stainless steel, ceramic plate, polyethylene, wood), recovery rates between 26 and 52% (norovirus) and between 10 and 58% (rotavirus) were determined. The results demonstrate the suitability of the swab sample method for virus detection on food and environmental surfaces. Compared to other methods, it is easy to perform and significantly time-saving, predestining it for routine testing.     

环境因素对病毒在水体中生存与传播的影响

目前,新型冠状病毒（2019-nCoV）在全球范围内广泛传播.2019-nCoV在粪便和污水中的检出揭示了病毒介水传播的可能性.了解各类环境因素对水源性病毒生存与传播的影响对介水传染病的防控及病毒风险评估具有重要意义.通过对国内外相关研究的文献调研,探究了影响病毒在水体中生存与传播的主要环境因素.研究表明,致病病毒在水环境中的存活和传播潜力与紫外线照射、水体温度、pH、盐度以及水体中存在的微生物和悬浮颗粒物等环境条件密切相关.即:①低温能够大幅延长病毒的存活时间,更有利于病毒在水体中的传播;高温能够加速病毒失活从而削弱病毒的传播潜力.②紫外线照射通过破坏病毒核酸能够有效去除和灭活病毒.③水体中的微生物能够产生对病毒颗粒不利的代谢物,或利用病毒衣壳作为营养来源,从而导致病毒失活.④各类水体中存在的大量悬浮颗粒物对病毒的吸附大大延长了病毒的存活时间,从而增强了病毒在水体中的潜在传播能力;此外,悬浮颗粒物能够促进或阻碍病毒在多孔介质中的传输与滞留.⑤pH通过改变病毒颗粒的表面电荷,影响病毒的团聚,从而影响病毒在水环境中的持久性.⑥无机离子通过影响病毒的团聚和吸附性能,从而改变其活性.环境因素对病毒存活的影响可能因不同的病毒特性而异,因此,应进一步探讨水环境中2019-nCoV存活能力的变异性,2019-nCoV在废水和饮用水处理过程中的赋存与归趋,并对废水、娱乐水域和饮用水中的2019-nCoV进行长期监测和定量风险评估.      

Semi-Direct Lysis of Swabs and Evaluation of Their Efficiencies to Recover Human Noroviruses GI and GII from Surfaces

Enteric viruses such as noroviruses (NoVs) continue to be the cause of widespread viral outbreaks due to person-to-person transmission, contaminated food, and contaminated surfaces. In order to optimize swabbing methodology for the detection of viruses on (food) contact surfaces, three swab elution/extraction strategies were compared in part one of this study, out of which, one strategy was based on the recently launched ISO protocol (ISO/TS 15216-1) for the determination of hepatitis A virus and NoV in food using real-time RT-PCR (RT-qPCR). These three swab elution/extraction strategies were tested for the detection of GI.4 and GII.4 NoV on high-density polyethylene (HD-PE) surfaces with the use of cotton swabs. For detection of GI.4 and GII.4, the sample recovery efficiency (SRE) obtained with the direct lysis strategy (based on ISO/TS 15216-1) was significantly lower than the SRE obtained with both other strategies. The semi-direct lysis strategy was chosen to assess the SRE of two common swabs (cotton swab and polyester swab) versus the biowipe (Biomérieux, Lyon, France) on three surfaces (HD-PE, neoprene rubber (NR), and nitrile gloves (GL)). For both surfaces, HD-PE and GL, no significant differences in SREs of GI.4 and GII.4 NoVs were detected between the three different swabs. For the coarser NR, biowipes turned out to be the best option for detecting both GI.4 and GII.4 NoV.     

病毒在无生命物体表面存活时间及其影响因素分析

2020年3月11日,世界卫生组织宣布新型冠状病毒肺炎（COVID-19,简称“新冠肺炎”）疫情全球大流行.病毒性传染病在人群中的发生需具备3个相互关联的条件,即传染源、传播途径和易感人群,其中传播途径是一个关键环节.从环境传播的角度,综述了包括SARS（重症急性呼吸综合征）病毒、MERS（中东呼吸综合征）病毒等病毒在无生命物体表面的存活时间及影响因素.结果表明:包括冠状病毒在内的一些具有传染性的病毒能在多种无生命物体表面存活一定时间,进而可能通过物体表面进行环境传播,造成潜在的健康风险;物体材质、温度、湿度以及病毒载量等因素是影响病毒在无生命物体表面存活的主要因素.在上述分析的基础上,对医疗废物和生活垃圾的收集、运输、处理等过程提出了针对性的防控措施,以期对新型冠状病毒（2019-nCoV）以及未来可能出现的其他病毒的环境传播防控对策提供参考.      

Capsid and Infectivity in Virus Detection

The spectacular achievements and elegance of viral RNA analyses have somewhat obscured the importance of the capsid in transmission of viruses via food and water. The capsid’s essential roles are protection of the RNA when the virion is outside the host cell and initiation of infection when the virion contacts a receptor on an appropriate host cell. Capsids of environmentally transmitted viruses are phenomenally durable. Fortuitous properties of the capsid include antigenicity, isoelectric point(s), sometimes hemagglutination, and perhaps others. These can potentially be used to characterize capsid changes that cause or accompany loss of viral infectivity and may be valuable in distinguishing native from inactivated virus when molecular detection methods are used.     

One-year Surveillance of Human Enteric Viruses in Raw and Treated Wastewaters,Downstream River Waters,and Drinking Waters

Human enteric viruses are a major cause of waterborne diseases, and can be transmitted by contaminated water of all kinds, including drinking and recreational water. The objectives of the present study were to assess the occurrence of enteric viruses (enterovirus, norovirus, adenovirus, hepatitis A and E virus) in raw and treated wastewaters, in rivers receiving wastewater discharges, and in drinking waters. Wastewater treatment plants’ (WWTP) pathogen removal efficiencies by adenovirus quantitative real-time PCR and the presence of infectious enterovirus, by cell culture assays, in treated wastewaters and in surface waters were also evaluated. A total of 90 water samples were collected: raw and treated wastewaters (treated effluents and ultrafiltered water reused for industrial purposes), water from two rivers receiving treated discharges, and drinking water. Nested PCR assays were used for the identification of viral DNA/RNA, followed by direct amplicon sequencing. All raw sewage samples (21/21), 61.9 % of treated wastewater samples (13/21), and 25 % of ultrafiltered water samples (3/12) were contaminated with at least one viral family. Multiple virus families and genera were frequently detected. Mean positive PCRs per sample decreased significantly from raw to treated sewage and to ultrafiltered waters. Moreover, quantitative adenovirus data showed a reduction in excess of 99 % in viral genome copies following wastewater treatment. In surface waters, 78.6 % (22/28) of samples tested positive for one or more viruses by molecular methods, but enterovirus-specific infectivity assays did not reveal infectious particles in these samples. All drinking water samples tested negative for all viruses, demonstrating the effectiveness of treatment in removing viral pathogens from drinking water. Integrated strategies to manage water from all sources are crucial to ensure water quality.     

Development of a Practical Method to Detect Noroviruses Contamination in Composite Meals

Various methods to detect foodborne viruses including norovirus (NoV) in contaminated food have been developed. However, a practical method suitable for routine examination that can be applied for the detection of NoVs in oily, fatty, or emulsive food has not been established. In this study, we developed a new extraction and concentration method for detecting NoVs in contaminated composite meals. We spiked NoV-GI.4 or -GII.4 stool suspension into potato salad and stir-fried noodles. The food samples were suspended in homogenizing buffer and centrifuged to obtain a food emulsion. Then, anti-NoV-GI.4 or anti-NoV-GII.4 rabbit serum raised against recombinant virus-like particles or commercially available human gamma globulin and Staphylococcus aureus fixed with formalin as a source of protein A were added to the food emulsion. NoV-IgG-protein A-containing bacterial complexes were collected by centrifugation, and viral RNA was extracted. The detection limits of NoV RNA were 10–35 copies/g food for spiked NoVs in potato salad and stir-fried noodles. Human gamma globulin could also concentrate other NoV genotypes as well as other foodborne viruses, including sapovirus, hepatitis A virus, and adenovirus. This newly developed method can be used as to identify NoV contamination in composite foods and is also possibly applicable to other foodborne viruses.     

Detection Method for Avian Influenza Viruses in Water

Recent events have shown that humans may become infected with some pathogenic avian influenza A viruses (AIV). Since soil and water, including lakes, rivers, and seashores, may be contaminated by AIV excreted by birds, effective methods are needed for monitoring water for emerging viruses. Combining water filtration with molecular methods such as PCR is a fast and effective way for detecting viruses. The objective of this study was to apply a convenient method for the detection of AIV in natural water samples. Distilled water and lake, river, and seawater were artificially contaminated with AIV (H5N3) and passed through a filter system. AIV was detected from filter membrane by real-time RT-PCR. The performance of Zetapor, SMWP, and Sartobind D5F membranes in recovering influenza viruses was first evaluated using contaminated distilled water. SWMP, which gave the highest virus recoveries, was then compared with a pre-filter combined GF/F filter membrane in a trial using natural water samples. In this study, the cellulose membrane SMWP was found to be practical for recovery of AIVs in water. Viral yields varied between 62.1 and 65.9% in distilled water and between 1 and 16.7% in natural water samples. The borosilicate glass membrane GF/F combined with pre-filter was also feasible in filtering natural water samples with viral yields from 1.98 to 7.33%. The methods described can be used for monitoring fresh and seawater samples for the presence of AIV and to determine the source of AIV transmission in an outbreak situation.     

Survival of Respiratory Viruses on Fresh Produce

In addition to enteric viruses of fecal origin, emerging zoonotic viruses such as respiratory coronaviruses and influenza viruses may potentially be transmitted via contaminated foods. The goal of this study was to determine the recovery efficiencies and the survival of two respiratory viruses, namely, adenovirus 2 (Ad2) and coronavirus 229E (CoV229E), on fresh produce in comparison to the enteric poliovirus 1 (PV1). Adenovirus was recovered with efficiencies of 56.5, 31.8, and 34.8 % from lettuce, strawberries, and raspberries, respectively. Coronavirus was recovered from lettuce with an efficiency of 19.6 % yet could not be recovered from strawberries. Poliovirus was recovered with efficiencies of 76.7 % from lettuce, but only 0.06 % from strawberries. For comparison purposes, the survival of Ad2, CoV229E, and PV1 was determined for periods up to 10 days on produce. The enteric PV1 survived better than both respiratory viruses on lettuce and strawberries, with only ≤1.03 log₁₀ reductions after 10 days of storage at 4 °C compared to CoV229E not being recovered after 4 days on lettuce and reductions of 1.97 log₁₀ and 2.38 log₁₀ of Ad2 on lettuce and strawberries, respectively, after 10 days. Nevertheless, these respiratory viruses were able to survive for at least several days on produce. There is therefore the potential for transfer to the hands and subsequently to the mucosa via rubbing the eyes or nose. In addition, some respiratory coronaviruses (e.g., severe acute respiratory syndrome coronavirus) and adenoviruses are also capable of replication in the gut and there is thus some potential for acquisition through the consumption of contaminated produce.