Search

Human noroviruses (HuNoV) are the leading cause of gastrointestinal illness and environmental monitoring is crucial to prevent HuNoV outbreaks. The recent development of a HuNoV cell culture assay in human intestinal enteroids (HIEs) has enabled detection of infectious HuNoV. However, this complex approach requires adaptation of HIEs to facilitate HuNoV replication from environmental matrixes. Integrating data from 200 experiments, we examined six variables: HIE age, HIE basement membrane compounds (BMC), HuNoV inoculum processing, HuNoV inoculum volume, treatment of data below limit of detection (LOD), and cutoff criteria for determining positive HuNoV growth. We infected HIEs with HuNoV GII.4 Sydney positive stool and determined 1.4 × 10 3 genome equivalents per HIE well were required for HuNoV replication. HIE age had minimal effect on assay outcomes. LOD replacement and cutoff affected data interpretation, with lower values resulting in higher estimated HuNoV detection. Higher inoculum volumes lead to minimal decreases in HuNoV growth, with an optimal volume of 250uL facilitating capture of low concentrations of HuNoVs present in environmental isolates. Processing of HuNoV inoculum is valuable for disinfection studies and concentrating samples but is not necessary for all HIE applications. This work enhances the HuNoV HIE cell culture approach for environmental monitoring. Future HIE research should report cell age as days of growth and should clearly describe BMC choice, LOD handling, and positive cutoff., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Contamination of fomites by human norovirus (HuNoV) can initiate and prolong outbreaks. Fomite swabbing is necessary to predict HuNoV exposure and target interventions. Historically, swab recovered HuNoV has been measured by molecular methods that detect viral RNA but not infectious HuNoV. The recent development of HuNoV cultivation in human intestinal enteroids (HIEs) enables detection of infectious HuNoV. It is unknown if the swabbing process and swab matrix will allow for cultivation of fomite recovered HuNoV. We used HIEs to culture swab-recovered HuNoV GII.4 Sydney from experimentally infected surfaces-a hospital bed tray (N = 32), door handle (N = 10), and sanitizer dispenser (N = 11). Each surface was swabbed with macrofoam swabs premoistened in PBS plus 0.02% Tween80. Swab eluate was tested for infectious HuNoV by cultivation in HIE monolayers. Infectious HuNoV can be recovered from surfaces inoculated with at least 10 5 HuNoV genome equivalents/3 cm 2 . In total, 57% (N = 53) of recovered swabs contained infectious HuNoV detected by HIEs. No difference in percent positive swabs was observed between the three surfaces at p = 0.2. We demonstrate that fomite swabbing can be combined with the HIE method to cultivate high titer infectious HuNoV from the environment, filling a significant gap in HuNoV detection. Currently, high titers of HuNoV are required to measure growth in HIEs and the HIE system precludes absolute quantification of infectious viruses. However, the HIE system can provide a binary indication of infectious HuNoV which enhances existing detection methods. Identification of infectious HuNoVs from swabs can increase monitoring accuracy, enhance risk estimates, and help prevent outbreaks., Competing Interests: Author JJ was employed by company Stantec. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Overbey, Zachos, Coulter, Jacangelo and Schwab.)

With increasing interest in peracetic acid (PAA) as a disinfectant in water treatment processes, this study determined PAA treatment effects on human noroviruses (hNoVs) genotype I (GI) and genotype II (GII) as well as effects on bacteriophage MS2 and murine norovirus (MNV) in relation to pH. Across all pH conditions, PAA achieved between 0.2 and 2.5 log 10 reduction of hNoVs over 120 min contact time in buffer solution as measured by reverse transcription-qPCR (RT-qPCR). The PAA treatments produced similar RT-qPCR reductions of MS2 and MNV, in the range of 0.2-2.7 log 10 . Infectivity assays achieved > 4 log 10 reduction of both MS2 and MNV in buffer solution after 120 min contact time. Comparing PAA activity across varying pH, disinfection at pH 8.5, in general, resulted in less reduction of infectivity and molecular signals compared to pH conditions of 6.5 and 7.5. This difference was most pronounced for reductions in infectivity of MNV and MS2, with as much as 2.7 log 10 less reduction at pH 8.5 relative to lower pH conditions. This study revealed that PAA was an effective disinfectant for treatment of hNoV GI and GII, MS2 and MNV, with greatest virus reduction observed for MS2 and MNV infectivity. RT-qPCR reductions of MS2 and MNV were lower than concurrent MS2 and MNV infectivity reductions, suggesting that observed hNoV RT-qPCR reductions may underestimate reductions in hNoV infectivity achieved by PAA. Although virus disinfection by PAA occurred at all evaluated pH levels, PAA is most effective at pH 6.5-7.5.

Peracetic acid (PAA) is a strong oxidant/bactericide that has been applied in various industries (e.g., food processing, pharmaceuticals, medical device sterilization, etc.) as a disinfectant. There is increasing interest in using PAA for wastewater disinfection because it does not form halogenated byproducts, and no post-treatment quenching is required. Previous studies have demonstrated good efficiency in controlling bacteria in wastewater, but limited information is available for viruses, especially those hosted by mammals (e.g., norovirus). Therefore, a study on the infectivity reduction of murine norovirus (MNV) was undertaken to evaluate the disinfection efficacy of PAA or UV alone and in combination with UV irradiation in undisinfected secondary effluent from a municipal wastewater reclamation facility (MWW) and phosphate buffer solution (PBS) at pH 7. Experiments employing MS2 bacteriophage were also performed in parallel for comparison purposes. MS2 infectivity reduction was found to be lower than MNV infectivity reduction for each condition studied - PAA, PAA + UV, and UV disinfection. These data suggest that MS2 may not be an appropriate surrogate to accurately predict the reduction of MNV infectivity. UV irradiation, in a dose range of 5-250 mJ/cm 2 , provided linear log inactivation (-log (N/N 0 )) with a regression slope (cm 2 mJ -1 ) of 0.031-0.034 and 0.165-0.202 for MS2 and MNV, respectively. UV irradiation provided similar inactivation for MS2 and MNV in both suspensions (PBS or MWW). Low infectivity reduction of MS2 was observed when PAA was used alone at a practical dose of 1.5 mg/L and below. A greater reduction of both MNV and MS2 was observed in PAA disinfection experiments using PBS as the microbial suspension medium, than in secondary effluent. Similar results were observed in PAA + UV experiments, in which greater synergistic effects were found in PBS than in MWW. Results of OH radical formation experiments suggest the presence of radical scavengers in MWW, which resulted in less opportunity for MNV and MS2 to encounter OHradicals. This study also demonstrated that the type of water can have a substantial impact on wastewater disinfection when employing PAA or PAA + UV treatment due to the matrix effect and the presence of radical scavengers, respectively. The results from this study could be employed to aid in the conceptual design of PAA and UV disinfection facilities, especially when norovirus is the organism of concern., (Copyright © 2018. Published by Elsevier Ltd.)

Noroviruses cause significant global health burdens and waterborne transmission is a known exposure pathway. Chlorination is the most common method of disinfection for water and wastewater worldwide. The purpose of this study was to investigate the underlying causes for discrepancies in human norovirus (hNoV) resistance to free chlorine that have been previously published, and to assess hNoV GI and GII persistence during disinfection of municipal secondary wastewater (WW) effluent. Our results reveal that choice of hNoV purification methodology prior to seeding the viruses in an experimental water matrix influences disinfection outcomes in treatment studies. Common hNoV purification processes such as solvent extraction and 0.45-μm filtration were ineffective in removing high levels of organics introduced into water or wastewater samples when seeding norovirus positive stool. These methods resulted in experimental water matrices receiving an additional 190 mg/L as Cl 2 of 15-s chlorine demand and approximately 440 mg/L as Cl 2 of 30-min chlorine demand due to seeding norovirus positive stool at 1% w/v. These high organic loads impact experimental water chemistry and bias estimations of hNoV persistence. Advanced purification of norovirus positive stool using sucrose cushion ultracentrifugation and ultrafiltration reduced 15-s chlorine demands by 99% and TOC by 93% for loose (i.e. unformed diarrhea) stools. Using these methods, hNoV GI and GII persistence was investigated during free chlorination of municipal WW. A suite five of kinetic inactivation models was fit to viral reverse transcription-qPCR reduction data, and model predicted CT values for 1, 2, and 3 log 10 reduction of hNoV GI in municipal WW by free chlorine were 0.3, 2.1, and 7.8 mg-min/L, respectively. Model predicted CT values for reduction of hNoV GII in WW were 0.4, 2.0, and 7.0 mg-min/L, respectively. These results indicate that current WW treatment plant disinfection practices employing free chlorine are likely protective for public health with regards to noroviruses, and will achieve at least 3-log reduction of hNoV GI and GII RNA despite previous reports of high hNoV resistance., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

One key assumption impacting data quality in viral inactivation studies is that reduction estimates are not altered by the virus seeding process. However, seeding viruses often involves the inadvertent addition of co-constituents such as cell culture components or additives used during preparation steps which can impact viral reduction estimates by inducing non-representative oxidant demand in disinfection studies and fouling in membrane assessments. The objective of this study was therefore to characterize a mammalian norovirus surrogate, murine norovirus (MNV), and bacteriophage MS2 at sequential stages of viral purification and to quantify their potential contribution to artificial oxidant demand and non-representative membrane fouling. Our results demonstrate that seeding solvent extracted and 0.1 micron filtered MNV to ~10 5 PFU/mL in an experimental water matrix will result in additional total organic carbon (TOC) and 30 min chlorine demand of 39.2 mg/L and 53.5 mg/L as Cl 2 , respectively. Performing sucrose cushion purification on the MNV stock prior to seeding reduces the impacts of TOC and chlorine demand to 1.6 and 0.15 mg/L as Cl 2 , respectively. The findings for MNV are likely relevant for other mammalian viruses propagated in serum-based media. Thus, advanced purification of mammalian virus stocks by sucrose cushion purification (or equivalent density-based separation approach) is warranted prior to seeding in water treatment assessments. Studies employing bacteriophage MS2 as a surrogate virus may not need virus purification, since seeding MS2 at a concentration of ~10 6 PFU/mL will introduce only ~1 mg/L of TOC and ~1 mg/L as Cl 2 of chlorine demand to experimental water matrices.

Human noroviruses (hNoVs) are a known public health concern associated with the consumption of leafy green vegetables. While a number of studies have investigated pathogen reduction on the surfaces of leafy greens during the postharvest washing process, there remains a paucity of data on the level of treatment needed to inactivate viruses in the wash water, which is critical for preventing cross-contamination. The objective of this study was to quantify the susceptibility of hNoV genotype I (GI), hNoV GII, murine norovirus (MNV), and bacteriophage MS2 to free chlorine in whole leaf, chopped romaine, and shredded iceberg lettuce industrial leafy green wash waters, each sampled three times over a 4-month period. A suite of kinetic inactivation models was fit to the viral reduction data to aid in quantification of concentration-time (CT) values. Results indicate that 3-log 10 infectivity reduction was achieved at CT values of less than 0.2 mg · min/liter for MNV and 2.5 mg · min/liter for MS2 in all wash water types. CT values for 2-log 10 molecular reduction of hNoV GI in whole leaf and chopped romaine wash waters were 1.5 and 0.9 mg · min/liter, respectively. For hNoV GII, CT values were 13.0 and 7.5 mg · min/liter, respectively. In shredded iceberg wash water, 3-log 10 molecular reduction was not observed for any virus over the time course of experiments. These findings demonstrate that noroviruses may exhibit genogroup-dependent resistance to free chlorine and emphasize the importance of distinguishing between genogroups in hNoV persistence studies. IMPORTANCE Postharvest washing of millions of pounds of leafy greens is performed daily in industrial processing facilities with the intention of removing dirt, debris, and pathogenic microorganisms prior to packaging. Modest inactivation of pathogenic microorganisms (less than 2 log 10 ) is known to occur on the surfaces of leafy greens during washing. Therefore, the primary purpose of the sanitizing agent is to maintain microbial quality of postharvest processing water in order to limit cross-contamination. This study modeled viral inactivation data and quantified the free-chlorine CT values that processing facilities must meet in order to achieve the desired level of hNoV GI and GII reduction. Disinfection experiments were conducted in industrial leafy green wash water collected from a full-scale fresh produce processing facility in the United States, and hNoV GI and GII results were compared with surrogate molecular and infectivity data., (Copyright © 2017 American Society for Microbiology.)

The objective of this study was to characterize human norovirus (hNoV) GI and GII reductions during disinfection by peracetic acid (PAA) and monochloramine in secondary wastewater (WW) and phosphate buffer (PB) as assessed by reverse transcription-qPCR (RT-qPCR). Infectivity and RT-qPCR reductions are also presented for surrogate viruses murine norovirus (MNV) and bacteriophage MS2 under identical experimental conditions to aid in interpretation of hNoV molecular data. In WW, RT-qPCR reductions were less than 0.5 log 10 for all viruses at concentration-time (CT) values up to 450 mg-min/L except for hNoV GI, where 1 log 10 reduction was observed at CT values of less than 50 mg-min/L for monochloramine and 200 mg-min/L for PAA. In PB, hNoV GI and MNV exhibited comparable resistance to PAA and monochloramine with CT values for 2 log 10 RT-qPCR reduction between 300 and 360 mg-min/L. Less than 1 log 10 reduction was observed for MS2 and hNoV GII in PB at CT values for both disinfectants up to 450 mg-min/L. Our results indicate that hNoVs exhibit genogroup dependent resistance and that disinfection practices targeting hNoV GII will result in equivalent or greater reductions for hNoV GI. These data provide valuable comparisons between hNoV and surrogate molecular signals that can begin the process of informing regulators and engineers on WW treatment plant design and operational practices necessary to inactivate hNoVs.

Ribavirin is a pharmaceutical antiviral used for the treatment of RNA virus infections including norovirus, hepatitis C virus, hepatitis E virus, Lassa virus, respiratory syncytial virus, and rhinovirus. Despite the drug's history and documented efficacy, the antiviral mechanism of Ribavirin remains unclear. Mechanisms proposed include depletion of the intracellular GTP pool, immunomodulatory effects, induction of error catastrophe, inhibition of viral polymerase activity, and/or inhibition of viral capping. In the present study, we leveraged deep sequencing data to demonstrate that Ribavirin increases murine norovirus (MNV-1) viral diversity. By serial passaging MNV-1 in RAW 264.7 cells for twenty generations in the presence of Ribavirin, we demonstrated statistically significant increases in both the number of unique haplotypes and the average pairwise difference (APD). Based on statistically significant differences in the probability of nucleotide mutations based on Roche 454 sequencing, we also demonstrated that single nucleotide substitutions are increased in the presence of Ribavirin. Finally, we demonstrated Ribavirin's impact on statistically significantly reducing the relative proportion of the dominant sequence within the quasispecies., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Norovirus infections are one of the most prominent public health problems of microbial origin in the U.S. and other industrialized countries. Surveillance is necessary to prevent secondary infection, confirm successful cleanup after outbreaks, and track the causative agent. Quantitative mass spectrometry, based on absolute quantitation with stable-isotope labeled peptides, is a promising tool for norovirus monitoring because of its speed, sensitivity, and robustness in the face of environmental inhibitors. In the current study, we present two new methods for the detection of the norovirus genogroup I capsid protein using electrospray and matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. The peptide TLDPIEVPLEDVR was used to quantify norovirus-like particles down to 500 attomoles with electrospray and 100 attomoles with MALDI. With MALDI, we also demonstrate a detection limit of 1 femtomole and a quantitative dynamic range of 5 orders of magnitude in the presence of an environmental matrix effect. Due to the rapid processing time and applicability to a wide range of environmental sample types (bacterial lysate, produce, milk, soil, and groundwater), mass spectrometry-based absolute quantitation has a strong potential for use in public health and environmental sciences., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Surface disinfection, as part of environmental hygiene practices, is an efficient barrier to gastroenteritis transmission. However, surface disinfectants may be difficult to obtain in remote, resource-limited, or disaster relief settings. Electrochemical oxidants (ECO) are chlorine-based disinfectants that can be generated using battery power to electrolyze brine (NaCl) solutions. Electrolysis generates a mixed-oxidant solution that contains both chlorine (HOCl, OCl(-)) and reactive oxygen species (e.g., ·OH, O3, H2O2, and ·O2-) capable of inactivating pathogens. One onsite generator of ECO is the Smart Electrochlorinator 200 (SE-200, Cascade Designs, Inc.). In a laboratory study, we assessed ECO surface disinfection efficacy for two gastrointestinal virus surrogates: bacteriophage MS2 and murine norovirus MNV-1. We quantified both infectivity and nucleic acid inactivation using culture-dependent and independent assays. At free available chlorine concentrations of 2,500 ppm and a contact time of 30 s, ECO inactivation of infective MS2 bacteriophage exceeded 7 log10 compared to MNV-1 disinfection of approximately 2 log10. Genomic RNA inactivation was less than infective virus inactivation: MS2 RNA inactivation was approximately 5 log10 compared to MNV-1 RNA inactivation of approximately 1.5 log10. The results are similar to inactivation efficacy of household bleach when used at similar free available chlorine concentrations. Our work demonstrates the potential of ECO solutions, generated onsite, to be used for surface disinfection.

Human noroviruses (NoVs) are a significant cause of nonbacterial gastroenteritis worldwide, with contaminated drinking water a potential transmission route. The absence of a cell culture infectivity model for NoV necessitates the use of molecular methods and/or viral surrogate models amenable to cell culture to predict NoV inactivation. The NoV surrogates murine NoV (MNV), feline calicivirus (FCV), poliovirus (PV), and male-specific coliphage MS2, in conjunction with Norwalk virus (NV), were spiked into surface water samples (n = 9) and groundwater samples (n = 6). Viral persistence was monitored at 25 degrees C and 4 degrees C by periodically analyzing virus infectivity (for all surrogate viruses) and nucleic acid (NA) for all tested viruses. FCV infectivity reduction rates were significantly higher than those of the other surrogate viruses. Infectivity reduction rates were significantly higher than NA reduction rates at 25 degrees C (0.18 and 0.09 log(10)/day for FCV, 0.13 and 0.10 log(10)/day for PV, 0.12 and 0.06 log(10)/day for MS2, and 0.09 and 0.05 log(10)/day for MNV) but not significant at 4 degrees C. According to a multiple linear regression model, the NV NA reduction rates (0.04 +/- 0.01 log(10)/day) were not significantly different from the NA reduction rates of MS2 (0.05 +/- 0.03 log(10)/day) and MNV (0.04 +/- 0.03 log(10)/day) and were significantly different from those of FCV (0.08 +/- 0.03 log(10)/day) and PV (0.09 +/- 0.03 log(10)/day) at 25 degrees C. In conclusion, MNV shows great promise as a human NoV surrogate due to its genetic similarity and environmental stability. FCV was much less stable and thus questionable as an adequate surrogate for human NoVs in surface water and groundwater.

Mass spectrometry (MS) represents a rapid technique for the identification of microbial monocultures, and its adaptation to the detection of pathogens in real-world samples is a public health and homeland security priority. Norovirus, a leading cause of gastroenteritis in the world, is difficult to monitor because it cannot be cultured outside the human body. The detection of norovirus capsid protein was explored using three common MS-based methods: scanning of intact proteins, peptide mass fingerprinting, and peptide sequencing. Detection of intact target protein was limited by poor selectivity and sensitivity. Detection of up to 16 target peptides by peptide mass fingerprinting allowed for the reproducible and confident (P < 0.05) detection of the 56-kDa norovirus capsid protein in the range of 0.1 x 10(-12) to 50 x 10(-12) mol in authentic standards of recombinant norovirus virus-like particles (VLPs). To explore assay performance in complex matrixes, a non-gel-based, rapid method (2 to 3 h) for virus extraction from human stool was evaluated (72% +/- 12% recovery), and additional analyses were performed on norovirus-free stool extracts fortified with VLPs. Whereas peptide mass fingerprinting was rendered impractical by sample interferences, peptide sequencing using nanospray tandem MS facilitated unambiguous identification of > or =250 fmol of capsid protein in stool extracts. This is the first report on MS-based detection of norovirus, accomplished by using structurally identical, noninfective VLPs at clinically relevant concentrations. It represents an important milestone in the development of assays for surveillance of this category B bioterrorism agent.

Background: The role of environmental surface contamination in the propagation of norovirus outbreaks is unclear. An outbreak of acute gastroenteritis was reported among residents of a 240-bed veterans long-term-care facility., Objectives: To identify the likely mode of transmission, to characterize risk factors for illness, and to evaluate for environmental contamination in this norovirus outbreak., Methods: An outbreak investigation was conducted to identify risk factors for illness among residents and employees. Stool and vomitus samples were tested for norovirus by reverse transcription polymerase chain reaction (RT-PCR). Fourteen days after outbreak detection, ongoing cases among the residents prompted environmental surface testing for norovirus by RT-PCR., Results: One hundred twenty-seven (52%) of 246 residents and 84 (46%) of 181 surveyed employees had gastroenteritis. Case-residents did not differ from non-case-residents by comorbidities, diet, room type, or level of mobility. Index cases were among the nursing staff. Eight of 11 resident stool or vomitus samples tested positive for genogroup II norovirus. The all-cause mortality rate during the month of the outbreak peak was significantly higher than the expected rate. Environmental surface swabs from case-resident rooms, a dining room table, and an elevator button used only by employees were positive for norovirus. Environmental and clinical norovirus sequences were identical., Conclusion: Extensive contamination of environmental surfaces may play a role in prolonged norovirus outbreaks and should be addressed in control interventions.

Traveler's diarrhea (TD) is the most common infectious illness acquired by visitors to developing nations. The purpose of this study was to utilize molecular diagnostic techniques to determine the prevalence of norovirus (NoV) in TD occurring among visitors from the United States to Guatemala and Mexico. Stool samples (n = 54) were collected from 34 TD cases and analyzed for NoV by reverse transcription-PCR and oligoprobe confirmation. The overall prevalence of NoV was 65%. Interestingly, all NoV-positive stool samples were identified as genogroup I NoVs, and time spent at travel destinations was found to be an important factor in determining the frequency of infection (P = 0.003). Eleven NoV-positive stool samples also tested positive for enterotoxigenic Escherichia coli, indicating that dual infections with this leading bacterial cause of TD were very common. Results of this study suggest that NoV infection is a frequent occurrence among travelers to Mexico and Guatemala who experience episodes of TD. In addition, the simple molecular detection method utilized here will serve to facilitate more in-depth epidemiological studies of this emergent viral pathogen in travelers and other at-risk populations.

Noroviruses (NoVs) are a leading cause of gastroenteritis worldwide and are recognized as the foremost cause of foodborne illness. Despite numerous efforts, routine cell cultures have failed to yield replicating NoV. This paper describes methods used to try to grow NoV in vitro in two laboratories. Cells (A549, AGS, Caco-2, CCD-18, CRFK, CR-PEC, Detroit 551, Detroit 562, FRhK-4, HCT-8, HeLa, HEC, HEp-2, Ht-29, HuTu-80, I-407, IEC-6, IEC-18, Kato-3, L20B, MA104, MDBK, MDCK, RD, TMK, Vero and 293) were cultured on solid or permeable surfaces. Differentiation was induced using cell culture supplements such as insulin, DMSO and butyric acid. In some cases, the cells and the NoV-containing stool samples were treated with bioactive digestive additives. Variables evaluated in cultivation experiments included the method of preparation of the virus inoculum, the genotype of the virus, conditions for maintenance of cell monolayers, additives in the maintenance medium and the method of inoculation of the cells. Serial blind passage studies were performed routinely. In addition to evaluation for CPE, evidence of virus replication was sought using immunofluorescent assays to detect newly produced viral capsid antigen and RT-PCR assays to detect the viral genome. Although some infected cultures remained NoV positive by RT-PCR for up to five passages and an occasional cell in a monolayer showed evidence of specific immunofluorescence, no reproducible NoV-induced CPE was observed and all RT-PCR results that were positive initially were negative following continued passaging. Thus, attempts to develop a method for the cultivation of NoV were unsuccessful.

BackgroundNorovirus is a leading cause of acute gastroenteritis worldwide. Routine norovirus diagnosis requires stool collection. The goal of this study was to develop and validate a noninvasive method to diagnose norovirus to complement stool diagnostics and to facilitate studies on transmission.MethodsA multiplex immunoassay to measure salivary immunoglobulin G (IgG) responses to 5 common norovirus genotypes (GI.1, GII.2, GII.4, GII.6, and GII.17) was developed. The assay was validated using acute and convalescent saliva samples collected from Peruvian children

Background: Healthcare-associated norovirus outbreaks place a large burden on healthcare staff. Environmental service workers (ESWs), however, remain understudied despite high contact with potentially contaminated surfaces. Understanding the magnitude of the risk of norovirus infection in healthcare ESWs can protect workers and improve infection control.Aim: This study simulated the risk of norovirus infection for unprotected ESWs after a single fomite contact, assuming no disinfection or protective equipment, in norovirus-positive patient rooms. In addition, the risk of secondary surface transmission from norovirus-exposed ESWs was simulated.Methods: A quantitative microbial risk assessment employing two-dimensional Monte Carlo simulation with parameters extracted from the literature was used to estimate norovirus infection from multiple fomite contact scenarios defined by: norovirus source (patient vomit/diarrhoea), location (bathroom/patient room) and target outcome (ESW/secondary illness).Findings: Unprotected ESWs have a maximum estimated risk of norovirus infection of 33% (1:3) for a single fomite contact in a room where a norovirus-positive patient had a diarrhoeal event. Patient vomit events lead to fomite contact risk estimates that are four orders of magnitude lower than those for diarrhoeal events. The estimated risk of secondary illness from touching a common surface is as high as 25% (1:4) after single fomite exposure following a diarrhoeal event.Conclusions: A single fomite contact may lead to sizable risk of norovirus infection in ESWs if personal protective equipment and disinfection are not used appropriately. ESWs can also transfer virus to secondary surfaces, initiating further infections. Interventions are needed to reduce fomite transfer of norovirus, and protect patients and staff from nosocomial infections. [ABSTRACT FROM AUTHOR]