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Wash water from fresh vegetables and root vegetables is an important vehicle for foodborne virus transmission. However, there is lack of assessing rapid viral inactivation strategies in wash water characterized by a high soil content at the post-harvest stage. Considering the significance of food safety during the washing stage for fresh and root vegetable produce prior to marketing, we assessed the inactivation efficacy by using chlorine dioxide (ClO 2 ) and peracetic acid (PAA) against a surrogate of human norovirus (murine norovirus 1, MNV-1) and hepatitis A virus (HAV), in wash water containing black soil and clay loam. The results indicated that MNV-1 and HAV were reduced to the process limit of detection (PLOD), with reductions ranging from 4.89 to 6.35 log 10 PFU, and 4.63 to 4.96 log 10 PFU when treated with ClO 2 at 2.5 ppm for 10 mins. Comparatively, when treated with 500 ppm of PAA for 10 mins, MNV-1 and HAV were maximum reduced to 1.75 ± 0.23 log 10 PFU (4.50 log 10 PFU reduction) and 2.13 ± 0.12 log 10 PFU (2.72 log 10 PFU reduction). This demonstrated the efficacy of ClO 2 in eliminating foodborne viruses in soil-rich wash water. When we validated the recovery of the virus from two types of wash water, the pH (9.24 ± 0.33 and 5.95 ± 0.05) had no impact on the recovery of MNV-1, while the recovery of HAV was less than 1 %. By adjusting the pH to a neutral level, recovery of HAV and its RNA levels was increased to 15.94 and 3.89 %. Thus, this study emphasized the critical role of pH in the recovery of HAV from the complex soil-rich aqueous environment, and the efficacy of ClO 2 serving as a pivotal reference for the development of control strategies against foodborne viruses in the supply chain of fresh and root vegetables., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

The objective of this study was to generate small interfering RNA (siRNA) to knockdown antiviral chemokine-related genes in fetal rhesus monkey kidney (FRhK-4) cells. We generated siRNA duplexes to suppress antiviral chemokines like CXCL10 and CCL4 in FRhK-4 cells by downregulating interferon regulatory factor (IRF) 3 and IRF7. Three siRNA duplexes (si-F-IRF3-1, si-F-IRF3-2, and si-F-IRF3-3) targeting IRF3, and one siRNA duplex (si-F-IRF7) targeting IRF7 were generated. A nontarget siRNA duplex was used as the negative control. The nontarget or target siRNA duplexes (si-F-IRF3-1, si-F-IRF3-2, si-F-IRF3-3, and si-F-IRF7) were transfected into FRhK-4 cells using transfection reagents, and they were then incubated at 37°C for 6 h with 5% CO 2 . After 6 h, the medium was removed, and fresh medium was added to each cell, and they were then incubated at 37°C for 48 h with 5% CO 2 . The transfected FRhK-4 cells were infected with hepatitis A virus (HAV) HM-175/18f (viral titer: 10 5  PFU/mL) and incubated at 37°C for 3 h with 5% CO 2 for HAV propagation. The expression levels of chemokines, including CXCL10 and CCL4, under the regulation of IRF3 and IRF7 in the transfected FRhK-4 cells were measured using quantitative real-time polymerase chain reaction after 3 h of HAV infection. The results indicated that CXCL10 and CCL4 expression levels were decreased in FRhK-4 cells transfected with si-F-IRF3-1, si-F-IRF3-3, or si-F-IRF7 (p < 0.05) compared to those in the negative control. These results indicate that si-F-IRF3-1 and si-F-IRF3-3, and si-F-IRF7 successfully knocked down IRF3 and IRF7 in FRhK-4 cells, respectively and suppressed antiviral chemokines. These siRNAs could be used to suppress antiviral chemokines in FRhK-4 cells., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

The performance of dishwashers in removing live viruses is an important informative value in practical applications. Since foodborne viruses are present in contaminated food surfaces and water environments. Insufficient washing of dishes typically makes a carrier of foodborne viruses. Dishwashers have shown excellent performance in removing bacterial pathogens, but very limited reports related to eliminate foodborne viruses on contaminated dish surfaces. Here, murine norovirus 1 (MNV-1), hepatitis A virus (HAV), and human coronavirus 229E (HCoV-229E) were experimentally inoculated on the dish surfaces (plate, rice bowl, and soup bowl). Plaque assay, 50% tissue culture infectious dose (TCID 50 ), and real-time quantitative polymerase chain reaction (RT-qPCR) were conducted to determine their removal efficiency of them through the general wash program of household dishwashers. Using titration assay, MNV-1 and HAV were reduced by 7.44 and 6.57 log10 PFU/dish, and HCoV-229E was reduced by 6.43 log10 TCID 50 /dish through the general wash program, achieving a ≥ 99.999% reduction, respectively. Additionally, RT-qPCR results revealed that viral RNA of MNV-1 and HCoV-229E reduced 5.02 and 4.54 log10 genome copies/dish; in contrast, HAV was not detected on any dish surfaces. This study confirmed the performance of household dishwashers in removing pathogenic live viruses through the general wash program. However, residual viral RNA was not sufficiently removed. Further studies are needed to determine whether the viral RNA can be sufficiently removed using combination programs in household dishwashers., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

A carrier (stainless steel disc as a default carrier) testing method is very needed for use in the actual food-processing fields by following the standard guideline. Here, we aimed to compare the virucidal efficacy of four commercial liquid disinfectants, including sodium hypochlorite (NaOCl), chlorine dioxide (ClO 2 ), and peracetic acid (PAA) against hepatitis A virus (HAV) following the OECD guideline protocol based on the quantitative carrier testing method and compared carrier testing results with the suspension testing results. The OECD method specifies a test for establishing whether a chemical disinfectant or a microbicide has a virucidal activity on hard non-porous surfaces. The antiviral efficacy was evaluated by plaque assays, and disinfectants were considered effective if the virus reduction was greater than or equal to 3 log 10 (99.9% decrease) for carrier or 4 log 10 (99.99% decrease) for suspension tests. Results indicated that ClO 2 above 500 ppm and 50% ethanol were effective in the carrier test method. In contrast, more than 200 ppm NaOCl and 50 ppm ClO 2 for all exposure times and 70% ethanol with contact for more than 5 min were effective in suspension tests. Treatment with PAA (80-2500 ppm) were not effective in carrier or suspension tests. Therefore, we recommend the use of more than 500 ppm ClO 2 or 50% ethanol with exposure for 10 min to disinfect surfaces that may be contaminated with HAV. Thus, these results could be effective in establishing official antiviral efficacy testing methods and basic data., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Bivalve molluscan shellfish, such as oysters, clams, and cockles, are well-recognized as vectors that concentrate foodborne pathogens by filter feeding. The objective of this study was to investigate the distribution and persistence of hepatitis A virus (HAV) in experimentally contaminated oysters that were either fed or not fed with algae. Oysters were experimentally contaminated with HAV and maintained in depuration conditions. qRT-PCR, immunohistochemistry (IHC), and in situ hybridization (ISH) were performed on oyster samples collected at 0, 1, 3, 5, and 7 days post-inoculation. When HAV-contaminated oysters were depurated for 7 days, HAV was detected in 91.1-97.8% of the digestive glands and gills. While the high viral load in the digestive glands in oysters did not change significantly regardless of algae-feeding, the viral load of the gills gradually decreased in both groups during the depuration. HAV antigen and RNA were detected in the digestive diverticula and connective tissues by both IHC and ISH. HAV was detected in the stomach, intestine, and gills by only ISH. The distribution of HAV in various oyster tissues may explain the persistence of contamination in oysters during the depuration process., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Red ginseng has a variety of bioactive functions and is widely used as an oriental medicinal herb and food ingredient. The aim of this study was to investigate the antiviral effect of red ginseng extract and ginsenosides against hepatitis A virus (HAV). To examine the antiviral effect against HAV, 0 to 10 μg/mL of red ginseng and purified ginsenoside Rb1 and Rg1 were pre-treated or co-treated on FRhK-4 cells. The HAV titer decreased significantly in all groups pretreated with red ginseng or purified ginsenosides. The reduction of HAV was significant in FRhK-4 cells pre-treated only with red ginseng. Our results showed that red ginseng and ginsenoside Rg1 and Rb1 could decrease HAV titers., (© 2013 Institute of Food Technologists®)

Hepatitis A is a major infectious disease epidemiologically associated with foodborne and waterborne outbreaks. Molecular detection using real-time RT-PCR to detect the hepatitis A virus (HAV) in contaminated vegetables can be hindered by low-virus recoveries during the concentration process and by natural PCR inhibitors in vegetables. This study evaluated three virus concentration methods from vegetables: polyethylene glycol (PEG) precipitation, ultrafiltration (UF), and immunomagnetic separation (IMS). UF was the most efficient concentration method, while PEG and IMS were very low for the recovery rate of HAV. These results demonstrate that UF is the most appropriate method for recovering HAV from contaminated vegetables and that this method combined with the real-time RT-PCR assay may be suitable for routine laboratory use.

This study aimed to develop a specific and sensitive duplex reverse transcription polymerase chain reaction enzyme-linked immunosorbent assay (duplex RT-PCR-ELISA) for hepatitis A virus (HAV) and hepatitis E virus (HEV). Duplex RT-PCR-ELISA could detect and differentiate HAV and HEV with specific probes. When ELISA technique was used to detect probe-bound RT-PCR products, duplex RT-PCR-ELISA could detect as little as 0.1 ng/μL HAV and HEV from clinical samples. Human norovirus, enterovirus, poliovirus, murine norovirus and feline calicivirus were used for the specificity test; all were negative. Therefore duplex RT-PCR-ELISA can be used for the simultaneous detection of HAV and HEV in contaminated fecal samples., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Fresh root vegetables are one of the main causes of viral foodborne diseases; however, the associated risk has not been fully considered. Therefore, the prevalence of five foodborne viruses including norovirus (NoV), hepatitis A virus (HAV), rotavirus (RotaV), adenovirus (AdV), and astrovirus (AstroV) in pre- and post-washed root vegetables (potato, carrot, sweet potato, and radish) were investigated by RT-qPCR or qPCR, nested RT-PCR, and sequencing. AdV and NoV GII were only detected in pre-washed samples. NoV GII was found in 1% and 2% of the potatoes and carrots, respectively and AdV in 4% and 10% of the potatoes and radishes, respectively. HAV was detected in 5.8% and 1.3% of the pre-and post-washed sweet potatoes, respectively and in 8% and 2% of the post-washed carrots and radishes, respectively. NoV GI and AstroV was not detected in any of the root vegetables. Sequencing and phylogenetic analysis revealed that the genotypes of AdV, NoV, and HAV were belonged to human AdV 41, NoV GII.3, and HAV IA and IB, respectively. This molecular surveillance study clearly demonstrated that human pathogenic viruses are found in root vegetables, and it is possible that they may serve as a potential reservoir for such viruses. Therefore, further research is necessary to confirm this hypothesis. Nevertheless, washing produce can lower the risks of contracting diseases due to certain root vegetables. • HAV was detected in both pre-and post-washed root vegetables regardless of washing. • AdV and NoV GII were detected only in pre-washed root vegetables. • AstroV was not detected in any root vegetables. • Detected viruses were subsequently confirmed and characterized by sequencing. • AdV, NoV, and HAV genotypes belonged to human AdV 41, NoV GII.3, and HAV IA and IB. [ABSTRACT FROM AUTHOR]

Plant- and soil-originated polymerase chain reaction (PCR) inhibitors like polyphenols, polysaccharides, and humic acid (HA) can impact the sensitivity of foodborne virus detection at a low concentration in post-washing water (PWW). This study aims to efficiently detect each virus by inoculating 102 PFU/mL of murine norovirus 1 (MNV-1) and hepatitis A virus (HAV) into post-washing water (PWW) containing various polymerase chain reaction (PCR) inhibitors, including humic acid (HA), pectin, starch, and gallic acid. To optimize the detection of these viruses, a paper filtration-coupled ultrafiltration (PFC-UF) method was used in combination with the polyaluminum chloride (PAC) and polyvinylpyrrolidone (PVP) RNA extraction method, pectinase treatment, and various commercial kits. The PFC-UF with the OneStep PCR Inhibitor Removal Kit can effectively remove HA in PWW when the HA content was less than 1.0 mg/L. However, PFC-UF with the PVP RNA extraction, pectinase treatment, and chloroform/butanol separation were not effective to improve viruses detection. PFC-UF with OneStep PCR Inhibitor Removal Kit successfully recovered MNV-1 (17.58 ± 7.28%) and HAV (7.37 ± 4.06%) from all 9 samples with high soil content and PCR inhibitors. This study successfully addressed the challenge of recovering viruses under extreme conditions of high soil content and PCR inhibitors. The findings provide a promising platform for validating the surveillance of foodborne viruses in washing water at farm fields or washing facilities. • Examined the recovery of viruses in the PWW containing PCR inhibitors, separately. • Effectively detected MNV-1 and HAV from soil-rich PWW containing PCR inhibitors. • It provides a feasible way to assess foodborne viruses at a low concentration in PWW. • Higher concentrations of PCR inhibitors increased the difficulty in virus recovery. [ABSTRACT FROM AUTHOR]

Herbal plants have long been used as traditional medicines to treat diseases caused by microbial pathogens. The hepatitis A virus (HAV) causes acute liver infection through the fecal–oral route. Although the antimicrobial activities of herbal extracts against bacterial and some viral pathogens have been extensively studied, their antiviral properties against HAV have not been investigated thus far. This study was designed to investigate the inhibitory effect of 16 herbal extracts against HAV. Significant inhibition of HAV was observed only when HAV was co-treated with extracts. Ten out of the 16 herbal extracts demonstrated significant virucidal activity against HAV. Alnus japonica extract at a concentration of 50 μg/mL reduced HAV titer by 3.43 ± 0.24 logs. Artemisia annua, Allium sativum, Allium fistulosum, and Agrimonia pilosa extracts showed 2.33 ± 0.43, 2.10 ± 0.41, 2.07 ± 0.60, and 2.03 ± 0.26-log reductions, respectively. Pleuropterus multiflorus, Eleutherococcus senticosus, Coriandrum sativum, Ginkgo biloba, and Torilis japonica extracts reduced HAV titer by 1.02 ± 0.21 to 1.90 ± 0.33 logs. Among the 10 herbal extracts, Alnus japonica extract was the most potent in inhibiting HAV without exhibiting cytotoxicity. [ABSTRACT FROM AUTHOR]

Abstract: Foodborne viruses such as norovirus (NoV), hepatitis A virus (HAV), hepatitis E virus (HEV), and rotavirus (RoV) are transmitted through water and food contaminated with stool. The purpose of this study was to examine the prevalence of foodborne viruses in shellfish collected in South Korea using real-time RT-PCR. Virus was eluted from the stomach and the digestive diverticula of 152 shellfish (51 oysters, 51 Manila clams, and 50 mussels) and concentrated with polyethylene glycol precipitation. The detection rate of NoV genogroup II, NoV genogroup I, HAV, HEV, and RoV was 21.7%, 5.9%, 0.7%, 0%, and 0% of shellfish, respectively. Although the geographic distribution of NoV was statistically significant, prominent seasonal variation in NoV was not observed in this study. In order to reduce norovirus food poisoning in the public, it is important to prevent the contamination of NoV in shellfish in South Korea. [Copyright &y& Elsevier]

The survival of murine norovirus (MNV) and hepatitis A virus (HAV) in bottled drinking water, strawberries, and oysters stored at different temperatures for various durations was investigated. In drinking water, over 6.4 and 1.6 log 10 plaque-forming units (PFU/mL) reductions of viable MNV and HAV were observed after 20 days of incubation at 35 °C. After 160 days of incubation at 20 °C, there was a sharp decrease in the MNV (>6.4 log 10 PFU/mL) and HAV (>2.9 log 10 PFU/mL) load but only a slight reduction at 4 °C. In fresh strawberry, the MNV and HAV titers reduced by 1.0 and 0.9 log 10 PFU/g after incubation for 2 days at 25 °C, by 0.85 and 1.22 log 10 PFU/g after 3 days of incubation at 15 °C, and by 0.7 and 1.1 log 10 PFU/g after 7 days of incubation at 4 °C, respectively. After 3 days of incubation at 4, 15, and 25 °C in oyster reductions of over 0.3, 0.4, and 0.6 log 10 PFU/mL in the MNV load and 0.16, 0.28, and 0.52 log 10 PFU/mL in the HAV load were observed, respectively. Even though, survival of the two viruses fluctuated with temperature and food commodity. Therefore, our results indicate that major enteric foodborne viruses such as MNV and HAV can survive for long periods, with only a partial reduction in their titers. Owing to the persistence of foodborne viruses, preventive measures need to be taken to prevent food contamination. • MNV and HAV survival in drinking water, strawberries, and oysters was investigated. • MNV and HAV survivability varies with temperature and food type. • Enteric foodborne viruses can persist for a long time at 4 °C. • Study results could lay the foundation for the development of food hazard assessment models. [ABSTRACT FROM AUTHOR]

Abstract: This study’s aim was to develop a specific and sensitive reverse transcription polymerase chain reaction enzyme-linked immunosorbent assay (RT-PCR-ELISA) for hepatitis A virus (HAV). Three concentration methods were compared using samples from cabbage, lettuce, and sesame leaves that were artificially contaminated with HAV. The specificity of the assay was tested against human norovirus, hepatitis E virus, rotavirus, enterovirus, and FCV. The sensitivity and specificity of RT-PCR-ELISA that were targeted to the 5′NCR and VP1 regions of HAV were compared. Nonspecific reactions were not observed. An optimal primer and probe set for RT-PCR-ELISA was selected for each VP1 and 5′NCR. The detection limit of the RT-PCR-ELISA was enhanced by 10–100 fold more than nested RT-PCR. Our new RT-PCR-ELISA was successfully optimized to screen vegetables for HAV contamination with high sensitivity in large samples. [Copyright &y& Elsevier]