The effect of cranberry juice and cranberry proanthocyanidins on the infectivity of human enteric viral surrogates

The effect of cranberry juice (CJ) and cranberry proanthocyanidins (PAC) on the infectivity of human enteric virus surrogates, murine norovirus (MNV-1), feline calicivirus (FCV-F9), MS2(ssRNA) bacteriophage, and phiX-174(ssDNA) bacteriophage was studied. Viruses at high (∼7 log₁₀ PFU/ml) or low (∼5 log₁₀ PFU/ml) titers were mixed with equal volumes of CJ, 0.30, 0.60, and 1.20 mg/ml final PAC concentration, or water and incubated for 1 h at room temperature. Viral infectivity after treatments was evaluated using standardized plaque assays. At low viral titers, FCV-F9 was undetectable after exposure to CJ or the three tested PAC solutions. MNV-1 was reduced by 2.06 log₁₀ PFU/ml with CJ, and 2.63, 2.75, and 2.95 log₁₀ PFU/ml with 0.15, 0.30, and 0.60 mg/ml PAC, respectively. MS2 titers were reduced by 1.14 log₁₀ PFU/ml with CJ, and 0.55, 0.80, and 0.96 log₁₀ PFU/ml with 0.15, 0.30, and 0.60 mg/ml PAC, respectively. ?-X174 titers were reduced by 1.79 log₁₀ PFU/ml with CJ, and 1.95, 3.67, and 4.98 log₁₀ PFU/ml with PAC at 0.15, 0.30, and 0.60 mg/ml, respectively. Experiments using high titers showed similar trends but with decreased effects. CJ and PAC show promise as natural anti-virals that could potentially be exploited for foodborne viral illness treatment and prevention.     

Antiviral effects of cranberry juice and cranberry proanthocyanidins on foodborne viral surrogates – A time dependence study in vitro

Cranberry juice (CJ) and cranberry proanthocyanidins (PAC) are widely known for their antibacterial, antiviral, and pharmacological activities. The effect of CJ and cranberry PAC on the infectivity of foodborne viral surrogates, murine norovirus (MNV-1), feline calicivirus (FCV-F9), MS2 (ssRNA) bacteriophage, and ?X-174 (ssDNA) bacteriophage after 0 min to 1 h at room temperature was evaluated. Viruses at titers of ∼5 log₁₀PFU/ml were mixed with equal volumes of CJ at pH 2.6, CJ at pH 7.0, 0.30 mg/ml CJ PAC, 0.60 mg/ml PAC, or water and incubated for 0, 10, 20, 30, 40, 50 min, and 1 h at room temperature. Infectivity was determined using standard plaque assays. The viral reduction rates of the four tested viruses were found to vary considerably. Among the tested viruses, FCV-F9 titers were decreased the most by ∼5 log₁₀PFU/ml within 30 min. MS2 titers were decreased the least by only ∼1 log₁₀PFU/ml after 1 h with CJ at pH 2.6 and 0.30 mg/ml PAC, and ∼0.5 log₁₀PFU/ml with CJ at pH 7.0 and 0.15 mg/ml PAC. MNV-1 and ?-X174 showed comparable titer reductions which was between that of FCV-F9 and MS2. In most cases, viral reduction within the first 10 min of treatment accounted for ≥50% of the total reduction. Transmission electron microscopy on FCV-F9 treated with CJ and PAC revealed structural changes. This study shows potential of using natural bioactive compounds for controlling foodborne viral diseases. Further studies are necessary to elucidate the mechanism of action of CJ components and to understand the differences in viral titer reduction profiles.     

Grape seed extract for foodborne virus reduction on produce

Grape seed extract (GSE) is reported to have antibacterial properties with few current studies on antiviral activity. Recently, we reported the effects of GSE against foodborne viral surrogates in vitro. This study evaluated the application of GSE (commercial Gravinol-S) against hepatitis A virus (HAV) and human norovirus surrogates, feline calicivirus (FCV-F9) and murine norovirus (MNV-1), on model produce. Washed and air-dried lettuce (3 × 3 cm²) and jalapeno peppers (25–30 g) were inoculated with FCV-F9, MNV-1, or HAV at high (∼7 log₁₀ PFU/ml) or low (∼5 log₁₀ PFU/ml) titers, and treated with 0.25, 0.5, 1 mg/ml GSE or water for 30 s to 5 min. Treatments were stopped/diluted with cell-culture media containing 10% heat-inactivated fetal bovine serum and evaluated using plaque assays. At high titers, FCV-F9 was reduced by 2.33, 2.58, and 2.71 log₁₀ PFU on lettuce; and 2.20, 2.74, and 3.05 log₁₀ PFU on peppers after 1 min using 0.25, 0.50, and 1 mg/ml GSE, respectively. Low FCV-F9 titers could not be detected after 1 min at all three GSE concentrations. Low titer MNV-1 was reduced by 0.2–0.3 log₁₀ PFU on lettuce and 0.8 log₁₀ PFU on peppers, without reduction of high titer. GSE at 0.25–1 mg/ml after 1 min caused 0.7–1.1 and 1–1.3 log₁₀ PFU reduction for high and low HAV titers, respectively on both commodities. Instrumental color analysis showed no significant differences between treated and untreated produce. GSE shows potential for foodborne viral reduction on produce as part of hurdle technologies.     

Comparison of reduction in foodborne viral surrogates by high pressure homogenization

With the increasing global spread of human noroviral infections and the emergence of highly virulent noroviral strains, novel inactivation methods are needed to control foodborne outbreaks. High pressure homogenization (HPH) is a novel method that can be applied for foodborne virus reduction in fluids being continuously processed. Our objective in the present study was to compare the titer reduction by HPH between feline calicivirus strain F9 (FCV-F9) and murine norovirus 1 (MNV-1) as surrogates for human noroviruses, and MS2 (single-stranded F-RNA coliphage) and somatic coliphage φX174 (single-stranded DNA) as indicators of fecal contamination. Duplicate experiments with each virus in phosphate-buffered saline were carried out with homogenization pressures of 0, 100, 200, 250, and 300 MPa, with exposure temperatures of 24, 46, 63, 70, and 75°C, respectively, for <2 s. FCV-F9 was found highly susceptible to HPH treatment pressures of 300 MPa, with a reduction of >4.95 log PFU/ml. Lower pressures of 250, 200, and 100 MPa resulted in reductions of 1.61, 0.60, and 0.18 log PFU/ml of FCV-F9, respectively, while MNV-1 was not reduced at these lower pressures. Coliphage φX174 showed no significant reduction at 300 MPa or lower homogenization pressures in comparison with MS2, which did show 3.3-log PFU/ml reduction at 300 MPa. Future studies using juices for industrial application of HPH to determine microbial inactivation with simultaneous retention of sensory and nutritional value of foods are needed.     

Survival of human norovirus surrogates in milk, orange, and pomegranate juice, and juice blends at refrigeration (4 °C)

Fresh fruits, juices, and beverages have been implicated in human noroviral and hepatitis A virus outbreaks. The purpose of this study was to determine the survival of human norovirus surrogates (murine norovirus, MNV-1; feline calicivirus, FCV-F9; and bacteriophage MS2) in juices (orange and pomegranate juices), juice blends (pomegranate and orange juice) and milk over 0, 1, 2, 7, 14, and 21 days at refrigeration (4 °C). Juices, juice blends, and milk were inoculated with each virus over 21 days, serially diluted in cell culture media, and plaque assayed. MNV-1 showed no reduction in titer after 21 days in orange juice and milk, but moderate reduction (1.4 log) in pomegranate juice from a titer of 5 log(10) PFU/ml. However, MNV-1 was completely reduced after 7 days in the orange and pomegranate juice blend. FCV-F9 from a titer of 6 log(10) PFU/ml was completely reduced after 14 days in orange as well as pomegranate juice and by ～ 3 logs after 21 days in milk at 4 °C. Interestingly, FCV-F9 was completely reduced after 1 day in the orange and pomegranate juice blend at 4 °C. MS2 was reduced by ～ 1.28 log after 21 days in orange juice from a titer of 6 log(10) PFU/ml, and <1 log after 21 days in milk or pomegranate juice, with juice blends showing minimal reduction (<1 log) after 21 days at 4 °C. These results show the survival pattern of noroviruses that aid in the transmission of foodborne viral outbreaks. The data obtained can be used in quantitative viral risk assessment studies and to develop improved measures to prevent virus survival towards controlling outbreaks.     

Efficacy of a levulinic Acid plus sodium dodecyl sulfate-based sanitizer on inactivation of human norovirus surrogates

Human noroviruses are the most common etiologic agent of foodborne illness in the United States. The inability to culture human noroviruses in the laboratory necessitates the use of surrogate viruses such as murine norovirus (MNV-1) and feline calicivirus (FCV) for inactivation studies. In this study, a novel sanitizer of organic acid (levulinic acid) plus the anionic detergent sodium dodecyl sulfate (SDS) was evaluated. Viruses were treated with levulinic acid (0.5 to 5%), SDS (0.05 to 2%), or combinations of levulinic acid plus SDS (1:10 solution of virus to sanitizer). MNV-1 inoculated onto stainless steel also was treated with a 5% levulinic acid plus 2% SDS liquid or foaming solution. Log reductions of viruses were determined with a plaque assay. Neither levulinic acid nor SDS alone were capable of inactivating MNV-1 or FCV, resulting in a ≤0.51-log reduction of the infectious virus titer. However, the combination of 0.5% levulinic acid plus 0.5% SDS inactivated both surrogates by 3 to 4.21 log PFU/ml after 1 min of exposure. Similarly, MNV-1 inoculated onto stainless steel was reduced by >1.50 log PFU/ml after 1 min and by >3.3 log PFU/ml after 5 min of exposure to a liquid or foaming solution of 5% levulinic acid plus 2% SDS. The presence of organic matter (up to 10%) in the virus inoculum did not significantly affect sanitizer efficacy. The fact that both of the active sanitizer ingredients are generally recognized as safe to use as food additives by the U.S. Food and Drug Administration further extends its potential in mitigating foodborne disease.     

Comparison of hydrostatic and hydrodynamic pressure to inactivate foodborne viruses

The effect of high hydrostatic pressure (HPP) and hydrodynamic pressure (HDP), in combination with chemical treatments, was evaluated for inactivation of foodborne viruses and non-pathogenic surrogates in a pork sausage product. Sausages were immersed in distilled water, 100-ppm EDTA, or 2% lactoferrin, and then inoculated with feline calicivirus (FCV), hepatitis A virus (HAV) or bacteriophage (MS2, phiX174, or T₄). Each piece was packaged individually and subjected to pressure by either HDP, HPP (500 MPa, 5 min, 4 °C), or control (no pressure). On sausages immersed in water, HPP and HDP significantly (P < 0.05) reduced titers of FCV by 2.89 and 2.70 log₁₀ TCID₅₀/ml, and HAV by log₁₀ 3.23 and 1.10, respectively, when compared to non-pressure-treated controls. Titers of T₄ (1.48 and 1.10 log₁₀ PFU/g) and MS2 (1.46 and 0.96 log₁₀ PFU/g) were also significantly reduced by HPP and HDP treatments, respectively, in combination with water. Inoculation of viruses and bacteriophage on a meat product may have protected viruses from complete inactivation by pressure treatments.

Industrial relevance

This is the first study to directly compare hydrostatic and hydrodynamic pressure technologies to inactivate microorganisms. This is also the first study to examine the inactivation of viruses and bacteriophages by pressure technology in a deli meat product. This study shows that viruses attached to meat surfaces may be protected from complete inactivation by hydrostatic and hydrodynamic pressure treatments, and these findings require more investigation into the survival of viruses in deli meat products.     

Inactivation of hepatitis A virus and norovirus surrogate in suspension and on food-contact surfaces using pulsed UV light (pulsed light inactivation of food-borne viruses)

This study was conducted to evaluate the inactivation of murine norovirus (MNV-1) and hepatitis A virus (HAV) by pulsed ultraviolet (UV) light. MNV-1 was used as a model for human norovirus. Viral suspensions of about 10⁶ PFU/ml were exposed to pulses of UV light for different times and at different distances in a Xenon Steripulse device (model RS-3000C). Inactivation studies were also carried out on 1-cm² stainless steel and polyvinyl chloride disks with 10⁵ PFU/ml. Inactivation of MNV-1 and HAV at 10.5 cm from the UV source was greater on inert surfaces than in suspension. The presence of organic matter (fetal bovine serum) reduced the effectiveness of pulsed light both in suspension and on surfaces. However, 2-s treatment in the absence of FBS completely inactivated (5 log reduction) the viral load at different distances tested, whether in suspension (MNV-1) or on disks (MNV-1 and HAV). The same treatment in the presence of fetal bovine serum (5%) allowed a reduction of about 3 log. This study showed that short duration pulses represent an excellent alternative for inactivation of food-borne viruses. This technology could be used to inactivate viruses in drinking water or on food-handling surfaces.     

Comparison of the murine norovirus-1 inactivation in cabbage Kimchi with two different salinities during storage

Enteric noroviruses are occasionally detected in Kimchi, which is a traditional dish made of fermented vegetables. This study was aimed at examining the effects of two levels of salt concentrations on the survival of murine norovirus-1 (MNV-1), a human norovirus surrogate, in experimentally contaminated cabbage Kimchi stored at 5 °C for 10 weeks. The number of total aerobic bacteria (TAB) and lactic acid bacteria (LAB), MNV-1 titer pH, and acidity were measured every week. The titers of MNV-1 in both low (1.17%) and normal (2.22%) salinity cabbage Kimchi were significantly (P < 0.05) decreased with increase in storage time. The overall reduction was 1.75 log₁₀ plaque-forming unit (PFU)/mL in normal salinity cabbage Kimchi and 1.24 log₁₀ PFU/mL in low salinity cabbage Kimchi. The time required to reduce the titer by > 1 log₁₀ PFU/mL in normal and low salinity cabbage Kimchi were 4 and 8 weeks, respectively. The pH value under both salinities significantly (P < 0.05) decreased until 4 weeks. The maximum acidity was 0.83% and 0.79% in normal and low salinity cabbage Kimchi, respectively, during the 10 weeks. The population of TAB and LAB reached up to 7.33 log₁₀ colony-forming unit (CFU)/g as a maximum population during the storage period of 3 weeks in normal salinity cabbage Kimchi. However, the population of TAB and LAB in low salinity cabbage Kimchi reached to 6.99 and 7.04 log₁₀ CFU/g at 5 and 4 weeks, respectively. Through these findings, fermentation factors such as TAB, LAB, pH, and acidity of cabbage Kimchi were influenced by salt concentration. The inactivation of MNV-1 in normal salinity cabbage Kimchi was much faster than that in low salinity cabbage Kimchi because the fermentation in normal salinity cabbage Kimchi progressed more quickly than that in low salinity cabbage Kimchi. However, both salinity cabbage Kimchi were able to infect cells for 70 days even though the MNV-1 was reduced over 1 log₁₀ during fermentation. Therefore, the way to protect cabbage Kimchi from norovirus must be considered.     

High hydrostatic pressure processing of murine norovirus 1-contaminated oysters inhibits oral infection in STAT-1(-/-)-deficient female mice

We have previously demonstrated that high pressure processing (HPP) is effective in preventing in vitro replication of murine norovirus strain 1 (MNV-1), a human norovirus surrogate, in a monocyte cell line following extraction from MNV-1-contaminated oysters. In the present study, the efficacy of HPP to prevent in vivo replication within mice fed HPP-treated MNV-1-seeded oyster extracts was evaluated. Oyster homogenate extracts seeded with MNV-1 were given 5-min, 400-MPa (58,016-psi) treatments and orally gavaged into immunodeficient (STAT-1(-/-)) female mice. Mice orally gavaged with HPP-treated MNV-1 showed significant (P ≤ 0.05) weight loss leading to enhanced morbidity, whereas those given 100 and 200 PFU of HPP-treated MNV-1 were comparable to uninfected controls. MNV-1 was detected, via real-time PCR, within the liver, spleen, and brain of all mice fed non-HPP-treated homogenate but was not detected in the tissues of mice fed HPP-treated homogenates or in uninfected control mice. Hepatocellular necrosis and lymphoid depletion in the spleen were observed in non-HPP-treated MNV-1 mice only. These results clearly show that HPP prevents MNV-1 infection in vivo and validates that viral inactivation by HPP in vitro is essentially equivalent to that in vivo. Further, the data suggest that HPP may be an effective food processing intervention for norovirus-contaminated shellfish and thus may decrease risk to both immunocompromised and immunocompetent individuals who consume shellfish.     

Removal of bacteriophages MS2 and phiX174 during transport in a sandy anoxic aquifer

The objectives of our study were to determine (i) removal of bacteriophage MS2 and phiX174, as surrogates for human pathogenic viruses, in an anoxic aquifer and (ii) the safe length of the microbial protection zone in anoxic aquifers. 3.5 Log units of MS2 were removed by adsorption and inactivation during 63 days residence time, which was 1.4 log units lower than removal of phiX174. These removal rates were considerably lower than previously reported for MS2 and phiX174 in oxic aquifers and consequently longer protection zones around anoxic aquifers might be needed. Therefore, the observed log removal of MS2 was used in a risk assessment approach to determine the required safe length of the microbial protection zone. In case of a leaking sewer in the vicinity of a well in an anoxic aquifer, the risk assessment demonstrated that a microbial protection zone of 110 m may be needed to meet a risk of infection of 10(-4) persons per year. This length can be two to three times larger than the length of the protection zone currently used in a number of countries.     

A combined treatment of UV-assisted TiO2 photocatalysis and high hydrostatic pressure to inactivate internalized murine norovirus

Human norovirus (HuNoV) is a major cause of foodborne illness associated with shellfish consumption. A solidified agar matrix (SAM) was experimentally prepared using agar solution for inactivation of murine norovirus (MNV-1) as a surrogate for HuNoV in a simulation model approach. MNV-1 was injected inside the SAM for virus internalization, and the effects of single and combined UV-assisted TiO₂ photocatalysis (UVTP) and high hydrostatic pressure (HHP) treatments were determined. The internalized MNV-1 were reduced by 2.9-log₁₀ and 3.5-log₁₀, respectively, after single treatments of UVTP (4.5 mW/cm², 10 min) and HHP (500 MPa, 5 min, ambient temperature). However, the internalized MNV-1 was reduced by 5.5-log₁₀ (below the detection limit) when UVTP was followed by HHP, indicating a synergistic inactivation effect. Analysis of viral morphology, proteins, and genomic RNA allowed elucidation of mechanisms involved in the synergistic antiviral activity of combined treatments, which appeared to disrupt the MNV-1 structure and damage both the capsid protein and genomic RNA.Industrial relevanceHHP treatment of raw oysters has proved commercially successful, but there is a less evidence available regarding the potential of HHP for inactivation of localized viruses present inside foods. A sequential combination of UV-assisted TiO₂ photocatalysis (UVTP) and high hydrostatic pressure (HHP) achieved significantly higher inactivation of localized virus compared to individual treatments due to a synergistic mechanism. An experimentally prepared model food system was found useful to simulate foods with morphological variations and unpredictable viral internalization patterns. This UVTP-HHP combined treatment for inactivation of localized MNV-1 can be useful for disinfection of raw oysters and other similar foods.     

Electron-beam inactivation of a norovirus surrogate in fresh produce and model systems

Norovirus remains the leading cause of foodborne illness, but there is no effective intervention to eliminate viral contaminants in fresh produce. Murine norovirus 1 (MNV-1) was inoculated in either 100 ml of liquid or 100 g of food. The inactivation of MNV-1 by electron-beam (e-beam), or high-energy electrons, at varying doses was measured in model systems (phosphate-buffered saline [PBS], Dulbecco's modified Eagle's medium [DMEM]) or from fresh foods (shredded cabbage, diced strawberries). E-beam was applied at a current of 1.5 mA, with doses of 0, 2, 4, 6, 8, 10, and 12 kGy. The surviving viral titer was determined by plaque assays in RAW 264.7 cells. In PBS and DMEM, e-beam at 0 and 2 kGy provided less than a 1-log reduction of virus. At doses of 4, 6, 8, 10, and 12 kGy, viral inactivation in PBS ranged from 2.37 to 6.40 log, while in DMEM inactivation ranged from 1.40 to 3.59 log. Irradiation of inoculated cabbage showed up to a 1-log reduction at 4 kGy, and less than a 3-log reduction at 12 kGy. On strawberries, less than a 1-log reduction occurred at doses up to 6 kGy, with a maximum reduction of 2.21 log at 12 kGy. These results suggest that a food matrix might provide increased survival for viruses. In foods, noroviruses are difficult to inactivate because of the protective effect of the food matrix, their small sizes, and their highly stable viral capsid.     

Resistance of phages lytic to pathogenic Escherichia coli to sanitisers used by the food industry and in home settings

Phages are potentially useful as antimicrobial agents in food‐processing environments, provided they can remain active upon extended storage and in the presence of sanitisers. Survival of six phages lytic against enteropathogenic (EPEC) and shiga‐toxigenic (STEC) Escherichia coli strains was assessed upon storage at 4 °C, ?20 °C and ?70 °C in phosphate‐buffered‐saline (PBS) and Tris‐magnesium‐gelatine buffer (TMG) for up to 1 year. The phages were also exposed to ethanol, sodium hypochlorite, peracetic acid, quaternary ammonium chloride (biocide A), hydrogen peroxide/peracetic acid/peroctanoic acid (biocide B), p‐toluensulfonchloroamide (biocide D) and alkaline chloride foam (biocide C). Plaque‐forming units remained unchanged when the phages were stored at 4 °C in both buffers tested, but decreased by 3.5 and 5.7 log₁₀ PFU when stored in PBS at ?20 and ?70 °C, respectively. Moreover, TMG seems to be the most protective suspension medium with decreasing temperature because a 1?log₁₀ PFU reduction was observed at ?20 and ?70 °C. Incubation in 100% ethanol for 24 h reduced plaque counts only by 2.5 log₁₀ PFU. In 10 ppm of sodium hypochlorite and in biocide B (0.13%), the counts decreased by ~5 and ~6 log₁₀ PFU after 15 min. Finally, biocide A and D reduced counts by 4 and 2–4 log₁₀ PFU after 30 min and 8 h of incubation, respectively. Phages were completely inactivated only by peracetic acid and biocides C and E. Therefore, the phages evaluated might be potentially applied together with classical sanitisers such as ethanol and industrial biocides A, B and D, in disinfection programs against pathogenic STEC and EPEC strains.     

Surrogates for the study of norovirus stability and inactivation in the environment: aA comparison of murine norovirus and feline calicivirus

Human noroviruses (NoVs) are the leading cause of food- and waterborne outbreaks of acute nonbacterial gastroenteritis worldwide. As a result of the lack of a mammalian cell culture model for these viruses, studies on persistence, inactivation, and transmission have been limited to cultivable viruses, including feline calicivirus (FCV). Recently, reports of the successful cell culture of murine norovirus 1 (MNV-1) have provided investigators with an alternative surrogate for human NoVs. In this study, we compared the inactivation profiles of MNV-1 to FCV in an effort to establish the relevance of MNV-1 as a surrogate virus. Specifically, we evaluated (i) stability upon exposure to pH extremes; (ii) stability upon exposure to organic solvents; (iii) thermal inactivation; and (iv) surface persistence under wet and dry conditions. MNV-1 was stable across the entire pH range tested (pH 2 to 10) with less than 1 log reduction in infectivity at pH 2, whereas FCV was inactivated rapidly at pH values < 3 and > 9. FCV was more stable than MNV-1 at 56 degrees C, but both viruses exhibited similar inactivation at 63 and 72 degrees C. Long-term persistence of both viruses suspended in a fecal matrix and inoculated onto stainless steel coupons were similar at 4 degrees C, but at room temperature in solution, MNV-1 was more stable than FCV. The genetic relatedness of MNV-1 to human NoVs combined with its ability to survive under gastric pH levels makes this virus a promising and relevant surrogate for studying environmental survival of human NoVs.     

Comparison of chlorine and peroxyacetic-based disinfectant to inactivate Feline calicivirus, Murine norovirus and Hepatitis A virus on lettuce

In recent years, raw fruits and vegetables have frequently been involved in foodborne transmission to humans of enteric viruses, particularly noroviruses and hepatitis A virus (HAV). Although viral contamination can occur during all steps of food processing, primary production is a critical stage on which prevention measures must be focused to minimize the risk of infection to consumers. Postharvest sanitation may be a valid technological solution for decreasing the bacterial load on fresh raw material, but there is a lack of data concerning the effectiveness of this process on enteric viruses. In this study, we compared the survival of two human norovirus surrogates, the feline calicivirus (FCV), and the murine norovirus (MNV-1), and of HAV on lettuce after water washing with bubbles and with or without ultrasound, and washing with bubbles in the presence of active chlorine (15 ppm) or peroxyacetic acid-based disinfectant (100 ppm). Cell culture and quantitative RT-PCR assays were used to detect and quantify the viruses on the surface of the lettuce after the sanitizing treatments. Levels of viral inactivation on the lettuce leaves were not significantly different between washing with bubbles and washing with bubbles plus ultrasound and were not dependant on the quantification method. A simple washing without disinfectant resulted in a decrease of approximately 0.7 log units in the quantity of virus detected for HAV and FCV and of 1.0 log unit for MNV-1.In the experimental set-up including a washing step (with or without ultrasound) followed by washing for 2 min in the presence of disinfectants, 15 ppm of active chlorine was found more effective for inactivating FCV (2.9 log units) than HAV and MNV-1 (1.9 log units and 1.4 log units, respectively) whereas 100 ppm of peroxyacetic-based biocide was found effective for inactivating FCV (3.2 log units) and MNV-1 (2.3 log units), but not HAV (0.7 log units). Quantitative RT-PCR results indicated that the presence of viral RNA did not correlate with the presence of infectious viruses on disinfected lettuce, except for MNV-1 processed with chlorine (15 ppm). In comparison with water washing, a substantial additional decrease of genomic FCV titer (1.1 log units) but no significant reduction of the genomic titers of HAV and MNV-1 were found on lettuce treated with chlorine (15 ppm). No significant effect of the disinfection step of lettuce with peroxyacetic-based biocide (100 ppm peracetic acid) was found by qRT-PCR on all genomic viral titers tested. This study illustrates the necessity of determining the effectiveness of technological processes against enteric viruses, using a relevant reference such as HAV, in order to reduce the risk of hepatitis and gastroenteritis by exposure to vegetables.     

Efficacy of cold atmospheric plasma for inactivation of viruses on raspberries

In this study, the effectiveness of cold atmospheric plasma (CAP) in inactivating murine norovirus (MNV/human norovirus surrogate) and hepatitis A virus (HAV) on aerosol-inoculated dark red Willamette raspberries was explored. Pulsed positive corona discharge system fed by synthetic air was used for the production of CAP. Raspberries were treated for 1, 3, 5, 7, and 10 min at 25 W. Application of CAP enabled a 4 log₁₀ infectivity reduction in <5 min for MNV and approximately 10 min for HAV (from starting level of 6.91 and 7.84 log₁₀ PFU/mL, respectively). Viral genome copies reduction of 3.18 log₁₀ for MNV and 4.32 for HAV were found from starting level of 5.76 and 6.47 log₁₀ gc/μL, respectively. CAP treatment did not result in significant degradation of fruit color, an important quality attribute. The study demonstrated CAP as an efficient post-harvest decontamination method to reduce viral load in raspberries without significantly affecting its quality parameters.Industrial relevanceDue to the fast-processing paces required in the raspberry industry, it is difficult to assure the complete microbiological safety of this fruit. Cold atmospheric plasma is a practical, environmentally-friendly, non-thermal tool for the effective reduction of microbial pathogens. The model developed in this study demonstrated that CAP treatment of fresh raspberries not only inactivated hazardous enteric viruses in a short time (10 min) but also unaffected fruit color stability. The simplicity of described CAP design and low-cost inputs (air and electricity) enable the commercial application of inexpensive plasma chambers for continuous surface decontamination of large volumes of raspberries without bringing processing to a standstill.     

Antiviral effect of multipurpose contact lens disinfecting solutions against coronavirus

PurposeTo evaluate the antiviral potential of five multipurpose disinfecting solutions against coronavirus (mouse hepatitis virus, a surrogate for SARS-CoV-2 human corona virus).MethodsTest solutions (Biotrue, renu Advanced [Bausch and Lomb], ACUVUE RevitaLens [Johnson and Johnson Vision], cleadew [Ophtecs corp.] or AOSept Plus [Alcon]) were mixed with the coronavirus mouse hepatitis virus at 10⁴ plaque forming units (PFU)/mL as the final concentration and incubated at room temperature for the specified disinfection time. Surviving virus from each sample was then quantified by standard plaque forming unit assay and the reduction of PFU for each disinfectant was compared to the phosphate buffer saline (PBS) treated negative control. A regimen test was also conducted using Biotrue.ResultsThe three multipurpose disinfecting solutions Biotrue (containing PHMB and polyquaternium-1), renu Advanced (PHMB, polyquaternium-1 and alexidine) and ACUVUE RevitaLens (polyquaternium-1 and alexidine) did not kill the coronavirus at the manufacturers recommended disinfection time in the stand alone test. After treatment, the virus’s titer (3.8 ± 0.2 log₁₀ for Biotrue, 3.7 ± 0.1 log₁₀ for renu and 3.7 ± 0.2 log₁₀ for RevitaLens) was similar to the negative control (3.7 ± 0.1 log₁₀; p ≥ 0.996). AOSept Plus (hydrogen peroxide) and cleadew (povidone iodine) significantly (p < 0.001) reduced the numbers of coronaviruses to below the detection limit (i.e. killed 3.7 ± 0.1 log₁₀ viruses compared to control). However, there was a significant reduction (p = 0.028) in numbers of coronaviruses attached to lenses when using the regimen test with Biotrue.ConclusionsThis study shows that oxidative contact lens disinfecting solutions (i.e. those containing povidone-iodine or hydrogen peroxide) provide superior antiviral activity against a coronavirus surrogate of SARS-CoV-2, unless the full regimen test (rub, rinse, disinfect) is used.