Virucidal effect of high power ultrasound combined with a chemical sanitizer containing peroxyacetic acid for water reconditioning in the fresh-cut industry

The use of chlorine-based sanitizers is widespread throughout the fresh produce industry in the process water to maintain microbial safety of produce, avoid cross-contamination and recycle water. In this study, alternative disinfection technologies were investigated due to the negative reports showing chlorine instability in the presence of organic matter and the undesirable by-product residues. Tsunami^® (15.2% peroxyacetic acid and 11.2% hydrogen peroxide) and high-power ultrasound (HPU, 0.56 kW/l, 20 kHz), alone or in combination, were evaluated in lettuce process water analyzing their ability to reduce MNV during either produce washing or water recycling. PAA concentrations of 6.4, 12.8 and 16 mg/l as well as HPU were insufficient for application in the produce washing tank where a rapid MNV inactivation is required. In contrast, a complete rapid inactivation of MNV (7.4 log TCID₅₀) was recorded when 80 mg/l of PAA was used. For recycled water reconditioning, HPU was applied and no virus reduction was observed. Two PAA concentrations (6.4 and 12.8 mg/l) that can be found in the recycled water were also tested. The MNV inactivation was faster at 12.8 mg/l than at 6.4 mg/l of PAA with 2.6 and 2 log reductions after 20 min contact. One additional log reduction was achieved 20 and 40 min later, respectively. MNV inactivation fitted the Bipashic model well. A contact time of 95 and 115 min would be required to obtain the complete elimination of MNV (4.4 log TCID₅₀) by 6.4 and 12.8 mg/l of PAA. PAA efficacy was not enhanced by the combination with HPU in contrast to the inactivation of pathogenic bacteria tested in other studies. This indicates that viruses are more resistant to this hurdle technology than pathogenic bacteria such as Escherichia coli O157:H7, Salmonella and other HPU conditions such as higher frequencies and combination of HPU with other physical treatments need to be explored.     

上海市诺如病毒感染性腹泻疾病负担的评估

目的对上海市诺如病毒感染性腹泻的疾病负担进行综合评估,为管理部门制定预防和控制诺如病毒感染的措施提供依据。方法结合上海市哨点医院诺如病毒感染性腹泻监测和诺如病毒感染暴发事件报告,采用伤残调整寿命年(DALYs)、发病率、就诊率和住院率等指标对上海市诺如病毒感染性腹泻的疾病负担进行评估。结果 2015年上海市感染性腹泻发病人数为483.05万人,其中诺如病毒感染性腹泻的发病人数为123.23万人,住院病例为370人,所产生的疾病负担为2 938.98 DALYs,即0.12 DALYs/千人。结论上海市诺如病毒感染性腹泻的疾病负担相对较高,需要加强预防控制。     

Norovirus and MS2 inactivation kinetics of UV-A and UV-B with and without TiO2

Germicidal ultraviolet, such as 254-nm UV-C, is a common method of disinfection of pathogenic enteric viruses. However, the disinfection efficacies of UV-A or -B in terms of inactivating waterborne viruses such as norovirus have not been characterized. We evaluated the inactivation kinetics of MS2 bacteriophage and murine norovirus (MNV), a surrogate of human norovirus (NoV), by UV-A and -B. In addition to UV disinfection, we further investigated whether the presence of TiO₂ could enhance the virus inactivation kinetics of UV-A and -B. Both MS2 and MNV were highly resistant to UV-A. However, the addition of TiO₂ enhanced the efficacy of UV-A for inactivating these viruses. UV-A dose of 1379 mJ/cm² resulted in a 4 log₁₀ reduction. In comparison, UV-B alone effectively inactivated both MS2 and MNV, as evidenced by the 4 log₁₀ reduction by 367 mJ/cm² of UV-B. The addition of TiO₂ increased the inactivation of MS2; however, it did not significantly increase the efficacy of UV-B disinfection for inactivating MNV. When these treatments were applied to field water such as groundwater, the results were generally consistent with the laboratory findings. Our results clearly indicated that UV-B is useful for the disinfection of waterborne norovirus. However, MNV was quite resistant to UV-A, and UV-A effectively inactivated the tested viruses only when used in combination with TiO₂.     

Discharge-based QMRA for estimation of public health risks from exposure to stormwater-borne pathogens in recreational waters in the United States

This study is the first to report a quantitative microbial risk assessment (QMRA) on pathogens detected in stormwater discharges-of-concern, rather than relying on pathogen measurements in receiving waters. The pathogen concentrations include seven “Reference Pathogens” identified by the U.S. EPA: Cryptosporidium, Giardia, Salmonella, Norovirus, Rotavirus, Enterovirus, and Adenovirus. Data were collected from 12 sites representative of seven discharge types (including residential, commercial/industrial runoff, agricultural runoff, combined sewer overflows, and forested land), mainly during wet weather conditions during which times human health risks can be substantially elevated. The risks calculated herein therefore generally apply to short-term conditions (during and just after rainfall events) and so the results can be used by water managers to potentially inform the public, even for waters that comply with current criteria (based as they are on a 30-day mean risk). Using an example waterbody and mixed source, pathogen concentrations were used in QMRA models to generate risk profiles for primary and secondary water contact (or inhalation) by adults and children. A number of critical assumptions and considerations around the QMRA analysis are highlighted, particularly the harmonization of the pathogen concentrations measured in discharges during this project with those measured (using different methods) during the published dose–response clinical trials. Norovirus was the most dominant predicted health risk, though further research on its dose–response for illness (cf. infection) is needed. Even if the example mixed-source concentrations of pathogens had been reduced 30 times (by inactivation and mixing), the predicted swimming-associated illness rates – largely driven by Norovirus infections – can still be appreciable. Rotavirus generally induced the second-highest incidence of risk among the tested pathogens while risks for the other Reference Pathogens (Giardia, Cryptosporidium, Adenovirus, Enterovirus and Salmonella) were considerably lower. Secondary contact or inhalation resulted in considerable reductions in risk compared to primary contact. Measurements of Norovirus and careful incorporation of its concentrations into risk models (harmonization) should be a critical consideration for future QMRA efforts. The discharge-based QMRA approach presented herein is particularly relevant to cases where pathogens cannot be reliably detected in receiving waters with detection limits relevant to human health effects.     

Norovirus contamination on French marketed oysters

Contaminated shellfish have been implicated in gastroenteritis outbreaks in different countries. As no regulation has been set up yet regarding viral contamination of food, very few data are available on the prevalence of contaminated products on the market. This study presents data obtained from oysters collected on the French market in one producing area over a 16 month period of time. Noroviruses were detected in 9% of samples with a seasonal impact and influence of climatic events. Contamination levels were low and, surprisingly, oysters sampled directly from the producer were found to have less contamination than oysters from supermarkets.     

水中诺如病毒富集效果及核酸提取方法的比较

目的比较诺如病毒检测时3种核酸提取方法的效率,并对水中诺如病毒的检测方法进行初步研究。方法采用Trizol法、硅胶柱法及磁珠法分别提取含有诺如病毒的粪便样本的核酸,Real-time PCR检测其Ct值,比较3种方法提取核酸的效率。将含有诺如病毒的粪便加入纯水中,采用MCE-PEG法富集后,Real-time PCR检测其Ct值,评价MCE-PEG法富集病毒的效果。结果硅胶柱法的Ct平均值最小,在3种方法中提取诺如病毒RNA效率最高,磁珠法提取效果次之,Trizol法效果最差;MCE-PEG法富集病毒效果良好。结论磁珠法提取诺如病毒RNA效率虽不是最高,但可实现自动化,节省工作时间,降低了人力成本,是3种方法中基层实验室提取诺如病毒RNA最适合的一种;MCE-PEG法可以用来富集水中的诺如病毒。     

北京市市售扇贝中诺如病毒监测分析

目的监测北京市市售扇贝中诺如病毒和A组轮状病毒污染状况,分析其基因特征。方法 2014年11月至2015年10月,采集北京市3个水产品出售市场新鲜扇贝72份。取扇贝消化组织,用聚乙二醇(PEG)8000沉淀法进行病毒富集后提取核酸,用实时荧光逆转录-聚合酶链式反应(RT-PCR)法检测诺如病毒和A组轮状病毒核酸。用半巢式RT-PCR方法扩增诺如病毒GⅠ/GⅡ组衣壳蛋白区基因,对PCR产物进行测序,用BioEdit 7.0.9.0软件进行序列比对,用MEGA 6.06软件构建进化树。结果 72份扇贝样品均未检出A组轮状病毒;诺如病毒检出率为27.8%(20/72),其中GⅡ组16份,GⅠ组2份,GⅠ和GⅡ组混合2份。冬季检出率最高,为61.1%(11/18);夏季未检出。8份诺如病毒核酸阳性样品测序结果为GⅡ.17型,属于GⅡ.17型ClusterⅢb分枝。有6株GⅡ.17型毒株与2015年中国人源诺如病毒、2016和2017年日本人源诺如病毒、2017和2018年韩国水中诺如病毒以及2015年日本牡蛎中诺如病毒序列相似性为100.0%。结论北京市市售扇贝中存在诺如病毒污染,食用扇贝有引起病毒性急性胃肠炎的风险。     

2015~2016年河北省贝类水产品中诺如病毒污染状况研究

目的 对2015~2016年河北省6个设区市的市售牡蛎、海虹、血蚶样品中的诺如病毒进行连续一年的检测, 了解河北省贝类水产品中诺如病毒污染状况。方法 分离贝类水产品消化腺, 用蛋白酶K消化后进行前处理, 提取RNA, 用一步法实时荧光逆转录聚合酶链式反应法检测GⅠ型和GⅡ型诺如病毒。对河北省贝类水产品的诺如病毒污染状况进行分析。结果 共采集691份贝类样品进行检测, 其中仅检出GⅠ型阳性样品16份, 阳性率2.32%; 仅检出GⅡ型阳性样品22份, 阳性率3.18%; 同时检出GⅠ和GⅡ型的阳性样品4份, 阳性率0.58%; 总阳性率6.08%。2016年6~7月阳性率最高。沿海地区样品阳性率为内陆地区的2.35倍, 有统计学差异(χ2=8.58, P<0.01)。结论 河北省市售贝类水产品中部分存在诺如病毒污染, 需要加强对贝类水产品的诺如病毒污染检测和控制, 特别是沿海地区, 降低诺如病毒引起的食源性腹泻的疾病负担。     

GI/GII型诺如病毒两联装甲RNA标准样品的研制

针对目前缺乏适配多项检测标准、稳定、安全的诺如病毒RNA标准样品的问题,研制基于MS2噬菌体内含常见GI/GII型诺如病毒检测靶标两联装甲RNA标准参考样品。人工合成MS2噬菌体成熟酶基因、衣壳蛋白基因、包装位点及GI/GII型诺如病毒靶标基因,克隆于表达载体pET-28a(+)中,构建重组质粒pET-MS2-NoV。经大肠杆菌BL21诱导表达,先后利用PEG6000、酶处理和丙烯葡聚糖凝胶层析柱纯化表达产物。SDS-PAGE和透射电镜鉴定产物大小及结构,荧光定量PCR检测有无残留核酸。之后对纯化的病毒样颗粒（Virus-like particles,VLPs）开展定值、均匀性和短期稳定性研究。SDS-PAGE结果表明重组质粒在BL21中表达出了目的蛋白,大小在10~15 ku之间,与预期一致;纯化后的VLPs无杂蛋白和残留核酸;透射电镜下呈结构完整、大小均一的球状,直径约25 nm。纯化后AR-NoV中GI型和GII型靶标定值结果分别为(4.04±0.62)×107 copies/μL和(6.16±0.30)×107 copies/μL。单因素方差检验证实样品均一性良好,F相似文献