生菜共生菌对诺如病毒灭活的影响
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摘要
目的了解生菜共生菌对诺如病毒灭活的影响及机理。方法从农田和超市采集生菜样品,分离鉴定共生菌;加共生菌的诺如病毒液为实验组,不加共生菌为对照组。采用高温、紫外线、含氯消毒剂灭活,分析实验组与对照组病毒量比值,评价共生菌对诺如病毒抗灭活能力的影响;通过检测共生菌对紫外线破坏病毒衣壳蛋白的影响和对病毒吸附能力的影响研究共生菌作用机理。结果共鉴定11株生菜共生菌,均为杆菌,以假单胞菌属为主。10株生菜共生菌能提高诺如病毒抗高温灭活能力,其中稻田叶螨微杆菌、台湾贪铜菌(SC061204)、古川假单胞菌、烟粉虱肠杆菌和食树脂假单胞菌(SC061211)较为显著;11株菌均能提高诺如病毒抗紫外线灭活能力,其中恶臭假单胞菌、稻田叶螨微杆菌和烟粉虱肠杆菌较为显著;恶臭假单胞菌能提高诺如病毒抗二氧化氯灭活能力,危害等级为Ⅰ级。恶臭假单胞菌、稻田叶螨微杆菌、烟粉虱肠杆菌能显著降低紫外线对诺如病毒衣壳蛋白的破坏作用。9株共生菌促进诺如病毒吸附,吸附促进率为1.04%～46.73%;恶臭假单胞菌和食树脂假单胞菌(SC061211)抑制诺如病毒吸附,吸附促进率分别为-6.50%和-19.85%。结论生菜共生菌可能通过保护诺如病毒衣壳蛋白和促进病毒吸附来提高诺如病毒抗灭活能力,在灭活诺如病毒的过程中应控制共生菌的存在。
Objective To demonstrate the effects of symbiotic bacteria from lettuce on inactivation of norovirus(NV). Methods Symbiotic bacteria were isolated from the lettuces sampled from farmlands and supermarkets. NV mixed with symbiotic bacteria was set as the experimental group,without symbiotic bacteria as the control group. After the inactivation by high temperature,ultraviolet light(UV) and chlorine dioxide, the ratio of NV amount in the experimental group and the control group was calculated to evaluate the effects of symbiotic bacteria. The mechanism of symbiotic bacteria was revealed by detecting their effects on the protection of viral capsid protein from UV and on the adsorption of NV. Results Eleven symbiotic bacteria were identified from lettuces,all of which were bacilli,mainly Pseudomonas. Ten symbiotic bacteria could improve the heat-resistant ability of NV,with Microbacterium oryzae,Cupriavidus taiwanensis(SC061204),Pseudomonas furukawaii,Enterobacter tabaci and Pseudomonas resinovorans(SC061211) more significant. Eleven symbiotic bacteria could improve anti-UV ability of NV,with Pseudomonas putida, Microbacterium oryzae and Enterobacter tabaci more significant. Only one strain of Pseudomonas putida could improve anti-chlorine dioxide ability of NV(Class I hazard). Pseudomonas putida, Microbacterium oryzae and Enterobacter tabaci could significantly reduce the damage of NV capsid protein. Nine symbiotic bacteria could promote NV adsorption into lettuces,with the promotion rates ranged from 1.04% to 46.73%;while Pseudomonas putida and Pseudomonas resinovorans(SC061211) could restrain NV absorption, with the promotion rates of-6.50% and-19.85%. Conclusion Symbiotic bacteria from lettuce may enhance the anti-inactivation of NV by protecting capsid protein and promoting adsorption of NV. It is recommended to control the presence of symbiotic bacteria in the process of inactivating NV.
Objective To demonstrate the effects of symbiotic bacteria from lettuce on inactivation of norovirus(NV). Methods Symbiotic bacteria were isolated from the lettuces sampled from farmlands and supermarkets. NV mixed with symbiotic bacteria was set as the experimental group,without symbiotic bacteria as the control group. After the inactivation by high temperature,ultraviolet light(UV) and chlorine dioxide, the ratio of NV amount in the experimental group and the control group was calculated to evaluate the effects of symbiotic bacteria. The mechanism of symbiotic bacteria was revealed by detecting their effects on the protection of viral capsid protein from UV and on the adsorption of NV. Results Eleven symbiotic bacteria were identified from lettuces,all of which were bacilli,mainly Pseudomonas. Ten symbiotic bacteria could improve the heat-resistant ability of NV,with Microbacterium oryzae,Cupriavidus taiwanensis(SC061204),Pseudomonas furukawaii,Enterobacter tabaci and Pseudomonas resinovorans(SC061211) more significant. Eleven symbiotic bacteria could improve anti-UV ability of NV,with Pseudomonas putida, Microbacterium oryzae and Enterobacter tabaci more significant. Only one strain of Pseudomonas putida could improve anti-chlorine dioxide ability of NV(Class I hazard). Pseudomonas putida, Microbacterium oryzae and Enterobacter tabaci could significantly reduce the damage of NV capsid protein. Nine symbiotic bacteria could promote NV adsorption into lettuces,with the promotion rates ranged from 1.04% to 46.73%;while Pseudomonas putida and Pseudomonas resinovorans(SC061211) could restrain NV absorption, with the promotion rates of-6.50% and-19.85%. Conclusion Symbiotic bacteria from lettuce may enhance the anti-inactivation of NV by protecting capsid protein and promoting adsorption of NV. It is recommended to control the presence of symbiotic bacteria in the process of inactivating NV.
引文
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[3]吴晨,吴昊澄,丁哲渊,等.浙江省学校突发公共卫生事件特征分析[J].预防医学,2018,30(1):16-21.
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[6]廖巧红,冉陆,靳淼,等.诺如病毒感染暴发调查和预防控制技术指南(2015版)[J].中国病毒病杂志, 2015, 5(6):448-458.
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[9]李楠,王佳慧,李凤琴,等.检测鲜草莓中GⅡ型诺如病毒的两种富集方法比较[J].中国食品卫生杂志,2015,27(3):242-246.
[10] RAJKO-NENOW P,KEAVENEY S,FLANNERY J,et al. Characterisation of norovirus contamination in an Irish shellfishery using real-time RT-qPCR and sequencing analysis[J]. International Journal of Food Microbiology,2012,160(2):105-112.
[11] DE ABREU C A,MIAGOSTOVICH M P. Optimization of an adsorption-elution method with a negatively charged membrane to recover norovirus from lettuce[J]. Food and Environmental Virology,2013,5(3):144-149.
[12] ALMAND E A, MOORE M D, OUTLAW J, et al. Human norovirus binding to select bacteria representative of the human gut microbiota[J]. PLo S One,2017,12(3):0173124.
[13] MULCAHY L R, ISABELLA V M, LEWIS K.Pseudomonas aeruginosabiofilms in disease[J]. Microbial Ecology, 2014, 68(1):1-12.
[14]谢朝云,熊芸,孙静,等. 91株恶臭假单胞菌流行病学特征及耐药性[J].中国感染控制杂志,2017,16(12):1185-1188.
[15] TAN M,HUANG P,MELLER J,et al. Mutations within the P2domain of norovirus capsid affect binding to human histo-blood group Antigens:evidence for a binding pocket[J]. Journal of Virology,2003,77(23):12562-12571.
[16] GAO X, ESSEILI M A, LU Z, et al. Recognition of histo-blood group antigen-like carbohydrates in lettuce by human GII.4 norovirus[J]. Applied and Environmental Microbiology, 2016, 82(10):2966-2974.
[17] LI D,BREIMAN A,LE PENDU J,et al. Binding to histo-blood group antigen-expressing bacteria protects human norovirus from acute heat stress[J]. Frontiers in Virology,2015,6:659.
[18] AMARASIRI M, HASHIBA S, MIURA T, et al. Bacterial histo-blood group antigens contributing to genotype-dependent removal of human noroviruses with a microfiltration membrane[J].Water Research,2016,95:383-391.
[19] YAMAOKA H,NAKAYAMA-IMAOHJI H,HORIUCHI I,et al.Tetramethylbenzidine method for monitoring the free available chlorine and microbicidal activity of chlorite-based sanitizers under organic-matter-rich environments[J]. Letters in Applied Microbiology,2016,62(1):47-54.
[20] LOPEZ-GALVEZ F,TRUCHADO P,S魣NCHEZ G,et al. Occurrence of enteric viruses in reclaimed and surface irrigation water:relationship with microbiological and physicochemical indicators[J].Journal of Applied Microbiology,2016,121(4):1180-1188.
[2]刘慧,马丽萍,周德庆.生鲜食品中诺如病毒污染与作用受体的研究进展[J].食品安全质量检测学报,2017,8(1):1-5.
[3]吴晨,吴昊澄,丁哲渊,等.浙江省学校突发公共卫生事件特征分析[J].预防医学,2018,30(1):16-21.
[4] JONES M K, WATANABE M, ZHU S, et al. Enteric bacteria promote human and mouse norovirus infection of B cells[J].Science,2014,346(6210):755-759.
[5] DENG W, ALMEIDA G, GIBSON K E. Co-culture with Enterobacter cloacae does not enhance virus resistance to thermal and chemical treatments[J]. Food and environmental virology,2019:1-9.
[6]廖巧红,冉陆,靳淼,等.诺如病毒感染暴发调查和预防控制技术指南(2015版)[J].中国病毒病杂志, 2015, 5(6):448-458.
[7] JACKSON C R,RANDOLPH K C,OSBORN S L,et al. Culture dependent and independent analysis of bacterial communities associated with commercial salad leaf vegetables[J]. BMC Microbiology,2013,13(1):1-12.
[8] ISO. Microbiology of food and animal feed-horizontal method for determination of hepatitis A virus and Norovirus in food using real-time RT-PCR. Part 1:method for quantification,ISO 15216-1:2017(E)[S]. Switzerland:International Organization for Standardization,2017:24-25.
[9]李楠,王佳慧,李凤琴,等.检测鲜草莓中GⅡ型诺如病毒的两种富集方法比较[J].中国食品卫生杂志,2015,27(3):242-246.
[10] RAJKO-NENOW P,KEAVENEY S,FLANNERY J,et al. Characterisation of norovirus contamination in an Irish shellfishery using real-time RT-qPCR and sequencing analysis[J]. International Journal of Food Microbiology,2012,160(2):105-112.
[11] DE ABREU C A,MIAGOSTOVICH M P. Optimization of an adsorption-elution method with a negatively charged membrane to recover norovirus from lettuce[J]. Food and Environmental Virology,2013,5(3):144-149.
[12] ALMAND E A, MOORE M D, OUTLAW J, et al. Human norovirus binding to select bacteria representative of the human gut microbiota[J]. PLo S One,2017,12(3):0173124.
[13] MULCAHY L R, ISABELLA V M, LEWIS K.Pseudomonas aeruginosabiofilms in disease[J]. Microbial Ecology, 2014, 68(1):1-12.
[14]谢朝云,熊芸,孙静,等. 91株恶臭假单胞菌流行病学特征及耐药性[J].中国感染控制杂志,2017,16(12):1185-1188.
[15] TAN M,HUANG P,MELLER J,et al. Mutations within the P2domain of norovirus capsid affect binding to human histo-blood group Antigens:evidence for a binding pocket[J]. Journal of Virology,2003,77(23):12562-12571.
[16] GAO X, ESSEILI M A, LU Z, et al. Recognition of histo-blood group antigen-like carbohydrates in lettuce by human GII.4 norovirus[J]. Applied and Environmental Microbiology, 2016, 82(10):2966-2974.
[17] LI D,BREIMAN A,LE PENDU J,et al. Binding to histo-blood group antigen-expressing bacteria protects human norovirus from acute heat stress[J]. Frontiers in Virology,2015,6:659.
[18] AMARASIRI M, HASHIBA S, MIURA T, et al. Bacterial histo-blood group antigens contributing to genotype-dependent removal of human noroviruses with a microfiltration membrane[J].Water Research,2016,95:383-391.
[19] YAMAOKA H,NAKAYAMA-IMAOHJI H,HORIUCHI I,et al.Tetramethylbenzidine method for monitoring the free available chlorine and microbicidal activity of chlorite-based sanitizers under organic-matter-rich environments[J]. Letters in Applied Microbiology,2016,62(1):47-54.
[20] LOPEZ-GALVEZ F,TRUCHADO P,S魣NCHEZ G,et al. Occurrence of enteric viruses in reclaimed and surface irrigation water:relationship with microbiological and physicochemical indicators[J].Journal of Applied Microbiology,2016,121(4):1180-1188.