冷诱导金黄色葡萄球菌环境胁迫耐受性及生物被膜控制
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摘要
为研究金黄色葡萄球菌经冷诱导后对其他环境胁迫耐受性的变化,将耐冷冻能力较强的分离株S.aureus W3与标准菌株S.aureus CICC10201分别在4℃下冷胁迫5 h后,测定其在高温、酸碱、氯化钠、过氧化氢、酒精的胁迫下存活率变化。同时对分离株S.aureus W3的生物被膜形成及控制进行了研究。结果表明,经-20℃冷藏8周后,S.aureus W3的存活率显著高于S.aureus CICC10201 (p <0.05)。经冷诱导后,S.aureus W3在45～60℃环境下存活率提升至18.5%。冷诱导使金黄色葡萄球菌对酸敏感性增强但对碱产生交叉抗性。冷诱导后的两株菌在氯化钠中的存活率均降低,S.aureus W3最低下降了24.09%。另外,冷诱导后金黄色葡萄球菌对酒精的耐受性增强。分离株S.aureus W3在冷诱导前后过氧化氢耐受性没有明显变化(p> 0.05),但标准菌株S.aureus CICC10201敏感性增强,存活率显著下降(p <0.05)。分离株S.aureus W3较S.aureus CICC10201具有更强的生物被膜形成能力,运用1%84消毒水能有效清除其生物被膜。本研究为冷冻食品中金黄色葡萄球菌的安全控制提供理论依据。
In order to study the effect of cold induction on other environmental stress tolerance of Staphylococcus aureus,the S.aureus W3 which had strong resistance to freezing and standard strain S.aureus CICC10201 were measured at high temperatures,pH,NaCl,hydrogen peroxide and alcohol after 5 h of cold stress at 4 ℃.Meanwhile,the biofilm formation ability and control method were studied.The results showed that after 8 weeks of storage at-20 ℃,the survival rate of S.aureus W3 was significantly higher than that of S.aureus CICC10201 (p < 0.05). After cold induction,the survival rates of S. aureus W3 increased by 18.5% at 45 ～60 ℃. Cold induction enhanced the sensitivity of S. aureus to acidic environment,but it produced cross resistance to alkaline environment. After cold induction,the survival rate of two strains in hypertonic environment decreased,and the lowest decreased by 24.09%.In addition,the tolerance of S.aureus to alcohol increased after cold induction.For hydrogen peroxide,there was no significant (p > 0.05) change in the tolerance of S.aureus W3 before and after induction.But the sensitivity of the S.aureus CICC10201 was enhanced after cold induction,and the survival rate decreased.S.aureus W3 has stronger biofilm formation ability than S.aureus CICC10201,and 1% 84 disinfection can effectively remove its biofilm. This study provides a theoretical basis for the safety control of S.aureus in frozen foods.
In order to study the effect of cold induction on other environmental stress tolerance of Staphylococcus aureus,the S.aureus W3 which had strong resistance to freezing and standard strain S.aureus CICC10201 were measured at high temperatures,pH,NaCl,hydrogen peroxide and alcohol after 5 h of cold stress at 4 ℃.Meanwhile,the biofilm formation ability and control method were studied.The results showed that after 8 weeks of storage at-20 ℃,the survival rate of S.aureus W3 was significantly higher than that of S.aureus CICC10201 (p < 0.05). After cold induction,the survival rates of S. aureus W3 increased by 18.5% at 45 ～60 ℃. Cold induction enhanced the sensitivity of S. aureus to acidic environment,but it produced cross resistance to alkaline environment. After cold induction,the survival rate of two strains in hypertonic environment decreased,and the lowest decreased by 24.09%.In addition,the tolerance of S.aureus to alcohol increased after cold induction.For hydrogen peroxide,there was no significant (p > 0.05) change in the tolerance of S.aureus W3 before and after induction.But the sensitivity of the S.aureus CICC10201 was enhanced after cold induction,and the survival rate decreased.S.aureus W3 has stronger biofilm formation ability than S.aureus CICC10201,and 1% 84 disinfection can effectively remove its biofilm. This study provides a theoretical basis for the safety control of S.aureus in frozen foods.
引文
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[29]Vasudevan P,Nair M K,Annamalai T,et al.Phenotypic and genotypic characterization of bovine mastitis isolates of Staphylococcus aureus for biofilm formation[J].Veterinary Microbiology,2003,92(1-2):179-183.
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[2]崔莹,张秀丽,胡巅,等.速冻食品中金黄色葡萄球菌及肠毒素污染状况调查[J].中国卫生检验杂志,2013(3):764-766.
[3]王晓杰.冷冻胁迫下金黄色葡萄球菌亚致死及失活规律研究[D].郑州:河南农业大学,2013.
[4]王晓杰,王娜,宋金丽,等.冷冻胁迫下金黄色葡萄球菌的亚致死及失活规律[J].食品科学,2013,34(19):5-9.
[5]索标,居冉,王娜,等.冷冻胁迫对金黄色葡萄球菌亚致死及clf A,msr R毒力基因转录的影响[J].中国食品学报,2015,15(11):154-159.
[6]Duval B D,Mathew A,Satola SW,et al.Altered growth,pigmentation,and antimicrobial susceptibility properties of Staphylococcus aureus due to loss of the major cold shock gene csp B[J].Antimicrobial Agents and Chemotherapy,2010,54(6):2283-2290.
[7]Onyango L A,Dunstan R H,Gottfries J,et al.Effect of low temperature on growth and ultra-structure of Staphylococcus spp[J].Plos One,2012,7(1):29031-20037.
[8]Alreshidi M M,Dunstan R H,Macdonald M M,et al.Metabolomic and proteomic responses of Staphylococcus aureus,to prolonged cold stress[J].Journal of Proteomics,2015,121:44-55.
[9]Shah J,Desai P T,Chen D,et al.Preadaptation to cold stress in Salmonella enterica serovar Typhimurium increases survival during subsequent acid stress exposure.[J].Applied&Environmental Microbiology,2013,79(23):7281-7283.
[10]Cebrián G,Sagarzazu N,Pagán R,et al.Development of stress resistance in Staphylococcus aureus after exposure to sublethal environmental conditions[J].International Journal of Food Microbiology,2010,140(1):26-33.
[11]Singh M.,Mullins H.R.,Simpson,et al.Effect of acid adaptation on thermal tolerance of Escherichia coli O157∶H7 and Salmonella enterica in meat serum[J].Journal of Food Safety,2010,30(1):111-123.
[12]Doulgeraki A I,Ciccio P D,Ianieri A,et al.Methicillinresistant food-related Staphylococcus aureus:A review of current knowledge and biofilm formation for future studies and applications[J].Research in Microbiology,2016,168(1):1-5.
[13]梁晏瑞.金黄色葡萄球菌生物被膜形成特性及抑制清除方法研究[D].华南理工大学,2016.
[14]Stepanovic S,Vukovic D,Dakic I,et al.A modified microtiter-plate test for quantification of staphylococcal biofilm formation[J].J Microbiol Methods,2000,40(2):175-179.
[15]Liebeke M,Lalk M.Staphylococcus aureus metabolic response to changing environmental conditions-A metabolomics perspective[J].International Journal of Medical Microbiology,2014,304(3-4):222-227.
[16]Juneja V K,Klein P G,Marmer B S.Heat shock and thermotolerance of Escherichia coli O157∶H7 in a model beef gravy system and ground beef[J].Journal of Applied Microbiology,1998,84(4):677-684.
[17]Mcmahon C M M,Byrne C M,Sheridan J J,et al.The effect of culture growth phase on induction of the heat shock response in Yersinia enterocolitica and Listeria monocytogenes[J].Journal of Applied Microbiology,2000,89(2):198-206.
[18]Shenoy K,Murano E A.Effect of Heat Shock on the thermotolerance and protein composition of Yersinia enterocolitica in brain heart Infusion broth and ground pork[J].Journal of Food Protection,1996,59(4):360-364.
[19]Lindquist S.The heat-shock response[J].Annu Rev Biochem,1985,55(1):1151-1191.
[20]Laury A M,Alvarado M V,Nace G,et al.Validation of a lactic acid-and citric acid-based antimicrobial product for the reduction of Escherichia coli O157∶H7 and Salmonella on beef tips and whole chicken carcasses[J].Journal of Food Protection,2009,72(10):2208-2211.
[21]Park S H,Choi M R,Park J W,et al.Use of organic acids to inactivate Escherichia coli O157∶H7,Salmonella Typhimurium,and Listeria monocytogenes on organic fresh apples and lettuce[J].Journal of Food Science,2011,76(6):293-298.
[22]Olaimat A N,Al-Nabulsi A A,Osaili T M,et al.Survival and inhibition of Staphylococcus aureus,in commercial and hydrated tahini using acetic and citric acids[J].Food Control,2017,77:179-186.
[23]PanéfarréJ.,et al.The sigma B regulon in Staphylococcus aureus and its regulation[J].International Journal of Medical Microbiology,2006,296(4):237-258.
[24]陈俊,张惠萍,朱希贵,等.金黄色葡萄球菌耐盐生长的形态观察及机理的初步研究[J].云南农业大学学报(自然科学),2010,25(4):517-520.
[25]Kuroda M,Tanaka Y,Aoki R,et al.Staphylococcus aureus giant protein Ebh is involved in tolerance to transient hyperosmotic pressure[J].Biochemical&Biophysical Research Communications,2008,374(2):237-241.
[26]Chiang M L,Chen H C,Wu C,et al.Effect of acid adaptation on the environmental stress tolerance of three strains of Vibrio parahaemolyticus[J].Foodborne Pathogens&Disease,2014,11(4):287.
[27]Horsburgh M J,Clements M O,Crossley H,et al.Per Rcontrols oxidative stress resistance and iron storage proteins and is required for virulence in Staphylococcus aureus[J].Infection&Immunity,2001,69(6):3744-3450.
[28]Rodrigues D,Cerca N,Teixeira P,et al.Listeria monocytogenes and Salmonella enterica enteritidis biofilms susceptibility to different disinfectants and stress-response and virulence gene expression of surviving cells.[J].Microbial Drug Resistance,2011,17(2):181-189.
[29]Vasudevan P,Nair M K,Annamalai T,et al.Phenotypic and genotypic characterization of bovine mastitis isolates of Staphylococcus aureus for biofilm formation[J].Veterinary Microbiology,2003,92(1-2):179-183.
[30]Abel M.Biofilm formation and dispersal of Staphylococcus aureus under the influence of oxacillin.[J].Microbial Pathogenesis,2013,62(4):66-72.