肠炎沙门菌C50336ZinT基因缺失株构建及其相关生物学特性
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摘要
为研究锌调控蛋白ZinT在肠炎沙门菌致病中的作用,本试验利用λ-Red同源重组系统构建了肠炎沙门菌(C50336)ZinT基因缺失突变株(C50336ΔZinT),并对其生物学特性进行分析。结果显示,与野生型菌株和回复菌株相比,缺失株C50336ΔZinT在LB培养基中生长速度无明显差异,生化特性亦无明显变化,但其在Minimal培养基中生长明显变慢。此外缺失株C50336ΔZinT对H_2O_2处理和高渗环境的适应能力显著下降,但在巨噬细胞内的存活率无明显降低。动物试验表明,ZinT基因缺失突变株的毒力和野生型菌株相比仅有轻微下降。本试验探讨了ZinT与肠炎沙门菌锌摄取和毒力间的联系,为进一步阐释肠炎沙门菌感染致病机制奠定基础。
In this study,we examined the role of ZinT in Salmonella enteritidis caused infection with ZinT deleted and tested the biochemical characteristics.The data showed that,the ZinT-knockout did not affect the growth of C50336 strains in LB and change of biochemical characteristics were not observed in ZinT-deletion mutant.Compared to the wild type and complemented strains,the ZinT-deficient mutant displayed sharply impaired survival in minimal medium and significantly increased sensitivity to H_2O_2 and hypertonic treatment.The similar survival in macrophage and slightly attenuated virulence was observed.This study determined the close relationship of ZinT and Salmonella's survival under Zn~(2+)-deficient condition,and provided insight into the underlying mechanism of Salmonella caused infection.
In this study,we examined the role of ZinT in Salmonella enteritidis caused infection with ZinT deleted and tested the biochemical characteristics.The data showed that,the ZinT-knockout did not affect the growth of C50336 strains in LB and change of biochemical characteristics were not observed in ZinT-deletion mutant.Compared to the wild type and complemented strains,the ZinT-deficient mutant displayed sharply impaired survival in minimal medium and significantly increased sensitivity to H_2O_2 and hypertonic treatment.The similar survival in macrophage and slightly attenuated virulence was observed.This study determined the close relationship of ZinT and Salmonella's survival under Zn~(2+)-deficient condition,and provided insight into the underlying mechanism of Salmonella caused infection.
引文
[1] OH J H,PARK M K.Recent Trends in Salmonella outbreaks and emerging technology for biocontrol of Salmonella using phages in foods:a review[J].J Microbiol Biotechnol,2017,27(12):2075-2088.
[2] 杨保伟,张秀丽,曲东,等.2007—2008陕西部分零售畜禽肉沙门菌血清型和基因型[J].微生物学报,2010(5):654-660.
[3] RUBY T,MCLAUGHLIN L,GOPINATH S,et al.Salmonella long-term relationship with its host[J].FEMS Microbiol Rev,2012,36(3):600-615.
[4] LIM J,LEE K M,KIM S H,et al.YkgM and ZinT proteins are required for maintaining intracellular zinc concentration and producing curli in enterohemorrhagic Escherichia coli (EHEC) O157:H7 under zinc deficient conditions[J].Int J Food Microbiol,2011,149(2):159-170.
[5] GABBIANELLI R,SCOTTI R,AMMENDOLA S,et al.Role of ZnuABC and ZinT in Escherichia coli O157:H7 zinc acquisition and interaction with epithelial cells[J].BMC Microbiol,2011,11:36.
[6] ILARI A,ALALEONA F,TRIA G,et al.The Salmonella enterica ZinT structure,zinc affinity and interaction with the high-affinity uptake protein ZnuA provide insight into the management of periplasmic zinc[J].Biochim Biophys Acta,2014,1840(1):535-544.
[7] PETRARCA P,AMMENDOLA S,PASQUALI P,et al.The Zur-regulated ZinT protein is an auxiliary component of the high-affinity ZnuABC zinc transporter that facilitates metal recruitment during severe zinc shortage[J].J Bacteriol,2010,192(6):1553-1564.
[8] LIU Y,ZHANG Q,HU M,et al.Proteomic analyses of intracellular Salmonella enterica Serovar Typhimurium reveal extensive bacterial adaptations to infected host epithelial cells[J].Infect Immun,2015,83(7):2897-2906.
[9] DATSENKO K A,WANNER B L.One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products[J].Proc Natl Acad Sci USA,2000,97(12):6640-6645.
[10] 张志强,李永慧,吴同垒,等.致泥鳅点状出血征温和气单胞菌的分类鉴定[J].中国兽医学报,2017,37(6):1070-1076.
[11] CHI F,WANG Y,GALLAHER T K,et al.Identification of IbeR as a stationary-phase regulator in meningitic Escherichia coli K1 that carries a loss-of- function mutation in rpoS[J].J Biomed Biotechnol,2009,2009:520283.
[12] DARFEUILLE-MICHAUD A,BOUDEAU J,BULOIS P,et al.High prevalence of adherent-invasive Escherichia coli associated with ileal mucosa in Crohn-s disease[J].Gastroenterology,2004,127(2):412-421.
[13] KURTZ J R,GOGGINS J A,MCLACHLAN J B.Salmonella infection:Interplay between the bacteria and host immune system[J].Immunol Lett,2017,190:42-50.
[14] FU J,QI L,HU M,et al.Salmonella proteomics under oxidative stress reveals coordinated regulation of antioxidant defense with iron metabolism and bacterial virulence[J].J Proteomics,2017,157:52-58.
[15] MONACK D M.Salmonella persistence and transmission strategies[J].Curr Opin Microbiol,2012,15(1):100-107.
[16] GRADASSI M,PESCIAROLI M,MARTINELLI N,et al.Attenuated Salmonella enterica serovar Typhimurium lacking the ZnuABC transporter:an efficacious orally-administered mucosal vaccine against salmonellosis in pigs[J].Vaccine,2013,31(36):3695-3701.
[17] BOBROV A G,KIRILLINA O,FOSSO M Y,et al.Zinc transporters YbtX and ZnuABC are required for the virulence of Yersinia pestis in bubonic and pneumonic plague in mice[J].Metallomics,2017,9(6):757-772.
[18] FEIGL G,LEHOTAI N,MOLNAR A,et al.Zinc induces distinct changes in the metabolism of reactive oxygen and nitrogen species (ROS and RNS) in the roots of two Brassica species with different sensitivity to zinc stress[J].Ann Bot,2015,116(4):613-625.
[2] 杨保伟,张秀丽,曲东,等.2007—2008陕西部分零售畜禽肉沙门菌血清型和基因型[J].微生物学报,2010(5):654-660.
[3] RUBY T,MCLAUGHLIN L,GOPINATH S,et al.Salmonella long-term relationship with its host[J].FEMS Microbiol Rev,2012,36(3):600-615.
[4] LIM J,LEE K M,KIM S H,et al.YkgM and ZinT proteins are required for maintaining intracellular zinc concentration and producing curli in enterohemorrhagic Escherichia coli (EHEC) O157:H7 under zinc deficient conditions[J].Int J Food Microbiol,2011,149(2):159-170.
[5] GABBIANELLI R,SCOTTI R,AMMENDOLA S,et al.Role of ZnuABC and ZinT in Escherichia coli O157:H7 zinc acquisition and interaction with epithelial cells[J].BMC Microbiol,2011,11:36.
[6] ILARI A,ALALEONA F,TRIA G,et al.The Salmonella enterica ZinT structure,zinc affinity and interaction with the high-affinity uptake protein ZnuA provide insight into the management of periplasmic zinc[J].Biochim Biophys Acta,2014,1840(1):535-544.
[7] PETRARCA P,AMMENDOLA S,PASQUALI P,et al.The Zur-regulated ZinT protein is an auxiliary component of the high-affinity ZnuABC zinc transporter that facilitates metal recruitment during severe zinc shortage[J].J Bacteriol,2010,192(6):1553-1564.
[8] LIU Y,ZHANG Q,HU M,et al.Proteomic analyses of intracellular Salmonella enterica Serovar Typhimurium reveal extensive bacterial adaptations to infected host epithelial cells[J].Infect Immun,2015,83(7):2897-2906.
[9] DATSENKO K A,WANNER B L.One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products[J].Proc Natl Acad Sci USA,2000,97(12):6640-6645.
[10] 张志强,李永慧,吴同垒,等.致泥鳅点状出血征温和气单胞菌的分类鉴定[J].中国兽医学报,2017,37(6):1070-1076.
[11] CHI F,WANG Y,GALLAHER T K,et al.Identification of IbeR as a stationary-phase regulator in meningitic Escherichia coli K1 that carries a loss-of- function mutation in rpoS[J].J Biomed Biotechnol,2009,2009:520283.
[12] DARFEUILLE-MICHAUD A,BOUDEAU J,BULOIS P,et al.High prevalence of adherent-invasive Escherichia coli associated with ileal mucosa in Crohn-s disease[J].Gastroenterology,2004,127(2):412-421.
[13] KURTZ J R,GOGGINS J A,MCLACHLAN J B.Salmonella infection:Interplay between the bacteria and host immune system[J].Immunol Lett,2017,190:42-50.
[14] FU J,QI L,HU M,et al.Salmonella proteomics under oxidative stress reveals coordinated regulation of antioxidant defense with iron metabolism and bacterial virulence[J].J Proteomics,2017,157:52-58.
[15] MONACK D M.Salmonella persistence and transmission strategies[J].Curr Opin Microbiol,2012,15(1):100-107.
[16] GRADASSI M,PESCIAROLI M,MARTINELLI N,et al.Attenuated Salmonella enterica serovar Typhimurium lacking the ZnuABC transporter:an efficacious orally-administered mucosal vaccine against salmonellosis in pigs[J].Vaccine,2013,31(36):3695-3701.
[17] BOBROV A G,KIRILLINA O,FOSSO M Y,et al.Zinc transporters YbtX and ZnuABC are required for the virulence of Yersinia pestis in bubonic and pneumonic plague in mice[J].Metallomics,2017,9(6):757-772.
[18] FEIGL G,LEHOTAI N,MOLNAR A,et al.Zinc induces distinct changes in the metabolism of reactive oxygen and nitrogen species (ROS and RNS) in the roots of two Brassica species with different sensitivity to zinc stress[J].Ann Bot,2015,116(4):613-625.
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