多位点序列分型和脉冲场凝胶电泳技术在肠炎沙门菌暴发事件病原学鉴定中的应用
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摘要
目的探讨脉冲场凝胶电泳(pulse-field gel electrophoresis,PFGE)和多位点序列分型(multilocus sequence typing,MLST)技术在沙门菌鉴定中的应用价值。方法对分离自一起食物中毒事件中的27株沙门菌,用肠杆菌科鉴定系统API 20E和ATB ID 32E进行生化鉴定并血清分型,以脉冲场凝胶电泳(PFGE)和多位点序列分型(MLST)进行分子分型。结果 27株沙门菌的生化鉴定结果为沙门菌属细菌,其中25株血清分型结果为肠炎沙门菌,抗原模式(9,12:g,m);其余两株菌H相抗原除g和m因子凝集阳性外,d因子也有凝集反应。PFGE结果显示27株菌株中26株的带型完全一致,另1株与其他26株仅有1条条带的差异,相似度为96.3%。27株菌的MLST型别均为ST11型,提示为肠炎沙门菌。结论分子分型技术,特别是MLST技术有助于沙门菌的辅助鉴定。
Objective To assess the application of multi-locus sequence typing(MLST) and pulse-field electrophoresis(PFGE) in the identification of Salmonella.Methods Twenty-seven isolates collected from Salmonella foodborne disease outbreak were identified by API 20 E together with ID 32 E system for bacterial serotyping.PFGE and MLST were performed for molecular genotyping.Results All 27 isolates were classified as Salmonella genus by biochemistry,and the serotypes of 25 strains were found to be Salmonella enterica serovar enteritidis with the antigenic formula of 9,12:g,m,while,the other 2 isolates also showed an agglutination reaction of d factor in H antigen.PFGE revealed that 26 out of 27 stains had identical banding patterns,while the other one isolate only had 1 band difference with 96.3% similarity.All 27 strains were identified as ST11 by MLST,indicating of Salmonella enterica serovar enteritidis.Conclusions Molecular typing techniques especially MLST are helpful in the identification of Salmonella enterica.
Objective To assess the application of multi-locus sequence typing(MLST) and pulse-field electrophoresis(PFGE) in the identification of Salmonella.Methods Twenty-seven isolates collected from Salmonella foodborne disease outbreak were identified by API 20 E together with ID 32 E system for bacterial serotyping.PFGE and MLST were performed for molecular genotyping.Results All 27 isolates were classified as Salmonella genus by biochemistry,and the serotypes of 25 strains were found to be Salmonella enterica serovar enteritidis with the antigenic formula of 9,12:g,m,while,the other 2 isolates also showed an agglutination reaction of d factor in H antigen.PFGE revealed that 26 out of 27 stains had identical banding patterns,while the other one isolate only had 1 band difference with 96.3% similarity.All 27 strains were identified as ST11 by MLST,indicating of Salmonella enterica serovar enteritidis.Conclusions Molecular typing techniques especially MLST are helpful in the identification of Salmonella enterica.
引文
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[14]汪皓秋,潘劲草,郑伟,等.多位点序列分型和脉冲场凝胶电泳技术辅助鉴定肠炎沙门菌[J].中华预防医学杂志,2011,45(8):765-767.
[15]周国明,汪皓秋,俞骅,等.多位点序列分型技术在沙门菌鉴定中的应用研究[J].中国卫生检验杂志,2013,23(1):120-126.
[2]Swaminathan B,Gerner SP,Barrtt T.Focus on Salmonella[J].Foodborne Pathog Dis,2006,3(2):154-155.
[3]Scallan E,Hoekstra RM,Angulo FJ,et al.Foodborne Illness Acquired in the United States-Major Pathogens[J].Emerg Infect Dis,2011,17(1):7-15.
[4]朱超,许学斌.沙门菌属血清型诊断[M].上海:同济大学出版社,2009:1.
[5]Wattiau P,Boland C,Bertrand S.Methodologies for Salmonella enterica subsp.enterica subtyping:gold standards and alternatives.[J].Appl Environ Microbiol,2011,77(22):7877-7885.
[6]PulseNet USA.Standard operating procedure for PulseNet PFGE of Escherichia coli O157:H7,Escherichia coli nonO157(STEC),Salmonella serotypes,Shigella sonnei and Shigella flexneri.(April 2013)[EB/OL].[2014-07-01].http://www.cdc.gov/pulsenet/PDF/ecoli-shigella-salmonella-pfge-protocol-508c.pdf.
[7]Thong KL,Cordano AM,Yassin RM,et al.Molecular analysis of environmental and human isolates of Salmonella typhi[J].Appl Environ Microbiol,1996,62(1):271-274.
[8]Tenover FC,Arbeit RD,Goering RV,et al.Interpreting chromosomal DNA restriction patterns produced by pulsedfield gel electhophoresis:criteria for bacterial strain typing[J].Clin Microbiol,1995,33(9):2233-2239.
[9]Foley SL,Lynne AM,Nayak R.Molecular typing methodologies for microbial source tracking and epidemiological investigations of Gram-negative bacterial foodborne pathogens.[J].Infect Genet Evol,2009,9(4):430-40.
[10]娄静,刁保卫,李杰,等.沙门菌脉冲场凝胶电泳分型与血清型的对应关系.[J].中华流行病学杂志,2013,34(6):618-621.
[11]Maiden MC,Bygraves A,Feil E,et al.Multilocus sequence typing:A portable approach to the identification of clones within populations of pathogenic microorganisms[J].Proc Natl Acad Sci USA,1998,95(3):3140-3145.
[12]Maiden M.Multilocus sequence typing of bacteria[J].Annu Rev Microbiol,2006,60(1):561-588.
[13]Achtman M,Wain J,Weill FX,et al.Multilocus sequence typing as a replacement for serotyping in Salmonella enterica[J].PLoS Pathog,2012,8(6):e1002776.
[14]汪皓秋,潘劲草,郑伟,等.多位点序列分型和脉冲场凝胶电泳技术辅助鉴定肠炎沙门菌[J].中华预防医学杂志,2011,45(8):765-767.
[15]周国明,汪皓秋,俞骅,等.多位点序列分型技术在沙门菌鉴定中的应用研究[J].中国卫生检验杂志,2013,23(1):120-126.
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