Objective To establish and compare the pulsed-field gel electrophoresis （PFGE）, multiple-locus variable number tandem repeat analysis （MLVA） and automated ribotyping for subtyping of Citrobacter strains. Methods P...Objective To establish and compare the pulsed-field gel electrophoresis （PFGE）, multiple-locus variable number tandem repeat analysis （MLVA） and automated ribotyping for subtyping of Citrobacter strains. Methods PFGE protocol was optimized in terms of plug preparation procedure, restriction enzymes and configuration of electrophoretic parameters. MLVA method was evaluated by finding variable number tandem repeats in two genomes of Citrobacter strains. The ribotyping was performed by using the automated RiboPrinter system. Results We optimized the plug preparation procedure, focused on the cell suspension concentration （turbidity of 2.5 to 3.5）, SDS addition （no SDS needed） and lysis time （1 h）, and selected the appropriate restriction enzyme （Xbal） and the electrophoretic parameters （1.0 s-20.0 s for 19 h） of PFGE. There was nearly no discriminatory power of MLVA between Citrobacter strains. For 51 Citrobacter strains, automated ribotyping gave a D-value of 0.9945, while PFGE gave a D-value of 0.9969. Both PFGE and automated ribotyping clustered strains from the same sources （with the same species from the same place at the same time identified as the same source） and divided strains from different sources （from different years, places and hosts） into different subtypes. Conclusion PFGE protocol established in this paper and automated ribotyping are suitable for application in Citrobacter subtyping.展开更多
目的了解辽宁地区炭疽芽胞杆菌的流行特征及菌型基因特征。方法通过多位点可变数目串联重复序列(Multiple locus variable numbers of tandem repeats analysis,MLVA)分型实验,对2001—2011年辽宁省分离到的6株炭疽芽胞杆菌分离株DNA进...目的了解辽宁地区炭疽芽胞杆菌的流行特征及菌型基因特征。方法通过多位点可变数目串联重复序列(Multiple locus variable numbers of tandem repeats analysis,MLVA)分型实验,对2001—2011年辽宁省分离到的6株炭疽芽胞杆菌分离株DNA进行检测,DNA指纹图谱使用BioNumerics 4.0软件进行统计分析,得出聚类分析结果。结果聚类分析发现,6株炭疽芽胞杆菌株可分为2个基因型。对于炭疽暴发而言,其可变数目串联重复序列遗传标记具有高度相似性。结论炭疽芽胞杆菌基因组中的串联重复序列可作为炭疽芽胞杆菌基因分型的指标,在炭疽暴发事件中的病原体溯源上具有重要的意义。展开更多
目的建立针对食品来源的单核细胞增生李斯特氏菌(Listeria monocytogenes,Lm)分离株的多位点串联重复序列分型(Multiple-Locus Variable number tandem repeat Analysis,MLVA)方法,为暴发确认和溯源检测提供实验室支持。方法对2005—201...目的建立针对食品来源的单核细胞增生李斯特氏菌(Listeria monocytogenes,Lm)分离株的多位点串联重复序列分型(Multiple-Locus Variable number tandem repeat Analysis,MLVA)方法,为暴发确认和溯源检测提供实验室支持。方法对2005—2014年间分离自食品的91株Lm进行14个可变数目串联重复序列(Variable Number of Tandem Repeats,VNTR)位点的检测,评估最优检测位点组合并分析检测结果。结果通过采用软件分析,由LMV1、LMV2、LMV7、Lm10、Lm11、Lm23、LM-TR6、TR3和Lm15等9个VNTR位点组成的位点组合为最优MLVA检测位点,可以将91株Lm分离株分为70个型别,分型能力达到0.987 1。结论本研究建立的基于全自动毛细管电泳的由9个检测位点组成的Lm的MLVA分型方法,具有操作简便、快速、结果客观、操作标准化、易于在不同实验室间比较的优势,可作为一线检测方法用于李斯特菌病的暴发确认和溯源检测。展开更多
基金supported by the project (grant 2005CB522904 and 2005CB522905) from the Ministry of Scientific Technologythe project (grant 2008ZX10004-001, 2008ZX10004-008, and 2009ZX10004-101) from the Ministry of Scientific Technology and the Ministry of Health of the People’s Republic of China
文摘Objective To establish and compare the pulsed-field gel electrophoresis （PFGE）, multiple-locus variable number tandem repeat analysis （MLVA） and automated ribotyping for subtyping of Citrobacter strains. Methods PFGE protocol was optimized in terms of plug preparation procedure, restriction enzymes and configuration of electrophoretic parameters. MLVA method was evaluated by finding variable number tandem repeats in two genomes of Citrobacter strains. The ribotyping was performed by using the automated RiboPrinter system. Results We optimized the plug preparation procedure, focused on the cell suspension concentration （turbidity of 2.5 to 3.5）, SDS addition （no SDS needed） and lysis time （1 h）, and selected the appropriate restriction enzyme （Xbal） and the electrophoretic parameters （1.0 s-20.0 s for 19 h） of PFGE. There was nearly no discriminatory power of MLVA between Citrobacter strains. For 51 Citrobacter strains, automated ribotyping gave a D-value of 0.9945, while PFGE gave a D-value of 0.9969. Both PFGE and automated ribotyping clustered strains from the same sources （with the same species from the same place at the same time identified as the same source） and divided strains from different sources （from different years, places and hosts） into different subtypes. Conclusion PFGE protocol established in this paper and automated ribotyping are suitable for application in Citrobacter subtyping.
文摘目的了解辽宁地区炭疽芽胞杆菌的流行特征及菌型基因特征。方法通过多位点可变数目串联重复序列(Multiple locus variable numbers of tandem repeats analysis,MLVA)分型实验,对2001—2011年辽宁省分离到的6株炭疽芽胞杆菌分离株DNA进行检测,DNA指纹图谱使用BioNumerics 4.0软件进行统计分析,得出聚类分析结果。结果聚类分析发现,6株炭疽芽胞杆菌株可分为2个基因型。对于炭疽暴发而言,其可变数目串联重复序列遗传标记具有高度相似性。结论炭疽芽胞杆菌基因组中的串联重复序列可作为炭疽芽胞杆菌基因分型的指标,在炭疽暴发事件中的病原体溯源上具有重要的意义。
文摘目的掌握江苏省2018年人感染布鲁氏菌主要流行株的种型和基因型。方法运用普通PCR及AMOS多重PCR确认分离株的生物种型;采用多位点序列分型(Multilocus sequence analysis,MLSA)和多位点串联重复序列分析(Multiple-locus variable number tandem repeat analysis,MLVA)鉴定基因型,并与国内外流行株进行聚类分析。结果2018年共分离到56株布鲁氏菌,MLSA分型显示一株菌为猪种布鲁氏菌ST(Sequence type)17型,其他均为羊种布鲁氏菌ST8型。MLVA将56株菌分为47个基因亚型(46个羊种,1个猪种),聚类显示羊种布鲁氏菌全部为“东地中海簇”。结论2018年江苏省人感染布病主要为“东地中海簇”的ST8型羊种布鲁氏菌,并首次发现一例人感染ST17型猪种布鲁氏菌。
文摘目的建立针对食品来源的单核细胞增生李斯特氏菌(Listeria monocytogenes,Lm)分离株的多位点串联重复序列分型(Multiple-Locus Variable number tandem repeat Analysis,MLVA)方法,为暴发确认和溯源检测提供实验室支持。方法对2005—2014年间分离自食品的91株Lm进行14个可变数目串联重复序列(Variable Number of Tandem Repeats,VNTR)位点的检测,评估最优检测位点组合并分析检测结果。结果通过采用软件分析,由LMV1、LMV2、LMV7、Lm10、Lm11、Lm23、LM-TR6、TR3和Lm15等9个VNTR位点组成的位点组合为最优MLVA检测位点,可以将91株Lm分离株分为70个型别,分型能力达到0.987 1。结论本研究建立的基于全自动毛细管电泳的由9个检测位点组成的Lm的MLVA分型方法,具有操作简便、快速、结果客观、操作标准化、易于在不同实验室间比较的优势,可作为一线检测方法用于李斯特菌病的暴发确认和溯源检测。
