部分茶饮料原辅料中优势菌-克罗诺杆菌(原阪崎肠杆菌)的分离和鉴定
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摘要
克罗诺杆菌(原阪崎肠杆菌)是新兴的条件致病菌,主要通过污染的婴儿配方奶粉引起2岁以下婴幼儿严重疾病甚至死亡。在对茶饮料原辅料192份样品进行细菌总数测定和优势菌16S rDNA序列分析时,发现2批次绿茶和1批次果葡糖浆中优势菌疑似条件致病菌-克罗诺杆菌,而且其数量较高(4.0×102cfu/g~104cfu/g)。进而通过rpoB序列分析,将4个分离株鉴定为阪崎克罗诺杆菌Cronobacter sakazakii(原阪崎肠杆菌Enterobacter sakazakii)。结果表明在常见的食品原料-果葡糖浆和绿茶中存在克罗诺杆菌的污染,rpoB分型方法在对克罗诺杆菌属细菌的鉴定到种上表现出高度特异性,灵敏而准确。
Cronobacter is an emerging opportunistic bacterial pathogen which could cause serious neonatal diseases by contaminated powdered infant formula.During the investigation of microbiology in tea-drink materials from 2010-2011,192 samples were analyzed by routine colony counts and 16 S rDNA sequence analysis of predominant bacteria.Three samples were found predominated by 4.0×102->104cfu/g Cronobacter like bacteria.And it was confirmed by 16S rDNA and rpoB sequence analysis that those four isolates were C.sakazakii(formerly E.sakazakii).It were showed that normal food materials-green tea and fructose syrup could also contaminated by Cronobacter,and rpoB sequence analysis is capable to differential several species within this genus.
Cronobacter is an emerging opportunistic bacterial pathogen which could cause serious neonatal diseases by contaminated powdered infant formula.During the investigation of microbiology in tea-drink materials from 2010-2011,192 samples were analyzed by routine colony counts and 16 S rDNA sequence analysis of predominant bacteria.Three samples were found predominated by 4.0×102->104cfu/g Cronobacter like bacteria.And it was confirmed by 16S rDNA and rpoB sequence analysis that those four isolates were C.sakazakii(formerly E.sakazakii).It were showed that normal food materials-green tea and fructose syrup could also contaminated by Cronobacter,and rpoB sequence analysis is capable to differential several species within this genus.
引文
[1]Food and Agriculture Organization and World Health Organization.JointFAO/WHO workshop on Enterbacter sakazakii and other microorganismsin powdered infant formula[C].Geneva,2004.
[2]PANGALOS G.Sur un bacille chromogène isolépar hémoculture C Rsoc biol[J].Comptes Rendus Seances Soc Biol,1929,100:1097.
[3]FARMER JJIII,ASBURY MA,HICKMAN FW,et al.The Enterobacteri-aceae study group.Enterobacter sakazaki:a new species of“Enterobacte-riaceae”isolated from clinical specimens[J].Int J Syst Evol Microb,1980,30:569-584.
[4]IVERSEN C,MULLANE N,MCCARDELL B,et al.a new genus to ac-commodate the biogroups of Enterobacter sakazakii,and proposal ofCronobacter sakazakii gen.nov.,comb.nov.,Cronobacter malonaticussp.nov.,Cronobacter turicensis sp.nov.,Cronobacter muytjensii sp.nov.,Cronobacter dublinensis sp.nov.,Cronobacter genomospecies I,and ofthree subspecies,Cronobacter dublinensis subsp.Dublinensis subsp.nov.,Cronobacter dublinensis subsp.lausannensis subsp.nov.and Cronobacterdublinensis subsp.lactaridi subsp.nov[J].Int J Syst Evol Microb,2008,58:1442-1447.
[5]JOSEPH S,CETINKAYA E,DRAHOVSKA H,et al.Cronobacter condi-menti sp.nov.,isolated from spiced meat and Cronobacter universalis sp.nov.,a novel species designation for Cronobacter sp.genomospecies 1,recovered from a leg infection,waterand food ingredients[J].Int J SystEvol Microb,2012,62(6):1277-1283.
[6]CRUZ A,XICOHENCATL-CORTES J,GONZALEZ-PEDRAJO B,etal.Virulence traits in Cronobacter species isolated from different sources[J].Can J Microbiol,2011,57(9):735-744.
[7]赵贵明,袁飞,杨海荣,等.阪崎肠杆菌的分类编号及新种属生物学特性[J].卫生研究,2010,2(3):248-250.
[8]STOOP B,LEHNER A,LVERSEN C,et al.Development and evaluationof rpoB based PCR systems to differentiate the six proposed species with-in the genus Cronobacter[J].Int J Food Microb,2009,136(2):165-168.
[9]周胜利,黄佳,晁爱敏,等.基于rpoB基因的阪崎克罗诺杆菌分子检测体系的建立及其应用[J].安全与环境学报,2012,12(4):158-161.
[10]GB4789.2-2010食品微生物学检验菌落总数测定[S].
[11]KANDHAI MC,REIJ MW,GORRIS LC,et al.Occurrence of Enter-obacter sakazakii in food production environments and households[J].Lancet,2004,63(9402):39-40.
[12]Leuschner RG,BEW J.A Medium for the Presumptive Detection of En-terobacter sakazakii Infant Formula:Interlaboratory Smdy[J].J AOACInt,2004,87(3):604-613.
[13]黄忠梅,王快翀,田延河.从进口饼干、巧克力和方便面中检出阪崎肠杆菌[J].中国食品卫生杂志,2007,19(2):137-13.
[14]王艳,焦彦朝,罗阿东,等.辣椒制品中阪崎肠杆菌的检测与鉴定[J].贵州农业科学,2012,40(6):198-200.
[15]陆幸儿,郑悦康,吴灿权.熟肉制品受染阪崎肠杆菌情况调查[J].中国卫生监督杂志,2011,2(2):168-169.
[16]TURCOVSK I,KUNIKOVK,DRAHOVSKH,et al.Biochemicaland molecular characterization of Cronobacter spp.(formerly Enterobac-ter sakazakii)isolated from foods[J].Anton Van Leeuw,2011,99(2):257-269.
[17]KIM KP,LOESSNER MJ.Enterobacter sakazakii invasion in human in-testinal Caco-2 cells requires the host cell cytoskeleton and is enhancedby disruption of tight junction[J].Infect Imm,2008,76(2):562-570.
[2]PANGALOS G.Sur un bacille chromogène isolépar hémoculture C Rsoc biol[J].Comptes Rendus Seances Soc Biol,1929,100:1097.
[3]FARMER JJIII,ASBURY MA,HICKMAN FW,et al.The Enterobacteri-aceae study group.Enterobacter sakazaki:a new species of“Enterobacte-riaceae”isolated from clinical specimens[J].Int J Syst Evol Microb,1980,30:569-584.
[4]IVERSEN C,MULLANE N,MCCARDELL B,et al.a new genus to ac-commodate the biogroups of Enterobacter sakazakii,and proposal ofCronobacter sakazakii gen.nov.,comb.nov.,Cronobacter malonaticussp.nov.,Cronobacter turicensis sp.nov.,Cronobacter muytjensii sp.nov.,Cronobacter dublinensis sp.nov.,Cronobacter genomospecies I,and ofthree subspecies,Cronobacter dublinensis subsp.Dublinensis subsp.nov.,Cronobacter dublinensis subsp.lausannensis subsp.nov.and Cronobacterdublinensis subsp.lactaridi subsp.nov[J].Int J Syst Evol Microb,2008,58:1442-1447.
[5]JOSEPH S,CETINKAYA E,DRAHOVSKA H,et al.Cronobacter condi-menti sp.nov.,isolated from spiced meat and Cronobacter universalis sp.nov.,a novel species designation for Cronobacter sp.genomospecies 1,recovered from a leg infection,waterand food ingredients[J].Int J SystEvol Microb,2012,62(6):1277-1283.
[6]CRUZ A,XICOHENCATL-CORTES J,GONZALEZ-PEDRAJO B,etal.Virulence traits in Cronobacter species isolated from different sources[J].Can J Microbiol,2011,57(9):735-744.
[7]赵贵明,袁飞,杨海荣,等.阪崎肠杆菌的分类编号及新种属生物学特性[J].卫生研究,2010,2(3):248-250.
[8]STOOP B,LEHNER A,LVERSEN C,et al.Development and evaluationof rpoB based PCR systems to differentiate the six proposed species with-in the genus Cronobacter[J].Int J Food Microb,2009,136(2):165-168.
[9]周胜利,黄佳,晁爱敏,等.基于rpoB基因的阪崎克罗诺杆菌分子检测体系的建立及其应用[J].安全与环境学报,2012,12(4):158-161.
[10]GB4789.2-2010食品微生物学检验菌落总数测定[S].
[11]KANDHAI MC,REIJ MW,GORRIS LC,et al.Occurrence of Enter-obacter sakazakii in food production environments and households[J].Lancet,2004,63(9402):39-40.
[12]Leuschner RG,BEW J.A Medium for the Presumptive Detection of En-terobacter sakazakii Infant Formula:Interlaboratory Smdy[J].J AOACInt,2004,87(3):604-613.
[13]黄忠梅,王快翀,田延河.从进口饼干、巧克力和方便面中检出阪崎肠杆菌[J].中国食品卫生杂志,2007,19(2):137-13.
[14]王艳,焦彦朝,罗阿东,等.辣椒制品中阪崎肠杆菌的检测与鉴定[J].贵州农业科学,2012,40(6):198-200.
[15]陆幸儿,郑悦康,吴灿权.熟肉制品受染阪崎肠杆菌情况调查[J].中国卫生监督杂志,2011,2(2):168-169.
[16]TURCOVSK I,KUNIKOVK,DRAHOVSKH,et al.Biochemicaland molecular characterization of Cronobacter spp.(formerly Enterobac-ter sakazakii)isolated from foods[J].Anton Van Leeuw,2011,99(2):257-269.
[17]KIM KP,LOESSNER MJ.Enterobacter sakazakii invasion in human in-testinal Caco-2 cells requires the host cell cytoskeleton and is enhancedby disruption of tight junction[J].Infect Imm,2008,76(2):562-570.