增菌-PCR法与传统方法检测禽肉中沙门菌的比较研究  

Comparison study of enrichment-PCR and traditional method for detection of Salmonella in poultry

作　　者：闫琳[1] 王晓英[1] 郭云昌[1] 裴晓燕[1] 余东敏[1] 

机构地区：[1]中国疾病预防控制中心营养与食品安全所,北京100021

出　　处：《卫生研究》2011年第3期348-351,354,共5页Journal of Hygiene Research

基　　金：国家自然科学基金资助项目(No.30571575);科技部科研院所技术开发研究专项(No.2009EG150293)

摘　　要：目的比较增菌-PCR法与传统方法检测禽肉中沙门菌的敏感性、特异性和时效性。方法以沙门菌invA基因为基础,选择特异性引物,采用30株沙门氏菌和18株非沙门菌建立并验证PCR法的特异性。选用肠炎沙门菌CMCC50041,参照GB4789.4—2010进行人工染菌实验,染菌量分别为每25g样本1、10、102、103、104、105和106 CFU。分别在增菌0、4、8、12和18h取1ml培养液,提取DNA进行PCR检测,并同时划线接种选择性平板,用传统方法进行分离鉴定,比较两种方法检测的敏感性和特异性。采集16份市售整鸡样本,用这两种方法进行检测,进一步比较两者的阳性检出率。结果 30株沙门菌都检出阳性,而非沙门菌都检测为阴性,说明本研究建立的PCR法特异性好。人工染菌的样本在增菌12h后,PCR法检出限可达每25g禽肉样本1CFU,传统方法检出限为每25g禽肉样本10CFU。增菌-PCR法整个流程只需要1天,比传统方法需时提前4～6天。16份市售样本,增菌-PCR法的阳性率为43.75%(7/16),高于传统方法31.25%(5/16)。结论增菌-PCR法具有快速简便、敏感特异等优点,较传统方法大大缩短了检出时限,提高了沙门菌的检出率。Objective To compare the sensitivity,specificity and timeliness of Salmonella in poultry detected by enrichment-PCR and traditional method.Method Salmonella specific primers（PsalF-PsalR） were chosen on the basis of invA gene.PCR method was developed and its specificity was proved by 30 Strains of Salmonella standard and isolates,and 18 strains of non-Salmonella.Refering to GB4789.4—2010,artificially contaminated experiment was done,six artificially-inoculated samples containing final concentration of salmonella CMCC50041（1,10,102,103,104,105 and 106 CFU /25g poultry samples） were preparated respectively,meanwhile one sample without inoculation was as control of background value.All the samples were incubated at 37℃ for 0,4,8,12 and 18 h,and 1ml of culture solutions were taken from the samples to extract DNA,respectively.PCR and traditional methods were applied to detect Salmonella bacteria.Sixteen samples of retail whole poultry were collected from markets in Beijing and detected by the above two methods after enriching for 12 h in SC at 37℃.Results PCR method established was stable and specific,as 30 Salmonella were detected positive and 18 non-Salmonella were negative.PCR detection limit for artificially contaminated samples after enriching for 12h was 1 CFU/25g sample.Traditional method detection limit was 10 CFU / 25g samples.The time limits of enrichment-PCR were 4-6 days,shorter than those of traditional method.The positive rate out of total 16 market-sold samples was 43.75%（7/16）for enrichment-PCR,while the rate was 31.25%（5/16）for traditional method.Conclusion Enrichment-PCR was simple,rapid,sensitive and specific in comparition with traditional method.It could be applied in fast diagnosis of food poisoning and actual food samples screening.

关 键 词：增菌-PCR法 禽肉 沙门菌 检测 

分 类 号：R155.51[医药卫生—营养与食品卫生学] R155.31[医药卫生—公共卫生与预防医学]