动物医学进展
動物醫學進展
동물의학진전
PROGRESS IN VETERINARY MEDICINE
2013年
12期
145-148,149
,共5页
温肖会%翟少伦%丘婷%吕殿红%贾春玲%袁洁%黄忠%周秀蓉%魏文康
溫肖會%翟少倫%丘婷%呂殿紅%賈春玲%袁潔%黃忠%週秀蓉%魏文康
온초회%적소륜%구정%려전홍%가춘령%원길%황충%주수용%위문강
blaNDM-1 基因%耐药细菌%聚合酶链反应
blaNDM-1 基因%耐藥細菌%聚閤酶鏈反應
blaNDM-1 기인%내약세균%취합매련반응
blaNDM-1 gene%drug-resistant bacteria%PCR
为了调查养殖场和农贸市场中的耐药细菌是否携带 blaNDM-1基因,根据 GenBank 已公布的blaNDM-1基因序列设计 PCR 引物,优化 PCR 反应体系和反应条件,建立了 blaNDM-1基因的 PCR 检测方法,以人工合成的 blaNDM-1基因作为阳性质控品,以大肠埃希菌、肺炎克雷伯菌、阴沟肠杆菌和铜绿假单胞菌等4株标准菌株为阴性对照,验证耐药细菌 blaNDM-1基因 PCR 检测方法的特异性、敏感性、重复性,并对229株大肠埃希菌分离株和31株链球菌分离株进行检测。结果表明,blaNDM-1基因 PCR 检测方法的特异性、敏感性、重复性试验均成立,该方法扩增的目的基因片段为241 bp,能检出的最小基因组 DNA 浓度为9.18×10-7μg/μL;229株大肠埃希菌分离株和31株链球菌分离株 blaNDM-1基因 PCR 检测结果均没有出现目的条带,表明从广东省部分地区分离到的细菌分离株中没有携带 blaNDM-1基因。该方法可作为耐药细菌 blaNDM-1基因的早期检测、快速筛查手段在兽医临床耐药细菌检测中应用。
為瞭調查養殖場和農貿市場中的耐藥細菌是否攜帶 blaNDM-1基因,根據 GenBank 已公佈的blaNDM-1基因序列設計 PCR 引物,優化 PCR 反應體繫和反應條件,建立瞭 blaNDM-1基因的 PCR 檢測方法,以人工閤成的 blaNDM-1基因作為暘性質控品,以大腸埃希菌、肺炎剋雷伯菌、陰溝腸桿菌和銅綠假單胞菌等4株標準菌株為陰性對照,驗證耐藥細菌 blaNDM-1基因 PCR 檢測方法的特異性、敏感性、重複性,併對229株大腸埃希菌分離株和31株鏈毬菌分離株進行檢測。結果錶明,blaNDM-1基因 PCR 檢測方法的特異性、敏感性、重複性試驗均成立,該方法擴增的目的基因片段為241 bp,能檢齣的最小基因組 DNA 濃度為9.18×10-7μg/μL;229株大腸埃希菌分離株和31株鏈毬菌分離株 blaNDM-1基因 PCR 檢測結果均沒有齣現目的條帶,錶明從廣東省部分地區分離到的細菌分離株中沒有攜帶 blaNDM-1基因。該方法可作為耐藥細菌 blaNDM-1基因的早期檢測、快速篩查手段在獸醫臨床耐藥細菌檢測中應用。
위료조사양식장화농무시장중적내약세균시부휴대 blaNDM-1기인,근거 GenBank 이공포적blaNDM-1기인서렬설계 PCR 인물,우화 PCR 반응체계화반응조건,건립료 blaNDM-1기인적 PCR 검측방법,이인공합성적 blaNDM-1기인작위양성질공품,이대장애희균、폐염극뢰백균、음구장간균화동록가단포균등4주표준균주위음성대조,험증내약세균 blaNDM-1기인 PCR 검측방법적특이성、민감성、중복성,병대229주대장애희균분리주화31주련구균분리주진행검측。결과표명,blaNDM-1기인 PCR 검측방법적특이성、민감성、중복성시험균성립,해방법확증적목적기인편단위241 bp,능검출적최소기인조 DNA 농도위9.18×10-7μg/μL;229주대장애희균분리주화31주련구균분리주 blaNDM-1기인 PCR 검측결과균몰유출현목적조대,표명종광동성부분지구분리도적세균분리주중몰유휴대 blaNDM-1기인。해방법가작위내약세균 blaNDM-1기인적조기검측、쾌속사사수단재수의림상내약세균검측중응용。
To investigate whether the blaNDM-1 gene was carried by drug-resistant bacteria in the farms and farmer′s markets,the PCR primers were designed according to the sequence of blaNDM-1 gene from GenBank and the PCR system and reaction conditions were optimized.The specificity,sensitivity,repeatability of PCR detection method for blaNDM-1 gene was verified by positive control of synthetic blaNDM-1 gene and nega-tive control of 4 reference strains including of E.coli,Klebsiella pneumoniae ,Enterobacter cloacae and Pseudomonas aeruginosa .229 isolates of E.coli and 31 isolates of Streptococcus were detected by PCR method.The results showed that the specificity,sensitivity,repeatability of PCR method for blaNDM-1 gene was established and the target gene was about 241 bp.The minimal detectable DNA concentration was 9. 18×10-7 μg/μL.There was no target fragment on agarose gel for 229 isolates of E.coli and 31 isolates of Streptococcus.It showed that blaNDM-1 gene was not carried by bacterial isolates from some area of Guang-dong province.The PCR method can be used on monitoring of drug-resistant bacteria as an early diagnosis and rapid screening method for the drug-resistant bacteria carrying blaNDM-1 gene in veterinary clinic.