临床致病性不动杆菌的分子生物学鉴定

目的对临床致病性不动杆菌进行种属鉴定,并比较3种方法(44℃生长试验、特异性引物PCR扩增和16SrRNA测序)对鲍曼不动杆菌鉴定的特异性、准确性及应用价值。方法收集临床致病不动杆菌56株,首先接种在胰蛋白胨大豆肉汤中44℃进行培养,观察是否可以生长;其次,提取56株不动杆菌基因组DNA,用两组不同的特异性引物进行PCR扩增鉴定;最后,用通用引物对56株不动杆菌的16SrRNA进行测序鉴定。结果 56株不动杆菌中有17株可在44℃生长。经特异性引物PCR扩增鉴定,发现8株鲍曼不动杆菌和3株13TU型不动杆菌均可在44℃中较早较快生长。金标准16SrRNA测序确定56株不动杆菌中含有9个种,其中鲍曼不动杆菌和13TU型不动杆菌的鉴定结果与特异性引物PCR扩增法相同。结论应用分子生物学方法鉴定鲍曼不动杆菌具有准确、高效且可重复性高的优点,尤其是特异性PCR扩增法,可作为较难诊断的鲍曼不动杆菌的首选鉴定方法。

Improved identification of 56 pathogenic Acinetobacter strains isolated from clinical cases

Objective To identify the species of Acinetobacter isolates collected from clinical cases and compare the accuracy between the methods of 44 ℃ growth test and molecular ways including specific-PCR amplification and 16S rRNA sequencing in identification of Acinetobacter baumannii.Methods Fifty-six Acinetobacter clinical isolates were collected from hospitals and cultured in TSB at 44 ℃.DNAs of these 56 isolates were extracted and PCR was performed using two sets of specific-primers.16S rRNA was then sequenced by universal primers to identify the species of these Acinetobacter strains.Results Seventeen out of 56 clinical strains could grow at 44 ℃,in which,however,eight and three isolates were confirmed as A.bacumannii and genomic sp.13TU respectively via specific-primer PCR.A total of 9 species were discovered out of 56 Acinetobacter strains through 16S rRNA sequencing,the standard method in identifying bacteria,and the results obtained for A.baumannii and A.genomicsp.13TU were the same as those obtained by specific-primer PCR.Conclusion The molecular methods such as specific-primer PCR and 16S rRNA sequencing are much more reliable in identifying A.baumannii.Specific-primer PCR is especially recommended in confirming A.baumannii for its accuracy and high efficiency.