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目的了解鲍曼不动杆菌中Ⅰ类整合子与ISCR1分布情况及其所携带耐药基因的种类,从而分析其结构及其与耐药性的关系。方法采用煮沸法提取鲍曼不动杆菌基因组DNA,设计引物进行PCR反应,扩增产物用1.2%琼脂糖凝胶电泳检测并记录结果,根据Ⅰ类整合子可变区扩增产物图谱进行RFLP分型,最后进行产物的序列分析。结果PCR扩增51株鲍曼不动杆菌,Ⅰ类整合酶基因intⅠ扩增阳性45株,其中包括可变区阳性32株。并且Ⅰ类整合子的可变区扩增出的目的片段大小各不相同,有28株扩增片段约为2.3kb,2株扩增片段约为2.4kb,2株扩增片段为3kb;ISCR1扩增阳性22株,其下游扩增阳性5株,扩增片段大小约为3kb。同时携带Ⅰ类整合子和ISCR1扩增阳性菌目的片段大小约为2kb。酶切图谱显示,大小为2.3kb的Ⅰ类整合子可变区产物为同一带型;序列分析显示,大小不同的Ⅰ类整合子的可变区各片段所含的基因盒各不相同;耐药基因序列分析显示,5株ISCR1的下游扩增片段为qnrA1-ampR。同时显示Ⅰ类整合子和ISCR1两种元件以串联的形式存在于鲍曼不动杆菌中。结论Ⅰ类整合子与ISCR1大多以串联形式同时存在于骨髓炎患者感染的鲍曼不动杆菌中,该结构对耐药基因在不同鲍曼不动杆菌菌株间的水平传播可能起介导作用。
Objective To understand the distribution of class Ⅰ integron and ISCR1 and the types of drug resistance genes carried by Acinetobacter baumannii in order to analyze its structure and its relationship with drug resistance. Methods The genomic DNA of Acinetobacter baumannii was extracted by boiling method. The primers were designed for PCR reaction. The amplified products were detected by 1.2% agarose gel electrophoresis and the results were recorded. Based on the amplified product map of class Ⅰ integron variable region, Type, the final product of the sequence analysis. Results 51 strains of Acinetobacter baumannii were amplified by PCR, and 45 strains of int Ⅰ gene of type Ⅰ integrase were amplified, including 32 strains of variable region. The size of the target fragment amplified by the variable region of the class I integrons varied from 28 isolates to 2.3 kb, 2 isolates to 2.4 kb and 2 isolates to 3 kb. ISCR1 22 were positive for amplification and 5 were positive for downstream amplification. The amplified fragment size was about 3kb. At the same time carrying the class Ⅰ integron and ISCR1 amplified positive bacteria the purpose of the fragment size of about 2kb. The restriction enzyme digest showed that the class Ⅰ integron variable region products of the size of 2.3kb belonged to the same band. Sequence analysis showed that the gene cassettes contained in the variable region of each class Ⅰ integrons of different sizes contained different gene cassettes. The analysis of drug gene sequence showed that the downstream amplification of 5 ISCR1 was qnrA1-ampR. At the same time, two types of integrons, type I and ISCR1, were found in Acinetobacter baumannii in series. Conclusion Most of class Ⅰ integrins and ISCR1 coexist in Acinetobacter baumannii infected with osteomyelitis in series. This structure may play an important role in mediating the horizontal transmission of drug resistance genes among different strains of Acinetobacter baumannii.