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目的评价脉冲场凝胶电泳(pulsed field gel electrophoresis,PFGE)技术用于肺炎克雷伯菌(Klebsiella pneumoniae)的分型能力。方法选用220株肺炎克雷伯菌,对PFGE方法的分型力、可重复性、分辨力、流行病学一致性及其与多位点序列分型(MLST)的分型一致性进行评价。挑选全部220株菌株进行实验,评价PFGE的分型力;挑选18株菌株重复2次实验评价PFGE的可重复性;分别挑选3组无流行病关联的碳青霉烯耐药临床株、腹泻患者肠道分离株、食品分离株,使用Simpson差异指数(D)评价PFGE的分辨力;挑选同一次暴发期间分离的34株菌株,评价PFGE的流行病学一致性及其与MLST对暴发菌株分型结果的一致性;挑选64株不同来源的菌株,评价PFGE和MLST分型对流行病学不相关菌株的分型一致性。用Bio Numerics软件对所有实验菌株的PFGE图谱进行分析。结果 220株肺炎克雷伯菌中有216株通过PFGE能够获得有效的图谱,其分型力为98.18%(216/220);18株菌株重复2次实验的图谱完全一致;PFGE对无流行病关联的50株碳青霉烯耐药临床株、34株腹泻患者肠道分离株、48株食品分离株分型的D值分别为0.9910、0.9964、1.0000;针对34株暴发期间分离菌株,PFGE能够很好地甄别出暴发菌株,高度相关菌株和不相关菌株,并且分型结果与菌株分离病区、分离时间具有一致性;针对暴发菌株,PFGE能够将相同MLST型别的菌株聚成一簇;针对流行病学不相关菌株,PFGE不能将相同MLST型别的菌株分成相同或相似的带型或者聚成一簇。结论 PFGE对肺炎克雷伯菌分型具有很好的分型力、可重复性和分辨力;针对暴发菌株,PFGE能够将相同MLST型别的菌株聚成一簇;针对流行病学不相关菌株,PFGE和MLST分型一致性差。
Objective To evaluate the typing ability of Klebsiella pneumoniae by pulsed field gel electrophoresis (PFGE). Methods 220 strains of Klebsiella pneumoniae were selected to evaluate the typing accuracy, repeatability, resolution, epidemiological consistency and classification consistency of multilocus sequence typing (MLST). All 220 strains were selected for the experiment to evaluate the typing ability of PFGE. 18 strains were selected to repeat the experiment twice to evaluate the repeatability of PFGE. Three strains of carbapenem-resistant clinical isolates without epidemics, Gut isolates and food isolates, and the discriminatory power of PFGE was evaluated using the Simpson difference index (D); 34 strains isolated during the same outbreak were selected to evaluate the epidemiological consistency of PFGE and its association with the outbreak strain of MLST Results of the consistency; selection of 64 strains from different sources, evaluation of PFGE and MLST typing on epidemiologically unrelated strains of typing consistency. PFGE profiles of all experimental strains were analyzed using Bio Numerics software. Results 216 strains of 220 strains of Klebsiella pneumoniae were able to obtain effective maps by PFGE, and their typing power was 98.18% (216/220). The profiles of 18 strains were identical in 2 replicates. 50 strains of carbapenem-resistant clinical isolates, 34 strains of diarrhea patients with intestinal isolates, 48 food isolates typing D values were 0.9910, 0.9964, 1.0000; for the 34 strains isolated during the outbreak, PFGE can Outbreak strains, highly related strains and unrelated strains were well screened, and the typing results were consistent with the isolates of the strains and the time of isolation. In the case of outbreak strains, PFGE was able to cluster strains of the same MLST type; In epidemiologically unrelated strains, PFGE can not classify strains of the same MLST type into the same or similar bands or cluster into clusters. Conclusion PFGE has good typing, repeatability and resolving power for the typing of Klebsiella pneumoniae. For the outbreak strains, PFGE can cluster strains of the same MLST type. For strains not related to epidemiology, PFGE and MLST typing poor consistency.