耐碳青霉烯酶肺炎克雷伯菌的分离鉴定及生物学特性分析

以临床尿路感染患者尿液为研究对象，通过分离培养及16s rDNA进行肺炎克雷伯菌的分离鉴定；采用Kirby-Bauer纸片法进行药敏实验，并对分离株进行碳青霉烯酶表型筛选；通过PCR检测常见的碳青霉烯酶耐药基因、荚膜血清型和毒力基因分布情况；对分离的肺炎克雷伯菌株进行了生物被膜形成能力及其对小鼠致病力的分析。本研究从采取的86例尿液标本中分离检出11株耐碳青霉烯酶肺炎克雷伯菌，分离率为12.79%。耐碳青霉烯酶基因PCR检测结果显示，blaNDM、blaVIM、blaIMP和blaKPC基因在分离株中均呈现不同程度的分布。耐药性分析发现11株肺炎克雷伯菌对氨苄西林耐药率为90.91%，对头孢噻肟耐药率为63.64%，对链霉素、庆大霉素、氯霉素和诺氟沙星的耐药率较低。耐碳青霉烯类肺炎克雷伯菌荚膜分型结果显示，分离的11株细菌中10株均为强毒力型菌株（90.9%），其中K57血清型4株（36.4%），K1血清型1株（9.1%），K2血清型1株（9.1%），K5血清型1株（9.1%），K20血清型3株（27.3%），提示该批分离株具有较强的致病力。此外，毒力因子分布结果显示，其毒力因子rmpA（54.5%）、Aerobactin F（54.5%）在菌株中分布较为广泛。生物被膜形成能力检测及小鼠致病性试验结果显示，9株分离株均有较强的生物被膜形成能力，菌株致病力可能与荚膜血清型及生物被膜形成能力相关。综上所述，临床分离的致尿路感染病原肺炎克雷伯菌呈现多重耐药特征，强毒力菌株以K57荚膜型为主，K1、K2均有分布，提示对尿路感染病原应加强耐药监测，合理使用抗菌药物，有效防控多重耐药和强毒力菌株的感染与流行。

ISOLATION, IDENTIFICATION AND BIOLOGICAL CHARACTERISTICS ANALYSIS OF CARBAPENEMASE-RESISTANT KLEBSIELLA PNEUMONIAE

Klebsiella pneumoniae was isolated and identified by 16s rDNA in urine of patients with urinary tract infection. Kirby-Bauer disk method was used for drug sensitivity test and carbapenemase phenotype screening was performed on the isolates. The distribution of common carbapenemase resistance genes, capsule serotypes and virulence genes were detected by PCR. The biofilm formation ability and pathogenicity of isolated K. pneumoniae strains in mice were analyzed. In this study, 11 strains of carbapenemase-resistant K. pneumoniae were isolated from 86 urine samples, and the isolation rate was 12.79%. The PCR results of carbapenemase-resistant genes showed that blaNDM, blaVIM, blaIMP and blaKPC genes were distributed in different degrees in the isolates. Resistance analysis showed that the resistance rates of 11 strains of K. pneumoniae to ampicillin were 90.91%, the resistance rate to cefotaxime was 63.64%, the resistance rate to streptomycin, gentamicin, chloramphenicol and norfloxacin was low. The results of carbapenemase-resistant K. pneumoniae capsule typing showed that 10 of the 11 isolates were virulent strains (90.9%), including 4 isolates of K57 serotype (36.4%), 1 isolate of K1 serotype (9.1%), 1 isolate of K2 serotype (9.1%), and 1 isolate of K5 serotype (9.1%). There were 3 isolates (27.3%) of serotype K20, suggesting that this batch of isolates had strong pathogenicity. In addition, the results of virulence factor distribution showed that virulence factor rmpA (54.5%) and Aerobactin F (54.5%) were widely distributed in the bacterial strains. The results of biofilm formation ability detection and pathogenicity test in mice showed that all the 9 isolates had strong biofilm formation ability, and the pathogenicity of the strains might be related to the serotype of the capsule and biofilm formation ability. In summary, the clinically isolated pathogens K. pneumoniae which causing urinary tract infection showed the characteristics of multiple drug resistance, and the highly virulent strains were mainly K57 capsular strains, while K1 and K2 were distributed, suggesting that drug resistance monitoring and rational use of antibiotics should be strengthened for the pathogens of urinary tract infection.