耐碳青霉烯类抗菌药物的肠杆菌科细菌耐药基因型分析及治疗

Resistantgenotype analysis of carbapenem resistant enterobacteriaceae bacteria and treatment

目的通过对齐齐哈尔市第一医院碳青霉烯类抗生素耐药的肠杆菌科细菌(CRE)耐药基因型的检测及耐药机制分析,为临床抗生素的合理应用和感染的防控提供科学依据。方法收集2018年5月—2020年1月齐齐哈尔市第一医院检验科住院患者的痰液、创口分泌物、肺泡灌洗液、尿液及血液等临床病房患者合格标本中分离到的肠杆菌科细菌,应用梅里埃Vitek2-compact全自动微生物细菌鉴定与药敏仪及K-B纸片扩散法鉴定和筛选CRE菌株;采用改良Hodge试验初步判断CRE菌株产碳青霉烯酶情况;应用聚合酶链式反应PCR法检测CRE菌株的耐药基因型以及膜孔蛋白相关基因型。结果38株CRE中,26株肺炎克雷伯菌,9株大肠埃希菌及3株阴沟肠杆菌,标本主要来源于痰,主要分布于ICU、神经外科等科室。检测了CRE菌株对该院临床常用的19种抗生素的耐药性:头孢菌素类药物,除头孢吡肟耐药率89.5%、头孢他啶耐药率94.8%外,头孢唑啉、头孢呋辛及头孢曲松耐药率都为100%;β-内酰胺类抑制剂复合制剂类药物氨苄西林/舒巴坦、头孢哌酮/舒巴坦及哌拉西林/他唑巴坦耐药率分别为100%、84.2%及81.6%;喹诺酮类药物环丙沙星及左氧氟沙星耐药率分别为94.7%及81.6%;碳青霉烯类药物厄他培南及亚胺培南耐药率分别为97.4%及84.2%;氨基糖苷类药物庆大霉素、妥布霉素及阿米卡星耐药率分别为60.5%、42.1%及26.3%;氨曲南耐药率为84.2%;复方新诺明耐药率为57.9%;未检出对替加环素耐药的菌株。改良Hodge试验阳性菌株都为产KPC型碳青霉烯酶的肺炎克雷伯菌。PCR方法检测菌株耐药基因型:SHV有24株,CTX有23株,KPC有15株,NDM有5株,其中IMP、TEM、VIM、GES、OXA基因型未检出,检出膜孔蛋白OmpC有36株,膜孔蛋白OmpF、OmpK35、OmpK36均未检出,有15株菌株检测出两种或两种以上耐药基因型。结论该院CRE菌株主要为肺炎克雷伯菌,主要来源于临床患者痰液标本,CRE感染常发生于ICU、神经外科等患者基础疾病较多、抗生素应用以及机械通气较频繁的科室;CRE菌株对临床常用药物耐药情况严重,使CRE在治疗中面临用药选择的困难;改良Hodge试验可以用来初筛菌株是否产KPC型碳青霉烯酶;该院CRE菌株耐药机制以产碳青霉烯酶为主,其次为β-内酰胺酶和/或AmpC酶基因的表达同时合并膜孔蛋白的突变或缺失。

Objective To detect the genotypeof carbapenem-resistant enterobacteriaceae(CRE)in the first hospital of Qiqihar City and analyze the resistance mechanism,in orderto provide scientific basis for the rational application of clinical antibiotics and the prevention and control of infection.Methods Enterobacteriaceae that isolated from the qualified samples of sputum,wound secretions,alveolar lavage fluid,urine and blood in the laboratory department of the first hospital of Qiqihar which collected from hospitalized patients from May 2018 to January 2020 were collected.Merieievitek 2-Compact automatic microbial bacterial identification,drug sensitivity instrument and K-B disk diffusion method were used to identify and screen CRE strains.The carbapenemases production of CRE strain was preliminarily determined by modified Hodge test.Polymerase chain reaction(PCR)was used to detect the resistance genotype and membrane pore protein related genotype of CRE strain.Results Among the 38 CRE strains,there were 26 strains of Klebsiella pneumoniae,9 strains of Escherichia coli and 3 strains of Enterobacter clocloe.The samples were mainly from sputum and mainly distributed in neurosurgery department and ICU.The resistance of CRE strains to 19 kinds of antibiotics commonly used in clinic in the hospital was detected.For cephalosporins,the drug resistance rates of cefazolin and cefuroxime and ceftriaxone were 100%,except for cefepime(89.5%)and ceftazidime(94.8%).The drug resistance rates of β-lactam inhibitor compound drugs ampicillin/sulbactam,cefoperazone/sulbactam and piperacillin/tazobactam were 100%,84.2%and 81.6%,respectively.The drug resistance rates of ciprofloxacin and levofloxacin those belongs to quinoloneswere 94.7%and 81.6%,respectively.The drug resistance rates of ertapenem and imipenem of carbapenems were 97.4%and 84.2%,respectively.The drug resistance rates of gentamicin,tobramycin and amikacin were 60.5%,42.1%and 26.3%,respectively.The drug resistance rate of aztreonam was 84.2%.The resistance rate of cotrimoxazole was 57.9%.No strains resistant to tegecycline were detected.All the positive strains according to the Hodge test were KPC-producing klebsiella pneumoniae.Drug-resistant genotype detected by PCR:There were 24 strains of SHV,23 strains of CTX,15 strains of KPC,and 5 strains of NDM,among which IMP,TEM,VIM,GES,and OXA genotypewere not detected.36 strains of Omp C of membrane pore protein related genotypewere detected,none of OmpF,Omp K35,and Omp K36 were detected.15 strains were detected with two or more drug resistant genotype in our hospital.Conclusions THE CRE strains were mainly Klebsiella pneumoniae,mainly from sputum specimens of clinical patients.CRE infections were often found in departmentssuch as neurosurgery department and ICU where patients with basic diseases and received antibiotics and frequent mechanical ventilation.The resistance of CRE strains to commonly used drugs is serious,which makes it difficult to choose drugs in treatment of CRE.The modified Hodge test could be used to screen whether the strain produces KPC carbapenemase.The resistance mechanism of CRE strains in our hospital was mainly carbapenase production,followed byβ-lactamase and/or AMPCase gene expression combined with membrane pore protein mutation or deletion.