寧夏某三甲醫院28例尿路感染耐碳青霉烯類腸桿菌目細菌分子流行特征及耐藥性分析

中圖分類號：R446.5 文獻標志碼：A

Molecular epidemiological characteristics and antimicrobial resistance of carbapenem-resistant Enterobacterales in 28 cases of urinary tract infection in a grade A tertiary hospital in Ningxia

Yan Lixin1.2， Zhu Chenyu，Tao Jia^2，4 ，HongWei1.2，Hu Xinxin1.2，andJia Wei4.4 （1TheFirstCliicalMedicalCollgeofNngxiaMedicalUniversityYnchuan75oo4;2ingxiaKeyLaboratoryofliicald Pathogenic Mcrobiologyichuan7o4;3ushanecondPeople'sHospitalzushan3o;4Laboratoryedicalte General Hospital ofNingxia Medical University， Yinchuan 750004）

AbstractObjectiveTo explore the molecular epidemiological characteristics，antimicrobial resistance，and homogeneityof carbapenem-resistant Enterobacteriaceae （CRE）in patients with urinary tractinfections at the General HospitalofNingxia Medical University，providing areference for their treatment.MethodsFrom December 2013 to November 2022，a total of28 strains ofCRE fromurinary tract infection inpatients were collected.WHONET5.6 software was used to analyze the distribution and antimicrobial susceptibilityof the strains.Polymerase chain reaction （PCR） was used to detect four carbapenemase genes （ [bla_KPC. bla_NDM，bla_OXA-48 and bla_IMP） ， and Sanger sequencing was used to analyze subtypes.Multilocus sequence typing （MLST） was employed to determine the clonal types of the strains.The transferabilityof resistance genes was verified through conjugation experiments. ResultsAmong the 28 CRE strains， 11 （39.30%） wereKlebsiellapneumoniae（ K. pneumoniae） and 9 （32.14%） wereEscherichiacoli， mainly isolated from emergency department inpatients.Drug susceptibility results showed that more than half of the bacteria testedwersstant torousommonatiboticdtedetedesistaegsere bla_?NDM and bla_KPC， with bla_NDM-5 （ 77.27% being predominant. Conclusionexperiments revealed that 7 strains of E . coli carrying the bla_NDM-5 gene and 3 strains of K. pneumoniae carrying the bla_NDM-5 gene could successfully transfer teir carbapenem resistance phenotype to the recipient strain E .coli EC600.MLST results indicated that 11K. pneumoniae strains and 9E . coli stains had 18 different sequence types （STs），suggesting a diverse range ofclone types. Cluster analysis demonstrated close genetic relationships among strains within the same ward. Conclusions The CRE strains causing urinary tract infections at the hospital are predominantly K. pneumoniae and E .coli， and their resistance mechanism to carbapenems is mainly associated with the production of NDM-5 carbapenemase.The bla_NDM-5 gene can horizontally spread through plasmids，emphasizing the need for close monitoring of CRE strains to prevent further disemination ofresistance genes and potential hospital outbreaks.

Key WordsCarbapenem-resistant Enterobacterales; Urinary tract infection; NDM-5;Multilocus sequence typing; Conjugation of plasmids

尿路感染（urinarytractinfections，UTIs）是醫院感染中僅次于呼吸道感染的第二大感染性疾病[1]，其中腸桿菌目細菌，特別是大腸埃希菌是最常見的致病菌[2]。隨著抗菌藥物的廣泛使用，病原菌的耐藥率不斷增加。碳青霉烯類抗生素因其廣泛的抗菌譜和強大的抗菌活性，曾是治療嚴重細菌感染的首選藥物。近年來碳青霉烯類耐藥腸桿菌目（carbapenem-resistantEnterobacterales，CRE）的細菌的檢出率不斷增加，給UTI的臨床治療帶來了極大的困難。本研究旨在調查寧夏醫科大學總醫院2013年12月—2022年11月臨床分離的來自尿路感染CRE的患者信息、耐藥種類、耐藥基因及傳播情況等，并對分離獲得的大腸埃希菌和肺炎克雷伯菌耐藥性及同源性進行分析，為臨床患者的診斷和治療提供參考依據。

1材料與方法

1.1材料

1.1.1 菌株來源

收集2013年12月—2022年11月寧夏醫科大學總醫院住院患者尿路感染的CRE非重復菌株28株。大腸埃希菌ATCC25922為藥物敏感性質控菌株；大腸埃希菌EC600作為接合實驗受體菌株。

1.1.2 儀器與試劑

基質輔助激光解吸電離飛行時間質譜儀（法國BioMérieux公司），紫外凝膠成像及分析系統儀、麥氏比濁儀、高壓水平電泳儀和電泳槽（美國Bio-Rad公司），PCR儀（德國Eppendorf公司），藥敏紙片（英國Oxoid公司），Taq2PCRMasterMix、DNAMarkerDL2000（日本Takara公司），50xTAE和MH瓊脂平板（北京索萊寶）。

1.2方法

1.2.1 菌種鑒定及體外藥敏試驗

基質輔助激光解吸電離飛行時間質譜儀（MALDI-TOFMS）進行細菌鑒定，VITEK2系統進行藥敏試驗，結果判定參照2021年美國臨床和實驗室標準化協會（CLSI）M100（第31版）標準[3]。

1.2.2 耐藥基因檢測

采用高溫水煮裂解法提取細菌DNA，通過PCR擴增碳青霉烯酶基因（ 、 bla_oxA-48 、 bla_?NDM 和bla_IMP） 。……