【摘要】：目的調查天津醫(yī)科大學總醫(yī)院臨床分離肺炎鏈球菌對常用抗菌藥物的敏感性,檢測其常見血清型及毒力基因的分布情況,并對其耐藥機制進行初步分析,為合理使用抗生素治療肺炎鏈球菌感染提供依據(jù)同時控制該耐藥菌株在醫(yī)院內的傳播。方法收集2012年10月至2014年4月天津醫(yī)科大學總醫(yī)院臨床分離的非重復肺炎鏈球菌72株,應用Vitek-2 compact全自動細菌分析系統(tǒng)及配套鑒定卡和相應微量肉湯稀釋試驗分別進行菌種鑒定和藥敏試驗;應用聚合酶鏈反應(PCR)方法檢測大環(huán)內酯類抗菌藥物耐藥基因,包括紅霉素核糖體甲基化酶基因(erm B)、主動外排基因(mef A、mef E),四環(huán)素耐藥基因(tet M)以及轉座子整合酶基因(int Tn);常見血清型基因(3、6A/B、14、15A、19A、19F、23F)及毒力基因自溶素基因(lyt A)、溶血素基因(ply)及神經氨酸酶基因(nan A)。PCR擴增的基因陽性產物進行測序并與Gen Bank數(shù)據(jù)庫比對。結果72株肺炎鏈球菌藥敏結果顯示,對萬古霉素、利福平均敏感,對左氧氟沙星、頭孢噻肟、氯霉素、阿莫西林、青霉素敏感率大于76%,其中對青霉素耐藥率PRSP為1.4%,中介率PISP為22.2%、敏感率PSSP為76.4%。對紅霉素、克林霉素、四環(huán)素敏感率低于10%。72株肺炎鏈球菌攜帶紅霉素耐藥基因erm B 65株(90.3%)、mef E 17株(23.6%)、mef A 9株(12.5%)、四環(huán)素耐藥基因tet M 63株(87.5%),轉座子整合酶基因int Tn 62株(86.1%),其中erm B+tet M+int Tn 38株(52.8%)。對四環(huán)素耐藥的肺炎鏈球菌tet M基因陽性率是93.8%,高于四環(huán)素敏感菌株(20%);對四環(huán)素耐藥的菌株,其對紅霉素和克林霉素的耐藥率亦高于四環(huán)素敏感的菌株;int Tn基因陽性組的紅霉素、克林霉素和四環(huán)素的耐藥率較int Tn基因陰性組高。72株肺炎鏈球菌常見血清型依次為19F(15,20.8%)、3(12,16.7%)、23F(9,12.5%)、6A/B(8,11.1%)、14(6,8.3%)、19A(4,5.6%)和15A(2,2.8%)。毒力基因檢測結果為lyt A 65株(90.3%)、ply 67株(93.1%)、nan A 52株(72.2%),其中51株(70.8%)同時攜帶lyt A、ply和nan A三種基因,12株(16.7%)同時攜帶lyt A和ply基因,1株(1.4%)同時攜帶ply和nan A基因。PCR擴增產物經測序并與Gen Bank數(shù)據(jù)庫比對后均為目的基因。結論本研究的臨床分離肺炎鏈球菌對萬古霉素、利福平敏感率均為100%,左氧氟沙星、氯霉素、頭孢噻肟敏感率較高,對紅霉素、克林霉素和四環(huán)素耐藥率較高。臨床分離的肺炎鏈球菌同時攜帶多種耐藥基因,其中最常見的耐藥基因組合方式為erm B+tet M+int Tn,erm B基因和tet M基因分別是引起大環(huán)內酯類抗菌藥物和四環(huán)素類抗菌藥物耐藥的主要機制,int Tn基因可能是介導肺炎鏈球菌多重耐藥的重要機制,臨床醫(yī)師應注意合理用藥。血清型19F、3和23F是主要常見的血清型,本地區(qū)人群接種肺炎鏈球菌疫苗時應首選含有這三種血清型的PCV13莢膜多糖抗原疫苗。毒力基因lyt A和ply檢出率較高,為進一步研究毒力因子的作用機制提供一定基礎,并為研制抗肺炎鏈球菌新藥和預防肺炎鏈球菌感染的新型疫苗提供幫助。
[Abstract]:Objective To investigate the sensitivity of Streptococcus pneumoniae to the common antibiotics in the general hospital of Tianjin Medical University, to detect the distribution of the common serotypes and virulence genes, and to analyze the mechanism of drug resistance. To provide a basis for the rational use of antibiotics for the treatment of Streptococcus pneumoniae infection, the transmission of the drug-resistant strain within the hospital is also controlled. Methods 72 strains of non-repeated Streptococcus pneumoniae were collected from October, 2012 to April, 2014 in the general hospital of Tianjin Medical University, and the strain identification and drug sensitivity test were carried out by using the Vitk-2 compact full-automatic bacterial analysis system and the matched identification card and the corresponding micro-broth dilution test. The drug-resistant genes of macrolides are detected by polymerase chain reaction (PCR) method, including the erythromycin ribosomal methylase gene (erm B), the active efflux gene (mef A, mef E), the tetracycline resistance gene (tet M) and the transposon integrase gene (int Tn); Common serotype genes (3, 6A/ B, 14, 15A, 19A, 19F, 23F) and virulence gene autolysin gene (lyt A), hemolysin gene (ply), and neuraminidase gene (nan A) are common. The PCR-amplified gene-positive product was sequenced and compared to the Gen Bank database. Results The drug sensitivity of 72 strains of Streptococcus pneumoniae showed that both vancomycin and rifampin were sensitive, and the sensitivity of levofloxacin, cefixime, chloramphenicol, amoxicillin and penicillin was more than 76%. The resistance rate of penicillin to penicillin was 1.4% and the intermediate rate of PISP was 2.2%. The sensitivity rate of PSSP was 76.4%. The sensitivity of erythromycin, klincomycin and tetracycline was lower than 10%. 72 strains of S. pneumoniae carried the erythromycin-resistant gene erm B 65 (90.3%), mef E 17 (23. 6%), mef A 9 (12.5%) and tetracycline-resistant gene tet M 63 (87.5%). The total of the transposon integrase gene int Tn 62 (86.1%), of which the erm B + tet M + int Tn 38 (52.8%). The positive rate of the Tet M gene of the streptococcus pneumoniae resistant to tetracycline is 93.8%, which is higher than that of the tetracycline-sensitive strain (20%); the resistance rate of the tetracycline-resistant strain is higher than that of the tetracycline-sensitive strain; and the erythromycin of the int Tn gene positive group, The common serotypes of the 72 strains of S. pneumoniae were 19F (15, 20. 8%), 3 (12, 16. 7%), 23F (9, 12.5%), 6A/ B (8, 11.1%), 14 (6, 8. 3%), 19A (4, 5.6%) and 15A (2, 2.8%). The results of virulence gene detection were lyt A 65 (90.3%), pldy 67 (93.1%), and nan A 52 (72.2%), of which 51 (72.8%) simultaneously carried lyt A, The three genes of ply and nan A, 12 (16. 7%) carried the lyt A and the ply genes, and 1 (1.4%) simultaneously carried the ply and nan A genes. The PCR products were sequenced and compared with the Gen Bank database. Conclusion The sensitivity of the clinical isolates of Streptococcus pneumoniae to vancomycin and rifampin is 100%, levofloxacin, chloromycetin and cemedoxime are more sensitive to vancomycin, and the resistance to erythromycin, klincomycin and tetracycline is high. The clinically isolated streptococcus pneumoniae carries a plurality of drug resistance genes at the same time, wherein the most common drug resistance gene combination mode is the erm B + tet M + int Tn, the erm B gene and the tet M gene are the main mechanisms for causing the drug resistance of the macrolide antibiotics and the tetracycline antibiotics, The int Tn gene may be an important mechanism to mediate the multiple drug resistance of S. pneumoniae, and the clinician should pay attention to the rational use of the drug. Serotypes 19F, 3, and 23F are the main common serotypes, and the population of the region is vaccinated with a Streptococcus pneumoniae vaccine that is preferred to contain the three serotypes of the PCV13 membrane polysaccharide antigen vaccine. The detection rate of the virulence genes lyt A and ply is high, which provides a basis for the further study of the mechanism of the virulence factors, and is a new type of vaccine for the development of a new drug for the streptococcus pneumoniae and the prevention of the infection of the streptococcus pneumoniae.
【學位授予單位】：天津醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2015
【分類號】：R446.5

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