G群链球菌的分子鉴定及耐药特征分析

doi:10.3969/j.issn.1673-8640.2016.09.019

摘要/Abstract

摘要：

目的探讨16S rDNA测序法鉴定β-溶血性G群链球菌的价值,通过体外药物敏感性试验确定G群链球菌对青霉素、红霉素和克林霉素等抗菌药物的耐药性,并确定红霉素耐药菌株的耐药表型及基因型。方法采用API 20 Strep链球菌鉴定系统、兰氏抗原血清分型和16S rDNA测序法对复旦大学附属华山医院临床分离的50株β-溶血性G群链球菌进行鉴定。采用琼脂稀释法对该50株G群链球菌进行体外药物敏感性试验,并通过纸片法D试验测定红霉素耐药菌株的耐药表型。通过聚合酶链反应（PCR）检测其大环内酯类耐药基因ermB和mefA。结果50株链球菌经手工API 20 Strep链球菌鉴定系统鉴定,均为停乳链球菌。16S rDNA PCR扩增及测序,42株为停乳链球菌,8株为咽峡炎链球菌,鉴定概率>97%。以测序结果为标准,API 20 Strep链球菌鉴定系统鉴定的准确率为84%。兰氏抗原血清分型均为G群。药物敏感性试验结果显示,50株G群链球菌中没有对左氧氟沙星、头孢曲松、万古霉素和青霉素耐药的菌株。红霉素耐药菌株共28株（56%）,其中cMLs型耐药菌株26株（52%）,Ms型耐药菌株2株（4%）,没有发现iMLs型耐药菌株,红霉素中介菌株4株（8%）。克林霉素耐药菌株共31株（62%）,其中红霉素敏感而克林霉素耐药菌株3株（6%）,红霉素中介而克林霉素耐药菌株2株（4%）。cMLs型耐药菌株中均检测到ermB基因,2株Ms型耐药由mefA基因介导,4株红霉素中介菌株中检出ermB和mefA基因各1株。结论16S rDNA测序法能准确鉴定G群链球菌。G群链球菌对红霉素和克林霉素的耐药率有所提高,但对青霉素敏感,β-内酰胺类药物可以作为治疗G群链球菌感染的首选药物。

关键词: G群链球菌, 药物敏感性试验, 16S rDNA, ermB, mefA

Abstract:

Objective To investigate the significance of 16S rDNA sequencing method in identifying beta-hemolytic Streptococcus G and determine the drug resistance of beta-hemolytic Streptococcus G to antibiotics,like penicillin,erythromycin and clindamycin,by in vitro drug susceptibility test,in order to determine the drug resistance phenotypes and genotypes of erythromycin-resistant isolates. Methods A total of 50 beta-hemolytic Streptococcus G isolates were collected from Fudan University Huashan Hospital,and they were identified by API 20 Strep Streptococcus identification system,Lancefield's streptococcal grouping test and 16S rDNA sequencing method. The in vitro drug susceptibility test was performed by agar dilution method. Erythromycin-resistant phenotypes were determined by D test. The drug resistance genes,ermB and mefA,were determined by polymerase chain reaction（PCR）. Results The 50 beta-hemolytic Streptococcus G isolates were identified by API 20 Strep Streptococcus identification system asStreptococcus dysgalactiae. For 16S rDNA sequencing method,there were 42 isolates of Streptococcus dysgalactiae and 8 isolates of Streptococcus anginosus,and the identification rate was >97%. 16S rDNA sequencing method was used as standard,and the identification accuracy of API 20 Strep Streptococcus identification system was 84%. The results of Lancefield's streptococcal grouping test were all group G. The results of drug susceptibility test showed that there was no levofloxacin-,ceftriaxone-,vancomycin- and penicillin-resistant isolates. Erythromycin resistance was found in 28 isolates（56%）,among them,there were 26 isolates（52%） with cMLs,2 isolates（4%） with Ms and no isolate with iMLs. Furthermore,4 isolates （8%） were intermediately resistant to erythromycin. A total of 31 isolates（62%） were clindamycin-resistant,among them,3 isolates（6%） were sensitive to erythromycin and resistant to clindamycin,and 2 isolates（4%） were intermediately resistant to erythromycin and resistant to clindamycin. All cMLs-resistant isolates carried ermB,and 2 Ms-resistant isolates carried mefA. There were 2 isolates of 4 isolates with intermediate resistance to erythromycin carried ermB and mefA,respectively. Conclusions 16S rDNA sequencing method could identify Streptococcus G. The drug resistance rates to erythromycin and clindamycin increase. However,all the isolates are sensitive to penicillin. The beta-lactam antibiotic may be the first choice for the treatment of Streptococcus G infection.

Key words: Streptococcus G, Drug susceptibility test, 16S rDNA, ermB, mefA

中图分类号:

R446.5

何丽华, 倪丽君, 刘红, 王蓓, 田月如, 郭建, 吴文娟. G群链球菌的分子鉴定及耐药特征分析[J]. 检验医学, 2016, 31(9): 820-824.

HE Lihua, NI Lijun, LIU Hong, WANG Bei, TIAN Yueru, GUO Jian, WU Wenjuan. Identification of Streptococcus G and its drug resistance[J]. Laboratory Medicine, 2016, 31(9): 820-824.

图/表 2

参考文献 15

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抗菌药物	MIC解释标准（μg/mL）
抗菌药物	敏感	中介	耐药
红霉素	≤0.25	0.5	≥1
克林霉素	≤0.25	0.5	≥1
青霉素	≤0.12	-	-
万古霉素	≤1	-	-
左氧氟沙星	≤2	4	≥8
头孢曲松	≤0.5	-	-

抗菌药物	MIC解释标准（μg/mL）
抗菌药物	敏感	中介	耐药
红霉素	≤0.25	0.5	≥1
克林霉素	≤0.25	0.5	≥1
青霉素	≤0.12	-	-
万古霉素	≤1	-	-
左氧氟沙星	≤2	4	≥8
头孢曲松	≤0.5	-	-

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