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目的:应用多位点序列分型(MLST)分析38株热带假丝酵母菌的亲缘关系,体外比较热带假丝酵母菌对常见抗真菌药物的敏感性,检测耐药基因的表达水平。方法:通过对6对等位基因进行测序,用e BURST分组和非加权配对算术平均法(UPGMA)聚类分析菌株的进化和变异。微量肉汤稀释法进行体外抗真菌药物敏感试验,实时荧光定量聚合酶链反应(q-RT-PCR)检测耐药基因CDR1、CDR2、ERG1、ERG3、ERG6、ERG9和ERG11表达。结果:38株菌株中获得30个二倍体序列(DST),其中14个新DST,11个新等位基因,27个DST为单体型。e BURST分组,主要分成2组,大多数DST属于单体型。聚类分析总共分18个进化枝。阿尼芬净、米卡芬净、卡泊芬净、氟康唑、伏立康唑、伊曲康唑、两性霉素B、氟胞嘧啶、咪康唑、布康唑、克霉唑、特康唑、特比萘芬和制霉菌素14种抗真菌药物的MIC90分别为0.030μg/ml、0.015μg/ml、0.500μg/ml、1.000μg/ml、0.125μg/ml、0.250μg/ml、1.000μg/ml、0.250μg/ml、2.000μg/ml、2.000μg/ml、0.250μg/ml、0.250μg/ml、256.000μg/ml和8.000μg/ml。绝大多数耐药基因表达量较高的菌株可能在进化枝11和14。结论:热带假丝酵母菌MLST变异较大,菌株之间遗传距离较远。热带假丝酵母菌对特比萘芬普遍耐药,对目前其他常用的抗真菌药物多为敏感。
OBJECTIVE: To analyze the genetic relationship of 38 strains of Candida tropicalis by multilocus sequence typing (MLST). To compare the susceptibility of Candida tropicalis to common antifungal agents in vitro and to detect the expression of drug resistance genes. Methods: Six pairs of alleles were sequenced and the evolution and variation of the strains were clustered by e BURST grouping and UPGMA (Unweighted Pairwise Arithmetic Average Method). In vitro antifungal susceptibility test was carried out by using the broth microdilution method. The expression of CDR1, CDR2, ERG1, ERG3, ERG6, ERG9 and ERG11 were detected by real-time quantitative polymerase chain reaction (qRT-PCR). RESULTS: Thirty diploid sequences (DSTs) were obtained from 38 isolates, of which 14 new DST, 11 new alleles and 27 DST haplotypes. e BURST groupings are divided into 2 groups, most of which belong to haplotypes. Cluster analysis is divided into a total of 18 clades. Anidine net, micafungin, caspofungin, fluconazole, voriconazole, itraconazole, amphotericin B, flucytosine, miconazole, butoconazole, clotrimazole, , Terbinafine and nystatin were 0.030 μg / ml, 0.015 μg / ml, 0.500 μg / ml, 1.000 μg / ml, 0.125 μg / ml, 0.250 μg / ml and 1.000 μg ml, 0.250 μg / ml, 2.000 μg / ml, 2.000 μg / ml, 0.250 μg / ml, 0.250 μg / ml, 256.000 μg / ml, and 8.000 μg / ml. Strain 11 and 14 may be the most resistant strains. Conclusion: MLST variation of Candida tropicalis is large, and the genetic distance between strains is far. Candida tropicalis is generally resistant to terbinafine and is more susceptible to other commonly used antifungal agents.