阴道白念珠菌毒力因子表达水平及与耐药的相关性

目的 了解复发性念珠菌性阴道炎（RVVC）患者与阴道白念珠菌定植者的阴道白念珠菌的生物膜、水解酶和菌丝壁蛋白1（HWP1）基因等毒力因子的体外表达水平及其耐药性,初步探究白念珠菌致病性和耐药性与毒力因子之间的关系,为进一步探明白念珠菌引起的RVVC的致病和耐药机制提供资料。方法 前瞻性收集分离来源于成年女性RVVC患者和阴道白念珠菌定植者的100株阴道白念珠菌,采用酵母样真菌药物敏感性试剂盒检测对5种常用抗真菌药物的敏感性,采用结晶紫染色法检测其生物膜形成能力,采用牛血清白蛋白平板、卵黄琼脂平板和三丁酸甘油脂平板检测阴道白念珠菌分泌型天冬氨酸蛋白酶（SAP）、磷脂酶（PL）和脂肪酶（Lip）的活性,采用实时荧光PCR检测HWP1基因的表达。结果 5种常用抗真菌药物中至少1种耐药的白念珠菌RVVC患者来源有13株,阴道定植者来源有10株;RVVC患者来源菌株的生物膜、3种分泌型水解酶和HWP1基因体外表达水平均高于阴道定植者来源菌株（均P<0.05）,两组在生物膜形成强弱、3种水解酶表达高低等定性角度,差异均有统计学意义（均P<0.05）;耐药性菌株生物膜表达水平高于非耐药性菌株外（P<0.05）,对3种分泌型水解酶和HWP1基因体外表达水平上两组间没有差异（均P>0.05）;Pearson相关性分析结果显示,除HWP1基因和生物膜-OD之间呈弱相关性（r=0.211,P=0.039）外,其他各项指标之间没有相关性（均|r|<0.2,P>0.05）。结论 白念珠菌的毒力因子生物膜、SAP、 PL、Lip和HWP1都可能在RVVC的致病过程中发挥了重要作用,HWP1与生物膜的形成呈弱相关性,生物膜的形成与阴道白念珠菌的耐药性产生可能存在关联。

Expression level of virulence factors of vaginal Candida albicans and its correlation with drug resistance

Objective To understand resistance and in vitro expression levels of virulence factors such as biofilm, hydrolytic enzyme and hyphal wall protein1 (HWP1) gene of Candida albicans in patients with recurrent vulvovaginal candidiasis (RVVC) and vaginal Candida albicans colonizer. The preliminary exploration of the relationship between the pathogenicity and drug resistance of Candida albicans and provide information for further understanding of the pathogenicity and drug resistance mechanism of RVVC caused by Candida were explored. Methods Prospective collection of 100 vaginal Candida albicans from adult female RVVC patients and colonizers were conducted, the sensitivity to five commonly used antifungal agents was detected by yeastlike fungal drug sensitivity kit, and the biofilm-forming ability was detected by crystal violet staining,the activities of secreted aspartic protease (SAP), phospholipase (PL) and lipase (LIP) were detected by bovine serum albumin (BSA) plate, yolk AGAR plate and tributyrin plate,the expression of HWP1 gene was detected by real-time fluorescent PCR. Results Among the Candida albicans resistant to at least one of the five commonly used antifungal drugs, 13 strains were from RVVC patients and 10 from vaginal colonizers; the expression levels of biofilm in vitro, three hydrolytic enzyme and HWP1 genes in the strains from RVVC patients were significantly higher than those from vaginal colonization (all P<0.05), there were statistically significant differences between the two groups in terms of the qualitative aspects such as the ability of biofilm formation and the expression level of three hydrolytic enzyme (all P<0.05); the biofilm expression level of drug-resistant strains was significantly higher than that of non-resistant strains (P<0.05), and there was no significant difference in the in vitro expression levels of the three secreted hydrolases and HWP1 gene between the two groups (all P>0.05); Pearson correlation analysis showed that, except for the weak correlation between HWP1 gene and biofilm-OD (r=0.211, P=0.039), there was no obvious correlation between other indicators (all |r|<0.2 , P>0.05). Conclusions These virulence factors of Candida albicans may have played an important role in the pathogenesis of RVVC, including biofilm, SAP, PL, Lip and HWP1, there was a weak correlation between HWP1 and the formation of biofilm,the formation may be related to the resistance of vaginal Candida albicans.