| Objectives: ①To investigate Candida species and in vitro antifungal susceptibility pattern of Candida isolates obtained from vulvovaginal candidiasis patients against seven antifungal agents.②To investigate Candida genotype distribution and relation of genotype with antifungal minimal inhibitory concentration (MICs) and susceptibility among acute Vulvovaginal candidiasis (VVC) and recurrent VVC (RVVC).Methods and material:①Candida isolates were collected at obstetrics and gynecology clinic of first affiliated hospital of Kunming medical university from VVC and RVVC patients. VITEK(?)2 Compact system and chromogenic media (CHROMagar and Autobio) were used for identification of the Candida species. ②Antifungal susceptibility test was carried out according to the test method outlined in CLSI document M27-A3. According to the minimum inhibitory concentration (MIC) of the drugs, the suscepitibility of drug was determined. ③DNA extraction was performed using AxyPrep Blood Genomic DNA miniprep kit. Candida genotype was investigated on the basis of 25S rDNA. SPSS version 17.0 was used for statistical analysis, Kruskal-Wallis H test, Mann-Whitney U test and Fisher’s Exact test was performed, P<0.05 was considered statistically significant. Frequency tables and graphs were used to summarize the data.Results:C. albicans (335) was the most common species isolated among 384 Candida isolates followed by Candida glabrata (22), Candida tropicalis (15) and Candida krusei (12). C. albicans was predominant in both the cases VVC and RVVC. Out of 229 VVC isolates,196 (85.6%) were identified as C. albicans and among 155 RVVC isolate, C. albicans identified was 139 (89.7%). In VVC cases, C. albicans was followed by C. glabrata (7.4%), C. tropicalis (3.9%) and C. krusei (3.1%). But in RVVC cases, C. tropicalis (3.9%) was predominant among non-albicans and was followed by C. glabrata (3.2%), and C. krusei (3.2%).Candida albicans was also the predominant species in pregnant (88.7%) as well as non-pregnant group (87.0%). The following non-albicans species isolated were; Candida glabrata (6.8%), Candida tropicalis (4.0%) and Candida krusei (2.2%) in non-pregnant group. On the contrary, Candida krusei was predominant non-albicans species followed by Candida tropicalis (3.2%) and none of the non-albicans isolated belonged to Candida glabrata in pregnant group.The susceptibility result among C. albicans and non-albicans demonstrated 85.4%(286/335) of the C. albicans was susceptible compared to 28.6%(14/49) of non-albicans for fluconazole (P<0.001). Similarly,76.1%(255/335) of the C. albicans was susceptible compared to 61.2% (30/49) of non-albicans towards clotrimazole (P=0.011). On the contrary, against miconazole, non-albican was more susceptible (P<0.001), with 95.9%(47/49) being susceptible compared to 72.5%(243/335) of C. albicans. Also for itraconazole, non-albicans was more susceptible (P=0.003), with 63.3%(31/49) being susceptible compared to 35.2%(118/335) of C. albicans.The susceptibility result among non-albicans showed significant obvious difference in susceptibility for fluconazole, C. tropicalis (14/15,93.3%) being more susceptible (P<0.001) than C. glabrata. Majority of all three non-albicans (C. glabrata, C. krusei and C. krusei) exhibited high susceptibility towards 5-fluorocytosine, miconazole and amphotericin B.Among WC and RVVC, C. albicans in VVC group was found to be significantly more susceptible (P=0.04) against FCA (174/335,88.8%) than in RVVC group (112/335,80.6%). However demonstrated significantly lower susceptibility against MCZ (P=0.001) and AMB (P=0.007) in VVC group. Other two antifungal agents,5-fluorocytosine and clotrimazole demonstrated high susceptibility among both VVC and RVVC group with>80% isolates being susceptible to these drugs. In contrast, nystatin and itraconazole demonstrated less susceptibility in both the groups with resistance rate being >40%.Isolates from pregnant patients were found to be more susceptible than from non-pregnant patients towards MCZ (P<0.001) and AMB (P=0.028) in VVC group, and MCZ (P=0.018), ITR (P=0.013) and NYS (P=0.006) in RVVC group.Out of 335 C. albicans isolates,309 (92.2%),19 (5.6%) and 7 (2%) isolates were identified as genotype A (450 bp), B (840 bp) and C (450 bp and 840 bp) respectively. None of the PCR products in this study were 1040 bp and 1080 bp long that corresponds to C. dubliniensis. Genotype A was prevalent in both VVC and RVVC (P=0.061). Out of 196 VVC cases,89.8% (176/196) was genotype A and 10.2%(20/196) was type B and C. In 139 RVVC cases,95.7% (133/139) was due to genotype A and 4.3%(6/139) was due to type B and C. All non-albicans isolates produced 450 bp PCR products except for 1 C. glabrata and 2 C. krusei, which gave 840 bp long products.Analysis of the correlation between genotypes and antifungal susceptibility demonstrated genotype B and C (16/26,61.5%) was more susceptible than genotype A (S=103/309,33.3%) against itraconazole (P=0.003). To 5-fluorocytosine, all isolates with 840 bp products including three non-albicans were susceptible.The MICs of genotype B and C against 5-fluorocytosine (P=0.008) and itraconazole (P=0.003) were observed to be lower than genotype A. The lowest MICs observed for genotype A was against clotrimazole and amphotericin B (MIC90=1μg/ml) while for other two types was against 5-fluorocytosine (MIC90=0.475μg/ml). The highest MICs observed was against nystatin, miconazole and itraconazole for type A (MIC90=16μg/ml) while for type B and C, it was against nystatin (MIC90=10.4μg/ml).Conclusion:①C. albicans was the most commonly isolated yeast responsible for VVC and RVVC in pregnant and non-pregnant women.②Fluconazole and clotrimazole may be less effective in non-albicans infection as for significant in-vitro resistance found in non-albicans. However, miconazole and itraconazole may be effective for non-albicans than C. albicans infection. ③Among VVC and RVVC, susceptibility in non-albicans did not show any difference. C. albicans in RWC showed decreased susceptibility compared to WC for fluconazole, which warns us its future use. On the other hand, miconazole and amphotericin B may be less effective in WC than RVVC due to C. albicans. ④Compared to non-pregnant patients, Nystatin, amphotericin B, miconazole and itraconazole were significantly more susceptible in isolates from pregnant patients. Except for itraconazole, all other azoles showed high susceptibility and can be effective in pregnancy.⑤Genotype A was the predominant C. albicans strain followed by type B and C. Genotype showed no association with VVC type however showed to have impact on drug MICs and susceptibility. Presence of group 1 intron may be the most important factor contributing to 5-fluorocytosine susceptibility as all 840 bp products of C. albicans as well as non-albicans, which actually determines that the strain contains group 1 intron, were all susceptible to 5-fluorocytosine. |
PDF Full Text Request