Analysis Of ERG11 Gene Mutation In ITR-resistance Candida Krusei

Invasive fungal infections(IFIs)have increased dramatically, following the widespread application of broad-spectrum antibiotics, immunodepressants, therapeusis of intervention and organ transplantation. Candida albicans remains the most common conditional pathogenic fungi isolated from clinical samples, however, there has been a significant increase in the frequency of non-albicans Candida species isolation in recent years. And the utilization of antifungal agents promotes the process of resistance formation. Drug-resistant strain is rising in non-albicans Candida species isolated from clinical samples. non-albicans Candida species resistance is an arising medical problem. More and more researchers pay their attention to the molecular mechanisms of non-albicans Candida species resistance to antifungal agents,especially to azoles.ERG11 gene is the encoding gene of 14 alpha-demethylase(Erg11p),Erg11p is a key enzyme in the ergosterol synthesis pathway of Candida species. Ergosterol is essential for maintaining the integrity and function of Candida species membrane. The study is more about Candida albicans,there was no report concerning identification of resistant isolates of Candida krusei with ERG11.Objective:To study the relationship between itraconazole (ITR)-resistance of clinical Candida krusei isolates and the mutation of ERG11 gene encoding ITR-targeted enzyme. azoles can bind with the enzyme and block ergosterol synthesis. If one or more mutations in ERG11 result in changes in the Erg11p spatial configuration, a decrease in the affinity between the azole and protein occurs. This altered phenotype often makes isolates resistant to azole.Methods:From June 2009 to October 2010, Three strains of ITR-susceptible and six ITR-resistent C. krusei were isolated from the respiratory tract,urethra and vagina samples, in the Second Hospital of Shanxi Medical University. ERG11 gene was amplified by PCR using C. krusei genomic DNA extracts as the templates and the DNA sequences of the PCR products were determined and compared using BLAST softwares.Results:The comparison of ERGll gene sequences identified mutations at 7 sites in 10 strains,including 7 same-sense mutations.Base substitutions at the sites of 447bp may take place in both drug-resistant and drug-susceptible strains.The point mutation at the site of 12bp,327bp,429bp,552bp take place in ITR-resistant strain. The point mutation at the site of 696bp,927bp take place in ITR-susceptible strain.Conclusions:The results indicate that there is Sequence Polymorphism at DNA level in clinical strains of C. krusei which are resistant to itraconazole.ERG11 gene mutations are not found to be involved in the development of itraconazole resistance in C. krusei in this study.