Combination Of Itraconazole And Tacrolimus Against Candida Krusei And Analysis Of Mutations In ERG11 Gene Of Candida Krusei By Itraconazole Resistance

Objective The use of the antifungal agent "Itraconazole" and the calcineurin inhibitor "Tacrolimus" alone,together with the above two drugs,was evaluated in vitro for growth of Candida krusei.Whether or not the combination is more effective than the antifungal alone can inhibit the growth of Candida krusei.Methods According to the guidelines of the American Institute of Clinical and Laboratory Standards,12 Candida krusei strains were divided into blank control group,itraconazole group,tacrolimus group,and combination group by microdilution method,and then 12 The Candida krusei strain was incubated for 24 h and its growth was observed.Results In comparison with the alone use of isotriconazole in Candida krusei,the combined use of itraconazole,its minimal inhibitory concentration was much less than that of alone,with a maximum multiple of 32 fold,a minimum multiple up to 4 times;the same,In comparison with the alone use of tacrolimus in Candida krusei,the combined use of tacrolimus,its maximum multiple of 32 times,the lowest multiple of 2 times.Conclusion The combined use of itraconazole and tacrolimus in 12 strains of Candida krusei displayed a powerful synergistic effect in vitro.Objective To investigate the relationship between ERG11 gene mutation and its resistance to itraconazole in ergosterol synthesis pathway in Candida krusei.Methods From January 2014 to December 2016,12 clinical isolates of Candida krusei were collected from the clinical laboratory of Anhui Medical University First Affiliated Hospital.The strains were derived from sputum,urine,feces and secretions.There were 5 strains of itraconazole-resistant,2 strains of drug-dependent and 1 strain of susceptible strain.Then,Candida krusei was broken to extract DNA,and PCR was performed on the ERG11 gene.DNA sequencing was performed on the PCR.Finally,the sequencing results of PCR were analyzed and compared using BLAST software.Results After sequencing of the ERG11 gene amplified by Candida krusei,we found that 3 synonymous codons were substituted in ERG11 in C.krusei strains(including T289 C,T649C,and A983T).Of the 3 synonymous mutations,T649 C and A983 T appeared only in itraconazole-resistant strains,whereas T289 C appeared in non-resistant itraconazole-resistant strains and resistant Candida krusei strains.Conclusion In our study,no missense mutations were found in Candida krusei isolates,but three new synonymous codon substitutions were found in the ERG11 gene of Candida krusei strain,of which T649 C and A983 T only appeared at traconazole-resistant strains,suggesting that silent mutations might be involved in resistance mechanisms krusei strains.