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Candida krusei  ( C. krusei ) has been recently recognized as an important pathogen involved in mycotic mastitis of cows. The phenotypic and molecular characteristics of 15  C. krusei  clinical isolates collected from cows with clinical mastitis in three herds of Yinchuan, Ningxia, were identified by matrix-assisted laser desorption ionization–time of flight mass spectrometry. In addition to sequencing analysis, the  ERG11  gene that encodes 14α-demethylases, the expression of the  ERG11  gene, and efflux transporters  ABC1  and  ABC2  in itraconazole-susceptible (S), itraconazole-susceptible dose dependent (SDD), and itraconazole-resistant (R)  C. krusei  isolates was also quantified by a quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) assay. Sequencing analysis revealed three synonymous codon substitutions of the  ERG11  gene including T939C, A756T, and T642C in these  C. krusei  clinical isolates. Among them, T642C and T939C mutations were detected in itraconazole-resistant and -susceptible  C. krusei  isolates, but the A756T substitution was found only in itraconazole-resistant isolates. Importantly, the expression of the  ERG11  gene in itraconazole-resistant isolates was significantly higher compared with itraconazole-SDD and itraconazole-susceptible isolates ( p  = 0.052 and  p  = 0.012, respectively), as determined by the qRT-PCR assay. Interestingly, the expression of the  ABC2  gene was also significantly higher in itraconazole-resistant isolates relative to the itraconazole-SDD and itraconazole-susceptible strains. Notably, the expression of  ERG11  was positively associated with resistance to itraconazole ( p  = 0.4177 in SDD compared with S,  p  = 0.0107 in SDD with R, and  p  = 0.0035 in S with R, respectively). These data demonstrated that mutations of the  ERG11  gene were involved in drug resistance in  C. krusei . The A756T synonymous codon substitution of the  ERG11  gene was correlated with an increased expression of drug-resistant genes including  ERG11  and  ABC2  in itraconazole-resistant  C. krusei  isolates examined in this study.

The A756T Mutation of the ERG11 Gene Associated With Resistance to Itraconazole in Candida Krusei Isolated From Mycotic Mastitis of Cows