title: Overexpression of Both ERG11 and ABC2 Genes Might Be Responsible for Itraconazole Resistance in Clinical Isolates of Candida krusei.
abstract: OBJECTIVE
  To study the main molecular mechanisms responsible for itraconazole resistance in clinical isolates of Candida krusei.
  
  
  METHODS
  The 14α-demethylases encoded by ERG11 gene in the 16 C.krusei clinical isolates were amplified by polymerase chain reaction (PCR), and their nucleotide sequences were determined to detect point mutations. Meanwhile, ERG11 and efflux transporters (ABC1 and ABC2) genes were determined by quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) for their expression in itraconazole-resistant (R), itraconazole-susceptible dose dependent (SDD) and itraconazole-susceptible (S) C.krusei at the mRNA level.
  
  
  RESULTS
  We found 7-point mutations in ERG11 gene of all the C.krusei clinical isolates, including 6 synonymous mutations and 1 missense mutation (C44T). However, the missense mutation was found in the three groups. The mRNA levels of ERG11 gene in itraconazole-resistant isolates showed higher expression compared with itraconazole-susceptible dose dependent and itraconazole-susceptible ones (P = 0.015 and P = 0.002 respectively). ABC2 gene mRNA levels in itraconazole-resistant group was significantly higher than the other two groups, and the levels of their expression in the isolates appeared to increase with the decrease of susceptibility to itraconazole (P = 0.007 in SDD compared with S, P = 0.016 in SDD with R, and P<0.001 in S with R respectively). While ABC1 gene presented lower expression in itraconazole resistant strains. However, the mRNA levels of ERG11, ABC1 and ABC2 in a C.krusei (CK10) resistant to both itraconazole and voriconazole were expressed highest in all the itraconazole-resistant isolates.
  
  
  CONCLUSIONS
  There are ERG11 gene polymorphisms in clinical isolates of C.krusei. ERG11 gene mutations may not be involved in the development of itraconazole resistance in C.krusei. ERG11 and ABC2 overexpression might be responsible for the acquired itraconazole resistance of these clinical isolates.
authors: Department of Hematology, Tianjin First Central Hospital, The First Central Clinical College of Tianjin Medical University, Tianjin, China.
authors: Xiaoyuan
authors: X
authors: He
authors: Department of Hematology, Tianjin First Central Hospital, The First Central Clinical College of Tianjin Medical University, Tianjin, China.
authors: Mingfeng
authors: M
authors: Zhao
authors: Department of Clinical Laboratory, Tianjin First Central Hospital, Tianjin, China.
authors: Jinyan
authors: J
authors: Chen
authors: Department of Hematology, Tianjin First Central Hospital, The First Central Clinical College of Tianjin Medical University, Tianjin, China.
authors: Rimao
authors: R
authors: Wu
authors: Department of Clinical Laboratory, Tianjin First Central Hospital, Tianjin, China.
authors: Jianlei
authors: J
authors: Zhang
authors: Department of Hematology, Tianjin First Central Hospital, The First Central Clinical College of Tianjin Medical University, Tianjin, China.
authors: Rui
authors: R
authors: Cui
authors: Department of Hematology, Tianjin First Central Hospital, The First Central Clinical College of Tianjin Medical University, Tianjin, China.
authors: Yanyu
authors: Y
authors: Jiang
authors: Department of Hematology, Tianjin First Central Hospital, The First Central Clinical College of Tianjin Medical University, Tianjin, China.
authors: Jie
authors: J
authors: Chen
authors: Department of Hematology, Tianjin First Central Hospital, The First Central Clinical College of Tianjin Medical University, Tianjin, China.
authors: Xiaoli
authors: X
authors: Cao
authors: Department of Hematology, Tianjin First Central Hospital, The First Central Clinical College of Tianjin Medical University, Tianjin, China.
authors: Yi
authors: Y
authors: Xing
authors: Department of Hematology, Tianjin First Central Hospital, The First Central Clinical College of Tianjin Medical University, Tianjin, China.
authors: Yuchen
authors: Y
authors: Zhang
authors: Department of Hematology, Tianjin First Central Hospital, The First Central Clinical College of Tianjin Medical University, Tianjin, China.
authors: Juanxia
authors: J
authors: Meng
authors: Department of Hematology, Tianjin First Central Hospital, The First Central Clinical College of Tianjin Medical University, Tianjin, China.
authors: Qi
authors: Q
authors: Deng
authors: Department of Hematology, Tianjin First Central Hospital, The First Central Clinical College of Tianjin Medical University, Tianjin, China.
authors: Tao
authors: T
authors: Sui
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000378:metabolism*; D000935:Antifungal Agents / Q000494:pharmacology; D002175:Candida / Q000187:drug effects / Q000254:growth & development / Q000302:isolation & purification* / Q000378:metabolism*; D002177:Candidiasis / Q000188:drug therapy / Q000378:metabolism / Q000382:microbiology*; D025141:Drug Resistance, Fungal; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D015966:Gene Expression Regulation, Fungal; D006801:Humans; D017964:Itraconazole / Q000494:pharmacology*; D008826:Microbial Sensitivity Tests; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D018528:ATP-Binding Cassette Transporters; D000935:Antifungal Agents; D005656:Fungal Proteins; D012333:RNA, Messenger; D017964:Itraconazole