ABSTRACT
To investigate the effect of CDR1/CDR2 or MDR1 genes overexpression on oxidative stress in Candida albicans,we evaluated the effect of H2O2 on cell viability in C.albicans overexpressing genes CDR1 /CDR2 or MDR1 and their parent strains.After establishing an oxidative stress model with H2O2,we detected reactive oxygen species (ROS),mitochondrial membrane potential (Δψm) and the expression of oxidative stress response-related genes (CAP1 and GRP2) and ROS clearance related-genes (SOD2 and SOD5).The results showed that C.albicans growth were inhibited by 100％ after the treatment of 5 mmol/L H2O2.HeO2 caused more ROS accumulation and Δψm reduction in parent strains than in CDR1/CDR2 or MDR1 genes overexpressed strains (P＜0.05).Compared to parent strains,the up-regulated expression of CAP1 and GRP2 were relatively less in CDR1/CDR2 or MDR1 genes overexpressed strains,moreover,the down-regulated expression of SOD2 and SOD5 were also relatively less in CDR1/CDR2 or MDR1 genes overexpressed strains (P＜0.05).In conclusion,the overexpression of CDR1/CDR2 and MDR1 genes could reduce the oxidative stress response and enhance the adaptability of C.albicans to oxidative stress.
ABSTRACT
Introduction: The commensal yeast Candida albicans, can cause superficial or systemic candidiasis in susceptible hosts. In Chile, azole antifungals are the most widely used drugs in the treatment of candidiasis. In a previous study performed at our center, 2.1 and 1.6% of clinical isolates of C. albicans were found to be resistant to fluconazole and voriconazole, respectively. Objective: To characterize the resistance mechanisms involved in azoles resistance in Chilean clinical isolates. Methodology: Eight resistant, nine susceptible-dose dependent (SDD) and 10 susceptible strains (n: 27) were selected according to the Clinical Laboratory Standards Institute (CLSI) M27-S3 criteria, from vaginal and urine samples. Mutations in the 408-488 region of the ERG11 gene were studied by sequencing, and the relative expression of ERG11 gene and efflux pump genes CDR1, CDR2 and MDR1, was evaluated by quantitative real-time PCR (q-PCR). Results: No mutations were detected in the ERG11 gene and its overexpression was found only in 12.5% of the resistant strains (1/8). The most prevalent mechanism of resistance was the over-expression of efflux pumps (62.5%; 5/8). Conclusion: The study of the expression of efflux pumps by q-PCR could be a useful diagnostic tool for early detection of azole resistance in C. albicans.
Introducción: Candida albicans es una levadura comensal capaz de causar una infección oportunista en hospederos susceptibles denominada candidiasis, que puede ser superficial o sistémica. En Chile, los antifúngicos más utilizados para el tratamiento de las candidiasis son los azoles. En un estudio previo en nuestro centro, se detectó que 2,1 y 1,6% de cepas clínicas de C. albicans fueron resistentes a fluconazol y voriconazol, respectivamente. Objetivo: Caracterizar los mecanismos de resistencia involucrados en la resistencia a azoles en cepas clínicas chilenas. Metodología: Según los criterios del Clinical Laboratory Standards Institute (CLSI) M27-S3, se seleccionaron ocho cepas resistentes, nueve cepas susceptibles dosis dependiente (SDD) y 10 cepas sensibles (n: 27), aisladas de flujo vaginal y orina. Se evaluó la presencia de mutaciones en la región 408-488 del gen ERG11 por secuenciación y la expresión relativa del gen ERG11 y de los genes de bombas de eflujo CDR1, CDR2 y MDR1 por RPC en tiempo real cuantitativa (q-PCR). Resultados: No se encontraron mutaciones en el gen ERG11 y la sobre-expresión de éste sólo se presentó en 12,5% de las cepas resistentes (1/8). El mecanismo prevalente en la cepas resistentes fue la sobre-expresión de bombas de eflujo encontrándose en 62,5% de las cepas resistentes (5/8). Conclusión: El estudio de la expresión bombas de eflujo por q-PCR podría ser una herramienta diagnóstica útil para la detección temprana de resistencia a azoles en C. albicans.
Subject(s)
Female , Humans , Antifungal Agents/pharmacology , Candida albicans/drug effects , Fluconazole/pharmacology , Voriconazole/pharmacology , Chile , Candida albicans/genetics , Candida albicans/isolation & purification , Drug Resistance, Fungal , Gene Expression Regulation, Fungal , Genes, Fungal/genetics , Real-Time Polymerase Chain Reaction , RNA, Fungal/geneticsABSTRACT
ATP binding cassette transporter (ABCT),a membrane transporting protein existing in mammalian, bacterial, fungal and many other types of cells, is correlated with multidrug resistance in many cells. To date 10 ABCT have been described in Candida albicans , but only CDR1 and CDR2 genes were associated with multidrug resistance. Moreover, their encoding proteins Cdr1p and Cdr2p have many functions such as efflux pump and phospholipid translocator. The progress in the study of Cdr1p and Cdr2p proteins mediating multidrug resistance in Candida albicans was summarized in the paper.