Elias Khajeh1, Seyed Javad Hosseini Shokouh2, Masoumeh Rajabibazl3, Maryam Roudbary4, Sajad Rafiei3, Peyman Aslani5, Zohreh Farahnejad5. 1. a School of Medicine , Tehran University of Medical Sciences , Tehran , Iran. 2. b Department of Infectious Diseases, School of Medicine , AJA University of Medical Sciences , Tehran , Iran. 3. c Department of Clinical Biochemistry , Shahid Beheshti University of Medical Sciences , Tehran , Iran. 4. d Department of Parasitology and Mycology, School of Medicine , Iran University of Medical Sciences , Tehran , Iran. 5. e Department of Medical Mycology and Parasitology , AJA University of Medical Sciences , Tehran , Iran.
Abstract
BACKGROUND: Several studies examined the effect of the Echinophora platyloba extract in treatment of azole-resistant Candida albicans clinical isolates. OBJECTIVE: We investigated the effect of E. platyloba extract on expression of CDR1 and CDR2 genes in fluconazole-resistant clinical isolates of C. albicans using real-time PCR. MATERIALS AND METHODS: The crude extract of E. platyloba was obtained using percolation method. Using serial dilution method, different concentrations of extract were achieved. Two hundred microlitres of fungal suspension (10(6) CFU/ml) was added to the media and cultured with different concentrations and then incubated at 37 °C for 48 h. The concentration of extract in the first tube, which inhibited the growth of C. albicans, was recorded as the Minimal Inhibitory Concentration (MIC). In order to analyse the expression of CDR1 and CDR2 genes, RNA was extracted from C. albicans isolates before and after treatment with MIC of E. platyloba using glass beads and the denaturing buffer agents in an RNase-free environment and then the cDNA was synthesised and used for real-time PCR assay. RESULTS: Twenty of total of 148 isolates were resistant to fluconazole. The MIC and MFC for the alcoholic extract of E. Platyloba were 64 mg/ml and 128 mg/ml, respectively. Real-time PCR results revealed that the mRNA levels of CDR1 and CDR2 genes significantly declined after incubation with E. Platyloba (both p values < 0.001). CONCLUSION: E. Platyloba is effective in reducing CDR1 and CDR2 expression which in turn plays an important role in fluconazole resistance in Candida species.
BACKGROUND: Several studies examined the effect of the Echinophora platyloba extract in treatment of azole-resistant Candida albicans clinical isolates. OBJECTIVE: We investigated the effect of E. platyloba extract on expression of CDR1 and CDR2 genes in fluconazole-resistant clinical isolates of C. albicans using real-time PCR. MATERIALS AND METHODS: The crude extract of E. platyloba was obtained using percolation method. Using serial dilution method, different concentrations of extract were achieved. Two hundred microlitres of fungal suspension (10(6) CFU/ml) was added to the media and cultured with different concentrations and then incubated at 37 °C for 48 h. The concentration of extract in the first tube, which inhibited the growth of C. albicans, was recorded as the Minimal Inhibitory Concentration (MIC). In order to analyse the expression of CDR1 and CDR2 genes, RNA was extracted from C. albicans isolates before and after treatment with MIC of E. platyloba using glass beads and the denaturing buffer agents in an RNase-free environment and then the cDNA was synthesised and used for real-time PCR assay. RESULTS: Twenty of total of 148 isolates were resistant to fluconazole. The MIC and MFC for the alcoholic extract of E. Platyloba were 64 mg/ml and 128 mg/ml, respectively. Real-time PCR results revealed that the mRNA levels of CDR1 and CDR2 genes significantly declined after incubation with E. Platyloba (both p values < 0.001). CONCLUSION:E. Platyloba is effective in reducing CDR1 and CDR2 expression which in turn plays an important role in fluconazole resistance in Candida species.