Development of a new real-time TaqMan PCR assay for quantitative analyses of Candida albicans resistance genes expression.
Candida albicans is an important opportunistic pathogen that can cause serious fungal diseases in immunocompromised patients including cancer patients, transplant patients, and patients receiving immunosuppressive therapy in general, those with human immunodeficiency virus infections and undergoing major surgery. Its emergence spectrum varies from mucosal to systemic infections and the first line treatment is still based on fluconazole, a triazole derivate with a potent antifungal activity against most of C. albicans strains. Nevertheless the emergence of fluconazole-resistant C. albicans strains can lead to treatment failures and thus become a clinical problem in the management of such infections. For that reason we consider it important to study mechanisms inducing azole resistance and the possibilities to influence this process. In this work we give a short report on a real-time PCR (TaqMan) assay, which can be used for quantitative analyses of gene expression levels of MDR1, CDR1 and ERG11, genes supposed to contribute to development of the resistance mechanisms. We show some results achieved with that assay in fluconazole susceptible and resistant strains that confirm results seen earlier in experiments using Northern blot hybridisation and prove that the comparative DeltaCt method is valid for our system.
Kofla
Grzegorz
G
Humboldt University Berlin, Department of Medicine 2, Division of Oncology/Hematology, Charité Campus Mitte, Chariteplatz 1, 10117 Berlin, Germany. grzegorzkof@yahoo.de
Ruhnke
Markus
M
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000737:chemistry / Q000235:genetics; D000935:Antifungal Agents / Q000494:pharmacology*; D002176:Candida albicans / Q000187:drug effects / Q000235:genetics* / Q000254:growth & development / Q000302:isolation & purification; D002180:Candidiasis, Oral / Q000382:microbiology*; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry / Q000235:genetics; D025141:Drug Resistance, Fungal / Q000235:genetics*; D015725:Fluconazole / Q000494:pharmacology*; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics; D006801:Humans; D026901:Membrane Transport Proteins / Q000737:chemistry / Q000235:genetics; D008826:Microbial Sensitivity Tests; D010088:Oxidoreductases / Q000737:chemistry / Q000235:genetics; D012331:RNA, Fungal / Q000737:chemistry / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D058886:Sterol 14-Demethylase
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000935:Antifungal Agents; C094693:CDR1 protein, Candida albicans; C548418:CYP51A1 protein, human; D005656:Fungal Proteins; D026901:Membrane Transport Proteins; D012331:RNA, Fungal; D015725:Fluconazole; D003577:Cytochrome P-450 Enzyme System; D010088:Oxidoreductases; D058886:Sterol 14-Demethylase