Identification of Candida spp. by Randomly Amplified Polymorphic DNA Analysis and Differentiation between Candida albicans and Candida dubliniensis by Direct PCR Methods

Abstract

Because Candida species have innately highly variable antifungal susceptibilities, the availability of a fast and reliable species identification test is very important so that suitable and effective therapeutic measures may be taken. Using three oligonucleotide primers, we established a randomly amplified polymorphic DNA (RAPD) analysis method that enabled direct identification of the most common opportunistic pathogenic Candida species. RAPD analysis revealed a characteristic molecular fingerprint for each Candida species. Differences between the profiles for Candida albicans and C. dubliniensis were evident. RAPD analysis is a relatively easy, reproducible, and reliable technique that can be useful in providing genetic fingerprints for the identification of strains. In addition, a collection of different C. albicans strains was identified by a specific PCR based on multiple secreted aspartic proteinase (SAP) genes and the dipeptidyl aminopeptidase (DAP2) gene. Our findings demonstrate that PCR based upon the SAP and DAP2 sequences is a simple, rapid, clear, and direct technique for the identification and differentiation of C. albicans and C. dubliniensis.

Phylogeny and evolution of the aspartyl protease family from clinically relevant Candida species

“…SAP gene distribution among C. dubliniensis (Gilfillan et al 1998), C. tropicalis (this work) and C. albicans populations seems to be universal (Bautista et al 2003); however, a differential distribution of SAP1-3 and SAP4-6 gene families in C. albicans strains isolated from vaginal and systemic hematogenous infections has been observed (Kalkanci et al 2005). The clonal structure of reproduction of C. albicans populations is typical of an asexual fungus and limits their capacity to generate diversity (Lott & Effat 2001, Tavanti et al 2004); thus, gene duplication and rapid differentiation must be mechanisms that promote diversity.…”

Candida nivariensis Isolated from an Indonesian Human Immunodeficiency Virus-Infected Patient Suffering from Oropharyngeal Candidiasis

“…Further physical examination revealed rales in both lungs, with no wheezing and no lymph node enlargement. Laboratory investigations revealed values for hemoglobin of 6.7 g/dl, a leukocyte count of 6,500/mm 3 , a platelet count of 220,000/mm 3 , and a CD4 count of 104 cells/mm 3 . The patient died before undergoing complete examination and having proper treatment.…”

The Proteolytic System of Candida dubliniensis

“…The C. dubliniensis strains were confirmed to the species level using molecular methods described by Bautista et al [10] . They were routinely maintained on YEPD plates (1% yeast extract, 2% peptone, 2% glucose, and 2% agar) at 28ºC.…”

“…Hybridization and immunological detection were performed as recommended by the supplier. SAP DNA probes to be used in hybridization experiments were generated for PCR from C. albicans SAP genes: SAP1, SAP2, SAP3, or SAP4-6 as described by Bautista-Muñoz et al [10] . Genomic DNA from C. albicans ATCC 10231 was used as template.…”