Comparison of four molecular typing methods for evaluating genetic diversity among Candida albicans isolates from human immunodeficiency virus-positive patients with oral candidiasis

Abstract

Candida albicans strain delineation by karyotyping, NotI restriction pattern analysis, hybridization with specific probe 27A, and PCR fingerprinting with the phage M13 core sequence were performed with 30 isolates from the oral cavities of 30 human immunodeficiency virus (HIV)-infected patients and 8 reference strains. Within the panel of clinical isolates, 20 were geographically related, although 10 isolates were susceptible to fluconazole and 10 isolates were resistant to fluconazole. The remaining isolates used in this study were fluconazole resistant and geographically unrelated. A composite DNA type was defined for each of the strains as the combination of types obtained by the four molecular methods. By this procedure, a great diversity of DNA types was found among isolates from the oropharynges of HIV-infected individuals with oral candidiasis. This diversity was not reduced when isolates were evaluated on the basis of whether they came from the same geographical locale and whether they were fluconazole resistant. These data refute the idea of a clonal origin for fluconazole-resistant strains among HIV-positive patients. Karyotyping was the least discriminatory method, yielding 19 DNA types among the 38 strains analyzed. Conversely, hybridization with the 27A probe showed a unique DNA pattern for each of the strains examined in this study. Our results demonstrate that at least two different molecular methods are needed for Candida albicans typing and that there is a great deal of strain variation within the species, irrespective of place of origin or antifungal resistance patterns.

Molecular heterogeneity of fluconazole‐resistant and ‐susceptible oral Candida albicans isolates within a single geographic locale

“…3). In a preliminary run, we found that the conditions used were optimal for separation of high, but not low‐molecular‐weight chromosomes as reported by previous workers (7). Thus, in order to achieve better discrimination, electrophoretic parameters were amended as described in the Methods.…”

“…A recent study indicated genotypic clustering among FLZ‐resistant geographically related isolates from HIV‐infected patients, suggesting a common origin of the strains (9). In contrast, other workers who used four independent molecular typing methods (electrophoretic karyotyping, macrorestriction analysis of chromosomal DNA with Not I, hybridization of EcoR I‐digested DNA with the 27A repeat sequence of C. albicans, and PCR fingerprinting) were unable to demonstrate a clonal origin of the FLZ‐resistant strains amongst HIV‐positive patients (7). Therefore, the main objective of the current study was to determine whether geographically related FLZ‐resistant C. albicans isolates in a single locale have a clonal origin.…”

“…Emergence of drug resistance in C. albicans is now a universal problem, especially in patients with immunosuppressive conditions associated with acquired immune deficiency syndrome (AIDS), organ transplantation, and cancer chemotherapy (4–9). A particularly distressing aspect of antifungal therapy is the rapid emergence of C. albicans strains that are resistant to fluconazole (FLZ).…”

Antifungal activity of human salivary mucin‐derived peptide, MUC7 12‐mer, in a murine model of oral candidiasis

“…Abbreviations: AIDS, acquired immune deficiency syndrome; ALP, alkaline phosphatase; ALT, alanine aminotransferase; ANOVA, analysis of variance; AST, aspartate aminotransferase; BUN, blood Introduction Oral candidiasis, as a result of infection by the opportunistic fungus, Candida albicans, is a major cause of oral and esophageal infection in immunocompromised individuals (1,2), including up to 90% of AIDS affected patients (3). Despite excellent pharmacokinetic characteristics and low toxicity of known efficacious antifungal agents like fluconazole, treatment failures are still encountered because of the emergence of antifungalresistant C. albicans (3)(4)(5). The need to develop alternate therapeutic agents has hence soared tremendously.…”

A molecular epidemiological study of sequential oral isolates of Candida albicans from terminally ill patients

“…Theoretically, genotypic approaches should offer more stability and have, in practice, proven to be more discriminatory than phenotypic methods (10). Electrophoretic karyotyping (EK) involves the separation of chromosomalsized DNA molecules by pulsed-field gel electrophoresis (PFGE) and has been used successfully in candidal typing (14,15). An alternative approach is the use of polymerase chain reaction (PCR) based methods such as RAPD-PCR, amplification of hypervariable microsatellite DNA sequences and inter-repeat PCR [IR-PCR; (16,17)].…”