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Comparison of four molecular approaches to identify Candida parapsilosis complex species.

ABSTRACT: Since the description of Candida orthopsilosis and C. metapsilosis in 2005, several methods have been proposed to identify and differentiate these species from C. parapsilosis sensu stricto. Species-specific uniplex polymerase chain reaction (PCR) was performed and compared with sequencing of the D1/D2 region of the LSU 28S rDNA gene, microsatellite typing of C. parapsilosis sensu stricto, and PCR-restriction fragment length polymorphism patterns in the ITS1-5.8S-ITS2 region of the rDNA gene. There was agreement between results of testing of 98 clinical isolates with the four PCR-based methods, with 59 isolates identified as C. parapsilosis sensu stricto, 37 as C. orthopsilosis, and two as C. metapsilosis.

SUBMITTER: Barbedo LS

PROVIDER: S-EPMC5319372 | biostudies-literature | 2017 Mar

REPOSITORIES: biostudies-literature

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Comparison of four molecular approaches to identify Candida parapsilosis complex species.

Barbedo Leonardo Silva LS Figueiredo-Carvalho Maria Helena Galdino MH Muniz Mauro de Medeiros MM Zancopé-Oliveira Rosely Maria RM

Memorias do Instituto Oswaldo Cruz 20170216 3

Since the description of Candida orthopsilosis and C. metapsilosis in 2005, several methods have been proposed to identify and differentiate these species from C. parapsilosis sensu stricto. Species-specific uniplex polymerase chain reaction (PCR) was performed and compared with sequencing of the D1/D2 region of the LSU 28S rDNA gene, microsatellite typing of C. parapsilosis sensu stricto, and PCR-restriction fragment length polymorphism patterns in the ITS1-5.8S-ITS2 region of the rDNA gene. Th ...[more]

PMID: 28225905

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Project description:The Candida parapsilosis group encompasses three species: C. parapsilosis, C. orthopsilosis, and C. metapsilosis. Here, we describe the incidence and echinocandin susceptibility profile of bloodstream isolates of these three species collected from patients admitted to an Italian university hospital from 2007 to 2014. Molecular identification of cryptic species of the C. parapsilosis complex was performed using polymerase chain reaction amplification of the gene encoding secondary alcohol dehydrogenase, followed by digestion with the restriction enzyme BanI. Minimum inhibitory concentrations were determined using the broth microdilution method according to European Committee for Antimicrobial Susceptibility Testing (EUCAST EDef 7.2) and Clinical Laboratory Standards Institute (CLSI M27-A3) guidelines, and the results were compared with those obtained using the E-test and Sensititre methods. Of the 163 C. parapsilosis complex isolates, 136 (83.4%) were identified as C. parapsilosis, and 27 (16.6%) as C. orthopsilosis. The species-specific incidences were 2.9/10,000 admissions for C. parapsilosis and 0.6/10,000 admissions for C. orthopsilosis. No resistance to echinocandins was detected with any of the methods. The percent essential agreement (EA) between the EUCAST and E-test/Sensititre methods for anidulafungin, caspofungin, and micafungin susceptibility was, respectively, as follows: C. parapsilosis, 95.6/97.8, 98.5/88.2, and 93.4/96.3; C. orthopsilosis, 92.6/92.6, 96.3/77.8, and 63.0/66.7. The EA between the CLSI and E-test/Sensititre methods was, respectively, as follows: C. parapsilosis, 99.3/100, 98.5/89.0, and 96.3/98.5; C. orthopsilosis, 96.3/92.6, 100/81.5, and 92.6/88.9. Only minor discrepancies, ranging from 16.9% (C. parapsilosis) to 11.1% (C. orthopsilosis), were observed between the CLSI and E-test/Sensititre methods. In conclusion, this epidemiologic study shows a typical C. parapsilosis complex species distribution, no echinocandin resistance, and it reinforces the relevance of using commercially available microbiological methods to assess antifungal susceptibility. These data improve our knowledge of the national distribution of species of the psilosis group, as there are very few studies of these species in Italy.

Project description:Background and Purpose:Candida parapsilosis complex isolates are mainly responsible for nosocomial catheter-related infection in immunocompromised patients. Biofilm formation is regarded as one of the most pertinent key virulence factors in the development of these emerging infections. The present study aimed to compare in vitro antifungal susceptibility patterns and biofilm-related genes expression ratio in planktonic and biofilm's cells of clinically C. parapsilosis complex isolates. Materials and Methods:The current study was conducted on a number of 17 clinical C. parapsilosis complex (10 C. parapsilosis sensu stricto, 5 C. orthopsilosis, and 2 C. metapsilosis). The antifungal susceptibility patterns of amphotericin B, ?uconazole, itraconazole, voriconazole, posaconazole, and caspofungin in planktonic and biofilm forms were closely examined using CLSI M27-A3 broth microdilution method. The expression levels of biofilm-related genes (BCR1, EFG1, and FKS1) were evaluated in planktonic and biofilm's cells using Real-time polymerase chain reaction (PCR) technique. Results:The obtained results indicated that all C. parapsilosis complex isolates were able to produce high and moderate amounts of biofilm forms. In addition, the sessile minimum inhibitory concentrations were reported to be high for fluconazole (? 64 µg/ml), itraconazole, voriconazole, and posaconazole (? 16 µg/ml), as compared to planktonic minimum inhibitory concentrations. Moreover, a significant difference was observed between antifungal susceptibility patterns for all azole antifungal agents (P<0.05). Furthermore, the BCR1 overexpression was considered significant in biofilms with regard to planktonic cells in C. parapsilosis species complex (P=0.002). Conclusion:C. parapsilosis complex isolates were found susceptible to most of the tested antifungal drugs, while biofilms demonstrated a noticeable resistant to azoles. The marked discrepancy noted in antifungal susceptibility patterns among these species should be highlighted to achieve effective therapeutic treatment.

Dataset Information

Comparison of four molecular approaches to identify Candida parapsilosis complex species.

Publications

Comparison of four molecular approaches to identify Candida parapsilosis complex species.

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