Phenotyping by Fourier-transform infrared spectroscopy versus restriction- and amplification-based genotyping of Candida tropicalis isolates: a comparison
Abstract number: 902_p1782
Al Masri A.S.
Invasive candidiasis has increased dramatically over the past decades. The contribution of non-albicans Candida spp. including C. tropicalis to invasive infections is rising, yet little is known of their molecular epidemiology. Therefore, effective typing systems are required for tracing epidemiological distribution patterns.
In addition to restriction endonuclease analysis of genomic DNA (REAG) followed by pulsed-field gel electrophoresis (PFGE), different amplification-based methods were studied: Arbitrarily primed (AP)-PCR under low-stringency conditions with a set of 38 different 10mer arbitrary oligonucleotides (G + C content: 3090%), interrepeat (IR)-PCR based on micro- and minisatellite sequences and eucaryotic telomeric motifs using eight different primers. These results were compared with those obtained by phenotyping the isolates with Fourier-transform infrared spectroscopy (FTIR). For fingerprinting, 60 C. tropicalis reference strains and clinical isolates from different European and Asian centers were included into the study.
The rank order of discriminatory ability among the genotyping methods was as follows: REAG using BssH II >> AP-PCR > IR-PCR. In contrast to other restriction enzymes used, BssH II revealed a series of distinct bands in the region from 100300 bp to approximately 23 kb. Regarding AP-PCR fingerprinting, random primers with a G + C content of 50% showed the best discriminatory power if prolonged ramp times (5 min) were applied. In contrast, IR-PCR was shown to be not suitable for genotyping C. tropicalis isolates. Using the Ward algorithm and the first derivates of the spectra, FTIR showed a main branching into two groups, which was further divided in several subgroups depending on the heterogeneity level selected. In general, FTIR was less discriminatory than REAG, but easier to perform. The classifications derived from REAG and FTIR were not completely congruent. Strains representing the same major profile but being epidemiologically unrelated appeared to be widespread in several centres included.
These findings indicate that REAG followed by PFGE is the most useful molecular method for the investigation of inter-strain variation within the species C. tropicalis. Dependent upon primer design, AP-PCR may offer a fairly well alternative molecular fingerprinting method. In addition, the whole-organism fingerprinting by FTIR provides a tool with sufficient resolving power to distinguish isolates of this Candida species."
|Session name:||XXIst ISTH Congress|
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