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Production of labeled DNA probes for the rapid diagnosis of disseminated candidiasis in immunocompromised patients Cheung, Lori


The increasing incidence of disseminated (invasive) candidiasis is probably attributable to iatrogenic factors and to improved pre and postmortem evaluation. Premortem diagnosis of such infections have seldom been made early enough for successful treatment. In order to increase the likelihood of successful antifungal chemotherapy, rapid diagnosis of such infections is vital. However, present diagnostic procedures for invasive candidiasis are insensitive and often do not reliably differentiate superficial from invasive infections. This study was undertaken to produce DNA probes and to optimize conditions for rapid and efficient detection of Candida DNA. Seven random Candida albicans DNA fragments (2-7 kbp) were cloned into plasmid pACYC 184. These recombinant plasmids were labeled with either ³²p or biotin and used as probes. Two of the four recombinant plasmids tested were genus specific. The other two were slightly cross reactive with other yeasts (Saccharomyces cerevisiae and Hansenula anomala). Probes labeled with ³²p were twice as sensitive as the biotin probes. One ³²p labelled recombinant (#66) detected 7 Pg of target DNA , which corresponds to approximately 2 X 10⁵ C.albicans cells. With refined simple DNA extraction procedures for C.albicans (in serum), these recombinant probes could possibly be suitable for clinical application.

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