Design and Evaluation of Peptide Nucleic Acid Probes for Specific Identification of Candida albicans

Author:

Kim Hyun-Joong,Brehm-Stecher Byron F.

Abstract

Candida albicansis an important cause of systemic fungal infections, and rapid diagnostics for identifying and differentiatingC. albicansfrom otherCandidaspecies are critical for the timely application of appropriate antimicrobial therapy, improved patient outcomes, and pharmaceutical cost savings. In this work, two 28S rRNA-directed peptide nucleic acid-fluorescencein situhybridization (PNA-FISH) probes, P-Ca726 (targeting a novel region of the ribosome) and P-CalB2208 (targeting a previously reported region), were evaluated. Hybridization conditions were optimized by using both fluorescence microscopy (FM) and flow cytometry (FCM), and probes were screened for specificity and discriminative ability against a panel ofC. albicansand various nontargetCandidaspp. The performance of these PNA probes was compared quantitatively against that of DNA probes or DNA probe/helper combinations directed against the same target regions. Ratiometric analyses of FCM results indicated that both the hybridization quality and yield of the PNA probes were higher than those of the DNA probes. In FCM-based comparisons of the PNA probes, P-Ca726 was found to be highly specific, showing 2.5- to 5.5-fold-higher discriminatory power forC. albicansthan P-CalB2208. The use of formamide further improved the performance of the new probe. Our results reinforce the significant practical and diagnostic advantages of PNA probes over their DNA counterparts for FISH and indicate that P-Ca726 may be used advantageously for the rapid and specific identification ofC. albicansin clinical and related applications, especially when combined with FCM.

Publisher

American Society for Microbiology

Subject

Microbiology (medical)

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