DETECTION AND DIFFERENTIATION OF CAUSATIVE FUNGI OF ONYCHOMYCOSIS USING PCR AMPLIFICATION AND RESTRICTION ENZYME ANALYSIS

Background Onychomycosis, a fungal nail infection, has become one of t he most important dermatophytoses, Unfortunately a predictably success ful diagnostic approach to onychomycosis does not yet exist. Objective The purpose of this study was to develop a deoxyribonucleic acid (DNA )-based diagnostic method to improve the sensitivity and specificity o f the detection and differentiation of the pathogenic fungi of onychom ycosis. Methods We attempted to detect fungi in the nail using polymer ase chain reaction (PCR) primer systems that were designed in conserve d sequences of the small ribosomal subunit 18S-rRNA genes shared by mo st fungi, and differentiated between species by restriction enzyme ana lysis of the amplified product. Results Fragments of the gene coding f or 18S-rRNA were amplified successfully from medically important fungi species, but not from normal nails. Restriction fragment length polym orphism patterns using HaeIII endonuclease were sufficiently different to allow the recognition of individual species. Conclusions The PCR-r estriction enzyme analysis method appears to be a more sensitive detec tion and identification technique for onychomycosis than conventional methods, and has considerable diagnostic value.