Genetic relationships and isozyme profile of dermatophytes and Candida strains from Egypt and Libya

Abstract

Three molecular techniques random amplified polymorphic DNA (RAPD), inter-simple sequence repeat (ISSR) and restriction fragment length polymorphism (RFLP) were employed for identification and to study the genetic relationship among six species of dermatophytes and three species of yeasts isolated from Egyptian and Libyan patients with skin mycosis. Each species was represented by two isolates, one from Egyptian patients and the second from Libyan. RAPD in which four random 10-mer primers and two ISSR primers were used to amplify the DNA fragments of target fungi and RFLP in which two universal primers (ITS1 and ITS4) were used to amplify the internal transcribed spacer (ITS) regions of the ribosomal (rRNA) gene in fungal isolates followed by digestion with HinfI and HaeIII endonucleases was carried out. Three molecular marker techniques showed considerable potential for identifying and discriminating dermatophytes and Candida species and the achieved results confirmed identification based on conventional morphological methods. Results of RAPD and ISSR markers revealed 78.7% genetic similarity (GS) between Microsporum canis and other tested fungi reflecting a relatively longer genetic distance from other isolates of dermatophytes and yeasts. Candida krusei and Candida tropicalis were closely related showing 93.3% GS. C. albicans showed 90.9% similarity with other species of Candida. Epidermophyton floccosum was easily separated from all Trichophyton species showing 87.3% similarity. Unique bands were displayed by certain fungi and can be taken as a positive marker for isolate identification and discrimination. RFLP technique revealed differences in the number (1 to 5) and size (8 to 378 base pairs) of DNA fragments depending on the fungal isolate and restriction enzyme used. Within each fungal species, different isolates of dermatophytes and Candida from Egypt and Libya showed close relationship. Seven isozyme systems namely esterase, peroxidase, malate dehydrogenase, acid phosphatase, glutamate-oxalo-acetate transaminase, Urease and protease were studied to detect the gene expression and genetic variability among the different isolates of dermatophytes and Candida.

Keywords: Random amplified polymorphic DNA (RAPD), inter-simple sequence repeat (ISSR), restriction fragment length polymorphism (RFLP), dermatophytes, Candida, isozymes

African Journal of Biotechnology Vol. 12(29), pp. 4554-4568