Comparision of identification methods for the differentiation of trichophyton rubrum from trichophyton mentagrophytes

Nowadays a rapid and striking change is observed in the taxonomy of dermatophytes. They are either re-grouped or re-named, in addition some varieties remain out of nomenclature giving origin to new species. The gold standard in laboratory diagnosis is sequencing analysis. But dermatophyte species are identified at the level of species according to colony characteristics, microscopic morphologies, growth requirements, physiological and biochemical characteristics due to infer-structural needs and high cost of molecular based researches. As changes are observed for a single dermatophyte species both for its morphology (colony pattern, pigments and growth rate) or physiology (in vitro hair perforation) and for its genotype (rRNA and mating patterns); the reliability of the method is questioned. Many micro-organisms belonging to Trichophyton rubrum complex are renamed i.e., T. fischeri, T. kanei, T. raubitschekii and similar are identified. Trichophyton interdigitale, and other micro-organisms pertaining to Trichophyton mentagrophytes sensu lato are re-classified by sequencing analysis. But one of the principal problems observed in mycology laboratory is the differentiation of T. rubrum from T. mentagrophytes using classical diagnostic methods. This study aimed to discuss the efficacy of morphological, physiological and biochemical diagnostic tests used in the laboratory identification of micro-organisms pertaining to both complexes before the current classification, in addition to interrogate their reliability in routine laboratory practice. In the study, T. fischeri CBS 100081, T. fluvimunionse CBS 592.68, T. kanei CBS 289.86, T. kuryangei CBS 422.67, T. kuryangei CBS 517.63, T. kuryangei CBS 518.63, T. pervesii CBS 303.38, T. raubitschekii CBS 202.88, T. raubitschekii CBS 287.86, T. raubitschekii CBS 100084, T. raubitschekii CBS 102856, T. rodhainii CBS 376.49, T. rubrum CBS 302.60, T. rubrum CBS 363.53, T. rubrum CBS 363.62, T. rubrum CBS 392.58, T. rubrum var. nigricans CBS 100237, T. megninii CBS 389.58, T. asteroides CBS 424.63, T. langeronii CBS 764.84, T. mentagrophytes CBS 318.56, T. mentagrophytes var. mentagrophytes CBS 110.65, T. mentagrophytes var. mentagrophytes CBS 160.66, T. papillosum CBS 347.55, T. quinckeanum CBS 572.75, Arthroderma benhamiae (MT-) CBS 807.72, A. benhamiae (MT+) CBS 808.72, A. simii (MT-) CBS 417.65, A. simii (MT+) CBS 448.65, A. vanbreuseghemii CBS 428.63 and A. vanbreuseghemii CBS 558.66 obtained from Centraalbureau voor Schimmelcultures (CBS), Utrecht, the Netherlans were used. Urease reaction (in broth, in tube-agar and Petri agar), in vitro hair perforation test (blond child hair, sheep and goat hair), growth in corn-meal agar and bromcresol purple-milk solids-glucose agar (BCPMSG), Tween opacity test, xiv sorbitol assimilation, salt tolerance, growth in 1 to 7 set of Trichophyton agars were evaluated. Additionally, the presence of macro- and micro- conidia were investigated by inoculation into brain-heart infusion agar (BHIA), malt extract agar (MEA), Lowenstein-Jensen agar (LJ), potatoe dextrose agar (PDA), oat-meal agar (OA), cornmeal dextrose agar (CMDA), Oxoid chromogenic Candida agar (OCCA), Christensen urea agar and Trichophyton agars 1 to 7, and by confronting their performance at days 5, 10 and 15. It was found that urease reaction had provided more rapid and more powerful identification at the level of species when urea media prepared in Petri dishes had been used. Hair perfororation test resulted useful in the differentiation of the two complexes only in the first ten days. Macroconidia production was more prevalent in BHIA and LJ media (P < 0.05). Tween opacity, sorbitol assimilation and salt tolerance tests provided no clue. Additionally, T.rubrum complex produced red pigments in CMDA and BCPMSG, while T. mentagrophytes complex produced none. For example, T. raubitschekii was differentiated from T. rubrum complex by being urease positive, producing red pigments around colonies in BCPMSG, later shifting pH towards alcaline, being unable to perforate hair, producing opaque zone around colonies in Tween opacity test in three days, presenting abundant macro- and micro- conidia in culture. It was also differentiated from T. mentagrophytes complex by producing red pigments in CMDA. It can be concluded that by using over-cited tests in the context of an algorithm T. rubrum complex could be differentiated from T. mentagrophytes complex, in addition to the identification of micro-organisms pertaining to both complexes at the level of species.