Abstract
The genus Madurella comprising four species, M. fahalii, M. mycetomatis, M. pseudomycetomatis, and M. tropicana, represents the prevalent cause of eumycetoma worldwide. The four species are phenotypically similar and cause an invariable clinical picture, but differ markedly in their susceptibility to antifungal drugs, and epidemiological pattern. Therefore, specific identification is required for optimal management of Madurella infection and to reveal proper epidemiology of the species. In this study, a novel multiplex real-time PCR targeting the four Madurella species was developed and standardized. Evaluation of the assay using reference strains of the target and non-target species resulted in 100% specificity, high analytical reproducibility (R2 values >0.99) and a lowest detection limit of 3 pg target DNA. The accuracy of the real-time PCR was further assessed using biopsies from eumycetoma suspected patients. Unlike culture and DNA sequencing as gold standard diagnostic methods, the real-time PCR yielded accurate diagnosis with specific identification of the causative species in three hours compared to one or two weeks required for culture. The novel method reduces turnaround time as well as labor intensity and high costs associated with current reference methods.
Highlights
Mycetoma, a neglected tropical disease of the subcutaneous tissue, is characterized by progressive tumefaction and formation of grains and multiple draining sinuses, leading to massive soft tissue destruction and disfigurement [1, 2]
Madurella mycetomatis is the most prevalent species worldwide. Other species such as M. fahalii, M. pseudomycetomatis, and M. tropicana can cause mycetoma and have a different susceptibility towards the drug used for treating mycetoma patients
In Sudan, cases that show presence of black grains in histology or fine needle aspiration cytology are usually diagnosed as M. mycetomatis eumycetoma [6], but M. fahalii causes infection with black grains
Summary
A neglected tropical disease of the subcutaneous tissue, is characterized by progressive tumefaction and formation of grains and multiple draining sinuses, leading to massive soft tissue destruction and disfigurement [1, 2]. E. sinuses in the skin and grain discharge, mycetoma can be recognized from other skin diseases, while proper diagnosis with identification of the etiologic agent requires a combination of diagnostic tools. Madurella fahalii was described in 2012 from a eumycetoma patient in Sudan, a country that is highly endemic for mycetoma [8]. In Sudan, cases that show presence of black grains in histology or fine needle aspiration cytology are usually diagnosed as M. mycetomatis eumycetoma [6], but M. fahalii causes infection with black grains. Occurrence of marked differences in antifungal susceptibility between Madurella species and other agents of black grain eumycetoma necessitate the development and implementation of a species-specific identification system [7, 10]. Correct identification will assist in the administration of appropriate antifungal therapy, and will help to elucidate the epidemiology and distribution of agents of eumycetoma
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