Molecular detection and identification of agents of eumycetoma: detailed report of two cases

Abstract

We describe two cases of eumycetoma in the legs. The infections could not be adequately diagnosed by classical mycology, but the causative agents were successfully identified as Madurella mycetomatis by species-specific PCR and DNA sequencing.

FIG. 1.

(A) Direct microscopy of 8-week-old cultures of M. mycetomatis (isolate P-1) on soil extract agar. Only vegetative hyphae with some chlamydospore-like cells (arrows) were seen. (B) M. mycetomatis-specific PCR for species identification using primers 26.1A and 28.3A (1). Lanes 1 and 6 contain a 100-bp molecular size marker. Lanes 2 to 5 show the PCR amplicons of P-1, P2, positive control, and negative control, respectively. Both P-1 and P-2 were positive.

FIG. 2.

(A) Massive subcutaneous mass involving the inner part of the right knee. (B) T1-weighted magnetic resonance image after gadolinium injection, showing a large heterogeneous soft tissue mass. Bone and knee joint are healthy and not involved. (C) During surgical excision, the lesion appeared to be well capsulated, and it was excised completely. Note the massive lesion, which measures about 16 cm. (D) Four-day-old culture of the black grains on chocolate blood agar. The fungus starts presenting as white mycelium and later become olivaceous or brown. (E) The lesion consisted of dense fibrous tissues containing multiple cavities full of thick exudates containing pus, blood, and grains. (F) Gomeri-Grocott staining showing a fungal grain surrounded by inflammatory cells. One can clearly see the fungal hyphae growing toward the periphery of the grain.