Revision of agents of black-grain eumycetoma in the order Pleosporales

Abstract

Eumycetoma is a chronic fungal infection characterised by large subcutaneous masses and the presence of sinuses discharging coloured grains. The causative agents of black-grain eumycetoma mostly belong to the orders Sordariales and Pleosporales. The aim of the present study was to clarify the phylogeny and taxonomy of pleosporalean agents, viz. Madurella grisea, Medicopsis romeroi (syn.: Pyrenochaeta romeroi), Nigrograna mackinnonii (syn. Pyrenochaeta mackinnonii), Leptosphaeria senegalensis, L. tompkinsii, and Pseudochaetosphaeronema larense. A phylogenetic analysis based on five loci was performed: the Internal Transcribed Spacer (ITS), large (LSU) and small (SSU) subunit ribosomal RNA, the second largest RNA polymerase subunit (RPB2), and translation elongation factor 1-alpha (TEF1) gene. In addition, the morphological and physiological characteristics were determined. Three species were well-resolved at the family and genus level. Madurella grisea, L. senegalensis, and L. tompkinsii were found to belong to the family Trematospheriaceae and are reclassified as Trematosphaeria grisea comb. nov., Falciformispora senegalensis comb. nov., and F. tompkinsii comb. nov. Medicopsis romeroi and Pseudochaetosphaeronema larense were phylogenetically distant and both names are accepted. The genus Nigrograna is reduced to synonymy of Biatriospora and therefore N. mackinnonii is reclassified as B. mackinnonii comb. Nov. Mycetoma agents in Pleosporales were phylogenetically quite diverse despite their morphological similarity in the formation of pycnidia, except for the ascosporulating genus Falciformispora (formerly in Leptosphaeria). Most of the species diagnosed from human mycetoma were found to be related to waterborne or marine fungi, suggesting an association of the virulence factors with oligotrophism or halotolerance.

Keywords: Madurella; Pleosporales; Trematosphaeriaceae; mycetoma; taxonomy.

Fig. 1

Phylogram of combined dataset (LSU, SSU, RPB2, TEF1) obtained by Bayesian analysis and maximum likelihood (values of ≥ 0.70 for Bayesian probability and ≥ 70 % for maximum likelihood shown in bold branches). Species causing mycetoma shown in red, with obsolete names between inverted commas. Dothidea sambuci and Phaeosclera dematioides were used as outgroup.

Fig. 1

Phylogram of combined dataset (LSU, SSU, RPB2, TEF1) obtained by Bayesian analysis and maximum likelihood (values of ≥ 0.70 for Bayesian probability and ≥ 70 % for maximum likelihood shown in bold branches). Species causing mycetoma shown in red, with obsolete names between inverted commas. Dothidea sambuci and Phaeosclera dematioides were used as outgroup.

Fig. 2

Phylogenetic tree resulting from Bayesian analysis and maximum likelihood for the ITS gene (values of ≥ 0.8 for Bayesian probability and ≥ 80 % for maximum likelihood are shown with bold branches). Medicopsis romeroi was used as outgroup.

Fig. 3

Average of colony diameters of studied species after 2 wk of incubation at various temperatures ranging from 15 to 36 °C, with 3 °C intervals, including 10 °C, 37 °C, and 40 °C.

Fig. 4

Identification scheme of eumycetoma causative agents based on physiological properties of studied species.

Fig. 5

Falciformispora senegalensis (CBS 196.79). Colonies after 2 wk of incubation on: a. MEA; b. OA; c. cleistothecia; d, e. asci; f. ascospore; g. ascospore surrounded by thin sheath. — Scale bars = 10 μm.

Fig. 6

Medicopsis romeroi (CBS 252.60). Colonies after 2 wk of incubation on: a. MEA; b. OA; c–e. pycnidia; f. conidia; g. conidiophores. — Scale bars = 10 μm.

Fig. 7

Pseudochaetosphaeronema larense(CBS 640.73). Colonies after 2 wk of incubation on: a. MEA; b. OA; c–e. pycnidia; f. conidia; g. conidiophores. — Scale bars = 10 μm.

Fig. 8

Trematosphaeria grisea (a–c. CBS 332.50, d–g. CBS 120271). Colonies after 2 wk of incubation on: a. MEA; b. OA c. ABTS media; d, e. pycnidia; f. conidia; g. conidiophores. — Scale bars = 10 μm.

Fig. 9

Trematosphaeria pertusa (CBS 122368). Colonies after 2 wk of incubation on: a. MEA; b. OA. c, d. pycnidia; e. verrucose hyphae and conidia; f. conidia; g. conidiophores. — Scale bars = 10 μm.