Coniochaeta massiliensis sp. nov. Isolated from a Clinical Sampl28

Abstract

The genus Coniochaeta belongs to the class Ascomycota and the family Coniochaetaceae. Some of the Coniochaeta species are plant and animal pathogens, while others are known to be primarily involved in human diseases. In the last few decades, case reports of human infections with Coniochaeta have increased, mainly in immunocompromised hosts. We have described and characterised a new species in the genus Coniochaeta, here named Coniochaeta massiliensis (PMML0158), which was isolated from a clinical sample. Species identification and thorough description were based on apposite and reliable phylogenetic and phenotypic approaches. The phylogenetic methods included multilocus phylogenetic analyses of four genomic regions: ITS (rRNA Internal Transcribed Spacers 1 and 2), TEF-1α (Translation Elongation Factor-1alpha), B-tub2 (β-tubulin2), and D1/D2 domains (28S large subunit rRNA). The phenotypic characterisation consisted, first, of a physiological analysis using both EDX (energy-dispersive X-ray spectroscopy) and Biolog^TM advanced phenotypic technology for fixing the chemical mapping and carbon-source oxidation/assimilation profiles. Afterwards, morphological characteristics were highlighted by optical microscopy and scanning electron microscopy. The in vitro antifungal susceptibility profile was characterised using the E-test^TM exponential gradient method. The molecular analysis revealed the genetic distance between the novel species Coniochaeta massiliensis (PMML0158) and other known taxa, and the phenotypic analysis confirmed its unique chemical and physiological profile when compared with all other species of this genus.

Keywords: Coniochaeta; biolog phenotypic technology; energy-dispersive X-ray spectroscopy; genotype; multilocus DNA sequencing; one new taxon; yeast.

Figure 1

Culture growth on Sabouraud Dextrose Agar + gentamicin and chloramphenicol after five days of incubation at 25 °C. The colour of both the recto and verso of the colonies was white/beige to salmon. (A) Coniochaeta massiliensis PMML0158. (B) Coniochaeta hoffmannii DSM 2693. (C) Coniochaeta mutabilis DSM 10716.

Figure 2

Morphology of Coniochaeta massiliensis (PMML0158). (A,B) Formation of conidiogenous cells (adelophialides) on hyphae and presence of collarette (arrow). (C–F) Conidia aggregating in clusters. (G) Conidia aggregating along the sides of the hyphae. (H) Phialoconidia assembled at the phialide tip. (I,J) Cylindrical conidia with thin and smooth walls. (K,L) Wide hyphae. Optical microscopy (magnification ×1000). Scale bars: 50 μm. Scanning electron microscopy (15 KeV lens mode 4). Scale bars: J = 10 μm; B,F,H = 30 μm; D = 40 μm; L = 50 μm.

Figure 3

Principal component analysis (PCA) processed with the XLSTAT software of the energy-dispersive X-ray spectroscopy chemical mapping profile, performed for the novel species Coniochaeta massiliensis (PMML0158) and two reference strains in the genus. The principal components F1 and F2 explained 100% of the chemical mapping profile variance.

Figure 4

Heat map computed with the XLSTAT software for carbon-source oxidation by the Biolog^TM system for the novel species Coniochaeta massiliensis (PMML0158) and two reference strains in the genus. Colour-gradient interpretation: the most-oxidized substrates are shown in light orange and the least-oxidized substrates are shown in red.

Figure 5

Heat map computed with the XLSTAT software for carbon-source assimilation by the Biolog^TM system for the novel species Coniochaeta massiliensis (PMML0158) and two reference strains. Colour-gradient interpretation: the most-assimilated substrates are shown in light orange and the least-assimilated substrates are shown in red.

Figure 6

Multilocus phylogenetic tree of the newly isolated species Coniochaeta massiliensis PMML0158 (indicated with red dots) and 11 reference strains (type strains are indicated with blue dots), based on the concatenated ITS, B-tub2, and D1/D2 sequences. Phialemonium obovatum CBS 279.76 was used as an outgroup. The maximum-parsimony tree was generated using the MEGA 11 software, with 1000-replication bootstrap values.

Figure 7

Four single-locus phylogenetic trees using the ITS (A), D1/D2 (B), B-tub2 (C), and TEF-1α (D) genomic regions. The species included in each tree differ because the sequences for each locus were not available for all strains. The red dots indicate the new species Coniochaeta massiliensis PMML0158 and blue dots indicate type strains. Phialemonium obovatum CBS 279.76 was used as the outgroup. The maximum-parsimony tree was generated using the MEGA 11 software, with 1000-replication bootstrap values.