Molecular phylogeny of the Pseudallescheria boydii species complex: proposal of two new species

Abstract

Pseudallescheria boydii (anamorph Scedosporium apiospermum) is the species responsible for human scedosporiosis, a fungal infection with a high mortality rate and which is difficult to treat. Recently, it has been demonstrated that high genetic variation exists within this species. We have performed a morphological and molecular study involving numerous strains of clinical or environmental origins and from different countries. The analysis of partial sequences of the beta-tubulin (two loci) and calmodulin genes and the internal transcribed spacer region of the rRNA gene has demonstrated that P. boydii is a species complex. The combined analysis of the sequences of the four loci of 60 strains has showed the presence of 44 haplotypes in the in group. Three species morphologically related to P. boydii sensu stricto, i.e., Pseudallescheria angusta, Pseudallescheria ellipsoidea, and Pseudallescheria fusoidea, which had previously been considered synonyms, could be differentiated genetically from P. boydii in our study. It is relevant that two of the three strains now included in P. ellipsoidea have caused invasive infections. The species Pseudallescheria minutispora and Scedosporium aurantiacum are clearly phylogenetically separated from the other species studied and are here proposed as new. Morphological features support this proposal. All the strains included in S. aurantiacum species have a clinical origin, while those included in P. minutispora are environmental. Further studies are needed to demonstrate whether all the species included in the P. boydii complex have different clinical spectra and antifungal susceptibility.

FIG. 1.

One of the 12 most-parsimonious trees obtained from heuristic searches based on BT2 sequence. Bootstrap support values above 70% are indicated at the nodes. Type strains are indicated with boldface type. P. africana and S. prolificans were used as outgroups. Pb, P. boydii; Pe, P. ellipsoidea; Pf, P. fusoidea; Pa, P. angusta; Pm, P. minutispora; Sa, S. aurantiacum; Paf, P. africana; Sp, S. prolificans; CI, consistency index; RI, retention index; HI, homoplasy index.

FIG. 2.

One of the 420 most-parsimonious trees obtained from heuristic searches based on TUB sequence. Bootstrap support values above 90% are indicated at the nodes. Type strains are indicated with boldface type. P. africana and S. prolificans were used as outgroups. Pb, P. boydii; Pe, P. ellipsoidea; Pf, P. fusoidea; Pa, P. angusta; Pm, P. minutispora; Sa, S. aurantiacum; Paf, P. africana; Sp, S. prolificans; CI, consistency index; RI, retention index; HI, homoplasy index.

FIG. 3.

The single most-parsimonious tree obtained from heuristic searches based on CAL sequence. Bootstrap support values above 70% are indicated at the nodes. Type strains are indicated with boldface type. P. africana and S. prolificans were used as outgroups. Pb, P. boydii; Pe, P. ellipsoidea; Pf, P. fusoidea; Pa, P. angusta; Pm, P. minutispora; Sa, S. aurantiacum; Paf, P. africana; Sp, S. prolificans; CI, consistency index; RI, retention index; HI, homoplasy index.

FIG. 4.

One of the 5,000 most-parsimonious trees obtained from heuristic searches based on ITS sequence. Bootstrap support values above 80% are indicated at the nodes. Type strains are indicated with boldface type. P. africana and S. prolificans were used as outgroups. Pb, P. boydii; Pe, P. ellipsoidea; Pf, P. fusoidea; Pa, P. angusta; Pm, P. minutispora; Sa, S. aurantiacum; Paf, P. africana; Sp, S. prolificans; CI, consistency index; RI, retention index; HI, homoplasy index.

FIG. 5.

One of the 2,496 most-parsimonious trees obtained from heuristic searches based on analysis produced from the combined data set. Bootstrap support values of 100% are indicated at the nodes. Type strains are indicated with boldface type. P. africana and S. prolificans were used as outgroups. Pb, P. boydii; Pe, P. ellipsoidea; Pf, P. fusoidea; Pa, P. angusta; Pm, P. minutispora; Sa, S. aurantiacum; Paf, P. africana; Sp, S. prolificans; S, source; T, teleomorph; G, Graphium anamorph; CI, consistency index; RI, retention index; HI, homoplasy index; c, clinical; e, environmental; +, presence; −, absence.

FIG. 6.

Scedosporium aurantiacum isolates (A, D, E) IHEM 15458 and (B, C) FMR 8630. (A) Colony growing on PDA after 14 days of incubation at 25°C. (B, C) A conidiogenous cell and conidia from solitary conidiophores. (D) A synnema of the Graphium anamorph. (E) Apical part of a synnema producing conidia.

FIG. 7.

Pseudallescheria minutispora strain FMR 4072. (A) Colony growing on PDA after 14 days of incubation at 25°C. (B) Conidiogenous cells and conidia of the Scedosporium anamorph. (C) Apical part of a synnema of the Graphium anamorph producing conidia. (D) Ascoma. (E) Conidia borne on undifferentiated hyphae. (F) Ascospores.