Clonal species Trichoderma parareesei sp. nov. likely resembles the ancestor of the cellulase producer Hypocrea jecorina/T. reesei

Abstract

We have previously reported that the prominent industrial enzyme producer Trichoderma reesei (teleomorph Hypocrea jecorina; Hypocreales, Ascomycota, Dikarya) has a genetically isolated, sympatric sister species devoid of sexual reproduction and which is constituted by the majority of anamorphic strains previously attributed to H. jecorina/T. reesei. In this paper we present the formal taxonomic description of this new species, T. parareesei, complemented by multivariate phenotype profiling and molecular evolutionary examination. A phylogenetic analysis of relatively conserved loci, such as coding fragments of the RNA polymerase B subunit II (rpb2) and GH18 chitinase (chi18-5), showed that T. parareesei is genetically invariable and likely resembles the ancestor which gave raise to H. jecorina. This and the fact that at least one mating type gene of T. parareesei has previously been found to be essentially altered compared to the sequence of H. jecorina/T. reesei indicate that divergence probably occurred due to the impaired functionality of the mating system in the hypothetical ancestor of both species. In contrast, we show that the sexually reproducing and correspondingly more polymorphic H. jecorina/T. reesei is essentially evolutionarily derived. Phenotype microarray analyses performed at seven temperature regimens support our previous speculations that T. parareesei possesses a relatively high opportunistic potential, which probably ensured the survival of this species in ancient and sustainable environment such as tropical forests.

FIG. 1.

Trichoderma parareesei. (a to c) Cultures after 4 days on CMD (a), SNA (b) and PDA (c); (e) conidiophore without cover glass (9 days); (f) young conidiophore (3 days); (g) young conidiophore with sterile elongation (3 days); (h) conidiophore of a shrub on growth plate (2 days); (i to n) conidiophores after 3 days (i and n) and after 9 days (j to m); (o and r) phialides at 2 days (o) and 8 days (r); (p and q) chlamydospores (7 days); (s and t) conidia (8 days); (e to t) growth on CMD; (a to c, p, and q) growth at 30°C; (e and j to m) growth at 15°C; (f to i, n, o, and r to t) growth at 25°C; (a to c, f, g, i, and n to t) strain C.P.K. 717; (e, h, and j to m) strain C.P.K. 661; (d and u) Hypocrea jecorina (Trichoderma reesei); (d) culture on PDA after 4 days at 30°C (C.P.K. 917); (u) conidia (C.P.K. 1127, SNA, 25°C, 4 days). Scale bars: 15 mm (a to d), 30 μm (e and h), 15 μm (f, g, i, k, l, p, and q), 10 μm (j, m, n, and r to u), and 5 μm (o).

FIG. 2.

Temperature-dependent growth of T. parareesei and H. jecorina on different carbon sources. (A) Temperature-dependent growth rates of both species calculated for the 16 best carbon sources (cluster I, l-arabinose, d-arabitol, d-cellobiose, dextrin, i-erythritol, d-fructose, gentobiose, α-d-glucose, maltotriose, d-mannitol, d-mannose, d-melezitose, d-trehalose, d-xylose, γ-aminobutyric acid), as estimated by Druzhinina et al. (2) for H. jecorina. The complete carbon source utilization profiles are given in Table S1 in the supplemental material. (B) Temperature-dependent growth of T. parareesei and H. jecorina on individual carbon sources. Vertical bars correspond to standard deviations, calculated on the basis of the profiles of six and five strains for T. parareesei and H. jecorina, respectively. Colored labels highlight the differences.

FIG. 3.

Bayesian circular phylogram inferred from the concatenated data set of partial exons of rpb2 and chi18-5 phylogenetic markers. Symbols at the nodes correspond to PPs of >94%. Arrows point to clades/nodes of phylogenetic species. (A) Phylogenetic position of the Reesei subclade with respect to other species from Trichoderma section Longibrachiatum; (B) phylogenetic relations between species of the Reesei subclade (enlarged from panel A). The ex type strain of T. reesei QM 6a (C.P.K. 917) is underlined.