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Abstract
Analysis of a worldwide collection of strains of Trichoderma asperellum sensu lato using multilocus genealogies of four genomic regions (tef1, rpb2, act, ITS1, 2 and 5.8s rRNA), sequence polymorphism-derived (SPD) markers, matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) of the proteome and classical mycological techniques revealed two morphologically cryptic sister species within T. asperellum, T. asperellum, T. asperelloides sp. nov. and a third closely related but morphologically distinct species. T. yunnanense. Trichoderma asperellum and T. asperelloides have wide sympatric distribution on multiple continents; T. yunnanense is represented by a single strain from China. Several strains reported in the literature or represented in GenBank as T. asperellum are re-identified as T. asperelloides. Four molecular SPD typing patterns (I–IV) were found over a large geographic range. Patterns I–III were produced only by T. asperellum and pattern IV by T. asperelloides and T. yunnanense. Pattern I was found in North America, South America, Africa and Europe and Asia (Saudi Arabia). Pattern III was found in Africa, North America, South America and Asia, not in Europe. Pattern II was found only in Cameroon (central Africa) and Peru. Pattern IV was found in all continents. All SPD II pattern strains formed a strongly supported subclade within the T. asperellum clade in the phylogenetic tree based on rpb2 and MLS (combined multilocus sequence). The diversity of DNA sequences, SPD markers and polypeptides in T. asperellum suggests that further speciation is under way within T. asperellum. MALDI-TOF MS distinguished T. yunnanense from related taxa by UPGMA clustering, but separation between T. asperellum and T. asperelloides was less clear.
Over many years several individuals have contributed strains of Trichoderma to our ongoing project of Trichoderma taxonomy. We thank the following who contributed strains used in the present study: Drs S. Agbeniyi (CRIN, Abidjan, Nigeria), A. Akrofi (CRIG, Tafo, Ghana), Enrique Arevalo (ICT, Tarapoto, Peru), Le Dinh Don (Long Nam University, Ho Chi Minh City, Vietnam), Harry Evans (CABI-BioScience, UK), T. Gromovykh (Siberian State Technological University, Krasnoyarsk, Russia), L. Kolombet (SRCAM, Obolensk, Russia), T. Okuda (formerly Nippon Roche, Kanagawa, Japan), C. Suarez (INIAP, Pichilingue, Ecuador), P.R. Tondje (IRAD, Yaounde, Cameroon), Giovanni Vanacci (University of Pisa, Italy), F. Vega (USDA-ARS, Beltsville, Maryland). Latonia Miller (BPI) and Corinne Hurard (EBCL) provided excellent technical assistance. Drs C. Benagli and M. Tonolla (Cantonal Institute of Microbiology, Bellinzona, Switzerland) provided helpful advice and assistance in running MALDI-TOF MS analyses. Dr Priscila Chaverri (Univ. Maryland, USA) provided helpful comments on the phylogenetic analyses and taxonomy.