The present study includes the molecular characteristics of Trichoderma pleurotum and Trichoderma pleuroticola isolates collected from green moulded cereal straw substrates at 47 oyster mushroom farms in Poland. The screening of the 80 Trichoderma isolates was performed by morphological observation and by using the multiplex PCR assay. This approach enabled specific detection of 47 strains of T. pleurotum and 2 strains of T. pleuroticola. Initial identifications were confirmed by sequencing the fragment of internal transcribed spacer regions 1 and 2 (ITS1 and ITS2) of the rRNA gene cluster and the fragment including the fourth and fifth introns and the last long exon of the translation-elongation factor 1-alpha (tef1) gene. ITS and tef1 sequence information was also used to establish the intra- and interspecies relationship of T. pleurotum and T. pleuroticola originating from the oyster mushroom farms in Poland and from other countries. Comparative analysis of the ITS sequences showed that all T. pleurotum isolates from Poland represent one haplotype, identical to that of T. pleurotum strains from Hungary and Romania. Sequence analysis of the tef1 locus revealed two haplotypes ("T" and "N") of Polish T. pleurotum isolates. The "T" type isolates of T. pleurotum were identical to those of strains from Hungary and Romania. The "N" type isolates possessed a unique tef1 allele. Detailed analysis of the ITS and tef1 sequences of two T. pleuroticola isolates showed their identicalness to Italian strain C.P.K. 1540.

Institute of Plant Genetics Polish Academy of Sciences Strzeszyńska 34 Poznań Poland

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Błaszczyk L, Popiel D, Chełkowski J, Koczyk G, Samuels GJ, Sobieralski K, Siwulski M. Species diversity of Trichoderma in Poland. J Appl Genetics. 2011;52:233–243. doi: 10.1007/s13353-011-0039-z. PubMed DOI PMC

Carbone I, Kohn LM. A method for designing primer sets for speciation studies in filamentous ascomycetes. Mycologia. 1999;91:553–556. doi: 10.2307/3761358. DOI

Chełkowski J, Golka L, Stępień Ł. Application of STS markers for leaf rust resistance genes in near-isogenic lines of spring wheat cv. Thatcher. J App Genet. 2003;44:323–338. PubMed

Doohan FM, Parry DW, Jenkinson P, Nicholson P. The use of species-specific PCR-based assays to analyse Fusarium ear blight of wheat. Plant Pathol. 1998;47:197–205. doi: 10.1046/j.1365-3059.1998.00218.x. DOI

Druzhinina IS, Kopchinskiy AG, Komon M, Bissett J, Szakacs G, Kubicek CP. An oligonucleotide barcode for species identification in Trichoderma and Hypocrea. Fungal Genet Biol. 2005;42:813–828. doi: 10.1016/j.fgb.2005.06.007. PubMed DOI

Gea FJ. First report of Trichoderma pleurotum on oyster mushroom crops in Spain. J Plant Pathol. 2009;91:504.

Hatvani L, Antal Z, Manczinger L, Szekeres A, Druzhinina IS, Kubicek CP, Nagy A, Nagy E, Vágvölgyi C, Kredics L. Green mould diseases of Agaricus and Pleurotus spp. are caused by related but phylogenetically different Trichoderma species. Phytopathology. 2007;97:532–537. doi: 10.1094/PHYTO-97-4-0532. PubMed DOI

Jaklitsch WM, Komon M, Kubicek CP, Druzhinina IS. Hypocrea voglmayrii sp. nov. from the Austrian Alps represents a new phylogenetic clade in Hypocrea/Trichoderma. Mycologia. 2005;97:1365–1378. doi: 10.3852/mycologia.97.6.1365. PubMed DOI

Komon-Zelazowska M, Bisset J, Zafari D, Hatvani L, Manczinger L, Woo S, Lorito M, Kredics L, Kubicek CP, Druzhinina IS. Genetically closely related but phenotypically divergent Trichoderma species cause green mold disease in oyster mushroom farms worldwide. Appl Environ Microbiol. 2007;73:7415–7426. doi: 10.1128/AEM.01059-07. PubMed DOI PMC

Kopchinskiy A, Komon M, Kubicek CP, Druzhinina IS. TrichoBLAST: a multilocus database for Trichoderma and Hypocrea identifications. Mycol Res. 2005;109:657–660. doi: 10.1017/S0953756205233397. PubMed DOI

Kredics L, Hatvani L, Antal Z, Manczinger L, Druzhinina IS, Kubicek CP, Szekeres A, Nagy A, Vágvölgyi C, Nagy E. Green mould disease of oyster mushroom in Hungary and Transylvania. Acta Microbiol Immunol Hung. 2006;53:306–307.

Kredics L, Kocsube S, Nagy L, Komon-Zelazowska M, Manczinger L, Sajben E, Nagy A, Vagvolgyi C, Kubicek CP, Druzhinina IS, Hatvani L. Molecular identification of Trichoderma species associated with Pleurotus ostreatus and natural substrates of the oyster mushroom. Microb Lett. 2009;300:58–67. doi: 10.1111/j.1574-6968.2009.01765.x. PubMed DOI

Muthumeenakshi S, Brown AE, Mills PR. Genetic comparison of the aggressive weed mould strains of Trichoderma harzianum from mushroom compost in North America and the British Isles. Mycol Res. 1998;102:385–390. doi: 10.1017/S0953756297004759. DOI

Park MS, Bae KS, Yu SH. Molecular and morphological analysis of Trichoderma isolates associated with green mold epidemic of oyster mushroom in Korea. J Huazhong Agric Univ. 2004;23:157–164.

Park MS, Bae KS, Yu SH (2004b) Morphological and molecular analysis of Trichoderma species associated with green mold epidemic of oyster mushroom in Korea. New Challenges in Mushroom Science. Proceedings of the 3rd Meeting of Far East Asia for Collaboration on Edible Fungi Research, Suwon, Korea, pp 143–158

Park MS, Bae KS, Yu SH. Two new species of Trichoderma associated with green mold of oyster mushroom cultivation in Korea. Mycobiology. 2006;34:111–113. doi: 10.4489/MYCO.2006.34.3.111. PubMed DOI PMC

Samuels GJ, Dodd SL, Gams W, Castlebury LA, Petrini O. Trichoderma species associated with the green mold epidemic of commercially grown Agaricus bisporus. Mycologia. 2002;94:146–170. doi: 10.2307/3761854. PubMed DOI

Szekeres A, Kredics L, Antal Z, Hatvani L, Manczinger L, Vágvölgyi C. Genetic diversity of Trichoderma strains isolated from winter wheat rhizosphere in Hungary. Acta Microbiol Immunol Hung. 2005;52:156. doi: 10.1556/AMicr.52.2005.2.2. PubMed DOI

Sharma SR, Vijay B. Yield loss in Pleurotus ostreatus spp. caused by Trichoderma viride. Mushroom Res. 1996;5:19–22.

Thompson JD, Higgins DG, Gibson TJ. CLUSTALW: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res. 1994;22:4673–4680. doi: 10.1093/nar/22.22.4673. PubMed DOI PMC

White TJ, Bruns T, Lee S, Taylor JW. Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In: Innis MA, Gelfand DH, Shinsky JJ, White TJ, editors. PCR protocols: a guide to methods and applications. San Diego: Academic; 1990. pp. 315–322.

Woo SL, Di Benedetto P, Senatore M, Abadi K, Gigante S, Soriente I, Ferraioli S, Scala F, Lorito M. Identification and characterization of Trichoderma species aggressive to Pleurotus in Italy. J Zhejiang Univ Agric Life Sci. 2004;30:469–470.