One possibility, testing of which NVP-BSK805 solubility dmso is beyond the scope of current work, is that the one-step lactonohydrolase evolved as a neofunctionalisation (present within filamentous fungi of Leotiomycetes/Sordariomycetes orders) of the two-step detoxification
mechanism retained by T. mycotoxinivorans. If so, the original mechanism can still exist in select extant lineages (within filamentous Ascomycota) in varying degrees (dependent on selection pressure towards one-step detoxification). Conclusions Our research shows the first finding of a functional zearalenone lactonohydrolase in mycoparasitic Trichoderma aggressivum (an activity earlier characterised in the Clonostachys rosea strains). Based on the combined screening of over ninety isolates of Trichoderma/Clonostachys and in silico investigation of origins of the enzyme activity (through phylogeny reconstruction and homology modelling) we were able to provide click here supporting evidence for its evolutionary origins,
as well as monophyly of functional lactonohydrolase homologs in both genera. The supporting evidence for presence and activity of functional enzyme homologs is based on chemical analyses, gene expression patterns, homology models showing conservation of key structural features and a marked reduction of zearalenone content in cultured samples (containing both medium and mycelium). Methods Fungal isolates Fungal isolates originated from culture collections of the Institute of Plant Genetics (Polish Academy of Sciences, Poznan, Poland); Institute of Science of Food Production (Bari, Italy; ITEM), Institute of Food Technology (Poznan Fenbendazole University of Life Sciences, Poznan, Poland), Department of Forest Pathology (Poznan University of Life Sciences, Poznan, Poland), Research Institute
of Vegetable Crops, (Skierniewice, Poland) and Rothamsted International UK. The isolates were derived from soil, compost, wood, cultivated mushroom and cereal grain samples. All 98 isolates were identified using both morphological  and molecular methods (ITS 4-5 and tef1 markers) (Additional file 1: Table S1). Isolation of pure cultures Fungal isolates investigated in this study were collected from pieces of decaying wood, cultivated mushroom compost, samples of soil and cereal grain. The samples were plated on salt water nutrient agar (SNA)  and incubated at 20°C for 6 days. Putative Trichoderma and Clonostachys colonies were purified on potato dextrose agar (PDA, Oxoid). Pure culture were transferred to the tubes containing SNA medium and stored at -20°C for further study. Isolation of DNA Mycelium used for DNA extraction was obtained by inoculating Czapek-Dox broth (Sigma-Aldrich) with Yeast Extract (Oxoid) and streptomycin sulphate (50 mg/L-1, AppliChem) and after incubation at 25°C for 21 days on a rotary shaker (120 rpm). Mycelium was collected on filter paper in a Büchner funnel, was held with sterile water, frozen at -20°C, and freeze – dried.