Identification of Tr80437 as a Key Gene for Furfural Resistance and Enhanced Cellulase Production in Trichoderma reesei by Bulked Segregant Analysis and Cross-Species Screening

Trichoderma reesei is a primary fungal strain for industrial cellulase production but is strongly inhibited by furfural, a cytotoxic byproduct generated during lignocellulosic biomass pretreatment. To overcome this limitation, a furfural-tolerant T. reesei mutant was generated via atmospheric and room-temperature plasma (ARTP) mutagenesis and backcrossing. Bulked Segregant Analysis (BSA) of F1 progeny with contrasting tolerance phenotypes identified a quantitative trait locus on chromosome X containing 172 candidate genes. Comparative analysis with Neurospora crassa and functional screening using its knockout library identified a furfural tolerance-related gene, NCU04116. Disruption of Tr80437, the T. reesei orthologue of NCU04116, conferred enhanced furfural tolerance and increased cellulase activity under furfural stress compared with the wild-type strain. These findings identify Tr80437 as a promising genetic target for engineering robust T. reesei strains for industrial cellulase production.