## 2 Results and Discussion  
### Genome Mining and Bioinformatic Analysis  
The genus _Trichoderma_ is of great importance to agriculture and industry, and it has been a hot issue in research and development. Of the 158 _Trichoderma_ genomes published in the NCBI database, six sequencing projects have targeted _T. afroharzianum_ strains through Illumina NovaSeq or Sanger technologies since 2021. In this study, Illumina NovaSeq/PacBio Sequel platforms were used to sequence the whole genome of ACCC 33109, which was fine-mapped and submitted to GenBank under BioProject PRJNA1206798. Similar to the genomes of the _T. afroharzianum_ strains that are 38.4-44.1 Mb in size, with the G+C contents of 46.5-50.5% (Supplementary Table S1), ACCC 33109 is 40.7 Mb in size, with the G+C content of 47.0%, and encodes 11,022 genes, including 9827 genes involved in the KEGG metabolic pathway, 54 secondary metabolic gene clusters (645 genes), and 10 families of cytochrome P450 (176 genes).  
Genome mining is an efficient and reliable strategy for the discovery of novel terpenids from filamentous fungi. Using this strategy, [PERSON] et al. [21] identified a biosynthetic gene cluster of terpene synthase \(BgtA\) in the genome of _Trichoderma erinaceum_ F1-1 and discovered eight new bergamotene-derived sesquiterpenoids. As for ACCC 33109, 35 genes were mapped to the putative KEGG biosynthesis pathway of the terpenoid skeleton (Table 1). Acetyl CoA generated through glycolysis, the tricarboxylic acid cycle, and other pathways is catalyzed by seven putative enzymes (A03930, A05584, A02030, A07965, A02844, A03938, and A10372) of the prevalent acid pathway (MVA) to form the terpene precursor isopentenyl pyrophosphate (IPP; C5), which is further catalytically isomerized by A03978 to produce dimethylallyl pyrophosphate (DMAPP; C5) or condensed by six putative isopentenyl transferases (PTs; A01536, A03044, A03290, A05498, A05610, and A06847) to produce geranyl/farnesyl/geranyl/farnesylfarnesy/hexaprenyl pyrophosphate (GPP, FPP, GGPP, FFPP, and HexPP; C10-C30). These long carbon skeletons are further cyclized by nine putative terpene cyclases (TCs; A01534, A01643, A09488, A04687, A10648, A06313, A03322, A04858, and A09872) to produce the terpenoid skeleton. Through secondary enzyme reactions such as dehydrogenation, hydroxylation, methylation, and acylation catalyzed by, i.e., A01608, A02633, A02814, A02831, A02842, A05170, A06568, A06978, A07199, A08871, A09359, A03646, etc., the side-chains of the terpenoid backbones are decorated to generate various terpenoid compounds with diverse bioactivities. Moreover, gene clusters of ten _TaTS_ genes involving 57 genes were also identified (Supplementary Table S2), which also contribute to the diversification of terpenoid compounds.  
Phylogeny analysis of the deduced proteins indicated that the 15 putative _TaTS_ genes were mainly confined to Class I TC and Class I PT, and they shared 25.82-79.54% identities with the Swiss-Prot homologs (Figure 1). Of them, A05610 and A04858 represent the ancestors of ACCC 33109 TSs and TCs, respectively, and A03044 and A03290 are bifunctional PT-TC. Further phylogeny analysis divided the 133 TCs from _Trichoderma_ spp. into seven clades, and nine TCs from ACCC 33109 are evenly distributed (Supplementary Figure S1).