Colonization of Trichoderma spp. is essential for exerting their beneficial functions on the plant. However, the interactions between Trichoderma spp. and plant roots are still not completely understood. The aim of this study was to investigate how TgSWO affect Trichoderma guizhouense to establish themselves in the plant rhizosphere and promote plant growth. In this study, we deeply analyzed the molecular mechanism by which the functional characterization of the TgSWO by expressing different functional region deletion proteins (FRDP) of TgSWO. Root scanning analysis results showed that TgSWO could dramatically increase root density and promote growth. In addition, we also found that TgSWO could expand root cell walls, subsequently increase root colonization. Moreover, knockout of TgSWO mutants (KO) or overexpression of TgSWO mutants (OE) produced greatly reduced or increased the number of cucumber root, respectively. To clarify the molecular mechanism of TgSWO in plant-growth-promotion, we analyzed the ability of different FRDP to expand the root cell wall. The root cell wall architecture were considerably altered when treated by ΔCBD protein (the TgSWO gene of lacking in the CBD domain was cloned and heterologously expressed), in correlation with the present YoaJ domain of TgSWO. In contrast, neither the expansion of cell walls nor the increase of roots was detectable in ΔYoaJ protein. Our results emphasize the YoaJ domain is the most critical functional area of TgSWO during the alteration of cell wall architecture. Simultaneously, the results obtained in this study also indicate that TgSWO might play a plant-growth-promotion role in the Trichoderma-plant interactions by targeting the root cell wall.

Endo-1,4-β-mannanase is an enzyme that can catalyze the random hydrolysis of β-1, 4-mannosidic linkages in the main chain of mannans, glucomannans and galactomannans and has a number of applications in different biotechnology industries. Penicillium oxalicum is a powerful hemicellulase-producing fungus (Bioresour Technol 123:117-124, 2012); however, few previous studies have focused on the cloning and expression of the endo-1,4-β-mannanase gene from Penicillium oxalicum. A gene encoding an acidophilic thermostable endo-1,4-β-mannanase (E.C. 3.2.1.78) from Penicillium oxalicum GZ-2, which belongs to glycoside hydrolase family 5, was cloned and successfully expressed in Pichia pastoris GS115. A high enzyme activity (84.4 U mL-1) was detected in the culture supernatant. The recombinant endo-1,4-β-mannanase (rPoMan5A) was tagged with 6 × His at its C-terminus and purified using a Ni-NTA Sepharose column to apparent homogeneity. The purified rPoMan5A showed a single band on SDS-PAGE with a molecular mass of approximately 61.6 kDa. The specific activity of the purified rPoMan5A was 420.9 U mg-1 using locust bean gum as substrate. The optimal catalytic temperature (10 min assay) and pH value for rPoMan5A are 80°C and pH 4.0, respectively. The rPoMan5A is highly thermostable with a half-life of approximately 58 h at 60°C at pH 4.0. The K m and V max values for locust bean gum, konjac mannan, and guar gum are 7.6 mg mL-1 and 1425.5 μmol min-1 mg-1, 2.1 mg mL-1 and 154.8 μmol min-1 mg-1, and 2.3 mg mL-1 and 18.9 μmol min-1 mg-1, respectively. The enzymatic activity of rPoMan5A was not significantly affected by an array of metal ions, but was inhibited by Fe3+ and Hg2+. Analytical results of hydrolytic products showed that rPoMan5A could hydrolyze various types of mannan polymers and released various mannose and manno-oligosaccharides, with the main products being mannobiose, mannotriose, and mannopentaose. Our study demonstrated that the high-efficient expression and secretion of acid stable and thermostable recombinant endo-1, 4-β-mannanase in Pichia pastoris is suitable for various biotechnology applications.

Trichoderma spp. is a cosmopolitan group of soil fungi which plays a remarkable role in stimulating plant growth after interacting with plant roots and has good application prospects in intensive agriculture. In this study, rice straw and amino acids improved the population of Trichoderma guizhouense NJAU4742 under solid-state fermentation and helped us develop a new type of organic fertilizer. The effects of this biological organic fertilizer were evaluated in the growth of peppers (Capsicum annuum L.) for two seasons under sandy and mountain soils. In the first season, the yields in T6 (0.06% solid fermentation products in soil) and AT6 (added 0.06% solid fermentation products and 1% amino acid organic fertilizer in soil) treatments were increased by 41.8% and 52.3% in sandy soil and by 51.6% and 46.5% in mountain soil, respectively, compared with chemical fertilizer. During the second season, the same trend was obtained in both sandy and mountain soils. Soil peroxidase activity (125.2 μmol·g⁻¹ dw), urease activity (58.7 μmol·g⁻¹ dw) and invertase activity (13.11 mg·g⁻¹ dw) reached their highest levels in biological organic fertilizer compared to the treatments with chemical fertilizer and solid fermentation products. Redundancy analysis showed that crop yield was positively correlated with enzyme activities, soil organic carbon, total nitrogen, and available phosphorus. Thus, we demonstrated that NJAU4742-enriched biological organic fertilizer could accelerate the transformation of nutrients and promote pepper growth.