Study On High-density Fermentation Of Trichoderma Viride Spores And Its Regulation On Plant Growth

Using the green Trichoderma strain Tv-1511(CGMCC No.16800)preserved by the Institute of Biology of Qilu University of Technology(Shandong Academy of Sciences)as the material,this study aims at some problems,such as the function of bio-organic fertilizer is single,the fertilizer effect is not significant,and the growth-promoting mechanism is not clear.The optimization of fermentation formula and the control of fermentation conditions,the industrial high-density fermentation sporulation process of Trichoderma was developed to reduce the cost of bacterial production.Trichoderma fungus was applied to the growth process of Arabidopsis thaliana and peppermint.The growth-promoting effect and mechanism of green Trichoderma Tv-1511 on Arabidopsis thaliana and peppermint were studied by mass spectrometry and q RT-PCR.Seven kinds of synthetic additives were used in the liquid medium of sporulation fermentation of Trichoderma viride Tv-1511,five of which could increase the spore yield of Trichoderma viride.When the synthetic additive G was used,the effective spore number could reach 4.0×10~7cfu/m L.On the basis of additive G,the addition amount of glucose as auxiliary carbon source in the culture medium was optimized,and the number of effective spores reached 4.4×10~7cfu/m L when glucose was 3%.Through the microscopic observation of the changes of bacteria in the process of fermentation,it was found that the yield of monomer chlamydospore was the highest when it was fermented for 138-147 h.In the actual production,it is recommended to use 3%artificial additive G,3%glucose and fermentation for 6 days to produce green Trichoderma Tv-1511 spore liquid.MPK6 and plasma membrane H~+-ATPase regulate the role of Trichoderma viride Tv-1511 in leaf colonization and growth promotion of Arabidopsis thaliana,which provides a theoretical basis for the wide application of Trichoderma fungicide in the field.Spraying green Trichoderma Tv-1511 on Arabidopsis thaliana leaves,it was observed that Trichoderma could colonize Arabidopsis thaliana leaves,effectively promote the growth of Arabidopsis thaliana,increase biomass and promote nutrient absorption.Plasma membrane H~+-ATPase plays an important role in regulating Trichoderma viride Tv-1511 in plant leaf colonization,and plays a major regulatory role in AHA1 gene.Through western blot and q RT-PCR,it was found that Trichoderma viride Tv-1511 treatment increased the H~+-ATPase activity of Arabidopsis plasmalemma by 1.24 times and the expression of AHA1 gene by 2.19 times.Trichoderma viride Tv-1511 could activate MAPK6,and increase its enzyme activity and protein expression intensity by 2.35 times and 1.11 times,respectively.Trichoderma viride Tv-1511 mediates the expression and activity of plasma membrane H~+-ATPase through MPK6,thus colonizing leaves and promoting plant growth.MAPK and NADPH oxidase regulated the colonization of Trichoderma viride Tv-1511 in peppermint root and promoted the yield and quality of peppermint essential oil,which provided a theoretical basis for the application of Trichoderma fungicide in peppermint cultivation.When Trichoderma viride Tv-1511 was applied to the root of mint,it was observed that Trichoderma colonized in the root,and significantly increased the plant growth index(leaf area,primary root length,lateral root length,lateral root density),biomass(fresh weight,dry weight)and photosynthesis index(net photosynthetic rate,chlorophyll content),which effectively promoted the growth of peppermint.Peppermint inoculated with Trichoderma viride Tv-1511 significantly increased the contents of menthone(31.5%),menthol(59.75%)and prenone(1.01%)in essential oils and increased the expression of PR and MR by 2.78 and 2.33 times,respectively.Through western blot and q RT-PCR,it was found that MPK6 regulated the metabolism of peppermint essential oil at the transcriptional level and affected the activities of related enzymes.The activity of NADPH oxidase in peppermint seedlings treated with ASA and DPI decreased by 1.62 times and 2.62 times respectively.It was found that NADPH oxidase mediates the production of reactive oxygen species and participates in the activation of MAPK.Trichoderma viride Tv-511 regulates the genes related to the metabolism of peppermint essential oil through potential MAPK signal pathway,thus improving the quantity and quality of peppermint essential oil.In summary,the high-density fermentation process of sporulation of Trichoderma viride Tv-1511 was developed in this study,which provides a reference for reducing the industrial production cost of Trichoderma fungicides.The promoting effect and mechanism of green Trichoderma Tv-1511 in the production of Arabidopsis thaliana and peppermint were explored.Results could provide theoretical basis for the targeted development of growth-promoting agents for specific crops and the development of Trichoderma bio-organic fertilizer.