Correlation Analysis Between Green Mold Disease Of Dictyophora Rubra And Soil Microbial Community And Screening Of Control Agents In Guizho

In China,edible fungi production has become the fifth largest agricultural industry after grain,oil,fruit,and vegetables,making China the first production sector of edible fungi in the world.Currently,the edible fungi industry also supports rural revitalization strategy and offers tremendous market potential and development opportunities in Guizhou province.Hongtuo Zhusun(Dictyophora rubrovalvata M.Zang)is a local specialty of mushroom in Guizhou,China.It is rich in proteins,amino acids,vitamins,and polysaccharides.Meanwhile,it provids various biological benefits,including immune regulation,anti-tumor properties,reduction of lipids and glucose,antioxidant effects,and anti-aging properties.Therefore,due to its high values in nutrition,medicine,and economy,the areas of its growing increased rapidly in last decade.The cultivation of D.rubrovolvata requires soil coverage to transition from mycelial growth to reproductive growth and form fruiting bodies for successful cultivation.However,in recent years,the prevalence of green mold disease has severely affected the yield and quality of these mushrooms,leading to a significant drop in economic yields.Thus,there is an urgent need for a comprehensive prevention and control system for this disease,as well as related technical research and support.This study investigated the occurrences of green mold disease in D.rubrovolvata from key growing areas in Guizhou Province,such as Guiyang City,Panzhou City,Zhenfeng County,Zijin County,Tongren City,Rongjiang County,Kaili City,and Nayong County.The study included isolating and identifying pathogens,determining the nature and pattern of disease occurrence,and studying the relationship between soil factors and green mold disease occurrence.Moreover,indoor screening of fungicides for dominant pathogens was performed to provide a theoretical basis for disease prevention and control.The key findings from this study include:1.Occurrence patterns and identification of causal agents of green mold disease on D.rubrovolvata in main growing areas in Guizhou province:The average incidence rate was 48.74% for green mold diseases in the investigated growing areas.A total of 150 rotten mushroom eggs were collected for pathogen isolation and purification.Based on morphological observations and phylogenetic tree analysis using ITS and TEF-αsequences,73 isolates of Trichoderma spp.and 10 other isolates,including Penicillium spp.,Chaetomium spp.,Fusarium spp.,Aspergillus spp.,Geotrichum spp.,and Mucor spp.,have been identified.Koch’s postulates confirmed the pathogenicity of representative strains and confirmed that Trichoderma virens,Trichoderma harzianum,and Trichoderma guizhouense are the primary pathogens causing green mold disease in D.rubrovolvata.The investigation also found that the disease occurs mainly during the fruiting body formation phase and spreads rapidly in the greenhouse through various means,including air currents,water,insects and fungi growers.The peak of green mold disease incidence coincides with periods of high temperature in summer,with rising temperatures exacerbating the incidence and severity of the disease.2.Physicochemical properties and soil enzyme activity of D.rubrovolvata soilThis study measured the physicochemical properties and soil enzyme activities of healthy and diseased D.rubrovolvata soils.The results showed no significant difference in the physicochemical properties between diseased and healthy soils.However,the healthy soils had significantly higher urease and catalase contents than the diseased soils,with values of 192.3 U/g and 29.85 U/g,respectively,compared to 145.89 U/g and 24.91 U/g.These findings suggest that soil enzyme activities may contribute to the health of D.rubrovolvata cultivation.3.Soil microbial communities of healthy and diseased D.rubrovolvata soilsTo investigate the disease occurrence mechanisms of green mold disease on D.rubrovolvata,based on microbiome approaches,we employed high-throughput sequencing to determine the soil microbial community structures in both healthy and diseased soils.The results showed that the diseased soils were predominantly enriched with Mortierellomycetes,Mortierellales,Mortierellaceae,Mortierella,Sordariomycetes,Hypocreales,Hypocreaceae,and Trichoderma.Furthermore,the relative abundance of Trichoderma in the diseased soil was 3.53 times greater than that in healthy soil,where fungal communities of Phallales and Phallaceae were mainly enriched.Therefore,the enrichment of Trichoderma pathogenic microorganisms in the soil could be the primary cause of green mold disease.The finding is in accordance with above causal agent isolation and identification studies.In this study,correlation studies have revealed that soil urease content positively correlates with the abundance of Phallaceae but negatively correlates with Saccharomycopsidaceae.Additionally,p H is positively correlated with the abundance of Cladosporiaceae and Strophariaceae,while available phosphorus in the soil positively correlates with Hypocreaceae.Moreover,soil p H,alkali-hydrolyzable nitrogen,available potassium,total nitrogen,and soil invertase content correlate positively with Aspergillaceae abundance.Chemical pesticides and biocontrol agents are important control resources for disease management.We further conducted indoor screening of fungicides and antagonistic bacterial strains agaisnt green mold disease in D.rubrovolvata.By using the mycelium growth rate method for indoor fungicide screening,we identified 23 fungicides that inhibit the growth of Trichoderma mycelium.The results showed that six chemical fungicides and two plant-derived fungicides exerted significant inhibitory effects on different Trichoderma spp.pathogens.The inhibitory effects of these fungicides on the three Trichoderma strains were comprehensively ranked from high to low as follows: 10% benzyl ether·methyl benzimidazole,400 g/L fluosilazole,450 g/L imazalil,25%pyraclostrobin·fomesafen,1.5% berberine·cnidilin,5% vanillin,75% carbendazim,and 80%thiophanate-methyl.We subsequently conducted sensitivity tests of these eight fungicides on the mycelial growth of D.rubrovolvata,which revealed that 5% vanillin,25%pyraclostrobin·fomesafen,and 75% carbendazim had inhibition rates below 5%,indicating their potential for safe application in D.rubrovolvata production.The comprehensive inhibitory effects of these fungicides on the three Trichoderma strains were ranked from high to low as follows: 10%benzyl ether·methyl benzimidazole > 400 g/L fluosilazole > 450 g/L imazalil > 25%pyraclostrobin·fomesafen > 1.5% berberine·cnidilin > 5% vanillin > 75% carbendazim > 80%thiophanate-methyl.Subsequently,sensitivity tests of these eight fungicides on the mycelial growth of D.rubrovolvata were conducted,revealing that 5% vanillin,25% pyraclostrobin·fomesafen,and75% carbendazim had inhibition rates below 5%,indicating their potential for safe application D.rubrovolvata production.Additionally,65 pure bacterial strains were initially isolated from the healthy soil used for cultivating D.rubrovolvata,and antagonistic experiments were conducted on the three Trichoderma pathogens that affect D.rubrovolvata,using the five-point method with bacterial suspension.The results indicated that four Bacillus strains exhibited strong antagonistic effects on the mycelial growth of the three Trichoderma strains,with strain B.amyloliquefaciens 1-1N exhibiting the highest inhibition rates: 79.20% against T.harzianum G11,75.14% against T.guizhouense LZ31,and 69.53% against T.virens T3.Therefore,strain 1-1N demonstrates the promising potential for the biological control of green mold disease in D.rubrovolvata.