The Effects Of Holotrichia Parallela Larva Damage On Rhizosphere Bacterial Community Of Peanut And Preliminary Exploration Of Its Mechanism

Subterranean insect pests,white grubs,have been recognized as major pests that are difficult to control due to their underground habitat and hidden damage.However,the variety of microbial agents used to control white grubs is limited,and new insecticidal strains and genetic resources need to be further explored.Recent studies have shown that when plants exposed to biotic and abiotic stresses,they enriched beneficial microorganisms to cope with stress.How do plants respond to the damage caused by white grubs,and what is the effect on the structure and function of rhizosphere bacterial communities? The clarification of these problems will help to understand the interaction relationship between “pest-plant-rhizosphere microbes” and lay a theoretical foundation for breaking through the existing screening mode of insecticidal strains and establishing a screening system for new and highly effective insecticidal strains.In this study,Holotrichia parallela larvae,peanuts and rhizosphere bacteria were used as the research objects.Through 16 S rRNA gene high-throughput sequencing and culturomics method,the effects on bacterial community structure and function of peanut rhizosphere were analyzed in the field trial.It was found that the feeding of white grubs caused significant changes in the rhizosphere bacterial community of peanut under field conditions.The abundances of Enterobacter spp.and Acinetobacter spp.increased;7 strains of Enterobacter spp.with insecticidal activity against Coleopteran pests were isolated in the rhizosphere,and these strains have no insecticidal activity against Lepidopteran Plutella xylostella.The main findings are as follows:In this study,a method for preserving rhizosphere soil samples for large-scale sampling in the field was established.The samples were supplemented with GTC buffer and then stored at room temperature for 1 day.The 16 S rRNA gene sequencing results showed that there were no obvious effects on the rhizosphere bacterial community structure and diversity.In the field trial,white grubs damage caused changes in the bacterial community structure of peanut rhizosphere.Using 16 S rRNA gene high-throughput sequencing technology,it was found that the abundance of Bacillus spp.,Enterobacter spp.,Acinetobacter spp.,etc.increased after grub treatment.After the feeding of white grubs 1 d and 7 d,the bacterial community structure of peanut rhizosphere changed significantly.Macrogenomic sequencing results showed that white grubs treatment caused significant changes in the function of peanut rhizosphere bacteria.Using culturomics and three-generation sequencing technology,the cultivable bacteria of peanut rhizosphere were isolated and identified after the feeding of grub.A total of 43 plates(96-well plates)mixed strains(4128 well mixed strains)were isolated by culturing with six mediums such as TSB.Using the Barcode labeling method,the 16 S rRNA full-length gene was sequenced by the PacBio sequencing platform,and a total of 29,823 sequences were obtained,which were annotated to 4 phylums: Proteobacteria,Bacteroidetes,Actinobacteria,Firmicutes;and annotated to 147 genera,such as Pantoea,Enterobacter.Strains with insecticidal activity were obtained from the cultivable bacteria of the peanut rhizosphere after grub damage.A total of 7 strains with insecticidal activity against Colaphellus bowringi,initially identified as strains of Enterobacter,were indentified by insecticidal activity assay.Strains with insecticidal activity against Plutella xylostella were not identified.The non-targeted metabolome technique was used to analyze the rhizosphere metabolites of peanut.The results showed that the metabolites of peanut roots changed significantly after the treatment.A total of 19 differential metabolites were identified,among which 11 metabolites such as campesterol,stigmasterol and palmitic acid increased,and 8 metabolites such as inositol and malic acid decreased.This study clarified the effects of grub damage on the structure and function of peanut rhizosphere bacterial community under field conditions.It lays a foundation for clarifying the interaction mechanism of “grub-peanut-rhizosphere bacteria” and mining rhizosphere insecticidal microbial resources.