Analysis Of Apple Rhizosphere Microbial Community Structure And Screening Of Plant Growth-promoting Rhizobacteria

The continuous cropping of apple trees has affected the growth and development of fruit trees,the yield and quality of apple.The physical and chemical methods can not effectively prevent the continuous cropping obstacles.The research shows that the method of biological control has an irreplaceable advantage in the prevention and control of plant continuous cropping obstacles.In this paper,the microbial community structure of apple rhizospherewas analyzed to provide the theoretical basis for the biological control measures.The screening of apple rhizosphere growth-promoting bacteria provide species resources for solving the apple continuous cropping obstacle.In this experiment,in order to control the continuous cropping obstacles of apple orchards,apple trees were selected planted for 5 years,10 years,15 years and 20 years from Yantai Qixia apple orchard,and each year was randomly selected with three repetitions.Soil samples of each repetition was selected with 10%,30%,50%,70% and 100% between the trunk to the projection of the crown in apple rhizosphere,a total of 60 soil samples.The soil total DNA was extracted from the above 60 samples.A total of 16671 OTU(97% similarity level)were measured by high-throughput sequencing results.Through the analysis of the years and sites,we find that the maximum OTU level in 10 years is 9193,and the OTU level of the sample near the trunk or near the projection of the crown is higher than that of the middle.According to the statistical analysis of classification,43 classes and 712 genus were obtained.The main classes were Proteobacteria,Acidobacteria,Planctomycetes,Actinobacteria,Verrucomicrobia,Bacteroidetes,Gemmatimonadetes,Chloroflexi and Firmicutes.Among them,the relative abundance of the core classes including Proteobacteria,Acidobacteria,Planctomycetes,Actinobacteria were about 62.33%.The main bacteria genus were Gemmatimonas,Sphingomonas,Gaiella,Planctomyces,Candidatus_Koribacter,Bryobacter,Gemmata,Pirellula,Nitrospira,Rhizomicrobium,Haliangium,Rhodanobacter,Blastocatella,Reyranella,Steroidobacter,Chryseolinea,Bacillus and Arthrobacter,which account for about17% of the total bacteria genus.Through the determination of total nitrogen,ammonium nitrogen,nitrate nitrogen,organic matter,available phosphorus and available potassium in all samples,it was found that the six nutrient contents in the 15-year samples were higher than the other three years.It is obvious that planting 15-year of apple trees has the best growth environment.Through the RDA analysis between microbial community and environmental factors,we can know that Bacillus,Rhodanobacter,Sphingomonas,Lactococcus,Bacillus,Mycobacterium,Rhizomicrobium,Dongia,Blastocatella,Pseudomonas,Acidisphaera can enhance the content of nutrient elements in the rhizosphere of apple trees,and contribute to the growth of apple trees.In contrast,Pirellula,Planctomyces,Ohtaekwangia,Gemmata,Nitrospira,Haliangium,Roseiflexus,Reyranella,Arthrobacter,Burkholderia,Candidatus_Koribacter,Aquicella can reduce the content of nutrients elements in the rhizosphere of apple trees,which is not conducive to the growth of fruit trees.The study screens out 15 excellent functional bacteria,which include four strains of producing protease bacteria,three strains of producing IAA bacteria,four strains of phosphate-solubilizing bacteria and four strains of bacteria that antagonize four kinds of Fusarium.Through the strains morphological observation,physiological and biochemical identification experiment and 16 S rDNA identification,the preliminary identification results was four strains of high-yield protease,which include YL17 Bacillus methylotrophicus,Y2 Bacillus cereus,YL15 Bacillus methylotrophicus and YS8 Pseudomonas auricularis,three strains of producing IAA,which include YS2 Enterobacter ludwigii,YH22 Enterobacter ludwigii,YD8 Klebsiella variicola,four strains of phosphate-solubilizing,which include YD28 Bacillus subtilis,YD11 Pseudomonas auricularis,YD6 Providencia rettgeri,YG7 Serratia nematodiphila,and four strains of antagonizing four kinds of Fusarium,which include Y9 Bacillus methylotrophicus,YK50 Bacillus methylotrophicus,Y4 Bacillus methylotrophicus,YA24 Paenibacillus polymyxa.In this study,a variety of apple rhizosphere growth-promoting bacteria were screened,which greatly enriched the resources of apple rhizosphere microorganisms.Through the combination of soil nutrient content and high-throughput sequencing technology,we found that the rule of growth and development of apple trees and the advantages and disadvantages of rhizosphere microbes,which provided theoretical support for the research of microbial fertilizer after us and laid the foundation for the study of apple continuous cropping obstacle.