Dynamic Analysis Of Dominant Microflora In Vermiculite Matrix During The Development Of Potato Commom Scab

Potato common scab is an important soil-borne and seed-borne disease caused by plant pathogen Steptomyces spp.,which has a great influence on potato production.At present,the pathogenesis of scab is still unclear.In this paper,from the perspective of etiology and dynamic composition of microorganisms,the infection characteristics of S.scabies and the diversity of surrounding microorganisms during the process of scab of micropotato plants on vermiculbsfite substrate were tracked and observed.The main research results were as follows:1.Fluorescent labeling and infection observation of S.scabies:the green fluorescent protein gene(GFP)was ligated to Streptomyces expression vector pSET-dCas9 carrying the strong promoter ermE*p by gene recombination method.The recombinant fluorescence expression vector pSET-GFP was obtained and transferred into S.scabies wild strain CPS-1 after conjugation.The labeled strain CPS-1-GFP was screened and obtained.The colony morphology of the labeled strain was basically the same as that of the wild strain,and the fluorescence signal of spores was stronger and the hyphae was weaker.The labeled strains were distributed in tuber,root,stolon,underground stem,aboveground stem and leaf of microtuber tissue culture plants inoculated in greenhouse.2.Dynamic analysis of dominant microflora in vermiculite matrix during the development of micropotato scab:vermiculite matrix samples were collected during the growth of microtuber,and Illumina Miseq high-throughput sequencing technology was used to analyze the diversity of microbial composition.A total of 1268024 effective sequences of bacteria and 1536390 effective sequences of fungi were obtained.The diversity of bacterial community in vermiculite matrix increased and the diversity of fungal community decreased during the whole growth period of microtuber.Analysis of the dominant flora of the top 50 abundances of bacteria and fungi showed that:the relatively stable dominant bacteria were Pseudarthrobacter,Acinetobacter,Nocardioides,Massilia and Dyella,the fungi were Plectosphaerella,Talaromyces,Ramophialophora,Trichoderma,Fusarium,Botrytis and Alternaria.There was no significant difference in the abundance of the dominant microflora in different periods.After comparing the top 10 bacterial communities with great differences in abundance in samples from different periods,the dominant flora with large differences in abundance were Leifsonia,Thermomonas,Streptomyces,Caulobacter,Jatrophihabitans,Rhodobacter,Phaeosphaeria,Aureobasidium,Setophaeosphaeria,Setomelanomma,Rhizopus and Symmetrospora.The abundance of Dyella,Ramophialophora,Mortierella,Rhodococcus,Streptomyces,Botrytis and Massilia increased significantly during the formation and expansion of microtubers,while the abundance of Actinoplanes was higher before the formation of tubers.The abundance of Talaromyces was relatively high and stable after the formation of tuber,while the abundance of Massilia increased gradually after 20-50 days of microtuber transplantation.Quantitative analysis of pathogens showed that the number of pathogens in per gram of vermiculite substrate increased gradually with the increase of transplanting time.There was no significant difference in the number of pathogens at 50 d and before 50 d,but the content of pathogens at 60 d and 70 d was significantly higher than that at the 40 d before.3.Analysis of culturable microflora in the spot after micropotato scab:morphological observation of the isolated bacteria,actinomycetes and fungi,and preliminary molecular identification of the strains using 16S rDNA and ITS sequences.A total of 29 strains of bacteria were obtained.Among them,Bacillus simplex,Bacillus gaemokensis,Bacillus cereus,Bacillus amyloliquefaciens,Bacillus aryabhattai,Agrobacterium tumefaciens,Variovorax paradoxus,Microbacterium thalassim were 8 species.27 strains of actinomycetes,including 7 strains of Streptomyces scabies,Streptomyces diastatochromogenes,Streptomyces rishiriensis.Streptomyces galilaeus.Streptomyces panaciterrae,Nocadioides albus,Streptomyces rochei.There were 17 strains of fungi,including 6 species,such as Fusarium tricinctum,Fusarium redolens,Fusarium solani,Clonostachys rosea,Alternaria solani and Plectosphaerella cucumerina.After comparative analysis of the dominant microflora of vermiculite matrix and culturable microorganisms in scab spots,the common culturable microflora were Fusarium,Plectosphaerella,Streptomyces,Alternaria and Nocardioides.