Isolation Of A Novel Biocontrol Myxococcus Sp.BS And The Myxobacterial Predation Mechanism Against Bacterial Soft Rot

Myxobacteria are a class of complex multicellular micro-organisms with social behavior which can slide on the solid surface and predate bacteria by means of group cooperation.As an indigenous microorganism with soil colonization ability,Myxobacteria plays an important role in regulating the composition and dynamic distribution of microbial communities in soil.At present,most of the research focus on their basic developmental characteristics,including the locomotion,fruit body formation and self-recognition mechanism.however,the research on controlling harmful microorganisms including plant pathogenic bacteria by using the predatory characteristics of Myxobacteria is relatively rare,so it is valuable to control pathogenic bacteria by using predatory Myxobacteria.In this study,a biocontrol bacterium BS identified as Myxococcus sp.was obtained by screening several Myxobacteria strains,which has the good ability to predate Pectobacterium carotovorum subsp.catotovorum,?Pec?.The biocontrol potential and mechanism of the bacterium BS were studied in detail in the paper.In this study,12 Myxobacteria strains were isolated and purified from the collected soil by inducing fruiting body with rabbit manure and yeast,and numbered with the initial letters of the collected sites.According to physiological and biochemical analysis,16S rDNA sequence alignment analysis and fruiting body morphology observation,the 12 strains were identified as genus,6 strains belong to the genus archaea sp.,4 strains belong to the genus Alyxococcus sp.,and 2 strains belong to the genus Corallococcus sp..The strains screened were co-cultured with Pcc,and the viable cells of Pcc were counted.Finally,Myxococczis sp.BS with strong ability to predate Pcc,was screened out.Strain BS had strong predation ability on phytopathogenic bacteria such as Erwinia amylovora,Ralstonia solanacearum,Dickeya fangzhongudai,Dickeya chrysanthenzum,Dickeya solani.Pot experiments showed that strain BS could effectively inhibit the infection rate of Pcc to potato and calla lily,and reduce the incidence of plant diseases.In the infection experiment of potato,the rot rate of potato tuber was 45.1%on the 5th day and 96.7%on the 10th day in control group,combination BS with Pcc group was 0 on the 5th day and 18.4%on the 10th day,the control effect was 81%and the control of streptomycin was 8.2%.The results of pot experiment of calla lily showed that the incidence and disease index of group treated with BS decreased from 75.0%to 19.8%and 62.5%to 6.3%respectively.At the same time,the increase of cormeles treated with BS was also observed.The results indicated that strain BS has good biocontrol potential against soft rot.Based on the good biocontrol effect of Myxocococus sp.BS to soft rot bacteria,the mechanism of-its predation on pathogenic bacteria was studied.The results showed that the effective predation of Pcc by strain BS was achieved through cell-cell contact.The cell contact between predator and food plays an important role in the process of Myxobacteria predating pathogenic bacteria.According to the literature,the material transport between cells mainly includes TypeVI secretion systems?T6SS?,vesicles,Type ?secretion systems?T4SS?,nanotubes?nanoubes?,autolysis and lipoprotein transfer.By analyzing the genome of Myxobacteria and combining its predation characteristics,we infer that T6SS and vesicles play an important role in the predation process of Myxobacteria.In this study.Myxococcus xanthus DK1622 was used as the material to study the T6SS gene cluster.it was found that there was only one synthetic gene cluster in the genome of strain DK1622,which consisted of 13 proteins.MXAN₄807-MXAN₄813 constituted the core element of T6SS structure.The results showed that the key gene inactivated did not affect the predation characteristics.Therefore,T6SS was not dominant in the predation of Myxobacteria.In order to further verify that role of vesicles in the predation of Myxobacteria,strain BS and Pcc were co-cultured,and the morphology of bacteria was observed by electron microscopy.the results showed that a large number of outer membrane vesicles were secreted around Myxobacteria,and some vesicles were attached to the periphery of Pcc cells.Furthermore,the extracellular vesicles of DK1622 were extracted and their ability to prey on Pcc was verified.it was found that the extracellular vesicles secreted by Myxococcus xanthus DK1622 had the ability to inhibit Pcc.The counting results showed that the number of viable Pcc bacteria treated by vesicles was only 37%to the original number.In combination with the reports,vesicles contain a variety of hydrolytic enzymes and secondary metabolites with antibacterial activity,suggesting that vesicles play an important role in the predation of bacteria by Myxobacteria.