Research On The Formation Mechanism Of Paenibacillus Polymyxa Biofilm And Its Colonization

Paenibacillus polymyxa CF05 strain is a bacterial strain isolated from the Tianmu Mountain ancient cedar with significant biocontrol effect on various plant diseases such as Chinese hickory trunk canker androot rot,as well as good research and development value.Effective colonization is a prerequisite for microbial pesticides to exhibit their biocontrol function.Biofilm is a special structure formed by bacteria accumulating on the surface of boundary phase to resist the external adverse environment,and plays an important role during bacterial stable colonization.In order to further determine the biocontrol mechanism of Paenibacillus polymyxa,the morphological and physiological characteristics of P.polymyxa CF05 biofilm formation were investigated,as well as biofilm formation and regulation.The results were described as follows.1.The formation process of biofilm on the solid surface of glass,etc.by CF05 strains was confirmed by scanning electron microscopy and other microscopic observation techniques.According to the bacterial morphology changes and polymer secretion characteristics,it was divided into the following five stages.Bacterial free period(Phase ?): bacterial cells were long rod-shaped,rounded at both ends,with smooth bacterial surface;initial contact period(phase ?): bacteria were attached to the contact surface,with viscous secretion;microstructure period(Phase ?): bacteria were mixed and aggregated to form a membrane structure;mature structure period(Phase ?): bacteria exhibited an irregular arrangement of three-dimensional stratification,with large gaps between the bacteria;aging shedding period(Phase ?): part of the cells fell off,and the biofilm was reconstruction.2.The analysis of determination of the bacterial biofilm formation ability of different sugars by solid phase adsorption was carried out.It was found that sucrose imposed the greatest influence on the ability of bacterial biofilm formation.The content of exopolysaccharide and the biomass of biofilm increased significantly After sucrose treatment,.3.Using sugar separation,metabolic full spectrum analysis and other techniques,the change of sugar component in CF05 biofilm stimulated by 1% sucrose was determined.It was found that some contents among the metabolites from the biofilm of P.polymyxa CF05,such as the glycosides of 8-D-Olivosyl-landomycin,Landomycin I,Netilmicin,N2'-Acetylgentamicin C1 a,varied dramatically.It is presumed that the metabolites produced by sucrose-treated P.polymyxa CF05 changed the microenvironment of the biofilm,thus affecting the bacterial activity in the biofilm.4.The GFP expression vector pPPGFP was constructed by fluorescent labeling technology to achieve the GFP labeling of P.polymyxa CF05.The GFP-labeled vector was used to observe the colonization of P.polymyxa CF05 in the cortex of Chinese hickory,providing materials for further study on the bacterial colonization inside the Chinese hickory cortex.In summary,this study focused on the formation and colonization of P.polymyxa biofilm,confirmed the morphological changes of biofilm formation,found the significant effect of sucrose on biofilm formation,and determined the sugar content in biofilm components,which realized the fluorescent labeling of Paenibacillus polymyxa and the colonization observation in the Chinese hickory cortex.The results provided an important reference for the study of biocontrol mechanism and microbial pesticide development of P.polymyxa CF05 strain...