Influencing Factors On Biofilm Formation Of Two Dickeya Spp.and Transcription Of Dickeya Zeae MS1

Bacterial soft rot caused by Dickeya spp.is one of the most devastating diseases worldwide.However,there is currently no effective treatment for these diseases.Bacterial biofilms were regonized as one of the pathogenic factors of Dickeya spp.and it is an important weapon for bacteria to resistant advers environment.Here,we evaluated the influencing factors on biofilm formation of two Dickeya spp.(Dickeya zeae MS1 and Dickeya dieffenbachiae PA1).And we also carried out comparative transcriptomic analysis between planktonic and biofilm phenotypes,using RNA-sequencing,in order to gain insight into the role of Dickeya zeae MS1 in the pathogenesis of bacterial soft rot disease.The results were as follows:1.The development biofilm of Dickeya zeae MS1 was studied.The results showed that Dickeya zeae MS1 biofilm could be categorized into four phases:attachment,development,maturation and detachment.And mature biofilm consisted of a highly organized architecture of both bacterial cells and a self-produced matrix of extracellular polysaccharides.2.The influencing factors affected Dickeya dieffenbachiae PA1 biofllm formation was also studied Dickeya dieffenbachiae PA1 had strong ability to form biofilm.And Dickeya dieffenbachiae PA1 could form ball-like colonies,and colonies networks in the biofilm development phase.Basides,the results showed that the most suitable carbon source for supporting the growth of Dickeya dieffenbachiae PA1 biofilm was sucrose,and at 30?,pH 8.0.When the temperature was lower or higher than 30?,pH values below 8.0 or above as well as glucose,lactose,maltose as a carbon source,biofilm formation was inhibited.Additionally,the susceptibility of biofilm to environmental stress was evaluated.Biofilm cells were less susceptible to heat,cold,UV light than their planktonic counterparts.3.The influencing factors of Dickeya zeae MS1 biofilm formation were clarified.The results showed that sucrose was the most suitable carbon source for supporting the growth of biofilm cells and that 32? and pH 7.0 were the most favorable of the temperatures and pH values examined of Dickeya zeae MS1.Furthermore,the addition of Ca2+,Fe2+,K+ and Na+ enhanced the formation of biofilm in minimal medium cultures,whereas 2.5 mM Cu2+ and Mn2+ was inhibitory,and Mg2+ had no effects on Dickeya zeae MS1.D-Methionine(D-Met)had the strongest inhibitory effect on biofilm formation of Dickeya zeae MS1 among the five D-amino acids.When the concentration of D-Met was 22.5?25 mM,22.5 mM D-Met significantly decreased the microbe's motility(i.e.,swirrmming,swarming,and twitching),reduced the cellular metabolic activity of D.ickeya zeae MS1 biofilm cells,and increased the activity of streptomycin sulfate.D-Met probable target the protein components of the biofilm matrix,and significantly inhibited biofilm formation.4.The results of comparative transcriptomic analysis between planktonic and biofilm phenotypes of Dickeya zeae MS1 showed that significant differences were found in the expression levels of 402 genes,of which 293 genes were significantly up-regulated,109 genes were significantly down-regulated.GO analysis of all differential genes associated with the bacterial biofilm reavled that transport processes,establishment of colonization processes and colonization processes were enrichment in biology processes;membrane components was enrichment in cellular process;transporter activity and antioxidant activity were enrichment in molecular biology function.KEGG pathway analysis revealed that the genes involved in pathways associated with bacterial biofilm were as follows:biosynthesis of secondary metabolites,ABC transporters,biosynthesis of amino acids,and microbial metabolism in diverse environments.Bacterial biofilm can protect the bacterial cells from adverse environment.However,the development of biofilm can also be affected by various physical,chemical,nutritional conditions,and genes involved in pathway associated with biosynthesis of secondary metabolites,ABC transporters,biosynthesis of amino acids,and microbial metabolism in diverse environments.In this study,we evaluated biofilm formation and its development mechanisms.We inferre that application of D-Met or other influencing factors that could inhibit biofilm fornation,to destroy bacterial biofilm and further control of soft rot diseases.