Comparative Proteome Analysis Of Pectobacterium Carotovorum Subsp.Carotovorum In The Interaction With The Host Plant Zantedeschia Elliotiana’black Magic’ In Different Ways And The Research Of Related Pathogenic Genes

Calla lilies (Zantedeschia sp., Family:Araceae) was known as "the flower star of twenty-first century", because of its flamboyant colors, elegant sharp. However, the development of calla lily has been greatly hindered due to serious occurrence of bacterial soft rot and has no ideal method of prevention. According to reports, the pathogen, isolated from colored calla lily, was Pectobacterium carotovorum subsp. carotovorum (Pcc). Pcc produces plant cell wall-degrading enzymes (PCWDEs), such as pectin and pectate lyases, polygalacturonases, cellulases, proteases and other virulence factors that contribute to causing soft rot disease of economically important plants, leading to significant economic losses.In order to analyzing the differentially expressed proteins of Pcc in the interaction with calla lilies. The protein expression profiles of Pcc grown in2YT medium (Pcc-2YT), Pcc grown in2YT+0.4%stalk extract of Zantedeschia elliotiana (Pcc-2YT-Ze) and Pcc recovered from stalk of the host plant calla lilies (Pcc-V-Ze) were analyzed by using two-dimensional electrophoresis (2-DE), mass technology and ImageMaster2D platinum5.0software. The results showed that,(1), There were18.13%of the total protein spots (70/386) in the gels indicated expression high differentially between Pcc-2YT and Pcc-2YT-Ze. Totally5proteins were identified by mass spectrometry analysis, it was found that in addition to beta-lactamase (upregulating exprssed proteins), antibiotic protein, was main pathogenic factor of Pcc, other four proteins were all belong to metabolic enzymes, among them, molecular chaperone DnaK and1-deoxy-D-xylulose-5-phosphate synthase (DXS) were proteins specifically expressed; ATP synthase subunit alpha and cysteine synthase were upregulating exprssed proteins.(2), Pcc-V-Ze was compared with Pcc-2YT and Pcc-2YT-Ze respectively. There were great differences in protein expression between Pcc-V-Ze and Pcc-2YT-Ze.68proteins were up-regulated expressed, the upregulation rate was17.09%;65proteins were down-regulated expressed, the downregulation rate was16.33%. Among them,53proteins were analyzed with mass technology.In order to studying the roles of these differentially expressed proteins in pathogenicity. Single exchange mutants were generated by biparental mating from the wild-type strain PccS1.17mutants was generated successfully by biparental mating. Among them,9mutants have no pathogenicity difference,8mutants,△PrmA、△IclR、△ClpP、△FlgK、△PotA、△MreB、△HsP33and△Oadh2Al, reduced maceration symptoms.To investigate functions of flgKpcc gene in Pectobacterium carotovorum subsp. carotovorum (Pcc), the gene deletion mutant AflgKpcc was generated by biparental mating from the wild-type strain PccS1. Non-flagellum, cell precipitation in the culture and significantly attenuated motility on0.3%semisolid medium were observed in△flgKpcc compared to those of PccSl.While there were no apparent alterations in bacterial growth in vitro, significant decreases in cellulase, protease activity and biofilm formation were found in△flgKpcc. The pathogenicity on Zantedeschia elliotiana was attenuated in AflgKpcc. AflgKpcc-KH, the complementation strain restored the phenotypes o f△flgKpcc partially or completely. As a result, the flgKpcc mutation not only influenced on flagellar motility, but also virulence-related factors in Pcc.