| Bacterial fruit blotch(BFB)caused by Acidovorax citrulli bacteria is a devastating bacterial disease to watermelon,melon and other cucurbitaceae crops.If BFB occurs,it will seriously affect the commodity value and edible value of melons,which brings a huge economic loss to the melon industry.However,the pathogenic mechanism of A.citrulli has not been fully identified.c-di-GMP is a second messenger of bacteria.It can be synthesized by GGDEF/GGEEF domain-containing guanylate cyclase(diguanylate cyclase)in bacteria,and can be degraded by phosphodiesterase(PDE)with EAL or HD-GYP domain.Research shows that,c-di-GMP is involved in regulating the pathogenic effects of many pathogenic bacteria.Currently,there is a lack of research on the regulation of c-di-GMP pathway in A.citrulli.In this study,we have carried out the research on identifying proteins related to c-di-GMP metabolism in A.citrulli.The main research progress as follow:1.The c-di-GMP metabolism related proteins in A.citrulli were identified.Based on bioinformatics analysis,total 48 proteins related to c-di-GMP metabolic exist in the A.citrulli AAC00-1 genome,where three proteins contained EAL domain,eight proteins contained HD-GYP domain,21 proteins contained GGDEF domain,and 16 proteins contained both EAL and GGDEF domain2.c-di-GMP involved in the regulation of A.citrulli virulence has been proved.In this study,the predicted genes related to c-di-GMP metabolism were individually knocked out.By testing the virulence of the mutant to melon seedlings,the pathogenicity of ΔAave＿4208to melon seedlings decreased significantly.In order to further study the effect of Aave＿4208on the pathogenic mechanism of A.citrulli,we constructed the Aave＿4208 absence mutant and its corresponding complementation strain,and determined the effect of Aave＿4208 on pathogenicity,colonization in vivo,biofilm,motility,hypersensitivity respose(HR),growth level in vitro and antioxidant stress.The results of phenotype showed that compared with the wild type,the absence of Aave＿4208 can reduce the pathogenicity,biofilm formation,swimming ability,colonization in the host and growth ability in vitro,and it delays the ability to elicite HR and improves the tolerance to hydrogen peroxide,while the phenotypes cause by complementation strain and overexpression strain are consistent with the wild-type stain.To learn more about the regulatory effect of Aave＿4208 on A.citrulli,in this study,we analyzed the transcriptome of Aave＿4208,and found that the absence of Aave＿4208 resulted in the 608 differential expression genes,among which the expression of T3SS-related genes hrc C,hrc V,hrc U,hrc R,hrc J and flagellum-related genes fil A,fil S decreased.These data are consistent with the above experimental results.All these results indicate that c-di-GMP pathway is involved in the regulation of A.citrulli virulence.This is the first systematic study on the regulation of c-di-GMP pathway of A.citrulli on its pathogenicity.All the proteins related to c-di-GMP metabolism in A.citrulli xjl12 were identified,and it was proved that the c-di-GMP pathway was involved in the regulation of A.citrulli virulence.This study provides a scientific basis for further understanding the pathogenesis of A.citrulli,and also lays a theoretical foundation to effectively control BFB.Fur has been studied in many pathogens,and studies have shown that in addition to regulating iron related genes,fur gene also regulates many important pathogenic factors,including quorum sensing,biofilm,exopolysaccharide production,swimming,anti-oxidative stress and some metabolic pathways.In order to further understand the transcriptional regulation function of fur A on A.citrulli,this study planned to conduct transcriptome analysis of fur A mutant of A.citrulli.The results showed that fur A negatively regulated the production of ferritin carriers(feo A and feo B),and positively regulated the ferritin associated genes(bfr).fur A negatively regulates iron-associated transporters to maintain iron balance in vivo.fur A deficiency resulted in decreased expression of flagella-related and chemotactic genes in the T3 SS,and decreased expression of catalase and superoxide dismutase,as well as tricarboxylic acid transporter... |
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