Project description:The Lon protein is a protease implicated in virulence of many pathogenic bacteria, including some plant pathogens. However, little is known about the role of Lon in bacteria from genus Dickeya. This group of bacteria include important potato pathogens, with the most aggressive species, D. solani. To determine the importance of Lon for pathogenicity and response to stress conditions of bacteria, we constructed a D. solani Δlon strain. The mutant bacteria showed increased sensitivity to certain stress conditions, in particular osmotic and high-temperature stresses. Furthermore, qPCR analysis showed an increased expression of the lon gene in D. solani under these conditions. The deletion of the lon gene resulted in decreased motility, lower activity of secreted pectinolytic enzymes and finally delayed onset of blackleg symptoms in the potato plants. In the Δlon cells, the altered levels of several proteins, including virulence factors and proteins associated with virulence, were detected by means of MS-SWATCH analysis. These included components of the type III secretion system and proteins involved in bacterial motility. Our results indicate that Lon protease is important for D. solani to withstand stressful conditions and effectively invade the potato plant.
Project description:To identify differently expressed proteins in tuber tissue of potato cultivars and diploid interspecific hybrids of Solanum, differing in resistance to Dickeya solani, comparative analysis was performed. Two highly resistant (Bea and Humalda) and three susceptible (Irys, Katahdin, Ulster Supreme) potato cultivars, as well as the highly resistant (DG 00-270) and the susceptible (DG 08-305) diploid clones, were studied. DG 00-270 exhibited higher resistance to D. solani than the cultivars Bea and Humalda. Proteins were extracted from wounded potato tubers inoculated with bacteria at an early symptomatic phase and from controls, i.e., intact tubers and wounded mock-inoculated tubers. Protein profiles were analyzed using nano-liquid chromatography coupled with tandem mass spectrometry (LC-MS-MS/MS).
Project description:The DsbA oxidoreductase is a crucial factor responsible for introduction of disulfide bonds to the extracytoplasmic proteins in bacteria. A lack of the proper disulfides frequently leads to instability and/or loss of protein function. In pathogens, numerous envelope and extracellular proteins play important roles in pathogenesis; therefore, their improper disulfide bonding may lead to avirulent phenotypes. The importance of the DsbA function in phytopathogens has not been extensively studied yet. Dickeya solani is a bacterium from the Soft Rot Pectobacteriaceae group which is responsible for very high economic losses mainly on potato. In recent years, D. solani became the most abundant potato pathogen among Dickeya species in Europe. In this work, using the D. solani dsbA mutant, we demonstrated that a lack of the DsbA function caused loss of virulence. Mutant bacteria were deficient in most secreted virulence determinants and were not able to develop disease symptoms in the natural host, the potato plant. The SWATH-MS-based proteomic analysis revealed that the dbsA mutation led to multifaceted effects in the D. solani cells. First of all, the levels of the majority of plant cell wall degrading enzymes and proteins related to motility and chemotaxis were severely reduced. Furthermore, the protein profiles suggested induction of the envelope and cytoplasm stress responses in the mutant cells. Finally, the outer membrane barrier seemed to be disturbed by the mutation. Our results clearly demonstrate that the function played by the DsbA oxidoreductase is indispensable for D. solani virulence and a lack of DsbA significantly disturbs cellular physiology. A thorough analysis of proteomic research suggests that a lack of virulence may result from both, abnormalities of the disulfide deprived virulence determinants and the envelope stress-dependent repression of the virulence genes in the dsbA mutant.
Project description:Lytic bacteriophages able to infect and kill Dickeya spp. can be readily isolated from virtually all Dickeya spp.-containing environments, yet little is known about the selective pressure those viruses exert on their hosts. Here, we identified two spontaneous phage-resistant D. solani IPO 2222 mutants, DsR34 and DsR207, resistant to infection caused by phage vB_Dsol_D5 (ΦD5) that expressed a reduced ability to macerate potato tuber tissues compared to the wild-type, phage-susceptible D. solani IPO 2222 strain. Genome sequencing revealed that mutants had point mutations in two genes encoding: secretion protein HlyD (mutant DsR34) and elongation factor Tu (EF-Tu) (mutant DsR207). Both mutations impacted the proteoms of D. solani grown in rich and minimal media. Furthermore, DsR34 and DsR207 were characterized for features essential for their ecological success in a plant environment, including the ability to use various carbon and nitrogen sources, production of plant cell wall degrading enzymes, ability to form biofilms, siderophore production, swimming and swarming motility and virulence in planta. Compared to the wild-type ΦD5-susceptible D. solani strain, mutants DsR34 and DsR207 expressed reduced ability to macerate chicory leaves and to colonize and cause symptoms in growing potato plants. The implications of the ΦD5 resistance on the ecological performance of D. solani are discussed.
2023-04-28 | PXD038825 | Pride
Project description:The gene expression profiling of potato in response to only Serratia plymuthica OR in combination with Dickeya solani in a time course RNA-Seq
Project description:Improved understanding of bacterial-fungal interactions in the rhizosphere should assist in the successful application of bacteria as biological control agents against fungal pathogens of plants, providing alternatives to chemicals in sustainable agriculture. To understand the functional response of the fungal phytopathogen Rhizoctonia solani to different bacteria and to elucidate whether the molecular mechanisms that the fungus exploits involve general stress or more specific responses, we performed a global transcriptome profiling of R. solani Rhs1AP anastomosis group 3 (AG-3) during interaction with the S4 and AS13 species of Serratia using RNA-seq. Transcriptome analysis revealed that approximately 10% of the fungal transcriptome was differentially expressed during challenge with Serratia. The numbers of S4- and AS13-specific differentially expressed genes (DEG) were 866 and 292 respectively, while there were 1035 common DEGs in the two treatment groups. Four hundred and sixty and 242 genes respectively had fold values exceeding 8x and for further analyses this cut-off value was used. Functional classification of DEGs revealed a general shift in fungal gene expression in which genes related to xenobiotic degradation, toxin and antioxidant production, energy, carbohydrate and lipid metabolism and hyphal rearrangements were subjected to transcriptional regulation. In conclusion, it was found out that most genes were regulated in the same way in the presence of both bacterial isolates, but there were also some strain-specific responses. The findings in this study will be beneficial for further research on biological control and in depth exploration of bacterial-fungal interactions in the rhizosphere.
Project description:Differential analysis of the potato-Rhizoctonia solani AG3 interaction. Samples were extracted from R. solani inoculated potato sprouts at two time points. R. solani is one of the most prominent fungal pests of potato and therefore of great economic relevance.
Project description:Volatiles of certain rhizobacteria can cause growth inhibitory effects on plants/ Arabidopsis thaliana. How these effects are initiated and which mechanisms are enrolled is not yet understood. Obviously the plant can survive/live with the bacteria in the soil, which suggest the existance of a regulatory mechanism/network that provide the possibility for coexistance with the bacteria. To shed light on this regulatory mechanism/network we performed a microarray anlaysis of Arabidopsis thaliana co-cultivated with two different rhizobacteria strains. In this study we used the ATH1 GeneChip microarray to investigate the transcriptional response of 4 to 5 days old Arabidopsis thaliana seedlings at 6 h, 12 h and 24 h exposure to volatiles of the rhizobacteria Serratia plymuthica HRO-C48 or Stenotrophomonas maltophilia R3089.
Project description:Rhizoctonia solani is a nectrotrophic fungal pathogen that causes billions of dollars of damage to agriculture worldwide and infects a broad host range including wheat, rice, potato and legumes. In this study we identify wheat genes that are differentially expressed in response to the R. solani isolate, AG8-1, using microarray technology. A significant number of wheat genes identified in this screen were involved in ROS production and redox regulation. Levels of ROS species were increased in wheat root tissue following R. solani infection as determined by NBT, DAB and titanium sulphate measurements/stainings. Pathogen/ROS related genes from R. solani were also tested for expression patterns upon wheat infection. TmpL, a R. solani gene homologous to a gene associated with ROS regulation in Alternaria brassicicola, and OAH, a R. solani gene homologous to oxaloacetate acetylhydrolase which has been shown to produce oxalic acid in Sclerotinia sclerotiorum, were highly induced in R.solani when infecting wheat. We speculate that the wheat germin-like protein (GLP) is induced to inactivate the oxalic acid that is produced by the R. solani OAH.
Project description:Previously, we investigated the effect of fungal VOCs on the behavior of phylogenetically different soil bacteria (Schmidt et al 2015). In these experiments we showed that VOCs emitted by several fungi can lead to phenotypical responses in bacteria, for example, by inducing a change in motility (Schmidt et al 2015). We observed that the plant pathogenic fungus Fusarium culmorum produced a unique cluster of VOCs consisting primarily of terpenes. When exposed to the VOCs emitted by this fungus, the rhizobacterium Serratia plymuthica PRI-2C responded with an induction of motility. It is plausible that in soil, microorganisms sense changes in their environments via shifts in VOCs blend and adapt their behavior accordingly (Garbeva et al 2014). Although several studies indicated that VOCs can be used as signaling molecules in microbial inter-species interactions, the following questions remain unanswered as how are VOCs perceived as signals by the microorganisms and which regulatory pathways and genes are involved in the response? To answer these questions, the rhizosphere isolate S. plymuthica PRI-2C was grown alone or exposed to VOCs emitted by F. culmorum. The bacterial transcriptome and proteome were analyzed under each situation to identify the molecular basis of the bacterial response to fungal VOCs.