Abstract
BackgroundRalstonia solanacearum, the causal agent of bacterial wilt, is a genetically diverse bacterial plant pathogen present in tropical and subtropical regions of the world that infects more than 200 plant species, including economically important solanaceous crops. Most strains of R. solanacearum are only pathogenic at temperatures between 25 to 30°C with strains that can cause disease below 20°C considered a threat to agriculture in temperate areas. Identifying key molecular factors that distinguish strains virulent at cold temperatures from ones that are not is needed to develop effective management tools for this pathogen. We compared protein profiles of two strains virulent at low temperature and two strains not virulent at low temperature when incubated in the rhizosphere of tomato seedlings at 30 and 18°C using quantitative 2D DIGE gel methods. Spot intensities were quantified and compared, and differentially expressed proteins were sequenced and identified by mass spectrometry (MS/MS).ResultsFour hundred and eighteen (418) differentially expressed protein spots sequenced produced 101 unique proteins. The identified proteins were classified in the Gene Ontology biological processes categories of metabolism, cell processes, stress response, transport, secretion, motility, and virulence. Identified virulence factors included catalase (KatE), exoglucanase A (ChbA), drug efflux pump, and twitching motility porin (PilQ). Other proteins identified included two components of a putative type VI secretion system. We confirmed differential expression of 13 candidate genes using real time PCR techniques. Global regulators HrpB and HrpG also had temperature dependent expression when quantified by real time PCR.ConclusionsThe putative involvement of the identified proteins in virulence at low temperature is discussed. The discovery of a functional type VI secretion system provides a new potential virulence mechanism to explore. The global regulators HrpG and HrpB, and the protein expression profiles identified suggest that virulence at low temperatures can be partially explained by differences in regulation of virulence factors present in all the strains.
Highlights
Ralstonia solanacearum, the causal agent of bacterial wilt, is a genetically diverse bacterial plant pathogen present in tropical and subtropical regions of the world that infects more than 200 plant species, including economically important solanaceous crops
Comparative protein profiles of Ralstonia solanacearum strains at 30°C and 18°C Comparison at 30°C and 18°C of cell-associated proteins from strain P597 incubated in rich media in absence of tomato plants identified 61 protein spots differentially expressed in a preliminary experiment
Cell-associated comparative protein profiles at two temperatures obtained from bacterial cultures of strains GMI1000, P597, P673 and UW551 incubated in contact with tomato seedlings rhizosphere revealed 872 differentially expressed protein spots, determined by statistically differential intensities for each strains at two temperatures
Summary
The causal agent of bacterial wilt, is a genetically diverse bacterial plant pathogen present in tropical and subtropical regions of the world that infects more than 200 plant species, including economically important solanaceous crops. Sudden changes in temperature can induce adaptive shock responses in bacteria, enabling them to colonize widely diverse environments. These adaptive responses involve major changes in the physiological state of the bacterial cells [1,2] allowing them to survive and function in the new environment. The risk of introducing such strains to new crop zones could be economically devastating These pathogenicity differences may involve changes in protein regulation or may be due to the presence of unique genes involved in enabling virulence at low temperatures. This is the case of the bacterial wilt pathogen Ralstonia solanacearum
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