Degradation of acyl-homoserine lactone signalling molecules in the plant pathogen Erwinia carotovora

Erwinia carotovora is a Gram-negative plant pathogen affecting several commercially important crops such as potatoes and tobacco. Virulence in many strains of E. carotovora is controlled by the quorum sensing signalling molecule N-(3-oxohexanoyl)-L-homoserine lactone (OHHL) which accumulates as the bacteria multiply and activates exoenzyme and secondary metabolite production. Mutant strains lacking OHHL are avirulent, suggesting a preventative treatment for infection. It had previously been observed in E. carotovora that OHHL accumulates in a cell density dependent fashion until stationary phase when the concentration falls in an equally rapid manner. This work shows that the stationary phase decline in OHHL is due to alkalisation of the growth media and that growing in buffered media can prevent this decline. Functional genomics was used to explore the possibility that E. carotovora subsp. atroseptica might possess an OHHL degrading enzyme similar to AttM from Agrobacterium tumefaciens. Four AttM orthologues were identified in the Eca strain 1043 genome, with one of these proteins (209-2), showing some degrading activity against synthetic HHL. Mutants were created in each of the orthologues but they had no effect on OHHL accumulation or exoenzyme production. The mutation in the 209-1 gene reduced motility, while mutation in the 91 gene reduced virulence in potato tubers. Further investigation using proteomic analysis found three of the mutants had reduced levels of Re1A, which is involved in regulation of the attM operon in A. tumefaciens. This study shows that E. carotovora would exhibit reduced virulence in basic environments due to increased AHL lability. Only one of the AttM orthologues identified possessed AHL degrading ability under the conditions tested but three did effect the expression of AttM, suggesting potential involvement in AHL regulation.EThOS - Electronic Theses Online ServiceGBUnited Kingdo