MUTANTS OF RALSTONIA (PSEUDOMONAS) SOLANACEARUM SENSITIVE TO ANTIMICROBIAL PEPTIDES ARE ALTERED IN THEIR LIPOPOLYSACCHARIDE STRUCTURE AND ARE AVIRULENT IN TOBACCO

Ralstonia solanacearum K60 was mutagenized with the transposon Tn5, an d two mutants, M2 and M88, were isolated, Both mutants were selected b ased on their increased sensitivity to thionins, and they had the Tn5 insertion in the same gene, 34 bp apart, Sequence analysis of the inte rrupted gene showed clear homology with the rfaF gene from Escherichia coli and Salmonella typhimurium (66% similarity), which encodes a hep tosyl-transferase involved in the synthesis of the lipopolysaccharide (LPS) core, Mutants M2 and M88 had an altered LPS electrophoretic patt ern, consistent with synthesis of incomplete LPS cores, For these reas ons, the R. solanacearum gene was designated rfaF. The mutants were al so sensitive to purified lipid transfer proteins (LTPs) and to an LTP- enriched, cell wall extract from tobacco leaves, Mutants M2 and M88 di ed rapidly in planta and failed to produce necrosis when infiltrated i n tobacco leaves or to cause wilting when injected in tobacco stems, C omplemented strains M2 and M88* were respectively obtained from mutan ts M2 and M88 by transformation with a DNA fragment harbouring gene rf aF. They had a different degree of wild-type reconstituted phenotype. Both strains retained the rough phenotype of the mutants, and their LP S electrophoretic patterns were intermediate between those of the wild type and those of the mutants.