The GDSL-Lipolytic Enzyme Lip1 Is Required for Full Virulence of the Cucurbit Pathogenic Bacterium Acidovorax citrulli

Bacterial fruit blotch caused by Acidovorax citrulli is a serious disease of cucurbit crops. Here we report characterization of a mutant strain of A. citrulli M6 defective in lip1, a gene encoding a lipolytic enzyme. The M6-lip1(-) mutant was detected in a mutant library screen aimed at identifying M6 mutants with altered levels of twitching motility. In this screen M6-lip1(-) was the only mutant that showed significantly larger twitching motility haloes around colonies than wild-type M6. Sequence analyses indicated that lip1 encodes a member of the GDSL family of secreted lipolytic enzymes. In line with this finding, lipolytic assays showed that the supernatants of M6-lip1(-) had lower lipolytic activity as compared with those of wild-type M6 and a lip1-complemented strain. The mutant was also affected in swimming motility and had compromised virulence on melon seedlings and on Nicotiana benthamiana leaves relative to wild-type and complemented strains. Lip1 contains a predicted N-terminal signal sequence for type II secretion. Evidence from our study confirms Lip1 is indeed secreted in a type II secretion-dependent manner, and this is required for full virulence of A. citrulli. To the best of our knowledge this is the first study reporting contribution of lipolytic activity to virulence of a plant-pathogenic Acidovorax species.