dltA gene mutation in the teichoic acids alanylation system of Lactococcus garvieae results in diminished proliferation in its natural host.

The dltABCD cluster is involved in the d-alanylation of teichoic acids in gram positive bacteria. In order to determine the role of this alanylation in the physiology and virulence of Lactococcus garvieae, a previously isolated dltA Delta Tn917 signature tagged mutagenesis (STM) clone was analyzed. RT-PCR results revealed that dltABCD genes form an operon. No major differences could be established between the parental and mutant strains with respect to growth rate, autolytic properties, and susceptibility to acid conditions, lysozyme treatment, anionic detergents, or oxidant agents. However, the dltA mutant was more susceptible to nisin than the parental strain, with minimum inhibitory concentration (MIC) values of 8 and 16 microg/ml, respectively. Less proliferation of the mutant was observed in in vivo competence index experiments (CI=0.08). Furthermore, the mutant strain had a 50% lethal dose (LD(50)) 3-fold that of the parental strain. These results, together with the fact that the dltA Delta Tn917 mutant was isolated as a STM clone, reveal that the dltA locus of Lactococcus garvieae is required for full growth and pathogenesis on rainbow trout.