Molecular evolution of virulence in natural field strains of Xanthomonas campestris pv. vesicatoria

The avrBs2 avirulence gene of the bacterial plant pathogen Xanthomonas camp estris pv, vesicatoria triggers disease resistance in pepper plants contain ing the Bs2 resistance gene and contributes to bacterial virulence on susce ptible host plants. We studied the effects of the pepper Bs2 gene on the ev olution of avrBs2 by characterizing the molecular basis for virulence of 20 X. campestris pv. vesicatoria field strains that were isolated from diseas e spots on previously resistant Bs2 pepper plants. All held strains tested were complemented by a wild-type copy of avrBs2 in their ability to trigger disease resistance on Bs2 plants. DNA sequencing revealed four mutant alle les of avrBs2, two of which consisted of insertions or deletions of 5 nucle otides in a repetitive region of avrBs2. The other two avrBs2 alleles were characterized by point mutations with resulting single amino acid changes ( R403P or A410D). We generated isogenic X. campestris pv. vesicatoria strain s by chromosomal avrBs2 gene exchange to study the effects of these mutatio ns on the dual functions of avrBs2 in enhancing bacterial virulence and ind ucing plant resistance by in planta bacterial growth experiments. The delet ion of 5 nucleotides led to loss of avrBs2-induced resistance on Bs2 pepper plants and abolition of avrBs2-mediated enhancement of fitness on suscepti ble plants. Significantly, the point mutations led to minimal reduction in virulence function of avrBs2 on susceptible pepper plants, with either mini mal (R403P allele) or an intermediate level of (A410D allele) triggering of resistance on Bs2 plants. Consistent with the divergent selection pressure s on avrBs2 exerted by the Bs2 resistance gene, our results show that avrBs 2 is evolving to decrease detection by the Bs2 gene while at the same time maintaining its virulence function.