Pigment and virulence deficiencies associated with mutations in the aroE gene of Xanthomonas oryzae pv. oryzae

Xanthomonadins are yellow, membrane-bound pigments produced by members of t he genus Xanthomonas, We identified an ethyl methanesulfonate-induced Xanth omonas oryzae pv. oryzae mutant (BXO65) that is deficient for xanthomonadin production and virulence on rice, as well as auxotrophic for aromatic amin o acids (Pig(-) Vir(-) Aro(-)). Reversion analysis indicated that these mul tiple phenotypes are due to a single mutation. A genomic library of the wil d-type strain was used to isolate a 7.0-kb clone that complements BXO65, By transposon mutagenesis, marker exchange, sequence analysis, and subcloning , the complementing activity was localized to a 849-bp open reading frame ( ORF), This ORF is homologous to the aroE gene, which encodes shikimate dehy drogenase in various bacterial species. Shikimate dehydrogenase activity wa s present in the wild-type strain and the mutant,vith the complementing clo ne, whereas no activity was found in BXO65. This clone also complemented an Escherichia coli aroE mutant for prototrophy, indicating that aroE is func tionally conserved in X. oryzae pv. oryzae and E. coil, The nucleotide sequ ence of the 2.9-kb region containing aroE revealed that a putative DNA heli case gene is located adjacent to aroE, Our results indicate that aroE is re quired for normal levels of virulence and xanthomonadin production in X ory zae pv, oryzae.