ERWINIA-CHRYSANTHEMI HRP GENES AND THEIR INVOLVEMENT IN SOFT-ROT PATHOGENESIS AND ELICITATION OF THE HYPERSENSITIVE RESPONSE

Unlike the bacterial pathogens that typically cause the hypersensitive response (HR) in plants, Erwinia chrysanthemi has a wide host range, rapidly kills and macerates host tissues, and secretes several isozymes of the macerating enzyme pectate lyase (Pel). PelABCE(-) and Out(-) (secretion-deficient) mutants were observed to produce a rapid necrosis in tobacco leaves that was indistinguishable from the HR elicited by the narrow-host-range pathogens E. amylovora Ea321 and Pseudomonas syringae pv. syringae 61. E. amylovora Ea321 hrp genes were used to identify hybridizing cosmids in a cosmid library of E. chrysanthemi EC16 DNA in Escherichia coli. A 16-kb BamHI fragment in one of these cosmids, pCPP2030, hybridized with E. amylovora hrp genes and was mutagenized with Tn10mini-kan. The mutations were introduced into the PelABCE(-) mutant CUCPB5006 by marker exchange. Two of the resultant hrp::Tn10mini-kan mutations were found to abolish the ability of CUCPB5006 to cause any necrosis in tobacco leaves unless complemented with pCPP2030. These two mutations were also marker-exchanged into the genome of wild-type strain AC4150. Analysis of DNA sequences flanking the hrp-2::Tn10mini-kan insertion revealed the mutated gene to be similar to a gene in E. amylovora Ea321 hrp complementation group VIII and to P. s. pv. syringae 61 hrpX. Neither of the hrp::Tn10mini-kan mutations affected the production or secretion of pectic enzymes by AC4150 or CUCPB5006. However, the hrp mutations reduced the ability of both AC4150 and CUCPB5006 to incite successful infections in witloof chicory leaves. The results indicate that E. chrysanthemi can elicit an Hrp-dependent HR and support the hypothesis that the Hrp system has a significant role in an early stage of pathogenesis by E. chrysanthemi.