Twenty years of molecular genetics with Erwinia amylovora: answers and new questions about eps-synthesis and other virulence factors.

摘要

Erwinia amylovora and many other plant-associated bacteria produce exopolysaccharides (EPS) of high molecular weight. Levan is a homopolymer of fructose synthesized by secreted levansucrase. During plant colonization, it may partially substitute for a more complex capsular EPS. E. amylovora synthesizes amylovoran, a high molecular weight EPS assembled from repeating units with galactose, glucuronic acid and often glucose. Genes for amylovoran synthesis are located in the ams region. The encoded proteins act as sugar transferases or participate in transport and polymerization of the repeating units. E. amylovora ams mutants were not virulent in plant assays. Similar to degradation of the EPS capsules with a viral EPS depolymerase, cells without capsules are exposed to plant defense mechanisms interfering with pathogen colonization. Amylovoran synthesis is regulated by a two-component system with genes rcsA, rcsB and rcsC or by the global regulator hns. The expression of levansucrase depends on the activator proteins RlsA, RlsB, and RISC. Detection of E. amylovora from plant samples is done by plating on semi-selective agar plates, and by PCR with primers derived from plasmid pEA29 and the ams-region on the chromosome. Recently, we applied plasmid primers to real-time PCR. Differentiation of E. amylovora strains can efficiently be done by PFGE analysis, and dendrograms were also constructed from amino acid sequences derived from hrpN genes of E. amylovora strains and the Asian pear pathogens..