Genetic Analysis of a Pathogenic Erwinia sp. Isolated from Pear in Japan

Abstract

Four Erwinia strains, originally isolated in Japan from pear trees with bacterial shoot blight symptoms, were analyzed to determine their genetic relationship with Erwinia amylovora and E. pyrifoliae. When genomes were characterized with amplified fragment length polymorphism markers and by comparative groEL sequence analysis, the Japanese Erwinia sp. and South Korean E. pyrifoliae strains were placed in the same group, which was phylogenetically distinct from a group of 15 strains of E. amylovora. Sequencing of the 29,593-bp plasmid pEJ30 from Erwinia strain Ejp556 revealed that this plasmid was nearly identical to plasmid pEP36 from E. pyrifoliae and was closely related to the nontransferable ubiquitous plasmid pEA29 from E. amylovora. Twenty-one presumptive genes and their order in pEP36 were highly conserved in pEJ30; however, transposon Tn5394, which was present in pEP36, was not found in pEJ30. Short-sequence DNA repeats were conserved between pEJ30 and pEP36, and were different from short-sequence repeats in pEA29. Despite base-pair mismatches, primer pairs used in pEA29 polymerase chain reaction assays for E. amylovora amplified plasmid DNA from the Japanese Erwinia Ejp556 and Ejp562. Like E. pyrifoliae and a few strains of E. amylovora, Japanese Erwinia Ejp617 contained plasmids related to E. pyrifoliae ColE1-related plasmid pEP2.6. Based on these genetic analyses, we conclude that the Erwinia pathogen of pear in Japan is closely related to E. pyrifoliae and that both of these pathogens are demonstrably distinct from E. amylovora.

Nucleotide Sequences, Genetic Organization, and Distribution of pEU30 and pEL60 from Erwinia amylovora

“…We selected 54 strains for plasmid analysis (Table 1) from a collection of 167 E. amylovora strains from the United States and Canada (8,11), and Lebanon (17) that were previously uncharacterized for plasmid content. Plasmid isolations were done using the method of Kado and Liu (6) as modified by Sundin et al (19).…”

“…1-to 2.5-kb fragments that were excised and purified using a Nucleotrap gel extraction kit (Clontech Laboratories) according to the manufacturer's instructions except that the DNA was eluted with water at 80°C. Fragment ligations into pGem7zf-(Promega, Madison, Wis.) and subsequent preparative steps for high-throughput sequencing were performed as previously described (8). All sequencing reactions were run at the Michigan State University Genomics Technology Support Facility.…”

Genetic Differences between Blight-Causing Erwinia Species with Differing Host Specificities, Identified by Suppression Subtractive Hybridization

“…JE1 contained ejp19, a putative membrane ABC transporter previously identified in Ejp556 (16), and JE2, JE3, and JE4 also contained gene sequences with similarity to hypothetical transmembrane proteins. Previous work has reported that differences between Rubus strains and other isolates of E. amylovora include several membrane proteins and an ABC transporter (2).…”

“…are unable to infect apple and pear (2,9). The Rubus-infecting strains of E. amylovora are closely related to the apple-and pear-infecting strains, as evidenced by similar amplified fragment length polymorphism and PCR fingerprints, species-level total DNA-DNA homology, and nearly identical sequences in pathogenicity and virulence genes (7,11,(16)(17)(18). Recently, other blight-causing Erwinia sp.…”

Caracterización fenotípica y genotípica de aislados españoles de Erwinia amylovora.

“…Aunque como patógeno no es tan bien conocida, se han identificado tres grandes grupos de genes implicados en su patogenicidad: los genes ams, responsables de la biosíntesis del exopolisacárido amilovorano; los genes hrp (hypersensitive response and pathogenicity), necesarios para el poder patógeno y la inducción de la respuesta hipersensible; y los genes dsp (disease specific), implicados en el desarrollo de la enfermedad. Se ha identificado, de forma adicional, otro factor que interviene en la virulencia de E. amylovora: la producción del sideróforo desferrioxamina E. Asimismo, se ha propuesto también un papel en la virulencia para el plásmido de 29 kb de E. amylovora (pEA29) .Uno de los rasgos más característicos y sorprendentes de esta especie es su gran homogeneidad, tanto a nivel fenotípico como genotípico (Vanneste, 1995 Maxon-Stein, 2003). Al margen de estas dos excepciones, la profundización en el conocimiento de la diversidad del resto de cepas de este patógeno sería de gran utilidad en el seguimiento de la propagación de E. amylovora en el espacio y en el tiempo.…”