Instability of short-sequence DNA repeats of pear pathogenic Erwinia strains from Japan and Erwinia amylovora fruit tree and raspberry strains

Abstract

An array of short-sequence DNA repeats (SSRs) occurs in the plasmid pEA29 of the fire blight pathogen Erwinia amylovora. A large number of "fruit tree" strains, mainly from Central and Western Europe, were screened for their SSR numbers, and the analyses were extended to five raspberry strains from North America and six pear pathogenic Erwinia strains from Japan. The repeat ATTACAGA present in all E. amylovorastrains was found to be reiterated 3 to 15 times. The Japanese strains contained the major repeat sequence GGATTCTG, which was reiterated 16 to 24 times. ATTACAGG, which resembles the SSR of E. amylovora, was reiterated two or three times. In a novel approach, sequencing gels were used to visualize the rare occurrence of shorter arrays (down to three repeats) in E. amylovoraand the Japanese Erwinia strains. Changes in the repeat numbers in E. amylovora were observed repeatedly when the bacteria had been exposed to stress conditions. The repeat structures of homo- and heteroduplices of PCR-amplified repeats were also analyzed by cleavage of annealed molecules with the single-strand-specific endonuclease from bacteriophage T4. Not only heteroduplexes, but also homoduplexes showed non-matching regions in the SSRs, which could arise from transient formation of loops due to strand slippage during the assays.