Development of strain specific simple sequence repeats and assessment of genetic diversity in Erwinia amylovora from marker selection to phylogenetic analysis

Erwinia amylovora is a highly destructive phytopathogenic bacterium that causes fire blight in Rosaceae plants, particularly apple and pear trees. Due to its status as a quarantine organism in many countries, extensive epidemiological and genotypic studies have been conducted to better understand and manage this pathogen. This study presents a novel workflow for identifying strain-specific simple sequence repeats (SSRs) in bacterial genomes, focusing on E. amylovora. Leveraging the abundance of available genome data, the workflow involves identifying single-copy genes via BLASTX searches, followed by SSR identification using the MISA tool and validation through sequence alignment. Utilizing the genome of E. amylovora strain CFBP 1430 as a reference, 27 SSRs within 26 single-copy genes were identified, displaying diverse repeat patterns and lengths. Five genes containing SSRs were selected, offering valuable markers for strain differentiation. Phylogenetic analysis of 114 E. amylovora strains revealed three distinct groups: widely prevalent, Western NA, and Eastern NA clades, in line with previous findings. A comparative analysis with clustered regularly interspaced short palindromic repeat (CRISPR) analysis and multiple-locus variable-number tandem repeat analysis (MLVA) highlighted differences in marker selection strategies and underscored the need to validate findings using multiple genotyping approaches. While limitations remain due to incomplete nucleotide sequences in public databases, this study advances our understanding of the genetic diversity, population structure, and evolutionary dynamics of E. amylovora strains, and positions SSR-based methods as a complementary tool alongside other established techniques.