Transposition of the heat-stable toxin astA gene into a gifsy-2-related prophage of Salmonella enterica serovar Abortusovis

Abstract

The horizontal transfer and acquisition of virulence genes via mobile genetic elements have been a major driving force in the evolution of Salmonella pathogenicity. Serovars of Salmonella enterica carry variable assortments of phage-encoded virulence genes, suggesting that temperate phages play a pivotal role in this process. Epidemic isolates of S. enterica serovar Typhimurium are consistently lysogenic for two lambdoid phages, Gifsy-1 and Gifsy-2, carrying known virulence genes. Other serovars of S. enterica, including serovars Dublin, Gallinarum, Enteritidis, and Hadar, carry distinct prophages with similarity to the Gifsy phages. In this study, we analyzed Gifsy-related loci from S. enterica serovar Abortusovis, a pathogen associated exclusively with ovine infection. A cryptic prophage, closely related to serovar Typhimurium phage Gifsy-2, was identified. This element, named Gifsy-2AO, was shown to contribute to serovar Abortusovis systemic infection in lambs. Sequence analysis of the prophage b region showed a large deletion which covers genes encoding phage tail fiber proteins and putative virulence factors, including type III secreted effector protein SseI (GtgB, SrfH). This deletion was identified in most of the serovar Abortusovis isolates tested and might be dependent on the replicative transposition of an adjacent insertion sequence, IS1414, previously identified in pathogenic Escherichia coli strains. IS1414 encodes heat-stable toxin EAST1 (astA) and showed multiple genomic copies in isolates of serovar Abortusovis. To our knowledge, this is the first evidence of intergeneric transfer of virulence genes via insertion sequence elements in Salmonella. The acquisition of IS1414 (EAST1) and its frequent transposition within the chromosome might improve the fitness of serovar Abortusovis within its narrow ecological niche.

FIG. 1.

Gene organization within the “right” end regions of Gifsy-2 (serovar Typhimurium) and Gifsy-2AO (serovar Abortusovis) prophages. Black arrows show genes with a role in production of phage particles, named according to their phage lambda orthologs. Gray arrows show genes within the b region. Sequence data are from reference and this study. IR, IS1414 inverted repeated sequences.

FIG. 2.

Relative levels of invasiveness of S. enterica serovar Abortusovis strains SS44 (wild type), SSM2992 (ΔGifsy-2AO), and SSM916 (ΔinvH) in ovine ileal loops; each bar represents the mean ± the standard error of the mean of results from six loops tested. Three samples from each loop were analyzed. For the results for SS44 versus those for SSM916, the P value was <000.1.

FIG. 3.

Southern blot hybridization of serovar Abortusovis epidemic strains with the IS1414 (astA) probe. Multiple genomic copies of IS1414 (astA) were detected in the chromosomes of serovar Abortusovis epidemic isolates from Albania (strains SSM0074 [lane 1] and SSM0075 [lane 2]), Sardinia (strains SSM0078 [lane 3], SSM0088 [lane 4], and SSM0096 [lane 5]), Iran (strains SSM2045 [lane 6], SSM2046 [lane 7], SSM2026 [lane 8], and SSM2027 [lane 9]), and Russia (strain SSM0041 [lane 10]). Serovar Typhimurium strain ATCC 14028s (lane 11) and ETEC strain SSM3422 (lane 12) were used as negative and positive controls, respectively.