Methods: Bacterial isolates were tested for Stx expression using the ImmunoCard STAT! EHEC assay. Phage were induced by addition of 0.5µg/mL mitomycin C to pure cultures. Phage genomic DNA was purified from induced cultures by phage particle precipitation followed by anion exchange chromatography. Genomic DNA from the phage and lysogenized host were sequenced using the Illumina MiSeq platform, and the phage genome was assembled using SPAdes v3.5. Phylogenetic analysis of the phage genome was performed using Neighbor-joining tree inference with 100 bootstrap iterations.
Results: We describe two Stx-producing S. sonnei isolates from patients who presented to our hospital with diarrheal illnesses, both with history of travel to Baja California, Mexico. PCR analysis was performed on both isolates, revealing the presence of the stx1a gene. Treatment of clinical isolate cultures with mitomycin C induced bacterial lysis and the release of a stx1a-harboring phage. Deep sequencing of DNA from purified phage particles revealed a novel 62kb phage, named ɸSs-VASD. Mapping of the phage genomic sequence to the lysogenized host chromosome revealed the integration site to be the wrbA gene in both isolates, and phylogenic analysis revealed a close relationship to stx2-converting phages of STEC.
Conclusion: Our findings demonstrate the emergence of shiga-toxigenic S. sonnei in North America, and implicate phage-mediated horizontal gene transfer from STEC as the mechanism of toxin acquisition.
W. W. Chiu, None
D. Looney, None
M. Strain, None
S. Mehta, None