Genomic characterization of an atypical hydrogen sulfide-negative Salmonella enterica serovar Senftenberg strain lacking somatic antigen expression isolated from cooked mussels

Atypical Salmonella enterica strains that evade conventional detection pose significant challenges to food safety surveillance. A hydrogen sulfide (H2S)-negative and serologically untypable S. enterica strain (SF1060) was detected by qPCR from cooked farmed mussels in Galicia, Spain, and characterized using phenotypic and genomic approaches. Despite typical biochemical profiles, SF1060 failed to produce black colonies on XLD agar and lacked detectable somatic antigens by conventional serotyping. Hybrid genome assembly using Nanopore and Illumina sequencing yielded a closed chromosome and five plasmids. In silico analyses identified the strain as S. Senftenberg ST14. Comparative genomics revealed a chromosomal inversion at the rfb operon (encoding enzymes needed to synthesize deoxysugars and O antigens) mediated by IS5-family transposase ISEc68, which truncated the rfbD gene and separated the reminding rfb genes at rfbD, disrupting O-antigen biosynthesis, explaining the serotyping failure. The phs operon responsible for H2S production lacked premature stop codons, suggesting the H2S-negative phenotype results from an alternative mechanism. This study demonstrates how whole-genome sequencing resolves identification of atypical strains that fail culture-based detection and emphasizes the critical need for molecular surveillance methods in seafood safety programs, particularly in regions where atypical S. enterica variants may be endemic. Importance StatementPathogen surveillance is in a constant race against microbial evolution. The phenotypic methods used to detect and isolate foodborne bacteria like Salmonella enterica from foods are effective, but only if the pathogens retain their expected characteristics. This study provides a clear genetic snapshot of how Salmonella enterica can adapt to evade detection. This study characterizes a strain from a marine environment that had undergone genetic rearrangement, leading to the concurrent loss of a key biochemical marker and the somatic antigens essential for serological typing. Crucially, only genomic analysis could provide a definitive explanation for this serotyping failure; the underlying mechanism would have remained unknown using conventional methods alone. This work demonstrates that relying on a fixed set of phenotypic traits is an increasingly fragile strategy and highlights why genomic surveillance must become a routine component of food safety programs to keep pace with pathogen evolution.