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Aeromonas hydrophila  is a globally occurring, potentially virulent, gram-negative opportunistic pathogen that is known to cause water and food-borne diseases around the world. In this study, we use whole genome sequencing and  in silico  analyses to identify 14 putative O antigen gene clusters (OGCs) located downstream of the housekeeping genes  acrB  and/or  oprM . We have also identified 7 novel OGCs by analyzing 15 publicly available genomes of different  A .  hydrophila  strains. From the 14 OGCs identified initially, we have deduced that O antigen processing genes involved in the  wzx / wzy  pathway and the ABC transporter ( wzm / wzt ) pathway exhibit high molecular diversity among different  A .  hydrophila  strains. Using these genes, we have developed a multiplexed Luminex-based array system that can identify up to 14  A .  hydrophila  strains. By combining our other results and including the sequences of processing genes from 13 other OGCs (7 OGCs identified from publicly available genome sequences and 6 OGCs that were previously published), we also have the data to create an array system that can identify 25 different  A .  hydrophila  serotypes. Although clinical detection, epidemiological surveillance, and tracing of pathogenic bacteria are typically done using serotyping methods that rely on identifying bacterial surface O antigens through agglutination reactions with antisera, molecular methods such as the one we have developed may be quicker and more cost effective. Our assay shows high specificity, reproducibility, and sensitivity, being able to classify  A .  hydrophila  strains using just 0.1 ng of genomic DNA. In conclusion, our findings indicate that a molecular serotyping system for  A .  hydrophila  could be developed based on specific genes, providing an important molecular tool for the identification of  A .  hydrophila  serotypes.

Identifying genetic diversity of O antigens in Aeromonas hydrophila for molecular serotype detection