Cell SELEX-based DNA aptamer selection targeting Streptococcus agalactiae using high-throughput sequencing: toward multifunctional applications in aquaculture.

Streptococcus agalactiae is a Gram-positive bacterial pathogen accountable for disease outbreaks in tilapia, resulting in substantial economic losses in aquaculture. In this study, a 10-round whole-cell Systemic Evolution of Ligands by Exponential Enrichment procedure was conducted to isolate and characterize single-stranded DNA aptamers with specific binding affinity to S. agalactiae. The enriched single-stranded DNA libraries were sequenced by high-throughput sequencing, yielding a total of 180,987 reads, of which 126,933 included unique aptamer sequences. The most frequent sequences were selected for further analysis. Among them, SA1 had the highest frequency, with 231 reads, whereas SA2 showed a lower number of 16 reads. These findings underscored the validity of combining whole-cell Systemic Evolution of Ligands by Exponential Enrichment, high-throughput sequencing and aptamer-structure analysis to explore the interaction between aptamers and bacterial cell surfaces. The selected aptamers not only showed promise for the detection of S. agalactiae but also served as molecular probes for identifying surface-exposed proteins, studying host-pathogen interactions and guiding targeted drug delivery in aquaculture-related applications.