Aeromonas salmonicida aptamer selection and construction for colorimetric and ratiometric fluorescence dual-model aptasensor combined with g-C3N4 and G-quadruplex

Aeromonas salmonicida (A. salmonicida) is an important opportunistic pathogen to aquatic animals that causes severe economic losses to aquaculture, which makes its rapid detection and prevention are critical. In this work, a single-stranded DNA (ssDNA) aptamer (A.s-2) with high specificity to the bacteria was selected by Systematic Evolution of Ligands by Exponential Enrichment (SELEX). The selected aptamer was confirmed with high binding ability and specificity (Kd = 32 ± 8 nM). Furthermore, a novel dual-model colorimetric and ratiometric fluorescent aptasensor was constructed based on the G-quadruplex-modified aptamer and g-C3N4 for sensitive, reliable, and visual detection of the diseased bacteria in fishes. The quantitative detection was achieved in the linear range of 103-107 CFU mL-1 with a detection limit of 1.9 × 102 CFU mL-1. Meanwhile, the semi-quantitative detection can also be performed visually through fluorescence or color changes of the solution, which is suitable for the early diagnosis of pathogen infection in grassroots farms. Moreover, the developed aptasensor was successfully applied to detect A. salmonicida infection in zebrafish samples with satisfactory results. This work provides a framework for the rapid detection of pathogens in aquaculture, indicating its great prospects in food safety.