Hyperinvasive locus A gene-based electrochemical nanobiosensor for rapiddetection of Salmonella enterica in chicken eggshell matrices

Salmonella enterica can penetrate inside eggshells through horizontal transmission; thus, a rapid detection method on this matrix is needed. This study aimed to develop an electrochemical nanobiosensor prototype based on the hyperinvasive locus A (hilA) gene of Salmonella enterica for rapid (< 8 hrs) detection of the foodborne pathogen in this matrix. The biosensor was characterized in terms of sensitivity, selectivity and linear detection range as well as its capability to rapidly detect artificially spiked S. enterica in chicken eggshells. The presence of the target hilA gene was also validated through PCR. The hilA genosensor had a linear detection range of 0.30 – 30 ng/µL or 6.84 – 684.5 nM with R2 = 0.8647 at p<0.01 as well as having sensitivity as low as 0.3 ng/µL or 6.84 nM. The nanobiosensor is specific to S. enterica with significantly higher current responses for S. enterica subsp. enterica serotype Enteritidis (6.13±0.69 µA) and S. enterica subsp. enterica serotype Typhimurium (3.59±0.26 µA) as compared with Shigella spp. (1.08±0.23 µA), E. coli O157:H7 (0.88±0.58 µA) and blank (0.67±0.08 µA). Artificially spiked S. enterica serovar Enteritidis (8×104 CFU/mL) in chicken eggshell samples were successfully detected, without DNA amplification, with peak current response of 4.69±1.38 µA against the blank (0.47±0.18 µA). With rapid analysis time (1.5 hrs target hybridization and detection excluding sample processing and 2.5 hrs offline screen-printed carbon electrode preparation), preliminary results show that the developed nanobiosensor could be used to monitor eggs for horizontal Salmonella contamination and improve food safety monitoring in layer farms.