Abstract
Salmonella enterica serovar Gallinarum is a non-motile serovar and is the causative agent of fowl typhoid, and poses a major challenge to poultry production, particularly where rapid diagnostics are lacking. Existing methods are either time-consuming or fail to distinguish motile from non-motile serotypes. We developed a dual-target colorimetric LAMP that detects Salmonella spp. via invA and discriminates S. Gallinarum via TRX (a taxon-restricted sequence), using two separate singleplex reactions. Specificity testing confirmed 100% accuracy, with exclusive amplification of S. Gallinarum through TRX. Analytical sensitivity was comparable to real-time PCR, detecting down to 2.41 CFU/µL (invA) and 1.65 CFU/µL (TRX). Applied to cloacal swabs from experimentally infected chickens (n = 12), the assay consistently outperformed bacteriological culture, detecting up to 25% more positives during early infection when bacterial loads were low or cells were non-culturable. This dual-target LAMP provides a rapid, sensitive, and serovar-discriminating diagnostic tool with strong potential for point-of-care use and real-time surveillance in poultry farms, thereby improving sanitary control of fowl typhoid and reducing associated economic losses.