A Colorimetric LAMP Assay for Salmonella spp. Detection: Towards a DNA Extraction-Free Approach for Pathogen Screening

As of today, bacteriological identification and the molecular approach PCR are considered the gold standards for <i>Salmonella</i> spp. detection. However, these methods are time-consuming and costly due to the requirements for enrichment and nucleic acid extraction. In this study, we evaluated the reliability of a developed colorimetric loop-mediated isothermal amplification (cLAMP) assay targeting the <i>hilA</i> gene, using Phenol Red as an amplification indicator. Given that Phenol Red is pH-dependent, and to develop an extraction-free test, we evaluated chicken meat pretreatment and thermal treatment. First, we assessed the reliability of this test using a pure culture of <i>Salmonella</i> spp. and then in 50 chicken samples pretreated with optimal NaOH concentrations under standardized conditions. Samples representing extreme pH values were artificially contaminated and subjected to DNA extraction and a heat-treatment protocol. Serial dilutions of these products served as templates for LAMP reactions. The assay sensitivity was estimated to be around 3.9 CFU/µL of pure bacterial culture. In contrast, in biological samples, we detected up to 10 CFU/µL using DNA extraction, while heat treatment successfully amplified the initial solution and even some dilutions up to 10³ CFU/µL. In conclusion, our cLAMP assay demonstrated good sensitivity and provided clear evidence of its potential for in-field use without relying on prior enrichment steps and DNA extraction.