Loop-mediated isothermal amplification (LAMP) assays for rapid on-site detection of Lacticaseibacillus casei, Lacticaseibacillus paracasei, and Lacticaseibacillus rhamnosus in probiotic products

We developed the colorimetric loop-mediated isothermal amplification (LAMP) and a real-time LAMP with direct DNA extraction for the on-site detection of the three closely related Lacticaseibacillus species, such as Lacticaseibacillus casei, Lacticaseibacillus paracasei, and Lacticaseibacillus rhamnosus in probiotic products. Species-specific LAMP primers were designed targeting a unique gene in each species through comparative genomic analysis. Each LAMP primer set amplified target species specifically, distinguishing between the closely related species within the Lacticaseibacillus species. Based on the designed LAMP primers, two LAMP-based assays with direct DNA extraction were compared; their limit of detection was 10³ CFU/mL for the Lacticaseibacillus species. In addition, both methods were applied to 44 commercial probiotic products and obtained identical results; it took about 45 min from sample preparation to confirming the presence or absence of the Lacticaseibacillus species. Thus, these methods can be rapid and simple in-field tests to detect L. casei, L. paracasei, and L. rhamnosus in probiotic products.