Pangenome-based design of strain-specific primers enables precise monitoring of bacteria in human microbiome intervention trials

Lactic acid bacteria (LAB), including many well-known beneficial bacteria, are increasingly used in diverse applications, including probiotics. Among these, strains of Lactobacillaceae are extensively researched. Monitoring the survival and persistence of specific strains in across niches remains a challenge, as selective techniques at strain level are often lacking. Here, we present a robust pangenome-based approach for detecting unique target genes to develop strain-specific primers. We designed selective and specific primers for six strains across different LAB species. Primers for the widely used probiotic strains Lacticaseibacillus rhamnosus GG and Lactiplantibacillus plantarum WCFS1 were validated using in-house samples from three placebo-controlled human intervention studies. These strains were specifically tracked from body sites such as the skin and the upper respiratory tract, with clear differences between treatment and control samples. This gene-based qPCR method enables sensitive strain detection and can be readily extended to other bacterial strains for diverse research and industrial applications.