Pangenome-based design of strain-specific primers allows the specific monitoring of engraftment in different habitats

Abstract Lactic acid bacteria (LAB), including many well-known beneficial bacteria, have seen a rise in the number of applications with specific strains across various areas, including live biotherapeutic products (LBPs). The most extensively researched strains belong to the Lactobacillaceae. Assessing the survival and persistence of specific strains in different niches is still an important challenge, while selective monitoring techniques are often lacking at strain level. Here, we show a robust pangenome-based approach for detecting singletons, which can be used to develop strain-specific primers. We developed selective and specific primers for six strains across different LAB species. The primers for the widely-used probiotic L. rhamnosus GG and L. plantarum WCFS1 were validated in in vivo studies and showed that these strains can persist in and on other habitats such as the human skin, upper respiratory tract and fermented vegetables. In conclusion, the selection of unique genes derived from the pangenome of a species resulted in a specific and sensitive method based on qPCR to detect and monitor strains in different habitats. This approach can be readily extended to other bacterial strains on other families for any type applications in research and industry.