Lactic acid bacteria (LAB) of the genus Lactiplantibacillus have been explored as potential mucosal vaccine vectors due to their ability to elicit an immune response against expressed foreign antigens and to their safety. However, tools for monitoring LAB distribution and persistence at the mucosal surfaces are needed. Here, we characterize Lactiplantibacillus plantarum bacteria expressing the infrared fluorescent protein IRFP713 for exploring their in vivo distribution in the mucosa and potential use as a mucosal vaccine vector. This bacterial species is commonly used as a vaginal probiotic and was recently found to have a niche in the human nose. Three different fluorescent L. plantarum strains were obtained using the nisin-inducible pNZRK-IRFP713 plasmid which contains the nisRK genes, showing stable and constitutive expression of IRFP713 in vitro. One of these strains was further monitored in BALB/c mice using near-infrared fluorescence, indicating successful colonization of the nasal and vaginal mucosae for up to 72 h. This study thus provides a tool for the in vivo spatiotemporal monitoring of lactiplantibacilli, allowing non-invasive bacterial detection in these mucosal sites. Key points • Stable and constitutive expression of the IRFP713 protein was obtained in different L. plantarum strains. • IRFP713+ L. plantarum 3.12.1 was monitored in vivo using near-infrared fluorescence. • Residence times observed after intranasal and vaginal inoculation were 24–72 h. Graphical abstract