Around weaning, piglets are susceptible to infection by bacterial pathogens, leading to increased morbidity and mortality. We identified isolates of Rothia nasisuis in the upper respiratory tract of weaned healthy piglets that produce valinomycin in vitro and in vivo via a giant multimodule non-ribosomal peptide synthase (NRPS) enzyme complex. Valinomycin is an antiviral and antibiotic ionophore that shuttles potassium ions across membranes and is capable of inflammasome activation and apoptosis in LPS-primed macrophages. R. nasisuis inhibited growth of Streptococcus and Rothia species inhabiting the same niche. To investigate the potential for the valinomycin-producing Rothia nasisuis strain to colonize new-born piglets we performed a colonization study in newborn litters of piglets. Briefly, the newborn piglets of 3 sows farrowing on the same day were randomized between the three sows to avoid genetic bias in the experiment. The sows were housed in separate maternity pens and the piglets marked so they could be identified during the study. 9 of the 12 piglets nursed by sows A and B (further referred to as litters A and B) were orally administered approximately 10exp9 CFU of valinomycin-producing R. nasisuis, using a syringe on d2, d19 and d27 after birth. Three 3 piglets in litters A and B were not inoculated with R. nasisuis to see if they became colonized by horizontal transfer. As a control group all the piglets housed with the sow C (further referred to as litter C) were not inoculated with R. nasisuis.