Simple SummaryThe use of probiotic bacteria in the aquaculture of salmonids, which are very sensitive to stress, represents a possibility to increase their resistance to diseases. We have developed a new, low-cost application form ensuring the rapid revitalization of probiotic bacteria. We tested continuous and cyclic feeding regimes with regard to their effect on the intestinal immune response and microbiota of rainbow trout. We found that continuous application stabilizes the intestinal microbiota in favor of beneficial lactic acid bacteria and does not cause unnecessary excessive stimulation of immunity, which can lead to an unwanted increase in energy demand. Cyclic application of probiotic feed has the same positive effect on the intestinal microbiota but provides the opportunity to stimulate the intestinal immune system of trout, for example, in periods of increased stress.AbstractIntensive fish farming is associated with a high level of stress, causing immunosuppression. Immunomodulators of natural origin, such as probiotics or phytoadditives, represent a promising alternative for increasing the immune function of fish. In this study, we tested the autochthonous trout probiotic strainL. plantarumR2 in a newly developed, low-cost application form ensuring the rapid revitalization of bacteria. We tested continuous and cyclic feeding regimes with regard to their effect on the intestinal immune response and microbiota of rainbow trout. We found that during the continuous application of probiotic feed, the immune system adapts to the immunomodulator and there is no substantial stimulation of the intestinal immune response. During the cyclic treatment, after a 3-week break in probiotic feeding and the reintroduction of probiotics, there was a significant stimulation of the gene expression of molecules associated with both cellular and humoral immunity (CD8, TGF-β, IL8, TLR9), without affecting the gene expression for IL1 and TNF-α. We can conclude that, in aquaculture, this probiotic feed can be used with a continuous application, which does not cause excessive immunostimulation, or with a cyclic application, which provides the opportunity to stimulate the immunity of trout, for example, in periods of stress.