Bacillus subtilis spores influence the capacity of rainbow trout leukocytes to respond to a posterior viral encounter

Although the positive effects of probiotics on the general immune status of aquacultured fish has been widely demonstrated using different probiotic strains and fish species, whether probiotics specifically stimulate antiviral responses has only been scarcely addressed. To provide more insights into this matter, we have investigated the capacity of Bacillus subtilis to modulate the transcription of a range on genes related to antiviral responses in rainbow trout (Oncorhynchus mykiss) through two independent experiments. In the first one, fish were fed a B. subtilis-supplemented diet or a control diet and after 15 or 45 days the levels of transcription of these genes evaluated in kidney and spleen. In the second experiment, fish were fed either of the diets and after 30 days, sacrificed and kidney and spleen leukocytes isolated. These leukocytes were then exposed in vitro to either Poly I:C (an analogue of double stranded RNA) or inactivated viral hemorrhagic septicemia virus (VSHV) (or left untreated). After 24 h of incubation, RNA was extracted from these cultures and the levels of transcription of the antiviral genes analyzed. Our results demonstrate that although the supplemented diet had a faint direct effect on the levels of transcription of these genes in spleen and kidney, it significantly increased the transcriptional response of leukocytes to posterior viral stimuli, pointing to the second method as a more suitable approach to screen for antiviral effects and providing novel insights on the capacity of this probiotic strain to up-regulate antiviral responses.