Novel chitinolytic Bacillus spp. increase feed efficiency, feed digestibility, and survivability to Vibrio anguillarum in European seabass fed with diets containing Hermetia illucens larvae meal

Insect meals (IM) are being pointed out as a promising sustainable protein source. However, chitin in IM may negatively interfere with fish performance and nutrient availability. Given that carnivorous fish have a low or inexistent ability to digest chitin, novel strategies are needed to overcome chitin-imposed bottlenecks. Carbohydrolytic sporeforming probiotics offer a competitive industrial approach, being able to resist the harsh feed manufacturing process, transport, storage, and animal´s gastrointestinal tract, while being able to increase the digestibility of otherwise indigestible components. Aiming to increase IM chitin digestibility, two chitinolytic sporeforming fish isolates (Fish isolates 645 (FI645) and 658 (FI658)), closely related to Bacillus licheniformis, were isolated from European sea bass (ESB) gastrointestinal tract, and incorporated (individually or as a mixture) in ESB diets with high defatted Hermetia illucens larvae meal (HM) levels (30% inclusion). FI were evaluated regarding their effects on ESB growth performance and digestibility. The spores were able to maintain their viability throughout the 180 days of diet storage at room temperature.Dietary inclusion of FI645 led to higher chitin digestibility. Concomitantly, the digestibility of dry matter, protein, and energy, overall leading to higher feed efficiency and protein efficiency ratio. Dietary inclusion of FI645 led to increased plasma Nacetylglucosamine (GlcNAc) levels. This is the first evidence for diet-mediated modulation of plasma GlcNAc concentrations. Dietary inclusion of FI645 also increased the expressions of the genes coding for N-acetylglucosamine kinase (nagk) and GlcNAc phosphomutase (pgm3), key enzymes from the GlcNAc salvage pathway and the hexosamine biosynthetic pathway (HBP). On the other hand, no differences were found for the expression of N-acetyl-D-glucosamine-6-phosphate deacetylase (amdhd2), a precursor enzyme for GlcNAc redirection to glycolysis. These results hint that free GlcNAc resulting from increased chitin digestibility might be preferentially redirected to protein O-GlcNAcylation processes through HBP in fish cells. Additionally, when challenged with Vibrio anguillarum, and after being feed for one month with the same diets, D645 also increased ESB survival from 52.5% to 77.5%, when compared to the control. Together, these results establish the potential of using FI645 as a probioticfor enhancing chitin digestion and acting as a prophylactic agent for ESB protection against V. anguillarum infection. This strategy opens novel possibilities for using commodities, including high chitin levels in aquafeeds, and reducing the use of antibiotics in aquaculture