Analysis of Antimicrobial Activity of Hemolymph Derived from Salmonella Challenged Black Solider Fly Larva

Abstract

Black soldier fly larvae (BSFL) have gained increasing attention as a sustainable feed ingredient in animal agriculture, not only for their nutritional value but also for their potential functional benefits. BSFL is known to grow well on different kinds of organic waste that contains pathogens. For example, there are previous studies in the literature that document BSFL growth on animal manure, poultry slaughterhouse wastes, etc. These organic wastes are known to contain different types of food-borne pathogens such as Escherichia coli, Salmonella sp., Campylobacter sp. etc. Many of the previous studies in the literature documented only the bioconversion potential of the BSFL grown on contaminated waste but failed to understand the fate of pathogens or induction of immune response by the BSFL exposed to pathogens. This study evaluated BSFL growth performance, Salmonella dynamics, and antimicrobial activity of larval hemolymph under controlled pathogen exposure. Bioconversion trials showed that larvae efficiently converted food waste in both control and pathogen-inoculated treatments, with bioconversion efficiency of 13.57 - 14.84%, substrate reduction of 65.32 - 74.93%, biomass gain of 135.72 - 148.48 g, and identical feed conversion ratios (4.85). These results indicate that the presence of Salmonella did not have effect on feed utilization. Microbial analysis showed that Salmonella persisted and multiplied in the substrate and frass of inoculated treatments though it was undetectable in harvested larvae and hemolymph after processing. Hemolymph inhibition assays demonstrated strong antimicrobial activity particularly the physically stressed (pricked) larvae showed the highest suppression at early stage. Hemolymph from the larvae immunized through pathogen infection exhibit complete inhibition of Salmonella at higher concentration. Survivability tests showed that survival of intensely infectious pricked larvae was more than 93% within 24 hours. Proteomic analysis identified several antimicrobial peptides including defensins, attacins, and cecropin-like peptides in the hemolymph of pricked and challenged larvae. The presence of these peptides supports the strong antibacterial activity observed in the treated groups. These results suggest that immune stimulation improves the antimicrobial properties of BSFL hemolymph without affecting their ability to efficiently convert waste. Overall, the study aims that immune activated BSFL could be a safer and more beneficial insect-based feed ingredient for poultry production.