Diet-Driven Modulation of Antibiotic Resistance Genes and Microbial Risk During the Bioconversion of Agro-Industrial Residues by Hermetia illucens

Background: Hermetia illucens larvae provide a sustainable bioconversion pathway that transforms agro-industrial residues into protein- and nutrient-dense biomass and frass, suitable for animal feed and soil amendment, respectively. Nevertheless, the potential spread of antibiotic resistance (AR) genes and pathogenic microorganisms poses biosafety concerns. This study examined the impact of four residue-based diet formulations; peas and chickpea (D1), peas and wheat (D2), onion and wheat (D3), and wheat with digestate (D4), on microbial safety during the bioconversion process. Methods: Enterococcus spp. (viable counts), Salmonella spp. (presence/absence), and 13 AR genes associated with resistance to tetracyclines, macrolide-lincosamide-streptogramin B, β-lactams, vancomycin, and aminoglycosides were quantified in single substrates, diets, larvae, and frass using qPCR. Results: Principal component analysis revealed diet-driven AR gene profiles. D1 lowered the levels of the greatest number of tested AR genes, particularly erm(B), tetracycline, and β-lactam genes in frass, as well as tet(O) and vanB in mature larvae. In contrast, D2 increased the AR gene levels in frass. All diets except D4 eliminated Salmonella spp. Enterococcus spp. loads varied by diet and larval stage, with D2 reducing counts in frass. Conclusions: Diet composition directly shapes microbial dynamics and AR gene dissemination, indicating that legume-based substrates may enhance biosafety in bioconversion systems.