Probiotic Modulation of Gut Microbiota Enhances Immunity and Nutrition in SIT Ceratitis Capitata Sterile Males.

The gut microbiome significantly influences insect physiology, immunity, and ecological fitness. However, mass-rearing and sterilization processes in Sterile Insect Technique (SIT) programs disrupt this natural balance. The resulting dysbiosis is a critical factor reducing the performance of sterile males. This study demonstrates that strategically administered probiotic supplementation can restore gut homeostasis and enhance the nutritional status and immunity of Ceratitis capitata sterile males. Our selected three-strain probiotic consortium L.E.K-PC (Lactococcus lactis, Klebsiella oxytoca, and Enterobacter sp.) was administered during, larval (L+), adult (A+) and both life stages (AL⁺), followed by a challenge with the entomopathogenic bacteria Pseudomonas sp. Full-length 16 S rRNA Nanopore sequencing revealed that AL⁺ males maintained a diverse and balanced gut microbiota with elevated levels of beneficial bacteria, whereas control flies exhibited marked dysbiosis. Probiotic supplementation significantly increased protein, carbohydrate, and lipid reserves in both life stages, upregulated the expression of antimicrobial peptide genes (Cecropin, Attacin and PGRP), enhanced phenoloxidases activity, and strengthened the antibactericidal capacity of hemolymph. Consequently, AL⁺ males showed the highest survival rate following infection. These results indicate that probiotic supplementation acts as “immunobiotic” coordinating nutritional support and immune activation through gut microbiota modulation. Incorporating such probiotic strategies into SIT programs could enhance sterile male resilience and pathogen resistance, offering a sustainable approachto optimizing mass-rearing and strengthening ecological pest management.