Lactococcus lactis NZ3900/pNZ8149-ChIL-4-ChIL-2 alleviates compound stress in chicks via reshaping immune homeostasis, gut microbiota and intestinal health

The poultry industry faces significant challenges due to multiple concurrent stressors, such as heat stress (HS) and live Eimeria vaccination. This study aimed to evaluate the efficacy of Lactococcus lactis NZ3900/pNZ8149-ChIL-4-ChIL-2 (RLIL4/2) as a trained immunity inducer in mitigating the adverse effects of combined stressors in young chicks, with a focus on systemic immune balance, growth performance, and intestinal health. Four-day-old ISA White pullets were randomly allocated into 4 groups with 3 biological replicates (20 birds/replicate): the C group (mild HS, 72≤THI≤79, 12h/day, basic crude protein diet [CP ≥23%], no Eimeria vaccine, no RLIL4/2), the A group (moderate HS [MHS], low crude protein diet [CP ≤17%], Eimeria vaccine), the R group (MHS, CP 17% diet with oral RLIL4/2), and the T group (same as group A plus oral RLIL4/2 1 h pre-vaccination). Compared with the control group, compound stress suppressed chick growth and immune‑organ development, triggering immune response imbalance, gut microbiota dysbiosis, intestinal barrier injury, and a marked expansion of pathogenic bacteria. RLIL4/2 could mitigate these pathological alterations and preserve host health, immune function, and microbiota homeostasis. Moreover, RLIL4/2 is capable of inducing trained immunity in macrophages, thereby enabling a more rapid immune response upon challenge. These results showed that RLIL4/2 relieves compound stresse-induced gosling enteritis by maintaining cytokine, tight junctions, and immunoglobulin relatively stable and improving the disorder of intestinal flora. RLIL4/2, an compound regulator, boosts disease preventive efficacy in chicks subjected to compound stress through multiple molecular pathways, and its application provides a novel intervention approach for avian disease control.