Effects of a Combined Antimicrobial Peptide and Candida utilis Supplementation on Rumen Development and Microecology of Weaned Hu Lambs

Rumen development and microecological homeostasis are critical for post-weaning health in lambs; however, effective nutritional strategies combining probiotics and antimicrobial peptides remain underexplored. This study investigated the effects of graded doses of a combined additive (Candida utilis + antimicrobial peptide (AMP)) on rumen development and microecology in weaned Hu lambs. Forty healthy pre-weaning male lambs (initial body weight: 15.77 ± 1.25 kg; age: 42.98 ± 1.80 d) were randomly assigned to four groups (n = 10): a control group (CK, fed milk/basal diet), and three treatment groups receiving the same milk/basal diet supplemented with graded doses of the combined additive, based on feed intake (LCB: AMP 0.05 g/kg + C. utilis 0.1 g/kg, MCB: AMP 0.05 g/kg + C. utilis 0.8 g/kg, HCB: AMP 0.05 g/kg + C. utilis 1.5 g/kg) over a 40-day period starting from day 6. At the end of the trial, six lambs per group were randomly selected for sampling for analyses of rumen morphology, fermentation parameters, 16S rRNA amplicon sequencing, and untargeted metabolomics. Results showed that, compared with CK, (1) MCB and HCB increased papilla height, and HCB also increased papilla width (p < 0.05); papilla width and muscular layer thickness showed significant linear increases with increasing additive dose (p < 0.05). (2) All treatments linearly reduced ruminal pH (p < 0.05), although all values remained within the normal physiological range; meanwhile, all treatment groups showed elevated levels of microbial protein (MCP), butyrate, valerate, and isovalerate; MCP also increased linearly with dose (p < 0.05); LCB decreased the acetate-to-propionate ratio(A:P), and this ratio exhibited a significant quadratic trend across treatment groups (p < 0.05). (3) HCB increased the α-diversity indices and reduced Methanobacteriota abundance; LCB elevated Bacillota; and all treatments decreased Spirochaetota, Synergistota, and Cyanobacteriota but increased Actinomycetota, Verrucomicrobiota, and Thermodesulfobacteriota (p < 0.05). (4) The number of differential metabolites and KEGG pathway enrichments gradually increased with increasing doses of the combined additive, with HCB exerting the most extensive modulatory effects on signaling and neural pathways. (5) Volatile fatty acids, Bacteroidota, and Xylanibacter were positively correlated with rumen development parameters (p < 0.05). Collectively, dietary supplementation with the combined additive improved rumen fermentation, microbial community structure, and metabolic profiles, with the high-dose (HCB) regimen showing the most effects on select parameters, whereas the low-dose (LCB) regimen exhibited advantages in specific fermentation traits. These findings provide preliminary evidence for the potential application of this combined additive in post-weaning lamb diets.