Dietary lactoferrin supplementation enhances immune function, improves intestinal integrity and reshapes gut microbiota in kittens

Lactoferrin is a natural iron-binding glycoprotein with barrier-protective, immunomodulatory, and antimicrobial activities, which is widely applied in human nutritional supplements and infant food additives. The present study aimed to evaluate the effects of dietary lactoferrin supplementation on immune parameters, intestinal barrier function, serum metabolome, and fecal microbiota in kittens. In this study, a total of 24 healthy kittens were divided into three groups and fed a commercial basal diet (CON) and the basal diet supplemented with 500 or 1000 mg/kg lactoferrin (L-LF or H-LF), respectively, for 42 days. Blood samples were collected on day 21 and 42 for measurement of immune and intestinal barrier markers. A repeated-measures mixed-effects model was used for data analysis. Serum and fecal samples were collected on day 42 for untargeted metabolomics and metagenomic analysis, respectively. Compared with CON, serum IgA level in the H-LF group was significantly higher on day 21 and day 42 ( p < 0.05). The H-LF group showed higher serum IgM and lower IL-6 levels than CON on day 21, and both lactoferrin groups reduced IL-1β concentration on day 42 ( p < 0.05), in contrast with the CON. Additionally, serum lipopolysaccharide and zonulin were decreased by H-LF relative to the CON on day 42 ( p < 0.05). Untargeted metabolomics analysis revealed that lactoferrin altered serum metabolite profiles and increased levels of proline betaine, L-argininium(1+), and carnitine in H-LF ( p < 0.05). Metagenomic analysis indicated that both L-LF and H-LF had a higher relative abundance of Bacteroidota but lower Escherichia coli than CON ( p < 0.05). The H-LF group also increased the abundance of Prevotella , Bifidobacterium and Segatella copri in the feces of kittens ( p < 0.05). Furthermore, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways of biosynthesis of secondary metabolites, biosynthesis of various nucleotide sugars, and sphingolipid metabolism were significantly enriched in kittens fed with H-LF ( p < 0.05). Overall, dietary lactoferrin enhanced immune function, improved intestinal barrier integrity, and remodeled the gut microbiota of kittens, with the higher dose (1000 mg/kg) showing more pronounced benefits.