Intestinal mucin offers a physical barrier to maintain host–commensal homeostasis. Glycosylation is essential for the appropriate functioning of mucin. Galacto-oligosaccharides (GOS) have been used as a prebiotic with proven intestinal benefits, while their regulatory mechanism on mucin remains unclear. This study employed an antibiotic-treated rat model to mimic gut dysbiosis and attempted to restore gut dysbiosis using GOS. The gut microbiome and intestinal mucus O-glycosylations (O-glycans) in the small intestine were profiled by high-throughput sequencing and glycomics. The sialic acid phenotype at the end of O-glycans was further validated with lectin staining. Expressions of key enzymes in sialic acid metabolism and epithelial morphology were determined as well. Antibiotics significantly increased the relative abundance of Escherichia/Shigella and decreased the relative abundance of Lactobacillus. This was accompanied by decreased microbial sialidase activity and increased sialic acid in the digesta, as well as an increase in epithelial sialidase activity. Analysis of key sialylation enzymes showed the upregulation of α 2,6 sialylation (e.g. ST6GALNACs) and downregulation of α 2,3 sialylation (e.g. ST3GALs) after antibiotic treatment. The glycomics results revealed that antibiotics increased core 4 and α 2,6 sialylated O-glycans and decreased core 1, core 3 and α 2,3 sialylated O-glycans in the intestinal mucus of rats, which was further confirmed by lectin staining. Intestinal histology results demonstrated that antibiotic treatment led to the dysbiosis of intestinal mucus homeostasis. To further test the role of microbiota in regulating intestinal mucus sialylation, we supplemented GOS with antibiotics. The results showed that GOS reversed the effects of antibiotics on the gut microbiota and intestinal mucus O-glycans (especially sialylated O-glycans), characterized by an increase of Lactobacillus and α 2,3 sialylated O-glycans and a decrease of Escherichia/Shigella and α 2,6 sialylated O-glycans. What's more, GOS reduced the stimulation of the intestinal mucosa by lipopolysaccharide (LPS) by increasing α 2,3 sialylated intestinal alkaline phosphatase (IAP) to enhance IAP activity, thereby restoring intestinal mucus homeostasis. Overall, GOS counteracts antibiotic-induced mucin deficiency by remedying the gut ecology and changing the mucin sialylation pattern, as reflected by the increase of α 2,3 sialylated O-glycans and the decrease of α 2,6 sialylated O-glycans.

Background: Studies have shown that some viral infections cause structural changes in the intestinal microflora, but little is known about the effects of tumorigenic viral infection on the intestinal microflora of chickens. Results: A 29-week commercial layer flock positive for avian leukosis virus-J (ALV-J), Marek’s disease virus (MDV) and avian reticuloendotheliosis virus (REV) was selected, and fresh fecal samples were collected and examined for the composition of the gut microflora by Illumina sequencing of the V3-V4 region of the 16S rRNA gene. The operational taxonomic units (OTUs) of the fecal microbiota differentiated the chickens infected with only ALV-J and those coinfected with ALV-J and MDV or REV from infection-negative chickens. The enrichment and diversity of cloacal microflora in chickens infected with ALV-J alone were slightly different from those in the infection-negative chickens. However, the diversity of cloacal microflora was significantly increased in chickens coinfected with both ALV-J and MDV or REV. Conclusions: The intestinal microbiota was more strongly disturbed in chickens after coinfection with ALV-J and MDV or REV than after infection with ALV-J alone, and there may be underlying mechanisms by which the capacity for the stabilization of the intestinal flora was impaired due to viral infection and tumorigenesis.

The mucin barrier is essential for postnatal gut homeostasis, but whether galacto-oligosaccharides (GOS) regulate mucin glycosylation remains unclear. Here, we investigated the effects of early-life GOS supplementation on mucin O-glycans and intestinal barrier development in piglets using UPLC, MALDI-TOF-MS, immunofluorescence, lectin staining, and histomorphological analyses. GOS supplementation significantly increased jejunal villus width, mucosal thickness, goblet cell numbers, and MUC2 expression during early life. Glycomic profiling revealed selective remodeling of mucin O-glycans, characterized by increased core 1/3 and α2,3-sialylated glycans and reduced core 2/4 and α2,6-sialylated structures. These alterations were accompanied by upregulation of C1GALT1, GALNTs, and ST3GALs, and downregulation of ST6GALNACs and neuraminidases. Lectin staining further confirmed enhanced α2,3-sialylation and reduced α2,6-sialylation. Collectively, these results demonstrate that dietary GOS promotes intestinal barrier development and selectively modulates the composition and sialylation of mucin O-glycans, providing novel mechanistic insight into how prebiotics support gut health in early life.