Mucin glycans govern pathogen adhesion, growth and virulence. We analyzed the O-glycome from six Atlantic salmon cohorts grown under various conditions from Sweden, Norway and Australia using LC-MS. The low interindividual variation identified may be a concern because interindividual variation is considered a population based defense against infection. The 169 identified structures represent a library for identifying structures important for host-pathogen interactions, understanding population differences of salmon mucin glycosylation in resistance to diseases and during breeding and selection of strains.

Campylobacter jejuni is a common cause of diarrheal disease worldwide. Human infection typically occurs through the ingestion of contaminated poultry products. We previously demonstrated that an attenuated Escherichia coli live vaccine strain expressing the C. jejuni N-glycan on its surface reduced the Campylobacter load in more than 50% of vaccinated leghorn and broiler birds to undetectable levels (responder birds), whereas the remainder of the animals was still colonized (non-responders). To understand the underlying mechanism, we conducted three vaccination and challenge studies using 135 broiler birds and found a similar responder/non-responder effect. Subsequent genome-wide association studies (GWAS), analyses of bird sex and levels of vaccine-induced IgY responses did not correlate with the responder versus non-responder phenotype. In contrast, antibodies isolated from responder birds displayed a higher Campylobacter-opsonophagocytic activity when compared to antisera from non-responder birds. No differences in the N-glycome of the sera could be detected, although minor changes in IgY glycosylation warrant further investigation. As reported before, the composition of the microbiota, particularly levels of OTU classified as Clostridium spp., Ruminococcaceae and Lachnospiraceae are associated with the response. Transplantation of the cecal microbiota of responder birds into new birds in combination with vaccination resulted in further increases in vaccine-induced antigen-specific IgY responses when compared to birds that did not receive microbiota transplants. Our work suggests that the IgY effector function and microbiota contribute to the efficacy of the E. coli live vaccine, information that could form the basis for the development of improved vaccines targeted at the elimination of C. jejuni from poultry.

Disease outbreaks are limiting factors for an ethical and economically sustainable aquaculture industry. The first point of contact between a pathogen and a host occurs in the mucus, which covers the epithelial surfaces of the skin, gills and gastrointestinal tract. Increased knowledge on host-pathogen interactions at these primary barriers may contribute to development of disease prevention strategies. The mucus layer is built of highly glycosylated mucins, and mucin glycosylation differs between these epithelial sites. We have previously shown that A . salmonicida binds to Atlantic salmon mucins. Here we demonstrate binding of four additional bacteria, A . hydrophila , V . harveyi , M . viscosa and Y . ruckeri , to mucins from Atlantic salmon and Arctic char. No specific binding could be observed for V . salmonicida to any of the mucin groups. Mucin binding avidity was highest for A . hydrophila and A . salmonicida , followed by V . harveyi , M . viscosa and Y . ruckeri in decreasing order. Four of the pathogens showed highest binding to either gills or intestinal mucins, whereas none of the pathogens had preference for binding to skin mucins. Fluid velocity enhanced binding of intestinal mucins to A . hydrophila and A . salmonicida at 1.5 and 2 cm/s, whereas a velocity of 2 cm/s for skin mucins increased binding of A . salmonicida and decreased binding of A . hydrophila . Binding avidity, specificity and the effect of fluid velocity on binding thus differ between salmonid pathogens and with mucin origin. The results are in line with a model where the short skin mucin glycans contribute to contact with pathogens whereas pathogen binding to mucins with complex glycans aid the removal of pathogens from internal epithelial surfaces.

The skin barrier consists of mucus, primarily comprising highly glycosylated mucins, and the epithelium. Host mucin glycosylation governs interactions with pathogens and stress is associated with impaired epithelial barrier function. We characterized Atlantic salmon skin barrier function during chronic stress (high density) and mucin O-glycosylation changes in response to acute and chronic stress. Fish held at low (LD: 14–30 kg/m3) and high densities (HD: 50-80 kg/m3) were subjected to acute stress 24 h before sampling at 17 and 21 weeks after start of the experiment. Blood parameters indicated primary and secondary stress responses at both sampling points. At the second sampling, skin barrier function towards molecules was reduced in the HD compared to the LD group (Papp mannitol; p < 0.01). Liquid chromatography–mass spectrometry revealed 81 O-glycan structures from the skin. Fish subjected to both chronic and acute stress had an increased proportion of large O-glycan structures. Overall, four of the O-glycan changes have potential as indicators of stress, especially for the combined chronic and acute stress. Stress thus impairs skin barrier function and induces glycosylation changes, which have potential to both affect interactions with pathogens and serve as stress indicators.

Glycosylation is the most common post-translational modification of proteins, and glycosylation changes at cell surfaces are frequently associated with malignant epithelia including head and neck squamous cell carcinoma (HNSCC). In HNSCC, 5-year survival remains poor, averaging around 50% globally: this is partly related to late diagnosis. Specific protein glycosylation signatures on malignant keratinocytes have promise as diagnostic and prognostic biomarkers and as therapeutic targets. Nevertheless, HNSCC-specific glycome is to date largely unknown. Herein, we tested six established HNSCC cell lines to capture the qualitative and semi-quantitative N-glycome using porous graphitized carbon liquid chromatography coupled to electrospray ionisation tandem mass spectrometry. Oligomannose-type N-glycans were the predominant features in all HNSCC cell lines analysed (57.5–70%). The levels of sialylated N-glycans showed considerable cell line-dependent differences ranging from 24 to 35%. Importantly, α2-6 linked sialylated N-glycans were dominant across most HNSCC cell lines except in SCC-9 cells where similar levels of α2-6 and α2-3 sialylated N-glycans were observed. Furthermore, we found that HPV-positive cell lines contained higher levels of phosphorylated oligomannose N-glycans, which hint towards an upregulation of lysosomal pathways. Almost all fucose-type N-glycans carried core-fucose residues with just minor levels (< 4%) of Lewis-type fucosylation identified. We also observed paucimannose-type N-glycans (2–5.5%), though in low levels. Finally, we identified oligomannose N-glycans carrying core-fucose residues and confirmed their structure by tandem mass spectrometry. This first systematic mapping of the N-glycome revealed diverse and specific glycosylation features in HNSCC, paving the way for further studies aimed at assessing their possible diagnostic relevance.