Gill Mucus and Gill Mucin O-glycosylation in Healthy and Amebic Gill Disease-Affected Atlantic Salmon.

John Benktander,George Birchenough,János T Padra,Sara K Lindén,James W Wynne,Russel Mcculloch,Ben Maynard,Kristina Sundell,Sinan Sharba,Henrik Sundh,Natasha A Botwright

doi:10.3390/microorganisms8121871

Abstract

Amoebic gill disease (AGD) causes poor performance and death in salmonids. Mucins are mainly comprised by carbohydrates and are main components of the mucus covering the gill. Since glycans regulate pathogen binding and growth, glycosylation changes may affect susceptibility to primary and secondary infections. We investigated gill mucin O-glycosylation from Atlantic salmon with and without AGD using liquid chromatography–mass spectrometry. Gill mucin glycans were larger and more complex, diverse and fucosylated than skin mucins. Confocal microscopy revealed that fucosylated mucus coated sialylated mucus strands in ex vivo gill mucus. Terminal HexNAcs were more abundant among O-glycans from AGD-affected Atlantic salmon, whereas core 1 structures and structures with acidic moieties such as N-acetylneuraminic acid (NeuAc) and sulfate groups were less abundant compared to non-infected fish. The fucosylated and NeuAc-containing O-glycans were inversely proportional, with infected fish on the lower scale of NeuAc abundance and high on fucosylated structures. The fucosylated epitopes were of three types: Fuc-HexNAc-R, Gal-[Fuc-]HexNAc-R and HexNAc-[Fuc-]HexNAc-R. These blood group-like structures could be an avenue to diversify the glycan repertoire to limit infection in the exposed gills. Furthermore, care must be taken when using skin mucus as proxy for gill mucus, as gill mucins are distinctly different from skin mucins.

Highlights

All mucosal epithelia are covered by a continuously secreted mucus layer and mucins are the main components of the mucus [1,2]
Mucins are comprised of 50–90% carbohydrate in the form of O-glycans, and substantial differences in glycosylation have been demonstrated in epithelia within and between species [3,4]
We have previously demonstrated that these two methods of mucin isolation yield similar O-glycan profiles using liquid-chromatography–mass spectrometry (LC-MS), with differences between O-glycan profiles being of a similar magnitude to those observed among technical replicates within each extraction method [3]

Summary

Introduction

All mucosal epithelia are covered by a continuously secreted mucus layer and mucins are the main components of the mucus [1,2]. Mucins are comprised of 50–90% carbohydrate in the form of O-glycans, and substantial differences in glycosylation have been demonstrated in epithelia within and between species [3,4]. The inter-individual variation in glycosylation between Atlantic salmon (ATS, Salmo salar) is very low compared to that of pigs and humans [3,4,5,6,7]. Since the large inter-individual variation in humans is considered a population-based defense against infection, the low variation in ATS glycosylation may indicate that they are vulnerable to infection at a population level [3,4]. Since glycans regulate pathogen binding and growth [8,9], glycosylation changes may affect susceptibility to both primary and secondary infections

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Methods

Results