Carbohydrate-binding IgM: an alternative for lectins in the serum of Vietnamese striped catfish?

Background and objectives:
Vietnamese striped catfish (Pangasianodon hypophthalmus) is intensively farmed in the Mekong delta. Our ongoing project aims at better understanding the defence mechanisms of this teleost against stress, pollution and attack by pathogenic microorganisms.
Methods:
Serum of P.hypophthalmus was investigated for the presence of lectins and other sugar-binding molecules. The serum was fractionated by affinity chromatography using twelve different carbohydrate-Sepharose columns and proteins were eluted with the corresponding sugars.
Results:
Proteins were found to bind to each of the affinity columns in a Ca-dependent way, and were identified to be IgM. Based on the observation that, besides being adsorbed to carbohydrates, these IgM also bind to several glycosylated and non-glycosylated proteins and to E.coli DNA, they are classified as natural antibodies (Nabs). All P.hypophthalmus IgMs have an H-chain of 72 kDa and an L-chain that displays heterogeneity, with three bands of 24, 26 and 28-29 kDa occurring in different combinations. Gel filtration and non-denaturing non-reducing PAGE shows that all IgMs form tetramers (H2L2)4, with no trace of dimers, monomers or halfmers. In non-reducing SDS-PAGE both monomers and halfmers are obtained in about equimolar amounts, indicating that some H-chains are connected by disulfide bonding and some are not. Microheterogeneity in the bands obtained is indicative for differences in the overall shape of the molecules. Cross-reactivity using polyclonal rabbit anti-Gal-IgM confirms this finding. All carbohydrate-binding IgM strongly agglutinate the pathogenic bacteria Aeromonas hydrophila and Edwardsiella ictaluri.
Conclusion:
Besides lectins (e.g. the short pentraxins SAP and CRP), P.hypophthalmus also developed a battery of carbohydrate-binding natural IgMs in their serum that may be involved in protecting the fish against various pathogens.