A holistic view of the dynamisms of teleost IgM: a case study of Streptococcus iniae vaccinated rainbow trout (Oncorhynchus mykiss).

To date, little is known about how trout IgM, the primary antibody of fish, varies in titer, specificity, disulfide cross-linking, and affinity following immunization with a pathogen. Work using defined antigens has demonstrated that the disulfide cross-linking structure of IgM becomes increasingly more polymerized during an immune response, coinciding with an increase in affinity, but it is unknown if this has relevance to aquatic pathogens. Understanding how IgM varies following vaccination with an aquatic pathogen is of considerable importance as effector functions allocated to multiple antibody isotypes in mammals are essentially relegated to this single molecule. To gain insights into the dynamism of IgM, rainbow trout were immunized with Streptococcus iniae and individual serum titers, their specificity and affinity to S. iniae, and the disulfide cross-linking pattern of both total-serum and specific Ig were analyzed over a period of 37 weeks. We found that in vaccinated animals titer increased by a factor of ≈100 from starting levels, affinity increased 10-fold, and diversity of S. iniae proteins recognized by trout antibody increased at least 5-fold. Most intriguing, though less cross-linked IgM predominated early in response, by week 5, the fully tetramerized antibody comprised 50% of total specific protein. We propose that this is a mechanism to optimize efficacy of carrying out effector functions and recognizing a wide array of epitopes with higher affinity.

Discovery and characterization of secretory IgD in rainbow trout: secretory IgD is produced through a novel splicing mechanism.

The gene encoding IgH δ has been found in all species of teleosts studied to date. However, catfish (Ictalurus punctatus) is the only species of fish in which a secretory form of IgD has been characterized, and it occurs through the use of a dedicated δ-secretory exon, which is absent from all other species examined. Our studies have revealed that rainbow trout (Oncorhynchus mykiss) use a novel strategy for the generation of secreted IgD. The trout secretory δ transcript is produced via a run-on event in which the splice donor site at the end of the last constant domain exon (D7) is ignored and transcription continues until a stop codon is reached 33 nt downstream of the splice site, resulting in the production of an in-frame, 11-aa secretory tail at the end of the D7 domain. In silico analysis of several published IgD genes suggested that this unique splicing mechanism may also be used in other species of fish, reptiles, and amphibians. Alternative splicing of the secretory δ transcript resulted in two δ-H chains, which incorporated Cµ1 and variable domains. Secreted IgD was found in two heavily glycosylated isoforms, which are assembled as monomeric polypeptides associated with L chains. Secretory δ mRNA and IgD(+) plasma cells were detected in all immune tissues at a lower frequency than secretory IgM. Our data demonstrate that secretory IgD is more prevalent and widespread across taxa than previously thought, and thus illustrate the potential that IgD may have a conserved role in immunity.