Identification and characterization of differentially expressed channel catfish IgM transcripts after vaccination with antigens of virulent aeromonas hydrophila

Authors: Zhang, Dunhua; Lange, Miles; Shoemaker, Craig; Beck, Benjamin

Submitted to: Fishes
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/11/2022
Publication Date: 1/19/2022
Citation: Zhang, D., Lange, M.D., Shoemaker, C.A., Beck, B.H. 2022. Identification and characterization of differentially expressed channel catfish IgM transcripts after vaccination with antigens of virulent aeromonas hydrophila. Fishes. 2022(7):24. https://doi.org/10.3390/fishes7010024.
DOI: https://doi.org/10.3390/fishes7010024

Interpretive Summary: Previously, we observed that serum from channel catfish vaccinated with extracellular proteins (ECP) prepared from virulent Aeromonas hydrophila (vAh) recognized specific vAh proteins and aggregated vAh cells, resulting in immune protection of fish against the establishment of pathogenesis. IgM proteins that elicited agglutination of both ECP and cells of vAh were isolated from the antiserum, revealing that there were versatile IgM proteins targeting numerous antigens (including virulence associated proteins). The aim of this study was to analyze IgM transcripts (cDNA) that were differentially expressed in tissues (kidney and liver) of immunized- and mock-immunized catfish with a particular emphasis on the variable domains of IgM heavy chain. Our results indicated that fish produced specific IgM transcripts in response to vAh immunogens. Comparative analysis of these antigen-driven IgM genes would facilitate future research for decoding the mechanism of tailoring antigen receptor specificity.

Technical Abstract: Channel catfish (Ictalurus punctatus) is the top species produced in US aquaculture and motile Aeromonas septicemia, caused by virulent Aeromonas hydrophila (vAh), is one of the most severe diseases that afflict catfish farms in recent years. Previously, vaccination of fish with extracellular proteins (ECP) of vAh was shown to produce a robust antibody-mediated immune response against vAh infection. In this study, we analyzed IgM transcripts that were differentially expressed in the kidney and liver of ECP-immunized and mock-immunized (control) fish with emphasis on variable domain of heavy chain. Quantitative PCR analysis indicated immunized fish produced significantly more IgM transcripts than control fish. Full-length IgM heavy chain cDNA was cloned, which encoded typical IgM peptide, including signal peptide, variable domain (VH), constant domain (CH), and carboxyl terminal peptide. Great sequence diversity was revealed in VH segment, with the third complementarity diversity region (CDR3) being most variable. Using germline VH gene grouping method, variants (clones) of VH characterized in this study belonged to nine VH families. Most unique variants (approximately 49%) were found in VH2 family. Vaccinated fish apparently had more unique variants than in control fish. There were 62% and 79% unique variants in kidney and liver of vaccinated fish, respectively, while 44% and 27% unique variants in kidney and liver of control fish, respectively. Among the unique variants in VH2 family, approximately 87% of them were found in vaccinated fish. Two-dimensional gel electrophoresis of semi-purified IgM protein confirmed that matured IgM protein was as variable as IgM transcripts identified in this study, with isoelectric points crossing from 6 to 10. Results of this study provided insight into the molecular and genetic basis of antibody diversity and enriched our knowledge of the complex interplay between antigens and antibodies in Ictalurid catfish.