Fish Physiology and Biochemistry

, Volume 37, Issue 1, pp 135–152

Identification of histones as endogenous antibiotics in fish and quantification in rainbow trout (Oncorhynchus mykiss) skin and gill

  • Edward J. Noga
  • Paul J. Borron
  • Jeffrey Hinshaw
  • William C. Gordon
  • Linda J. Gordon
  • Jung-Kil Seo
Article

DOI: 10.1007/s10695-010-9422-7

Cite this article as:
Noga, E.J., Borron, P.J., Hinshaw, J. et al. Fish Physiol Biochem (2011) 37: 135. doi:10.1007/s10695-010-9422-7

Abstract

Antimicrobial polypeptides (AMPPs) are increasingly recognized as a critical component of innate host defense. Among the AMPPs, polypeptides related to histones have been identified from many animals. Using peptide mapping, we further confirm the identity of two histone-like proteins from fish as members of the H2B (sunshine bass) and H1 (rainbow trout) histone groups. We optimized the conditions for measuring rainbow trout HLP-1/H2B via sandwich ELISA. We used two antibodies, one to the amino terminus and one to the carboxyl terminus, of trout histone H2B, as the capture antibodies, and we used peroxidase-labeled antibody raised to calf histone H2B as the secondary antibody. Specificity of the detecting antibody was confirmed by specific reactivity with histone H2B in tissue extracts via western blotting. The test was reproducible and capable of detecting as little as 5 ng of histone H2B (0.05 μg/ml). Histone H2B levels expressed in gill tissue of juvenile, healthy rainbow trout were well within concentrations that are lethal to important fish pathogens. However, there was a significant, age (size)-dependent decline in histone H2B concentrations as fish matured, until levels became virtually undetectable in market-size fish. In contrast, levels in skin appeared to remain high and unchanged in small versus large fish. Antibacterial activity in skin and gill tissues was closely correlated with histone H2B concentration measured via ELISA, which supports our previous finding that histones are the major AMPPs in rainbow trout skin and gill.

Keywords

Antimicrobial polypeptidesHistonesELISAImmunoassay

Supplementary material

10695_2010_9422_MOESM1_ESM.doc (62 kb)
Supplementary material 1 (DOC 62 kb)
10695_2010_9422_MOESM2_ESM.tif (1.1 mb)
Supplementary Figure 1. MS/MS spectra of the enzymatic digest mixture of sunshine bass HLP-1. (TIFF 1129 kb)

Copyright information

© Springer Science+Business Media B.V. 2010

Authors and Affiliations

  • Edward J. Noga
    • 1
    • 2
  • Paul J. Borron
    • 2
    • 5
  • Jeffrey Hinshaw
    • 3
  • William C. Gordon
    • 4
    • 6
  • Linda J. Gordon
    • 4
    • 6
  • Jung-Kil Seo
    • 1
  1. 1.Department of Clinical Sciences, College of Veterinary MedicineNorth Carolina State UniversityRaleighUSA
  2. 2.Norcarex Bio CorporationRaleighUSA
  3. 3.Department of Biology, College of Agriculture and Life SciencesNorth Carolina State UniversityMills RiverUSA
  4. 4.Bethyl LaboratoriesMontgomeryUSA
  5. 5.RTI InternationalResearch Triangle ParkUSA
  6. 6.Alpha Diagnostics International, Inc.San AntonioUSA