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United States Department of Agriculture

Agricultural Research Service

Research Project: Integrated Aquatic Animal Health Strategies

Location: Aquatic Animal Health Research

Title: Chicken-type lysozyme in channel catfish: expression analysis, lysozyme activity, and efficacy as immunostimulant against Aeromonas hydrophila infection

Authors
item Wei Pridgeon, Yuping
item Klesius, Phillip
item Dominowski, Paul -
item Yancey, Robert -
item Kievit, Michele -

Submitted to: Fish and Shellfish Immunology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: July 9, 2013
Publication Date: August 23, 2013
Repository URL: http://handle.nal.usda.gov/10113/58424
Citation: Wei Pridgeon, Y., Klesius, P.H., Dominowski, P.J., Yancey, R.J., Kievit, M.S. 2013. Chicken-type lysozyme in channel catfish: expression analysis, lysozyme activity, and efficacy as immunostimulant against Aeromonas hydrophila infection. Fish and Shellfish Immunology. 35:680-688.

Interpretive Summary: The transcriptional levels of lysozyme-c in skin, gut, liver, spleen, posterior kidney, and blood cells in healthy channel catfish was compared to that in channel catfish infected with Aeromonas hydrophila. Quantitative analysis revealed that the transcription levels of lysozyme c in infected catfish were significantly induced in all tissues tested. Recombinant lysozyme c exhibited significant lytic activity against Gram-positive and Gram-negative bacteria. When recombinant lysozyme c DNA was over-expressed in catfish gill cells G1B, the over-expression of lysozyme c offered significant protection to gill cells against A. hydrophila infection. When channel catfish were intraperitoneally injected with formulated recombinant lysozyme c DNA and challenged with a highly virulent A. hydrophila strain at 1-, 2-, 14-, and 28- days post treatment, the recombinant lysozyme c DNA offered 75%, 100%, 60%, and 77% protection to channel catfish, respectively. Macrophages of fish treated with lysozyme c DNA produced significantly higher amounts of reactive oxygen species and nitric oxide than that of fish treated with pcDNA vector alone. Taken together, our results suggest that recombinant lysozyme c could be used as a novel immunostimulant to protect channel catfish against A. hydrophila infection.

Technical Abstract: To understand whether chicken-type lysozyme (Lys-c) in channel catfish was induced by infection of Aeromonas hydrophila, the transcriptional levels of Lys-c in skin, gut, liver, spleen, posterior kidney, and blood cells in healthy channel catfish was compared to that in channel catfish infected with A. hydrophila by bath immersion. Quantitative PCR revealed that the transcription levels of Lys-c in infected catfish were significantly (P < 0.05) induced in all five tissues tested as well as in blood cells. Recombinant CC-Lys-c produced in Escherichia coli expression system (R-CC-Lys-c) exhibited significant (P < 0.05) lytic activity to Gram-positive Micrococcus lysodeikticus and Gram-negative A. hydrophila. When pcDNA3.2-vectored recombinant channel catfish lysozyme-c (pcDNA-Lys-c) was transfected in channel catfish gill cells G1B, the over-expression of pcDNA-Lys-c offered significant (P < 0.05) protection to G1B against A. hydrophila infection. When channel catfish were intraperitoneally injected with QCDCR adjuvant formulated pcDNA-Lys-c and challenged with a highly virulent A. hydrophila strain AL-09-71 at 1-, 2-, 14-, and 28- days post treatment, pcDNA-Lys-c offered 75%, 100%, 60%, and 77% protection to channel catfish, respectively. Macrophages of fish treated with pcDNA-Lys-c produced significantly (P<0.05) higher amounts of reactive oxygen species and nitric oxide than that of fish treated with pcDNA vector alone. Taken together, our results suggest that pcDNA-Lys-c could be used as a novel immunostimulant to protect channel catfish against A. hydrophila infection.

Last Modified: 8/19/2014