Serological diagnosis of equine influenza using the hemagglutinin protein produced in a baculovirus expression system.
Abstract: The hemagglutinin (HA) protein of an equine influenza strain, A/equine/La Plata/1/93 (LP/93), was produced using a baculovirus expression system. Silkworm larvae inoculated with recombinant baculovirus expressed high quantities of the HA protein which was then purified to greater than 95% purity by fetuin-affinity chromatography. Purified HA protein was used subsequently in an ELISA for detection of antibodies in horse sera. Two hundred serum samples from vaccinated racehorses were reacted on ELISA plates coated with 40.0 ng/ml of purified HA protein. Subsequent optical density (OD) levels revealed titers which correlated highly with respective hemagglutinin inhibition (HI) antibody titers which ranged from <1:8 to 1:256 (correlation coefficient among them was 0.850). ELISA OD levels and HI titers increased at 5 and 7 days post-inoculation, respectively, in a horse inoculated intranasally with LP/93. Respective antibody levels were observed to change in an essentially parallel manner during a period of 1 month. Similarly, ELISA OD levels correlated with HI titers in horses during a period of 6 weeks following intramuscular inoculation with inactivated single-strain vaccines containing LP/93, A/equine/Kentucky/1/81 (H3N8) or A/equine/Rome/5/91 (H3N8). A similar pattern was also observed in eight horses throughout a 10-week period following inoculation with a commercially available inactivated trivalent vaccine containing A/equine/Newmarket/1/77(H7N7), A/equine/Kentucky/81 and LP/93. From these results, it is suggested that this ELISA system could be used for disease diagnosis and surveillance of HI antibody titers among vaccinated horses.
Publication Date: 2001-09-07 PubMed ID: 11543878DOI: 10.1016/s0166-0934(01)00332-9Google Scholar: Lookup
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- Comparative Study
- Journal Article
Summary
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The research paper explores the use of a baculovirus expression system to produce the hemagglutinin protein of the equine influenza strain, for the purpose of diagnosing the disease in horses. The protein formed was then used in an Enzyme-Linked Immunosorbent Assay (ELISA) to detect antibodies in horses’ blood samples. This method proved effective and correlates with existing hemagglutinin inhibition (HI) antibody diagnosis methods, suggesting potential use in disease diagnosis and surveillance among vaccinated horses.
Production of Hemagglutinin Protein
- The hemagglutinin (HA) protein of the equine influenza strain A/equine/La Plata/1/93 (LP/93) was produced using a baculovirus expression system.
- The baculovirus system was used to infect silkworm larvae, resulting in high quantities of the HA protein.
- The HA protein was then purified to over 95% purity using fetuin-affinity chromatography. This process led to a sufficient quantity of high-quality protein for subsequent tests.
ELISA for Disease Detection
- The purified HA protein was used in ELISA tests to detect antibodies in horse blood serum.
- In a test of 200 serum samples from vaccinated racehorses, each sample was reacted on an ELISA plate coated with 40.0 ng/ml of purified HA protein.
- The resulting optical density (OD) levels had a strong correlation with existing hemagglutinin inhibition (HI) antibody titer test results.
Use in Inoculation Trials and Vaccine Monitoring
- The ELISA system was used to track responses to vaccination and inoculation with equine influenza strains.
- These responses were tracked over a period of one month or up to ten weeks, depending on the strain and method of inoculation used.
- These tests revealed a parallel trend between ELISA OD levels and HI titer levels, reinforcing the reliability of this diagnosis method.
Potential Use in Disease Diagnosis and Surveillance
- The results suggest that the ELISA system using the HA protein produced via a baculovirus expression system could be used for disease diagnosis in horses.
- Given the high correlation to existing diagnosis methods, the system could also be used in monitoring vaccinated horses for HI antibody titers which help assess the efficacy of vaccination and monitor potential outbreaks of equine influenza.
Cite This Article
APA
Sugiura T, Sugita S, Imagawa H, Kanaya T, Ishiyama S, Saeki N, Uchiyama A, Tanigawa M, Kuwano A.
(2001).
Serological diagnosis of equine influenza using the hemagglutinin protein produced in a baculovirus expression system.
J Virol Methods, 98(1), 1-8.
https://doi.org/10.1016/s0166-0934(01)00332-9 Publication
Researcher Affiliations
- Epizootic Research Station, Equine Research Institute, The Japan Racing Association, 1400-4 Shiba, Kokubunji-chou, Shimotuga-gun, 329-0412, Tochigi, Japan. sugiurat@epizoo.equinst.go.jp
MeSH Terms
- Animals
- Antibodies, Viral / immunology
- Baculoviridae / genetics
- Bombyx / virology
- Enzyme-Linked Immunosorbent Assay
- Genetic Vectors
- Hemagglutination Inhibition Tests
- Hemagglutinins, Viral / genetics
- Hemagglutinins, Viral / immunology
- Horse Diseases / diagnosis
- Horse Diseases / immunology
- Horses
- Influenza A virus / immunology
- Orthomyxoviridae Infections / veterinary
- Recombinant Proteins / immunology
- Serology / methods
- Time Factors
Citations
This article has been cited 2 times.- Kato T, Kajikawa M, Maenaka K, Park EY. Silkworm expression system as a platform technology in life science. Appl Microbiol Biotechnol 2010 Jan;85(3):459-70.
- Nwe N, He Q, Damrongwatanapokin S, Du Q, Manopo I, Limlamthong Y, Fenner BJ, Spencer L, Kwang J. Expression of hemagglutinin protein from the avian influenza virus H5N1 in a baculovirus/insect cell system significantly enhanced by suspension culture. BMC Microbiol 2006 Feb 24;6:16.
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