Controlling equine influenza: traditional to next generation serological assays (original) (raw)
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Vaccines
Equine influenza is a major respiratory disease of horses that is largely controlled by vaccination in some equine populations. Virus-neutralising antibodies, the mainstay of the protective immune response, are problematic in assaying for equine influenza virus, as most strains do not replicate efficiently in cell culture. Surrogate measures of protective antibody responses include the haemagglutination inhibition (HI) test and single radial haemolysis (SRH) assay. For this study, a pseudotyped virus, bearing an envelope containing the haemagglutinin (HA) from the Florida clade 2 equine influenza virus strain A/equine/Richmond/1/07 (H3N8), was generated to measure HA-specific neutralising antibodies in serum samples (n = 134) from vaccinated or experimentally-infected ponies using a pseudotyped virus neutralization test (PVNT). Overall, the results of PVNT were in good agreement with results from the SRH assay (100% sensitivity, 68.53% specificity) and HI test (99.2% sensitivity, 49...
The use of equine influenza pseudotypes for serological screening
Standard assays used for influenza serology present certain practical issues, such as inter-laboratory variability, complex protocols and the necessity for handling certain virus strains in high biological containment facilities. In an attempt to address this, avian and human influenza HA pseudotyped retroviruses have been successfully employed in antibody neutralization assays. In this study we generated an equine influenza pseudotyped lentivirus for serological screening. This was achieved by co-transfection of HEK293T cells with plasmids expressing the haemagglutinin (HA) protein of an H3N8 subtype equine influenza virus strain, HIV gag-pol and firefly luciferase reporter genes and harvesting virus from supernatant. In order to produce infective pseudotype particles it was necessary to additionally co-transfect a plasmid encoding the TMPRSS2 endoprotease to cleave the HA. High titre pseudotype virus (PV) was then used in PV antibody neutralization assays (PVNAs) to successfully distinguish between vaccinated and non-vaccinated equines. The sera were also screened by single radial haemolysis (SRH) assay. There was a 65% correlation between the results of the two assays, with the PVNA assay appearing slightly more sensitive. Future work will extend the testing of the PVNA with a larger number of serum samples to assess sensitivity/specificity, inter/intra-laboratory variability and to define a protective titre.
Veterinary Microbiology, 2011
New Zealand is free from equine influenza and has never experienced an incursion in its horse population. As part of New Zealand's preparedness to an incursion of an exotic animal disease, it was considered necessary to select the most accurate test for equine influenza (EI) from the array of those available. Four readily available blocking/ competitive enzyme-linked immunosorbent assays (ELISA), originally developed and marketed for the detection of antibodies against the avian influenza virus, were evaluated using serum samples from New Zealand non-infected, non-vaccinated horses (n = 365), and Australian field infected (n = 99) and experimentally infected horses (n = 3). Diagnostic specificities (DSP) and diagnostic sensitivities (DSE) were determined as follows: ELISA-1 = 98.1%/99.0%; ELISA-2 = 90.1%/99.0%; ELISA-3 = 98.1%/96.0%; ELISA-4 = 95.3%/99.0%. For ELISA-1, DSP and DSE results were comparable to previously published data on a larger sample number from Australian horses .
Effect of influenza A/Equine/H3N8 isolate variation on the measurement of equine antibody responses
Canadian journal of veterinary research = Revue canadienne de recherche vétérinaire
This study has tested the effect of using homologous or heterologous equine influenza A virus isolates to evaluate serum antibody levels to influenza A virus in vaccinated and naturally-infected horses. In addition, the potential effect of antigenic selection of virus variants in egg versus tissue culture propagation systems was studied. Serum antibody levels in samples from horses recently infected with a local influenza A virus isolate (A/equine 2/Saskatoon/1/90) or recently vaccinated with a prototype isolate (A/equine 2/Miami/1/63) were assessed by hemagglutination inhibition and by single radial hemolysis using cell or egg-propagated A
Frontiers in microbiology, 2018
Among all the emerging and re-emerging animal diseases, influenza group is the prototype member associated with severe respiratory infections in wide host species. Wherein, Equine influenza (EI) is the main cause of respiratory illness in equines across globe and is caused by equine influenza A virus (EIV-A) which has impacted the equine industry internationally due to high morbidity and marginal morality. The virus transmits easily by direct contact and inhalation making its spread global and leaving only limited areas untouched. Hitherto reports confirm that this virus crosses the species barriers and found to affect canines and few other animal species (cat and camel). EIV is continuously evolving with changes at the amino acid level wreaking the control program a tedious task. Until now, no natural EI origin infections have been reported explicitly in humans. Recent advances in the diagnostics have led to efficient surveillance and rapid detection of EIV infections at the onset ...
Journal of Clinical Microbiology, 2004
Four seronegative foals aged 6 to 7 months were exposed to an aerosol of influenza strain A/Equi/2/ Kildare/89 at 10 6 50% egg infective doses (EID 50 )/ml. Nasopharyngeal swabs were collected for 10 consecutive days after challenge. Virus isolation was performed in embryonated eggs, and the EID 50 was determined for all positive samples. The 50% tissue culture infective dose was determined using Madin-Darby canine kidney (MDCK) cells. Samples were also tested by an in vitro enzyme immunoassay test, Directigen Flu A, and by reverse transcription-PCR (RT-PCR) using nested primers from the nucleoprotein gene and a single set of primers from the matrix gene. RT-PCR using the matrix primers and virus isolation in embryonated eggs proved to be the most sensitive methods for the detection of virus. The Directigen Flu A test was the least sensitive method. The inclusion of 2% fetal calf serum in the viral transport medium inhibited the growth of virus from undiluted samples in MDCK cells but was essential for the maintenance of the virus titer in samples subjected to repeated freeze-thaw cycles.
Anti-human influenza protector antibody detected in horses as a zoonotic viruses
Brazilian Journal of Veterinary Research and Animal Science, 2004
This study aimed to investigate the incidence of the influenza virus, and its interspecies transmission cycle among horses. A comparative serological survey was performed using horse sera following challenge with both specific (equine) and non-specific (human) influenza virus strains. Bleedings of the 22 horses were performed during the years 1999 and 2000. Following treatment with kaolin (20%), added in rooster erythrocytes suspension (50%), for removal of non-specific antibodies, sera were titered against both Human and Equine Influenza virus, by the Hemagglutination Inhibition Assay(HI), recommended by WHO. The HI results of horse serological responses demonstrate cross reaction between both the specific strain, A/Eq1 (H7N7) ( 62.75%) and A/Eq2 (H3N8) (60.65%), and the non-specific strains, type A (H1N1) (79.5%) and A (H3N2) (94.45%) and type B (77.75%). It was noteworthy the high percentage of protection responses in equine sera aginst the non-specific strains, as compared with the specific strains. This finding suggests direct interspecies transmission of influenza virus as zoonotic viruses, particulary for the type B strain which is considered restricted to humans. It was the first report, in Brazil. Further studies are required to achieve a complete understanding of the incidence of influenza in our environment.
Analysis of Different Diagnostis Methods of Influenza in Horses
Acta Scientiae Veterinariae
Background: Equine Influenza is a serious, acute respiratory illness with characteristical clinical signs. The disease is caused by family of Orthomyxoviridae, genera Influenza virus A by two subtypes H7N7 and H3N8. Currently, there is believe that H7N7 has been replaced as a predominant subtype with the H3N8. Horse infection with influenza virus can be detected by serological tests on paired sera using HI test. Commercial rapid tests could be used for the detection of influenza virus. Recently it is widely use a PCR method as fast and more specific methods.Materials, Methods & Results: Fifty horses and one pony, age between one and 22 years have been included in experiment. Horses were of different race, sex, and age and vaccination status. Ten out of total 51 (10/51) have been regularly vaccinated against EI. Prior to initiation of these study epidemiological survey has been performed. The clinical examination has been followed by blood sampling for blood cell and serum extraction...
2017
Equine influenza is an acute respiratory infection of horses, donkeys, mules and zebras caused by two distinct subtypes (H7N7, formerly equi-1, and H3N8, formerly equi-2) of influenza A virus within the genus Influenza virus A of the family Orthomyxoviridae. They are related to but distinct from the viruses that cause human and avian influenza. In fully susceptible equidae, clinical signs include pyrexia and a harsh dry cough followed by a mucopurulent nasal discharge. Vaccinated infected horses can still shed the virus and serve as a source of virus to their cohorts. Characteristically, influenza spreads rapidly in a susceptible population. The disease is endemic in many countries with substantial equine populations like Australia, Japan, South Africa. to date New Zealand and Iceland are reported to be free of equine influenza virus. This review paper consists of introduction ,identification of the agent, etiology and epidemiology, transmission, clinical signs, diagnosis, serological tests ,treatment, prognosis, requirements for vaccines ,vaccination against equine influenza, prevention and conclusion.