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EHV-1 Risk Evaluation Assay

View the EHV-1 entry in our Test and Fee Database

Summary

Equine Herpesvirus-1 (EHV-1) causes respiratory disease, abortion, or neurologic disease known as equine herpesvirus myeloencephalopathy (EHM) in equids. In some horses EHM is fatal. Others can recover from neurologic disease. Ongoing EHV-1 and EHM outbreaks support the need for better protection of horses against EHV-1. The Serology/Immunology laboratory at the AHDC developed and offers a novel EHV-1 Risk Evaluation assay to determine the EHV-1 immune status and risk of EHV-1 infection for an individual horse. The assay’s interpretation is based on experimental vaccination and challenge studies performed at Cornell and revealed that specific antibodies (anti-EHV-1 IgG4/7 antibodies) highly correlate with protection from EHV-1 infection, disease, viral shedding and cell-associated viremia.1,2 These protective IgG4/7 antibodies have high neutralizing abilities for EHV-1 and immediately inactivate the virus at the respiratory entry site.3 We and others have also noticed that horses respond individually different to vaccination. Responses of horses to EHV vaccination can vary in both, magnitude and duration of the immune response. The EHV-1 Risk Evaluation assay is designed to address this variation by providing a tool to optimize immunity against EHV-1 infection in each individual horse. EHV-1 antibodies measured by the Risk evaluation assay distinguish if a horse is at low, medium or high risk of infection and disease if exposed to EHV-1. Based on the antibody values at the time of sampling, the assay also provides a recommendation when vaccination is next needed to maintain the ‘low risk status’ in a horse or other equid. Combining EHV-1 Risk Evaluation assay results with an individually optimized vaccination strategy for each horse provides optimal protection against EHV-1 infection. During an EHV-1/EHM outbreak, the EHV-1 Risk Evaluation assay can also provide a valuable management tool, informing about the host immune status against EHV-1 in horses that may have been exposed to the virus.4

Background

Despite frequent use of EHV vaccination in the US horse population, EHV-1 continues to be responsible for neurological EHM outbreaks at racetracks, equine competition grounds, horse sale events, veterinary teaching hospitals, and other locations with equine traffic or new equids being introduced. Many horses are latently infected with EHV-1. Periods of stress can reactivate the virus which can then lead to an EHV-1 or EHM outbreak. Observations during the past years suggested that only a single or very few horses developed EHM during EHV-1 outbreaks in regularly vaccinated horse populations, e.g. at United States Equestrian Federation (USEF) competitions. In contrast, several horses can develop EHM if an outbreak includes a larger number of horses without sufficient immunity against EHV-1. Quarantine and event closures take place after a first EHM case is confirmed by an EHV-1 positive PCR result.5,6 Breeding farms can experience outbreaks of EHV-1 abortions, which can also involve signs of EHM in some affected horses.7,8 EHV-1 outbreaks have major animal welfare impacts and put high economic constraints on the equine industry.

Recommended prevention strategies against EHV-1 infection include vaccination and biosecurity practices. While vaccines reduce clinical disease and shedding9,10 some EHV cases have occurred in frequently vaccinated horses.7,8 Available inactivated vaccines are licensed for prevention of respiratory disease, or both respiratory disease and abortion. None of the currently approved EHV vaccines claims to prevent EHM. In addition to several inactivated vaccines, a modified-live vaccine is available in North America and Europe and provides protection against fever and other clinical signs of disease,8,9 but is not approved for use in pregnant mares. For prevention of abortion, repeated vaccination with an inactivated EHV vaccine is recommended by vaccine suppliers. The EHV vaccination guidelines of the American Association of Equine Practitioners (AAEP) recommend frequent vaccination for many brood mares resulting in 3-4 EHV vaccinations per year for many years. Recent data have shown that frequent vaccination of pregnant mares can be counter-productive and reduce antibody values.11 For competition horses, EHV vaccination is currently recommended every 6 months by AAEP12 and USEF competition rules require documentation that the horse has been vaccinated within the past six months preceding an event.13 A recent study confirmed that intramuscular vaccination results in both, systemic and nasal mucosal EHV-1 specific IgG4/7 antibodies.14 These local mucosal antibodies rapidly neutralize EHV-1 in the upper respiratory track after infection3 and directly correlate with protection against disease caused by EHV-1. 

What distinguishes the EHV-1 Risk Evaluation assay and EHV-1 antibody testing from conventional methods such as EHV-1 serum neutralization tests?

Antibody responses to EHV vaccination or exposure to EHV-1 have commonly been measured by serum neutralization (SN) assays, complement fixation, or ELISA.10 These assays provide quantitative titers or values on antibodies against EHV but they neither distinguish antibody subtypes (isotypes), nor do they provide a risk evaluation or vaccination recommendation based on the current antibody titer.

In the EHV-1 Risk Evaluation assay, specific antibodies (total antibodies and IgG4/7) against EHV-1 are determined. These EHV-1 antibodies highly correlate with protection from infection and disease induced by EHV-1,1-3 and enable the use of the EHV-1 Risk Evaluation assay for infection risk evaluation at the individual horse level or in populations of horses. More specifically, the EHV-1 Risk Evaluation assay can be used to:

  • determine the risk of infection (including risk for developing clinical signs of disease, nasal virus shedding and cell-associated viremia) following EHV-1 exposure in non-pregnant horses;
  • identify horses and other equids at high risk of EHV-1 infection (if exposed to EHV-1, these horses will get infected and, likely, develop clinical signs of disease, shed and transmit virus and develop cell-associated viremia, the latter is the pre-requisite for EHM);
  • make informed decisions on EHV vaccination based on the current antibody results in non-pregnant horses and equids of all risk groups;
  • adjust vaccination protocols in protected horses, e.g. by extending vaccination intervals in the EHV-1 low risk group with high antibody amounts, or by vaccinating ‘low responders’ more frequently;
  • inform when vaccination is required in horses with a history of side-effects to vaccination;
  • obtain supportive information about the EHV-1 host immune status during an EHV-1 outbreak (early in the course of an outbreak when a horse is found to be at risk of exposure to EHV-1, antibody testing can be performed on serum and nasal secretion samples to evaluate immunity or lack thereof and may help determine if a horse has become exposed4). 

Why is the EHV-1 immune status of your horse important?

Horses develop a wide range of responses to EHV vaccination. Antibody responses after vaccination vary individually. They can range from very high to low depending on the individual horse’s genetically determined immune response, and upon the way the horse’s immune system recognizes vaccine components. Although current vaccine formulas have been optimized to overall result in very good immunity and antibody titers, a small proportion of horses usually produce low and/or short-lasting antibodies. In the case of EHV-1, this can result in loss of protection in individual horses and explains the observation that EHV-1 and EHM cases can sometimes occur in vaccinated horses. In the opposite case, some horses can have high and long-lasting EHV-1 antibodies after many years of regular vaccination. These horses can show local side-effects, such as pain and swelling at the vaccine injection site. For these horses, the EHV-1 Risk Evaluation provides quantitative antibody values for informed decision making regarding the need for booster vaccination, and simultaneously gives ‘peace of mind’ that the horse is well protected.

Which protective antibodies does the EHV-1 Risk Evaluation assay measure?

Herpesviruses have several highly antigenic envelope glycoproteins. EHV-1 glycoprotein C (gC) is used as the antigen in the EHV-1 Risk Evaluation assay. It is involved in cell entry, has been used as target for vaccine development, and shown to induce B-cell responses and neutralizing antibodies against human herpesviruses and EHV-1. Besides its function in cell entry, EHV gC can interfere with the activation of the complement cascade by binding to the complement component C3. Previous research has shown that EHV-1 infection or vaccination induces a broad immune response against several EHV-1 glycoproteins and that gC-specific antibodies are a sensitive indicator for the broad anti-viral immune response.1-3,11,14

Does the assay distinguish immunity against EHV-1 and EHV-4?

EHV-1 gC shares high sequence homology with the gC antigen of EHV-4 and serological responses cross-react between EHV-1 and EHV-4 gC. For immune protection it does not matter if the antibodies were induced by EHV-1 or EHV-4 antigen. Neutralizing ability and protection against EHV-1 are influenced by the antibody isotype. Anti-gC IgG4/7 antibodies neutralize the virus and are protective. Anti-gC IgG4/7 antibodies also represent the long-lasting antibody response against EHV-1.1,2,13

EHV-1 Risk Evaluation assay interpretation

The EHV-1 Risk Evaluation assay quantifies protective antibodies in serum or plasma. These antibodies distinguish if a horse is at low, moderate or high risk of getting infected and developing disease following EHV-1 infection. The assay’s antibody result also provides a recommendation when EHV vaccination is next needed to maintain or improve protection of the horse against EHV-1 infection and disease.

EHV-1 risk categories and vaccination recommendations

Anti-gC total Ig <3,000 MFI & anti-gC IgG4/7 <400 MFI

The horse (equid) is at high risk of EHV-1 infection and, if exposed to the virus, has a high likelihood of developing disease. Immediate EHV vaccination is needed to increase anti-gC total Ig and IgG4/7 antibodies and to protect the horse from EHV-1 infection.

Anti-gC total Ig <3,000 MFI & anti-gC IgG4/7 ³400 MFI; or anti-gC total Ig ³3000 MFI & anti-gC IgG4/7 <400 MFI

The horse (equid) is at moderate risk of EHV-1 infection and, if exposed to the virus, has some likelihood of developing disease. Vaccination is recommended within the next month to boost the EHV-1 antibodies.

Anti-gC total Ig >3,000 MFI & anti-gC IgG4/7 >400 MFI

The horse (equid) is at low risk of EHV-1 infection and of developing disease. EHV vaccination is not needed at this time. Based on the EHV-1 antibody values, the horse should be vaccinated in the next 3-6 months.

Anti-gC total Ig >12,000 MFI & anti-gC IgG4/7 >10,000 MFI

The horse (equid) is at very low to no risk of EHV-1 infection and of developing disease. EHV vaccination is not needed for the next 12 months to maintain the EHV-1 immune biomarker values in the low-risk category. High EHV-1 antibodies in these MFI ranges are often seen in horses after curing a recent EHV-1 infection and can also occur in some well-vaccinated horses.

Can the EHV-1 Risk Evaluation assay help to manage horses and infection risk in outbreak situation?

During an EHV-1/EHM outbreak, the EHV-1 Risk Evaluation assay together with testing of intranasal EHV-1 antibodies and inflammatory markers can inform about the immune and EHV-1 exposure status of a horse.4 For optimal interpretation, it is important to take the samples as early as possible after the EHV-1 index case is confirmed. For more information on combined blood and nasal swab sample submissions and on testing during an outbreak, please contact the AHDC.

References

  1. Perkins G, Babasyan S, Stout AE, Freer H, Rollins A, Wimer CL, Wagner B. 2019. Intranasal IgG4/7 antibody responses protect horses against equid herpesvirus-1 (EHV-1) infection including nasal virus shedding and cell-associated viremia. Virology, 531: 219-232.
  2. Schnabel CL, Babasyan S, Rollins A, Heather Freer H, Wimer CL, Perkins GA, Fahad R, Osterrieder N, Wagner B. 2019. An equine herpesvirus type 1 (EHV-1) Ab4 open reading frame (ORF)2 deletion mutant provides immunity and protection from EHV-1 infection and disease. J. Virology, 93(22):e01011-19.
  3. Eady NA, Holmes C, Schnabel C, Babasyan S, Wagner B. 2024. Equine herpesvirus type-1 (EHV-1) replication at the upper respiratory entry site is inhibited by neutralizing EHV-1 specific IgG1 and IgG4/7 mucosal antibodies. J. Virol., 98: e0025024. doi: 10.1128/jvi.00250-24.
  4. Perkins GA, Wagner B, Rollins A, Sfraga H, Pearson E, Cercone M. 2025. Antibody testing to detect viral exposure in contact horses during an equine herpesvirus myeloencephalopathy outbreak. AJVR, doi.org/10.2460/ajvr.25.03.0106.
  5. Henninger RW, Reed SM, Saville WJ, Allen GP, Hass GF, Kohn CW, Sofaly C. 2007. Outbreak of neurologic disease caused by Equine Herpesvirus-1 at a University Equestrian Center.  J Vet Intern Med;21;157-65.
  6. Traub-Dargatz JL, Pelzel-McCluskey AM, Creekmore LH, Geiser-Novotny S, Kasari TR, Wiedenheft AM, Bush EJ, Bjork KE. 2013. Case-control study of multistate equine herpesvirus myeloencephalopathy outbreak.  JVIM;27:339-46.
  7. Walter J, Seeh C, Fey K, Bleul U, Osterrieder N. 2013. Clinical observations and management of a severe equine herpesvirus type 1 outbreak with abortion and encephalomyelitis. Acta Vet Scand;55:19.
  8. Damiani AM, de Vries M, Reimers G, Winkler S, Osterrieder N. 2014. A severe equine herpervirus type 1 (EHV-1) abortion outbreak caused by a neuropathogenic strain at a breeding farm in northern Germany. Vet Microbiol;172:555-62.
  9. Goehring LS, Wagner B, Bigbie R, Hussey SB, Rao S, Morley PS, Lunn DP. 2010. Control of EHV-1 viremia and nasal shedding by commercial vaccines. Vaccine;28:5203-11.
  10. Goodman LB, Wagner B, Flaminio MJ, Sussman KH, Metzger SM, Holland R, Osterrieder N. 2006. Comparison of the efficacy of inactivated combination and modified-live virus vaccines against challenge infection with neuropathogenic equine herpesvirus type 1 (EHV-1). Vaccine;24: 3636-45.
  11. Wagner B, Goodman LB, Babasyan S, Freer H, Torsteinsdóttir S, Svansson V, Björnsdóttir S, Perkins GA. 2015. Antibody and cellular immune responses of naïve mares to repeated vaccination with an inactivated equine herpesvirus vaccine. Vaccine;33:5588-97.
  12. Equine Herpesvirus (Rhinopneumonitis); Vaccination guidelines; AAEP.
  13. GR845 Equine vaccination rule USEF.
  14. Wagner B, Schnabel CL, Rollins A. 2025. Increase in virus-specific mucosal antibodies in the upper respiratory tract following intramuscular vaccination of previously exposed horses against equine herpesvirus type-1/4. Vaccines, 13(3), 290.