Frequently asked questions about Equine Infectious Anemia (EIA)

What is Equine Infectious Anaemia (EIA)?

It is a disease caused by a virus that affects members of the equidae family (horses, zebras, donkeys, and mules).

The only defenses are prevention measures, laboratory diagnoses, and proper management.

There is legislation in place governing official equine health control programs in all countries, as it is a mandatory reporting disease. The reference serum test is Agar Gel Immunodiffusion Test (AGID), considered by the World Organisation for Animal Health (OIE) as the ‘gold’ standard test for Equine InfectiousAnaemia.

Etiology of EIA:

Equine Infectious Anaemia (EIA) is a persistent viral infection caused by a virus RNA genome, highly mutagenic, which integrates into the host genome.

The EIA virus belongs to the Lentivirus genus of the family Retroviridae, a sub-family of the Orthoretrovirinae, and is genetically and antigenically related to other lentivirus of the genus, such as caprine arthritis encephalitis virus (CAEV), bovine immunodeficiency (BIV), feline immunodeficiency (FIV), human immunodeficiency 1, human immunodeficiency 2, and visna-maedi virus.

EIA Symptoms

  1. The horse may be recently infected with acute symptoms of the disease and a potential life expectancy of 2 or 3 weeks. However, the disease is only identified through the detection of an increase in body temperature that may go unnoticed by the caretaker. In general, however, the horse recovers and continues within the team. IEA is only detected through a routine AGID test.
  2. If the horse survives the chronic stage, it will present recurring symptoms of fever, spots on the mucosa, anaemia, edemas on the lower limbs, weight loss, anorexia.
  3. However, most horses are symptomless carriers. Yet, as most equines continue in service, this condition may transform into chronic or acute symptoms and when subject to excessive work, it  results in an acute attack and death.

How is the disease diagnosed?

Due to the persistence of EIAV in infected equidae, detection of EIAV antibodies confirms the infection. For this purpose, the p26 protein of the viral core is employed for diagnosis purposes.

To clarify and by way of example, the virion contains:

gp90 virus envelope protein, between 60% and 70% of mutations in the env gene are located in the sequences that codify gp90, given a quantity of mutations 3 times greater than the changes in gp45; by contrast to gp90, the envelope sequence of gp45  is highly conserved in all isolated samples of the EIA virus.

At the same time, with regard to the structural proteins of the predominant proteins in lentivirus up to 90% of the protein structure is codified by the gag gene, of which the p26 protein is the largest protein in the viral core. It is highly conserved in all isolated samples of the Equine Infectious Anaemia virus in the world. As such, all diagnostic tests are based on detection of antibodies for the p26 protein.

p26 is more antigenically stable in isolated EIAV than gp45 and gp90 glycoproteins (OIE Terrestrial Manual 2013 Chapter 2.5.6-2.1.1) and the one used in commercial diagnosis reagents.

While the ELISA can detect antibodies slightly earlier and in smaller concentrations than the AGID, a positive test result by ELISA should be retested using the AGID test to confirm the diagnosis because some false-positive results have been noted with the ELISA, which do not occur in immunodiffusion procedures because of the specificity of the identity reaction (OIE Terrestrial Manual 2013 Chapter 2.5.6, page 2).

How is the immunodiffusion reaction test (AGID) accomplished?

After 24–48 hours the precipitation reactions are examined.

The reference lines should be clearly visible at 24 hours.

A weakly positive reaction may take 48 hours to form and is indicated by a slight bending (OIE Terrestrial Manual 2013 Chapter 2.5.6 Equine Infectious Anaemia 2.1.3 iv, and regional legislation).

Larger test times will invalidate the AGID technique.  Weak positive sera  should be read within the time specified by the OIE technical standard and regional standards.

Can horses with Equine Infectious Anaemia (EIA) cease to be positive?


The Equine Infectious Anaemia Virus is a persistent infection.

Once the animal has been infected, it will remain a carrier throughout its life.

It may transmit the disease through the blood to other equidae, principally through hematophagic insect bites, during mating or gestation, or through contaminated instruments that come into contact with open wounds or the blood of an animal (syringes, surgical implements, spurs, etc).

For laboratory diagnosis, it is recommended that veterinarians always include in the tests a weak positive serum  with a minimum number of antibodies to enable comparative analysis of bending in relation to test weak positive serum and to complete the analysis of bending within the 48 hours prescribed in the applicable national and international legislation (OIE).

What are the product’s storage and transportation temperatures?

Storage Temperature:

For daily use, the CRM EIA should be stored at 5oC ±3oC and when not used for prolonged periods, frozen at -15oC e -20oC or less.

The CRM EIA  is stable when stored at temperatures below 8oC.

Transportation Temperature:

When transported, the product may be refrigerated or frozen depending on the distance traveled.

Validation of the transport temperature indicates that small variations in temperature above 8oC to 14oC are permitted during scheduled transport times of 3-5 days, which will not result in product non-conformity.

What is the Certified Reference Material for EIA diagnosis by AGID?

Reference material refers to material sufficiently homogeneous and stable with reference to specified properties, which has been established to be fit for its intended use in a measurement or in an examination of nominal properties. (VIM 2012-5.13- International Vocabulary of Metrology).

Certified reference material (CRM) refers to reference material accompanied by documentation issued by an authoritative body and providing one or more specified property values with associated uncertainties and traceabilities, using valid procedures. (VIM 2012-5.14- International Vocabulary of Metrology).

The International Laboratory Accreditation Cooperation (ILAC) defines the elements required to confirm metrological traceability as an uninterrupted traceability chain related to an international standard or national standard, documented calibration procedures, and recognized technical competence. (VIM 2012-2.41- International Vocabulary of Metrology).

Bruch certified reference  material for EIA Antigen and standard serum PMRC 0008  is intended for accredited Laboratories in conformity with the international quality standard ABNT NBR ISO/IEC 17025:2005 – General requirements for the competence of testing and calibration laboratories, Validation of Methods (requirement 5.4.5), and Reference Materials (requirement

Bruch certified reference  material for EIA Antigen and standard serum PMRC 0008,for the diagnosis of EIA by Agar Immunodiffusion (AGID) uses p26 antigen produced in cell cultures and equine serum obtained from animals with a history of disease. Standardization is in conformity to regional standards and to the international OIE standard. The Ag p26 materials and standard serum for AGID were produced as per ISO Guide 34 and submitted to homogeneity, stability, and characterization studies, pursuant to Guides 34 and 35. Characterization by qualitative attributes using immunodiffusion. The certified value has guaranteed traceability through the characterization of the CRM, being the traceability conducted by a qualitative property value.

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