EQUINE VETERINARY EDUCATION Equine vet. Educ. (2022) 34 (7) 381-388 doi: 10.1111/eve.13491
Review Article
A practitioner’s guide to understanding infectious disease diagnostics in the United Kingdom. Part 2: Serological diagnostic testing methods and diagnostic test result interpretation
F. M. Whitlock* and J. R. Newton
British Horseracing Authority, London, UK *Corresponding author email:
fleurwhitlock1@gmail.com Keywords: horse; diagnostics; infectious disease; laboratory
Summary This article is the second in a review series about infectious disease diagnostics and summarises serological testing techniques, diagnostic test validation and result interpretation. It highlights the requirement for results to be interpreted with testing limitations in mind and in conjunction with the case’s clinical presentation. With these foundations, the treating clinician should always be well placed to deal with equine infectious diseases.
Introduction
A good understanding of laboratory-based sample analysis techniques and their validation is necessary for optimal diagnostic test result interpretation. Part 1 in this series discussed the importance of agent detection testing in infectious disease outbreaks, how this is achieved and the different testing methods available for agent detection (Whitlock and Newton 2022). This second section will summarise serological testing techniques, test validation and results’ interpretation. A more extensive guide to international infectious disease outbreak handling and eradication can be found in the HBLB International Codes of Practice (
https://codes.hblb.org.uk/). Additional information on laboratory diagnostics for equine infectious diseases endemic to North America can be found in the AAEP Infectious Disease Guidelines (
https://aaep.org/ guidelines/infectious-disease-control/using-guidelines).
Immunological response to infection diagnostic testing methods
Serology is used to analyse samples for the presence of pathogen-specific antibodies. Their presence demonstrates an immune response (at some point) to the specific pathogen or to vaccination against the specific pathogen (if vaccine is not a marker vaccine that can be readily differentiated from infection). Although this is not an exhaustive list, the most commonly used serological testing methods for equine infectious diseases present in the UK will be discussed and are summarised in Table 1.
Enzyme-linked immunosorbent assay
An enzyme-linked immunosorbent assay (ELISA) detects antigen–antibody complexes by an enzyme–substrate reaction. Viral antigen is bound to the plate, the sample serum is added and if the sample contains the target antibody (primary antibody conjugate) to the test, it will bind
to the viral antigen with a resultant colour change in a direct ELISA (Fig 1). Indirect, sandwich and competitive ELISAs utilise secondary antibodies labelled with an enzyme, and if any primary antibody is present, complexes will form with a resultant colour change. An example of when an ELISA may be used is when testing to determine whether a sample contains Streptococcus equi (S. equi)-specific antibodies. Currently, there are two different ELISAs available and a clinician is advised to familiarise themselves with the merits of each of these, to assist in choice of assay and result interpretation. In short, an ELISA based on the SeM protein was initially developed to identify horses that may be at risk of sequelae following S. equi infection, such as metastatic abscessation, or from developing purpura haemorrhagica following receipt of an SeM-containing vaccine (Sweeney et al. 2005). Genetically, S. equi is very similar to Streptococcus zooepidemicus (S. zooepidemicus) and it was found that this test had the propensity to produce false- positive results due to this cross-reactivity (Kelly et al. 2006). Another assay was developed utilising proteins that were more specificto S. equi, to avoid the issues of cross-reactivity with S. zoopidemicus (Robinson et al. 2013). As previously discussed in Part 1 of this series, infectious disease diagnostic testing must be utilised and interpreted in light of the clinical scenario, whilst also having an awareness of the accuracy and limitations of each available diagnostic test for a disease. Again, S. equi serological testing is a good example for this. The most optimal application of the ELISA test has been reported to be following a recent confirmed strangles outbreak, to determine a horse’s exposure status, with those exposed being at risk of being persistently infected. This then enables more targeted agent detection diagnostics of those cases with serological evidence of being exposed (Newton et al. 2000). Clinicians should have an awareness of the limitations of utilising the ELISA as a screening test when previous potential exposure status to S. equi is unknown. This is as S. equi carriers may not maintain detectable antibodies, despite being chronically infected (Bowen et al. 2020). It is commonplace that infectious disease diagnostics may require various sample types, multiple sampling time points and a series of different diagnostic tests, in order to obtain a correct diagnosis.
Advantages of ELISA • High sensitivity and specificity • Quick to run • Capacity for high volume and automated sample throughputs
© 2021 EVJ Ltd
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