Indirect Fluorescent Antibody Test and Surface Antigen ELISAs for Antemortem Diagnosis of Equine Protozoal Myeloencephalitis

Abstract

Background: Recent research suggests that serum : CSF titer ratios could provide the most accurate antemortem diagnosis of equine protozoal myeloencephalitis (EPM) caused by Sarcocystis neurona.Objectives: The purpose of this study was to assess the utility of two commercially available tests, the indirect fluorescent antibody test (IFAT) and the surface antigen 2, 4/3 ELISA (SAG2, 4/3 ELISA), using archived paired serum and CSF samples.Animals: Samples were obtained from 4 types of clinical patients. Confirmed positive cases (n = 9 horses; 11 sample sets) had neurologic deficits and postmortem lesions consistent with EPM. Confirmed negative cases (n = 28) had variable clinical signs and postmortem lesions consistent with another disease. Suspected positive cases (n = 6) had neurologic deficits consistent with EPM, marked improvement after treatment, and exclusion of other diseases. Suspected negative cases (n = 14) had variable signs with a strong presumptive diagnosis of another disease.Methods: For each test, descriptive statistics were calculated using serum results alone, CSF results alone, and a serum : CSF titer ratio.Results: Overall accuracy was highest for SAG2, 4/3 ELISA titer ratio at 0.97 (95% CI 0.88-0.99) with sensitivity = 0.88 (95% CI 0.66-0.97) and specificity = 1 (95% CI 0.92-1). IFAT CSF and titer ratio results also showed high accuracy at 0.88 (95% CI 0.77-0.94), but lower sensitivity = 0.65 (95% CI 0.41-0.83).Conclusions and Clinical Importance: Using serum results alone was least accurate for both test types. The more accurate methods, such as the SAG2, 4/3 ELISA serum : CSF titer ratio, should be utilized.

Equine Protozoal Myeloencephalitis: An Updated Consensus Statement with a Focus on Parasite Biology, Diagnosis, Treatment, and Prevention

“…This study also showed that minor blood contamination of the CSF sample (ie, up to 10,000 red cells per μL) will not confound the assay results. Subsequently, 2 additional studies examining a more extensive collection of horses with neurologic disease showed that a simple serum:CSF antibody titer ratio was sufficient in many cases for an accurate diagnosis of EPM caused by S. neurona 71, 72. Although use of a serum:CSF titer ratio should be equally effective for diagnosis of EPM caused by N. hughesi , an optimal serum:CSF titer ratio cut‐off needs to be established.…”

“…Descriptions of testing options and reported test performance are shown in Table 1 71, 72, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93. All tests can be performed on serum or CSF, and none is considered a gold standard.…”

“…The IFAT is a quantitative (end‐point titer) test for antibodies against culture‐derived whole merozoites. Although serum titers obtained with the IFAT have been used to predict the likelihood of EPM, with higher titers suggesting greater probability of disease, studies that have used diverse collections of neurologic disease cases have shown that a serum titer alone is a poor predictor of EPM 71, 72. As a quantitative test, the IFAT can be used to calculate a serum:CSF titer ratio.…”

“…These molecules have proven to be good serologic targets in the assays because of their high level of expression in the parasite and their immunogenicity in infected horses 94, 95, 96. The SnSAG2 ELISA and the SnSAG4/3 ELISA accurately detect antibodies against S. neurona in equine serum and CSF samples88, 91 and were used to demonstrate the value of detecting intrathecal antibody production for EPM diagnosis 71, 72. An ELISA based on the SnSAG1 surface protein has been described 86.…”

Biotechnological tools for diagnosis of equine infectious diseases

“…The sarcocystis neurona causes a dreadful disease, equine protozoal myeloencephalitis (EPM) in equines. The IFAT was successfully validated for CSF testing for confirmation of EPM in equines (Duarte et al, 2006;Johnson et al, 2013).…”