Changes

===Immunodiagnostic  tests===

===Immunodiagnostic  tests===

All of these tests aim to confirm exposure to the pathogens of EPM by detecting the presence of antibodies to these agents(FUrr). None of these tests is considered a gold standard and they are only supportive. Currently, a definitive diagnosis can only be obtained at postmortem.(IVIS 4)

All of these tests aim to confirm exposure to the pathogens of EPM by detecting the presence of antibodies to these agents(FUrr). None of these tests is considered a gold standard and they are only supportive. Currently, a definitive diagnosis can only be obtained at postmortem.(IVIS 4)

*'''Immunoblot analysis (Western blot) of serum and CSF''': cultured merozoites are used to detect antibodies versus ''S.neurona''-specific proteins.  The CSF test has over 90% specificity and sensitivity(86 in Furr).  The blood brain barrier is not prevent the passage of antibodies, thus the CSF concentration of a specific antibody will be directly related to its serum concnetration (87 in Furr).  This permeability is likely responsible for many of the weakly false-positive CSF immunoblot tests.  Blood contamination during CSF collection or bleeding within the CNS due to trauma or infection might also cause false positives.  The CSF titre will be greatly increased during CNS infection as there will be local production of the antibody.  One of the difficulties in interpreting immunoblot results is that many horses develop antibodies against ''S.neurona'' in the absence of neurological disease.(EPM4)  For this reason, testing CSF may be preferable to serum despite the impact that minor blood contamination may have on CSF results.(IVIS 4)False negative results may arise if horses fail to respond to the specific proteins recognised by the immunoblot.  Such cases are rare, so a negative immunoblot result tends to exclude the diagnosis of EPM.(Merck)  Cases that originally test negative should be re-tesed 14-21 days later.  In most instances, owing to a substantial incubation period, detectable levels of IgG are present prior to the emergence of clinical signs.(Furr)  

*'''Immunoblot analysis (Western blot) of serum and CSF''': senstivity around 90%, specificity 48-89%.(EPM4)  Cultured merozoites are used to detect antibodies versus ''S.neurona''-specific proteins.  The CSF test has over 90% specificity and sensitivity(86 in Furr).  The blood brain barrier is not prevent the passage of antibodies, thus the CSF concentration of a specific antibody will be directly related to its serum concnetration (87 in Furr).  This permeability is likely responsible for many of the weakly false-positive CSF immunoblot tests.  Blood contamination during CSF collection or bleeding within the CNS due to trauma or infection might also cause false positives.  The CSF titre will be greatly increased during CNS infection as there will be local production of the antibody.  One of the difficulties in interpreting immunoblot results is that many horses develop antibodies against ''S.neurona'' in the absence of neurological disease.(EPM4)  For this reason, testing CSF may be preferable to serum despite the impact that minor blood contamination may have on CSF results.(IVIS 4)False negative results may arise if horses fail to respond to the specific proteins recognised by the immunoblot.  Such cases are rare, so a negative immunoblot result tends to exclude the diagnosis of EPM.(Merck)  Cases that originally test negative should be re-tesed 14-21 days later.  In most instances, owing to a substantial incubation period, detectable levels of IgG are present prior to the emergence of clinical signs.(Furr)  

*'''Whole organism indirect fluorescent antibody test (IFAT)''': serum titres of more than 1:100 and CSF titres of more than 1:5 indicate an active infection. The IFAT is considered to have slightly improved diagnostic efficiency than the immunoblot test(92 in furr) but is unable to distinguish between ''S.neurona'' and other related nonpathogenic organsims such as ''S.fayeri''(94 in Furr).  This can lead to false positive results.  Compared with the immunblot test, CSF blood contamination has an insignificant effect on the IFAT.(11 in IVIS 4)  An IFAT for ''N.hughesi'' is also available from the Universty of California.(Furr)

*'''Whole organism indirect fluorescent antibody test (IFAT)''': sensitivity around 90%, specificity 97-100%.(EPM4)  Serum titres of more than 1:100 and CSF titres of more than 1:5 indicate an active infection. The IFAT is considered to have slightly improved diagnostic efficiency than the immunoblot test(92 in furr) but is unable to distinguish between ''S.neurona'' and other related nonpathogenic organsims such as ''S.fayeri''(94 in Furr).  This can lead to false positive results.  Compared with the immunblot test, CSF blood contamination has an insignificant effect on the IFAT.(11 in IVIS 4)  An IFAT for ''N.hughesi'' is also available from the Universty of California.(Furr)

*'''ELISA for antibodies to the snSAG-1 protein''': based on an immunodominant surfae antigen of S.neurona (SAG-1).(IVIS 4)  Serum titres more than 1:100 suggest an active infection.  False negatives are possible as not all ''S.neurona'' isolates produce the specific protein.(58 in Furr).  SAG-5 is an alternative surface antigen of ''S.neurona'' strains, which is mutually exclusive to SAG-1.(15 in IVIS 4) Therefore, the ELISA may only be of use where strains of ''S.neurona'' expressing SAG-1 predominate.(IVIS 4)

*'''ELISA for antibodies to the snSAG-1 protein''': based on an immunodominant surfae antigen of S.neurona (SAG-1).(IVIS 4)  Serum titres more than 1:100 suggest an active infection.  False negatives are possible as not all ''S.neurona'' isolates produce the specific protein.(58 in Furr).  SAG-5 is an alternative surface antigen of ''S.neurona'' strains, which is mutually exclusive to SAG-1.(15 in IVIS 4) Therefore, the ELISA may only be of use where strains of ''S.neurona'' expressing SAG-1 predominate.(IVIS 4)

Available evidence indicates that both the WB and IFAT have similar sensitivities (_90%). The estimated IFAT specificity is higher than the estimated WB specificity, and the IFAT CSF results are much less affected by blood contamination than WB CSF results. The

*'''CSF analysis''': to rule out other conditions as stated below.  Most horses with EPM have normal CSF (Furr). Rarely, increased totoal protein or white blood cell count is seen in severe cases(Furr). PCR can be used to detect ''S.neurona'' DNA in CSF. (EPM8)

SAG-1 ELISA has the least amount of evidence to support its use, and preliminary data yield a low sensitivity that will likely limit its use in some or most geographic regions.(IVIS 4)

*'''Diclazuril''': a positive response to treatment with diclazuril would firmly support a diagnosis of EPM, since the drug has no antimicrobial activity.(EPM 1)

 

*'''Blood gene expression biomarkers''': may be sensitive and specific indicators of early and active disease<ref>Eastman, E, Furr, M, McKenzie, H, Saville, W.J, Dubey, J.P (2005) Early diagnosis of Sarcocystis neurona infection  using bloodgene expression biomarkers.  In:  ''51st Annual Convention of the American Association of Equine Practitioners - AAEP'', Seattle, WA, USA.</ref>

To rule out other conditions as stated below.  Most horses with EPM have normal CSF (Furr). Rarely increased totoal protein or white blood cell count is seen in severe cases(Furr). PCR can be used to detct S Neorna DNA (EPM8)

 

The PCR could be useful in the postmortem diagnosis of EPM in cases in which there are only small numbers of protozoal pathogens.  In two of the horses, both apicomplexan pathogens were detected. The infection with N hughesi would have been missed, in the absence of detectable agent, by routine histology and by S neurona specific immunohistochemistry.(EPM6)

 

Diclazuril is believed to exhibit no antibacterial activity. A positive clinical response to the treatment with diclazuril would provide strong support of a diagnosis of EPM. Likewise, lack of response may add support to the presence of an alternative condition. (EPM 1)

 

 

The demonstration of a distinct gene expression biomarker signature for clinical EPM disease in equine peripheral blood leukocytes represents a significant advance over current methods for the diagnosis of EPM. The biomarker signature represents a specific and sensitive indicator of active and early disease.  Further studies on the applicability of this gene signature in clinical cases and in chronic disease are currently underway.

Eastman, E, Furr, M, McKenzie, H, Saville, W.J, Dubey, J.P (2005) Early diagnosis of Sarcocystis neurona infection  using bloodgene expression biomarkers.  In:  51 Annual Convention of the American Association of Equine Practitioners - AAEP, 2005 - Seattle, WA, USA