ECUN

ECUN Serological Testing

About the Assays

The antibody test is performed in the ELISA format and results are currently reported as a titer – meaning the sample is diluted and tested through the endpoint of its reactivity on the ELISA. All samples are screened at a 1:64 dilution for IgM and IgG E. cuniculi specific antibodies. CRP is a standardized automated assay providing quantitative results.

Sample Collection and Submission

Fresh non-hemolyzed serum or plasma samples are desirable. Freezing should be considered if delays greater than 2 days are present. All samples should be centrifuged and separated to a size appropriate transport tube even if a gel separator is present in the tube.

Interpretation of Results

It is common to have seropositive (ECUN IgG+) rabbits. However, those rabbits which are infected carry significantly higher titers of antibody. The mean titer of infected animals is 1:1324. In addition, they demonstrate significantly higher levels of gamma globulins. Notably, animals with ocular involvement vs. those with neurological or renal signs demonstrated lower amounts of antibody. Studies of experimentally and naturally infected rabbits have demonstrated a strong association of IgM titers with active infection. Studies have not yet been completed to address the longevity of these IgM titers at this time. The absence of IgM suggests exposure or previous infection; however, given our previous studies, high IgG titers without IgM seroconversion should still be strongly considered as infected animals.

In a study of IgM and IgG titers, using a cutoff titer of greater than or equal to 1:64 for IgM and greater than or equal to 1:512 for IgG, a positive predictive value of 92% was observed. More data regarding this finding can be found in the 2015 JEPM publication listed below.

CRP is a major acute phase protein in rabbits. This biomarker will increase with systemic inflammatory processes. While not diagnostic of ECUN infection, the elevation of this protein will be supportive of infection and results can be interpreted in conjunction with IgG and IgM titers. Abnormal IgM and IgG titers (see above) in conjunction with an elevated CRP level provides a positive predictive value of 100%. Acute phase proteins can provide key prognostic value. With a positive response to treatment, CRP levels can drop within 24 hours.

While positive titers can be difficult to interpret, it is notable that in our studies that only 3% of rabbits with clinical signs consistent with E. cuniculi infection were found to be seronegative. Seronegative status has been reported in especially young animals and in animals with other concurrent diseases.

References:

C. Cray, G. Arcia, S. Kelleher, R. Schneider, and K. Arheart. Application of ELISA and protein electrophoresis in the diagnosis of Encephalitozoon cuniculi infection in rabbits. American Journal of Vetertinary Research, 70(4):478-482, 2009.

C. Cray. New testing option for the diagnosis of Encephalitozoon cuniculi in rabbits. ExoticDVM, 11(2):27-28, 2009.

C. Cray, M. Rodriguez, and Y. Fernandez. Acute phase protein levels in rabbits with suspected Encephalitozoon cuniculi infection. Journal of Exotic Pet Medicine, 22:280-286, 2013.

S. Berger Baldotto, C. Cray, A. Turner Giannico, L. Reifur, and F. Montiani-Ferreira. Seroprevalence of Encephalitozoon cuniculi infection in pet rabbits in Brazil. Journal of Exotic Pet Medicine, 24:435-440, 2015.

C. Cray, S. McKenny, E. Perritt, and K.L. Arheart. Utility of IgM titers with IgG and C-reactive protein quantitation in the diagnosis of suspected Encephalitozoon cuniculi infection in rabbits. Journal of Exotic Pet Medicine, 24:356-360, 2015.

G. Desoubeaux, A. Pantin, R. Peschke, A. Joachim, and C. Cray. Application of Western blot analysis for the diagnosis of Encephalitozoon cuniculi infection in rabbits: example of a quantitative approach. Parasitology Research, 116:743-750, 2017.