Characterizing Agreement of Hemagglutination Inhibition Assays Using Four Different Ophidian Paramyxovirus Isolates in Wild Eastern Massasauga Rattlesnakes (Sistrurus catenatus catenatus) from Illinois
American Association of Zoo Veterinarians Conference 2007

Matt Allender1, DVM, MS; Mark A. Mitchell2, DVM, MS, PhD; Michael J. Dreslik3, MS, PhD, Christopher Phillips3, MS, PhD; Val R. Beasley2, DVM, PhD, DABVT

1Department of Small Animal Clinical Sciences, College of Veterinary Medicine, University of Tennessee, Knoxville, TN, USA; 2University of Illinois, College of Veterinary Medicine, Urbana, IL, USA; 3University of Illinois, Illinois Natural History Survey, Champaign, IL, USA


Hemagglutination inhibition (HI) assay is the sole serologic method currently available to detect exposure to ophidian paramyxovirus (OPMV) in snakes. HI tests are rapid, easy to perform, and do not require species-specific antibodies. The inherent limitations of HI assays are cross-reactivity between similar viruses and moderate specificity, which could result in false-positive samples.1 This could explain the variability in testing found between different OPMV isolates. Three laboratories perform these assays in the United States, each using a different isolate, with one laboratory using two distinct isolates. But to date, no study has evaluated agreement among the assays.

Blood was collected from 26 wild eastern massasauga rattlesnakes (Sistrurus catenatus catenatus) from Illinois in 2006. Each blood sample was divided into three aliquots, packaged similarly, shipped the same day to each laboratory, and assayed using the available OPMV isolate(s) at each laboratory. Findings from these assays included: San Lucan rattlesnake (SLR) isolate 100% positive (26/26), green tree python (GTP) isolate 100% positive (26/26), western diamondback rattlesnake (WDR) isolate 56% positive (14/25), and Aruba Island rattlesnake isolate zero positives (0/26). The kappa statistic for the SLR-GTP isolates demonstrated substantial agreement, the SLR-WDR and GTP-WDR isolates had slight agreement, and the AIR-SLR, AIR-GTP, and AIR-WDR had less than chance agreement. In light of the lack of agreement among the assays, we propose that at present the HI assay is not a reliable indicator of infection or exposure to OPMV in the eastern massasauga rattlesnake.


We gratefully thank the Morris Animal Foundation for support of this project. We thank Dr. Arturo Angelo at the Texas Veterinary Medicine Diagnostic Laboratory, Ms. April Childress at the University of Florida, and Dr. Melissa Kennedy at the University of Tennessee for their assistance in relevant methods descriptions.

Literature Cited

1.  Janeway, C. A., P. Travers, M. Walport, and J. D. Capra. 1999. The induction, measurement, and manipulation of the immune response. In: Immunobiology: The Immune System in Health and Disease. Elsevier Science Ltd., Garland Publishing, New York, NY; 34–48.


Speaker Information
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Matt Allender, DVM, MS
Department of Small Animal Clinical Sciences
College of Veterinary Medicine
University of Tennessee
Knoxville, TN, USA

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