Development and Validation of a Novel Quantitative Polymerase Chain Reaction Assay for Measurement of Delphinid Herpesvirus-4 in the Bottlenose Dolphin (Tursiops truncatus)
Abstract
Delphinid herpesvirus 4 (DeHV-4) has been implicated as one potential etiology for orogenital papillomatosis of adult bottlenose dolphins (Tursiops truncatus) in both wild and managed populations.1,2,3 Oral papillomas in bottlenose dolphins have been suggested to undergo malignant transformation into oral squamous cell carcinoma, with potential for metastasis to local lymph nodes and systemic disseminated disease.1,3 Current diagnosis of DeHV-4 is achieved through a standard nested polymerase chain reaction (nPCR) and sequencing. TaqMan assays, with probe hybridization, represent a more efficient option for herpesvirus identification and quantification. This study developed a quantitative PCR (qPCR) assay using a TaqMan probe-based assay, targeting the highly conserved polymerase region of the DeHV-4 genome. The qPCR assay was optimized and validated using orogenital and dermal tissue samples of bottlenose dolphins known to be DeHV-4 positive and negative based on nested polymerase reaction assays and sequencing, as well as samples known to be positive for other herpesvirus species. This optimized and validated qPCR assay can be used as a tool for further research into DeHV-4 and its potential role in oral papillomatosis in bottlenose dolphins.
Acknowledgements
The authors wish to thank Dr. Judy St. Leger of SeaWorld Parks and April Childress of the University of Florida, College of Veterinary Medicine.
* Presenting author
+ Student presenter
Literature Cited
1. Bossart GD, Ghim SJ, Rehtanz M, Goldstein J, Varela R, Ewing RY, Fair PA, Lenzi R, Joseph B, Hicks CL, Schneider LS, Mckinnie CJ, Reif JS, Sanchez R, Lopez A, Novoa S, Bernal J, Goretti M, Rodriguez M, Defran RH, Jenson AB. 2005. Orogenital neoplasia in Atlantic bottlenose dolphins (Tursiops truncatus). Aquatic Mammals. 31;4:473–480.
2. Van Elk CE, Van De Bildt MWG, De Jong AAW, Osterhaus ADME, Kuiken T. 2009 Genital herpesvirus in bottlenose dolphins (Tursiops truncatus): cultivation, epidemiology, and associated pathology. Journal of Wildlife Diseases. 45;4:895–906.
3. Smolarek Benson KA, Manire CA, Ewing RY, Saliki JT, Townsend FI, Ehlers B, Romero CH. 2006. Identification of novel alpha- and gamma herpesviruses from cutaneous and mucosal lesions of dolphins and whales. Journal of Virological Methods. 136;1–2:261–266.