Development of a Quantitative PCR Assay to Evaluate the Clinical Significance of Tursiops truncatus Herpesvirus 3
abstract
Herpesvirus infections are being reported with increased frequency in cetaceans. Currently, three alphaherpesviruses and one gammaherpesvirus have been reported in bottlenose dolphins. Herpesviruses are often host-restricted and tend to co-evolve with their host.1 Most vertebrate species investigated have yielded at least one herpesvirus, if not several.1 Although bottlenose dolphin herpesvirus 3 (TtHV-3) had originally been found in a skin lesion2, the clinical significance of this alphaherpesvirus in a clinically healthy bottlenose dolphin is unknown.
TtHV-3 was identified in the white blood cells (WBC) of a clinically healthy bottlenose dolphin using nested consensus polymerase chain reaction (PCR).3 Alphaherpesviruses are known to establish latency in the sensory ganglia1, so detection of virus in circulating WBC is indicative of active virus replication and infection. Therefore, a specific quantitative PCR (qPCR) assay was developed as a tool for determining the clinical significance of this finding. A series of 19 WBC samples, collected from the dolphin between October 2005 and December 2007, were analysed with qPCR. The qPCR assay was found to be more sensitive (detection limit: 10 virus copies/sample), robust and faster (up to 66 samples/day) than the nested consensus PCR. The association of circulating virus titers with clinical signs was explored using a logistic regression model. In addition, 55 buffy coat samples from clinically healthy dolphins were collected randomly between July and September 2007 and analyzed to establish the baseline prevalence of circulating TtHV-3 in a healthy population of collection animals.
References
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2. Smolarek Benson KA, Manire CA, Ewing RY, Saliki JT, Townsend FI, Ehlers B, Romero CH. 2006. Identification of novel alpha- and gammaherpesviruses from cutaneous and mucosal lesions of dolphins and whales. Journal of Virological Methods 136: 261-266.
3. VanDevanter DR, Warrener P, Bennett L, Schultz ER, Coulter S, Garber RL, Rose TM. 1996. Detection and Analysis of Diverse Herpesviral Species by consensus Primer PCR. Journal of Clinical Microbiology 34(7): 1666-1671