Kathryn Smith, a graduate student in the laboratory of Udeni Balasuriya, PhD, MS, BVSc, professor at the University of Kentucky Gluck Equine Research Center, has developed and validated a new real-time polymerase chain reaction (PCR) assay. This assay can detect and discriminate between neuropathogenic (causing or capable of causing disease of nervous tissue) and non-neuropathogenic equine herpesvirus-1 (EHV-1) strains in nasal swab samples.

All EHV-1 strains can potentially induce respiratory disease as well as abortion in mares, but only specific neuropathogenic strains can cause large-scale outbreaks of neurologic disease. Several years ago, researchers linked a single nucleotide mutation in viral DNA polymerase encoding gene (open reading frame 30 [ORF30]) to the neuropathogenic type of EHV-1. Identifying this particular mutation led to the Gluck Center's late George Allen, PhD, to develop a real-time PCR assay (E2) to diagnose EHV-1 infection in horses. The PCR assay (E2) distinguishes between potential neuropathogenic and non-neuropathogenic EHV-1 strains and identifies any viral DNA in the horse's nasal secretion revealing an active virus shedding. However, the assay was found to yield false negative results and, thus, the sensitivity was not as high as desired for detecting low amounts of virus particles in clinical samples.

"Now we have developed and validated an improved and more reliable new real-time PCR assay for diagnosing EHV-1," Balasuriya said.

The new and improved real-time PCR assay (E1) has been evaluated and compared to E2 assay using 76 archived EHV-1 isolates and 433 clinical specimens from cases of suspected EHV-1 infection. Scientists have developed the new assay (E1) by redesigning new primers and probes to detect and differentiate neuropathogenic and non-neuropathogenic EHV-1 strains, said Balasuriya.

Balasuriya says the E1 is 10 times more sensitive than the E2 and can detect down to 10 virus particles in one sample. The comparisons of the two tests indicate that E2 lacks adequate sensitivity for routine diagnostic applications and consequently generates more false negative results.

"The greater sensitivity and accuracy for detection of EHV-1 makes this improved test an important diagnostic tool to be used during outbreaks of EHV-1," Balasuriya said. "A rapid reliable diagnosis is crucial in order to ensure biosecurity and immediate quarantine measures when an outbreak occurs."

Of all the infectious viral diseases, EHV-1 is the one considered to be the most costly to the equine industry, giving rise to large-scale outbreaks and losses every year. EHV-1 can cause a variety of clinical signs including acute upper respiratory disease with fever, abortion, and neurologic disease. Most horses have been exposed to EHV-1 early in life and consequently establish a latent infection, sporadically shed virus, or exhibit only a few clinical signs. They become silent carriers showing no clinical signs of illness but can potentially cause new infections in foals and young horses. What makes EHV-1 tricky is the virus' unique ability to stay dormant and reappear later when stressed horses might shed infectious virus particles.

Shaila Sigsgaard is a contributing writer for the Bluegrass Equine Digest.

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