Hepacivirus qPCR

Equine Herpesviruses
Of the currently recognised herpesviruses in horses, EHV1, EHV3 and EHV4 are alpha-herpesviruses
and are of the greatest clinical importance. The gamma-herpesviruses EHV2 and EHV5 may be
associated with ocular and respiratory infections in horses although their importance is relatively
minor.
EHV 1 and 4
EHV-1 infection is widespread and it should be expected that all horses will be at risk of infection
resulting in subclinical infection or respiratory disease. Less frequently abortion (especially last
trimester), neonatal mortality and myeloencephalopathy (typically cauda equina signs in mature
horses) may occur. Most horses are inevitably exposed to EHV1 at some point and long-term latent
infection is then established with the possibility of recrudescence and shedding at times of stress
(e.g. foaling, sales, mixing, training, transport…).
EHV4 is a common cause of relatively mild respiratory disease and also a rare cause of abortion in
mares. Neurologic disease associated with EHV4, if it exists, is very rare indeed.
Diagnosis
Diagnosis is generally made by the presence of suggestive clinical signs (e.g. respiratory disease,
abortion, neurologic disease) alongside supportive laboratory data. As for most infectious diseases,
diagnosis is supported by seroconversion and/or detecting the infectious agent itself.

1. Serology (red top tubes)
   A high antibody titre (>1:80) suggests recent infection, but if the initial sample reveals a low titre
   then a second sample 2-3 weeks later looking for around a 4 x increase in titre indicates recent
   infection. Serologic tests don’t generally distinguish between antibodies to EHV1 and EHV4 although
   an ELISA targeting the C-terminal portion of glycoprotein G of both viruses is available for
   distinguishing serologic response to each virus, although is expensive and not widely available. Both
   the CFT and ELISA tests only reliably detect recent infections (<2-3 months previously) although
   there are some examples of seropositivity from vaccination more than a year previously. Antibody
   titres in CSF are not helpful in diagnosis of myeloencephalopathy due to leakage of serum from
   damaged blood vessels into the CSF.
2. Virus identification (nasal/nasopharyngeal swabs, buffy coat (green or purple top tubes),
   foetus/placenta)
    PCR allows for rapid identification of the presence of EHV1 or 4 infection and is generally
   the preferred method. It is advisable during an outbreak that in-contacts are also
   sampled (especially with signs of pyrexia) to increase the diagnostic rate. It can be
   performed on:
   o nasal/nasopharyngeal swabs indicate presence or absence of current shedding
   although only provides a snapshot of that current time and doesn’t guarantee that
   virus will not be shed later (or before).
   o buffy coat samples from heparinised blood indicate presence or absence of current
   viraemia although only provides a snapshot of that current time and doesn’t
   guarantee that viraemia will not occur later (or before).
   o aborted foetuses (or at least the foetal liver, lung, spleen, adrenal gland and thymus)

o placenta and, where possible, endometrial biopsies
o (CSF is rarely helpful due to such a low viral content)
 Virus isolation can be performed on similar samples to PCR but requires a higher viral
load for positive results and also takes several days. Gauze swabs and viral transport
media are required.
 Histopathology including immunohistochemistry following post mortem examination of
foetuses, placentae, endometrial biopsies, brain and spinal. Gives definitive evidence of
EHV related disease.

EHV 3
This the cause of coital exanthema, a contagious, venereal pustular/vesicular dermatitis affecting the
penis and vulva occasionally seen in breeding animals. Transmission might also occur from insects
and equipment. Signs appear about a week after infection and the clinical disease lasts for 2-3
weeks. Depigmented spots may be left after infection has cleared.
Diagnosis

1. Serology (red top tubes) – Paired samples 2-3 weeks apart looking for around a 4 x increase
   in titre (virus neutralisation test)
2. Virus identification (swabs from lesions) – PCR or Virus isolation
   EHV 2
   Conjunctivitis or keratitis might be seen associated with EHV2 infection. PCR can be performed on
   conjunctival/corneal scrapes or swabs.
   EHV 5
   Interest has been growing recently in the apparent association between EHV5 and the progressive
   inflammatory interstitial lung disease known as equine multinodular pulmonary fibrosis (EMPF).
   Bronchoalveolar lavage samples from many such cases have been found to be positive for EHV5 by
   PCR.