Re-emergent Threat of Equine

Herpesvirus-1 Neurologic Disease

Peter J. Timoney

Department of Veterinary Science

Gluck Equine Research Center
University of Kentucky, Lexington, KY 40546-0099
 Outcomes of EHV-1 Infection in Horses
Abortion

Respiratory

Neonatal Death
Neurological
(EHM)

Ack. Dr. G. Allen (2008)

 Equine Rhinopneumonitis
General features –
► Contagious disease of equids endemic in vast
majority of domesticated equid populations.
► Term encompasses range of syndromes caused by
either EHV-1 or EHV-4.
► Of 5 herpesviruses known to infect the horse, EHV-
1 & EHV-4 are the 2 of greatest veterinary medical
significance.
► Believed EHV-1 / EHV-4 have co-evolved with
horses over millions of years.
► Neither virus of public health significance.
3/14
 EHV-1 and EHV-4 Infections

EHV-1 Infection not only of respiratory tract

epithelium and associated lymphatic
glands but also vascular endothelium
especially of nasal mucosa, lung, adrenal,
thyroid and in the case of some strains,
CNS and endometrium.

EHV-4 Infection restricted primarily to respiratory

tract epithelium and associated lymph
glands. Some strains can set up a
leukocyte-associated viremia.
3/14
 Industry Concerns

► EHV-1 best known for its economic impact

as a cause of contagious abortion
worldwide.

► EHM of concern not only economically but

also from a welfare viewpoint because of
the distressing nature of the disease.

► Lack
of a commercial vaccine of proven
capability to prevent EHM.
3/14
Equine Herpesvirus Myeloencephalopathy

1966 -
First definitive association between EHV-1 and
myeloencephalopathy following isolation of the
virus from brain and spinal cord of a horse with
severe neurologic disease. (Saxegaard, F.,
Nord. Vet. Med., 1966).

3/14
Equine Herpesvirus Myeloencephalopathy
General features –
► Syndrome recorded with increasing frequency over
past 5-10 years, can be associated with high
morbidity & case fatality rate.
► Usually a sequel to a primary respiratory infection,
febrile episode or abortion.
► Can occur in horses of any age, breed or either
gender.
► Nature of illness dependent on location & severity
of lesions in CNS.
► Disease most frequently associated with infection
with neuropathogenic strains of EHV-1.
3/14
 Equine Herpesvirus-1
Myeloencephalopathy

► Many outbreaks of EHM associated with

venues / premises where significant
numbers of horses are congregated
together.

► Conditions at shows, etc, conducive to

respiratory transmission of EHV-1 by direct
/ indirect means.
3/14
Increase in Incidence of Outbreaks of EHV-1
Neurologic Disease, 1970 - 2006
No. of neurologic disease outbreaks
(US and UK) from which virus or
Time interval viral DNA were available
1970 – 75 1
1976 – 80 3
1981 – 85 4
1986 – 90 6
1991 – 95 5
1996 – 2000 6
2001 – 2006 33

Ack: Dr. G.P. Allen

Equine herpesvirus myeloencephalopathy
caused by the hypervirulent, mutant
(neuropathogenic) strain of the virus
designated by USDA a potentially
emergent disease of the horse.

(USDA: APHIS: VS: CEAH: Center for

Emerging Issues Information Sheet,
January 2007)

3/14
Association of Novel Genotype of EHV-1
with Neurologic Disease
► Majority of severe and sometimes
extensive EHM events associated with
novel virus genotype.

► Novel genotype characterised by single

point mutation on catalytic subunit of viral
DNA polymerase.

► Guanine substituted for adenine at position

2254.
3/14
 Nucleotide Substitution in Neuropathogenic Strains
of EHV-1
Replicase
EHV-1 DNA gene

(neutral)
Abortion Strains: -- GTC AAC TAC --
Asparagine

Paralytic Strains: -- GTC GAC TAC -- Aspartic acid

Ack: Dr. G.P. Allen (acidic)
Outbreaks of EHV-1 Neurologic Disease in
USA, 2000 - 2006
--- Genotype of Virus Isolates ---

No. of EHV-1
Time Span CNS Outbreaks Wild-Type Mutant

2000 – 2006 26 2 24

Wild-Type Outbreaks Mutant Outbreaks

• Low neurologic morbidity • High neurologic morbidity
• Low to zero neurologic mortality • High neurologic mortality
Ack: Dr. G.P. Allen
 Clinical Outcome in Relation to Virus
Genotype Involved
► Interms of both neurologic-attack and
case-fatality rates, clinical outcome can
vary depending on genotype of EHV-1.

► Outbreaks caused by G2254 tend to be

more extensive and clinically more severe.

► In comparison, A2254 strains associated

with lower neurologic-attack and case-
fatality rates.
3/14
Equine Herpesvirus Myeloencephalopathy
Characteristics of Vasculitis

► Perivascular cuffing with mononuclear cells and

neutrophils.
► Extensionof inflammatory cells from intima into
media and adventitia of vessel wall.
► Endothelial proliferation and necrosis.
► Necrosis of media.
► Occasionally, thrombin in vessel lumen.
3/14
EHV-1 Paralysis Results from Endothelial
Cell Infection
Spinal Cord Blood Vessel of Paralyzed Horse

EHV-1 infected
endothelial cells

Fibrin thrombus

Inflammatory
lymphocytes

Ack: Dr. G.P. Allen

 Neuropathogenic Strains of EHV-1
Summary of properties –

► Most frequently but not invariably associated

with a single point mutation in the catalytic
subunit of the gene (ORF30) encoding the viral
DNA polymerase gene.
► "Turbo-charged" versions of wild type virus.
► Total
body burden of mutant strains of EHV-1
much greater than wild type virus.
► No evidence of neurotropism.

3/14
 Viremia in Foals after Inoculation with G2254
Mutant or Wild Type Strains of EHV-1
n = 10 foals/group
400
Magnitude of Viremia

Mutant EHV-1
300

200

Wild Type EHV-1

100

0
5 10 15 20
Days Post-Inoculation with EHV-1 Ack: Dr. G.P. Allen
Replicative Capacities of A2254 and G2254
Genotypes of EHV-1

► A2254 and G2254 genotypes differ significantly in

their respective replicative capacities.

► Cell-associated viremia and duration of

respiratory shedding greater in cases of G2254
infection.

► Infectionwith G2254 strains results in vasculitis

in the CNS that is more severe and more
widespread.
3/14
Consequences of Mutation on Pathogenicity
of Genetic Variants of EHV-1

► Enhanced replicative capacity

► Elevated level of viremia

► More widespread vasculitis

► Greater severity of vasculitis

► Greater mortality from neurologic disease

Ack: Dr. G.P. Allen

Clinical Outcome following Infection with
Neuropathogenic Strains of EHV-1

► Infection with G2254 strains may not necessarily

result in development of neurologic disease.

► Individualanimal outcomes can be influenced

by age, innate immunity, acquired immunity,
challenge dose, hormonal status and
environmental factors.

3/14
Evidence that A2254 Nucleotide Substitution not
the Only Determinant of Neuropathogenicity

Report that 24% of the isolates from horses with

neurological disease possessed the A2254 and not
the G2254 genotype (Perkins et al., 2009).

Identification of viruses with nonsynonymous

nucleotide substitutions in ORF30 besides A2254 to
G2254 from horses without signs of neurologic
disease (Smith et al., 2010).
(continued)

Ack: Dr. U. Balasuriya (2011)

Evidence that A2254 Nucleotide Substitution not
the Only Determinant of Neuropathogenicity
(continued)

Sequence analysis of EHV-1 field strains has

identified other mutations outside of the small region
of ORF30 sequenced by Nugent et al. (2006).

Mutations found in same gene or genes encoding

proteins of viral elongation complex or viral envelope
proteins.

Ack: Dr. U. Balasuriya (2011)

 Factors Involved in the Epidemiology of
Equine Herpesvirus Myeloencephalopathy

► Virus strain.

► Modes of transmission.

► Immune status of individual animals / groups of

horses.
► Existence of the carrier state.

► Various management practices.

3/14
Ack: Dr. G.P. Allen
 EHV-1 and EHV-4 Infections
Latency –

► Latency of EHV-1, EHV-4 widespread (40-60%) in

adult equids.
► Individual animals may be carriers of one or both
viruses.
► Sites of latency of EHV-1 / EHV-4: lymphoreticular
tissues associated with the respiratory tract,
circulating CD3+ lymphocytes, and the trigeminal
ganglia (EHV-1).

(continued)
3/14
 EHV-1 and EHV-4 Infections
Latency (cont.) –

► Carrier state probably life-long.

► No infectious virus present unless latent virus has
been reactivated.
► Latent
virus can be reactivated by environmental /
pharmacological stimuli.

3/14
 Expansion in the Reservoir Size of the Latent G2254
Neuropathogenic EHV-1 Strains in Kentucky TB Mares

20%

n = 450 abortion isolates of EHV-1

Mutant Strain of EHV-1
(% of Total Isolates)

15%

10%

5%

1960’s 1970’s 1980’s 1990’s 2000’s

Decade
Smith, K. 2007. Master’s Thesis. University of Kentucky Ack: Dr. G.P. Allen
 Prevalence of Latent, G2254 Neuropathogenic
Strains of EHV-1 in TB Mare Population of Kentucky
Sub-maxillary lymph nodes collected from 132 necropsied TB
mares.
Tested for latent EHV-1 DNA by PCR.
46% of tested mares harbored latent wild-type EHV-1 DNA.
8% of tested mares harbored G2254, neuropathogenic
strains of EHV-1 (=18% of total latent reservoir of EHV-1).

8%
M

46%
EHV-1 DNA in
WT
SMLN tissue
of TB mares

132 TB broodmares Ack: Dr. G.P. Allen

 Development of Equine Herpesvirus
Myeloencephalopathy
Risk factors –

► Existinglevels of cytotoxic T-lymphocyte precursor

(CTLP) cells specific for EHV-1 critically important.

► Significantly greater risk in elderly horses (≥ 20 y.o.).

► Significantly
greater risk in horses exposed to
ORF30G2,254genotype of EHV-1.

► No significant correlation with pre-exposure levels of

serum neutralising antibodies to EHV-1).

(G.P. Allen, AJVR, 69:1595-1600, 2008) 3/14

Relationship Between EHV-1 Cellular Immunity
and Viremic Load
Viremic Load (Log10)

Cellular Immunity
(Pre-Infection CTLp Frequency per million PBMC)
Ack: Dr. G.P. Allen
Dr. Roger Doll, 1960’s
Equine Herpesvirus Myeloencephalopathy
Key points –

► One of 5 clinical syndromes caused by EHV-1

and infrequently, certain strains of EHV-4.
► An emergent disease of increasing veterinary
medical and economic significance since
2000.
► Usually a sequel to a primary herpesvirus
respiratory infection, febrile illness or abortion.
► Can occur in horses of any age, breed or
either gender.
(continued) 2/12
Equine Herpesvirus Myeloencephalopathy
Key points (cont.) –

► Nature of illness depends on location and

severity of lesions in CNS.
► Majority of outbreaks caused by hypervirulent,
neuropathogenic (mutant) strains of EHV-1.
► Neuropathogenic EHV-1 strains give rise to
much greater body burdens of virus than wild
type virus.
► Neuropathogenic EHV-1 strains cause higher
morbidity and case-fatality rates.
(continued)
2/12
Equine Herpesvirus Myeloencephalopathy
Key points (cont.) –

► Evidence of increasing prevalence of latent

infection with neuropathogenic strains of EHV-1.
► Risk factors associated with development of
EHM:
 Age (≥ 20 years old).
 Infection with neuropathogenic strain of EHV-1.
 Level of CTLP cells specific for EHV-1.

► Very doubtful current vaccines effective in

preventing EHM.
2/12