Full text of DOI:10.1080/00480169.2002.36300

New Zealand Veterinary Journal
ISSN: 0048-0169 (Print) 1176-0710 (Online) Journal homepage: http://www.tandfonline.com/loi/tnzv20
Equine respiratory viruses in foals in New Zealand
M Dunowska , CR Wilks , MJ Studdert & J Meers
To cite this article: M Dunowska , CR Wilks , MJ Studdert & J Meers (2002) Equine respiratory
viruses in foals in New Zealand, New Zealand Veterinary Journal, 50:4, 140-147, DOI:
10.1080/00480169.2002.36300
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140
New Zealand Veterinary Journal 50(4), 140-147, 2002
Scientific Article
Equine respiratory viruses in foals in New Zealand
M Dunowska*§, CR Wilks*†, MJ Studdert‡ and J Meers*#
and 16/23 foals showed serological evidence of recent EAdV-1
infection. None of the 67 serum samples tested were positive
for antibodies to EAV, reovirus 3 or PIV3. There was no clear
association between infection with any of the viruses isolated or
tested for and the presence of overt clinical signs of respiratory
disease.
Abstract
AIMS: To identify the respiratory viruses that are present
among foals in New Zealand and to establish the age at which
foals first become infected with these viruses.
METHODS: Foals were recruited to the study in October/
November 1995 at the age of 1 month (Group A) or in March/
April 1996 at the age of 4–6 months (Groups B and C). Nasal
swabs and blood samples were collected at monthly intervals.
Nasal swabs and peripheral blood leucocytes (PBL) harvested
from heparinised blood samples were used for virus isolation;
serum harvested from whole-blood samples was used for se-
rological testing for the presence of antibodies against equine
herpesvirus (EHV)-1 or -4, equine rhinitis-A virus (ERAV),
equine rhinitis-B virus (ERBV), equine adenovirus 1 (EAdV-1),
equine arteritis virus (EAV), reovirus 3 and parainfluenza virus
type 3 (PIV3). Twelve foals were sampled until December 1996;
the remaining 19 foals were lost from the study at various times
prior to this date.
CONCLUSIONS: There was serological and/or virological
evidence that EHV-1, EHV-2, EHV-5, EAdV-1 and ERBV
infections were present among foals in New Zealand. EHV-2
infection was first detected in foals as young as 3 months of age.
The isolation of EHV-2 from nasal swabs preceded serological
evidence of infection with other respiratory viruses, suggesting
that EHV-2 may predispose foals to other viral infections.
KEY WORDS: Equine respiratory disease, equine herpesviruses,
equine rhinitis viruses, equine adenovirus, infection, foal, horse
Introduction
Several viruses including equine influenza virus, EHV-1, EHV-4, ERAV
(formerly equine rhinovirus type 1), ERBV (formerly equine rhino-
virus type 2), reoviruses, PIV3, and EAV have been associated with
respiratory disease in horses (Mumford and Rossdale 1980; Jolly et
al 1986; Powell 1991; Mair 1996). The results of a recent virologi-
cal and serological survey of horses sampled following outbreaks of
respiratory disease, and yearlings from yearling sales, indicated that
EHV-2 and possibly ERBV might be involved in equine respiratory
disease in New Zealand (Dunowska et al 2002).
RESULTS: The only viruses isolated were EHV-2 and EHV-5.
EHV-2 was isolated from 155/157 PBL samples collected dur-
ing the period of study and from 40/172 nasal swabs collected
from 18 foals. All isolations from nasal swabs, except one, were
made over a period of 2–4 months from January to April (Group
A), March to April (Group B) or May to July (Group C). EHV-5
was isolated from either PBL, nasal swabs, or both, from 15
foals on 32 occasions. All foals were positive for antibodies to
EHV-1 or EHV-4, as tested by serum neutralisation (SN), on
at least one sampling occasion and all but one were positive for
EHV-1 antibodies measured by enzyme-linked immunosorbent
assay (ELISA) on at least one sampling occasion. Recent EHV-1
infection was evident at least once during the period of study in
18/23 (78%) foals for which at least two samples were collected.
SN antibodies to ERBV were evident in 19/23 (83%) foals on
at least one sampling occasion and 15/23 foals showed evidence
of seroconversion to ERBV. Antibodies to ERAV were only de-
tected in serum samples collected from foals in Group A and
probably represented maternally-derived antibodies. Haemag-
glutination inhibition (HI) antibody titres ≥1:10 to EAdV-1were
evident in 21/23 (91%) foals on at least one sampling occasion
It is generally accepted that foals are protected from viral in-
fections soon after birth by maternally-derived antibodies and
subsequently become infected when these maternal antibodies
decline at approximately 4–6 months of age (Allen and Bryans
DNA
Deoxyribonucleic acid
EAdV-1 Equine adenovirus 1
EAV
EHV
ELISA
ERAV
ERBV
H&E
HI
Equine arteritis virus
Equine herpesvirus
Enzyme-linked immunosorbent assay
Equine rhinitis-A virus
Equine rhinitis-B virus
Haematoxylin and eosin
Haemagglutination inhibition
Peripheral blood leucocyte(s)
Phosphate buffered saline, pH 7.0
Polymerase chain reaction
Parainfluenza virus type 3
Red blood cell(s)
* Institute of Veterinary, Animal and Biomedical Sciences, Massey University,
New Zealand
† Current address: School of Veterinary Science, University of Melbourne,
Parkville VIC 3052, Australia
‡ Centre for Equine Virology, University of Melbourne, Parkville VIC 3052,
Australia
# Current address: University of Queensland, St Lucia QLD 4072, Australia
§ Author for correspondence. Current address: Department of Pathology,
Colorado State University, Fort Collins CO 80523, United States of America.
Email: mdunowsk@lamar.colostate.edu
PBL
PBS
PCR
PIV3
RBC
RK13
SN
Rabbit kidney cell line
Serum neutralisation
Vero
African green monkey (cells)

Dunowska et al
New Zealand Veterinary Journal 50(4), 2002
141
1986; Powell 1991). However, some recent investigations have
shown that infections can occur earlier than 4–6 months of age,
often in the presence of maternal antibody. EHV-1 and EHV-2
infections have been described in foals as young as 30 days of age
(Fu et al 1986; Gilkerson et al 1999) and EAdV-1 was isolated
from a healthy Clydesdale foal at 3 days of age (Wilks and Stud-
dert 1972).
volved frequent (twice weekly) handling from an early age. Their
dams had been vaccinated with inactivated EHV-1/EHV-4 vac-
cine during pregnancy (Donald 1998).
Group B
Foals from Group B (B1 to B19) belonged to a large Thorough-
bred breeding farm in the Manawatu region. The foals were born
in October–December 1995 and samples were first collected in
March–April 1996, at the time when the foals were brought in
for weaning and branding at about 4–6 months of age. Samples
were collected from 19 foals during the first visit in March. Out
of these 19 foals, only 11 remained on the farm and were sampled
on the second sampling occasion. Four more foals were lost from
the study due to transfer to another farm before the last sampling
in December 1996.
It is useful to know the age at which foals first become at risk
from viral infection for successful prophylaxis, especially by vac-
cination. The purpose of this investigation was to establish which
respiratory viruses were present among foals and at what age foals
first became infected under New Zealand conditions.
Group C
Materials and Methods
Group C comprised five foals (C1 to C5) from a small stud in the
Manawatu region that bred both Thoroughbred and Standard-
bred horses. Foals C1 and C2 were Thoroughbreds and Foals C3,
C4 and C5 were Standardbreds. Foal C1 was born in late October
and the remainder were born in November 1995. The foals were
branded in March 1996, before the first sampling in April 1996.
Foals C3 and C5 were weaned in April 1996 before the first sam-
pling, and Foals C1, C2 and C4 were weaned in May at the time
of the second sampling.
Origin of samples
Three groups of foals were sampled on a monthly basis. The dates
of samplings and numbers of foals sampled on each occasion are
shown in Table 1. Handlers were asked to observe foals for the
clinical signs of respiratory disease such as nasal discharge, cough
or lack of appetite.
Group A
Group A comprised seven Thoroughbred foals (A1 to A7) that
were kept on a Massey University experimental farm in the Mana-
watu region in the North Island of New Zealand. The foals were
born in October–November 1995 and samples were first collected
within the foals’ first month of life. Foals were weaned at the end
of March and branded in April 1996. Foals were sampled until
December 1996. Six foals were sold after weaning, hence only
one was available for sampling in November and December 1996.
These foals were also utilised for another experiment, which in-
Collection of samples
Samples collected from each foal during each visit included a
nasal swab (Virocult, Medical Wire and Equipment Co Aus-
tralasia Pty Ltd), and blood. Blood samples were collected by
jugular venepuncture into plain and heparinised vacutainers for
the collection of serum and PBL, respectively, with the exception
of the first two samplings of Group A foals. From this group of
foals only nasal swabs and blood for serology were collected in
Table 1. Sample collection dates, average ages (months) at sampling and numbers (n) of foals sampled for each of the three study groups.
Group A
n
Group B
n
Group C
n
Date
Age
Date
Age
Date
Age
31/10/95
or 7/11/95
7
7
7
7
7
<1
1
27/11/95
9/01/96
9/02/96
6/03/96
3
4
5
29/03/96
a
or 3/04/96
19
5
a
a
3/04/96
6
3
6
7
29/04/96
25/05/96
11
10
6
7
3/04/96
5
5
5
5
5
5
4
4
5
6
a
7/05/96
16/05/96
10/06/96
16/07/96
15/08/96
14/10/96
14/11/96
16/12/96
7
2/07/96
15/08/96
11/10/96
8/11/96
2
2
2
1
1
9
10/07/96
20/08/96
14/10/96
18/11/96
17/12/96
9
9
9
8
7
8
8
10
12
13
14
9
9
11
12
13
11
12
13
12/12/96
a
First sampling after weaning

142
New Zealand Veterinary Journal 50(4), 2002
Dunowska et al
October and November 1995. Virus isolation from PBL was first
attempted from blood samples collected at the third sampling of
this group, in January 1996.
reference negative serum. Foals were regarded as recently infected
with EHV-1 if an increase in blocking activity from <60% to
>60% or if a 4-fold rise in blocking activity >60% was evident
using ELISA. Serological evidence for EHV-4 infection could not
be obtained, because EHV-4-specific antibody status could not
be determined using these methods in horses positive for EHV-1
antibodies. Serological testing for antibodies against EHV-2 and
EHV-5 was not performed.
All procedures involving the experimental use of animals were
approved by the Massey University Animal Ethics Committee
(Palmerston North, New Zealand).
Processing of samples
Samples were transferred to the laboratory within 2–6 h after col-
lection and processed. Blood samples were allowed to clot at 4°C
overnight, serum separated by centrifugation at 2,000 g for 20
min, and stored in aliquots at –70°C. Nasal swab samples were
processed according to the manufacturer’s instructions, filtered
and either inoculated on to monolayer cell cultures or stored at
–70°C. PBL were separated from heparinised blood as described
by Gleeson and Coggins (1985). Briefly, buffy-coat-rich plasma
was collected from blood that had been allowed to settle for 10–
20 min at room temperature and incubated with an equal volume
Antibodies to ERAV and ERBV were detected using SN as-
says that used ERAV.393/76 (Studdert and Gleeson 1977) and
ERBV.P1436/71 (Steck et al 1978), respectively, as reference
viruses. Test sera were regarded as positive if they showed titres
>1:100 (for ERAV) or >1:2 (for ERBV). Samples collected from
foals in Group A that were negative for antibodies to ERAV at a
dilution of 1:100 were retested at a dilution of 1:10. Antibodies
to PIV3 and EAdV-1 were detected using HI assays that used a
commercially available human PIV3 strain (Denka Seiken Co
Ltd, Japan) and a New Zealand EAdV-1 isolate (courtesy of GW
Horner, National Centre for Disease Investigation, Wallaceville,
New Zealand) as antigens, and 0.6% guinea pig RBC or 0.5%
human type-O RBC suspension, respectively, as indicator sys-
tems. The tests were conducted according to standard laboratory
procedures using 4 haemagglutination units of virus antigen per
well (Lennette et al 1988). The tests were conducted at room
temperature for PIV3 or at 37°C for EAdV-1. Non-specific in-
hibitory activity in serum samples was minimised by trypsin and
periodate treatment (Hsiung 1982) and non-specific agglutinins
were removed by prior adsorption with a 50% suspension of the
respective RBC for 1 h. Test sera were regarded as positive if the
titre was >1:10. A complement fixation test was used to detect an-
tibodies against reovirus 3, using commercially obtained human
reagents, according to the manufacturer’s instructions (Denka
Seiken Co Ltd, Japan). Antibodies to EAV were tested for using
a SN assay, and titres >1:4 were regarded as positive. Foals were
regarded as recently infected with ERAV, ERBV, EAdV-1, reovi-
rus 3 or EAV if there was a 4-fold rise in antibody titre between
successive samples.
of red blood cell (RBC) lysing buffer (0.85% NH Cl, 0.017M
4
Tris, pH 7.4) for 5–10 min. Cells were pelleted by centrifugation
at 250 g for 10 min, washed once in phosphate buffered saline
(PBS), pH 7.0, and resuspended in 2 ml of PBS.
Virus isolation and detection
A 200 μl quantity of resuspended PBL or nasal swab filtrate was
inoculated on to each of three cell cultures: rabbit kidney (RK13),
Vero and primary equine foetal kidney monolayers. Viruses were
initially identified based on cytopathic effect produced in cell
culture. Isolated herpesviruses were typed using a virus-specific
polymerase chain reaction (PCR) procedure (Dunowska et al
1999, 2002). PCR using primers specific for EHV-1 and EHV-4
was also performed on all PBL cultures on equine foetal kidney
cells, PBL cultures on RK13 cells from samples collected from
foals at the time they showed serological evidence of EHV-1 or
EHV-4 infection, all cell cultures positive for EHV-5, and directly
on all 172 nasal swab filtrates. Additionally, electron microscopy
of clarified cell culture lysates (pelleted at 100,000 g for 2 h),
and examination of monolayers stained with haematoxylin and
eosin (H&E) were performed on several occasions. Cell culture
lysates from inoculated Vero and RK13 cells were also tested for
haemagglutination activity using human type-O RBC (for Vero
cell cultures) or guinea pig RBC (for RK13 cell cultures), to test
for the presence of reovirus 3 and PIV3, respectively, that do not
produce an obvious cytopathic effect.
Results
Signs of respiratory disease
Except for two foals that showed a slight mucopurulent nasal
discharge on one sampling occasion in March or August 1996, all
other foals in Group A remained healthy throughout the period
of study. Foals in Group B showed clinical signs of respiratory dis-
ease during April and May. However, clinical data on individual
foals were not available. Foals in Group C showed nasal discharge
of varying severity in May and June. The two fillies (Foals C1
and C4) seemed to be more markedly affected than the colts
(Foals C2, C3 and C5), Foal C4 showing a white, copious nasal
discharge at the time of sampling in June. Also, Foals C1 and C2
showed slight nasal discharge at the time of the sixth sampling, in
October 1996.
Serology
All serological tests were performed at the same time, after all
samples had been collected. For each serological test, paired serum
samples from individual horses were processed preferentially on
the same plate or at least in the same batch to minimise between-
test variation. All serum samples were tested in duplicate. Testing
for EHV-1 SN, EHV-1 ELISA, ERAV, EAdV-1, and ERBV was
performed on 151, 164, 163, 163, and 162 sera, respectively,
from 23 foals. Testing for antibodies against EAV, reovirus 3 and
PIV3 was performed on 67 serum samples from 23 foals.
The SN test for antibodies to EHV-1 or EHV-4 used EHV-1
Durham (Horner 1981) as the reference virus. Sera were regarded
as positive to EHV-1 or EHV-4 if they showed a titre >1:2. Foals
were regarded as recently infected with EHV-1 or EHV-4 if they
showed a 4-fold rise in the EHV-1 SN titre. Type-specific an-
tibodies to EHV-1 were detected using a blocking ELISA (van
de Moer et al 1993; Donald 1998). Samples were regarded as
positive if they showed >60% blocking activity compared with a
Virus isolation and detection
The only viruses isolated were EHV-2 and EHV-5 (Figure 1).
EHV-2 was first isolated from either PBL or nasal swab samples
from foals as young as 3 months (Group A) or 4–6 months
(Groups B and C) of age (Figure 1). EHV-2 was isolated from
155/157 (99%) of the PBL samples tested and from 40/172 nasal
swabs from 18 foals. All isolations from nasal swabs, except one,

Dunowska et al
New Zealand Veterinary Journal 50(4), 2002
143
Figure 1. Viruses isolated from nasal swabs ( ■ ), peripheral blood leucocytes (PBL) ( ■ ) or both nasal swabs and PBL ( ■ ) from foals sampled on
a
a monthly basis: (a) Group A; (b) Group B, and; (c) Group C. Numbers above the x axis indicate numbers of foals sampled on each occasion. Virus
b
isolation was attempted from nasal swabs only; Virus isolation was attempted from PBL from four foals only.
were made over a period of 2–4 months from January to April
(Group A), March to April (Group B) or May to July (Group C)
(Figure 1). EHV-2 was not isolated from subsequent nasal swab
samples but continued to be isolated from PBL. EHV-5 was iso-
lated from either PBL or nasal swabs, or both, of 15 foals on 32
occasions. With one exception, all samples that were positive for
EHV-5 were also positive for EHV-2. All samples tested for the
presence of EHV-1 and EHV-4 using PCR, including 172 nasal
swab filtrates, yielded negative results.
at least one sampling occasion and all but one were positive for
EHV-1-specific antibodies measured by ELISA on at least one
sampling occasion. Serological evidence of recent EHV-1 infec-
tion was present at least once during the period of study in 18/23
(78%) foals from which at least two blood samples were collected.
Serological evidence of recent EHV-1 infection was first detected
between May and August in all 3 groups. Foals B8, B13 and
B19 showed serological evidence of EHV-1 or EHV-4 infections
twice during the period of study, 5–7 months apart (Figure 3).
Three foals (A3, B1 and B19) showed serological evidence of re-
cent EHV-1 infection without a corresponding 4-fold rise in the
EHV-1 SN titres (Figures 2 and 3). Foals A1 and A4 showed a rise
in ELISA titres without reaching the 60% cut-off value (from 7 or
Serology
The results of serological testing are presented in Figures 2–4.
All foals were positive for SN antibodies to EHV-1 or EHV-4 on

144
New Zealand Veterinary Journal 50(4), 2002
Dunowska et al
14% to 57 or 49%, respectively) at the last sampling in April. Foal
B16 showed a 4-fold rise in the EHV-1 SN titre, while its ELISA
titre remained below the 60% cut-off value, possibly indicating
EHV-4 infection (Figure 3).
Antibodies to ERAV were only detected in serum samples col-
lected from foals from Group A (Figure 2). In this group, 3/7
foals had high ERAV SN antibody titres at their first sampling,
in October–November. The sera of the remaining four foals were
negative at a 1:100 dilution and were retested at a 1:10 dilution,
at which 3/4 remained negative and 1/4 showed a titre of 1:40.
During the period November to March, the titres of Foals A1,
A2 and A6 gradually declined, indicating that the antibodies
were probably maternally derived. Antibodies to ERAV were not
detected in any of the serum samples collected from the foals in
April and all the foals sampled remained negative until the last
sampling in December.
SN antibodies to ERBV were evident in 19/23 (83%) foals on at least
one sampling occasion and 15/23 foals showed evidence of serocon-
version to ERBV during the study period. All foals in Group A had
SN antibodies to ERBV at the first sampling in October–November
(Figure 2). Initial titres ranged from 1:8–1:128. ERBV antibody titres
gradually declined during the following months, suggesting that the
antibodies detected were probably maternally derived. Subsequently,
between February and April, 3/7 foals in Group A showed a rise in
ERBV antibody titres, possibly indicating recent infection. Most of
the foals from Groups B and C were negative for ERBV antibodies at
the first sampling in March–April. During the following months, 9/11
(82%) foals from Group B seroconverted to ERBV, titres ranging from
1:4–1:128 (Figure 3). In Group C, 3/5 foals showed a rise in titre to
ERBV from <1:2 to 1:4 (Figure 4). None of the foals from this group,
however, developed an ERBV titre greater than 1:4 during the study
period, except for Foal C5 that had a titre of 1:16 in April and May.
Haemagglutination inhibition (HI) antibody titres to EAdV-1
≥1:10 were evident in 21/23 (91%) foals on at least one sampling
occasion and 16/23 foals showed serological evidence of recent
EAdV-1 infection. (Figures 2–4). None of the 67 serum samples
tested were positive for antibodies to EAV, reovirus 3 or PIV3.
There was no clear association between infection with any of the
viruses isolated or tested for and the presence of overt clinical
signs of respiratory disease.
Discussion
This study provides serological and/or virological evidence of EHV-
1, EHV-2, EHV-5, EAdV-1 and ERBV infection in all three groups
of foals studied. The prevalences of EHV-1 SN antibodies (100%)
and ERBV SN antibodies (83%) were higher than those reported
in an earlier study conducted in New Zealand, in which 67% of five
11-month-old foals were positive for SN antibodies to EHV-1 and
41% were positive for SN antibodies to ERBV (Jolly et al 1986).
The higher prevalences evident in our study probably reflect the fact
that we sampled foals over a period of several months, whereas the
prevalences reported by Jolly et al (1986) were calculated from single
serum samples. All foals were positive for EHV-2, as determined by
virus isolation, at least once during the current study period. Similar
results have been reported by others (Fu et al 1986; Murray et al
1996). Other viruses, including ERAV, reovirus 3 and PIV3, were
not detected among the foals included in this study, which corre-
sponds with previous findings (Jolly et al 1986).
Figure 2. Results of serological testing for the presence of (a) serum
neutralising (SN) and (b) ELISA antibodies to equine herpesvirus 1
(EHV-1); (c) SN antibodies to equine rhinitis-A virus (ERAV) and; (d)
equine rhinitis-B virus (ERBV); and (e) haemagglutination inhibition
(HI) antibodies to equine adenovirus 1 (EAdV-1) in foals in Group A.
Titres of individual foals (A1 through A7) are shown, expressed as
reciprocal serum dilutions. Consecutive bars represent consecutive
months from October 1995 through to December 1996. The rst
sampling in October-November is regarded as an October sampling.
Speci c dates of each sampling are shown in Table 1.

Dunowska et al
New Zealand Veterinary Journal 50(4), 2002
145
Figure 3. Results of serological testing for the presence of (a) serum neutralising (SN) and (b) ELISA antibodies to equine herpesvirus 1 (EHV-1);
(c) SN antibodies to equine rhinitis-B virus (ERBV) and; (d) haemagglutination inhibition (HI) antibodies to equine adenovirus 1 (EAdV-1) in foals in
Group B. Titres of individual foals (B1 through B19) are shown, expressed as reciprocal serum dilutions. Consecutive bars represent consecutive
months from October 1995 through to December 1996. The rst sampling in March-April is regarded as a March sampling. Speci c dates of each
sampling are shown in Table 1.

146
New Zealand Veterinary Journal 50(4), 2002
Dunowska et al
Despite serological evidence of infection with other viruses, only
EHV-2 and EHV-5 were isolated from the nasal swabs and PBL
samples collected in our study. It is possible that viruses other
than EHV-2 and EHV-5 were not present in the samples collect-
ed. The success of virus isolation depends to a large extent on the
timing of sampling in relation to infection, and many infections
are only diagnosed retrospectively from serology results. The fact
that none of the nasal swab filtrates tested directly for the presence
of EHV-1 or EHV-4 DNA using PCR methods yielded positive
results supports the view that neither EHV-1 nor EHV-4 were
present in the samples collected.
There are several possible explanations for failure to isolate viruses
other than EHV-2 or EHV-5 even if foals were shedding them at
the time of sampling. These include collection of nasal rather than
nasopharyngeal swabs, suboptimal processing of samples or cell
culture conditions, or interference between herpesviruses present
in the samples with the ability of other viruses to grow in vitro.
The inhibitory effect of EHV-2 on the propagation of EHV-1 in
cell culture has been reported by others (Dutta et al 1986; Welch
et al 1992). The cell culture conditions used in our study may
not have been optimal for the growth of rhinitis viruses. Despite
serological evidence of their presence, isolation of rhinitis viruses
has never been reported in New Zealand, suggesting that isola-
tion of these viruses in cell culture may be difficult. However,
19 and 28 ERBV isolates obtained during studies conducted in
America (Carman et al 1997) and Switzerland (Steck et al 1978),
respectively, were isolated using standard cell culture techniques.
It is also possible that rhinitis viruses may have been replicating
in the cell culture but not producing any visible cytopathic effect,
as has been demonstrated by others (Li et al 1997).
In all three groups of foals, the isolation of EHV-2 and EHV-5
from nasal swabs preceded serological evidence of infection with
other respiratory viruses. This is consistent with the notion that
infection with EHV-2 and EHV-5 predisposes horses to infection
with other viruses. Several recent investigations have provided
indirect evidence that the role of EHV-2 and possibly EHV-5
in causing respiratory disease in horses may have been underes-
timated in the past. A significantly greater rate of EHV-2 isola-
tion from tracheal aspirates of foals showing signs of respiratory
disease compared with healthy foals was reported by Murray et
al (1996). Borchers et al (1997) showed that PBL samples from
70% of horses showing signs of respiratory disease were positive
for EHV-2, whereas samples from only 42% of healthy horses
were positive. In a further study, an EHV-2 vaccine was shown
to protect foals from pneumonia due to Rhodococcus equi infection
(Nordengrahn et al 1996). The fact that EHV-2 is able to infect foals
in the presence of maternally-derived antibodies (Wilks and Studdert
1974; Pálfi et al 1978; Fu et al 1986; Murray et al 1996) and can be
isolated from horses with high anti-EHV-2 antibody titres (Borch-
ers et al 1997), provides further evidence that EHV-2 (and possibly
EHV-5) modulates the immune response of its host.
Alternatively, detection of EHV-2 and EHV-5 infections by vi-
rus isolation before infections with other viruses were detected
serologically could be explained by differences between these
diagnostic methods. Serological diagnosis, especially in young
animals, is less reliable than virus isolation because residual levels
of maternal antibodies can interfere with the animal’s ability to
mount an active humoral response. Three foals showed serologi-
cal evidence of recent EHV-1 infection using ELISA, without a
corresponding 4-fold rise in SN antibody titres. These infections
would not have been detected by a SN test alone. In addition,
Foals A1 and A3 showed a significant rise in ELISA titre with-
out reaching the 60% cut-off value used in our study, and their
corresponding SN titres remained low. The rise in ELISA titres
could represent a serological response to EHV-1 infection in the
presence of residual levels of maternal antibodies. Alternatively, it
could represent cross-reaction to EHV-4 antibodies and EHV-4
infection in these foals. In Foal B16, the 4-fold rise in EHV-1 SN
titre concomitant with a low ELISA titre is suggestive of infec-
Figure 4. Results of serological testing for the presence of (a) serum
neutralising (SN) and (b) ELISA antibodies to equine herpesvirus 1
(EHV-1); (c) SN antibodies to equine rhinitis-B virus (ERBV); and (d)
haemagglutination inhibition (HI) antibodies to equine adenovirus 1
(EAdV-1) in foals in Group C. Titres of individual foals (C1 through
C5) are shown, expressed as reciprocal serum dilutions. Consecutive
bars represent consecutive months from October 1995 through to
December 1996. Speci c dates of each sampling are shown in Table 1.

Dunowska et al
New Zealand Veterinary Journal 50(4), 2002
147
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tions with other viruses may also have gone undetected during the
first 4–6 months of life.
Some of the foals sampled showed serological evidence of infec-
tion with EHV-1, EHV-4, EAdV-1 or ERBV in April and May
(Groups A and B) or in June–July (Group C). The majority of
foals from Group A remained healthy, whereas most of the foals
from Groups B and C showed signs of respiratory disease at some
stage between April and June. Since there was serological and/or
virological evidence of the presence of the same viral infections
among these three groups of foals, the presence or absence of
clinical signs of respiratory disease could reflect some unidentified
differences in environmental or husbandry conditions between
these groups. It could also result from the fact that only limited
numbers of foals were sampled. Another possibility is that the
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ticularly foals from Group C, were due to bacterial rather than
viral infections. The data of Hoffman et al (1993) implicated the
predominant role of primary bacterial infections in respiratory
disease in foals, as these authors did not isolate any viruses from
101 cases of respiratory disease in foals. Also, seroconversion to
EHV-1 or EHV-4 was only detected twice among 47 randomly
selected, paired serum samples. These results are in contrast to the
results of our study. Although the occurrence of bacterial infec-
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Acknowledgements
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This work was supported by grants from the New Zealand
Equine Research Foundation and the Norman Cunningham
Trust. Financial support for MJ Studdert was supported by
Racing Victoria and a Special Virology Fund. We thank Nino
Ficorilli at the University of Melbourne for technical assistance in
serological testing for equine rhinitis viruses and staff at the Cen-
tral Animal Health Laboratory, Wallaceville, New Zealand, for
performing the EAV serological assays. We also thank the owners
and handlers of the foals who participated in the survey.
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Accepted for publication 17 April 2002