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The
Veterinary Journal
The Veterinary Journal 177 (2008) 159–168
www.elsevier.com/locate/tvjl
Review
Foot-and-mouth disease: A review of intranasal infection
of cattle, sheep and pigs
Robert Sellers a, John Gloster
b
b,*
a
4 Pewley Way, Guildford, Surrey GU1 3PY, UK
Met Office, based at the Institute for Animal Health, Pirbright Laboratory, Ash Road, Pirbright, Woking, Surrey, Guildford GU24 0NF, UK
Accepted 15 March 2007
Abstract
In an outbreak of foot-and-mouth disease (FMD) it is important to identify animals at risk from airborne virus. Investigations have
been carried out over the years to determine the dose required to infect cattle, sheep and pigs by the intranasal route. This paper reviews
the results of investigations for animals which have been infected by instillation or spraying a virus suspension into the nostrils or by
exposure to affected animals through a mask or by indirect contact.
The lowest doses were found by use of a mask. With virus from affected pigs given through a mask, doses of 18 infectious units (IU) in
cattle and 8 IU in sheep were found to cause infection and give rise to lesions. Overall, cattle required the least amount of virus followed
by sheep. Pigs required a dose of 22 IU to cause infection and a dose of 125 IU to give rise to lesions. In many experiments pigs failed to
become infected. With all three species the dose varied with the individual animal and the virus strain. For modelling previous outbreaks
and in real time, a dose of 8 IU or 10 and 50% infectious doses (ID50) could be used where cattle and sheep were involved. Experience in
the field, combined with the results from experiments involving natural infection, indicate that pigs are not readily infected by the intranasal route. However, for modelling purposes a dose of about 25 IU should be used with care.
Investigations are needed to determine doses for virus strains currently in circulation around the world. In addition, the nature of the
aerosol droplets needs to be analysed to determine how the respective amounts of infective and non-infective virus particles, host components and, in later emissions, the presence of antibody affect the survival in air and ability to infect the respiratory tract. Further work
is also required to correlate laboratory and field findings through incorporation of the doses into modelling the virus concentration
downwind in order that those responsible for controlling FMD are provided with the best available assessment of airborne spread.
Finally, the doses found for infection by the intranasal route could be applied to other methods of spread where virus is inhaled to assess
risk.
Crown Copyright 2007 Published by Elsevier Ltd. All rights reserved.
Keywords: Foot-and-mouth disease; Intranasal infection; Review of experimental results
Introduction
Foot-and-mouth disease (FMD) is a highly infectious
viral disease of cloven-hoofed animals both domestic and
wild. The disease spreads by contact between infected
and domestic animals, by animal products (milk, meat and
semen), by mechanical transfer on people, wild animals
*
Corresponding author. Tel.: +44 1483 231023.
E-mail address: [email protected] (J. Gloster).
and birds, by vehicles and fomites and by the airborne route.
Spread by airborne carriage on the wind has been considered
a possibility from the beginning of the 20th century especially
by Scandinavians (Bang, 1912; Donaldson, 1979; Penberthy,
1901) and its part in spread of disease was recognised in the
UK in the 1967–1968 and 2001 epidemics and in the spread
of FMD from Brittany to the Isle of Wight in 1981 and in
Hampshire in 1967 (Donaldson et al., 1982; Gloster et al.,
2003, 2005a,b; Hugh-Jones and Wright, 1970; Mikkelsen
et al., 2003; Sellers and Forman, 1973).
1090-0233/$ - see front matter Crown Copyright 2007 Published by Elsevier Ltd. All rights reserved.
doi:10.1016/j.tvjl.2007.03.009
160
R. Sellers, J. Gloster / The Veterinary Journal 177 (2008) 159–168
The airborne disease cycle can be divided into three
stages: emission, transport and inhalation. Before 1984,
the amounts of virus emitted by infected animals, the size
of the infective particles and their survival in air was generally established but the process of inhalation, especially the
dose required to initiate infection by the respiratory route,
remained to be investigated more fully. A review of aspects
of airborne spread of FMD is given in the paper on pathogenesis and diagnosis of FMD by Alexandersen et al.
(2003b).
The potential for the spread of FMD by the airborne
route can be determined by estimating the amount of virus
released into the atmosphere and establishing the meteorological conditions in the vicinity of infected animals. This
information is used as input into an atmospheric dispersion
model which calculates downwind concentrations of virus
and an inhaled dosage (Gloster et al., 2003; Ryall and Maryon, 1998; Sørensen et al., 2000, 2001). To determine the
area at risk from airborne infection the dosage to initiate
infection is also required.
This paper provides a review of the published experimental data involving cattle, sheep and pigs with a view
to giving guidance on dosage to those required to model
the airborne risk of disease spread and to those responsible
for controlling disease outbreaks. The dose determined can
also be used to assess the risk of infection of the respiratory
tract, where virus is inhaled from contact with affected animals, contaminated personnel, vehicles that have previously transported affected animals, aerosols from milk
spills and from contaminated fomites.
Experimental conditions
The experiments can be divided into four categories:1. Application by instillation of virus suspension into the nostrils – artificial method of infection with artificially prepared virus.
2. Application by a spray of virus suspension to nostrils –
artificial method of infection with artificially prepared
virus.
3. (a) Exposure of animals to the aerosols of infected
animals through a mask – artificial method of infection
with naturally produced virus. (b) Exposure of cattle
virus generated from a May spinning top through a mask –
artificial method of infection with artificially prepared
virus.
4. Indirect contact between infected animals and clean recipient animals – natural method of infection with naturally produced virus.
All of the experiments were carried out in isolation units
under disease controlled conditions with animals from
European breeds up to 3 years old. FMD strains of all
virus types except SAT1 and SAT3 were used. The type
of spray and the collecting apparatus varied between
experiments.
FMD virus was titrated by different methods: intradermal inoculation of the tongues of cattle (Graves
and Cunliffe, 1960; Henderson, 1952; Korn, 1957),
unweaned mice (Eskildsen, 1969; Terpstra, 1972), sheep,
lamb and pig kidney monolayer tissue cultures (Sutmoller et al., 1968; McVicar and Sutmoller, 1976, 1969;
Bouma et al., 2004; Brown et al., 1992, 1996) and
bovine thyroid monolayer (BTY) tissue cultures, which
from 1981 onwards were used in the majority of experiments. BTY cultures had been found to be the most
sensitive at detecting virus in air samples from affected
animals (Donaldson et al., 1970). Strains adapted to
pigs do not grow in BTY cultures and the pig kidney
IBRS2 cell line was used instead (Dunn and Donaldson,
1997). In experiments from 2002 onwards, viral RNA in
virus containing material was measured by the reversetranscriptase polymerase chain reaction (RT-PCR)
method.
The genome equivalents per millilitre were about
100–1000-fold higher than the titre of infective virus in
BTY cells, although in late infection this increased up
to 100-fold more, probably due to the presence of antibodies (Alexandersen et al., 2003a). This ratio between
genome equivalents and infective virus is similar to the
ratio of non-infective 25 nm particles and infective
25 nm particles (100–1000-fold to 1) found by electron
microscopy (EM) of virus suspensions (Report, 19561960; Bradish et al., 1960).
Evidence of infection was taken as the presence of clinical signs, viraemia and seroconversion, or viraemia and
seroconversion, or viraemia or seroconversion alone.
Some FMD strains do not give rise to detectable lesions
in cattle, pigs and especially in sheep (Alexandersen
et al., 2003b). Viraemia can be of short duration without
leading to the development of antibody (Garland, 1974;
Gibbs et al., 1975; Donaldson and Kitching, 1989). Where
animals were killed during the incubation period, the
presence of virus in more tissues than earlier in the incubation period was taken as evidence of infection. Animals
may also show transient levels of antibody (Alexandersen
et al., 2002; Alexandersen and Donaldson, 2002). Animals
with transient antibody levels 1 in 45 or greater were
regarded as being infected. Animals without detectable
lesions may pass on infection to others (Callens et al.,
1998).
Review of animal experiments
The experimental results by all methods of infection for
cattle, sheep and pigs are given in Tables 1–3 and summarised in Table 4 and Fig. 1. Experiments specifically
designed to estimate dose are indicated in the tables (D).
The remaining experiments were part of investigations on
the pathogenesis of FMD, the exposure of vaccinated animals to infection and the development of carriers, in which
the dose was also measured.
R. Sellers, J. Gloster / The Veterinary Journal 177 (2008) 159–168
161
Table 1
Recipient animals – cattle
Method
Virus type/
strain
Instillation
A4691
Virus
source
Dose (log IU)
Amount
2.0 PK/SK IU
4.0 PK/SK IU
6.0 PK/SK IU
PK/SK
PK/SK
PK/SK
PK/SK
IU
IU
IU
IU
Experimental
length
C
V
S
+ve
ve
No.
Fig. 1
Comments
5 mL suspension
5 mL suspension
5 mL suspension
2
2
2
2
2
2
2
2
2
4
0
0
Sutmoller et al. (1968)
2
2
5 mL
5 mL
5 mL
5 mL
suspension
suspension
suspension
suspension
1
2
4
3
1
2
4
3
1
2
4
3
1
2
4
3
1
0
0
0
McVicar and Sutmoller (1976)
5
6
6
0
Bouma et al. (2004). No end point
Instillation
01
3.0
4.0
5.0
7.0
Instillation
O/NET/
2001
3.0 Cattle IU
(=3.8 PK IU, 3.6
LK IU)
3 mL suspension
3
Spray
039 D
5.1 Cattle IU
2 mL spray
6
6
2
Henderson (1952)
Spray
A119 D
1.95 Cattle IU
4.95 Cattle IU
6.95 Cattle IU
2 mL spray
2 mL spray
2 mL spray
2
2
2
2
2
2
4
0
0
Henderson (1952)
Spray
AM1 D
5.85 Cattle IU
3 · 4 mL spray
1
1
1
Henderson (1952)
Spray
O1
BFS1860
0.8 BTY IU
0.01 mL fine spray
3
Burrows et al. (1981) 1? ?contact
infection
3.1 BTY IU
0.3 mL coarse
spray
0.01 mL fine spray
0.3 mL coarse
spray
0/
1?
4
3
3
0
0
Virus in tissues in incubation period
1
1
1
1
1
0
0
0
0
0
Eskildsen (1969)
Virus in tissues in incubation period
No end points
4
0
Korn (1957). Virus in tissues at 63 h
p.i. No end point
3.5 BTY IU
5.75 BTY IU
Spray
Spray
O
4.0
5.0
5.0
5.7
6.2
O2
Mouse
Mouse
Mouse
Mouse
Mouse
IU
IU
IU
IU
IU
3.6 Cattle IU
100 mL
100 mL
100 mL
100 mL
100 mL
4
2
3
3
spray
spray
spray
spray
spray
1
1
1
0.5 mL spray
0
Spray
A
8.3 LK IU
1 mL spray
1
1
0
Brown et al. (1996). No end point
Spray
Asia1
5.8 LK IU
2 mL spray
3
3
0
Brown et al. (1992). No end point
Mask
O1
BFS1860D
Aerosol
from
May
spinning
top
Mask
SAT2 SAR
3/79D
2 pigs
3.85 to 5.05 BTY
IU
1.25 to 2.45 BTY
IU
0.95, 1.25, 1.45,
2.25
BTY IU
1.05, 1.25 BTY
IU
1–1.5 min
5
5
6
6
0
1–2 min
4
4
6
6
0
1 min
2
2
4
4
1.45–1.75 BTY
IU
1.25, 1.55, 1.95
BTY IU
1.45, 1.55 BTY
IU
1.45, 1.6, 2.25
BTY IU
1.55, 1.55 BTY
IU
10 min
5
5
5
5
10 min
2
3
2
3
60 min
60 min
60 min
60 min
3
3
3
3
3
3
3
3
BTY
BTY
BTY
BTY
IU
IU
IU
IU
1 min
1
Donaldson et al. (1987)
No end point
No end point
2
10 min
0
1a
Donaldson et al. (1987)
2
5 min
3
3
5 min
2
Indirect
O1
BFS1860
2 pigs
2.25
2.35
3.25
4.25
Indirect
O
UKG2001
3 pigs
3.85 BTY IU
2h
1
1
Indirect
O
UKG2001
4 sheep
N/A
5h
2
1
3
3
3
3
0
0
0
0
2
Donaldson and Kitching (1989)
No end points
1
1
1
3
Aggarwal et al. (2002)
1
2
0
Aggarwal et al. (2002). No dose
given
Key: C = clinical signs, V = viraemia, S = seroconversion or carrier; PK = pig kidney monolayer tissue cultures, SK = sheep kidney monolayer tissue cultures, LK = lamb
kidney monolayer tissue cultures, BTY = bovine thyroid monolayer tissue cultures, IU = infectious units, p.i. = post infection, trans = transient, N/A = not available. The
dose is the amount of virus inhaled at the nostrils, D = experiment to determine dose, ID50s are expressed as IU (ID50 · 0.7 assuming a Poisson distribution). The number in
the penultimate column (no. Fig. 1) cross refers to the X-axis in Fig. 1.
162
R. Sellers, J. Gloster / The Veterinary Journal 177 (2008) 159–168
Table 2
Recipient animals – sheep
Method
Virus
type/
strain
Virus
source
Dose (log IU)
Amount
Instillation
Instillation
O Greece
23/94
O2
4.9 BTY IU
2 mL
suspension
0.2 mL
spray
0.2 mL
spray
0.2 mL
spray
0.2 mL
spray
4.0 PK/SK IU
Instillation
A1
4.0 PK/SK IU
Instillation
A4691
4.0 PK/SK IU
Instillation
CTdF
4.0 PK/SK IU
Instillation
O Greece
23/94
2.85 BTY IU
3.35 BTY IU
3.85 BTY IU
4.85 BTY IU
5.85 BTY IU
Mask
Indirect
Indirect
O1 BFS
1860 D
O1 BFS
1860
O UKG
2001
2 pigs
1.8, 2.1, 3.0
BTY IU 0.45
1.3 BTY IU
0.9, 0.9, 1.3, 1.8
BTY IU
0.95 BTY IU
1.8, 1.9, 2.0
BTY IU
0.7,1.7 BTY IU
1.7, 2.1, 2.3
BTY IU
1.0 BTY IU
0.9, 0.9, 0.9, 1.0,
1.7 BTY
IU
Experimental
length
2 mL
suspension
2 mL
suspension
2 mL
suspension
2 mL
suspension
2 mL
suspension
C
V
S
+ve
14
15
15
16
0
3
1
4
0
12
1
3
1
0
5
1
4
1
ve
4
5
5
0
4
5
5
0
4
5
5
0
15 min
3
3
3
3
15 min
10 min
2
4
4
4
No.
Fig. 1
Comments
Hughes et al. (2002). No end
point
McVicar and Sutmoller
(1969)
McVicar and Sutmoller
(1969). No end point
McVicar and Sutmoller
(1969)
McVicar and Sutmoller
(1969)
Hughes (2002)
4
Gibson and Donaldson
(1986)
Gibson et al. (1984). No end
point
Aggarwal et al. (2002). No
end point
2
10 min
1
10 min
10 min
1
3
3
3
10 min
10 min
2
10 min
5
5
5
5
0
2
3
3
3
1
4 pigs
3.05 BTY IU
2h
4
4
4
4
0
7
3 pigs
3.45 BTY IU
2h
3
4
4
4
0
8
4
4
5
5
0
2
2
1
5
Esteves et al. (2004)
6
J.-F. Valarcher et al.,
unpublished data
No end points
Indirect
O UKG
2001
4 sheep
N/A
>5 h
Indirect
O UKG
2001 D
3 sheep 1
per box
2.35 BTY IU
24 h
Indirect
O UKG
2001
2 sheep
3.15 BTY IU
2h
0
2
3.45
3.65
3.75
4.25
4h
6h
8h
25 h
0
0
0
0
2
2
2
2
BTY
BTY
BTY
BTY
IU
IU
IU
IU
Aggarwal et al. (2002)
No end point
No dose given
For key see Table 1.
Cattle
Most experiments were carried out with type O strains,
but type A, SAT 2 and Asia 1 strains were also used. In
some successful experiments no end point was determined
(Table 1). The lowest dose by instillation was found to be
log 2.0 IU (100 IU) (Sutmoller et al., 1968) and by spray
was log 1.95 IU (90 IU) (Henderson, 1952). In experiments
using a mask, log 1.25 IU (18 IU) from natural infection
and log 0.95 IU (9 IU) from spray from a May spinning
R. Sellers, J. Gloster / The Veterinary Journal 177 (2008) 159–168
163
Table 3
Recipient animals – pigs
Method
Virus
type/
strain
Instil
OD
Spray
O1
Mask
O1 Laus
Virus
source
2 pigs
SW/65
D
Mask
O1 Laus
3 pigs
SW/65
Dose (log IU)
Amount
>6.25 Cattle
IU
3.95 Mouse IU
1 mL
0
2
2 mL
2
0
1.35, 1.95, 2.1,
2.45, 2.45,
2.45, 2.55 BTY
IU 1.95,
2.35 BTY IU
Experimental
length
10 min
C
V
1
S
7
+ve
7
10 min
1.25, 1.35,
1.35, 1.55,
1.55, 2.05, 2.1
BTY IU
2 and 10 min
1.35, 1.55, 1.75
BTY IU
10 min
ve
No.
Fig. 1
Comments
Graves and Cunliffe (1960). No
infection or end point
Terpstra (1972). Virus in tissues at 72
and 96 h p.i., no end point
9
Alexandersen et al. (2002) 2/7
antibodies at 14, but not 21 dpi.
9
Alexandersen et al. (2002)
2
0
7
No end point
D
Mask
O1 Laus
3 pigs
0
3
9
SW/65
D
Mask
O1 Laus
No end point
3 pigs
SW/65
D
Mask
O1 Laus
3 pigs
SW/65
D
Mask
O1 Laus
3 pigs
SW/65
D
Mask
O SKR
Alexandersen et al. (2002)
3 pigs
1/2000
D
1.35, 1.45,
1.95, 1.95,
2.85 BTY IU
10 min
1.35, 1.95,
2.45, 2.45, 2.6
BTY IU
10 min
1.55, 2.85, 3.1,
3.1, 3.45
BTY IU
1.45, 2.95, 3.35
BTY IU
5 min
2.25,
2.35,
2.55,
2.85,
3.25,
IU
5 min
2.25,
2.35,
2.55,
2.95,
3.25 BTY
0
5
9
Alexandersen et al. (2002)
No end point
0
5
9
Alexandersen et al. (2002)
No end point
1
5
5
5 min
10
Alexandersen and Donaldson (2002) 4
trans antibodies
10a
Alexandersen and Donaldson (2002)
3
0
10
No end point
Indirect
O UKG
2001
3 pigs
>4.7 BTY IU
24–48 h
0
8
10b
Alexandersen and Donaldson (2002) 1
trans antibody
No end point
Indirect
O UKG
2001
3 pigs
3.1 BTY IU
2h
0
4
11
Aggarwal et al. (2002)
No end point
Indirect
O UKG
2001
4 sheep
N/A
5h
4
0
Indirect
O Taw
9/97
1 pig in 4
boxes
>3.25 BTY IU
24–48 h
0
8
12a
Alexandersen et al. (2003a) 1 trans
antibody
No end point
Indirect
C Nov
SW 73
1 pig in 4
boxes
>5.1 BTY IU
24–48 h
0
8
12
Alexandersen et al. (2003a)
No end point
4
4
Aggarwal et al. (2002)
No end point
For key see Table 1.
top (Mitchell and Stone, 1982) were the lowest doses. The
lowest dose to cause clinical lesions was log 1.25 IU (18 IU)
after natural infection or spraying through a mask. The
highest dose that failed to cause infection using a mask
was log 1.55 IU (35 IU) for natural infection and
log 1.25 IU (18 IU) from spray from a May spinning top
(Donaldson et al., 1987). The lowest dose for indirect contact infection was log 2.25 IU (180 IU) (Donaldson and
164
R. Sellers, J. Gloster / The Veterinary Journal 177 (2008) 159–168
Table 4
Minimum dose (IU) to initiate sub-clinical and clinical infection: highest dose to fail to cause infection
Instillation
Spray
1
Cattle
Sheep
Pigs
2
100
2250
>1,800,000
3
1
Indirect contact
2
3
90
7100
2250
9000
Mask
1
2
3
1
2
3
180
1120
>250,000
180
1120
>250,000
7100
17,800
250,000
9
8
22
18
8
125
35
50
2250
1, minimum dose to initiate sub-clinical infection; 2, minimum dose to initiate clinical infection; 3, highest dose not to initiate infection.
a dose of log 2.35 IU (225 IU) over 24 h was found to infect
(Esteves et al., 2004). Differing values were found in the
dose required to infect individual sheep after natural infection through a mask.
Kitching, 1989). Henderson (1952) found a difference
between strains for the lowest dose to cause infection
(Table 1), a finding that he correlated with differences in
reaction to contact infection. In the experiments with
masks the dose to cause infection varied with individual
animals.
Pigs
The strains used in the experiment were of type O apart
from C Noville. Instillation of log 6.25 IU (1,800,000 IU)
failed to give rise to lesions in pigs (Graves and Cunliffe,
1960). Virus was recovered from the tissues of pigs killed
72 and 96 h after receiving a dose of log 3.95 IU
(9000 IU) (Terpstra, 1972). The lowest dose to cause infection in pigs exposed to natural virus through a mask was
log 1.35 IU (22 IU), while the lowest dose to cause lesions
was log 2.1 IU (125 IU) (Alexandersen et al., 2002). The
highest dose to fail to cause infection after exposure to natural infection through a mask was log 3.35 IU (2250 IU)
(Alexandersen and Donaldson, 2002). Experiments where
pigs were exposed to natural infection with naturally pro-
Sheep
Strains of types O, A and C were used in the experiments
the majority being of type O. End points could not be demonstrated in every experiment, some showing successful
infection, one other failure to infect by indirect contact
(J.-F. Valarcher et al., unpublished data). The lowest dose
for instillation was log 3.35 IU (2250 IU) (Hughes, 2002).
In mask experiments the lowest dose was found to be
log 0.9 IU (8.0 IU), which was also the dose to cause clinical infection. The highest dose that failed to cause infection
was log 1.7 IU (50 IU) (Gibson and Donaldson, 1986). In
experiments where sheep were exposed by indirect contact
Cattle
6
M M
* P
I
P
Pigs
Sheep
M M I I
* P P P
M I
P S
I
S
M I I I
P P P S
M M M
P P P
I
P
I I I
PPP
M M
P P
5
Log IU
4
3
2
1
0
1 1a 3
No infection
1 1a 2 3
Infection
4 56
No infection
Key: Numbers = reference identification (Tables 1 to 3),
4 7 8 5
9 10 10a&b 111212a
Infection
M = Mask, I = Indirect,
No infection
P = Pig,
9 10
Infection
S = Sheep,
* = Spinning top
Fig. 1. Summary of results from experiments which involved exposure of cattle, sheep and pigs to artificially prepared virus through a mask, natural virus
through a mask or by indirect infection. The results from experiments involving cattle, sheep and pigs have been plotted separately. For each experiment
the source and method of administering the virus is given (C = cattle, P = pig, S = sheep. * = spinning top, M = mask and I = indirect contact). Where in
an experiment animals have been recorded as non-infected and infected the same symbol has been plotted (open = no infection, closed = infection). For
example Gibson and Donaldson (1986) exposed sheep to virus from a pig (P), using a mask (M) and estimated the dosage for a number of animals some of
which became infected (filled in squares) and others which remained uninfected (open squares).
R. Sellers, J. Gloster / The Veterinary Journal 177 (2008) 159–168
duced virus by a natural method of infection – indirect contact – were not successful (Alexandersen and Donaldson,
2002; Aggarwal et al., 2002; Alexandersen et al., 2003a).
There was a difference between strain O UKG2001 and
strain O SKR 1/2000 in the successful response and failure
to respond respectively to infection through a mask by natural virus (Alexandersen and Donaldson, 2002). In the
mask experiments there was variation in the responses of
individual pigs to the doses given (Alexandersen et al.,
2002; Alexandersen and Donaldson, 2002).
It can be seen in Fig. 1 that the lowest dose was found
using a mask (artificial method, natural virus). There was
little difference between cattle, sheep and pigs in the lowest
dose to cause infection; however with pigs a greater dose
than that for cattle was found to give rise to lesions. In
addition a dose 60-fold greater than that for cattle failed
to infect pigs (Alexandersen and Donaldson, 2002). With
instillation, spray and indirect contact the doses found to
infect cattle were considerably less than the doses required
to infect pigs. Sheep also required a higher dose than cattle
when infection was by instillation or indirect contact
(Table 4).
Discussion
It may be thought that the absence of any standardized
technique and the differences in housing, instrumentation
and methods of measurement make it impossible to draw
conclusions about the dose required by the intranasal
route. However, in the majority of experiments reported
from 1981 onwards, BTY tissue cultures (the most sensitive
system) were used for virus assay. In addition, such experiments were reported from one centre (BBSRC, Pirbright),
where from 1960 infected livestock were held in units under
negative pressure, although the nature of the ventilation
system changed over the years.
Instillation and spraying of virus preparations into the
nostrils were the methods used to measure the dose in earlier experiments in cattle. In cattle the lowest doses by
instillation (100 IU, Sutmoller et al., 1968), spray (90 IU,
Henderson, 1952) and also by coughing and sneezing
(100 ID50, Sellers et al., 1971) were similar. They were also
lower than the doses found for pigs (Graves and Cunliffe,
1960 – instillation; Terpstra, 1972 – spray). In later experiments the development of the use of a mask ensured that
virus from natural sources was introduced to the respiratory tract more effectively and the doses for cattle and pigs
were found to be lower than those found by instillation and
spray. As with instillation and spray in the earlier experiments the dose for cattle by administration through a mask
was lower than that for pigs. In the field pigs are less likely
to be infected through airborne spread than cattle. The
doses used in previous modelling were based on those
found in early experiments (Gloster et al., 1981; Donaldson
et al., 1982). In later modelling, the lowest doses found in
cattle and sheep by mask experiments (natural infection
by artificial means) were used.
165
In investigations of doses of FMD virus strains in the
future the use of a mask (artificial method of infection with
naturally produced virus) would be the method of choice.
The method most resembling that in the field (natural
method with natural virus – indirect contact) did not result
in lower doses and higher doses were found by artificial
infection (instillation and spray). It should be pointed out
that the majority of experiments by instillation, spray and
indirect contact were not carried out to measure dose.
The experimenters were using the method they found most
appropriate to infect the animals.
The dose to initiate infection varied with the virus strain
used and between individual animals. Some of this variation could be attributed to the fact that experiments were
carried out under different conditions and with differing
assay systems. Overall slight differences could be important
in the field, where at low doses some highly susceptible
individuals could become infected, whereas with other
strains other individuals may escape infection resulting in
the failure of disease spread.
Where exposure was by indirect contact (natural method
and natural virus) factors in addition to animal species,
individual animal and virus strain affected the success of
infection by different doses. Such factors included the construction of the building and the airflow in the isolation
unit. Successful infection by indirect contact was achieved
in some of the experiments described here as well as by
Fogedby et al. (1960), Burrows (1968), Garland (1974)
and Sellers et al. (1968). Failure to cause infection in some
of the experiments reviewed here was found by other investigators, such as Traub and Wittman (1957), where infection was directed from infected animals in a shed to
calves and pigs. Bouma et al. (2004) found no evidence
of infection when calves were exposed by indirect or direct
contact to calves infected and reacting to the FMD strain
of the 2001 Netherlands outbreak. However the lesser susceptibility of calves to the Netherlands virus as well as the
environment could have been responsible for the failures.
Hutber and Kitching (2000) analysed the spread of
FMD within a cattle herd in Saudi Arabia and concluded
that spread had occurred through aerosol between pens
housing calves. This report also emphasized the importance
of spatial or physical barriers in preventing cross-infection.
Differences in inhalation of FMD virus by people in experiments in different laboratories could be explained by the
presence or absence of air currents (Amass et al., 2003;
Donaldson and Sellers, 2003). Air currents were found to
assist the spread of viruses not known to cause airborne
spread over distance, for example, African swine fever
(ASF) and classical swine fever (CSF) (Dewulf et al.,
2000; Wilkinson et al., 1977).
Successful infection in cattle and sheep occurred 1–
10 min after doses of 8–18 IU given by mask. Sellers
et al. (1970) found virus in the nose 5 min after contact
with affected pigs. This indicated that if virus is present
in the air it can be inhaled rapidly and give rise to successful infection in susceptible animals. Any virus particles that
166
R. Sellers, J. Gloster / The Veterinary Journal 177 (2008) 159–168
fail to cause infection would be cleared from the respiratory tract and unlikely to build up until a critical dose is
reached.
Infective aerosol particles inhaled through the nostrils
pass through the nares and thence to sites in the respiratory tract, where the virus multiplies and spreads to other
parts of the body (Burrows, 1972). The site of initial multiplication after inhalation of particles from natural
sources has been found to be the pharyngeal area (Burrows et al., 1981; Alexandersen et al., 2003b). In the experiments described in this paper, particles from artificial
sources initiated infection in the nasal mucosa and lung
as well as in the pharyngeal area. In some papers the distribution of particles size between 2 and 10 lm is given,
but owing to the range of sizes inhaled or given by spray
it is not possible to determine from the results which size
of particle initiated infection and in which part of the
respiratory tract.
Gloster et al. (2006) decided that there was no need to
take into account the size of the particles when modelling
spread of disease in the field. However the size of aerosol
particles and their site of initial infection in the respiratory
tract are important in studying the pathogenesis of the disease. In addition, knowledge of particle size and likely site
of infection are required for designing vaccines or antiviral
substances for protection of the respiratory tract. The composition of individual aerosol particles may be important in
determining the ability to initiate infection in the respiratory tract. It could be assumed by correlation with EM
results that genome equivalents represent one infective
virus particle to 100 or more non-infective particles. An
aerosol particle may contain virus particles infective for
BTY tissue cultures, non-infective particles, host material
and, in the later stages of infection, antibody. Presence of
antibody and high content of non-infective particles emitted from animals later in the infection may impair the ability to infect animals by the respiratory route. Further
investigations of the nature of the aerosol particle are
required.
Investigations have been made by modelling of past
FMD outbreaks in UK in the Isle of Wight and in the
1967/68 and 2001 epidemics, where no cause other than airborne carriage of virus could be identified. The virus concentration at the site of the outbreak determined from
the models was found to be up to 7000-fold less than the
concentration that would lead to infection in cattle based
on the dose of 10 TCID50 (Sørensen et al., 2000; Donaldson et al., 2001; Gloster et al., 2003, 2005a,b). The discrepancy between laboratory results and field findings could be
due to lack of information on the output and timing of
infection in animals and transmission including meteorological factors as well as to the dose inhaled. Finding that
cattle and sheep can be infected by doses less than 18 IU
and 8 IU, respectively, is unlikely to increase the concentration downwind significantly. Future research would be better directed at examining virus output and meteorological
factors. The meteorological process, procedures and fac-
tors involved have been discussed elsewhere (Gloster
et al., in press). See reference section for latest situation.
The minimal doses derived from the experiments can
also be used to determine the risk through inhalation of
aerosols derived from secretions and excretions from
infected or contaminated animals, from contaminated people and contaminated vehicles, milk spills and fomites
(Donaldson, 1979). The amounts of virus emitted in secretions and excretions are given in Sellers (1971), Thompson
(1994) and Alexandersen et al. (2003b).
Conclusion
The experiments reported show that the lowest doses for
cattle and sheep after natural infection were 18 IU and
8 IU, respectively. These values could be used for modelling airborne spread given the present state of knowledge.
However investigations need to be carried out on the aerosol particle itself to define not only its components but also
its capability both for infecting the respiratory tract at different stages of infection in the donor and for its survival
over distance. The discrepancy between virus concentrations based on laboratory measurements of output and
concentrations found downwind in field outbreaks indicates that further investigations on virus output and downwind transport are required. The experiments, together
with field experience, indicate that spread within buildings
may vary due to a number of factors. They raise questions
about emission from and spread within buildings during an
outbreak. The doses determined can also be used in determining the risk of aerosol spread in the field when methods
of spread such as via contaminated vehicles and fomites are
involved.
Acknowledgements
The authors express thanks to former and present colleagues at the Institute for Animal Health and the Met Office. Defra are thanked for funding John Gloster’s
contribution to preparation of this paper (contract SE
2926). Alex Donaldson, Tony Garland and David Paton
are thanked for providing very helpful comments on the
draft text.
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