ADRENOCORTICAL RESPONSE OF CALVES TO TRANSPORT STRESS AS

MEASURED BY SALIVARY CORTISOL

L. R. FELL and D. A. SHUTT
Department of Agriculture, Hawkesbury AgriculturalResearchUnit, P.O. Bor217,
Richmond, N.S.W. 2753, Australia. Received 14 Nov. 1985, accepted 27 Mar.
t 986.
Fell, L. R. and Ssurr, D. A. 1986. Adrenocortical response of calves to trans-
port stress as measured by salivary cortisol. Can. J. Anim. Sci. 66:631-641 .
A new approach utilizing a "nonstressful" technique of saliva collection and de-
termination of salivary cortisol by radioimmunoassay was used to monitor the ad-
renocortical response of calves to repeated transport stress. Four standardized truck-
ing and sampling procedures were used with l9 calves previously trained to being
mustered into a race (chute) and having saliva collected. Basal mean salivary cor-
tisol levels in the 19 calves ranged from 0.3
-r
0.4 nmol L-\ to 3.4
-r
0.6 nmol
L
',
and after 30 min of trucking manoeuvres (starting, turning, reversing and stop-
ping), significant increases (P<0.01)
in mean salivary cortisol levels occurred on
all four occasions to values ranging from 10.5
+
1.4 nmol L
t
to 16.0
+
2.3 nmol
L
'.
After a further 2 h of normal road transport mean levels of salivary cortisol
were still significantly elevated (P<0.05) and ranged from 4.0
-r
0.9 nmol L
lto
11.3
-r
2.6 nmol L
'in
comparison with 0.5
+
0.2 nmol L
'to
1.4
+
0.3 nmol
L-' for nine control calves (not transported). Salivary cortisol levels returned to
pretreatment levels within 2 h post-transport. There was a tendency for Friesian
calves, less than I mo of age, to have a lower adrenocortical response to transport
than crossbred calves of the same age. No evidence was found for adaptation of the
adrenocortical response of the calves to transport.
Key words: Adrenocortical response, calves, transport stress, salivary cortisol
[R6action
corticosurr6nale des veaux au stress li6 au transport, telle que mesurde
par le cortisol salivaire.l
Titre abr6g6: Stress caus6 par le transport et cortisol salivaire chez les veaux.
Une nouvelle m6thode ayant recours i une technique "non stressante" de pr6ldve-
ment de salive et de d6termination du cortisol salivaire par radio-immunodosage a
6td utilis6e pour surveiller la r6action corticosurr6nale des veaux face au stress caus6
par un transport r.p,t6..
Quatre
essais normalis6s de transport par camion et d'6-
chantillonnage ont 6td appliqu6es ir 19 veaux pr6alablement habitu6s d 6tre re-
groupds dans un couloir (goulotte) et d subir un pr6lbvement de salive. Les con-
centrations moyennes de base du cortisol salivaire pr6lev6 sur 19 veaux se situaient
entre 0,3
-+
0,4 nmol L
I
et3,4
+
0,6 nmol L
'.
Aprds 30 min de transport par
camion (d6part, virage, marche arriEre et arr6t), on a observ6 au cours des quatre
essais une augmentation significative (P<0,01) des concentrations moyennes du
cortisol salivaire; les nouvelles valeurs variaient entre 10,5
+
1,4 nmol L
'
e 16,0
+
2,3 nmol L
r.
Aprds deux heures suppl6mentaires de transport normal sur route,
les concentrations moyennes du cortisol salivaire 6taient encore 6lev6es, de fagon
statistiquement significatives (P<0,05); elles se situaient entre 4,0
+
0,9 nmol L
'
e 11,3
-f
2,6 nmol L
'comparativement
e 0,5
+
0,2 nmol L-t etl,4
-r
0,3 nmol
L-' pour les neuf veaux t6moins (non transport6s). Moins de 2 h aprbs le transport,
les concentrations du coftisol salivaire sont redescendues d ce qu'elles 6taient avant
le traitement. Au cours du transport, les veaux Friesian, dgds de moins d'un mois,
Can. J. Anim. Sci. 662 637-641
(Sept. 1986)
637
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638
cANADTAN JOURNAL oF ANIMAL scrENCE
ont eu tendance i connaitre une 16action corticosurr6nale inf6rieure d celle des veaux
de croisement du m6me Age. Rien ne laisse croire que 1a r6action corticosurr6nale
subit des modifications par suite de l'adaptation des veaux au transport.
Mots cl6s: R6action corticosurr6nale, veaux, stress li6 au transport, cortisol salivaire
A method of assessing adrenocortical re-
sponse to stress, based on measurement of
the concentration of cortisol in saliva, was
developed (Fell et al. 1985) as partof a need
for re-evaluation of stress in farm animals
(see Dantzer and Mormdde
(1983)).
It also
avoids the adrenocortical effects of stress
associated with frequent blood sampling
(Bassett and Hinks 1969). This method al-
lows the monitoring of changes in biolog-
ically active
"free"
cortisol in plasma, but
not protein-bound cortisol as it does not en-
ter the saliva.
Calves are often exposed to stress when
they are transported from their home farm
(Kilgour and Mullord 1973; Carson and
Wood-Gush 1984), and plasma corticoste-
roids have been used as one index of stress
to evaluate some of the effects of transport
(Stephens
and Toner 1975; Johnston and
Buckland 1976; Crookshank et al. l9'79).
The present investigation was carried out to
extend the above observations by examin-
ing group and individual responses of
calves to repeated transport stress. This
study involved "nonstressful" saliva col-
lection and determination of changes in sal-
ivary cortisol to monitor stress.
MATERIALS AND METHODS
Twenty-eight calves were used in this experi-
ment, 14 Friesians and 14 mixed beef breeds
(predominantly Hereford, Aberdeen Angus), of
which 20 were male and eight female. The calves
were purchased locally and were reared from
about 1 wk old. The experimental design com-
prised four replicates of a standardized trucking
procedure (i.e. four trips), carried out with 19
of the calves (designated T1-Tl9), over a period
of 2 wk. Of these calves T4, T7, T11 and Tl9
were females and the other 15 were males. At
the mid-point of this period the age of the calves
ranged from 16 d (T19) to 60 d (Tl), and their
weights ranged from 40 to 80 kg.
The truck used had a floor space of 2.5 x 4.3
m which was totally enclosed by a slatted steel
crate with spaces (5 cm) between the slats. The
trucking procedure included moving the calves
from a paddock to a race for pretreatment sam-
pling of saliva (experimental period 1). The
calves were previously accustomed to this pro-
cedure. Nineteen of the calves were then driven
up a ramp for loading, from the rear, onto the
truck for 30 min of trucking manoeuvres, which
included reversing, turns to left and right, and
stopping and starting (experimental period 2). A
second salivary sample was then coilected on the
truck. The calves were then exposed to an ad-
ditional 2 h of normal road transport before re-
turning to the presampling area (experimental
period 3). After unloading down the ramp, the
calves were sampled for a third time in the race,
then released into their home paddock, and sam-
pled in the race 2h later (experimental period 4)
and24h later. Trips I and2 were carried out in
the same truck, but trips 3 and 4, unavoidably,
had to be carried out in another truck which was
the same size, but a different model. The second
truck appeared to have better suspension and
gave a smoother ride. Nine control calves were
sampled in the race with the calves that had been
transported (after experimental period 3).
Saliva samples from the calves were aspirated
from the side of the mouth with minimum re-
straint, and cortisol was determined by specific
radioimmunoassay as described by Fell et al.
(
r 985).
Salivary cortisol was found by Fell et al.
(1985) to give an accurate measurement of
plasma''free'' cortisol in the sheep, and this was
confirmed in five of these calves. When the five
calves were sampled after20 min transportation,
the mean salivary and plasma "free" cortisol
levels increased from 1.9
+
0.5 nmol L
'
and
0.8
+
0.1 nmol L
'to
9.0
-t-
1.3 nmol L
'and
9 .6
+
1.2 nmol L
' ,
respectively. Total cortisol
levels in blood plasma of the calves increased
from 11
+
2 nmol L-'to 100
+
ll nmol L
'
during the same period. There is normally an
increase in salivary flow rate during transport,
but cortisol concentration has been shown to be
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FELL AND SHUTT
-
TRANSPORT STRESS AND SALIVARY CORTISOL IN CALVES
independent of salivary flow rate (see Vining et
al. (1983),
Fell et al. (1985)). Differences be-
tween mean values in these exoeriments were
determined by Student's unpaired r-test. and re-
sults are expressed as the mean
+
SE.
RESULTS AND DISCUSSION
The space provided for the calves in the
truck (approximately
0.5 m,/calf) allowed
room for them to move about, but on each
trip the calves huddled together, particu-
larly during highway travelling at a steady
speed. Most faced towards the rear of the
truck. During the trucking manoeuvres
(turning,
stopping etc.), some calves lost
their footing temporarily, but, with one ex-
ception, all calves remained standing
throughout most of the treatment so none
was noticeably trampled. One calf (T5) was
lying down for most of the first trucking and
his salivary cortisol was in the middle of the
range for this trip.
With each successive trip, the calves
were noticeably easier to drive up the ramp
and onto the truck and seemed to be more
sure-footed during transport, indicating that
some degree of behavioral adaptation was
occurring. However, this was not reflected
in salivary cortisol. The mean salivary cor-
tisol levels for the 19 calves in resoonse to
four separate trips are shown in Fig. l. On
all four occasions after 30 min of trucking
manoeuvres, mean salivary cortisol levels
increased significantly from pretreatment
values of 0.3
-r
0.4 nmol L-'to 3.4
-r
0.6
nmol L
I
to values ranging from 10.5
-r
1.4 nmol L
I
to 16.0
-i-
2.3 nmol L-'
(P<0.01).
No significant differences were
found in salivary cortisol values between
trips 1 and 2 or between trips 3 and 4.
Somewhat lower values were found durins
trips 3 and 4 than during trips l and 2. bui
this was probably due to the second truck
being slightly more stable.
After a further 2 h of road transport.
levels of salivary cortisol were still signif-
icantly elevated (P<0.05) when all calves
were sampled back in the race. Mean levels
were 4.0
-r
0.9 nmol L
'to
11 .2
-r
2.6
nmol L
I
in comparison with values for
nine control calves (not transported) rang-
ingfrom0.5
r-
0.2nmolL
1to
1.4
-r
0.3
nmol L
'.
A further 2 h after transport sal-
ivary cortisol values had returned to the pre-
treatment range and no increases were noted
at 24 h post-transport.
Additional observations also indicated
that the calves did not show adrenocortical
adaptation to the transport procedures. One
month after the four experimental trips, 20
of the calves were transported on a trailer
to a new farm. The calves included six
calves that had previously acted as controls
and 14 calves that had participated in all
-3 -4
frfr
llnllil
"ll
[]
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rr
o
o24024
Hours
Fig. l. Changes in salivary cortisol levels in re-
sponse to transport. Values are mean salivary
cortisol levels (
+
SE) for 19 calves sampled be-
fore and after 30 min of trucking maneuvres and
2 h of road transport on four occasions. Trips I
and 2 were on the same truck and trips 3 and 4
were, unavoidably, on another truck ofthe same
dimensions, but somewhat smoother riding.
15
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CANADIAN JOURNAL OF ANIMAL SCIENCE
four transport treatments. Salivary cortisol
determinations in samples collected from
these two groups of calves, before trans-
port, at the end of 30-min transport to the
new farm, or 24 h after arrival at the new
farm, showed no significant differences.
Mean cortisol values before transport were
2.6
-r
0.8 nmolL-1(n:6)and3.2
+
0.7
nmol L
1
(n
:
14). After 30 min transport,
significantly higher (P<0.001) cortisol
levels of 15.7
-t
1.3 nmol L
'and
13.2
-t
2.3 nmol L
',
respectively, were found.
After 24 h on the new farm salivary cortisol
levels in both groups ofcalves had declined
It
5
o
15
10
5
o
Tzga 1234 1234
ExPerimental
Periods
Fig. 2. Changes in mean saiivary cortisol levels (+ SE) in 19 individual Friesian or beef breed
calves (Tl-Tl9) in response to four separate transportations. Experimental sampling periods cor-
respond to (1) pretreatment, (2) on truck after 30 min of trucking maneuvres, (3) in race after
completion of 2 h road transport, and (4) in race 2 h after transport. O, mixed beef breeds; *,
calves with a mean age of less than I mo of age at the mid-point of four transport runs over a period
of 2 wk. All other calves were 1.5-2 mo of ase.
20
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FELL AND SHUTT
-
TRANSPORT STRESS AND SALIVARY CORTISOL IN CALVES 641
to 1.3
-r
0.5 nmol L-r and 1.8
r-
0.4 nmol
L-1, respectively.
On the basis of mean salivary cortisol
levels of individual calves, obtained by
pooling the cortisol levels at period 2 from
the four trips, the individual calves were di-
vided into three groups (Fig2). Group I in-
cluded six calves with mean salivary cor-
tisol levels greater than 15 nmol L-
1.
Group
2 included six calves whose salivary corti-
sol levels were between 10 and 15 nmol
L
r,
and Group 3, seven calves with sali-
vary cortisol levels less than l0 nmol L
1.
Experimental period 2 was chosen for com-
parisons because peak salivary cortisol val-
ues after 30 min of variable transport were
obtained in 15 out of 19 ofthe calves at this
sampling period. It was noticeable (Fig. 2)
that all seven calves in Group 3 were rep-
resented by younger calves (mean age dur-
ing the four trips over a 2-wk period of less
than 28 d), while only four of the younger
calves were in Group 2 and only two were
in Group l Of interest, the younger calves
in Group 3 included five Friesian calves and
only two of the beef breeds.
In conclusion, it was found that all 19
calves responded to four separate transpor-
tations with increases in mean salivary cor-
tisol levels. This agrees with the findings
of Johnston and Buckland (19'76) who used
increases in plasma corticosteroids to in-
dicate changes in adrenocortical activity in
response to transport stress. However,
Johnston and Buckland (1916) found little
difference between the response of Holstein
bull calves of I wk of age and the same
calves at 3 mo of age. This may be ex-
plained in part by differences in experi-
mental design. For example, in our study
younger calves and older calves were trans-
ported together. We did find the salivary
cortisol response of individual calves to re-
peated transport to be sufficiently consis-
tent to classify them into three groups sug-
gesting that age and breed effects may be
worth investigating in another larger study.
ACKNOWLEDGMENTS
We are grateful to Hawkesbury Agricultural
College for providing facilities to run the calves,
to Cliff Bentley with the help of Ben Abbott and
Kristy Green for valuable assistance with the an-
imals. and to Ruth Connell for technical assis-
tance with the cortisol assays.
Bassett, J. M. and Hinks, N. T. 1969. Micro-
determination of corticosteroids in ovine periph-
eral plasma: elfects of venipuncture. corticotro-
pin, insulin and glucose. J. Endocrinol. 44:38'7-
403.
Carson, K. and Wood-Gush, D. G. M. 1984.
The behaviour of calves at market. Anim. Prod.
39 389-39'7.
Crookshank, H. R., Elissalde, M. H., White,
R. G., Clanton, D. C. and Smalley, H. E.
1979. Effect of transportation and handling of
calves upon blood serum composition. J. Anim.
Sci. 48: 430435.
Dantzer, R and Mormede, P. 1983. Stress in
farm animals: A need for revaluation. J. Anim.
Sci.57:6-18.
Fell, L. R., Shutt, D. A. and Bentley, C. J.
1985, Development of a salivary cortisol method
for detecting changes in plasma free cortisol
arising from acute stress in sheep. Aust. Vet. J.
62: 403-406.
Johnston, J.
D. and Buckland, R. B. 1976.
Response of male Holstein calves from seven
sires to four management stresses as measured
by plasma corticoid levels. Can. J. Anim. Sci.
56
'727-732.
Kilgour, R. and Mullord, M. 1973. Transport
of calves by road. N.Z. Vet. J. 2l:'7-lO.
Stephens, D. B. and Toner, J. N. 1975. Hus-
bandry influences on some physiological param-
eters of emotional responses in calves. Appl.
Anim. Ethol. l: 233-243.
Vining, R. F., McGinley, R. A. and Symons,
R. G. 1983. Hormones in saliva: Mode of entry
and consequent implications for clinical inter-
oretation. Clin. Chem. 29: 1752-1'7 56.
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