The effects of simulated transportation conditions on the core

body and extremity temperature, blood physiology, and behavior

of white-strain layer pullets

S. Lalonde,* K. Beaulac,* T. G. Crowe,y and K. Schwean-Lardner*,1

*Department of Animal and Poultry Science, University of Saskatchewan, Saskatoon, SK, Canada, S7N 5A8; and
y
Department of Mechanical Engineering, University of Saskatchewan, Saskatoon, SK, Canada, S7N 5A9

ABSTRACT Transportation of poultry is stressful. pressure of CO2, total CO2, bicarbonate, and glucose),
The transportation of broilers has been well studied, birds were slaughtered, and data loggers were retrieved.
while the transportation of layer pullets from rearing to Data were analyzed as a randomized complete block
laying facilities has not been thoroughly evaluated. This design via Proc Mixed (SAS 9.4) and significance was
experiment aimed to establish the effects of temperature declared at P  0.05. There were no interactions
(T)/RH combinations and duration (D) of transport, via observed for the T/RH and D combinations throughout
a 5 ! 2 factorial arrangement of simulated transport the study. The CBT and foot T were lowest in pullets
conditions using 5 T/RH combinations (21 C with 30% exposed to 215 compared with all other treatments.
RH [21/30], 21 C with 80% RH [21/80], 30 C with 30% Foot T was also highest in pullets exposed to 30/80
RH [30/30], 30 C with 80% RH [30/80], and 215 C with compared with 215, 21/30, and 21/80. There was no
uncontrolled RH [215]), and 2 exposure D (4 or 8 h). impact of T/RH on pullet blood physiology. Activity and
Pullets (18–19 wk; n 5 240) were obtained from 3 com- thermoregulatory behaviors were impacted by the T/RH
mercial farms (N 5 3 farms). Pretreatment, birds were combinations. Pullets exposed to 30/30 and 30/80 spent
orally administered a miniature data logger to record the most time panting. Pullets exposed to 30/80 also
core body temperature (CBT), an initial blood sample spent the least amount of time motionless. Duration had
was taken (5 birds/replicate), and initial foot T was minor impacts on pullet CBT, blood physiology, and
recorded. Behavior during exposure was video recorded. behavior. These data indicate that as a response to
Following exposure, a final blood sample was taken thermal stress, layer pullets were successful at imple-
(analyzed for heterophil to lymphocyte ratio, partial menting mechanisms to maintain homeostasis.
Key words: cold stress, heat stress, thermoregulation, hematology, welfare
2021 Poultry Science 100:697–706
https://doi.org/10.1016/j.psj.2020.10.077

INTRODUCTION transported from a rearing barn to a laying barn at

approximately 17 wk of age, and this transportation
Transportation is a fundamental component of the can be over either a short or long distance
poultry industry; however, it can also result in animal (Schwartzkopf-Genswein et al., 2012; NFACC, 2016).
welfare concerns. Many of the steps involved are Stressors such as microclimate (temperature [T] and
stressors for poultry, such as catching, loading, trans- RH), vibration, noise, and the duration (D) of transport
port, and unloading (Schwartzkopf-Genswein et al., can have a large impact on the welfare of birds during
2012). Unlike broilers and laying hens, pullets are typi- transportation (Mitchell and Kettlewell, 1998, 2009).
cally not feed-withdrawn before transportation, as they The majority of transportation studies have focused on
are not destined for slaughter. Rather, pullets are broiler chickens as they are most commonly transported
for slaughter at the end of their production cycle. The
stressor most frequently studied in relation to transport
Ó 2020 The Authors. Published by Elsevier Inc. on behalf of Poultry is thermal stress. Chickens are homeotherms, where the
Science Association Inc. This is an open access article under the CC BY- core body temperature (CBT) of inactive birds is approx-
NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
Received June 8, 2020.
imately 41 C to 42 C (Mount, 1979). As a first response
Accepted October 27, 2020. to thermal stress, birds will modify their behavior in order
1
Corresponding author: [email protected] to conserve or dissipate heat (Mount, 1979). Under cold

697
698 LALONDE ET AL.

stress, birds will huddle or burrow under one another in hyponatremia (Thomas et al., 2008). Furthermore, as
an attempt to minimize radiation to the environment a result of stress during transportation, corticosterone
and maximize conduction between conspecifics (Mount, in the body will increase, causing glucose to be mobilized
1979; Strawford et al., 2011). Furthermore, pteroerection from the fat stores in the body through glycogenolysis,
can be performed as a thermoregulatory mechanism and from protein stores through gluconeogenesis
against cold exposure, as feathers can aid in providing (Zhang et al., 2009; Sherwood et al., 2013). This glucose
insulation against exposure to the cold T, trapping air be- will then be used as energy to respond to stressors and
tween feather fibers to conserve body heat (Mount, 1979; thermoregulate against heat or cold stress. As a result
Strawford et al., 2011). Under heat stress, birds will move of evaporative heat loss during heat exposure, blood
away from conspecifics and lift their wings, as an attempt gases and the acid–base balance in the blood will change,
to maximize space usage within the crate to ensure that causing a shift toward a respiratory alkalosis
the maximum body surface area is exposed to the environ- (Koelkebeck and Odom, 1995; Lumb, 2000). Hyperventi-
ment (Mount, 1979; Weeks et al., 1997). If maximizing lation during heat exposure will increase the elimination
body surface area alone is not enough, birds will imple- of CO2, resulting in a decrease in the partial pressure of
ment evaporative heat loss mechanisms (panting) to carbon dioxide (pCO2) and total carbon dioxide (tCO2)
dissipate heat (Whittow et al., 1964; Mount, 1979). How- in the blood, causing an increase in the blood pH
ever, when the RH in the environment is high, panting (Ahmad and Sarwar, 2006; Sherwood et al., 2013).
may become ineffective at dissipating heat from the Finally, the H/L ratio has been shown to be a reliable in-
core body (Whittow et al., 1964; Richards, 1971). It has dicator of chronic stress in poultry (Felver-Gant et al.,
been shown in previous research that the CBT of broilers 2012). Stress will cause corticosterone to increase which
will significantly increase as a result of exposure to T will, in turn, result in either an increase or decrease in the
above 30 C for a 2 h D (Sandercock et al., 2001; number of heterophils and/or lymphocytes, ultimately
Menten et al., 2006). causing an increase in the H/L ratio (Blecha, 2000;
If the bird is still experiencing thermal stress after Elsayed, 2014). Therefore, changes in the blood physi-
implementing behavioral changes, then physiological ology of pullets during transport may indicate that
changes are likely to occur. Under cold stress, to further they were subjected to thermal stressors during trans-
conserve heat for the core body, blood vessels to the ex- port; thus this may indicate poor welfare.
tremities (feet and comb) will vasoconstrict to restrict Since most studies regarding poultry transportation
blood flow (Midtgard, 1989; Wellehan, 2014). However, have focused on broilers, very little is known regarding
if the ambient T becomes too cold, the extremities can the transportation of pullets. The objective of this study
be susceptible to frostbite, and hypothermia may occur was to quantify how exposure-simulated transport condi-
(Mount, 1979; Wellehan, 2014). Previous studies have tions (hot and neutral T, at a high and low RH, as well as
consistently found that the CBT of broilers will decrease cold T) over a D of 4 or 8 h affects pullet behavior, phys-
when exposed to T below 25 C for 3 h (Dadgar et al., iology, and welfare. This was achieved by evaluating
2011, 2012a). If thermoregulatory mechanisms against CBT, extremity T, behavior, and blood physiology.
cold stress are unable to maintain the bird’s CBT, their
welfare can be negatively impacted, and mortality may MATERIALS AND METHODS
occur. When under heat stress, blood vessels to the ex-
tremities (feet and comb) will vasodilate, causing The experimental protocol for this research was
increased blood flow, which can aid in dissipating heat approved by the University of Saskatchewan Animal
through convection and radiation through the bare skin Care Committee and was performed under the guide-
(Whittow et al., 1964; Richards, 1971; Midtgard, 1989). lines of the Canadian Council of Animal Care (1993,
Thermal stress can also alter the blood physiology of 2009).
birds. Measuring various blood physiology parameters
such as blood glucose, acid–base balance parameters, Experimental Design
blood gas concentration, dehydration markers, and the
heterophil to lymphocyte (H/L) ratio may give an indi- This study was a 5 ! 2 factorial arrangement with
cation of the degree of stress experienced by birds during 5 T/RH combinations and 2 D of simulated transporta-
transportation. Due to dehydration, the amount of fluid tion (4 or 8 h). A hot (130 C), and neutral (121 C) T
in the blood will decrease, increasing both the hemato- were chosen with a high (80%) and low (30%) RH, as
crit and hemoglobin values (Thomas et al., 2008; well as a cold T (215 C) with uncontrolled RH, which
Bergoug et al., 2013). However, it has also been hypoth- resulted in the following 5 combinations: 215 C uncon-
esized by previous research that the hematocrit and he- trolled RH (215), 121 C 30% RH (21/30), 121 C 80%
moglobin values may decrease through hemodilution, RH (21/80), 130 C 30% RH (30/30), and 130 C 80%
which is an adaptive response to evaporative heat loss RH (30/80).
where water is lost through the extracellular fluid rather
than through the plasma (Borges et al., 2004; Scanes, Birds and Housing
2016). In addition, the body will lose water (via dehydra-
tion and defecation) and electrolytes (through mainly A total of 240 white-feathered commercial layer pullets
defecation), resulting in low blood sodium levels and were obtained from 3 commercial farms (2 Lohmann
 SIMULATED TRANSPORT OF WHITE LAYER PULLETS 699
(3 birds/replicate) included blood glucose (mmol/L),
sodium (mmol/L), hematocrit (% packed cell volume),
hemoglobin (mmol/L), pH, base excess (BE), pCO2
(mmHg) and oxygen (pO2; mmHg), tCO2 (mmol/L), ox-
ygen saturation (sO2; %), and bicarbonate (HCO32;
mmol/L). Afterward, pullets were placed into 1 of 2
crates for a 15-min lairage period to obtain baseline read-
ings for CBT, and then transported to the environ-
mental chambers (College of Engineering, University of
Saskatchewan, Saskatoon, Canada). After transporta-
tion, an initial foot T reading was taken individually
from all pullets using a thermocouple attached to a mul-
timeter (Omega HH509; Omega Engineering Inc., Laval,
Canada), and a plastic clip to ensure the thermocouple
remained in place on the middle toe of the right foot.
Pullets were then transferred into the treatment crates
(stocking density 45.5 kg/m2) and placed in the environ-
mental chamber with 1 crate per treatment D.

During Chamber Exposure

Simulated transportation conditions were achieved us-
ing 2 environmental simulation chambers (2.1 m ! 3.4 m;
Figure 1. Image of the interior of one of the environmental simulation College of Engineering, University of Saskatchewan)
chambers used in this study. (Figure 1). Chamber conditions were monitored and
adjusted if needed in real time via a thermocouple and
LSL-Lite and 1 Dekalb White) within a 250-km radius of multimeter (Omega HH509) and RH sensor
Saskatoon, Saskatchewan. Pullets were obtained at 18 to (HM1500LF; Measurement Specialties, Inc., Toulouse,
19 wk of age, and were given a 3 to 5 d acclimatization France). In addition, there were 3 T and RH data loggers
period. Pullets were housed in 1 floor pen (iButton Hygrochron DS1923-F5#; Maxim Integrated)
(3.9 m ! 3.0 m) with straw litter, with feed and water in each chamber with one located on the wall to record
provided ad libitum. Feed was acquired from the farm chamber conditions and one in each crate at bird level
of origin and was provided via 3 aluminum tube feeders that were set to record T and RH every minute. Two
(38 cm pan diameter). Water was provided with 2 bell infrared camera systems (Panasonic WV-CF224FX;
drinkers (36 cm diameter). The lighting program from Panasonic Corporation of North America, Newark, NJ)
the farm of origin was maintained via incandescent bulbs. were placed on the wooden table inside each chamber,
with 1 camera observing each crate. Behavior was evalu-
Prior to Chamber Exposure ated from the recording at 5-min intervals via scan sam-
pling over the entire D of exposure (4 or 8 h). The
Prior to the simulated transport exposure, 8 pullets behaviors evaluated included activity (motionless, active,
were randomly assigned to one of the T/RH and D com- shuffle), thermoregulatory (burrow, shiver, pteroerection,
binations (N 5 40). Pullets were not feed-withdrawn panting, survey), pecking (object peck, bird peck), and
prior to chamber exposure. Each pullet was wing banded other actions (preen, head shake, gulp, scratch, unidenti-
for identification purposes, and was orally administered fiable) which are defined in the ethogram (Table 1).
a miniature data logger (iButton Thermochron
DS1922L; Maxim Integrated, San Jose, CA), which trav- Post Chamber Exposure
elled to the crop/gizzard and recorded CBT every min-
ute. An initial blood sample was taken from the At the end of the exposure D, the crate was removed
brachial vein into a dipotassium EDTA tube (5 birds/ from the chamber. A final foot T was taken for each pul-
replicate). All 5 samples were used for H/L analysis let and a second blood sample was collected from the
which was performed by preparing a blood smear. The same 5 birds per crate. Blood samples were again
slides were stained (PROTOCOL Hema 3; Fisher Scien- analyzed for H/L ratio (5 birds/replicate) and blood
tific, Ottawa, Canada) and read under 100! oil magni- physiology (3 birds/replicate). The pullets were then
fication (microscope B-290TB; Optika, Bergamo, Italy) euthanized via electric stunning and exsanguination.
by counting the number of heterophils and lymphocytes The data logger was retrieved from the crop/gizzard
within a field of view until a total of 100 cells was and the data were downloaded (OneWire Viewer;
reached. Immediately after collection, 3 blood samples Maxim Integrated) for further analyses. Delta (D)
were analyzed using a CG81 cartridge in the i-STAT1 CBT was calculated for every hour by considering the
handheld analyzer (Abbott Point of Care Inc., Prince- average hourly CBT during exposure and subtracting
ton, NJ). Blood physiology parameters analyzed the average baseline reading prior to exposure.
700 LALONDE ET AL.

Table 1. Ethogram of behavioral activities monitored during the study and the
criteria for each behavior; all behaviors are mutually exclusive, except for
panting (Webster and Hurnik, 1990; Hurnik et al., 1995; Webster, 2000;
Henrikson et al., 2018).
Category Behavior Description
Activity MotionlessBird may be standing or in a crouched position where
its body is in contact with the floor of the crate
(difficult to determine via videos). Bird is motionless
and may be in a collected posture with its head
retracted, and eyes may be open or closed
Activity Active Bird is moving its feet and/or wings and is changing
position/location within the crate
Activity Shuffle Bird is moving its feet while moving its body side to
side. Minimum of 1 series of side-to-side movements to
be considered. Bird resets itself near the same position
as when behavior began
Thermoregulatory Burrow Birds are actively attempting to burrow underneath
other birds
Thermoregulatory Shiver Repeated quivering of the wings and/or body in order
to produce body heat
Thermoregulatory Pteroerection Feathers are erect or being ruffled in an organized
manner
Thermoregulatory Panting Increase in breathing of the bird characterized by an
open mouth, polypnea, or increased thoracic
movements
Thermoregulatory Survey Movements of the bird’s head suggesting surveillance
of the bird’s environment
Pecking Bird peck Bird is using its beak, in short and quick forward
motions of the head, to make contact with other birds
Pecking Object peck Bird is using the beak, in short and quick forward
motions of the head, to make contact with either the
crate or the sensors on the crate
Other Preen Beak is used to manipulate the feathers on the bird’s
own body
Other Head shake Rapid side-to-side movements of the head. Bird is
immobile. Minimum of 2 series of side-to-side
movements to be considered
Other Gulp Head of bird is pointed vertically upward, beak is
opened, and bird takes a large gasp of air
Other Scratch Bird raises its leg over its wing and repeatedly rubs its
head with its talons
Other Unidentifiable Bird cannot be seen, or behavior cannot be
characterized

Statistical Analyses Core Body Temperature

Data were analyzed as a randomized complete block There were no interactions between the T/RH combi-
design with the farm of origin as the block, and crate nations and D on pullet CBT. The change in pullet CBT
was used as the experimental unit. A block design was over time when exposed to each T/RH group is shown in
chosen to control variability arising from the farm or ge- Figure 2 (not statistically analyzed). The pullets exposed
netic line. Data analyses were performed using SAS 9.4 to 215 for the 4 h D demonstrated an overall decrease in
(SAS Institute, Cary, NC). Data were verified for CBT (negative D CBT values), which persisted over the
normality prior to analyses using Proc Univariate and entire D. However, pullets exposed to 215 for the 8 h D
were log transformed when necessary. They were demonstrated a decrease in CBT initially (negative D
analyzed via one-way ANOVA using the Proc Mixed pro- CBT values), but by the end of the exposure D pullets
cedure with means separation conducted using the Tukey were able to stabilize CBT, returning close to baseline.
test. Treatment means and pooled SEM were determined Additionally, pullets exposed to 30/80 for both the 4
via Proc Means. Differences were considered significant at and 8 h D had an initial increase in CBT (positive D
P  0.05 and trends were noted when 0.05,P  0.10. CBT values); however, the CBT plateaued over time
returning close to baseline values. Lastly, pullets exposed
to 21/30 demonstrated an increase in their CBT (D CBT
RESULTS approximately 1.3 C) by the end of the 8 h D, whereas
Chamber and Crate Environment pullets exposed for the 4 h D remained close to baseline
readings. Final CBT values (averaged from the final
The chamber T and RH achieved are shown in hour of exposure D) are shown in Table 3. For the last
Table 2. The crate (bird level) T/RH conditions varied hour of exposure, pullets exposed to 215 had a lower
slightly from the chamber exposure conditions and CBT compared to all other treatments (P , 0.01). How-
have been provided in Table 2. ever, when D CBT was calculated there was only a
 SIMULATED TRANSPORT OF WHITE LAYER PULLETS 701
Table 2. Average crate and chamber T ( C) and RH (%) conditions during simulated
transport of white-feathered layer pullets exposed to 5 T/RH combinations (215 C
uncontrolled RH, 21 C 30% RH, 21 C 80% RH, 30 C 30% RH, and 30 C 80% RH) for 4 or
8 h duration.
T/RH treatments
215 21/30 21/80 30/30 30/80
Crate T RH T RH T RH T RH T RH
4h 213.13 75.06 24.17 41.55 23.37 73.42 31.77 29.10 32.58 74.53
8h 210.90 76.17 23.14 42.51 23.29 73.88 32.11 27.06 32.53 75.28
Chamber 214.72 69.72 22.05 38.43 21.88 81.39 31.25 29.36 31.49 77.95

Abbreviation: T, temperature.

tendency toward significance (P 5 0.08). No effect of D Blood Physiology

was observed for the final CBT of pullets. Pullet D CBT
was influenced by D with a minor decrease in the 4 h D Initial and D values for each blood physiology param-
(20.11) and a minor increase in the 8 h D (0.53). eter are shown in Table 4. There were no interactions be-
tween T/RH and D on pullet blood physiology. There
were no differences in the initial blood physiology pa-
rameters in relation to T/RH or D. No effect of T/RH
Foot Temperature
was noted on change (D values) in pullet blood glucose,
In this study there was no interaction of T/RH and D sodium, hematocrit, hemoglobin, pH, BE, pCO2, pO2,
on pullet foot T. Final foot T was highest for pullets tCO2, sO2, and HCO32. In addition, the H/L ratio was
exposed to the 30/80 treatment (35.20 C) and lowest not affected by the T/RH treatments. There was no
for pullets exposed to 215 (12.97 C),with those exposed impact of D on pullet blood glucose, sodium, hematocrit,
to 21/30, 21/80, and 30/30 treatments being intermedi- hemoglobin, pH, BE, pO2, or sO2. Increased exposure D
ate (Table 3). There was no impact of exposure D on the resulted in a change in pullet blood pCO2, tCO2, and
foot T of pullets. HCO32. Pullet blood D pCO2 demonstrated a larger
decrease with 8 h D (27.88) compared with 4 h D
(20.99) (P 5 0.03). In addition, the tCO2 (P 5 0.02)
A 2.0
and HCO32 values (P 5 0.02) demonstrated a larger in-
1.5 crease with 4 h D compared with 8 h D. There was no
impact of D on pullet H/L ratio.
∆ Core body temperature (°C)

1.0
0.5
0.0 -15°C
21°C 30% RH Behavior
-0.5
21°C 80% RH
-1.0
30°C 30% RH
There were no interactions between T/RH and D on
-1.5
30°C 80% RH pullet behavior during simulated transportation. Pullet
-2.0 behavior is represented as a percentage of the time birds
-2.5 spent performing their respective behavior and is pre-
-3.0 sented in Table 5. There was no effect of either T/RH
0.0 1.0 2.0 3.0 4.0
Duration (h) treatment on the percentage of time birds spent perform-
ing the following behaviors: shuffle, burrow, shiver, pter-
B 2.0
1.5
oerection, object peck, head shake, scratch, and
unidentifiable. Pullets exposed to 215 and 21/30 spent
∆ Core body temperature (°C)

1.0
0.5 the most time motionless, compared to those exposed
0.0 -15°C to 30/30 and 30/80 (P , 0.01). Pullets exposed to 30/
-0.5 21°C 30% RH 80 spent more time actively compared with pullets
-1.0
21°C 80% RH exposed to 215, with pullets exposed to 21/30, 21/80,
-1.5
30°C 30% RH
and 30/30 being intermediate (P 5 0.01). Panting
30°C 80% RH
-2.0 occurred more frequently in pullets exposed to 30/30
-2.5 and 30/80, compared with all other treatments
-3.0
0.0 1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0
(P , 0.01). In both the 30/30 and 30/80 treatments, pul-
Duration (h) lets spent more time surveying compared to those
exposed to 215 (P , 0.01). Bird pecking occurred
Figure 2. Change (D) in the CBT of white-feathered layer pullets more frequently in pullets exposed to 30/30 compared
during simulated transport exposed to 5 temperature/RH combinations with 215 and 21/80 (P 5 0.05). Pullets exposed to
(215 C uncontrolled RH, 21 C 30% RH, 21 C 80% RH, 30 C 30% RH,
and 30 C 80% RH) for a duration of (A) 4 or (B) 8 h. D CBT
215 spent less time preening compared with pullets
( C) 5 mean pullet CBT2mean baseline CBT. Abbreviation: CBT, exposed to 21/80 and 30/30, with pullets exposed to
core body temperature. 21/30 and 30/80 being intermediate (P 5 0.02). Gulping
702 LALONDE ET AL.

Table 3. CBT and extremity T ( C) of white-feathered layer pullets during simulated transport exposed to 5 T/RH combinations (215 C
uncontrolled RH, 21 C 30% RH, 21 C 80% RH, 30 C 30% RH, and 30 C 80% RH) for a D of 4 or 8 h.
T/RH combinations D
Parameter 215 21/30 21/80 30/30 30/80 P-value 4h 8h P-value Interaction P-value SEM1
Final CBT2 38.24b 40.00a 39.91a 40.61a 40.75a ,0.01 39.64 40.15 NS NS 0.24
D CBT3 20.62 0.53 0.21 0.43 0.56 0.08 20.11b 0.53a 0.03 NS 0.16
Final foot temperature 12.97d 30.18c 32.48b 34.45a,b 35.20a ,0.01 27.47 27.75 NS NS 2.02
D Foot temperature4 215.28 22.24 3.37 4.00 6.47 NS 22.53 21.43 NS NS 2.02
a–d
Means within a parameter with different superscripts are significantly different (P  0.05).
Abbreviations: CBT, core body temperature; D, duration; NS, not significant; T, temperature.
1
Pooled SEM.
2
Average CBT for the final hour of exposure.
3
Delta (D) CBT 5 mean pullet CBT (last hour of exposure)2mean baseline CBT (15 min prior to exposure).
4
Delta (D) 5 final foot temperature2initial foot temperature.

was observed more frequently in pullets exposed to both are similar to those reported by Dadgar et al. (2012b),
neutral treatments (21/30 and 21/80) compared with where it was noted that broilers (5–6 wk of age) exposed
pullets exposed to 30/80 (P 5 0.03). Exposure D only to 214 C for a 3 h D had a lower CBT compared to those
affected active behavior, with no effects on any other exposed to either 29 C or 21 C for the same D. Dadgar
behavior. Pullets exposed to the 4 h D were more active et al. (2010) also noted a lower CBT in broilers (5–6 wk
than those exposed for the 8 h D (P 5 0.03). of age) exposed to 27 C for a 3 to 4 h D, compared with
broilers exposed to T above 5 C. Additionally, in the
DISCUSSION current study, pullets exposed to 215 C demonstrated
a negative D CBT value, indicating that CBT decreased
The microclimate (combination of T and RH) within as a result of cold exposure; however, pullets in the 8 h
the transport trailer and specifically within each crate treatment were successful at stabilizing their CBT,
can influence the CBT of birds during transport, which compared to those exposed to 215 C for the 4 h D.
may negatively impact bird welfare. Data from the cur- This may suggest that 4 h is not a long enough period
rent experiment indicate that CBT was lower during the for pullets to achieve thermal homeostasis. It is also
last hour of exposure in the 215 treatment. These results important to consider the welfare of pullets exposed to

Table 4. Blood physiology of white-feathered layer pullets during simulated transport with 5 T/RH combinations (215 C
uncontrolled RH, 21 C 30% RH, 21 C 80% RH, 30 C 30% RH, and 30 C 80% RH) for 4 or 8 h D.
T/RH combinations D
Parameter 1
215 21/30 21/80 30/30 30/80 P-value 4h 8h P-value Interaction P-value SEM2
i Glucose 13.16 13.19 13.18 13.23 13.34 NS 13.29 13.16 NS NS 0.10
D Glucose 20.13 20.62 20.65 20.25 0.45 NS 20.24 20.25 NS NS 0.16
i Sodium 144.36 144.00 143.83 144.36 144.17 NS 144.05 144.23 NS NS 0.35
D Sodium 24.05 20.83 20.27 0.64 0.11 NS 20.41 21.35 NS NS 0.80
i Hematocrit 25.25 24.67 25.83 26.42 25.78 NS 25.63 25.64 NS NS 0.29
D Hematocrit 21.92 22.00 22.00 21.67 21.95 NS 22.04 21.75 NS NS 0.24
i Hemoglobin 5.35 5.26 5.46 5.57 5.45 NS 5.42 5.42 NS NS 0.06
D Hemoglobin 20.42 20.47 20.42 20.34 20.41 NS 20.44 20.36 NS NS 0.05
i pH 7.11 7.12 7.11 7.10 7.09 NS 7.11 7.11 NS NS 0.01
D pH 0.05 0.03 0.06 0.08 0.07 NS 0.05 0.05 NS NS 0.01
i BE 29.39 28.50 29.00 210.33 210.44 NS 210.00 29.07 NS NS 0.37
D BE 0.83 1.61 2.56 3.47 3.11 NS 3.06 1.58 0.06 NS 0.47
i pCO2 62.97 65.35 64.52 62.63 63.63 NS 61.55 66.09 0.07 NS 1.22
D pCO2 26.48 22.62 23.89 25.12 24.07 NS 20.99a 27.88b 0.03 NS 1.77
i pO2 67.94 67.28 66.72 69.31 66.94 NS 67.06 68.22 NS NS 0.85
D pO2 12.62 4.66 2.34 20.17 2.56 NS 2.77 6.02 NS NS 2.22
i tCO2 21.94 22.94 22.56 21.25 21.33 NS 21.39 22.62 0.07 NS 0.33
D tCO2 0.22 0.84 1.33 1.89 1.73 NS 2.07a 0.16b 0.02 NS 0.50
i sO2 84.97 84.56 84.28 84.53 83.11 NS 84.19 84.39 NS NS 0.43
D sO2 4.47 3.38 3.11 3.22 4.28 NS 3.47 3.92 NS NS 0.56
i HCO32 20.08 21.00 20.51 19.35 19.39 NS 19.53 20.60 0.10 NS 0.31
D HCO32 0.05 0.89 1.56 2.15 1.91 NS 2.14a 0.45b 0.02 NS 0.46
i H/L 0.41 0.38 0.39 0.46 0.47 0.10 0.44 0.40 NS NS 0.03
D H/L 0.19 0.13 0.14 0.04 0.19 NS 0.16 0.13 NS NS 0.03

Means within a parameter with different superscripts are significantly different (P  0.05).
a,b

Abbreviations: BE, base excess; D, duration; H/L, heterophil to lymphocyte ratio; NS, not significant; pCO2, partial pressure of carbon dioxide;
pO2, partial pressure of oxygen; sO2, oxygen saturation; T, temperature; tCO2, total carbon dioxide.
1
Delta (D) values 5 final2initial (i). Glucose (mmol/L), sodium (mmol/L), hematocrit (%), hemoglobin (mmol/L), BE (mmol/L), pCO2
(mmHg), pO2 (mmHg), tCO2 (mmol/L), sO2 (%), H/L.
2
Pooled SEM.
 SIMULATED TRANSPORT OF WHITE LAYER PULLETS 703
Table 5. Behavior (% of time) of white-feathered layer pullets during simulated transport exposed to 5 T/RH combinations (215 C
uncontrolled RH, 21 C 30% RH, 21 C 80% RH, 30 C 30% RH, and 30 C 80% RH) for a D of 4 or 8 h.
T/RH combinations D
Behavior1 215 21/30 21/80 30/30 30/80 P-value 4h 8h P-value Interaction P-value SEM2
Motionless 93.46a 93.35a 90.10a,b 79.80b 68.45c ,0.01 85.43 84.63 NS NS 2.03
Active 0.12b 0.49a,b 0.99a,b 0.63a,b 1.82a 0.01 1.07a 0.55b 0.03 NS 0.18
Shuffle 0.34 0.80 1.41 0.95 0.94 NS 0.82 0.95 NS NS 0.16
Burrow 0.07 0 0 0.02 0 NS 0 0.04 NS NS 0.02
Shiver 0.10 0 0.10 0.02 0 NS 0 0.09 NS NS 0.03
Pteroerection 0.07 0 0 0.02 0.09 NS 0.04 0.04 NS NS 0.02
Panting 0b 0.23b 0.30b 9.41a 18.22a ,0.01 5.07 6.20 NS NS 1.73
Survey 1.24b 2.15a,b 2.27a,b 3.18a 4.11a ,0.01 2.76 2.42 NS NS 0.25
Object peck 0.05 0.37 0.02 0.38 0.32 0.07 0.23 0.23 NS NS 0.06
Bird peck 0b 0.16a,b 0b 0.29a 0.16a,b 0.05 0.10 0.15 NS NS 0.04
Preen 0.29b 0.78a,b 1.46a 1.19a 0.82a,b 0.02 1.01 0.81 NS NS 0.12
Head shake 0.19 0.28 0.54 0.38 0.07 NS 0.32 0.26 NS NS 0.06
Gulp 0.10a,b 0.44a 0.43a 0.23a,b 0.05b 0.03 0.20 0.29 NS NS 0.06
Scratch 0 0.14 0.07 0.22 0.19 NS 0.10 0.15 NS NS 0.04
Unidentifiable 4.01 1.00 2.31 3.31 4.95 NS 2.93 3.30 NS NS 0.75
a–c
Means within a parameter with different superscripts are significantly different (P  0.05).
Abbreviations: D, duration; NS, not significant; T, temperature.
1
Category: activity behaviors include motionless, active, and shuffle; thermoregulatory behaviors include burrow, shiver, pteroerection, panting,
and survey; pecking behaviors include object peck and bird peck; other behaviors include preen, head shake, gulp, scratch, and unidentifiable.
2
Pooled SEM.

cold T. Previous work has indicated that the normal blood flow to the dorsal pedal artery of the giant petrel
CBT of a bird can fluctuate by 61 C (Donkoh, 1989; after 40 s of being submerged in ice water and noted pro-
Sandercock et al., 2001; Dadgar et al., 2010); however, nounced vasoconstriction (Millard, 1974). However, the
it is unknown whether a decrease of 2 C to 2.5 C, as giant petrel is an arctic species and may have greater
seen in this study, is of concern. There was a minor effect adaptation to cold environments compared with com-
of D on CBT in this study. This may suggest that a 4 h D mercial layer pullets, making comparison with this study
is not sufficient for pullets to thermoregulate, as D CBT difficult. Frostbite of human skin has been noted to
was negative after 4 h and positive, meaning CBT occur at a skin T of approximately 22.2 C due to the
increased slightly, after 8 h of simulated transport. salinity of body fluids (Nagpal and Sharma, 2004). The
Conversely, Menten et al. (2006) noted that broilers threshold for frostbite for poultry is unknown and the
exposed to 35 C with 85% RH for 90 and 120 min had final foot T for the pullets in the current study was
a significantly higher rectal T compared to broilers approximately 13 C, suggesting that although frostbite
exposed to the same conditions for 30 or 60 min. Howev- may not have occurred, the birds were likely experi-
er, this may be attributed to differences in the thermo- encing discomfort due to a decrease in foot T of approx-
regulation ability of both pullets and broilers, as imately 15 C.
broiler have higher metabolic heat production Behavioral mechanisms used by birds to dissipate
(Sandercock et al., 2006). heat, such as panting (evaporative heat loss), can cause
Another way to evaluate thermal stress is via blood changes in the levels of blood gases as well as the acid–
flow to the extremities. With heat exposure, vasodilation base balance in the body (Ait-Boulahsen et al., 1989;
within the extremities can aid in dissipating heat from Sherwood et al., 2013; Khosravinia, 2017). Although
the core body (Whittow et al., 1964; Richards, 1971). panting was increased in pullets exposed to both 30 C
With cold exposure, vasoconstriction within the extrem- conditions, exposure to these conditions did not impact
ities can aid in conserving heat for the core body levels of blood gases, or acid–base balance parameters
(Wellehan, 2014). In the present study, pullet foot T in the body, demonstrating the pullets’ ability to cope
were recorded to examine the impact of thermal stress with thermal stress. There was, however, an effect of
on the extremities. We observed higher foot T in pullets exposure D on the blood pCO2, tCO2, and HCO32. Pul-
subjected to heat exposure, suggesting that the pullets lets exposed to the conditions for 8 h likely spent a
were experiencing mild thermal stress resulting in an in- greater period of time panting, resulting in decreased
crease in blood flow to the extremities. On the other levels of pCO2. However, increased exposure D and
hand, lower foot T in the pullets subjected to cold expo- increased time spent hyperventilating were not enough
sure (215 C) suggests that vasoconstriction to the ex- to impact the blood pH, pO2, sO2, or BE. There is very
tremities did occur. There has been minimal research little information regarding the effect of transportation
on observing the changes in the extremity T of poultry D on the blood physiology of poultry. A study by
when experiencing thermal stress. While it is unclear if Sandercock et al. (2001) exposed 35- and 63-day-old
a change in the extremity T is a welfare concern, it is broilers to either heat stress (32 C, 75% RH) or neutral
important to consider that transportation under cold conditions (21 C, 50% RH) for a 2 h D, and found that at
conditions could potentially result in frostbite. One both ages, the pCO2 was lower in the birds exposed to
study conducted in avian species observed decreased heat stress rather than neutral conditions. Other
704 LALONDE ET AL.

behavioral modifications include increasing surface area restriction) to hot (25 C–35 C), neutral (10 C–20 C),
to dissipate heat through radiation and convection. In and cold conditions (25 C to 5 C), and transported
the current work, birds exposed to both 30 C treatments them for a distance of 130 km inside a commercial truck.
were observed to be surveying more often and were more The authors found that broilers exposed to 25 C to 35 C
active. Surveying (in this case, meaning stretching or during transportation had higher concentrations of blood
elongating the neck through the crate sides) can increase glucose (13.35 mmol/L), compared to those exposed to
body surface area exposed to the environment, which 10 C to 20 C (12.41 mmol/L). In the current study, there
can aid in dissipating heat through non-evaporative was no impact of T/RH on the change in blood glucose; the
heat loss mechanisms including radiation and convection range observed was 3.16 to 13.34 mmol/L which was
(Whittow et al., 1964; Richards, 1971; Mount, 1979). To similar to the hot treatment results in the study by
the authors’ knowledge, there has been limited research Vosmerova et al., 2010. However, pullets were not feed
on examining the effects of thermal stress on the perfor- restricted before simulated transport which may have
mance of behaviors associated with non-evaporative allowed for more available glucose to respond to thermal
heat loss, as most heat stress studies focus on behaviors stress, compared with broilers which are typically feed
such as panting. While an increase in activity may result withdrawn prior to transportation as a meat quality pre-
in more heat being produced, it is hypothesized that the caution. Transportation in conjunction with thermal
pullets in this study were likely trying to increase the stress may also lead to dehydration and can be evaluated
distance between themselves and conspecifics within by examining markers in the blood such as sodium and
the crate. hemoglobin concentrations, and hematocrit
When exposed to cold T, birds will typically huddle (Khosravinia, 2017; Minka and Ayo, 2017). Pullets in
and burrow under conspecifics to minimize the amount our study were provided water up until the pretreatment
of body surface area exposed to the environment, which procedures, and neither the T/RH conditions nor an
can conserve heat and energy (Mount, 1979; Strawford increased exposure D was enough to result in changes in
et al., 2011). Strawford et al. (2011) exposed male the dehydration markers.
(32 d of age) and female (33 d of age) broilers to An increase in the H/L ratio has been recognized as an
25 C, 210 C, and 215 C for a 3 h D while inside trans- indicator of chronic stress in poultry (Lin et al., 2006).
portation crates, and found that the overall space usage Aksit et al. (2006) exposed broilers to either 15 C,
inside the crates did not differ compared to when birds 22 C, or 34 C for 2 h and noted a higher H/L ratio in
were crated pre-exposure. Additionally, burrowing ac- the broilers exposed to 34 C (0.81) compared to the
tivity did not differ, but the broilers were observed to broilers exposed to 15 C (0.40) or 22 C (0.30). Further-
be moving away from locations in the crates where drafts more, a study conducted by Altan et al. (2010) measured
were present during exposure (Strawford et al., 2011). the change in the H/L ratio in broilers after exposure to
Pullets in the current study were able to conserve heat heat stress. In their study, broilers were exposed to 38 C
and energy by spending more time motionless. However, for 3 h in their home pen at 35 and 36 d of age (feed was
the percentage of time spent performing thermoregula- removed for the exposure period) (Altan et al., 2010).
tory behaviors associated with cold exposure such as These broilers were noted to have a higher H/L ratio
burrowing, pteroerection, and shivering was not signifi- compared to broilers not exposed to the heat stress
cantly different between the T/RH treatments, suggest- (Altan et al., 2010). In the present study, there were
ing that the pullets were able to cope with the cold stress no impacts of either the T/RH conditions or exposure
through minimal behavioral changes. In addition, pullets D on pullet H/L ratio, which may be attributed, again,
were not feed-withdrawn which likely allowed them to to differences between broilers and layer pullets, or it
continue producing metabolic heat through digestion. may relate to pullets being able to cope successfully
There is minimal literature examining the perfor- with the exposure conditions and maintain homeostasis.
mance of bird pecking or gulping behaviors with regard The results from this study indicate that exposure to
to thermal stress. Bird pecking has been suggested to various T/RH conditions can influence the CBT of pul-
be an aggressive, stereotypic, or exploratory behavior, lets. However, pullets are able to successfully implement
but this primarily pertains to studies conducted in mechanisms to maintain homeostasis resulting in minor
pens or cages, rather than during transportation changes in the blood physiology and behavior. Addition-
(Webster and Hurnik, 1990; Webster, 2000). To the au- ally, a D of up to 8 h had minimal impact on the physi-
thors’ knowledge, there has been limited research on ology and well-being of pullets during simulated
gulping behaviors in poultry; therefore, the purpose transport. While the changes observed in this study
behind performing this behavior is not well known. How- such as decreased CBT and foot T are minor, it is impor-
ever, both bird pecking and gulping behaviors in the tant to note that they may still result in stress. It is also
current study were observed in a low frequency; there- important to note that very little is known regarding the
fore, they do not appear to have any biological relevance transportation of pullets, and comparisons with broilers
to either of these behaviors. or end-of-lay hens is likely inaccurate as pullets will differ
Thermoregulation against heat or cold stress can be metabolically and in feather cover. Further studies
energy-demanding, which could impact the blood glucose focusing specifically on pullets and relating to crating
concentrations (Sherwood et al., 2013). Vosmerova et al. densities in relation to external T, following the pullets
(2010) exposed broilers (42-day-old, and unknown feed through the laying cycle, and other transport stressors
 SIMULATED TRANSPORT OF WHITE LAYER PULLETS 705
such as noise, vibration, and handling would provide Dadgar, S., T. G. Crowe, H. L. Classen, J. M. Watts, and
better insight on reducing transport-related stress. P. J. Shand. 2012a. Broiler chicken thigh and breast muscle re-
sponses to cold stress during simulated transport before slaughter.
Poult. Sci. 91:1454–1464.
Dadgar, S., E. S. Lee, T. G. Crowe, H. L. Classen, and
ACKNOWLEDGMENTS P. J. Shand. 2012b. Characteristics of cold-induced dark, firm, dry
broiler chicken breast meat. Br. Poult. Sci. 53:351–359.
The National Sciences and Engineering Research Donkoh, A. 1989. Ambient temperature: a factor affecting perfor-
Council of Canada, Egg Farmers of Canada, Olymel mance and physiological response of broiler chickens. Int. J. Bio-
meteorol. 33:259–265.
S.E.C., and Maple Lodge Farms are graciously acknowl- Elsayed, M. A. 2014. Effects of length of shipping distance and season
edged for their financial support. of the year temperature stress on death rates and physiological
condition of broilers on arrival to slaughterhouse. J. Nucl. Tech.
Appl. Sci. 2:453–463.
DISCLOSURES Felver-Gant, J., L. A. Mack, R. L. Dennis, S. D. Eicher, and
H. W. Cheng. 2012. Genetic variations alter physiological re-
The authors declare that they have no affiliations with sponses following heat stress in 2 strains of laying hens. Poult. Sci.
or involvement in any organization or entity with any 91:1542–1551.
Henrikson, Z. A., C. J. Vermette, K. Schwean-Lardner, and
financial interest (such as honoraria; educational grants; T. G. Crowe. 2018. Effects of cold exposure on physiology, meat
participation in speakers’ bureaus; membership, employ- quality, and behavior of Turkey hens and toms crated at transport
ment, consultancies, stock ownership, or other equity density. Poult. Sci. 97:347–357.
interest; and expert testimony or patent-licensing ar- Hurnik, J. F., A. B. Webster, and P. B. Siegel. 1995. Dictionary of Farm
Animal Behaviour. 2nd ed. Iowa State Univeristy Press, Ames, IA.
rangements), or non-financial interest (such as personal Khosravinia, H. 2017. Physiological responses of newly hatched broiler
or professional relationships, affiliations, knowledge or chicks to increasing journey distance during road transportation.
beliefs) in the subject matter or materials discussed in Ital. J. Anim. Sci. 14:519–523.
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