2003 Poultry Science Association, Inc.
Criteria for Evaluating Husbandry Practices to Alleviate Heat Stress in Broilers ¨ zkan,* M. C S. Yalc¸in,*,1 S. O ¸ abuk,† and P. B. Siegel‡ *Department of Animal Science, Ege University, Faculty of Agriculture, Izmir-Turkey; †Celal Bayar University, Akhisar Vocational Training School, Manisa-Turkey; and ‡Department of Animal and Poultry Science, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24060-0306
Primary Audience: Researchers, Broiler Managers, Extension Personnel SUMMARY This study was undertaken to investigate the effects of management techniques used under heat stress on growth as well as developmental instability, duration of tonic immobility (TI), and heterophil-to-lymphocyte ratios (H:L) of broilers. Three-hundred sixty cockerels obtained from a commercial hatchery were randomly assigned to 12 floor pens. When the cockerels reached 21 d of age, three pens were kept as controls. Broilers in the other nine pens were moved to a room heated from 32 to 35°C between 1000 and 1700 h each day from 21 to 42 d of age. Broilers in the heated room were randomized into three treatment groups consisting of 1) conditioned (chicks exposed to 36°C for 24 h at 5 d of age), 2) feed-restricted (during the heat stress, feed was withdrawn 2 h before the hot period, and chicks were fed between 1700 and 0800 h), 3) and heat-stressed only. Conditioned and feed-restricted broilers gained 3.2 and 2.8%, respectively, more BW than heat-stressed broilers. Relative asymmetry (RA) averaged across several bilateral traits for the prolonged heat stressor was more informative than the RA for a single bilateral trait. Corrrelations suggest that RA were not closely associated with duration of TI and H:L under the conditions of this experiment. Key words: broiler, heat stress, relative asymmetry, heterophil-to-lymphocyte ratio, tonic immobility 2003 J. Appl. Poult. Res. 12:382–388
DESCRIPTION OF PROBLEM Duration of tonic immobility (TI) has been used as a measurement for evaluating fearful behavior and may be used as a criterion for measuring well-being and levels of stress of chickens [1]. Because heterophil-to-lymphocyte ratios (H:L) increase under stressful conditions [2, 3, 4], the ratios have been used as an index 1
of the responses of hypothalamic-hypophysealadrenal axis to the stressors in chickens [2]. Although positive correlations between H:L and TI have been reported by several investigators [5, 6, 7], there was no correlation between H:L and TI from different preslaughter handling methods of broilers [8]. Developmental stability may also serve as a noninvasive means for assessing the degree to which an individual is able to buffer its development when stressed [9]. Al-
To whom correspondence should be addressed:
[email protected].
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TABLE 1. Mean BW (g), BW gain (g/d), and mortality (%) of broilers by age (d) and treatment BW
Treatment Control ConditionedA Food restrictionB Heat stressC SEM
BW gain
5d
7d
14 d
21 d
42 d
5 to 14 d
5 to 21 d
21 to 42 d
Mortality 0 to 42 d
** 92 88 — — 0.8
** 135 117 — — 1
NS 346 338 — — 4
NS 718 712 — — 7
** 2,159a 2,025b 2,018bc 1,961c 23
NS 12.1 11.9 — — 0.1
NS 39.1 38.9 — — 0.4
** 68.4a 62.4b 62.7bc 58.8c 0.9
NS 3.33 3.33 4.44 7.77 1.84
Means in the same column within a day with no common superscript differ significantly (P < 0.05). Broilers conditioned at 5 d of age, then heat stress given daily from 28 to 42 d of age. Food restriction was from 28 to 42 d of age during heat stress. C Heat stress was applied daily from 28 to 42 d of age. **P < 0.001. a–c A B
though developmental stability cannot be measured directly, asymmetry in terms of deviations from a perfect symmetry provides information on the level of responses of chickens to suboptimal environmental conditions [10]. Genetic [11, 12] and environmental stressors such as rearing density [13] and lighting regimens [10, 14] may increase asymmetry of bilateral traits in poultry. Because relative asymmetry (RA) may provide information on animal well-being [10, 14], relationships between different RA and with TI and H:L may exist. Reports, however, are contradictory. Although TI was positively correlated with RA in chickens reared under different environmental conditions [10, 13], comparisons of RA with TI and H:L led to the conclusion that an RA did not provide a general tool to assess fear or stress susceptibilities in chickens [15, 16]. Effects of heat stress as an environmental stressor on performance of broilers has been well-studied [e.g., 17, 18, 19]. It was suggested
[20] that H:L could be used as a criterion to select for resistance to heat stress and that TI was less in heat-stressed than in control hens [21]. Several management techniques have been used to alleviate the detrimental effects of heat stress on broiler performance [e.g., 22, 23, 24]. We measured developmental stability of bilateral traits in three broiler stocks when conditioned or feed-restricted under heat stress [25]. Conditioning to heat stress at 5 d of age lowered RA of P. major weight and appeared to decrease some of the adverse effects of heat stress at subsequent ages. Generally lacking, however, are comparisons within an experiment that evaluate different measurements to facilitate the choice of criteria for husbandry practices to alleviate heat stress in broilers. The experiment reported here was designed to compare several methods used to alleviate the detrimental effects of heat stress on RA, TI, and H:L in broilers. In
TABLE 2. Mean length and width of shank and length of face (mm) by age (d) and treatment Shank length
Treatment Control ConditionedA Food restrictionB Heat stressC SEM
Shank width
Face length
21 d
42 d
21 d
42 d
21 d
42 d
NS 66.2 66.1 — — 0.4
NS 95.9 95.0 97.7 97.4 1
NS 11.9 11.7 — — 0.1
* 15.3b 15.8ab 15.9a 16.2a 0.2
NS 20.5 20.7 — — 0.2
NS 25.1 24.6 24.1 24.3 0.3
Means in the same column within a day with no common superscript differ significantly (P < 0.05). Broilers conditioned at 5 d of age, then heat stress given daily from 28 to 42 d of age. Food restriction was from 28 to 42 d of age during heat stress. C Heat stress was applied daily from 28 to 42 d of age. *P < 0.05. a,b A B
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TABLE 3. Mean ± SEM for left-right differences of bilateral traits and types of asymmetry by age (d) and treatment Shank length (mm) Treatment Control ConditionedC Food restrictionE Heat stressF
21 d
42 d
Shank width (mm) 21 d
Face length (mm)
42 d
0.587 ± 0.194 −0.696 ± 0.419 0.117 ± 0.053 −0.190 ± DAA FAB DA FA 0.263 ± 0.343 −0.790 ± 0.551 0.054 ± 0.111 −0.133 ± FA FA FA FA — −0.452 ± 0.656 — −0.419 ± FA FA — −1.757 ± 0.530 — −4.619 ± FA FA
21 d
42 d
0.119 −0.004 ± 0.147 −0.152 ± 0.270 FA FA 0.112 −0.272 ± 0.205 −0.547 ± 0.282 ASD FA 0.168 — 0.100 ± 0.324 FA 0.109 — −0.214 ± 0.360 AS
A
Mean is different from zero with a normal distribution. Mean zero with a normal distribution. Broilers conditioned at 5 d of age, then heat stress given daily from 28 to 42 d of age. D Mean zero with a distribution not normal. E Food restriction was from 28 to 42 d of age during heat stress. F Heat stress was applied daily from 28 to 42 d of age. B C
addition, the relationships among these criteria were compared.
MATERIALS AND METHODS A total of 360 male broiler chicks obtained from a commercial hatchery were wingbanded, weighed, and randomly assigned to 12 floor pens, each with 30 birds. The brooding temperature was maintained at approximately 32°C for the first 3 d, then decreased 3°C weekly through 21 d. Water and feed were available ad libitum. When the cockerels reached 21 d of age, three pens were kept as controls. Broilers in the other nine pens were moved to a room heated from 32 to 35°C between 1000 and 1700 h each day from 21 to 42 d of age. The average nighttime temperature during this period was 26°C. Corresponding temperatures in the control room averaged 23°C during the day and 20°C during the night. During the 3-wk heat stress period, humidity ranged from 60 to 72% and from 47 to 55% in the heated and control rooms, respectively. Broilers in the heated room were randomized into three treatment groups consisting of 1) conditioned (chicks exposed to 36°C for 24 h at 5 d of age), 2) feed-restricted (during the heat stress, feed was withdrawn 2 h before the hot period, and chicks were fed between 1700 and 0800 h), and 3) heat-stressed only. There were three replicate pens per treatment. Broilers were fed a commercial starter diet in mash form with 23.5% CP and 3,120 kcal ME/kg from 0 to 10 d of age, a grower diet in
pellet form with 22.5% CP and 3,130 ME/kg from 11 to 21 d, and a finisher diet in pellet form with 20.5% CP and 3,240 kcal ME/kg from 22 to 42 d. The lighting regimen was 23L:1D throughout. Chicks were individually weighed at hatch after conditioning on the fifth day, and at 7, 14, 21, and 42 d of age. Daily BW gain was calculated for the periods 5 to 14, 5 to 21, and 21 to 42 d of age. Twenty-one broilers (7/replicate) from each group were tested for TI before heat stress at 21 d and after heat stress at 42 d of age. Broilers were carried to a separate room and subjected to TI. The procedure [8] involved placing the broiler on his back and restraining him for 15 s by placing one hand on its sternum. A stop watch was started to record latencies until he righted himself. If the broiler righted itself in less than 10 s the restraining procedure was repeated. When there was no righting response over the 10 min test period, a maximum score of 600 s was assigned [5]. Blood samples were taken from wing vein of the same broilers tested for TI. Blood smears were stained using May-Gru¨nwald and Giemsa stains, approximately 4 h after preparation with methyl alcohol fixation. Heterophils and lymphocytes were counted by the same person to 60 cells per individual and the H:L ratio calculated [2]. Left and right shank length, shank width, and face length of the same broilers were measured at the same ages (21 and 42 d) in millimeters using a digital caliper. Trait length
3.72 ± 4.90 ± 5.39 ± 4.70 ± 4.39
42 d
0.59 056 061 0.85
21 d
3.08 ± 0.19 2.94 ± 0.33 — — 0.17
Mean RA
0.49 0.50 0.79 0.50 2.99 ± 2.49 ± 4.31 ± 3.49 ± 4.29 B
Broilers conditioned at 5 d of age, then heat stress given daily from 28 to 42 d of age. Food restriction was from 28 to 42 d of age during heat stress. C Heat stress was applied daily from 28 to 42 d of age. *P < 0.05.
RESULTS AND DISCUSSION
A
3.04 ± 0.28 3.57 ± 0.59 — — 0.07 0.29 0.36 0.40 0.47 1.57 ± 2.22 ± 2.00 ± 2.22 ± 1.76 1.71 ± 0.20 1.94 ± 0.30 — — 0.33
21 d 42 d 21 d Treatment
Control ConditionedA Food restrictionB Heat stressC χ2
42 d
(%)
21 d
3.91 ± 0.59 3.30 ± 0.77 — — 0.80
Face length Shank width Shank length
TABLE 4. Means ± SEM relative asymmetry (RA) of shank length, shank width, and face length by age (d) and treatment
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and width was the mean of the left and right sides. Three categories were used for left-right differences. Fluctuating asymmetry (FA) was defined as mean zero and normal distribution, directional asymmetry (DA) was defined as mean not zero and normal distribution, and antisymmetry (AS) was defined as mean zero with a distribution that was not normal [26, 27]. Relative asymmetry was defined as the ratio of the absolute value of left-right divided by the value for the size of the trait. Mean RA was defined as the mean RA of the different traits. Data were subjected to one-way of ANOVA test. Shapiro-Wilk and t-statistics were used to test for normality of distribution with a mean of zero. The effects of treatments on RA were tested by Kruskall-Wallis one-way analysis [28]. H:L and TI were transformed to arc sine square root of the ratio and natural logarithms, respectively, before being subjected to ANOVA. All data are presented as untransformed. Relationships of RA with TI and H:L were assessed by product moment correlation analysis. Significance was considered at P < 0.05.
3.01 ± 3.21 ± 3.99 ± 3.85 ± 7.41*
42 d
0.26 0.22 0.27 0.38
YALC¸IN ET AL.: EVALUATING HEAT STRESS
Least-squares means of BW, gains in BW, and mortality are presented by age and treatment in Table 1. Although BW among treatment groups at hatch were similar, conditioning at 5 d of age resulted in a decrease in BW at the end of the fifth day and at d 7. These results were consistent with those of Yahav and Hurwitz [22], who reported exposure to high temperature at 5 d of age resulted in growth retardation. The differences between control and conditioned broilers, however, were transitory and not significant at 14 and 21 d. The compensatory growth observed on d 14 was earlier than that obtained by Yahav and Hurwitz [22]. Exposure to heat stress during a period from 21 to 42 d depressed BW by about 9%. This decrease in BW was less than that previously reported [19] and may be due to the period when the broilers were exposed to the heat stressed. Across treatments the lowest BW at 42 d of age was for heat-stressed broilers and heaviest for the controls. Conditioned and feedrestricted broilers gained 3.2 and 2.8%, respectively, more BW than those that were heatstressed. Thus, BW of these groups were brack-
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TABLE 5. Mean of durations of tonic immobility (TI), number of inductions needed to induce TI, and heterophilto-lymphocyte ratios (H:L) by age (d) and treatment Duration of TI (s)
Inductions (n)
H:L
Treatment
21 d
42 d
21 d
42 d
21 d
42 d
Control ConditionedA Food restrictionB Heat stressC SEM
NS 228 222 — — 38
* 175b 191b 200b 292a 40
NS 1.32 1.24 — — 0.08
NS 1.47 1.14 1.47 1.19 0.12
* 0.41b 0.51a — — 0.03
* 0.43b 0.49ab 0.41b 0.55a 0.04
Means in the same column within a day with no common superscripts differ significantly (P < 0.05). Broilers conditioned at 5 d of age, then heat stress given daily from 28 to 42 days of age. B Food restriction was from 28 to 42 d of age during heat stress. C Heat stress was applied daily from 28 to 42 d of age. *P < 0.05. a,b A
eted by the other two treatments (Table 1). Gains in BW between 21 and 42 d of age followed a similar pattern to that just justified. There was no difference between treatments for mortality. Shank length, shank width, and face length at 21 d of age were similar for all groups (Table 2). At 42 d there were no differences among treatments for shank length or face length. Shank width was greater in heat-stressed and feed-restricted than control broilers, with the conditioned group intermediate. Although all three types of asymmetry were observed for the bilateral traits measured, in 14 of 18 cases it was FA (Table 3). Relative asymmetries ranged from 1.57 to 5.39% (Table 4). Comparisons between treatments for the association between RA for any single trait were not significant; however, the mean RA for all traits measured at 42 d of age were different, being lowest for the control, followed by the conditioned group. Largest mean RA were for
the feed-restricted and heat-stressed broilers. These results demonstrate that an RA averaged across several bilateral traits for a prolonged stressor may be more informative than the RA for a single bilateral trait. This observation is logical because growth of body parts differ with age. On d 21 both duration of and number of inductions for TI were similar in control and conditioned broilers (Table 5). At 42 d of age, duration of TI was longer for heat stressed broilers than for those in the other groups, which did not differ. There were no differences among groups for numbers of induction to induce TI at 42 d. Conditioning resulted in an increased H:L at 21 d of age, showing that conditioning at 5 d was a stress that persisted to an older age. At 42 d H:L was higher for heat-stressed than for feed-restricted and control broilers, with conditioned broilers intermediate (Table 5).
TABLE 6. Correlation coefficients between durations of tonic immobility (TI) and heterophil-to-lymphocyte ratios (H:L) and mean relative asymmetries (RA) of bilateral traits at 21 and 42 d of age by treatment TI and H:L
TI and RA
H:L and RA
Treatment
21 d
42 d
21 d
42 d
21 d
42 d
Control ConditionedA Food restrictionB Heat stressC
0.05 0.41 — —
0.01 0.38 0.06 0.38
−0.04 −0.21 — —
0.01 0.09 0.07 0.11
0.01 −0.28 — —
−0.04 0.01 −0.15 0.04
A
Broilers conditioned at 5 d of age, then heat stress given daily from 28 to 42 d of age. Food restriction was from 28 to 42 d of age during heat stress. C Heat stress was applied daily from 28 to 42 d of age. B
YALC¸IN ET AL.: EVALUATING HEAT STRESS In the present study, high ambient temperatures between 21 and 42 d of age resulted in higher RA, greater H:L, and longer TI. Duration of TI and mean RA of different bilateral traits at 21 d of age were similar for control and conditioned broilers, indicating that conditioning at 5 d of age did not contribute to fearfulness and developmental instability of the bilateral traits measured. Moreover, the results suggest that conditioning resulted in less developmental instability and fearfulness at 42 d of age than that observed in heat-stressed broilers that were not conditioned. The higher H:L in conditioned broilers at 21 d of age indicated that conditioning modifies responses to stressors in broilers. Food restriction resulted in higher RA, but it yielded TI and H:L similar to controls. This result was consistent with
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findings that chickens adapted to repeated fasts [3]. No significant correlations were obtained between of TI, H:L, and mean RA within treatment groups (Table 6). All correlations between TI and H:L were positive with the one at 21 d for conditioned broilers approaching significance (P = 0.06). When all groups were taken into consideration at 42 d of age, there was a correlation of 0.21 between duration of TI and H:L (P = 0.01, n = 84 [data not shown]). Under the conditions of this experiment, it appears that RA may not be closely associated with TI and H:L, which is consistent with observations reported by Campo et al. [15]. The results also show that it is possible to reduce the effects of heat stress by conditioning and feed restriction.
CONCLUSIONS AND APPLICATIONS 1. Conditioning at 5 d of age did not influence BW gain between 5 and 14 or 5 and 21 d of age. Conditioning at 5 d of age or feed restricton during a daily heat stress from 21 to 42 d reduced the detrimental effects of heat stress on growth of broilers. 2. Conditioning and feed restriction resulted in a decrease in average RA of bilateral traits, TI, and H:L of broilers exposed to prolonged heat stress. 3. Based on the results of this experiment, RA may not be closely associated with TI and H:L. 4. Conditioning at a young age and feed restriction during a subsequent heat stress reduce the deleterious effects of heat stress.
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