Dietary Inclusion of Chromium to Improve Growth Performance and Immune-Competence of Broilers Under Heat Stress

Abstract

This study was conducted to investigate the effects of dietary supplementation with chromium chloride, CrCl₃.6H₂O (2mg kg^–1 basal diet) on the performance and immune response of broiler chickens under heat stress condition (25-43°C). A total of 80 one-day-old broiler chicks (Ross-308) were assigned to two treatment groups according to a completely randomized design. Each treatment consisted of four equal replicates, each contained ten chicks. Chicks were fed on basal diets supplemented with different concentrations of chromium (0 and 2 mg kg^–1 CrCl₃) from 1 to 35 days of age. Chromium supplementation as feed additives resulted in a slightly lower rectal temperature, and significantly (P<0.05) lower respiration rate for the broiler chickens received diet supplemented with chromium compared to the control (0 mg kg^–1 CrCl₃). Dietary chromium supplementation increased final body weight (BW) at the end of the production period (5 weeks). Average weight gain was significantly (P<0.05) higher in chickens fed on chromium supplemented diet. Feed intake was not influenced by dietary chromium supplementation, however, the efficiency of feed conversion was improved (P<0.05) in chromium supplemented chickens. Furthermore, dressing percentage was significantly (P<0.05) higher in Cr-treated chickens compared to control chickens. Chromium supplementation significantly (P<0.05) improved the immune response to Newcastle Disease Virus vaccine (NDV). The present results suggest that dietary chromium supplementation provides a good nutritional management approach to ameliorate heat stress induced depression in production performance and immune response of broiler chickens.

KeyWords:

• Broilers
• Chromium
• Growth performance
• Heat stress
• Immune response

Introduction

Poultry production in tropical countries faces a constant challenge. Heavy economic losses have been reported due to increased mortality and decreased productivity in response to the stress caused by the hot climate. Heat stress interferes with broiler comfort and suppresses productive efficiency (Al-Fataftah and Abu-Dieyeh, 2007). It has been postulated that heat stress reduces growth performance (Lara and Rostagno, 2013) and immune response (Bartlett and Smith, 2003). The improvement in poultry production has been established through new concepts of feeding. The immune system benefits greatly from proper nutrition of birds (Prieto and Campo, 2010). Researchers are challenged to find ways of increasing feed efficiency and lowering feed cost in order to lower the overall poultry cost. In response to these concerns and in order to give producers practical solution to the high production cost, many studies were carried out in many parts of the world focused on feed additives as a mean to promote growth and increase feed efficiency in poultry (Bartlett and Smith, 2003; Niu et al., 2009; Prieto and Campo, 2010). Chromium (Cr) is not currently considered an essential trace element for poultry; this micronutrient may play a nutritional and physiological role. Moreover, the National Research Council (NRC) has recommended 300 µg Cr kg^–1 diet for laboratory animals (NRC, 1995). Currently there are no NRC recommendations for Cr in poultry diets (NRC, 1994) and most poultry diets are basically composed of ingredients of plant origin, which have usually low content of chromium (Giri et al., 1990). Supplementation with an organic source of Cr such as chromium nicotinate (Toghyani et al., 2012) proved to be beneficial to broiler chickens under heat stress despite lowered feed consumption.

Two general forms of chromium have been used in supplementation trials, the inorganic form chromium chloride (CrCl₃) and organic forms that seem to have greater availability (Lukaski, 1999; Ghazi et al., 2012). However, because small concentrations are required, chromium chloride and not organic forms may be an economical source in many diets. Plant products contain a low content of chromium, implying that poultry and animals may have a deficiency of chromium because their diets consist either of all or a large proportion of plant ingredients (Gibson, 1990). Heat stress reduces growth rate, feed consumption, egg production and feed efficiency in poultry (Lara and Rostagno, 2013). Moreover, it has been reported that plasma chromium concentration was greatly decreased and its urinary excretion was increased during stress and diseases (Anderson et al., 1988; Morris et al., 1992; Uyanik et al., 2002). Therefore, the main objective of this study was to evaluate the effects of feeding a diet supplemented with chromium chloride (2 mg kg^–1) on production performance and immunity status of broiler chickens under heat stress conditions.

Materials and methods

This study was conducted on a total of 80 one-day-old broiler chicks (Ross-308) during the summer season (25 to 43°C). The chicks were kept in battery cages housed in an open-sided poultry house. The chicks were divided into two treatments group of 40 birds each (A and B). Each treatment group comprised of four equal replicates (10 birds/replicate). The birds were fed a basal diet (group A) or basal diet supplemented with 2 mg chromium chloride kg^–1 basal diet (group B).The birds were vaccinated against Newcastle disease virus (NDV). Live Newcastle disease vaccine (Lasota strain) was administrated via drinking water at 8 and 24 day of age.

Experimental feed

The feed was formulated according to National Research Council (NRC, 1994), to meet or exceed the nutrients requirements of broiler chickens. Ingredient and chemical compositions of the basal diets are shown in table 1 and 2, respectively. The birds were fed with starter diet until 21 days of age. Thereafter, they were fed with a finishing diet up to the end of the experiment. Feed and fresh clean tap water was offered ad libitum throughout the experiment.

Production performance measurements

Initial body weights for the bids were measured at the first experimental day using digital balance. Final body weight was measured on individual basis at the end of 5^th week, then averaged and expressed as g/bird/treatment. Daily average weight gain was calculated from the weight difference on individual basis, then averaged and expressed as g/day/bird/treatment. Weekly feed intake was determined by subtracting the weight of feed refusal from the weight of offered feed using digital balance. Feed intake was measured at weekly interval on replicate basis, and then average daily feed intake was expressed on individual basis as g/day/bird/treatment. Feed conversion ratio was calculated weekly computed from feed intake and weight gain of individual birds, and then expressed as average (FCR = Feed intake/Weight gain). All chickens were weighed before and after slaughter to calculate the dressing percentage. The internal organs, legs, head and feather were removed, and the carcass was weighed to calculate the dressing percentage as follow: Dressing (%) = Eviscerated carcass weight x 100/Live body weight.

Thermal and immunological measurements

Rectal temperature (T_r) and respiration rate (RR) were measured throughout the experimental period twice weekly at 14:00 h using digital thermometer and by visual counting of the movement of the vent per minute, respectively. The measurements were then expressed as average. Blood samples were collected weekly throughout the experimental period by brachial venipuncture in plain and EDTA-coated vacutainers. Heterophil/lymphocyte ratio was determined microscopically (immersion lens x 100) using fresh blood smear stained by Giemsa-May Günwald. Serum cortisol level was determined by commercial kit (125I-F Radioimmunoassay Kit- IMK- 484). Serum antibody titer against Newcastle disease virus was determined by haemagglutination test (HA). The test was performed on undiluted Lasota virus suspension. The last well showing HA was considered one HA unit and accordingly, the 4HA units were calculated. The virus suspension was diluted to contain 4HA units per 25 µL. Volumes of 25 µL were transferred to each well of the micro plate, then 25 µL of serum samples under test were placed in the first row of wells and two-fold serial dilutions were made. Volumes of 25 µL 4HA unit were placed in each well. Two rows of wells were left as controls. One row contained serial dilution of the antigen to confirm the 4HI units and the other row contained RBCs. The plates were then left for 30 min at room temperature for antigen/antibody reaction to take place. After that volume of 25 µL (1%) RBCs suspension was added to each well. The test was read after 30 min incubation at room temperature. Heamagglutination inhibition titer was expressed as the reciprocal of the highest dilution that gave 50% inhibition of agglutination (Allan et al., 1978).

Statistical analysis

The data of the study were arranged in a complete randomized design (CRD). The data were analyzed using the General Linear Models (GLM) procedure of SAS software (SAS, 2003). Significant differences (P<0.05) among the means were determined using Duncan’s test. The data are presented as Means ± standard deviations (SD) of the means.

Results

Climatic data

The climatic data recorded during the current study shows that broiler chickens were exposed to high average ambient temperature (T_a = 34.2±0.84ºC) with minimum value of 25°C and maximum value of 43ºC, and an average relative humidity of 43.6±13.28%. This resulted in a higher calculated average temperature humidity index (THI) of 30.73±0.80.

Thermoregulation

The average rectal (T_r) temperature throughout the study period (5 weeks) was slightly lower in Cr-supplemented broiler chickens (41.5ºC) compared to the control (41.6ºC) group (Figure 1a). However, the average respiration rate was significantly (P<0.05) lower in Cr-supplemented broiler chickens (64 breath/min) compared to the control (70 breath/min) group (Figure 1b).

Production performance

Dietary chromium supplementation resulted in a significantly (P<0.05) higher average final live body weight, average daily gain (ADG), feed conversion ratio (FCR) and carcass dressing percentage of Cr-supplemented broiler chickens compared to those of control group. However, the average daily feed intake was not influenced by chromium supplementation (Table 3).

Immune response

Inclusion of CrCl₃ (2 mg kg^–1) in the diet of broiler chickens has potentiate the immune response of the broiler chickens to vaccination. The average antibody titer to Newcastle Disease Virus (NDV) vaccine was significantly (P<0.05) improved in broiler chickens supplemented with chromium (721) compared to the control (593) chickens (Figure 2). Furthermore, the average heterophil/lymphocyte ratio (Figure 3a), and average serum cortisol level (Figure 3b) were significantly (P<0.05) lower in Cr-treated broiler chickens (0.44 and 5.6 nmol/L, respectively) compared to the control group (0.57 and 8.4 nmol/L, respectively).

Discussion

This investigation describes the effect of dietary inclusion of chromium chloride (CrCl₃) on the production performance, thermoregulatory responses and immune response of broiler chickens under heat stress conditions. The recorded climatic data indicated that broiler chickens were exposed to chronic heat stress throughout the experimental period (T_a = 34.2±0.84ºC; THI = 30.73±0.80).

As expected there was increase in rectal temperature during heat stress in chickens fed no supplemental chromium (data not shown). While there was no marked effect of dietary chromium on rectal temperature, there was a slight decrease in rectal temperature of Cr-supplemented heat-stressed broiler chickens. Similar but significant response was observed for respiration rate indicating that dietary Cr protects broiler chickens against heat stress.

The results of the current study revealed that Cr supplementation at the level of 2 mg kg^–1 basal diet could alleviate the negative effects of heat stress on broiler chickens’ production performance. It has been reported that growth rate and feed utilization efficiency of birds decrease when the ambient temperature increases over the thermoneutral zone of 19 to 23ºC (Geraert et al., 1996; Sahin et al., 2002). Broiler chickens fed Cr-supplemented diet had higher average daily gain and better feed conversion ratio, but average daily feed intake was not significantly improved by Cr-supplementation. Bona et al. (2011) reported that higher dietary chromium supplementation (1 mg Cr kg^–1) improves insulin sensitivity. Therefore, the higher dose of Cr used in this study could have increased insulin sensitivity of heat-stressed broiler chickens and consequently improved carbohydrate and lipid metabolism, which are known to be adversely affected under heat stress conditions (Nam et al., 1995). Thus, the impacts of heat stress on the production performance of the experimental broiler chickens were ameliorated by dietary chromium supplementation. Similar to the results reported in the current study, Sahin et al. (2002) observed that dietary organic Cr supplementation at the level of 200 to 1200 µg kg^–1 had increased body weight and feed efficiency in broilers reared under heat stress. On the other hand, Toghyani et al. (2012) reported that dietary organic Cr supplementation had increased body weight and feed intake without affecting feed efficiency in heat stressed broiler chickens. However, the results reported here are contradictory to those reported by Moeini et al. (2011) and Ghazi et al. (2012) who showed no differences in body mass and feed conversion ratio of Cr-supplemented heat stressed broiler chickens. The lack of consistency among these studies might be explained by the different chromium source and supplementation levels applied. Increased body fat deposition and decreased protein retention have been reported in heat stressed broiler chickens (Geraert et al., 1996). Accordingly, dietary chromium supplementation in the present study had improved the dressing percentage and carcass yield of broiler chickens. It has been shown in animal studies that Cr supplementation could increase muscle gain and decrease fat gain (Anderson, 1994; Zha et al., 2009). It is well established that Cr is involved in protein metabolism (Anderson, 1987) and regulate fat metabolism by potentiating insulin action (Vincent, 2000).

Dietary chromium supplementation had significantly reduced serum cortisol concentration and consequently elevated antibody response to NDV of heat-stressed broiler chickens. Suppression of antibody production in broilers under heat stress has been reported by Bartlett and Smith (2003) and Niu et al. (2009). This reduction in antibody production under heat stress could be due to the increase in inflammatory cytokines, with the subsequent stimulation of hypothalamic production of corticotrophin releasing hormone (Trout and Mashaly, 1994; Ogle et al., 1997), and corticosterone production from the adrenal cortex, which is known to inhibits antibody production (Gross, 1992). Therefore, the observed reduction in serum cortisol level of Cr-supplemented chickens in the present study (Figure 3b) could indicate that a reduction in serum cortisol level is one of the principal mechanisms by which chromium alleviate heat stress induced depression of immune response in broiler chickens.

It is well documented that heterophils are particularly sensitive to adrenocorticotropic hormone (ACTH) (Geraert et al., 1996). Furthermore, it has been shown that the number of heterophils had increased in the blood of corticosterone-fed chickens (Gross and Siegel, 1983). Therefore, the increase in lymphocytes and decrease in heterophils count, with the subsequent reduction of heterophil/lymphocyte ratio of Cr supplemented heat-stressed chickens observed in the present study could be related the Cr-induced reduction in serum cortisol (Figure 3b).

Figure 1. Effects of dietary chromium supplementation on rectal temperature (A) and respiration rate (B) of broiler chickens (mean ± SD; *P<0.05; Control, 0 mg Cr/kg basal diet; Cr-treated, 2 mg Cr/kg basal diet).

Figure 2. Immune response to Newcastle disease virus vaccine (NDV) of control and Cr-supplemented broiler chickens (*P<0.05; Control, 0 mg Cr/kg basal diet; Cr-treated, 2 mg Cr/kg basal diet).

Figure 3. Effects of dietary chromium supplementation on heterophil/lymphocyte ratio (A) and serum cortisol level (B) of broiler chickens (mean ± SD; *P<0.05; Control, 0 mg Cr/kg basal diet; Cr-treated, 2 mg Cr/kg basal diet).

Table 1. Basal feed ingredients in percentage.

	Starter (1-21 d)	Finisher (22-35 d)
Sorghum	59.80	62.00
Groundnut cakes	30.00	20.00
Wheat bran	2.00	9.58
Broiler concentrates^*	5.00	5.00
Salt (NaCl)	0.25	0.25
Methionine	0.09	0.06
Di-calcium phosphate	0.71	0.23
Calcium carbonate	0.28	0.50
Lysine	0.12	0.08
Vegetable oil	1.50	2.10
Premix	0.25	0.20

*Broiler concentrates from Provimi Holland: protein, 38%; metabolizable energy, 1825 kcal/kg; phosphorus, 7.5%; lysine, 10.58%; methionine, 4.25%; fat, 3.25%; crude fibre, 2%.

Table 2. Starter and finisher feed’s nutrient composition (basal feed)

	Starter (1-21 d)	Finisher (22-35 d)
Crude protein, %	23	21
Lysine, %	1.2	1.1
Methionine, %	0.5	0.45
Calcium, %	1.1	1.0
Phosphorus, %	0.5	0.4
Crude fibre, %	5.0	5.0
ME, kcal/kg	3100	3173

ME, metabolizable energy.

Table 3. Effect of dietary supplementation with chromium chloride (2 mg/kg) on the production performance of broiler chickens (mean ± SD).

	Control	Cr-treated
Initial body weight, g	35.52±1.84^a	35.50±0.68^a
Final body weight, g	1097.93±90.62^a	1239.75±30.59^b
Average daily gain, g	30.35±2.59^a	34.41±0.87^b
Average daily feed intake, g	50.85±4.89^a	53.40±3.19^a
Feed conversion ratio	01.68±0.03^a	01.55±0.03^b
Dressing, %	67.75±0.43^a	72.77±2.97^b

Control (0 mg Cr/kg basal diet); Cr-treated (2 mg Cr/kg basal diet).

^a,bMeans within the same row with different letters are significantly different at P<0.05.

Conclusions

Dietary inorganic chromium supplementation as feed additive (2 mg kg^–1 basal diet) has improved the production performance, and potentiated the immune-competence of heat-stressed broiler chickens. The obtained results indicate that dietary chromium supplementation offers a good nutritional management practice to ameliorate heat stress induced depression in production performance and immune response of broiler chickens.

Acknowledgments

The authors would like to extend their sincere appreciation to the Deanship of Scientific Research at King Saud University for its funding of this research through the Research Group Project No. RGP-VPP-171.