Chickens, experimental design and diets

One hundred and forty (140), 21-d-old, Arbor Acres broiler chickens were randomly distributed, keeping their initial body weights equal, in a straight-run, completely randomized experimental design among four treatment groups. Each treatment consisted of seven replicates and five male chickens per replicate. Each replicate was kept in battery brooders (35×25×30 cm length-width-height). During the experiment period (chickens 21-42 d old), the chickens were fed iso-caloric and iso-nitrogenous diets containing four levels of vegetable oils, 2.7% (27 g/kg diet) (basal diet), 4%(40 g/kg diet), 6 % (60 g/kg diet) and 8% (80 g/kg diet) (Table 1). The oil is refining type oil certified for human consumption in Egypt that produced and purchased from Extracted Oils Company and products (Damanhour, Egypt). The estimated composition (NRC, 1994) of major fatty acids contained palmitic acid (8.9 %), stearic acid (4.1 %), oleic acid (24.95 %) linoleic acid (56.05 %), and linolenic (5.3 %).

During the period when they were 25-27, 31-33, and 38-40 d old, the chickens were exposed to heat stress for 4 h a day (1000-1400 h) at 34 ºC, 70-75% relative humidity (RH) and, thereafter, during the unexposed period, the chickens were returned to a thermoneutral condition in which the average temperature and relative humidity were 24.9 °C and 66 % RH, respectively. The high temperature regimen was similar to those reported by Attia et al^3,12,21. The chosen period experienced the highest environmental temperature. The average outdoor temperature was 29.5 °C with 32 % RH. The housing conditions and control for the high environmental temperature was as reported by Attia and Hassan¹². During the period when they were 1-20 d old, the broilers were reared using common husbandry practices, according to the broiler management guide, and fed a commercial diet containing 22 % crude protein (CP), 3,060 kcal, 1 % Ca and 0.5 % available phosphorus (Table 1). The scientific and ethics committee of the Animal and Poultry Production Department, Faculty of Agriculture, Damanhour University approved the experimental protocol.

Broiler husbandry

During the pre-experimental period, when the chickens were 1-20 d old, the birds were kept under similar managerial and hygienic conditions. The chickens were fed corn-soybean meal feeds in mash form during d 1-20, as shown in Table 1. The husbandry of the broilers was done according to the Arbor Acres broiler husbandry guide. Mash feed and water were provided ad libitum. The vaccination regimen was Hitchiner + IB on d 8, avian influenza (AI)(H5N2) on d 9, Gumboro on d 14 and d 24, and Newcastle disease virus (NDV) via Lasota on d 14, 20 and 30. The chickens were provided with a 23:1 light-dark cycle.

Measurements

At 21 and 42 d of age, the broilers were weighed (g) and feed intake was recorded for the same period. Subsequently, their feed, protein and ME were calculated using the feed intake data, and the CP and ME values of the experiment. In addition, the feed conversion ratio (FCR), protein conversion ratio (PCR) and metabolizable energy conversion ratio (MECR) were calculated used the intakes of feed, protein and ME divided by body weight gain.

At 42 d of age, five chickens from each treatment, representing all treatment replicates, were slaughtered according to the Islamic method to determine carcass criteria and inner organs, including lymphoid organs, which were expressed as a percentage of the pre-slaughter weight.

A meat sample (n= 5 per treatment), consisting of 50 % of the deboned thigh meat and 50 % of the deboned breast meat of the slaughtered chickens, was collected on d 42. About 200 g of each sample was wrapped and frozen at -18 °C until used for chemical analyses. A part of each of the fresh meat samples was used to determine the physical characteristics of the meat (n= 5 samples per treatment). The method of Volvoinskaia²² was used to determine the water-holding capacity (WHC) and tenderness of the meat.

Colour intensity as the optical densities of the meat and drip were measured using the colorimetric method, and the pHs of the meat and drip were as reported by Husani et al²³ and Aitken et al²⁴, respectively. The chemical analyses of excreta samples (n= 5/treatment), and samples of breast and thigh meat (a mixture of 50 % breast and 50 % thigh meat) (n= 5/treatment), such as moisture, protein, ether extract and ash, were determined according to the Association Official Analytical Chemists (AOAC)²⁵ methods numbered 925.04, 990.3, 2003.06 and 942.05, respectively.

Five blood samples per treatment were collected on d 42, in unheparinised and heparinized tubes, to determine some of the hematological and biochemical constituents. Blood samples were centrifuged at 3,000 rpm for 20 min, and the plasma and serum were stored at -20 °C for further analyses. The blood’s hematological characteristics, such as hemoglobin (Hgb) and PCV, were determined based on Eilers method²⁶; red blood cells (RBCs) were determined as suggested by Hepler²⁷; and the blood mean cell volume, mean cell haemoglobin and mean cell haemoglobin concentration (MCHC) were calculated. The white blood cells (WBCs) and WBCs’ fractions were measured as described by Lucas and Jamroz²⁸; the phagocyte index (PI) and activity (PA) were measured as suggested by Leijh et al²⁹; and plasma glucose were determined according to the method of Trinder ³⁰, serum total protein were determined as cited by Weichselbaum³¹, serum albumin were measured according to the method of Doumas et al³², and serum globulin were determined according to Coles³³. In addition, the albumin-to-globulin ratio was calculated.

The serum aspartate amino transferase (AST) and alanine amino transferase (ALT) were gauged according to Reitman and Frankel’s³⁴ method. Renal function, creatinine and urea were assessed in the serum based on the suggestions of Bartles et al³⁵ and Sampson et al³⁶, respectively, and the urea-to-creatinine ratio was calculated. Alkaline phosphatase (ALP) enzymes were measured according to the method of Kind and King³⁷. The total plasma triglycerides, cholesterol, high-density lipoprotein, and low-density lipoprotein a were assessed according to the methods of Randrup et al³⁸, Watson³⁹, Friedwald et al⁴⁰ and Wieland and Seidel⁴¹, respectively. Whereas, the very-low-density lipoprotein was determined as plasma triglycerides/5⁴².

The methods of Koracevic et al⁴³ and Richard et al⁴⁴ were used to determine the total antioxidants capacity (TAC) and malondialdehyde (MDA), respectively. The serum antibody body titres for NDV and AI were measured as suggested by Takats⁴⁵, and Kai et al⁴⁶ respectively, and the infectious bursal disease (IBD) was determined according to Cosgrove’s method⁴⁷.

Statistical evaluation

An analysis of variance was done using a one-way analysis of variance, as described by SAS® ⁴⁸, using the following model:

where

Y= the dependent variable,

μ =the overall mean;

Fi =the effect of dietary oil treatments and

eij=the random error.

The replicate was the experimental unit. All percentages were transformed to log10 to normalize the data distribution before analysis. The mean difference at P≤0.05 was tested using the Student-Newman-Keuls test. The survival rate was analyzed using the chi-square test.

Growth performance

Results in Table 2 show a cubic-type pattern in bodyweight gain, FCR, PCR, MECR and EPI. In addition were significantly improved when feed was supplemented with 8% of oils during the high temperature period compared to the groups with 6% oil supplements. The feed, ME, and CP intakes were not significantly affected by the different oil concentrations. The survival rate was lower in 6% oils group than the other groups, but not significantly different (P=0.202).

Carcass characteristics and inner body organs

Table 3 shows the effects of dietary oil supplementation on the dressing percentage, abdominal fat and inner body organs of broiler chickens at 42 d of age. There was a significant effect on most of the traits studied except for the gizzard and pancreas percentage. Broilers fed a 6% DVO diet show a significantly lower dressing percentage than the control and those on the 8% DVO diets, but these groups did not differ from those fed 4% DVO. The percentage abdominal fat was increased and proventriculus was reduced in a dose dependent manner.

The intestine percentage was significantly increased feeding a 6% DVO diet as compared to the control and 8% DVO diets. In addition, the 4% DVO diets exhibited a significantly larger intestinal percentage than the control group (2.7% DVO). The liver percentage was significantly larger for the group fed 6% DVO than the other groups, but the heart percentage was significantly higher for groups fed 4% DVO than the other DVO groups.

Meat quality

There was no significant effect on most of the physical and chemical parameters of meat (Table 4), except for WHC, DM, protein and lipids in meat. The meat’s WHC was significantly decreased progressively as a result of feeding 6 and 8% DVO diets compared to the control (2.7% DVO) and 4% DVO diets. On the other hand, meat’s DM and lipid showed a stepwise significant increase with increasing DVO in broiler diets. Meat’s protein exhibited a significant increase with feeding 8% DVO diet compared to the control group (2.7%) while the result for groups fed 4% DVO was intermediate.

Blood biochemistry of liver and renal function indices

There was no significant effect on most of the biochemical constituents except for the plasma total protein, excreta nitrogen, excreta lipids, plasma total cholesterol, LDL and VHDL (Table 5). The plasma total protein was significantly lower for the groups fed 6% DVO than for the control and 8% DVO groups, while the result for groups fed 4% DVO was intermediate. In addition, the percentage of excreta nitrogen was significantly higher for the control group compared to those in the groups fed 6 and 8% DVO. The groups fed 4% DVO had significantly decreased excreta lipids compared to the other groups. The plasma cholesterol was similarly and significantly increased due to increasing the DVO levels in broiler diets, but the plasma LDL decreased. In addition, the VLDL significantly decreased in groups fed 8% DVO compared with the other groups.

The plasma AST was significantly decreased as a result of feeding 4 and 8% DVO compared to 6% DVO, but the other indices of liver function were not affected by the different DVO levels. In addition, the plasma urea was significantly lower for groups fed the control (2.7% DVO) and 8% DVO than that of group fed 6% DVO. There was no significant effect on plasma creatinine and the urea/creatinine ratio due to different levels of DVO.

Blood hematology

There was no significant effect on most of the hematological constituents, except for RBCs, PCV, MCV and MCH (Table 6). It was found that the value of RBCs was similarly and significantly decreased as a result of feeding 4 and 8% DVO compared to the control group, but the PCV was decreased as a result of feeding 4% DVO compared to the other DVO levels. In addition, the MCV and MCH were significantly increased for groups fed 8% DVO compared to the other groups and the control group, respectively.

In addition, there was no significant effect on most of the WBC parameters, except for lymphocytes and the stress index (Heterophile/ lymphocytes ratio; H/L ratio). Lymphocytes were significantly decreased for the 4, 6 and 8% levels of DVO groups compared to the control group. However, feeding 6% DVO significantly increased the H/L compared to the control group.

Immune status

The plasma TAC was significantly increased due to feeding diets containing 6 and 8% DVO compared to the control groups, but the MDA decreased compared to the control and 4% DVO groups (Table 7). Phagocyte activity was significantly increased from feeding diets containing 6 and 8% DVO compared to the control groups. However, PI was significantly higher of 4% DVO groups than the other groups.

The bursa weight and percentage were significantly increased for groups fed 8% DVO compared with the control and 4% DVO groups. The thymus weight and percentages for groups fed the control diet (2.7% DVO) and 6% DVO were significantly higher than the 4% and 8% DVO groups. There were no significant effects from different DVO concentrations on the spleen weight and percentages and antibody titre for NDV, AI and IBD.