ORIGINAL RESEARCH
published: 28 April 2020
doi: 10.3389/fvets.2020.00181
The Effects of Different Oil Sources
on Performance, Digestive Enzymes,
Carcass Traits, Biochemical,
Immunological, Antioxidant, and
Morphometric Responses of Broiler
Chicks
Youssef A. Attia 1*, Mohammed A. Al-Harthi 1 and Hayam M. Abo El-Maaty 2*
1
Arid Land Agriculture Department, Faculty of Meteorology, Environment, and Arid Land Agriculture, King Abdulaziz
University, Jeddah, Saudi Arabia, 2 Poultry Production Department, Faculty of Agriculture, Mansoura University, Mansoura,
Egypt
Edited by:
Kyung-Woo Lee,
Konkuk University, South Korea
Reviewed by:
Mahmoud M. Alagawany,
Zagazig University, Egypt
John Carragher,
University of Adelaide, Australia
*Correspondence:
Youssef A. Attia
yaattia@kau.edu.sa
Hayam M. Abo El-Maaty
hayam151@yahoo.com
Specialty section:
This article was submitted to
Animal Nutrition and Metabolism,
a section of the journal
Frontiers in Veterinary Science
Received: 24 January 2020
Accepted: 20 March 2020
Published: 28 April 2020
Citation:
Attia YA, Al-Harthi MA and
Abo El-Maaty HM (2020) The Effects
of Different Oil Sources on
Performance, Digestive Enzymes,
Carcass Traits, Biochemical,
Immunological, Antioxidant, and
Morphometric Responses of Broiler
Chicks. Front. Vet. Sci. 7:181.
doi: 10.3389/fvets.2020.00181
This research evaluate the influence of different oil sources, namely fish oil (FO),
coconut oil (CocO), canola oil (CanO), or a mixture of the three oils (MTO)—included
at 1.5% in broiler diets—compared to a no oil-supplemented diet. Hence, 250 unsexed,
1-day-old Cobb chicks were weighed and randomly allocated into five dietary treatment
groups of 50 chicks each and five replicates per group. Oil-supplemented diets
significantly increased the growth, improved the feed conversion ratio (FCR), and
decreased the abdominal fat percentage compared to the control diet. Amylase was
significantly elevated due to feeding the FO- or CocO-supplemented-diet compared to
the control diet, whereas lipase increased due to offering CocO- and CanO-enriched diet;
chymotrypsin increased due to different oil sources. High-density lipoprotein cholesterol
(HDL-C) increased markedly due to offering an oil-supplemented diet, but low-density
lipoprotein cholesterol (LDL-C), the LDL-C:HDL-C ratio, and malondialdehyde (MDA)
decreased. Blood plasma immunoglobulin (Ig) G and IgM significantly increased due to
feeding CocO, CanO, or MTO compared to the control group, whereas FO increased
IgG only. FO- and CanO-containing diets resulted in the highest increase in α2-globulin
and γ-globulin. The antibody titer to avian influenza (HIAI) and Newcastle disease (HIND)
were significantly elevated due to CocO supplementation compared to the control group.
The bursa follicle length and width and thymus cortex depth were increased considerably
due to the FO-supplemented diet compared to the control, but the follicle length:width
ratio decreased. The villus height:depth ratio was significantly elevated due to both the
CanO and MTO diets. The antioxidant status improved considerably due to the addition
of CocO and CanO. Both CanO and MTO similarly increased plasma T3, T4, and the
T3:T4 ratio. In conclusion, oil supplementations at 1.5% enhanced growth performance
and immune status, improved the blood lipid profile and antioxidants status, and the
effect of the oil sources depends on the criteria of response.
Keywords: fish oil, coconut oil, canola oil, broiler performance, immunological response
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INTRODUCTION
Coconut oil (CO) is a rich source of saturated fatty acids (SFA);
they comprise ∼90% of the total fatty acid content. Mediumchain fatty acids (MCFA, C6-C12) represent ∼60% of the entire
fatty acid content (27). These fatty acids are absorbed directly into
the portal circulation without re-esterification in the intestinal
cells (28). MCFA are burned exclusively and rapidly for energy
production (29). In contrast, the LCFA (28) are deposited in the
adipose tissue (30). MCFA reportedly decrease fat deposition in
the meat (31–33) and enhance blood lipid profiles in humans (34)
and rats (31). However, experiments with broilers indicated that
MCFA reduces the growth rate (35). Intriguingly, CocO enhances
fat digestion and growth performance of broilers during a
coccidiosis infection (36).
Currently, there is considerable interest in producing
functional food and/or value-added products such as eggs,
meat and milk (8). The sources of ω3 fatty acids are among
the most exciting potential applications in animal products
due to their health benefits (37, 38). Meat quality and
fatty acids are strongly affected by dietary fat/oil sources (8,
10, 39–41) and might significantly reduce the atherogenic,
thrombogenic, and cholesterol effects of animal products (37,
38). Although some information is available on the effect of
the use of individual oils on broiler performance, blood lipid
profile, and immunity (8, 42), the results on the impact of
the combination of different oil sources on the production,
pancreatic enzymes, meat quality, metabolic profiles, and
immune status in broilers are limited. Therefore, this study
evaluated the effect of feeding a diet that contained 1.5% of
FO, CocO, CanO, or a mixture of the three oils (MTO) on
the performance, digestive enzymes, carcass traits, biochemical,
immunological, antioxidant, and morphometry responses of
broilers chickens.
Poultry is a very important industry. Indeed, it is estimated that
in 2020, chicken will be the most consumed animal protein in
the world. Fat and oil are generally used in poultry diets to
increase the energy concentration. Fat-enhanced feeds increase
the efficiency of the feed energy and productivity in poultry (1, 2).
Moreover, oil improves the absorption of fat-soluble vitamins,
the palatability of diets, decreases the dustiness of feeds, and
reduces the passage rate of feed in the gut, which provides more
time for the sufficient absorption of nutrients (3–5). Besides, the
fatty acid profile of muscle tissue mirrors the dietary lipid profile
and can alter the blood levels of lipoproteins and triglycerides
(6–8). As a rule, the utilization of unsaturated fatty acids (UFAs)
in poultry diets improves the product quality (for example, ω3
and ω6). This improvement is concordant with the consumers’
interest (9, 10) and immune response (3, 4, 11–13).
Fish oil (FO) has high percentages of long-chain
polyunsaturated fatty acids (PUFA), particularly ω3 fatty
acids that increase oxidative damage and negatively affect the
flavor of animal products. FO is low in ω6 fatty acids and linoleic
acid (14, 15). Two types of ω3 fatty acids are found in fish meal:
eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA)
(15). Alpha-linolenic acid (ALA) is found in plant seeds and
can be transformed to EPA and DHA in the body (16). Corn
oil, animal fat, animal and vegetable fat blends, and FO do not
affect the feed consumption but improve the growth rate and
feed conversion rate (FCR) in broilers exposed to heat stress
compared to un-supplemented diets with heat stress (17).
Dietary ω3 PUFA sources, such as FO or linseed oil, have a
positive significant impact on humoral immunity, i.e., antibody
titers against the Newcastle disease virus (NDV), compared to
the control diet (18). FO has no negative effects on the immune
function of broilers (19). Furthermore, FO significantly increases
antibody titers and the relative weight percentages of the bursa
and spleen compared to a control diet (20). Birds fed a diet
with an ω6: ω3 ratio that exceeds the recommended levels show
damages to the intestinal epithelial cells (21). Further, a low ω6:
ω3 ratio increases malondialdehyde (MDA) in tissues, including
the muscle, and thus impairs meat quality.
Less than 2% of erucic acid (docosenoic acid, C22:1, ω-9) of
rapeseed variety is called canola oil (CanO) (22). High erucic acid
levels in the diet harm the growth, feed intake, and digestibility
of lipids (23). Furthermore, chicks fed diets that contain erucic
acid deposit less fat and use lipid energy less efficiently (23).
Feeding two varieties of CanO to female broilers increases the
growth rate compared to feeding tallow and acidulated soybean
oil soap stock. These data confirm the advantages of using CanO
as energy sources for birds (24). The better growth rates are a
result of the higher percentage of long-chain fatty acids (LCFA)
and high triglyceride content (24). Broilers fed a diet with poultry
fat, CanO, sunflower oil, corn oil, soybean oil, or lard exhibit
similar growth performance as well as cuts and carcass yields at
day 49 of age (25). Broilers fed corn oil and lard show higher
red-colored meat compared to those fed on CanO, soybean oil,
and sunflower oil, but the difference from the poultry fat is not
significant (26).
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MATERIALS AND METHODS
The experimental protocol was approved by the scientific
committee of the Poultry Production Department, Faculty
of Agriculture, Mansoura University. The care and
handling of the animals were performed to maintain their
rights, ensure their welfare, and cause minimal stress,
according to International Guidelines for research involving
animals (Directive2010/63/EU).
Experimental Design, Animals, Diets, and
Management
A total of 200 unsexed, 1-day-old Cobb chicks were weighed
and equally divided among four experimental groups (50 chicks
each). The chicks in each group were subdivided into five equal
replicates (10 chicks per replicate), and each replicate was housed
in 1 ×1 m floor pens in open-sides house furnished with wood
shaving for litter. Each pen was equipped with a tube pen feeder
and 5-L waterer. The first, second, third, and fourth groups were
fed a diet with 1.5% FO, CocO, CanO, or MTO (FO + CocO
+ CanO of 0.5% each), respectively. The oils were added to the
basal diet to isocaloric and isonitrogenous feeds to minimize the
interaction between dietary and supplemented fats.
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Oils and Broilers’ Performance and Immunity
TABLE 2 | The calculated composition and chemical analyses of the experimental
diets fed to broiler chicks from 21 to 42 weeks of age.
TABLE 1 | The calculated composition and chemical analyses of the experimental
diets fed to broiler chicks from 0 to 21 weeks of age.
Ingredients, %
FO1 -diet CocO2 -diet CanO3 -diet MTO4 -diet
Ingredients, %
FO1 -diet CocO2 -diet CanO3 -diet MTO4 -diet
Ground yellow corn
60.1
60.1
60.1
60.1
Ground yellow corn
66.35
66.35
66.35
66.35
Soybean meal 44%
18.0
18.0
18.0
18.0
Soybean meal 44%
16.0
16.0
16.0
16.0
Corn gluten meal 60%
15.5
15.5
15.5
15.5
Corn gluten meal 60%
11.5
11.5
11.5
11.5
Fish oil (FO)
1.50
–
–
0.50
Fish oil (FO)
1.5
–
–
0.50
–
1.50
–
0.50
Coconut oil (CocO)
–
1.5
–
0.50
Coconut oil (CocO)
Canola oil (CanO)
–
–
1.50
0.50
Canola oil (CanO)
–
–
1.5
0.50
Ground limestone
2.0
2.0
2.0
2.0
Ground limestone
2.0
2.0
2.0
2.0
Dicalcium phosphate
1.8
1.8
1.8
1.8
Dicalcium phosphate
1.8
1.8
1.8
1.8
Vitamin and mineral Premix5
0.30
0.30
0.30
0.30
Vitamin and mineral Premix5
0.30
0.30
0.30
0.30
Sodium chloride
0.30
0.30
0.30
0.30
Sodium chloride
0.30
0.30
0.30
0.30
–
–
–
–
–
–
–
–
L-Lysine-HCl
0.50
0.50
0.50
0.50
L-Lysine-HCl
0.25
0.25
0.25
0.25
Total
100
100
100
100
Total
100
100
100
100
DL-Methionine
DL-Methionine
Calculated analysis (as fed basis)
Calculated analysis (as fed basis)
Metabolizable energy MJ/kg
13.2
13.1
13.1
13.1
Metabolizable energy MJ/kg
13.2
13.2
31.2
13.2
Crude protein %
23. 1
23.1
23.1
23. 1
Crude protein %
20.0
20.0
20.0
20.0
Ether extract %
2.82
2.82
2.82
2.82
Ether extract %
2.94
2.94
2.94
2.94
Crude fiber %
2.78
2.78
2.78
2.78
Crude fiber %
2.73
2.73
2.73
2.73
Calcium %
1.20
1.20
1.20
1.20
Calcium %
1.19
1.19
1.19
1.19
Nonphytate P %
0.451
0.453
0.452
0.451
Nonphytate P %
0.45
0.45
0.45
0.45
Lysine %
1.19
1.19
1.19
1.19
Lysine %
0.922
0.921
0.923
0.922
Methionine %
0.453
0.451
0.452
0.453
Methionine %
0.392
0.391
0.392
0.391
Methionine + Cystine %
0.852
0.853
0.851
0.851
Methionine + Cystine %
0.743
0.742
0.741
0.741
90.7
Determined analysis (as fed basis)
Determined analysis (as fed basis)
Dry matter %
89.0
89.1
88.9
88.9
Dry matter %
91.0
90.9
90.9
Organic matter %
79.1
80.1
79.8
79.9
Organic matter %
81.7
81.7
81.7
81.5
Crude protein %
23.0
22.9
23.0
23.0
Crude protein %
22.0
22.0
22.0
22. 1
Ether extract %
3.17
3.16
3.17
3.17
Ether extract %
3.23
3.23
3.20
3.24
Crude fiber %
3.12
3.11
3.13
3.13
Crude fiber %
3.00
3.03
2.99
3.01
Ash %
9.89
9.05
9.13
9.07
Ash %
9.26
9.17
9.21
9.18
Nitrogen-free extract %
46.9
47.9
47.6
47.6
Nitrogen-free extract %
53.5
52.5
53.5
53.1
1 FO, Fish oil; 2 CocO, Coconut oil; 3 CanO, Canola oil; 4 MTO, Mixture of three oilds (FO +
CocO + CanO); 5 Each 3 kg of preMTO contained: vit. A 12,000,000 IU, vit. D3 3,500,000
IU, vit. E 20 g, vit. K3 3 g, vit. B1 3 g, vit. B2 8 g, vit. B6 3 g, vit. B12 15 mg, Ca pantothenate
12 g, Niacin 40 g, Folic acid 1.5 g, Biotin 50 mg, Choline chloride 600 g, Mn 80 g, Zn 75 g,
Fe 40 g, Cu 10 g, I 2 g, Se 0.3 g, Co 0.25 g, and CaCo3 as a carrier.
1 FO, Fish oil; 2 CocO, Coconut oil; 3 CanO, Canola oil; 4 MTO, Mixture of three oilds (FO +
CocO + CanO). 5 Each 3 kg of preMTO contained: vit. A 12,000,000 IU, vit. D3 3,500,000
IU, vit. E 20 g, vit. K3 3 g, vit. B1 3 g, vit. B2 8 g, vit. B6 3 g, vit. B12 15 mg, Ca pantothenate
12 g, Niacin 40 g, Folic acid 1.5 g, Biotin 50 mg, Choline chloride 600 g, Mn 80 g, Zn 75 g,
Fe 40 g, Cu 10 g, I 2 g, Se 0.3 g, Co 0.25 g, and CaCo3 as a carrier.
The experimental starter and growth basal diets were
composed mainly of maize, soybean meal, and corn gluten
meal and formulated using feedstuff composition recommended
by the National Research Council (43). The experimental diets
met the Cobb breeding guide. The ingredient composition and
calculated analysis of the basal diets are shown in Tables 1, 2. The
chicks were offered a starter diet—containing 23% crude protein
and ∼13.2 MJ/kg metabolizable energy (ME)—up to 21 days of
age. Subsequently, they were switched to the growth diets, which
contained ∼20% CP and 13.2 MJ/kg ME, from 21 to 42 days
of age.
All chicks were kept under similar managerial, hygienic, and
environmental. The vaccination and medical care program were
carried out under the supervision of a veterinarian. In brief, the
birds were vaccinated against IB+ NDV (Hitchner B1 strain) at
day 8, Gumboro at day 15, and the NDV LaSota strain at day 21.
Feed and water were offered ad libitum. Broilers were provided
with a 23 h light:1 h dark photoperiod. The indoor temperature
and relative humidity (RH) during the experimental period were
34◦ C and 43%, 32◦ C and 45%, and 30◦ C and 51% during weeks
1, 2, and 3 of age, respectively. The ambient temperature and RH
during weeks 4, 5, and 6 were 29.7◦ C and 55%, 30.2◦ C and 56%,
and 30.8◦ C and 55%, respectively.
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Criteria of Response
The body weight and feed intake of each replicate were recorded
at days 1, 21, and 42 of age. Records of live body weights (LBW)
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for at least 24 h. The samples were prepared and measured as
previously reported (54).
The proximate analyses of the experimental diets were
performed according to the following official methods of analysis
(55): dry matter, method number 934.01; crude protein, method
number 954.01; ether extract, method number 920.39; crude
fiber, method number 954.18; and ash, method number 942.05.
and feed intake (FI) were used to calculate the FCR, and (44) for
the European production index.
At 42 days of age, five birds from each treatment were
randomly selected to represent all treatment replicates; each
selected bird was around the average body weight for the
group. These specimens were weighed, fasted overnight, and
then slaughtered according to a previously reported Islamic
method (45). After complete bleeding, the feathers were plucked,
the carcasses eviscerated, and hot carcasses were weighed.
The weight of the abdominal fat, liver, gizzard, heart, giblets
(liver + gizzard + heart), and total edible parts (hot carcass
+ giblets) were recorded and expressed as a ratio to live
body weight.
Five blood samples per treatment—representative of all
replicates—were collected from the brachial vein with a
vacutainer tube using heparinized and non-heparinized tubes at
day 29 to determine the serum antibody titer and at day 42 to
determine the biochemical constituents of the blood. Serum and
plasma were separated by centrifugation at 1,500 g for 15 min.
The blood samples used for analyses were collected before the
start of all vaccinations (day 0) and after the end of the last
vaccination (day 29). Total antibody production specific for
the NDV vaccine was determined in serum using commercial
enzyme-linked immunosorbent assay (ELISA) kits (46). The
antibody response was measured by the hemagglutination
inhibition (HI) test (47). The assay is designed to measure IBD
antibody bound to influenza antigen-coated plates (48). The
Takatsy method (49) was used to determine the HI against NDV,
avian influenza (AI), and infectious bronchitis disease (IBB).
The biochemical markers were assayed using commercial
diagnostic kits (Spectrum Diagnostics, Obour City, Egypt) unless
otherwise stated. The total protein, albumin, triglyceride, total
cholesterol, and high-density lipoprotein cholesterol (HDLC) levels in the blood plasma were measured at day 42 of
age. The level of low-density lipoprotein cholesterol (LDLC) in blood plasma was estimated (50) as follows: LDLC = Total Cholesterol–(HDL-C + VLDL); where very-lowdensity lipoprotein (VLDL) was estimated as the concentration
of plasma triglycerides divided by five. In addition, the
activities of plasma superoxide dismutase (SOD), catalase (CAT),
malondialdehyde (MDA), alanine aminotransferase (ALT), and
aspartate aminotransferase (AST) as well as the total antioxidant
capacity (TAC) were determined. Alkaline phosphatase (AlkP),
immunoglobulin (Ig)G, IgM, and IgA, and thyroid hormones (T3
and T4) were also determined. In addition, α-, β- and γ-globulin
were measured according to a previously published method (51).
The contents of the duodenum, jejunum, and ileum were
qualitatively collected and then kept in equal quantities of saline
buffer. The mixture of each content was then centrifuged at
1,792 g for 15 min, and the supernatant was used to determine
the levels of some digestive enzymes. The amylase activity was
determined using the method described by Pinchasov et al. (51),
lipase activity according to Sklan and Halevy (52), and trypsin
and chymotrypsin activities according to Sklan et al. (53).
Representative samples of ilea, liver, thymus, and bursa (n =
5 per treatment) at 42 days of age were fastidiously dissected and
placed in a sufficient volume of 10% buffered formal saline (BFS)
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Statistical Analysis
The data were analyzed with the Statistical Analysis System (SAS)
software (56) using one-way analysis of variance (ANOVA) and
the generalized linear model (GLM) procedure. The StudentNewman-Keuls test (56) was used to predict differences among
the criteria; the effects were considered significant if P ≤ 0.05. All
percentages were log base 10 transformed before ANOVA and
then converted back to risk ratios for result presentation.
RESULTS AND DISCUSSION
Growth Performance
Table 3 presents the effect of different oil sources on the
performance of broilers during 1–42 days of age. Birds in this
experiment show signs of thermal stress as evidenced by panting,
lying on the floor and straightening the wing even the cooler
group were absent, but heat stress behavior was evident Oil
sources promoted positive effects on broiler chick growth during
days 1–21 and 1–42 of age. The results also revealed that CocO
source significantly increased chick growth during the 1–21-day
period (9.9%) compared the FO-diet. However, Wang et al. (57)
reported that a CocO-supplemented diet has no effect on weight
gain. In addition, MCFA decreases the growth rate (35), and
CocO enhances the digestion of fats and the performance index
during coccidiosis infection (36).
For the whole period, CanO significantly elevated growth
during days 1–42 compared to FO and CocO diets. The improved
growth performance of broilers on CanO could be due to
higher UFA and mono-unsaturated fatty acids (MUFA), and
the higher energy capacity of PUFAs (23). The growth rate of
female broilers fed two varieties of CanO is higher compared to
broilers fed tallow and acidulated soybean oil soap stock. The
increased growth is due to high LCFA and a high percentage of
triglycerides (24).
In previous studies, fat-enhanced feeds elevate the efficiency of
the consumed energy and productivity in poultry under normal
and hot climate condition, but the impact depends on fat/oil
source (2, 44, 45). Moreover, the oil improves the absorption
of fat-soluble vitamins, the palatability of diets, decreases the
dustiness of feeds and reduces the passage rate of feed in the gut,
which allows more time for the sufficient absorption of nutrients
particularly under high temperature due to impairing digestion
of feeds (1–5, 44). For examples, Zollitsch et al. (39) and Khatun
et al. (8) reported that broilers fed 6% soybean oil (mostly 84%
an unsaturated oil) or a combination of soybean oil and palm
oil (mostly 50% a saturated oil) exhibit significantly increased
growth during days 1–21 of age compared to a control diet
with 6% palm oil. Nobakht concluded that the growth rate of
broiler chickens decreases with dietary FO inclusion (14). The
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TABLE 3 | Effect of different dietary oil sources on growth performance of broiler chicks.
Traits
Dietary treatments
FO1 -diet
CocO2 -diet
CanO3 -diet
MTO4 -diet
44.5
44.5
44.6
44.5
RMSE
P-values
0.329
0.997
Body weight
Initial 1 day, g
Body weight gain
1–21days, g
537b
590a
583ab
575ab
28.6
0.022
1–42 days, g
2058b
2038b
2107a
2068ab
99.7
0.001
918a
853b
795b
845b
42.7
0.004
3156ab
115.9
0.015
Feed intake
21days, g
42 days, g
3290
a
3097
b
3020
b
Feed conversion rate
21days, kg/kg
42 days, kg/kg
1.71a
1.45bc
1.37c
1.47b
0.061
0.001
a
ab
b
1.53ab
0.075
0.022
1.60
1.52
1.43
European production index
1–42 days
308
319
350
324
2.82
0.148
0.129
Digestive enzymes activity, U/ml of intestinal content
Amylase
3.33
3.32
3.01
2.96
0.299
Lipase
11.7
12.7
12.8
12.5
1.65
0.712
Trypsin
28.2
28.9
28.4
24.8
2.65
0.089
Chymotrypsin
19.8
21.3
20.7
20.8
1.92
0.654
a,b,c Means
for each trait with different superscripts differ significantly at p < 0.05; 1 FO, Fish oil, 2 CocO, coconut oil, 3 CanO, canola oil, 4 MTO, mixture of three oils (FO + CocO + CanO).
that broilers fed a CocO-supplemented diet showed no difference
in FI compared to a FO-supplemented or control diet (14).
The broilers fed the CanO-supplemented diet from days 1–
21 and 1–42 exhibited the best change in FCR compared to the
other oil diet: 6.80–19.9% and 7.0–10.6%, respectively (Table 3).
The worst FCR was from the FO-supplemented feeds from days
1–21 and 1–42 of age. Broilers fed the CocO- or MTO-enriched
diet showed similar and better FCR than the FO during days
1–21 (15.2%). The present results indicate that CanO, which is
rich in linolenic and linoleic acids, enhances broiler growth and
FCR due to its high energy-yielding capacity compared to diets
rich in SFA. These enhancements were associated with increased
growth. The results also demonstrate that CanO used herein
had low erucic acid content and thus had no harmful effect
on broilers (23, 25). Likewise, oil supplementations improved
FCR of broiler under normal and high ambient temperature
(2, 44, 45). In literature, differences in FCR due to various oil
sources are exist for example, Wignjosoesastro et al. (61) revealed
that 10% CocO increases the rate of production and efficiency.
In addition, Zollitsch et al. (39) and Ayed et al. (40) observed
that broilers fed soybean oil present a significantly improved
FCR compared and palm oil diets. Besides, Khatun et al. (8)
revealed that a 6% PUFA-supplemented diet and a combination
of soybean and palm oils significantly enhance the FCR compared
to the 6% palm oil diet. However, broilers fed sunflower oil have
a better FCR compared to those fed beef tallow (61). On the other
hand, Wang et al. (57) reported that broilers fed a CocO-enriched
diet exhibit no difference in FCR compared to control. Nobakht
(14) concluded that the dietary FO level does not affect the FCR
of broilers.
contradiction among the above mentioned studies indicate that
the impact of the type of fat on growth performance depends on
the profile and levels of the utilized fatty acid(s) as well as the age
of the chickens and type of stress (58). Chickens fed an erucicacid-enriched diet exhibit a significantly lower growth rate, FI,
and lipid and fatty acid digestibility (23) and store less fat and
utilize energy less efficiency (23).
There was a significant effect on FI during days 1–21 and 1–
42 (Table 3). FO significantly increased FI compared to other
oils groups during days 1–21 of age. Furthermore, the FO diet
increased FI by 8.9% compared to the CanO diet from days 1–
42. The lowest FI was for CanO group from days 1–21 and 1–42
days of age. MTO supplementation presented an intermediate
effect on FI. The FO-mediated FI increase compared to CocO,
and CanO is in general agreement with Dawood and Mohammed
(59). Besides, 3% palm oil significantly increases feed intake
compared to soybean oil from days 17–38 of age but does not
affect FI from days 1–16 of age (40). However, 6% palm oil
or soybean oil or different combinations of the two does not
affect feed intake of broilers during days 1–21 days and 22–42
age (8). According to the present findings, the addition of SFA
increased the FI compared to UFA; this change depended on the
level and source of fats and age of chickens. In the literature, the
PUFA linolenic acid (18:3) and linoleic acid (18:2) decrease feed
intake compared to SFA, i.e., palmitic and stearic acids (60). The
decrease in FI observed herein of broilers fed CocO, and CanO
might be due to PUFAs and their high energy-yielding capacity.
In addition, the low digestibility of SFA compared to UFA was
cited Zollitsch et al. (39) and Ayed et al. (40), particularly during
the early weeks of life (3). However, Wang et al. (57) reported
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rather than stored in the body (27). Consumption of a feed
rich in MCFA increases upper body adiposity in overweight
men; hence, MCFA may be considered as a potential tool in the
prevention of weight gain and obesity (27, 32). Consistent with
the reduction in the dressing and total edible parts of broilers, the
FO-supplemented diet negatively affected carcass criteria (4, 64).
The observed increase in the liver percentage of the broilers
fed the CanO-enriched diet was due to additional fat deposition
in this organ rather than as abdominal fat. This phenomenon is
likely due to the higher UFA content in CanO (64.8%), of which
oleic is dominant. The ME of oils and fats depends on the number
of double bonds, the length of the carbon chain, the presence
or absence of ester bonds (free fatty acids or triglycerides), the
dietary composition (3). In addition, the specific configuration
of the UFA and SFA in the glycerol backbone, the free fatty
acid arrangement, the amount and the kind of the nutritional
triglyceride contents, the gut microflora, and the age and the sex
of the chicken (3, 39).
There were no significant differences in European production
Index and mortality under the present experiment condition.
Digestive Enzyme Activity
The activity of the digestive enzymes in the gut (proventriculus,
duodenum, jejunum, and ileum) as influenced by different
dietary oil sources is shown in Table 3. The measurements
estimated at day 42 of age after exposure of broilers to high
ambient temperature during 4–6 weeks of age indicate that
there were no significant differences due to different oil sources
on digestive enzyme activates. The inclusion of FO, and CocO
numerically increased intestinal amylase (10.5–12.5%) compared
to CanO and the combination of all three oils. In addition,
CocO-, CanO-, and MTO enriched diets numerically elevated
intestinal lipase 6.8–9.4% compared to the FO-supplemented
diet. However, there were a trend (P < 0.089) for trypsin
to be numerically higher (16.1%) in the groups that received
the individual fat sources compared to the MTO group.
Chymotrypsin activities numerically also increased (2.5%) due
to CocO compared to FO. A previous study indicated higher
activities of pancreatic trypsin, α-amylase, and intestinal maltase
due to oil supplementation (62). In general, the improved
digestive enzyme activities found herein are consistent with the
increased growth and feed use for growth under hot conditions
(Table 3). Along the same line, CocO improves fat digestion and
performance values during coccidiosis infection (36).
Plasma Lipid Profile
Table 5 presents the influence of different oil sources on the
plasma lipid profile of 42-day-old broilers. Plasma triglycerides,
total cholesterol, LDL-C, HDL-C, and the LDL-C:HDL-C ratio
and vLDL of broilers fed diets with different sources of oils
were similar. The total plasma lipids was significantly higher
in the FO and CanO groups compared to the MOT group,
which showed the greatest decrease (15.5%) in plasma total
lipids. The CocO groups presented intermediate values. The
plasma concentrations of triglycerides/HDL showed a trend
toward (P < 0.093) increased values (10.5–12.6%) of CanO and
MTO groups compared to FO and CocO groups. Overall, the
change in plasma lipid profiles, reflecting the dietary fatty acid
composition and correlated with the increased dietary PUFA
content, as well as elevated intestinal lipase activity. The positive
effect of CocO supplementation found herein on the plasma total
lipids, triglycerides/HDL indicates that oil is burnt for energy
and use for dissipate heat stress (27, 32, 44, 45). Similarly, the
MCFA improve serum lipid profiles in humans and rats (31, 34).
Researchers demonstrated that UFA and beneficial PUFA have
positive and healthy influences on plasma lipids due to its impact
on increasing HDL-C while decreasing LDL-C, the hazardous
lipoprotein segment (61, 65–67).
Carcass Characteristics
The dressing, liver, gizzard, heart, giblets, total edible part, and
abdominal fat after supplementation with the different oil sources
are shown in Table 4. The parameters measured at day 42 of age
indicate that broilers fed the CocO-, CanO-, and MTO-enriched
diets had a similar dressing percentage, and a resulted in 2.8–4.5%
increase in total edible parts compared to the FO-supplemented
feeds. Moreover, MTO supplementation had no additive effects
on carcass parameters and organs traits compared to induvial
oil sources. This indicates that individual supplementation of
oil source was adequate, and oil supplementations during hot
weather condition are beneficial (2, 44, 45). Broilers fed the
CocO, CanO-, and MTO-enriched diets had lower abdominal
fat percentage (32%) compared to the FO diets, but the liver
percentage was significantly higher (22.9%) with the CanOenriched diet compared to the CocO. The increased dressing
and total edible parts of broilers fed a PUFA-enriched diet
(CocO, CanO, or MTO) indicates that these animals had higher
energy availability for muscle growth, while the decrease in
abdominal fat in these groups shows a shift in energy use for
muscle growth rather than deposition in the abdominal cavity
(14, 15). In addition, Baião and Lara (3) and Nobakht et al.
(14); demonstrated that the inclusion of different PUFA sources
and levels in broiler diets significantly influenced carcass traits.
Moreover, including 5% CanO in broiler diets increases the breast
weight compared to the other groups (63). Similar performance
and carcass and cut yields were reported for broilers fed different
sources of fats (2, 25, 44, 45). The improved carcass yield and
the decrease in abdominal fat and liver percentage of broilers
fed the CocO-enriched diet indicates that oil is burnt for energy
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Plasma Thyroid Hormones
Thyroid hormones as affected by different types of oils are
shown in Table 6. The results show that CocO and MTO
supplementation promoted the greatest increases in T3 compared
to the FO and CanO diets. On the other hand, T4 and
T3 :T4 ratio were not affected by dietary fat sources. Thyroid
hormones decreased during hot weather and are involved in the
regulation of anabolic and catabolic pathways of protein, lipid
and carbohydrate metabolism (68, 69). A previous study reported
that a significant interaction between the type and level of fats
might have adverse effects on T3 and T4 levels (68, 69). The
dietary fatty acid profile reportedly affects deiodinase hepatic type
I activity (68, 69) and binding of T3 to nuclear receptors (70). This
finding partially contradicts the results of research that showed
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TABLE 4 | Effect of feeding different oil sources on carcass characteristics at 42 days (%).
Traits
Dietary treatments
RMSE
P-values
FO1 -diet
CocO 2 -diet
CanO3 -diet
MTO4 -diet
Dressing, %
71.3c
73.6b
74.6 a
74.9a
0.626
0.001
Liver, %
2.52ab
2.31b
2.84a
2.54ab
0.266
0.049
Gizzard, %
2.61
2.67
2.59
2.57
0.354
0.969
Heart, %
0.569
0.549
0.516
0.560
0.063
0.579
Giblets, %
5.70
5.53
5.94
5.67
0.320
0.280
Total edible part, %
77.0c
79.2b
80.5a
80.2a
0.725
0.001
Abdominal fat, %
2.42a
1.77b
1.73b
1.71b
0.261
0.002
a,b,c Means
for each trait with different superscripts differ significantly at p < 0.05; 1 FO, Fish oil, 2 CocO, coconut oil, 3 CanO, canola oil, 4 MTO, mixture of three oils (FO + CocO + CanO).
TABLE 5 | Effect of feeding different oil sources on some blood plasma lipid biochemical constituents; antioxidants indices; thyroid hormones and liver leakage indices.
Traits
Dietary treatments
RMSE
P-values
FO1 -diet
CocO2 -diet
CanO3 -diet
MTO4 -diet
Total lipids, mg/dl
560a
514ab
576a
487b
41.9
0.016
Triglycerides, mg/dl
144
144
150
157
10.3
0.206
Cholesterol, mg/dl
169
172
168
173
11.3
0.632
HDL, mg/dl
65.5
65.8
62.8
62.7
6.18
0.782
Triglycerides:HDL ratio
2.22
2.19
2.42
2.50
0.213
0.093
LDL, mg/dl
74.7
77.8
75.2
79.1
11.3
0.912
LDL: HDL ratio
1.17
1.20
1.21
1.26
0.258
0.948
vLDL, mg/dl
28.9
28.8
30.1
31.4
2.05
0.206
Plasma lipid metabolites
Plasma thyroid hormones
T4, ng/Ml
20.6
22.1
21.2
23.6
1.95
0.131
T3, ng/mL
4.51b
5.18a
4.59b
5.23 a
0.345
0.006
T3:T4 ratio
0.219
0.236
0.217
0.223
0.018
0.354
Plasma antioxidant enzymes
CAT, U/ml/h
61.5 b
74.1a
68.1ab
67.2ab
5.08
0.012
SOD, U/ml/h
75.5b
88.7a
84.2a
91.1a
4.89
0.001
TAC, nmol/ml
1.21b
1.36 a
1.40a
1.34a
0.091
0.023
MDA, nmol/ml
22.2a
17.2b
21.2ab
17.6b
2.69
0.021
MDA:TAC ratio
18.5a
12.7b
15.2b
13.1b
2.02
0.002
AST, U/dL
68.2b
57.9c
78.7a
65.7b
4.25
0.001
ALT, U/dL
26.1
a
b
a
24.1a
1.99
0.001
AlkP, U/dL
56.6a
65.4a
64.1a
5.75
0.001
Plasma liver leakage indices
18.8
24.3
43.3b
a,b,c Means for each trait with different superscripts differ significantly at p < 0.05; HDL, High density lipoprotein; LDL, Low density lipoprotein; vLDL, Very low density lipoprotein; CAT,
Catalase; SOD, super oxide dismutase; TAC, Total antioxidant capacity; MDA, Malondialdehyde; T4, Thyroxine; T3, Triiodothyronine; AST, Aspartate amino transferase; ALT, Alanine
amino transferase; 1 FO, Fish oil; 2 CocO, coconut oil; 3 CanO, canola oil; 4 MTO, mixture of three oils (FO + CocO + CanO).
diet (Table 4). CocO is a rich source of SFA and MCFA (6–12
carbon atoms) (3, 27), which can be absorbed directly into the
portal system without re-esterification in intestinal cells (28).
MCFA are exclusively and rapidly burned to produce energy
(29). By contrast, LCFA is commonly found in most diets
and are incorporated into chylomicrons after being absorbed
in the intestine, where they are subjected to re-esterification,
and then reach the bloodstream via the lymphatic system
(28). Most LCFA are stored in adipose tissue (30). In one
a reduction in T3 level of animals fed high-fat diets (71, 72).
However, it is difficult to explain the lack of effect of fat level on
the T3 concentration in rats fed palm and rapeseed oils.
The increase in T3 of broilers supplemented with CocO
diets under hot climate condition may be due to the elevated
energy availability and use for an anabolic processes for muscle
growth. It is evidenced by the rise in the growth rate and FCR
during 1–21 days of and the decrease in liver and fat deposition
in the abdominal cavity of broilers fed a CocO-supplemented
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TABLE 6 | Effect of feeding different oil sources on blood serum immune indices, and morphometric characteristics of Intestinal villi, bursa of Fabricius and thymus cortex.
Triats
Dietary treatments
RMSE
P-values
FO1 -diet
CocO 2 -diet
CanO3 -diet
MTO4 -diet
Total serum protein, g/dl
3.17a
2.14c
3.14a
2.64b
0.226
0.001
Albumin, %
32.2b
46.8a
52.3a
45.9a
7.07
0.003
α1-globulin, %
1.78b
8.48a
7.47a
3.70b
2.43
0.002
α2-globulin, %
5.84
3.87
5.55
6.17
1.91
0.268
β-globulin, %
8.92a
4.88b
9.33a
5.57b
2.25
0.012
γ-globulin, %
42.6ab
24.0c
35.9b
47.9a
6.09
0.001
1.20
1.34
1.40
1.26
0.192
0.405
IgG, mg/dL
460b
541a
542a
572a
23.9
0.001
IgM, mg/dL
191
b
a
a
244a
18.9
0.001
IgA, mg/dL
127
137
133
138
13.9
0.636
0.718
Serum protein metabolites
A:G ratio
Serum immunoglobulin
242
253
Serum antibody titer
HIBB, log2
4.24
3.62
3.23
4.21
1.08
HIBD, log2
3.62
2.89
3.11
3.34
1.14
0.451
HIAI, log2
3.66ab
4.23a
3.12b
3.17b
0.510
0.005
HIND, log2
5.61b
7.02a
6.23ab
6.07ab
0.688
0.037
Ville height, µm
623b
522c
661ab
703a
40.2
0.001
Crypt depth, µm
106
112
109
99.1
10.9
0.320
5.94a
4.76b
6.19a
7.18a
0.763
0.001
471b
474b
506a
512a
20.5
0.009
a
ab
b
304ab
36.5
0.033
Intestinal villi parameters
Ville height:depth ratio
Bursa of Fabricius follicle and thymus characteristics
Follicle length, µm
Follicle width, µm
355
310
278
Length: width ratio
1.34b
1.56ab
1.82a
1.70a
0.185
0.006
Thymus cortex depth, µm
112b
123b
155a
129b
13.93
0.001
a,b,c Means for each trait with different superscripts differ significantly at p < 0.05; α1-globulin, Alpha 1-globulin; α2-globulin, Aalpha 2 globulin; β-globulin, Beta-globulin; γ-globulin,
Gamma-globulin; IgG, Immunoglobulin G; IgM, Immunoglobulin M; IgA, Immunoglobulin A; HIBB, Hemagglutination-inhibition test for infectious bronchitis virus; HIBD, Hemagglutinationinhibition test for bursa disease; HIAI, Hemagglutination-inhibition test for avian Influenza; HIND, Hemagglutination-inhibition test for Newcastle disease virus; 1 FO, Fish oil; 2 CocO,
coconut oil; 3 CanO, canola oil; 4 MTO, mixture of three oils (FO + CocO + CanO).
(45, 65). Notably, Bhatnagar et al. (27) reported that CocO dietary
supplementation increases total tocopherols. Tocopherols are
essential antioxidants that protect the cell membrane from free
radicals (73). Furthermore, CocO is very stable against oxidation
and, consequently, not prone to peroxide formation. Therefore,
the incorporation of CocO enhances the constancy of the feeds
(27). Besides, CocO has unique antibacterial, antiprotozoal, and
antiviral effects, all of which control microbial rancidity (74, 75).
study, the total T4 levels are higher in Wistar rats fed 5 or
10% palm-oil-enriched compared with a rapeseed-oil-enriched
diet (68).
Plasma Antioxidant Status
The supplementation with CocO, CanO, or MTO significantly—
and similarly—increased SOD, and TAC activities compared
to the FO group (Table 6). In addition, CocO significantly
increased CAT compared to the FO. Besides, CocO- and MOTsupplemented diets significantly decreased plasma MDA and the
MDA: TAC ratio compared to the FO diet. In addition, CanO
substantially reduced MDA: TAC ratio compared to FO group,
and MAD to some extent. In general, CocO supplementation
induced the most substantial effect on the antioxidant profiles,
while FO enrichment promoted the smallest influence. It is well
known that MDA increased due feeding PUFA and particularly
during hot weather condition due to peroxidation process (44).
This beneficial effect of oils on the antioxidant profile shows the
essence of oil supplementation during hot weather condition,
particularly when a combined with antioxidant supplementation
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Hepatic Plasma Leakage Enzymes
Table 6 also shows the effect of different oil sources on the plasma
hepatic leakage enzymes of 42-day-old broilers. AST, ALT, and
AlkP were significantly lower in chickens fed the CocO-enriched
diet compared to those of the other oil groups. Furthermore,
broiler chicks fed FO and MTO presented considerably decreased
plasma AST activity compared to the CanO groups and raised
AlkP compared to the CocO group. These results indicate a
beneficial effect of PUFA on hepatic cell membrane integrity
that might be due to enriched phospholipids as an essential part
of cell membrane integrity containing two hydrophobic LCFA
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(4, 5, 65). The beneficial effects of CocO on liver leakage enzymes
as previously said might be due to its antioxidant (27) and
antimicrobial effects (74, 75). In literature, oil supplementation
improved liver function (44, 45). However, Yildirim et al. (66)
found that dietary fat sources, including cocoa butter, did not
affect plasma AST, ALT and AlkP in rats. The differences among
the studies might be due to the type of diets, levels and the kind
of lipids, strain, age and sex, and species of animals (73).
Serum Antibody Titer
None of the oil sources affected the HI test for infectious
bronchitis virus (HIBB) or bursa disease (HIBD) (Table 6). On
the other hand, broilers fed a diet supplemented with 50 g/kg
FO show a higher production of antibodies (IgM and IgG)
and globulins in the serum and maintain immune function
after vaccination compared to the control group (79). In the
present study, the addition of CocO resulted in a considerable
increase in the HIAI compared to the CanO and MTO groups. In
addition, the CocO-supplemented group exhibited significantly
increased HIND compared to the FO-group. In general, CocO
induced the highest antibody titers to HIAI and HIND. MCFA
can be absorbed through the portal circulation after ingestion.
Subsequently, they are metabolized by hepatocytes into ketones
and used as an energy source. This process maintains the
serum leptin level and promotes ketogenesis during shortterm fasting and energy restriction (80). Leptin can enhance
immune functions, including inflammatory cytokine production
in macrophages, granulocyte chemotaxis, and increased Th17
proliferation (81). The ketogenic diet activates a subset of T
cells in the lungs not previously associated with the immune
system’s response to influenza; this action enhances mucus
production by airway cells that can effectively trap the virus
(82). Nonetheless, the level and source of dietary fat did
not significantly influence antibody titer against NDV at 42
and 70 days of age (80, 83), but had a positive effect on
antibody in other experiments (44, 45). In humans, dietary
CocO normalize body lipids, protect the liver from alcohol
damage, and improve the immune system’s anti-inflammatory
response (27, 74, 75). Similarly, Yaqoob (12) suggested that
the impacts of MUFA on adhesion molecules are potentially
crucial, namely through their roles in the pathology of several
diseases involving the immune system. There is some evidence
that the olive oil effects on immune function in animal
studies are due to oleic acid rather than to trace elements or
antioxidants. Kelley and Daudu (84) reported that in animals
fats could inhibit and stimulate processes, depending upon the
species, the utilized fatty acids, and the index being examined.
These findings suggest that the absolute amounts or the ratios
between individual fatty acids or fatty acid classes are critical in
determining their effects on the immune response and need to be
investigated (12, 83).
It is worth noticing that, FO supplementation resulting in
increasing HIAI numerically and correlated with an increase
in γ-globin and a decrease in serum albumin. Dietary ω3
PUFA (FO or linseed oil) has a positive significant impact on
humoral immunity, i.e., antibody titers against NDV compared
to the control diet (18). Furthermore, FO supplementation
significantly increases antibody titers and the relative weight
percentages of the bursa and spleen compared to the control
under normal and hot climate condition (20, 44, 45). FO has
no adverse effects on the immune function of broilers (19).
On the other hand, birds fed a diet with an ω6: ω3 ratio that
exceeds the recommended levels exhibit damage to the intestinal
epithelial cells (21). Further, a low ω6: ω3 ratio in the diet
increases MDA in tissues, including the meat, and thus impairs
meat quality.
Immune Status
Serum Protein Profile
Table 6 also shows the influence of different oil sources on
serum immune indices of the broilers. The different oil sources
had no effect on α2-globulin or albumin to globulin ratio
measured at day 42 of age. The FO- and CanO-supplemented
diets similarly increased the total serum protein compared to
the CocO and MTO group. In general, the improvement in
protein use with FO and CanO supplementation found herein
can enhance the immune response, and the antibodies are
proteinic in nature. The increased serum protein of broilers fed
the FO- or CanO-enriched diet are in general agreement with
the effect of oils on improving nutrient absorption (including
protein) under hot condition (3, 4, 45). The present results
confirmed this effect because the different FO, and CanO oil
sources increased chymotrypsin activity. The increased enzyme
activity is associated with raising nutrient digestibility and thus,
nutrient absorption (1, 44, 65).
The FO-supplemented diet significantly decreased the
percentage of albumin (a non-specific immune protein)
compared to the other oil groups. Likewise, FO supplementation
decreased T- and B-cell proliferation and delayed type
hypersensitivity as measurements of cell-mediated immunity
(76, 77).
Serum Immunoglobulin
The CocO-, and CanO-enriched diets significantly increased α1globulin compared to the FO and MTO groups. The difference
in β-globulin due to the different oil sources showed that
FO and CanO induced substernal increase compared to CocO
and MTO. The γ-globulin was raised considerably by MTOenriched diet compared to the CocO, and CanO- enriched
groups. Immunoglobulins (γ-globulin) play a vital role in natural
and acquired immunity to infections (78). Globulins—a major
family of proteins—are an essential source of the protein present
in animal fluids, including the enzymes, antibodies, and fibrous
and contractile proteins usually found in the blood plasma (73).
α-and β-globulins are transport proteins, serve as substrates upon
which other substances are formed, and perform other diverse
functions (78).
Different oil sources did not meaningfully affect IgA, but IgG
and IgM were decreased considerably due to FO supplementation
compared to the other oil groups. However, Yildirim et al. (66)
revealed no marked changes in IgG among their experimental
groups. Besides, the IgM levels were significantly reduced
in cocoa butter alone and cocoa butter + sunflower oil
group as compared to the sunflower-oil-enriched diet and the
control group.
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Morphometric Measurements of the Villi,
Bursa of Fabricius and Thymus
is essential to optimize cellular and humoral immunity in
broilers (84).
In conclusion, oil supplementations at 1.5% enhanced growth
performance and immune status, improved the blood lipid
profile and antioxidants status, and the effect of the oil sources
depends on the criteria of response.
The villus height and villus height: depth ratio were significantly
affected by dietary oil supplementation (Table 6). The MTO
significantly increased the villus height compared to the FO
and CocO groups, and increased the villus height: depth ratio
compared to CocO group. These results indicate that FO, CanO
and MTO increase the absorption capacity of the intestine
and thus might be responsible for improving the growth and
immunity of broilers fed these diets. This is an essential influence
during hot weather condition due to impaired digestion and
absorption of nutrients (44, 45, 65, 73).
Follicle parameters and thymus cortex depth were
significantly affected by the source of oils. The addition of
CanO and MTO caused the highest increase in follicle length
compared to the other sources of oil, but FO significantly
increased the follicle width compared to the CanO group.
The follicle length: width ratio was significantly decreased in
broilers fed the FO-enriched diet compared to the CanO and
MTO groups.
The thymus cortex depth was significantly increased for
CocO-supplemented diets compared with the other oil groups.
Similarly, the source of fat had significant effects on the
weight and percentage of thymus and ratio of the bursa of
the Fabricius to body weight (4, 5). Nonetheless, the weight
of the bursa of Fabricius and weight and percentage of the
spleen, humoral immune response to SRBC injections or to
NDV, IBDV or IBV vaccinations were not affected by oil
treatments (4, 5). The addition of 2.5 or 5% soybean oil
to the diet increases the percentage of spleen and bursa,
respectively, compared to other groups (83). Taken together,
the fatty acid profile rather than the specific dietary lipid
DATA AVAILABILITY STATEMENT
The datasets generated for this study are available on request to
the corresponding author.
ETHICS STATEMENT
The experimental procedures were approved by Deanship
of Scientific Research, King Abdulaziz University, Jeddah,
Saudi Arabia under protocol number (DF-312-155-1441 H) that
recommends animal rights, welfare and minimal stress and did
not cause any harm or suffering to animals according to the Royal
Decree number M59 in 14/9/1431H.
AUTHOR CONTRIBUTIONS
All authors listed have made a substantial, direct and intellectual
contribution to the work, and approved it for publication.
FUNDING
This project was funded by the Deanship of Scientific Research
(DSR), King Abdulaziz University, Jeddah, under grant No. (DF312-155-1441 H). The authors, therefore, gratefully acknowledge
DSR technical and financial support.
REFERENCES
1. Chwen LT, Foo HL, Thanh NT, Choe D. Growth performance, plasma
fatty acids, villous height and crypt depth of preweaning piglets fed with
medium chain triacylglycerol. Asian Aust J Anim Sci. (2013) 26:700–4.
doi: 10.5713/ajas.2012.12561
2. Attia YA, Abd El-Hamid EAH, Nagadi SA, de Oliveira MC, Bovera F, Habiba
HI. Dietary distilled fatty acids and antioxidants improve nutrient use and
performance of Japanese male quails. Anim Sci Pap Rep. (2019) 37:65–74.
doi: 10.13140/RG.2.2.23000.03846
3. Baião NC, Lara LJC. Oil and fat in broiler nutrition. Braz J Poult Sci. (2005)
7:129–41. doi: 10.1590/S1516-635X2005000300001
4. Poorghasemi M, Seidavi A, Qotbi AAA, Chambers JR, Laudadio V, Tufarelli
V. Influence of dietary fat source on growth performance responses and
carcass traits of broiler chicks. Asian Aust J Anim Sci. (2013) 26:705–10.
doi: 10.5713/ajas.2012.12633
5. Poorghasemi M, Seidavi A, Qotbi AAA, Chambers JR, Laudadio V, Tufarelli V.
Effect of dietary fat source on humoral immunity response of broiler chickens.
Eur Poult Sci. (2015) 79. doi: 10.1399/eps.2015.92
6. Abdulla NR, Loh TC, Akit H., Sazili A., Foo HL, Mohamad R., et al. Fatty
acid profile, cholesterol and oxidative status in broiler chicken breast muscle
fed different dietary oil sources and calcium levels. S Afr J Anim Sci. (2015)
45:153–63. doi: 10.4314/sajas.v45i2.6
7. Alzawqari MH, Al-Baddany AA, Al-Baadani HH, Alhidary IA, Khan RU, Aqil
GM, et al. Effect of feeding dried sweet orange (Citrus sinensis) peel and lemon
grass (Cymbopogon citratus) leaves on growth performance, carcass traits,
Frontiers in Veterinary Science | www.frontiersin.org
8.
9.
10.
11.
12.
13.
14.
10
serum metabolites and antioxidant status in broiler during the finisher phase.
Environ Sci Pollut Res. (2016) 23:17077–82. doi: 10.1007/s11356-016-6879-7
Khatun J, Loh TC, Akit H, Foo HL, Mohamad R. Influence of different
sources of oil on performance, meat quality, gut morphology, ileal digestibility
and serum lipid profile in broilers. J Appl Anim Res. (2017) 46:479–85.
doi: 10.1080/09712119.2017.1337580
Raza T, Chand N, Khan RU, Shahid MS, Abudabos AM. Improving the
fatty acid profile in egg yolk through the use of hempseed (Cannabis
sativa), ginger (Zingiber officinale), and turmeric (Curcuma longa) in the
diet of Hy-Line White Leghorns. Arch Anim Breed. (2016) 68:183–90.
doi: 10.5194/aab-59-183-2016
Zaki EF, El Faham A, Nematallah GM. Fatty acids profile and quality
characteristics of broiler chicken meat fed different dietary oil sources
with some additives. Int J Health Anim Sci Food Saf. (2018) 5:40−50.
doi: 10.13130/2283-3927/9581
Galli C, Calder PC. Effects of fat and fatty acid intake on inflammatory and
immune responses: a critical review. Ann Nutr Metab. (2009) 55:123–39.
doi: 10.1159/000228999
Yaqoob P. Monounsaturated fats and immune function. Braz J Med BiolRes.
(1998) 31:453–65. doi: 10.1590/S0100-879X1998000400001
Swiatkiewicz S, Arczewska-Włosek A, Józefiak D. The relationship between
dietary fat sources and immune response in poultry and pigs: an updated
review. Livest Sci. (2015) 180:237–46 doi: 10.1016/j.livsci.2015.07.017
Nobakht A, Tabatbaei S, Khodaei S. Effects of different sources and levels of
vegetable oils on performance, carcass traits and accumulation of vitamin E in
breast meat of broilers. Cur Res J Biol Sci. (2011) 3:601–5.
April 2020 | Volume 7 | Article 181
Attia et al.
Oils and Broilers’ Performance and Immunity
15. Alagawany M, Elnesr SS, Farag MR, Abd El-Hack ME, Khafaga AF,
Taha AE, et al. Omega-3 and omega-6 fatty acids in poultry nutrition:
effect on production performance and health. Animals. (2019) 9:573.
doi: 10.3390/ani9080573
16. Shiels MI. Fatty acids and early human development. Early Hum Develop.
(2007) 83:761–6. doi: 10.1016/j.earlhumdev.2007.09.004
17. Smith M, Soisuvan K, Miller L. Evaluation of dietary calcium level and fat
source on growth performance and mineral utilization of heat-distressed
broilers. Poult Sci. (2003) 2:32–7. doi: 10.3923/ijps.2003.32.37
18. Ebeid T, Eid Y, Saleh A, Abd El-Hamid H. Ovarian follicular
development, lipid peroxidation, antioxidative status and immune
response in laying hens fed fish oil-supplemented diets to produce n3-enriched eggs. Animal. (2008) 2:84–91. doi: 10.1017/S17517311070
00882
19. Al-Khalifa HS, Givens D, Rymer C, Yaqoob P. Effect of n-3 fatty acids
on immune function in broiler chickens. Poult Sci. (2012) 91:74–88.
doi: 10.3382/ps.2011-01693
20. Jameel YJ, Sahib AM, Husain MA. Effect of dietary omega-3 fatty acid on
antibody production against Newcastle disease in broilers. Int J Sci Nat.
(2015) 6:23–7.
21. Konieczka P, Czauderna M, Smulikowska S. The enrichment of chicken meat
with omega-3 fatty acids by dietary fish oil or its mixture with rapeseed
or flaxseed-effect of feeding duration dietary fish oil, flaxseed, and rapeseed
and n-3 enriched broiler meat. Anim Feed Sci Technol. (2017) 223:42–52.
doi: 10.1016/j.anifeedsci.2016.10.023
22. López-Ferrer S, Baucells MD, Barroeta AC, Grashorn MA. n-3 enrichment
of chicken meat. 1. Use of very long-chain fatty acids in chicken diets
and their influence on meat quality: fish oil. Poult Sci. (2001) 80:741–52.
doi: 10.1093/ps/80.6.741
23. Lesson S, Summers JD. Nutrition of the Chickens. 4th Edn. Guelph, ON:
University Books (2001).
24. Thacker; PA, Petri D. Nutritional evaluation of canola protein concentrate
for broiler chickens. Asian Aust J Anim Sci. (2011) 24:1607–14.
doi: 10.5713/ajas.2011.11161
25. Andreotti MO, Junqueira OM, Cancherini LC, Rodrigues EA, Sakomura NK.
Valor nutricional de algumas fontes de gordura para frangos de corte. In:
Anais da 38◦ Reunião Anual da Sociedade Brasileira de Zootecnia. Piracicaba:
SBZ (2001).
26. Souza PA, Souza HBA, Oba A, Leonel FR, Pelicano ERL, Norkus EA, et al.
Características físicas e químicas da carne da coxa de frangos de corte
produzidos com diferentes fontes de óleo. In: 38◦ Reunião Anual da Sociedade
Brasileira de Zootecnia. Piracicaba: FEALQ (2001).
27. Piracicaba SP, Piracicaba, SBZ; Bhatnagar AS, Kumar P, Hemavathy J, Krishna
G. Fatty acid composition; oxidative stability; and radical scavenging activity
of vegetable oil blends with coconut oil. J Am Oil Chem Soc. (2009) 86:991–9.
doi: 10.1007/s11746-009-1435-y
28. Ferreira WM. Digestão e Metabolismo dos Lipídios. Belo Horizonte: Escola de
Veterinária da UFMG (1999). pp. 1–34.
29. Rubin M, Moser A, Vaserberg N, Greig F, Levy Y, Spivak H, et al.
Long-chain fatty acids; in long-term home parenteral nutrition: a
double-blind randomized cross-over study. Nutrition. (2000) 16:95–100.
doi: 10.1016/S0899-9007(99)00249-X
30. Rego Costa AC, Rosado EL, Soares-Mota M. Influence of the dietary
intake of medium chain triglycerides on body composition; energy
expenditure and satiety; a systematic review. Nutr Hosp. (2012) 27:103–8.
doi: 10.1590/S0212-16112012000100011
31. Han J, Hamilton JA, Kirkland JL, Corkey BE, Guo W. Medium-chain oil
reduces fat mass and down-regulates expression of adipogenic genes in rats.
Obes Res. (2003) 11:734–44. doi: 10.1038/oby.2003.103
32. St-Onge MP, Ross R, Parsons WD, Jones PJ. Medium chain triglyceri+des
increase energy expenditure and decrease adiposity in overweight men. Obes
Res. (2003) 11:395–402 doi: 10.1038/oby.2003.53
33. Takeuchi H, Noguchi O, Sekine S, Kobayashi A, Aoyama T. Lower weight
gain and higher expression and blood levels of adiponectin in rats fed
medium+-chain TAG compared with long-chain TAG. Lipids. (2006) 41:207–
12. doi: 10.1007/s11745-006-5089-3
34. Xie P, Wang Y, Wang C, Yuan C, Zou X. Effect of different fat sources in
parental diets on growth performance; villus morphology; digestive enzymes
Frontiers in Veterinary Science | www.frontiersin.org
35.
36.
37.
38.
39.
40.
41.
42.
43.
44.
45.
46.
47.
48.
49.
50.
51.
52.
53.
54.
11
and colorectal microbiota in pigeonsquabs. Arch Anim Nutr. (2013) 6:147–60.
doi: 10.1080/1745039X.2013.776329
Santos F, Donoghue AM, Venkitanarayanan K, Dirain ML, Reyes-Herrera
I, Blore PJ, et al. Caprylic acid supplemented in feed reduces enteric
campylobacter jejuni colonization in ten-day-old broiler chickens. Poult Sci.
(2008) 87:800–4. doi: 10.3382/ps.2007-00280
Adams BYC, Vahl HA, Veldman A. Interaction between nutrition and Eimeria
acervulina infection in broiler chickens: diet compositions that improve fat
digestion during Eimeria acervulina infection. Br J Nutr. (1996) 75:875–80.
doi: 10.1079/BJN19960193
Attia YA, Al-Harthi MA, Korish MM, Shiboob MM. Fatty acid and
cholesterol profiles and hypocholesterolemic; atherogenic; and thrombogenic
indices of table eggs in the retail market. Lipids Health Dis. (2015) 14:136.
doi: 10.1186/s12944-015-0133-z
Attia YA, Al-Harthi MA, Korish MM, Shiboob MM. Fatty acid and
cholesterol profiles; hypocholesterolemic; atherogenic; and thrombogenic
indices of broiler meat in the retail market. Lipids Health Dis. (2017) 16:40.
doi: 10.1186/s12944-017-0423-8
Zollitsch W, Knaus W, Aichinger F, Lettner F. Effects of different dietary fat
sources on performance and carcass characteristics of broilers. Anim Feed Sci
Technol. (1997) 66:63–73. doi: 10.1016/S0377-8401(96)01126-1
Ayed H.B, Attia H, and Ennouri M. Effect of oil supplemented diet on growth
performance and meat quality of broiler chickens. Adv Tech Biol Med. (2015)
4:156. doi: 10.4172/2379-1764.1000156
Wood JD, Richardson RI, Nute GR, Fisher AV, Campo MM, Kasapidou E,
et al. Effects of fatty acids on meat quality: a review. Meat Sci. (2003) 66:21–32.
doi: 10.1016/S0309-1740(03)00022-6
Velasco S, Ortiz LT, Alzueta C, Rebole A, Trevino J, Rodriguez ML.
Effect of inulin supplementation and dietary fat source on performance;
blood serum metabolites; liver lipids; abdominal fat deposition; and tissue
fatty acid composition in broiler chickens. Poult Sci. (2010) 89:1651–62.
doi: 10.3382/ps.2010-00687
NRC. Nutrient Requirement of Poultry. National Academy of Sciences. 9th ed.
Washington, DC: National Research Council (1994).
Attia YA, Al-Harthi MA, Elnaggar ASh. Productive, physiological and
immunological responses of two broiler strains fed different dietary
regimens and exposed to heat stress. Ital J Anim Sci. (2018) 17:686–97.
doi: 10.1080/1828051X.2017.1416961
Attia, YA, Hassan, SSh. Broiler tolerance to heat stress at various
dietary protein/energy levels. Europ Poult Sci. (2017) 81:2017.
doi: 10.1399/eps.2017.171
Grasse M, Rosenkrands I, Olsen A, Follmann F, Dietrich J. A flow cytometrybased assay to determine the phagocytic activity of both clinical and
nonclinical antibody samples against Chlamydia trachomatis. Cytometry A.
(2018) 93:525–32. doi: 10.1002/cyto.a.23353
King DJ, Seal BS. Biological and molecular characterization of Newcastle
disease virus (NDV) field isolates with comparisons to reference NDV strains.
Avian Dis. (1998) 42:507–16. doi: 10.2307/1592677
Cosgrove A. An apparently new disease of chickens: avian nephrosis. Avian
Dis. (1962) 6:385–9. doi: 10.2307/1587909
Takatsy G. The use of spiral loops in serological and virolegical micromethods.
Acta Microbiol Acad Sci Hung. (1956) 3:197.
Friedewald WT, Levy RI, Fredrickson DS. Estimation of the concentration
of low-density lipoprotein cholesterol in plasma; without use of
the preparative ultracentrifuge. Clin Chem. (1972) 18:499–502.
doi: 10.1093/clinchem/18.6.499
Pinchasov Y, Nir I, Nitsan Z. Metabolic and anatomical adaptations of heavy
bodies chicks to intermittent feeding. 2. pancreatic digestive enzymes. Br Poult
Sci. (1990) 31:769–77. doi: 10.1080/00071669008417307
Sklan; D., Halevy O. Protein digestion and absorption along
the ovine gastrointestinal tract. J Dairy Sci. (1985) 68:1676–81.
doi: 10.3168/jds.S0022-0302(85)81013-4
Sklan D, Hurwitz S, Budowski P, Ascarelli A. Fat digestion and absorption
in chicks fed raw or heated soybean meal. J Nutr. (1975) 105:57–63.
doi: 10.1093/jn/105.1.57
Attia YA, Al-Khalaifah HS, Abd El-Hamid HE, Al-Harthi MA, El-shafey
AA. Effect of different levels of multi-enzymes on immune response; blood
hematology and biochemistry; antioxidants status and organs histology of
April 2020 | Volume 7 | Article 181
Attia et al.
55.
56.
57.
58.
59.
60.
61.
62.
63.
64.
65.
66.
67.
68.
69.
70.
71.
Oils and Broilers’ Performance and Immunity
72. Vermaak WJ, Kalk WJ, Kuyl JM, Smit AM. Fatty acid induced
changes in circulating total and free thyroid hormones: in vitro effects
and methodological artefacts. J Endocrinol Invest. (1986) 9:121–6.
doi: 10.1007/BF03348081
73. Attia YA, Böhmer Barbara M, Roth-Maier Dora A. Responses of broiler chicks
raised under constant relatively high ambient temperature to enzymes, amino
acid supplementations, or a high-nutrient diet. Archiv Für Geflügelkunde.
(2006) 70:80–91.
74. Enig MG. Diet; serum cholesterol coronary heart disease. In: Mann GV,
editor. Coronary Heart Disease: The Dietary Sense Nonsense. London: Janus
Publishing (1993). pp. 36–60.
75. Enig MG. Lauric oils as antimicrobial agents: theory of effect; scientific
rationale; dietary applications as adjunct nutritional support for HIV-infected
individuals. In: Watson RR, editor. Nutrients Foods in AIDS. Boca Raton, FL:
CRC Press (1998). pp. 81–97.
76. Meydani SN, Ribaya-Mercado JD, Russell RM, Sahyoun N, Morrow FD,
Gershoff SN. Vitamin B-6 deficiency impairs interleukin 2 production and
lymphocyte proliferation in elderly adults. Am J Clin Nutr. (1991) 53:1275–80.
doi: 10.1093/ajcn/53.5.1275
77. Kramer IM, Koornneef I, de Laat SW, Adriana JM, Raaij AJM. TGFf1 induces phosphorylation of the cyclic AMP responsive element
binding protein in ML-CC164 cells. EMBO J. (1991) 10:1083–9.
doi: 10.1002/j.1460-2075.1991.tb08048.x
78. Attia YA, Al-Khalifa H, Ibrahim MS, Abd Al-Hamid AE, Al-Harthi MA, ElNaggar ASh. Blood hematological and biochemical constituents; antioxidant
enzymes; immunity and lymphoid organs of broiler chicks supplemented
with propolis; bee pollen and mannan oligosaccharides continuously or
intermittently. Poult Sci. (2017) 96:4182–92. doi: 10.3382/ps/pex173
79. Al-Mayah AAS. Effect of fish oil on humoral immunity of broiler chicks.
Basrah J Vet Res. (2009) 8:23–32. doi: 10.33762/bvetr.2009.56864
80. Leung YB, Cave NJ, Heiser A, Edwards PJB, Godfrey AJR, Wester T.
Metabolic and immunological effects of intermittent fasting on a ketogenic
diet containing medium-chain triglycerides in healthy dogs. Front Vet Sci.
(2020) 6:480. doi: 10.3389/fvets.2019.00480
81. Naylor C, Petri WA Jr. Leptin regulation of immune responses.
Trends Mol Med. (2016) 22:88–98. doi: 10.1016/j.molmed.2015.
12.001
82. Goldberg EL, Molony RD, Kudo E, Sidorov S, Kong Y, Dixit VD, et al.
Ketogenic diet activates protective γδ T cell responses against influenza virus
infection. Sci Immunol. (2019) 4:eaav2026. doi: 10.1126/sciimmunol.aav2026
83. Kelley DS, Daudu PA. Fat intake and immune response. Prog Food Nutr Sci.
(1993) 17:41−63.
84. Omidi S, Mohit A, Zadeh NGH. Effect of dietary fat level and source on
performance and immune system response of turkeys. Acta Sci Anim Sci.
(2020) 4:e46775. doi: 10.4025/actascianimsci.v42i1.46775
broiler chicks fed standard and low-density diets. Front Vet Sci. (2020) 6:510.
doi: 10.3389/fvets.2019.00510
AOAC. Official Methods of Analysis. Washington, DC: Association of Official
Analytical Chemists (2004).
SAS, Institute. SAS R User’s Guide for Personal Computer. Cary, NC: SAS
Institute Inc. (2004).
Wang J, Wang X, Li J, Chen Y, Yang W, Zhang L. Effects of dietary coconut
oil as a medium-chain fatty acid source on performance; carcass composition
and serum lipids in male broilers. Asian Aust J Anim Sci. (2015) 28:223–30.
doi: 10.5713/ajas.14.0328
Htin NN, Zulkifli I, Alimon AR, Loh TC, Hair-Bejo M. Effects of sources of
dietary fat on broiler chickens exposed to transient high temperature stress.
Arch Geflügelk. (2007) 71:74–80.
Dawood HY, Mohammed OE. Effect of the dietary fat sources and levels
on broiler performance and carcass-serum lipids. Am J Innov Res Appl Sci.
(2015) 1:318–31.
Atteh JO, Leeson S. Effects of dietary saturated or unsaturated fatty acids
and calcium levels on performance and mineral metabolism of broiler chicks.
Poult Sci. (1984) 63:2252–60. doi: 10.3382/ps.0632252
Wignjosoesastro N, Brooks CC, Herrick RB. The effect of coconut meal and
coconut oil in poultry rations on the performance of laying hens. Poult Sci.
(1972) 51:1126–1132. doi: 10.3382/ps.0511126
Jang IS, Ko YH, Kang SY, Lee CY. Effect of a commercial essential oil
on growth performance; digestive enzyme activity and intestinal microflora
population in broiler chickens. Anim Feed Sci Technol. (2007) 134:304–15.
doi: 10.1016/j.anifeedsci.2006.06.009
Nash DM, Hamilton RMG, Hulan HW. The effect of dietary herring
meal on the omega-3 fatty acid content of plasma and egg yolk lipids
of laying hens. Can J Anim Sci. (1995) 75:247–53. doi: 10.4141/cja
s95-036
Kiani A, Sharifi SD, Ghazanfari S. Influence of canola oil and
lysine supplementation diets on growth performance and fatty acid
composition of meat in broiler chicks. Int J Anim Vet Sci. (2017)
11:134−40.
Aggoor FAM, Attia YA, Qota EMA. A study on the energetic efficiency of
different fat sources and levels in broiler chick vegetable diets. Mansoura Univ
J Agric Sci. (2000) 25:801–20.
Yildirim E, Çinar M, Yalçinkaya I, Ekici H, Atmaca N, Güncüm E.
Effect of cocoa butter and sunflower oil supplementation on performance,
immunoglobulin, and antioxidant vitamin status of rats. BioMed Res Int.
(2014) 2014:606575. doi: 10.1155/2014/606575
Özdogan M, Akşit M. Effects of feeds containing different fats on carcass
and blood parameters of broilers. J Appl Poult Res. (2003) 12:251–6.
doi: 10.1093/japr/12.3.251
Lachowicz K, Koszela-Piotrowska I, Rosołowska-Huszcz D. Thyroid hormone
metabolism may depend on dietary fat. J Anim Feed Sci. (2008) 17:110–9.
doi: 10.22358/jafs/66475/2008
Lachowicz K, Koszela-Piotrowsk I, Rosołowska-Huszcza D. Dietary
fat type and level affect thyroid hormone plasma concentrations in
rats. J Anim Feed Sci. (2009) 18:541–50. doi: 10.22358/jafs/66430/
2009
Yamamoto N, Li QL, Mita S, Morisawa S, Inoue A. Inhibition of thyroid
hormone binding to the nuclear receptor mobilization of free fatty acids.
Hormone Metab Res. (2001) 33:131–7. doi: 10.1055/s-2001-14939
Otten MH, Hennemann G, Docter R, Visser TJ. The role of dietary fat
in peripheral thyroid hormone metabolism. Metabolism. (1980) 29:930–5.
doi: 10.1016/0026-0495(80)90035-9
Frontiers in Veterinary Science | www.frontiersin.org
Conflict of Interest: The authors declare that the research was conducted in the
absence of any commercial or financial relationships that could be construed as a
potential conflict of interest.
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