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Brazilian Journal of Poultry Science

Print version ISSN 1516-635XOn-line version ISSN 1806-9061

Braz. J. Poult. Sci. vol.21 no.2 Campinas  2019  Epub Nov 14, 2019

https://doi.org/10.1590/1806-9061-2018-0915 

Original Article

Effect of Dietary Inclusion of Sodium Bicarbonate on Digestibility of Nutrients and Immune Response in Caged Layers During the Summer

IDepartment of Animal Production, Riphah College of Veterinary Sciences, Lahore, Pakistan.

IIDepartment of Poultry Science, Institute of Animal and Dairy Sciences, University of Agriculture, Faisalabad, Pakistan.

IIIFaculty of Animal Production and Technology, Cholistan University of Veterinary and Animal Sciences Bahawalpur 63100, Pakistan.

IVEast Mediterranean Agricultural Research Institute, Adana, Turkey.

VDepartment of Physiology and Biochemistry, Cholistan University of Veterinary and Animal Sciences Bahawalpur 63100, Pakistan.

VIDepartment of Biology, Lahore Garrison University Lahore, Pakistan.

VIIDepartment of Veterinary Microbiology, Faculty of Animal Husbandry and Veterinary Sciences, Sindh Agriculture University, Tandojam, Sindh Province, Pakistan.


ABSTRACT

The aim of this project was to investigate the effect of dietary inclusion of sodium bicarbonate (NaHCO3) on nutrient digestibility and immune response of caged layers during summer when the temperature exceeds 40 ˚C. For immune response trial, White Leghorn layers (n=160; 24 weeks old) were purchased from a poultry farm and were divided into five treatment/diets groups (4 replicate/treatment). Diet A, was without NaHCO3 and served as control whereas, diets B, C, D, and E contained 0.5, 1.0, 1.5 and 2.0% NaHCO3, respectively. All these birds were vaccinated against Newcastle disease (ND) virus at the start of the experiment and thereafter with one-month intervals. Blood samples were collected from two birds/replicate at 10 days post vaccination each time to check antibody titer against ND virus. For digestibility trial, fecal samples were collected (6 layers/treatment group) at the start of the 37th week of age for two days, at 3 hours interval. Results revealed that immune response against ND virus 10 days post vaccination after 1st, 2nd and 3rd month was significantly (p<0.05) higher in layer birds fed diets containing NaHCO3. Digestibility of dry matter (DM), crude protein (CP), crude fiber (CF), ether extract (EE) and absorption of minerals were also found to be significantly (p<0.05) higher in groups treated with NaHCO3 and the birds fed diet containing 1% NaHCO3 showed the best results. In general, results revealed that 1% supplementation of NaHCO3 in layers’ diet have a beneficial impact in terms of immunity and diet digestibility.

Keywords: Heat stress; Layers; Newcastle disease; Nutrient digestibility; Sodium bicarbonate

INTRODUCTION

Birds are able to maintain their body temperature within narrow limits and an increase in body temperature due to higher ambient temperature or excessive metabolic activities may cause irreversible thermoregulatory events that could be harmful to the existence of the birds (Abbas et al., 2017). Different researchers have found good result on the performance by the supplementation of AA in chickens (Saeed et al., 2018a and Saeed et al., 2018b) and heat stress has shown detrimental effects on feed intake, growth and feed conversion ratio of the birds (Al-Hassani et al., 2001; Ekanayake et al., 2004; Li et al., 2015; Saeed et al., 2017; Mohammed et al., 2018). This reduction of performance might be explained by decreased digestibility of nutrients, increased heat production and reduced protein retention (Fouad et al., 2016; Orhan et al., 2018). Birds exposed to heat stress has shown reduced amylase and maltase activities, decreased protein and amino acid digestibility of complete diets and individual feed ingredients (Bayati et al., 2017) and reduced minerals absorption (Belhadj et al., 2016; Goff, 2018). The gastrointestinal size was also reported to decrease in heat-exposed chickens (Orhan et al., 2018).

Sodium bicarbonate (NaHCO3) is a white solid crystalline compound soluble in water, which is commonly used as an antacid to treat acid indigestion. It is commonly added as a simple solution for restoring the pH of water that has a high level of chlorine (Whiting et al., 1991). Sodium bicarbonate in feed or water has shown potential benefits on production performance (Ahmad et al., 2005; Khattak et al., 2012; Peng et al., 2013), egg characteristics (Kaya et al., 2004; Jiang et al., 2015) and blood profile (Kurtoglu et al., 2007) in poultry birds exposed to heat stress.

Sodium bicarbonate in the diet of layers may improve nutrient digestibility by increasing sodium ions concentration (Fethiere et al., 1994); improving electrolyte balance in poultry diets (Borges et al., 2003); meeting the requirements for the HCO3 - ions (Gorman & Balnave, 1994) and decrease the losses caused by heat stress (Abbas et al., 2017). It is cheap, easily available and easy to handle, therefore, can be safely incorporated in poultry diets to ameliorate the adverse effects caused by heat stress.

There is some evidence on the beneficial effect of NaHCO3 on immune response (Fouad et al., 2016) and nutrient digestibility (Lin et al., 2006), in broilers exposed to high ambient temperature. However, information on the effect of NaHCO3 on immune response and nutrient digestibility in caged layers during summer are scanty. Therefore, the present trial was carried out to study the effects of dietary inclusion of sodium bicarbonate on immune response against Newcastle disease virus and in vivo digestibility of DM, CP, CF, and EE. The effect of the addition of this compound in poultry diets was also studied on the absorption of some minerals i.e. calcium, phosphorus, sodium, potassium, and iron, in caged layers during the summer.

MATERIALS AND METHODS

Birds and housing

All the animal experimentation protocols were approved by the Directorate of Graduate Studies, University of Agriculture (UAF), Faisalabad (Pakistan). The experiment was conducted during the summer season when the temperature exceeds 40 ˚C. One hundred sixty commercial layers of 24 weeks of age having initial body weight as1328±14.3 of group A, 1310 ±7.0 of group B, 1318 ±16.4 of group C, 1324 ±11.0 of group D, 1312 ±12.1 of group E, were purchased from a commercial poultry farm. These layers were divided into 20 experimental units/replicates (8 layers/ replicate). These replicates were further allotted to five treatment groups (4 replicate/ treatment). Experimental birds were maintained in individual cages in a thoroughly cleaned and disinfected Poultry House of the Department of Parasitology, Faculty of Veterinary Sciences, UAF. These birds were maintained under similar managemental conditions like floor space, relative humidity, temperature and light in the open house.

Initially, these birds were reared in a group and were fed commercial layer ration during the 24 first weeks of age as an adaptation period. Thereafter, at the start of the 25th week, all the birds were individually weighed and transferred randomly to the individual cages using Completely Randomized Design. Each cage was supplied with a feeder and drinker line. The length, width and height of each cage were 41, 39 and 37 cm, respectively. Daily, 17 hours of light was provided to the birds throughout the experiment. A dry bulb thermometer was installed in the center of the house to record daily ambient temperature. Whereas, daily records of relative humidity inside the poultry house were maintained by using a digital hygrometer.

Experimental diets

Five experimental diets i.e. A (control, without Sodium bicarbonate), B (0.5%Sodium bicarbonate), C (1 % Sodium bicarbonate), D (1.5% Sodium bicarbonate) and E (2% Sodium bicarbonate) were used. Before the start of the experiment, all the diets were analyzed for their proximate chemical composition according to the technique described by AOAC (2010), in the Analytical Laboratory of the Institute of Animal Sciences, Faculty of Animal Husbandry, University of Agriculture, Faisalabad (Pakistan). Proportions of ingredients used in the experimental diets are shown in table 1. A weighed amount of the experimental diets was fed twice a day (morning and evening). All the diets were iso-nitrogenous (CP 16 %) and iso-caloric (ME 2700 Kcal/Kg diet) and were fed to the experimental birds (NRC, 1994), from 25-36 weeks of age (12 weeks).

Table 1 Proportion of the ingredients used in the experimental diets and their proximate composition. 

Ingredients (%) A
Basal diet
B
0.5% NaHCO3
C
1% NaHCO3
D
1.5% NaHCO3
E
2% NaHCO3
Maize 31.50 28.00 29.00 30.60 30.60
Rice broken 30.20 30.00 30.00 30.00 30.00
Fish meal CP 48% 3.60 5.50 7.00 7.00 7.00
Soybean meal, CP 45% 17.00 1.50 0.00 2.00 4.40
Canola meal, CP 35% 4.50 14.00 13.60 11.60 8.40
Rapeseed meal, CP 34% 3.10 3.00 3.00 3.00 3.00
Guar meal, CP 38.5% 0.00 2.50 3.00 3.00 3.00
Corn gluten 60% 0.00 2.00 2.00 2.00 2.00
Rice polishing 0.00 2.20 2.00 0.00 0.00
Dicalcium phosphate 0.50 0.00 0.00 0.00 0.00
Limestone 9.00 9.00 8.70 8.70 8.70
Mineraland vitamin Premix 0.30 0.30 0.30 0.30 0.30
DL-methionine 0.13 0.08 0.07 0.07 0.09
Lys. sulphate 65% 0.00 0.15 0.14 0.13 0.12
Salt 0.23 0.00 0.17 0.17 0.18
Sodium bicarbonate 0.00 0.50 1.00 1.50 2.00
Allzyme 0.015 0.015 0.015 0.015 0.015
Lincomix 0.02 0.02 0.02 0.02 0.02
Proximate composition
ME Kcal/Kg 2700 2700 2700 2700 2700
Crude protein (%) 17.00 17.00 17.00 17.00 17.00
Crude fiber (%) 3.29 3.8 3.74 3.44 3.39
Crude fat (%) 3.27 3.9 3.9 3.67 3.65
Crude ash (%) 11.97 11.9 12.01 11.85 11.89

Determination of antibody titer against Newcastle disease virus

At the end of the 36th week, five ml of blood was collected from healthy adult birds (wing-web) in a screw-top test tube having 1 mg/ml EDTA as an anticoagulant. The test tube was gently rotated for the mixing of blood and anticoagulant, but great care was taken to avoid hemolysis. Heamagglutination inhibitions (HI) test for determination of serum antibody titre was determined as described by Maff (1984).

Determination of nutrient digestibility and absorption of minerals

A digestibility trial was conducted during the experiment, at 36 weeks of age. For this purpose a separate group of 30 pullets was obtained from the same batch used for the performance trial. These layers were randomly allotted to five treatments (6 birds/treatment) such that each bird served as a replicate. These pullets were fed rations mixed with cellite (acid insoluble ash; AIA) at the rate of 1% as a marker.

The birds of these groups were fed their respective diets for one week (week 37) to assure that the passage of marker (AIA) in the feces of the birds was stabilized (Sales & Janssens, 2003) and during this period feces were not collected. After stabilization of the marker in the feces (week 38), all the birds were offered the same amount of their respective diets. The feed offered to the birds was divided into two equal portions and half of the feed was given at 9:00 am, and the rest at 9:00 pm. The feed not eaten was removed from the feeders and weighed at the end of the digestibility period.

Excreta collections, which started at the 8th experimental day, were made for a period of 48 hours (2 consecutive days) at two hours interval. Excreta samples were immediately frozen after each collection. The samples (feed and excreta) were analyzed thus collected were dried, finely ground and then analyzed for the determination of digestibility of dry matter (DM), crude protein (CP), ether extract (EE) and crude fiber (CF) contents using the method described by AOAC (2010). The samples (feed and excreta) were also analyzed for their mineral contents (Ca, P, Na, K, Fe and Mg) using atomic absorption spectrophotometer (Perkin Elmer, Beaconsfield, UK).

Digestibility of the nutrient was calculated by the following formula (Sales & Janssens, 2003):

D(%)=100AcidinsolubleashinfeedAcidinsolubleashinfeces×NutrientinfecesNutienteinfedd×100

Statistical analysis

The data thus collected were subjected to statistical analysis for interpretation of results using completely randomized design (CRD). Treatment means were compared by the Least Significance Differences test (Steel et al., 1997).

RESULTS

Antibody titer against Newcastle disease virus 10 days post vaccination during the 1st, 2nd and 3rd month is given in table 2. Findings of the study depicted that serum antibody titre against Newcastle disease virus of the birds 10 days post 1st vaccination was significantly influenced due to dietary inclusion of NaHCO3 in their diets. Birds of group C, which were fed diet containing 1% NaHCO3, showed maximum serum antibody titer against Newcastle disease virus when compared to the birds of other treated groups.

Table 2 Effect of dietary inclusion of sodium bicarbonate on immune response against Newcastle disease of caged layers. 

Variables Treatment
A Control B 0.5%NaHCO3 C 1%NaHCO3 D 1.5%NaHCO3 E 2%NaHCO3
1st Vaccination 43 d 86 cd 167a 145 ab 118 bc
2nd Vaccination 135 b 154 b 237 a 263 a 205 ab
3rd Vaccination 208 b 272 ab 368 a 288 ab 272 ab

Values within the same row which have different superscripts are significantly different (p<0.05).

Statistical analysis of the data revealed that birds using diets containing NaHCO3 exhibited significantly (p<0.05) higher serum antibody titer against Newcastle disease virus,10 days post the 2nd vaccination, when compared to those of the control group. Birds of group D, which were fed diet containing 1.5% NaHCO3, showed maximum serum antibody titer against Newcastle disease virus when compared to the birds of other treated groups. Findings of the study depicted that serum antibody titer against Newcastle disease virus of the birds 10 days post the 3rd vaccination was significantly influenced due to the dietary inclusion of NaHCO3 in their diets. Birds of group C, which were fed diet containing 1% NaHCO3, showed maximum serum antibody titer against Newcastle disease virus when compared to the birds of other treated groups.

Mean values regarding digestibility of DM, CP, CF and EE in birds fed diets with or without dietary inclusion of NaHCO3 are shown in table 3. The results revealed a significant (p<0.05) effect on DM digestibility due to the inclusion of NaHCO3 in the diets of layer when compared to those of the control group. The differences in DM digestibility values were also found to be significant among the treated groups. Birds using diet containing 1% NaHCO3 exhibited maximum digestibility followed by those of group D, B and E, whereas, the lowest DM digestibility was recorded in the control group.

Table 3 Effects of dietary inclusion of sodium bicarbonate on nutrient digestibility in layers. 

Variables Treatment
A Control B 0.5%NaHCO3 C 1%NaHCO3 D 1.5%NaHCO3 E 2%NaHCO3
Dry matter (%) 70.4± 6.09c 73.4±5.89b 77.1 ±5.79a 74.1± 6.34b 71.6±6.36c
Crude protein (%) 68.6±5.72c 72.7±4.65a 75±3.27a 72.2±3.38ab 69.2±5.44bc
Crude fibers (%) 29.0±2.81b 33.9±2.36ab 40.7 ±2.85a 33.2 ±2.1ab 30.9±2.95b
Ether extract (%) 82.0±5.57b 89.4± 4.02a 93.7± 5.46a 84.8±7.78b 82.7± 6.67b

Values within the same row which have different superscripts are significantly different (p<0.05)

The results showed a significant effect on CP digestibility due to the inclusion of NaHCO3 in the diets of layer when compared to those of the control group. Birds using diet containing 1% NaHCO3 exhibited maximum digestibility followed by those of group B, D and E, whereas, the lowest CP digestibility was recorded in the control group. Statistical analysis of the data depicted that the birds of treated groups, using diets containing NaHCO3 showed significantly (p<0.05) higher CF digestibility as compared to those of the control group. Birds using diet containing 1% NaHCO3 exhibited maximum digestibility followed by those of group B, D and E, whereas, the lowest CF digestibility was recorded in the control group. The results of the present study revealed a significant effect on EE digestibility due to the inclusion of NaHCO3 in the diets of the layer when compared to those of the control group. Birds using diet containing 1% NaHCO3 exhibited maximum digestibility followed by those of group B, D, and E, whereas, the lowest EE digestibility was recorded in the control group.

Mean values pertaining to the absorption of minerals i.e. calcium, phosphorous, iron, sodium and potassium in birds fed diets with or without the dietary inclusion of NaHCO3 are shown in table 4. The results revealed a significant effect on absorption of Ca due to the inclusion of NaHCO3 in the diets of the layer when compared to those of the control group. Birds using diet containing 1% NaHCO3 exhibited maximum absorption followed by those of group B, D and E, whereas, the lowest value was recorded in the controls.

Table 4 Effect of dietary inclusion of sodium bicarbonate on absorption (%) of the minerals in layers. 

Variables Treatment
A Control B 0.5%NaHCO3 C 1%NaHCO3 D 1.5%NaHCO3 E 2%NaHCO3
Calcium (%) 56.1± 3.29c 58.3±3.10b 60.3±3.20a 58.1± 3.28b 54.8 ±3.19c
Phosphorus (%) 50.8±2.32c 53.5±3.09b 57.6±3.27a 53±3.33b 49.6±2.40c
Iron (%) 50.5 ± 4.36d 53.1±3.38c 60.8 ±5.34a 55.6 ±5.2b 50±4.27d
Sodium (%) 51.6± 2.1c 54.1± 1.36b 59.2± 1.1a 53.5±1.11b 51.6.± 1.913b
Potassium (%) 51.8±2.10 c 53.5±2.47 b 56.3±3.64 a 53.5±3.64 b 52±3.47 c

Values within the same row which have different superscripts are significantly different (p<0.05)

The results revealed a significant effect on absorption of P due to the inclusion of NaHCO3 in the diets of layer when compared to those of the control group. The birds using diet containing 1% NaHCO3 exhibited maximum absorption followed by those of group B, D and E, whereas, the lowest P absorption was recorded in the control group. The results revealed a significant effect on absorption of Iron due to the inclusion of NaHCO3 in the diets of layer when compared to those of the control group. Birds using diet containing 1% NaHCO3 exhibited maximum absorption followed by those of group D, B and E, whereas the lowest iron absorption was recorded in the control group. The results revealed a significant effect on absorption of Na due to the inclusion of NaHCO3 in the diets of layer when compared to those of control group. Birds using diet containing 1% NaHCO3 exhibited maximum absorption followed by those of group B, D and E, whereas, the lowest Na absorption was recorded in the birds of the control group. The results revealed a significant effect on absorption of K due to the inclusion of NaHCO3 in the diets of layer when compared to those of the control group. Birds using diet containing 1% NaHCO3 exhibited maximum absorption followed by those of group B, D and E, whereas, the lowest digestibility of K was recorded in the control group.

DISCUSSION

Dietary inclusion of different levels of NaHCO3 depicted a significant increase in antibody titer against Newcastle disease in layers when compared to those fed diet without its addition. Environmental stressors have been known to affect immunity and innate resistance of the host directly or indirectly (Rakib et al., 2016). Therefore, increase in antibody titer against Newcastle disease virus in birds fed diets containing different levels of NaHCO3 may probably be due either to less heat stress upon these birds because of reduction in their body temperature or lower cortisol concentration as compared to those of control group, or both. Results of the present study are compatible with the findings of Khatak et al. (2012) who reported higher hemagglutination inhibition titer against Newcastle disease virus in birds consuming diets containing NaHCO3.

Borges et al. (2003) have observed that an increase in dietary electrolyte balance may cause a decrease in heterophil to lymphocyte ratio in blood, leading to an increase in antibody titer. Similarly, Santin et al. (2003) have reported a significant linear increase in Newcastle disease virus antibody titers with increasing DEB (40, 140, 240, 340mEq/kg), using NaCl, NaHCO3 and NH4Cl as supplements, Therefore, it may safely be concluded that the dietary addition of NaHCO3 may improve antibody titter against Newcastle disease virus in layers.

Diets containing NaHCO3 exhibited better digestibility of DM in layers. Increase in digestibility of dry matter (DM) of the treated groups may be due to more sodium ions concentration in the rations containing NaHCO3. A similar effect of increased sodium ions concentration in broilers has been observed by Fethiere et al. (1994). Dietary inclusion of NaHCO3 might have improved the electrolyte balance in the diet by creating favorable conditions for improvement in digestibility of nutrients (Borges et al., 2003; Mahmud et al., 2010).

Another probable explanation of better digestibility of DM in the birds fed diets containing NaHCO3 may be that pancreatic juices which are involved in digestion of most of the nutrients essentially contain NaHCO3. The presence of NaHCO3 in pancreatic juice, neutralizes the high acidity of chyme and raises it to be alkaline to prepare the chyme for the process of nutrient absorption, which takes place in the small intestine (Leeson & Summer, 2001). Therefore, increased digestibility of DM in treated groups may have been due to higher bicarbonate and sodium levels.

Heat stress may exert a negative influence on digestion and absorption of dietary nutrients as well as their metabolism (Puvadolpirod & Thaxton, 2000; Deraz, 2018), as has been observed in the birds of the control group. Therefore, decreased digestibility of DM in the control group may have been due to lower bicarbonate and sodium levels. On the other hand, the presence of NaHCO3 in the diets of treated birds might have improved their digestibility and prevented losses caused by heat stress (Mirsalimi et al., 1993). However, beneficial effects of NaHCO3 can be achieved only when its recommended optimum levels are incorporated in the diets. An excessive level of this chemical compound in the diet has been reported to be toxic in White Leghorn layers (Davison & Wideman, 1992). Therefore, this may be the probable reason for decreased digestibility of DM in group E, which was fed a diet containing 2% NaHCO3. Another reason for decrease in digestibility of dry matter in these birds might have been increased passage rate of digesta (Ravindran et al., 2008). However, Ahmad (1997) observed that DM digestibility in broilers was not influenced due to dietary inclusion of NaHCO3.

The birds fed diet without the inclusion of NaHCO3 (control) exhibited the lowest digestibility of protein. At an ambient temperature above 30°C, the thermoregulatory system is activated and causes an increase in blood flow to upper respiratory tract and other organs associated in heat excretion i.e., combs and wattles, which causes a decrease in blood flow to the digestive tract (Wolfenson, 1986). Consequently, activities of proteolytic enzymes in the upper part of the digestive system are decreased, ultimately leading to a decrease in protein digestibility. Considering the fact that heat stressed birds use glucogenic amino acids for glucose production (Nelson & Cox, 2000) during the process of gluconeogenesis in the birds, which is a metabolically expensive process (Nelson & Cox, 2000), provision of NaHCO3 in their diets can decrease glucose production from amino acids, which may lead to improved digestibility of protein during stress.

The addition of NaHCO3 in the diet of layers exhibited more digestibility of protein in these birds as compared to those of the control group. Protein consumed by the birds is broken down by the action of certain enzymes in the gastrointestinal tract to its constituent amino acids prior to absorption, and most of these amino acids require sodium (Leeson & Summer, 2001) for this process. Therefore, the increase in digestibility of protein of the treated groups may probably be due to the presence of more sodium ions concentration in the rations containing sodium bicarbonate. Sodium containing compounds such as sodium bentonite has been successfully used in sorghum containing diets to prevent deleterious effects of tannins present in it, on digestibility of protein (Pasha et al., 2008).

Dietary inclusion of different levels of NaHCO3 has depicted a significant increase in the digestibility of crude fibers in layers when compared to that fed diet without its addition. The increase in the digestibility of crude fibers of the treated groups may probably be due to the availability of more sodium and bicarbonate ions concentration in rations containing sodium bicarbonate. Dietary inclusion of NaHCO3 has also shown to improve electrolyte balance in poultry diets by creating physiological conditions favorable for improvement in digestibility of nutrients. Pasha et al. (2008) used different levels of sodium bentonite in broiler rations and found an improvement in nutrient digestibility as compared to the control group (without sodium bentonite). Similarly, Salari et al. (2006) have also observed improvement in nutrient digestibility because of the addition of sodium bentonite in broiler diets.

Digestibility of ether extract was found to be significantly better in the birds using diets containing NaHCO3 as compared to those of untreated group. Hyperthermia seems to be the most possible contributing factor for decreased digestibility and absorption of ether extract in the birds fed diet without sodium bicarbonate as have been observed by Koh & Macleod, (1999). Leeson & Summer, (2001), while discussing the factors affecting digestibility of fats, have also stated that fat digestibility is negatively affected in the birds exposed to heat stress. These findings are compatible with those observed in birds maintained under heat stress conditions.

Dietary inclusion of NaHCO3 significantly influenced the absorption of all the minerals (Ca, P, Fe, Na and K), which were studied in this trial. Birds using diets containing NaHCO3 exhibited better absorption of these minerals as compared to those of the untreated group. Minerals and trace elements are essential for optimum performance (Leeson & summers, 2001) of poultry birds. Therefore, increased mineral absorption in NaHCO3 fed birds may probably be due either to more availability of minerals as a result of increased feed intake or due to improved electrolyte balance or both (Borges et al., 2003).

CONCLUSIONS

The use of NaHCO3 proved to be a better choice to be included in the diets of commercial layers to reduce or at least ameliorate the harmful effects of heat stress on immune response against ND virus and nutrient digestibility during summer conditions.

ACKNOWLEDGMENTS

All the authors of the manuscript thank and acknowledge their respective Universities and Institutes and especially thankful to the Higher Education Commission of Pakistan (HEC) to support the scholar Ghulam Abbas for his project in Doctoral Degree.

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Received: October 03, 2018; Accepted: December 22, 2018

Corresponding author e-mail address Ghulam Abbas Department of Animal Production, Riphah College of Veterinary Sciences, Lahore, Pakistan. Phone: +90-322-3884500 Email: ghulamabbas_hashmi@yahoo.com

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These two authors equally contributed.

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