immune response

Glutamine represents approximately 30-35% of the total amount of nitrogen in the blood derived from amino acids. The role of glutamine as a nitrogen shuttle helps to protect against the toxic effects of high levels of ammonia in the blood. In addition, glutamine has vital and specific metabolic functions as a vehicle for carbon exchange between tissues, as a fuel for rapid cell division, and as a precursor of many naturally active molecules. It's a great example of how amino acids are used for different purposes in the metabolism and the immune system of the body. Glutamine, a conditionally essential amino acid, seems to be a key nutrient for the gut, as it may be a prominent source of vitality for the enterocytes. Glutamine appears to trigger an increase in the level of intestinal secretory immunoglobulin A, which is essential for mucosal defense. The investigation showed that the use of amino acid supplements in broiler diets improves the performance and the carcass characteristics. Glutamine is one of the most abundant amino acids in blood plasma and is involved in the building of muscle, and tissue, and gaining body weight. Researchers found that adding 10 g of L-glutamine per kg of food increased the weight gain and serum immunoglobulin A and G concentrations in broiler chickens. Therefore, an experiment was carried out in broiler chickens over 42 days of production to determine the effect of L-glutamine on broiler growth, carcass quality, and certain blood parameters.

The experiment was conducted using 300 one-day-old male Ross 308 broiler chickens in a completely randomized design with five treatments and five replicates from the age of 1 to 42 days. The experimental treatments included were: the addition of zero, 0.25, 0.5, 0.75, and 1% of L-glutamine in corn-soybean meal-based diets supplemented with dietary fat. During the experimental period, average daily feed intake (DFI), body weight gain (BWG), and feed conversion ratio (FCR) were measured. At the end of the experiment, two birds were selected from each replicate and slaughtered after weighing, and the weight of the internal organs (thigh, breast, heart, liver, gizzard, fat pad, pancreas, bursa, spleen, and thymus) was measured. Blood samples were collected from two birds per replicate to determine the number of white blood cells (WBC), and heterophile to lymphocyte ratio (H/L). Blood samples were taken from the jugular vein in tubes without anticoagulant and then centrifuged (2000×g for 10 min) to obtain a serum. On the 42nd day of rearing, serum cholesterol (total cholesterol, low-density lipoprotein (LDL), and high-density lipoprotein (HDL)) and triglycerides were determined by calorimetric examination.

Weight gain during the initial and growth periods was affected by L-glutamine supplementation (P<0.05). Despite higher feed intake for L-glutamine-supplemented diets during the initial period, this effect was significant only during the growth period (P<0.05). FCR and relative weight of internal organs, except liver weight, were not significantly different from the control group in the L-glutamine-supplemented chickens. The addition of 0.5% L-glutamine supplementation reduced cholesterol and triglyceride content and increased HDL in the blood of broilers (P<0.05). The H/L ratio in chickens that consumed L-glutamine was significantly lower than in the control group (P<0.05). L-glutamine has been reported to increase the activity of the intestinal enzyme Na-K ATPase, thereby indirectly increasing the absorption capacity of nutrients such as glucose and amino acids in the intestine. Researchers reported that L-glutamine increases the performance and feed intake of chickens by improving intestinal function and increasing digestibility. Expanding the villi of the small intestine can increase digestion and feed intake by increasing nutrient absorption and thus improving chicken performance. Improving the morphology of the digestive system is one of the possible mechanisms of the positive effect of L-glutamine on improving weight. Since glutamine stimulates anabolic conditions in the body and increases the amount of protein synthesis, therefore, along with muscle growth, it can increase the final body weight. Avian health is directly related to the immune system, and birds with adequate immune systems grow better. The majority of essential amino acids are recognized as critical resources for cytokine production and immune function.

Results from this study indicate that 0.5% dietary glutamine supplementation improved growth performance and had a positive effect on lipid metabolism-related blood parameters.

 Microelements in organisms have several structural, catalytic, and regulatory functions and play an important role in the functioning of the immune system. The trace mineral content of colostrum and milk is not optimal and in suckling calves, the inclusion of trace mineral supplements in the diet is necessary. Selenium (Se) is a basic mineral for humans and animals and has now been identified as an integral component of more than 35 selenoproteins which are involved in enzymatic, structural, or as yet unidentified functions. In livestock, Se is important for fertility and the prevention of diseases. Sufficient intake of Se increases fertility and enhances antioxidant protection systems and immunological potential. Se deficiency in cattle is associated with several problems, including delayed conception, muscular degenerative disease in calves, myocardial necrosis, heart failure, impaired immune function, increased risk of mastitis, abortion, perinatal mortality, and growth retardation in young calves. Based on the recommendation of the National Research Council, the daily requirements for Se and vitamin E in growing calves are 0.30 mg and 40 IU, respectively. To avoid Se deficiency and to promote animal health, inorganic sources of Se (sodium selenite and selenium) are routinely used as a supplement to the diet of farm animals. Zinc is involved in over 300 enzymes either as a component or as an activator. Zinc (Zn) is involved in immunity, metabolism, growth, and reproduction. It plays a role in the structure and function of antioxidant enzymes. Due to mineral interactions, the inorganic forms are less bioavailable in the gastrointestinal tract. The aim of this study was therefore to assess the effects of supplementation with sodium selenite and Zn oxide on growth performance, immune response, blood parameters, and antioxidant status in Holstein calves.

 The calves were kept in well-ventilated sheds with concrete floors and with individual feeding and watering facilities. The basal diet was prepared from locally available feed ingredients to meet the nutritional requirements of calves growing except for Zn and Se. Calves are fed milk at 10% of body weight in the morning and evening. The experiment started at four days of age and the calves were kept in individual pens for up to 42 days. The study used 36 Holstein calves with an average weight of 34.5±2 kg over 42 days in a completely randomized design. The basal diet (containing 40 mg Zn and 0.15 mg Se) was without Zn and Se supplementation (diet 1). Diet 2 contained 0.3 mg sodium selenite, diet 3 contained 40 mg Zn oxide, diet 4 contained 55 mg Zn oxide, diet 5 contained 0.3 mg sodium selenite + 40 mg Zn oxide, and diet 6 contained 0.3 mg sodium selenite + 55 mg Zn oxide. Feed intake was calculated daily, and calves were weighed weekly to calculate the feed conversion ratio and average daily gain. Blood samples were taken on days 20 and 42 to determine trace mineral levels in plasma and biochemical, enzymatic, hormonal, antioxidant, and hematological parameters. Data on growth and blood parameters were analyzed using the GLM procedure of the SAS program. Significant difference between treatments was determined by Duncan's test and the results were considered significant if the P-value was less than 0.05.

 The results showed that the use of sodium selenite and different levels of Zn oxide did not have a significant effect on feed intake, final weight, body weight gain, growth performance, and immune response (P>0.05). No significant differences in blood metabolite concentrations were observed between treatments (P>0.05). Supplementing milk with 0.3 mg sodium selenite and 40 and 55 mg Zn oxide significantly increased the activity of superoxide dismutase and decreased the levels of malondialdehyde and aspartate aminotransferase (P<0.05). Se plays a role in the defense against the accumulation of hydroperoxides from cellular metabolism. This biological function is mediated by selenoproteins such as glutathione peroxidase (GPx), iodothyronine deiodinases, and thioredoxin reductases, of which selenium is a structural component. Se also acts on enzymes involved in the production and regulation of thyroid hormones and is recognized as a factor in immunological function. The role of Zn as an antioxidant and its role in cell replication and proliferation are the two most directly related links between Zn and the immune system.

 Supplementing milk with Zn oxide and sodium selenite improved the antioxidant status and helped to relieve the stress experienced by suckling calves.

Recently, the inclusion of phytogenic feed additives in poultry for the purpose of supplementing gut health and positively impacting performance has received a great deal of attention and concern. Thyme is used in poultry nutrition in the form of herbal feed additive as it is known that its contents, such as thymol and carvacrol, have a positive impact on broiler performance and feed utilization, which in turn results in enhanced economic profits. This improvement in performance can be attributed to activation of the digestive system structure and function which causes an enhanced absorption and metabolism of the nutrient supplement and its ability to alter the gut microbiota.  Formulating diets based on ideal protein concept aim to reduce total N-excretions and ammonia emissionswhile meeting the nutritional needs of the animals. Poultry needs a specific quantity and balance of essential amino acids and nitrogen (for synthesis non-essential amino acids, NEAA) rather than crude protein. Chickens fed the standard levels of dietary protein can synthesize NEAA from excess essential amino acids. However, when low-protein diets are used, less EAA is available for NEAA synthesis. Reducing dietary protein in broiler feed is not a new subject. However, it is revealed that only few trials covered an entire production cycle with several feeding phases while majority of experiments focused on one phase only. Thyme improve protein afficiency

in this experiment, the effect two levels of Zataria multifolra (0 and 0.5 %) and two protein levels (100 (CP100) and 95 (CP95) % of Ross-308 broiler chickens’ requirement) on growth performance, physiological responses and meat quality were studied. In a completely randomized design with 2×2 factorial arrangement, 280 Ross-308 broiler chickens (one-d-old, mixed sexes) were allocated to 4 treatments, 5 replicates and 14 birds in each replicate. Growth performance, immune response, blood metabolites, blood cell count, meat quality and jejunum morphometry were considered.

Feed intake, BW and FCR of finisher and total period of production, carcass parameters except abdominal fat percentage were not affected by thyme poweder, protein levels or their interaction (P > 0.05) while inclusion of thyme powder on CP100 diet improved BW and BWG of broiler chickens during starter and grower periods (P<0.05). Inclusion of thyme powder on CP95 diet decreased serum cholesterol and LDL-cholesterol and increased HDL concentration. Feeding CP95 diet contatining 0.5 thyme poweder led to increase total protein concentration and decreased liver enzymes activity (P < 0.05). Also, feeding CP95 diet containing thyme powder decreased heterophile but increased lymphocyte count (P<0.05). Dietary treatments had no significant effects on humoral immune response, relative weight of lymphoid organs, breast and thigh meat pH and color, and jejunum morphometry.

In conclusion, apart from addition of thyme powder, 5 percent protein dilution in this experiment had no negative effects on growth performance, immune response and meat quality of broiler chickens and it maybe suggestible for broiler chickens.

Alpha-lipoic acid (α-LA) synthesized enzymatically in the mitochondrion from octanoic acid. Endogenous levels of α- lipoic acid have been reported in the micro molar range. Alpha-lipoic acid can easily pass through biological membranes due to its high lipophilicity and is converted into dihydro lipoic acid (DHLA) in cells. Alpha-lipoic acid and DHLA have been described as an ideal antioxidant couple, due to their very high protection ability against oxidative stress through multiple pathways. Alpha-lipoic acid and DHLA have metal chelating activity, acts as a coenzyme in glucose metabolism, and can produce other antioxidants such as ascorbate, vitamin E, and glutathione (GST). Alpha-lipoic acid has anti-inflammatory properties and its supplementation to broiler diets improves the expression of inflammation-related genes in the spleen. On the other hand, trivalent chromium has been recognized as an essential trace element for humans and other animals, which is part of the glucose tolerance factor (GTF). Chromium is essential for the metabolism of carbohydrates, proteins and lipids. Chromium may be present in the diet in the form of inorganic compounds or organic complexes. After absorption, chromium circulates in a free state and binds to transferrin or other plasma proteins (protein β-globulin). Trivalent chromium seems to play a role in the structure and expression of genetic information in animals. Chromium binding to nucleic acids is stronger than other metal ions. Chromium protects RNA against thermal denaturation. It is also clear that chromium is concentrated in the cell nucleus. By binding to chromatin, chromium participates in gene expression and increases the start sites and thus increases RNA synthesis. This increase is due to the induction of protein bound to the nucleus and the activation of nuclear chromatin, as a result, chromium effects gene function. Chromium supplementation has shown a positive effect on immune response of broiler chickens. The aim of this research was to investigate the simultaneous effect of dietary chromium propionate and alpha-lipoic acid supplementation on the expression of cytokines interleukin 1, 2, 4, and 10, as well as IFN-γ, TNF, and TGF β4 genes in the spleen tissue of broiler chickens.

For this purpose, an experiment was performed with 144 Ross 308 broiler chicks in complete randomized design (CRD) with a 3×2 factorial arrangement of treatments including six treatments, 3 replicates and 8 chickens in each replicate. There were 6 dietary treatments: three doses of chromium propionate (0, 750 and 1500 μg / kg) combined with two levels of alpha-lipoic acid (0 and 300 mg/kg) supplemented to the basal diets. At the end of the experimental period (42 days old), two chickens from each pen were randomly selected and killed. The sample was taken from the spleen and immediately frozen in liquid nitrogen and then stored at -80°C. The quantity and purity of the extracted RNA was checked using a Nanodrop device. In order to confirm the specific amplification of the desired genes in this study using primers, the PCR reaction was first performed. The expression of cytokine profile genes were quantified by quantitative real time PCR. In this way, 28S was used as a house-keeping gene to normalize the gene expression data. To analyze the obtained data, the method of difference in threshold degree changes (∆∆CT) was used. The method of investigating gene expression changes in this research was the 2-ΔΔCT method (comparative threshold) and the ratio of gene expression (28s).

The results of electrophoresis confirmed the expression of genes in the spleen cells of broiler chickens and they appeared as one band on 2% agarose gel. Electrophoresis of polymerase chain reaction products showed fragments of 86, 51, 82, and 88bp for interleukin 1, 2, 4, and 10, respectively. No bands were formed on agarose gel for IFN-γ, TNF and TGF β4 genes. The results showed that alpha-lipoic acid at the level of 300 mg/kg significantly decreased interleukin 1 gene expression than the control group (0 mg/kg). Also, supplementing the basal diet with the chromium propionate at the level of 1500 μg/kg significantly increased interleukin 1 and 4 genes expression compared to the control group and the chromium propionate at the level of 750 μg/kg. While the addition of alpha lipoic acid and chromium propionate had no effect on the expression of interleukin 2 and 10 genes.

Adding 300 mg/kg alpha lipoic acid to the diet can improve the immune response by reducing the expression of IL 1 gene in broilers. In addition, the addition of high amounts of chromium propionate (1500 μg) leads to a decrease in immunity (by increasing the expression of IL 1 and 4 genes). While the addition of chromium propionate up to the level of 750 μg/kg does not affect safety. Considering the results, it is recommended to add alpha lipoic acid at the level of 300 mg to the broiler diet. Addition of chromium propionate up to the level of 750 µg is fine if it has beneficial effects on yield and production characteristics.

During the winter months in most parts of Iran, the external temperature often drops below 0 °C. Typically, these temperatures fall below the optimal conditions for chick growth, resulting in cold stress and potential diseases. This can subsequently impact meat quality negatively and reduce breeding efficiency. The environmental temperature beyond the upper and lower limit of the thermoneutral zone is supposed to produce heat or cold stress in animals (Meltzer, 1983). The adverse climatic condition produces physiological stress which has profound economic influence on the productive efficiency including health and disease resistant capacity (Phuong et al., 2016). Exposure of poultry birds to extreme temperature stressor modulates the immune responsiveness and hematic-biochemical parameters of birds (Hangalapura et al., 2004). Among all the environmental stressors, cold stress induces physiological responses which are of high priority and energy demanding for homeotherms. Cold temperature can increase ascites susceptibility by increasing both metabolic oxygen requirements and pulmonary hypertension (Stolz et al., 1992). The biggest obstacle in raising broilers at high altitudes and cold conditions is the ascites syndrome. This condition can be characterized by an accumulation of fluid in the abdominal cavity and elevated mortality that tends to peak between 4-6 weeks of age (James, 2005). The International Biochar Initiative (2017) defines biochar as, “Solid material obtained from thermochemical conversion of biomass in an oxygenlimited environment.” Biochar is an ash substance that is produced from the burning of biological material via pyrolysis. This process heats the biological material in an anaerobic environment causing it to decompose into an ash form. Growth promotion and therapeutic antibiotics have been used to compensate for the high levels of stress, including cold stress,that can be present in intensive animal production. Stress can lower resistance to many of the microorganisms present in the environment; however, stress in general has been reported to have variable effects on the immune system and can both enhance and suppress responses (Siegel, 1995); cold stress has been shown to both stimulate and suppress chickens’ immune response (Regnier and Kelley, 1981; Hangalapura et al., 2006). Probiotics contain live microorganisms and spores which when administered in adequate amount, confer health benefits to the host. Bacillus subtilis (B. Subtilis) and Bacillus licheniformis (B. Licheniformis) are the two most widely used strains of probiotic bacteria in animal diets. Oral administration of B. Subtilis and B. Licheniformis can have a myriad of beneficial effects, such as improved growth and meat characteristics, optimized composition of intestinal microbiota, prevention of some diarrheal diseases, and reduced stresses. For such benefits, B. Subtilis and B. Licheniformis have attracted considerable attention as a potentially beneficial dietary supplement for animal health.  For the many negative effects of antibiotic drugs used in chick production, we want to compare the positive effects of some additives on performance of cold-stressed broiler chickens.

In this experiment, in order to examine the effect of solid waste biochar, probiotic and zeolite on improvement of performance, blood indices and small intestine morphology of broiler chickens reared under cold stress, in a completely randomized design, 315 Ross-308 broiler chickens (as hatched) were allocated to 7 treatments, 5 replicates and 9 birds in each in cold and warm houses. Treatments are including: 1) positive control (basal diet in recommended temperature), 2) negative control (basal diet + cold stress), 3-7) negative control + %0.5, %0.75 and 1% solid waste biochar, 0.02% probiotic Ecobacto-P and 1% zeolite, respectively. In cold stress groups, house temperature decreased to 17° C from 7d until end of the experiment (42d).

As compared to group reared in warm house, cold stress significantly decreased broiler chickens daily FI and BWG, serum total protein, albumin and globulin concentrations, thymus relative weight, lymphocyte percentage, breast and thigh meat pH, breast meat redness (b) and thigh meat yellowness (a) while increased total and ascitic mortality, red and white blood cell count, blood hemoglobin, heterophile percentage and heterophil to lymphocyte ratio, antibody titter against SRBC, breast and thigh meat whiteness and breast meat MDA concentration (P < 0.05).

In order to orthogonal contrast analysis between negative control group and additive groups, solid waste biochar, probiotic or zeolite couldn’t improve broiler chicken’s growth performance, breast and thigh meat quality parameters, blood metabolites concentration and cell count and did not ameliorate negative effects of cold stress.

To investigate the effects of using fennel root powder in the diet on the productive performance, immune responses, and meat quality of Arian broiler day-old chicks, an experiment was carried out in a completely randomized design with six treatments, four replicates and 20 (male and female) in each, for 42 days. The experimental groups include: 1- basic diet (control) 2- basic diet + 2 g/kg fennel root powder 3- basic diet + 4 g/kg fennel root powder 4- basic diet + 6 g/kg fennel root powder 5- basic diet + 250 mg/kg antibiotic and 6- basic diet + 200 mg/kg probiotic. The live weight , livability percentage , the production index and carcass characteristics were not affected by the experimental treatments during the whole period. In contrast, the average feed consumption in the whole period was affected by the treatments containing fennel and antibiotics (P<0.05). The antibodies against sheep red blood cells (SRBC) and immunoglobulin G were higher in the treatments using fennel root powder compared to the control treatment (P<0.05). The use of fennel root powder at all levels, reduced MDA after seven days of keeping meat in the refrigerator at 4 degrees Celsius (P<0.05). In conclution, regarding the positive effects of using fennel root powder on the immune responses and meat preservation of broilers, it is recommended to use it at the rate of 2 grams per kilogram of diets.

The use of medicinal plants and their derivatives can stimulate feed consumption, increase daily weight, feed conversion ratio, increase shelf life, improve the health and function of the digestive system. It seems that the use of an optimal mixture of several medicinal plants in the diet has positive effects on performance and carcass quality of broiler chickens in comparison to each one. Nowadays, with the popularization of ready-made feed in raising broiler chickens, many breeders tend to add food additives in drinking water. Therefore, it is a question that adding these compounds in feed or drinking water makes a difference. Three commercial plant additives that are used today are Coxan, O.X. Plant, and Entex. Coxan contains oregano (with the active ingredient of menthol) and garlic (with the active ingredients of allin and allicin), O.X. Plant contains savory (with the active ingredients of carvacrol and thymol), thyme (with the active ingredients of thymol and carvacrol) and red pepper oleoresin (with the active ingredient of capsaicin), Entex contains cinnamon (with the active ingredient of cinnamaldehyde), and garlic and eucalyptus (with the active ingredient of cineol). The aim of this research was to compare the effects of using these commercial plant additives (in water and feed) on growth performance, intestinal microflora and morphology and immune response of broilers.
A total of 336 Ross 308 broilers were examined in a completely randomized design with seven treatments, four replications, and 12 chickens per replication. Experimental treatments included: control (without phytogenic in feed or water), Coxan in feed (300 mg/kg) and in water (200 mL/1000 L), O.X. Plant in feed (200 mg/kg) and in water (135 mL/1000 L) and Entex in feed (500 mg/kg) and water (350 mL/1000 L). Daily weight gain, daily feed intake, daily water intake and feed conversion ratio were measured. At 42 days of age, two birds were selected from each experimental unit and after slaughter, the length of the intestinal components was measured separately (duodenum, jejunum, and ileum). To determine the microbial population, samples were taken from the ileum of chickens. EMB culture medium was used to determine Escherichia coli population and MRS culture medium was used for Lactobacillus bacteria. To check the humoral immune response, 0.1 mL of 25% sheep red blood cell solution in PBS was injected into the breast muscle of chickens on the 12th and 29th days of rearing. Blood was taken from the chickens on the 28th, 35th and 42nd days of breeding and the levels of Anti-SRBC, and immunoglobulins G and M were calculated. Antibody titer against Newcastle disease was measured by HI method.
The effect of additives in water and feed on average daily feed consumption, daily weight gain, feed conversion ratio and daily water consumption was not significant in the whole period. Since the amount of water consumed by the chickens did not change, it can be concluded that the additives added to the water in the examined amounts do not have a spicy or unpleasant taste that would cause the birds to refuse to drink water. The relative length of duodenum, jejunum, cecum and colon was not affected by experimental treatments. However, the relative length of the ileum was lower in the chickens that received Entex in the feed compared to the control group. No significant difference was observed in the relative length of different parts of the intestine in the treatments that received herbal additives either in feed or water. The relative length of duodenum, jejunum, cecum, and colon, villus length, villus area, crypt depth, thickness of lamina propria, thickness of muscular layer, and thickness of advantis layer were not affected by experimental treatments. The chickens that consumed 500 mg/kg of Entex herbal additive of feed had a lower villus width in the ileum region compared to the control group chickens. The population of Escherichia coli bacteria in the ileum of chickens that had received the addition of O.X. Plant and Entex in the feed decreased compared to the control group. The population of Escherichia coli bacteria in the Entex treatment in the feed was also reduced compared to the Coxan treatment in the feed. The population of Lactobacillus bacteria in the ileum of chickens that received Coxan in water, O.X. Plant in feed and water, and Entex in feed increased compared to the control group. There was no significant difference in the population of Escherichia coli and Lactobacillus bacteria in the treatments that received herbal additives either in feed or in water. The use of plant essential oils in poultry feed, while improving the microbial population by increasing the number of lactobacilli, by improving the morphological characteristics of the intestine, probably improves the ability of digestion and absorption in the digestive system and improves the growth efficiency of broiler chickens. Antibody levels against SRBC were higher at 28, 35, and 42 days in all chickens consuming herbal additives in water. The performance of animals is significantly influenced by the state of health and safety of the animal. A weak or stressed immune system causes weight loss when dealing with infectious diseases, so the use of immune system stimulating substances can increase performance by improving the immune status. In raising poultry, it is important to strengthen the immune system to prevent the occurrence of diseases.
The results of this research showed that the commercial plant additives of Coxan, O.X. Plant, and Entex had no significant effect on growth performance, relative length and intestinal morphology. The population of opportunistic bacteria Escherichia coli was significantly reduced compared to the control group with the addition of O.X. Plant and Entex in the diet. In general, it can be concluded that adding tested herbal additives not only did not have a negative effect on the drinking water consumption of broiler chickens, but also using them in water improved the immune responses of broilers more than adding them in feed. A total of 336 Ross 308 broilers were examined in a completely randomized design with 7 treatments, 4 replications, and 12 chickens per replication. Experimental treatments included: control (without phytogenic in feed or water), Coxan in feed (300 mg/kg) and in water (200 mL/1000 L), O.X. Plant in feed (200 mg/kg) and in water (135 mL/1000 L) and Entex in feed (500 mg/kg) and water (350 mL/1000 L). Daily weight gain, daily feed intake, daily water intake and feed conversion ratio were measured. At 42 days of age, 2 birds were selected from each experimental unit and after slaughter, the length of the intestinal components was measured separately (duodenum, jejunum, ileum). In order to determine the microbial population, samples were taken from the ileum of chickens. EMB culture medium was used to determine Escherichia coli population and MRS culture medium was used for Lactobacillus bacteria. To check the humoral immune response, 0.1 ml of 25% sheep red blood cell solution in PBS was injected into the breast muscle of chickens on the 12th and 29th days of rearing. Blood was taken from the chickens on the 28th, 35th and 42nd days of breeding and the levels of Anti-SRBC, immunoglobulin G and M were calculated. Antibody titer against Newcastle disease was measured by HI method. The effect of additives in water and feed on average daily feed consumption, daily weight gain, feed conversion ratio and daily water consumption was not significant in the whole period. Since the amount of water consumed by the chickens did not change, it can be concluded that the additives added to the water in the examined amounts do not have a spicy or unpleasant taste that would cause the birds to refuse to drink water. The relative length of duodenum, jejunum, cecum and colon was not affected by experimental treatments. However, the relative length of the ileum was lower in the chickens that received Entex in the feed compared to the control group. No significant difference was observed in the relative length of different parts of the intestine in the treatments that received herbal additives either in feed or water. The relative length of duodenum, jejunum, cecum, and colon, villus length, villus area, crypt depth, thickness of lamina propria, thickness of muscular layer, thickness of advantis layer were not affected by the experimental treatments. The chickens that consumed 500 mg/kg of Entex herbal additive of feed had a lower villus width in the ileum region compared to the control group chickens. The population of Escherichia coli bacteria in the ileum of chickens that had received the addition of O.X. Plant and Entex in the feed decreased compared to the control group. The population of Escherichia coli bacteria in the Entex treatment in the feed was also reduced compared to the Coxan treatment in the feed. The population of Lactobacillus bacteria in the ileum of chickens that received Coxan in water, O.X. Plant in feed and water and Entex in feed increased compared to the control group. There was no significant difference in the population of Escherichia coli and Lactobacillus bacteria in the treatments that received herbal additives either in feed or in water. The use of plant essential oils in poultry feed, while improving the microbial population by increasing the number of lactobacilli, by improving the morphological characteristics of the intestine, probably improves the ability of digestion and absorption in the digestive system and improves the growth efficiency of broiler chickens. Antibody levels against SRBC were higher at 28, 35 and 42 days in all chickens consuming herbal additives in water. The performance of animals is significantly influenced by the state of health and safety of the animal. A weak or stressed immune system causes weight loss when dealing with infectious diseases, so the use of immune system stimulating substances can increase performance by improving the immune status. In raising poultry, it is important to strengthen the immune system to prevent the occurrence of diseases. The results of this research showed that the commercial plant additives Coxan, O.X. Plant and Entex had no significant effect on growth performance, relative length and intestinal morphology. The population of opportunistic bacteria Escherichia coli was significantly reduced compared to the control group with the addition of O.X. Plant and Entex in the diet. In general, it can be concluded that adding tested herbal additives not only does not have a negative effect on the drinking water consumption of broiler chickens, but using them in water improves the immune responses of broilers more than adding them in feed.

In the past, vitamins were considered as unknown growth factors, but during the twentieth century, their structure and nature were gradually discovered. These compounds are necessary to maintain the integrity of the tissues and general health of the body. Broiler chickens are unable to synthesize vitamins (except vitamin C) or have the ability to make limited amounts of them (group B and K2). For this reason, vitamins should be added to broiler feed as a supplement. Vitamins interact with other nutrients in various ways. The main reference for vitamin requirements in broilers is the National Research Council (NRC, 1994). The recommended amounts ensure conditions where there are no severe deficiencies. Applying the NRC recommendations cannot guarantee the genetic potential of today's birds. Because these values were taken from old studies and using pure feeds in laboratory conditions. Due to the dependence of Iran's poultry industry on the import of vitamins, unfortunately, in recent years, few studies have been conducted in the country on the appropriate pattern of adding vitamins to the diet of broiler chickens. Therefore, it is necessary to examine the lower levels than the recommendation of the Ross 308 strain catalog (Aviagen, 2019) as the dominant strain of the country and to propose an optimal model for different regions that does not have a negative effect on the performance and safety traits and to reduce the need to import this strain. It will also follow the products.

The aim of this study was to determine the optimal pattern of vitamin supplementation in diets based on corn, wheat waste and soybean meal on production traits, carcass characteristics and immune responses of broilers from 1 to 42 days of age in Fars province. A total of 500 Ross 308 broilers with similar mean weight (mixed sex) were distributed in a completely randomized design with five treatments, five replications and 20 chickens per replication. Experimental treatments included five levels of vitamin supplementation: 1) control (100% of the recommended values of Ross 308, 2019 strain catalog), 2) 90% of the recommended values of Ross 308, 2019 strain catalog, 3) 80% of the recommended values of Ross 308, 2019 strain catalog, 4) 70% of the recommended values of Ross 308, 2019 strain catalog and 5) 60% of the recommended values of Ross 308, 2019 strain catalog. At the end of each period, the feed consumption and body weight of the birds were recorded with a digital scale with an accuracy of ±0.01. At the age of 39 days, blood was drawn from the vein under the wing of two birds from each experimental unit, with a syringe containing EDTA anticoagulant, and the blood sample was quickly transferred to the laboratory in a flask containing ice for differential counting of white blood cells. At the end of the 42nd day of rearing, two birds from each experimental unit were slaughtered to measure the relative weight percentage of carcass components. To measure the humoral immune response, the injection of 0.5 ml sheep antigen (SRBC) at the age of 21 days was used. CBH skin hypersensitivity test was also used to measure cellular immune response.

The highest and lowest mean body weight gain and feed intake belonged to the vitamin pattern of 100 and 60% of the recommendation of Ross 308 strain, respectively (P<0.01). The best feed conversion ratio and production index belonged to the vitamin pattern up to 90% of the Ross strain recommendation (P<0.01). The highest and lowest percentages of pectoralis muscle (22.2% and 19.2%) were related to the pattern of 100 and 60% of vitamin supplementation recommended by the Ross strain (P<0.01). The effect of reducing vitamin supplementation on cutaneous basophil hypersensitivity (CBH), serum antibody level against SRBC and percentage of heterophils, lymphocytes and heterophil to lymphocyte ratio was not significant.

There was no significant difference between the functional indices and most importantly the production index of treatment 1 (100% vitamin pattern) and treatment 2 (90% vitamin pattern). Carcass traits and indicators related to humoral and cellular immune systems did not show significant differences too. As a result, a 10% reduction in the level of vitamin supplementation compared to the recommended level of the Ross 308 catalog is possible and suggested for raising broiler chickens. The final result is that in the breeding conditions of Fars province poultry farms, it is possible to reduce the level of vitamin supplements in diets containing wheat waste up to 90% of the recommendation of Ross 308 strain compared to the control diet.

The use of antibiotics in poultry nutrition, in addition to their importance in preventing diseases, causes an increase in congenital anomalies, chronic diseases, increased microbial resistance and hundreds of other small and large complications, which is attributed to the current health problems of human society. The focus has always been on animal welfare, environmental care, limited use of medicines and the production of a healthy product without chemical residues that do not endanger human health. One of the suitable alternatives to antibiotics is bioactive peptides.

This experiment was used in a completely randomized design with 240 one-day old male commercial Ross 308 broilers in six treatments, five replications (eight chickens per replication). Experimental treatments include: basal diet, basal diet + 0.1% of vitamin E and 0.05% of maduramicin, basal diet + 0.05% of maduramicin, basal diet + 200 mg / kg superworm peptide, basal diet +400 mg / kg were superworm peptide and the basal diet was + 600 mg / kg superworm peptide. The effect of treatments on performance, microbial population, intestinal morphology and immune system response of broilers was investigated.

Addition of maduramicin (treatment 3) and superworm peptide at the levels of 200, 400 and 600 mg to the diet compared to the control group was able to reduce feed intake in the final period. On the other hand, treatments that used antibiotics, compared to treatment 5, which contained 400 mg of superworm peptide, were able to reduce the total bacterial population. Adding all levels of bioactive superworm peptide to the diet reduced the crypt diameter compared to the control group and also the diet containing 400 mg of the superworm peptide reduced the crypt depth compared to the control group. The results obtained in relation to immune factors can be stated that by adding antibiotics to the diet, it can increase immune factors such as immunoglobulin G at the age of 35 days compared to the control group, as well as the superworm peptide. At a concentration of 400 and 600 mg per diet, it was able to improve the total SRBC titer at 42 days of age compared to the control group.

The results of this experiment showed that the experimental treatments were able to improve the performance indices at the age of 11 to 42 days and some morphological features of the jejunum such as: crypt depth and diameter. Also, it can be said that the effect of experimental treatments on the total bacterial and lactobacilli population and the level of immunoglobulin G (day 35) and total SRBC titer (day 42) was significant.

This study was conducted to investigate the effects of sour tea powder on growth performance and immune responses of broiler chicks under conditions challenge with ascites. 400 broiler chicks were studied in arrangement of 4 experimental diets including: Treatment 1: Positive control group (without induction of ascites and without powder), treatment 2: Negative control group (induction of ascites without powder), Treatments 3 and 4) 1% and 2% sour tea powder groups with ascites induction, and inclusion of 1% and 2% tea powder. This study was conducted for 42 days and the induction of ascite was started from day 8. This study was conducted in form of a completely randomized design with 4 treatments and 5 replications. Performance parameters (body weight, feed intake and feed conversion ratio), immune responses against SRBC, weight of immune organs (spleen, thymus and bursa) and heart indexes were investigated. The results showed that the induction of ascites decreased body weight and increased feed conversion ratio in finisher period. The induction of ascites also decreased relative weight of heart indexes, immune organs and antibody titer (p<0.05). Dietary inclusion of 2% plant powder increased growth performance in finisher and total periods. It also improved relative weight of immune organs and antibody titer against SRBC. In conclusion, adding 2% Sour tea powder into diet is suggested to improve immune responses and growth performance under ascites condition.

In the recent years, many countries have banned or restricted the usage of antibiotics as growth-promoting, due to their undesirable effects on human and animal health. Using probiotics as an alternative to growth-promoting antibiotics play an important role in improving performance, health and increasing the quality of poultry products. Lactobacilli are the most popular probiotic bacteria that have beneficial effects on host’s performance and health. In this study, the effect of two native strains of Lactobacillus ruteri isolated from the gastrointestinal tract of native chickens in north-western and south-western of Iran were investigated on performance, carcass characteristics, blood parameters and immune response of broilers.

In this experiment, 300 Arbor-Acres® Plus+ male broiler chickens were divided in to 5 treatments, 4 replications (15 chickens per replication), in a completely randomized design. The experimental treatments include: 1- Basic diet as a control treatment, 2- Basic diet + Avilamycin antibiotic, 3- Basic diet + commercial probiotic (Bio-Poul®), 4- Basic diet + Lactobacillus reuteri isolate MG547731 (Plr2), and 5- Basic diet + Lactobacillus reuteri isolate MG547727 (Plr3).

Under the condition of this experiment, commercial probiotics, antibiotic and either Lactobacillus reuteri (Plr2) isolates, significantly improved daily weight gain in final period (p<0.05). Native isolates probiotics and commercial probiotics significantly (p<0.05) reduced the relative weight of gizzard and proventriculus, though no significant (p<0.05) differences were recorded for other carcass components in broiler chickens. After the first injection of SRBC, treatments include antibiotic and Lactobacillus reuteri (Plr2) isolate caused a significant increase of IgM, and after the second injection, Lactobacillus reuteri (Plr2) and commercial probiotic treatments, showed a significant (p<0.05) increase in immunoglobulin G and Total immunoglobulin. The antibody titer against Newcastle vaccine did not show a significant difference between treatments and the control group. There was no significant difference between serum glucose, cholesterol, triglyceride, VLDL and LDL between treatments and the control group. No significant difference was obtained in serum HDL level in probiotic treatments, while antibiotic treatment appeared with a significant decrease in HDL.

Addition of native Lactobacillus reuteri isolates to poultry diets can have beneficial effects similar to commercial probiotics, on performance, immune response and blood parameters of broilers.

The effect of chick initial body weight and dietary nutrients density on growth performance and immune response were investigated using 360 Ross 308 one-day-old straight run chicks (mixed sex) in a completely randomized design with 2×3 factorial arrangement of treatments, four replicates and 15 birds per replicate. The initial body weight of chicks was grouped as 42±1 g (standard weight); 38±1 g (light weight; 10% under standard weight); 46±1 g (heavy weight; 10% over standard weight) while the nutrients density of the diet was either 5% lower than the nutritional recommendation of Ross 308 (typical commercial diet in Iran), or as recommended by Ross 308 manual (high-density diet). Chickens fed the high-density diet had higher daily weight gain and lower conversion ratio than chickens fed the typical diets (P<0.05). The effect of dietary density and initial body weight of chicks on carcass components and weight of internal organs were not significant except for heart. Broilers fed the high-density diet and heavy weight had higher antibody titers to sheep red blood cell (P<0.05). Light chicks fed the high-density diets had greater total anti sheep red blood cell and immunoglobulin G titers than light chicks fed the typical diets (P<0.05). Results indicate that feeding light chicks with high-density diets increases their growth rate compared to the chicks with standard or heavy weights. Moreover, feeding diets with a high density of nutrients to light chicks can improve immune responses.

This research was carried out to evaluate the effect of sunflower seed meal fermented with Aspergillus niger and Saccharomyces cerevisiae on performance, nutrient digestibility, immune response, some blood parameters in 200 male Ross 308 broiler chicks in a completely randomized design. with five treatments. Experimental treatments were: 1- Diet containing sunflower seed meal fermented by Aspergillus niger 2- Diet containing sunflower seed meal fermented by Saccharomyces cerevisiae 3- Diet containing sunflower seed meal fermented by both Aspergillus niger and Aspergillus niger 4- Diet containing sunflower seed meal without processing 5-Control diet based on corn and soybean meal. Feed intake in control and treatment 1 were higher than the other treatments in starter and finisher periods of the experiment (P<0.05). Daily Feed intake in control group was higher than the other treatments in whole period of the experiment (P<0.05). Daily weight gain in control and treatment 1 were higher than the other group in starter and grower periods of the experiment (P<0.05). Feed conversion ratio in treatment 1 was better in finisher phase. In whole period of the experiment, feed conversion ratio in control and treatment 1 was better than the other treatments (P<0.05). Dry matter, crude protein, crude fat digestibility in the control group was higher than the other treatments (P<0.05). In conclusion, inclusion of sunflower seed meal fermented with Aspergillus niger, in broiler diet showed similar feed conversion ratio in compared to control group in whole period of the experiment and also lowered abdominal fat percentage.

Stress, as a response to adverse stimuli, is difficult to define and understand mostly due to its nebulous perception. As Selye (1976) pointed out, “stress is the nonspecific response of the body to meet any demand”, whereas stressor is defined as “an agent that produces stress at any time”. Hence, stress represents the reaction of the animal organism (i.e., a biological response) to stimuli that disturb its normal physiological equilibrium or homeostasis. Thanks to their antioxidants, nutrients and anti-inflammatory properties silver nanoparticles are used in many developed countries (Bhanja et al 2015). The list potential applications of nanotechnology are very wide and diverse, but it is undoubtedly one of its most valuable applications in the development of therapeutic and pharmaceutical cases. Also, the use of silver nanoparticles in the treatment of deadly diseases like avian influenza supports further research on biological systems (Xiang et al 2011). Among inorganic materials, metal oxides such as TiO2, ZnO, MgO CaO, and Ag are of particular interest because they are stable in hard conditions and generally considered to be safe substances for humans and animals.

A total number of 592 eggs with an average weight of 50 g were purchased from a commercial Hubbard F15 broiler breeder flock aged 50 wk. The eggs, allotted to 4 treatments of 4 replicates with 37 eggs each, were set on the same floor to provide similar incubation conditions. Treatments including 2 doses of silver nanoparticles (20 and 40 mg) was injected via the egg holes using 1 mL insulin syringes equipped with disposable needles (21 gauge). The control group did not receive any treatment, and the sham control was injected with 1 ml phosphate-buffered saline (Triplett et al 2018). The injection holes were immediately covered using paraffin. At the end of d 18 of incubation, the eggs were transferred to the hatching cabinet. On the 21d of incubation, the hatched chicks were taken out of the incubator and after counting (checking for hatching) and weighting, half of the chicks were killed on the same day to check Hemoral immunity (G and M),white blood cell count and study of the desired genes (TNF-α, IL-6 and TGF-β). Among other chickens, those that were more uniform in weight with the desired treatment were transferred to the breeding period for 42 day and again in 4 treatments (treatments applied during incubation) and 4 replications with 20 broilers per replication. During 42 day, broilers were provided with unlimited water and food. To induce oxidative stress during the breeding period, 500 μg/kg live weight Salmonella lipopolysaccharide injected at 12, 24 and 48 hours before killed (Xi et al 2000).

While ovo injection, silver nanoparticles was significantly decreased hatchability in comparison to the control group (P<0/05). In line with our results (Goel et al 2017) found that the hatchability of the eggs injected with nanoparticles was significantly lower than the control group. The reason can be attributed to the smaller size of younger embryos at the 7th ED, injection method, location injection, injection depth, injection time, genetics, hen age, egg size and hatching conditions (Sun et al 2018; Pilarski et al 2005). In case of ovo injection of 20 and 40 mg silver nanoparticles, significantly increased carcass percentage compared to the control group at hatching and post-hatch period, respectively (P<0/05). (Bhanja et al 2015; Pineda et al 2012) proved that silver nanoparticles had positive effects on embryo weight. Due to the significant amount of antioxidant in the silver nanoparticles inside the egg and its antioxidant effect on energy efficiency during embryonic life, silver nanoparticles receiving groups had a higher body weight at hatching time. Higher birth weight makes it possible to increase feed intake and weight gain in these treatments.Upon ovo administration of 20 mg silver nanoparticles, spleen was increased significantly compared to the control group at hatching period (P<0/05). This is in line with a previous study of Goal et al (2017), who observed higher liver and spleen weights in 40 mg/kg silver nanoparticles group. The development of B and T lymphocytes initiates during embryogenesis in the bursa of Fabricius and thymus, respectively, and matures in the spleen until post-hatch (Erf 1997). These organs play an important role in imparting immunity. The cells produced in these organs differentiate into cellular immunity and humoral immunity, thus imparting immunity against different pathogens. Therefore, increased liver and spleen weight indicates a better immunological health status of in ovo silver nanoparticles supplemented birds. Ovo treatments did not affect the concentrations of immunoglobulin (IgG), (IgM), WBC counts and H/L ratio (P>0/05). Our results are consistent with previous studies of Pineda et al (2012) who showed the concentration of IgG and IgM were not affected by ovo injection of 10 and 20mg/kg silver nanoparticles. (Salari et al 2016; Saki and Salari 2015) showed silver nanoparticles increased in serum IgM and IgE, and increased in blood neutrophilic granulocytes. Rezaei Zarchi et al (2012) reported, feeding the rats for 28 days at doses of 25, 50, 100 and 200 mg / kg of silver nanoparticles had no significant effect on WBC and H/L. Differences in results may be due to differences in injection method, location injection, injection depth, injection time, genetics, hen age, egg size and hatching conditions. In comparison with the control and 40 mg silver nanoparticles group at hatching, there was significant up-regulation of TNF-α, IL-6 and TGF-β gene expression in 20 mg silver nanoparticles injected embryos,. Tumor necrosis factor-α (TNF-α) is a key cytokine involved in inflammation and immunity and the pro-inflammatory cytokine IL-6 induces the final maturation of B cells into antibody-secreting plasma cells, thereby increasing the secretion of immunoglobulins (Balkwill 2009; Mosmann 1989). Silver nanoparticles can interact with the immune system by binding and reacting with cells or proteins, thereby modulating the immune response. In the present study, higher expression of TNF-α gene in the livers was observed in ovo injected silver nanoparticles embryos. These findings also support earlier studies by Khan et al (2013), who showed the gene expression of IL-6 and TNF-𝛼 were affected by 50nm GNPs in the kidneys of rats. Also Vadalasetty et al (2018) reported that expression of TNF-α and NF-kB at mRNA were significantly up-regulated in the 50ppm silver nanoparticles group.

The results of this study suggest that silver nanoparticles improve the immune response of broilers by improving growth and increasing the relative expression of genes involved in immune function.

Increasing the growth rate, shortening the breeding period and improving feed efficiency in broiler chickens has provided the basis for the occurrence of a number of metabolic disorders such as ascites syndrome. This experiment was carried out to investigate the effect of medicinal plants (ethanolic extracts of Olive leaf, Garlic, Roselle and a combination of them) on performance, blood parameters and immune response of broilers under ascites induction conditions.

A number of 360 male broiler chickens of Hubbard strain were used in a completely randomized design with six treatments and four replications and 15 chickens per each replication. Experimental treatments include 1) control diet, 2) diet containing 500 mg/kg vitamin C, 3) diet containing 500 mg/kg Olive leaf powder extract, 4) diet containing 500 mg/kg Garlic extract, 5) diet containing 500 mg/kg Roselle extract. 6) Diets contained a combination of three plant extracts. To induce ascites, all chickens were exposed to 15 °C cold stress from 15 to 42 days of age. During the breeding period, performance traits (body weight, feed intake) were recorded weekly. To measure blood biochemical parameters and thyroid hormones, at the end of the experiment, 2 blood samples were prepared from the chickens of each replication. To measure The Cutaneous Basophil Hypersensitivity response (CBH), Phytohemagglutinin-P (PHA-P) were injected intradermally into the wattle.

The results showed that the experimental treatments had no significant effect on body weight and feed consumption, but the treatments of Garlic extract, Roselle extract and vitamin C improved the feed conversion ratio (p<0.05). Visceral organs were not affected by the experimental treatments, but the ratio of the right ventricle to the whole ventricle increased as a result of cold stress (p<0.05) and the use of antioxidant compounds caused its relative improvement. Blood parameters of glucose, cholesterol and aspartate aminotransferase enzyme were not affected by experimental treatments. As a result of cold stress, blood albumin and triglyceride concentration decreased and blood uric acid concentration increased (p<0.05). The use of antioxidant compounds, especially vitamin C, decreased the lipid peroxidation index and increased the cutaneous basophilic sensitivity response (p<0.05). The use of the combination of three plant extracts reduced the serum concentration of T3 hormone (p<0.05).

The use of vitamin C and a combination of three extracts of Olive leaf powder, Garlic, and Roselle improves performance, blood and immune parameters under ascites induction conditions.

 This experiment was performed to evaluate the effects of peppermint extract, vitamins C, vitamin E, probiotics and antibiotics on performance, biochemical parameters and immune system of broilers under heat stress conditions. Peppermint (Mentha piperita) is considered as one of these medical plants and belongs to the Lamiaceae family. The essential oils and extracts of this plant are mainly made up of menthone, menthol and methyl acetate. Peppermint is traditionally used as an antiseptic, antispasmodic, mild tonic, antimicrobial.

 A total of 240 one-day-old broilers (Ross 308) were distributed to 8 treatments with 5 replications/treatment based on a randomized block design. Experimental diets consisted of base diet without feed additive and under standard temperature conditions (negative control), base diet without feed additive and under heat stress conditions (positive control), positive control supplemented with 28 mg/kg virginiamycin, 28 mg/kg probiotic protexin, 1 g/kg vitamin C, 1 g/kg vitamin E, 250 and 500 mg/kg peppermint extract, respectively. House temperature was initially set at 28°C for the second week and then reduced by 0.5°C per day until a temperature of 22°C was achieved at the end of the fourth week and then maintained constant thereafter. To induce heat stress, room temperature was raised to 34◦C during 10 AM to 16 PM from 15 to 42 days of age. A 23:1h light to darkness lightening regimen was followed throughout the experimentation period. Body weight gain and feed intake were recorded for days 10, 24 and 42 of age and data were used to calculate feed conversion ratio (FCR). At the end of the experiment (day 42), two birds from each replication with a body weight close to cage mean were selected and killed. Individual blood samples were collected form the slaughtered birds and centrifuged at 1800× g for 15 min. The collected sera samples stored at -20°C pending biochemical assessments. Concentrations of serum glucose, triglyceride, total cholesterol and low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), very- low-density lipoprotein cholesterol (VLDL-C), white blood cells (WBC), red blood cells (RBC), Hematocrit and Hemoglobin were determined by different recommended procedures. To assay the primary and secondary antibody responses against SRBC, 2 birds/replicate were immunized intramuscularly with 0.5 mL 10% SRBC in. Blood samples (1.5 mL/bird) were obtained from the brachial vein at 7d following each injection.

 The results showed that the highest feed intake and feed conversion ratio were observed in chickens fed with negative control diets (P <0.05). The greatest body weight was observed in the birds fed with the negative control and diets supplemented with probiotics and vitamin E (P <0.05). Broilers fed with diets containing antibiotics as same as positive control group could increase abdominal fat percentage and serum triglycerides concentration than other treatments (P <0.05). The broilers fed with diets supplemented with all feed additives decreased the heterophils to lymphocytes ratio compared to the positive control group (P <0.05). The probiotics and 500 mg peppermint extract-supplemented diets improved the antibody titer against Newcastle, Gumburo and influenza diseases, as well as anti-SRBC titer, compared to the positive control group (P <0.05). The results of this study are supported by other studies, indicating that the dietary supplementation of probiotic could improve physiological responses and immune system and thus performance of heat-stressed birds. Dietary supplementation of vitamin C is an effective strategy to reduce the harmful effects of heat stress in poultry. The results of previous studies have demonstrated that the antioxidant and inflammatory properties of peppermint contributes to the prevention and treatment of diseases associated with oxidative stress, through removing free radical. Medicinal plants with secondary metabolites have also reported to possess the positive effects on growth performance and thus increasing the immune function. The positive effects of peppermint could be due to its active ingredients such as carvacrol, flavonoids and menthol. Flavonoids with antioxidant effects have protective properties against free radicals. The mucous membrane of the gastrointestinal tract plays an important role in preventing antigens and harmful microorganisms from entering the organ and eliminating them, while also being effective in selective nutrient absorption. The results of the present experiment and the other reports also indicate that dietary supplementation of probiotic and vitamin E and C and peppermint extract improve the immune response and growth performance of broilers and could be account as an alternative to antibiotics.

 In conclusion, a high dose of peppermint (500 mg/kg) was as effective as probiotic, vitamin E and C in alleviating the negative effects of heat stress on growth performance, health and immune function.

Organic acids are one of the alternatives to antibiotics growth promotor which have beneficial effects on animal performance and gastrointestinal microbial population. In general, addition of organic acids into the feed lowers pH of the feed and gastrointestinal tract, stimulates growth, helps beneficial bacteria to overcome pathogenic bacteria, and reduces toxic metabolites produced by harmful bacteria.

A total of 240 day-old Ross 308 male broiler chicks were divided into 4 treatments, 4 replicates, and 15 broiler chickens per replicate in a completely randomized design. Experimental treatments were included 0, 200, 300 and 500 mL of acidifier in 1000 L drinking water.

Feed conversion ratio in broilers that received two levels of 200 and 500 mL of acidifier per 1000 liters in drinking water decreased compared to the control treatment (p<0.01). European efficiency index was higher in the whole acidifier levels than the control and a linear increasing trend was observed (p<0.05). Broilers received 500 mL of acidifier in water had the lowest FCR (1.64 vs. 1.80; p<0.01) and the highest EEF (394 vs. 332; p<0.01) compared with the control. The weight of bursa of Fabricius, spleen and pancreas in broilers received graded levels of acidifier was higher than the control (p<0.05). The pH of the proventriculus, duodenum and jejunum of broilers that received 500 mL of acidifier decreased compared to the control (p<0.05). Administration of graded levels of acidifier in drinking water had no effect on Lactobacillus, coliforms and Escherichia coli count in ileum (p>0.05). The thickness of the lamina propria layer increased in broilers received 300 mL of acidifier (p<0.05). Administration of acidifier in drinking water had no effect on antibody titer against SRBC and Newcastle.

The results indicate that adding 500 mL of acidifier in 1000 L drinking water improves growth performance by increasing the weight of digestive organs such as pancreas and acidifying the initial areas of the digestive tract.