Effects of feeding Isomaltooligosaccharides on the growth performance, carcass traits and immune response of broiler chickens

The research was conducted to study the effect of feeding broiler chickens on diets containing isomaltooligosaccharides on the growth performance, carcass traits and immune response. 90-one day old broiler chicks were used according to completely randomized two treatment groups and one control, 30 birds each. Birds fed ad-libitum on basal starter and grower-finisher diets for 35 day. Diets of treatment`s groups contained 0.5 g/Kg and 1 g/Kg of Isomaltooligosaccharides, while the control group fed on the basal diets without Isomaltooligosaccharides supplementation. Dietary supplementation of broiler chickens with Isomaltooligosaccharides improved body weight, feed conversion, carcass traits, two lymphoid organs weight and log antibody titer against avian flu vaccine. Most of the highest values were for birds fed low levels of Isomaltooligosaccharides. Feed intake decreases as Isomaltooligosaccharides level increases. Dietary supplementation with Isomaltooligosaccharides did not affect the lipids profile (triglycerides, total cholesterol, LDL and HDL), however the blood VLDL levels decreased with increased levels of Malondialdehyde and Glutathione reductase. Collectively, Dietary supplementation of broiler chickens with 0.5 g/Kg diet of Isomaltooligosaccharides improved growth performance, carcass traits and immune status.


Introduction
Oligosaccharides are composed of between two and nine monosaccharides linked through glycosidic bonds. Nondigestible oligosaccharides are oligosaccharides that are not hydrolyzed by digestive enzymes in the gastrointestinal tract. Isomaltooligosaccharides (IMO) are produced from glucose by enzymatic transgalactosylation [Hayashi et al., 1994;Vetere et al., 2000]. The mixture contains isomaltose (O-α-D-glucopyranosyl-(1-6)-D-glucopyranose), panose, isomaltotriose, and several other branched oligosaccharides composed of four or five glucose residues. More importantly, IMO have been used as a prebiotic similar to other nondigestible oligosaccharides [Crittenden, 1999;Zhang, 2000;Chen et al., 2001]. IMO was tested as prebiotic for poultry and stimulated the growth of Bifidobacterium and Lactobacillus and are not used by Salmonella or Escherichia coli [Chung and Day, 2004].This wasn't the aim of this paper. However, this role has been a matter of debate because prebiotics are a non-digestible feed ingredient that may benefit the host by selectively stimulating the growth or activity of one or a limited number of bacteria in the colon, resulting in improved host health and performance [Gibson and Roberfroid, 1995]. Although, IMO can be cleaved at the brush border of small intestinal mucosal cells [Hunziker et al., 1986;Najjar et al., 1991;Gu¨ nther and Heymann, 1998;Hertel et al., 2000], It enhance the growth performance of broilers [Chung and Day, 2004], potentially activate the immune system, thereby enhancing the resistance to diseases [Li et al., 2009]. Because of a very few investigations of IMO have been undertaken on broilers to date, the aim of this work was to study the influence of dietary supplementation of broilers with IMO on the growth performance, carcass traits and immune response.

Materials and Methods:
The experiment was carried out under the protocol approved by the Faculty of Veterinary Medicine, Department of Nutrition and Clinical Nutrition, Sadat City University Egypt. A total of 90 one day old chicken "cobb500" were randomly allotted to 3 floor pens 30 birds each. The pens were assigned to control and two treatment`s groups. The average body weights of the birds in groups were similar. Birds were raised to 35 days and fed corn-soya bean based diet (1-15 day of age) and grower-finisher diet (15-35 day of age) which formulated to guarantee the strain`s requirements or exceed recommended nutrient requirement [NRC, 1994]. One basal diet was mixed for each dietary period and the additives were blended in according to treatment distribution. The basal diets composition for all experimental phases is listed in Table 1. Basal diets (starter and grower-Finisher) were used without IMO supplementation for the control group. Supplemental IMO 93% was substituted for corn in the basal diets (starter and grower-Finisher) at inclusion rate of 0.5 g/kg and 1 g/kg for the two treatment`s groups (T1 and T2), respectively. The IMO product contained the following sugars: glucose (0.27%), maltose (7.4%), isomaltose (24.5%), panose (7.39%), and isomaltotriose (16.07%), and the remainder consisted of maltotriose and other oligosaccharides. Diets were antibiotic free and were provided in mash form.

Management and housing
Top-dressed litter with 2 inch of fresh pine wood shavings was used as bedding. The temperature was set at 30 to 33°C during the first week and was reduced by 2 °C per week until 23 °C was reached. Relative humidity was about 60 to 80%. The lighting program was 23L:1D. The chicks were vaccinated against Avial Flu disease. Access to feed and water was provided on an ad libitum basis.

Birds` performance measurements
Body weight and feed intake were monitored on a pen basis weekly and calculated individually, while weight gain, relative growth rate [Brody, 1945] and feed conversion ratio [Singh and Panda, 1992] values were calculated at the end of starting (0-14day of age ) and growing-finishing (14-35 day of age) periods. Mortality was also recorded on a daily basis in each pen. Chickens were killed by cervical dislocation at the end of the trial. Six birds per treatment group were randomly selected for determining carcass traits. They were de-feathered, eviscerated and dressed. Liver, gizzard, proventriculus, heart, thigh, breast, spleen and bursa of fabricius were collected, weighed and calculated as percentage of live weight.

Sampling and methods of analysis
Representative samples of mixed rations were taken for chemical analysis [A.O.A.C 2000]. At the end of 5 th week of age, blood samples were obtained from wing veins and directly liquated into 2-mL sterile vials, and allowed for one hour at room temperature (23Cº) for complete coagulation and for 3h in refrigerator (4Cº) for complete separation of serum before centrifugation at 1500 rpm, 6Cº for 20 min. The serum samples were used for determination of triglycerides [Fassati and Principe 1982], total cholesterol [Barnes et al., 1972], high density lipoprotein HDL [Najjar et al., 1991], low density lipoprotein LDL [Lee and Nieman, 1996], very low density lipoprotein VLDL [Lee and Nieman, 1996] and malondialdehyde [Wong et al 1987], glutathione reductase [Anderson, 1996] using Spectrophotometer and commercially available kits (Biosystem S.A, Costa Brava, 30, Barcelona, Spain) according to manufacturer's instructions.

Results and Discussion
Many investigations proved the role of IMO as prebiotic for human [Chen et al., 2001, Fu et al., 1999, animals [Crittenden, 1999, Kaneko et al., 1995, and poultry [Zhang, 2000 andZhang et al. 2003]. However, very few investigations were conducted on the role of IMO in the poultry as growth promoter or immune stimulant. In the present study, birds fed on diets supplemented with IMO had numerically the heavier body weight than the birds of control group. The heaviest significant body weights were for the birds fed on diet supplemented with 0.5g/Kg diet IMO compared with the control group but no difference when compared with that fed on diets contained 1g IMO/Kg of diet (Table 2). This effect was observed all over the experimental periods as reflex for improved relative growth rate for the treatment`s groups. This O c t o b e r 05, 2 0 1 4 effect agreed with the finding of Zhang et al. [2002] who showed enhanced body weights for birds fed on diets contained 0.1%, 0.2%, and 0.4% of IMO compared with control group and with those of Saminathan et al. [2014] who found an improved growth performance of broiler chickens fed on diets contained IMO. On the other hand, these results weren't agreed with the work of Dagcutan Broce [2013] who found no effect for feeding ration containing IMO on the growth performance of broilers. The overall Feed conversion ratio of the treatment`s groups were improved compared to the control group. Birds fed on diets supplemented with 0.5 g IMO/Kg diet have had the best significant feed conversion values. This improvement was observed at the ends of starting and finishing periods. Moreover, the feed intakes values were numerically decreased as the IMO increased (Table 2). This effect was agreed with the finding of Thitaram et al. This effect was demonstrated as a result of more aspects, the first was due to the stimulated glycolytic bacteria in the crop, because the level of amylase in chicken saliva is very low [Jerrett and Goodge, 1973], and only a trace of maltose or glucose is produced in the crop [Pritchard, 1972]. Secondly, production of more sugars from the cleavage of IMO at brush border of the intestinal mucosal cells [Hunziker et al., 1986;Najjar et al., 1991;Gu¨ nther and Heymann, 1998;Hertel et al., 2000]. Both aspects might enhanced the feed utilization of the treatment`s groups. On the other hand, Lactobacillius spp. is the most dominant bacterial species in the crop [Fuller, 1977;Mead, 1997;He et al., 2000]. It can inhibit growth of pathogens [Fu et al., 1999] by maintaining normal microbial balance in the crop [Fuller, 1973], which is important for broilers because the crop is the gateway for exotic bacteria to enter the intestines of birds. Also, Lactobacillius spp. is the only organism normally present in the duodeum and small intestine at levels significantly above 103 per gram in chickens [Barnes et al., 1972]. Most of the organic acids produced from fermentation of Lactobacillius spp. (such as L. fermentum, L. acidophilus, and L. plantarum) are lactic and acetic acids [Mitsuhashi et al., 1983] in addition to other organic acids, which initiate the prebiotic effect of IMO. Collectively, these suspected modes of action were included as causes for an improved growth performance. This increased growth performance proved the positive impacts for dietary IMO supplementation for broiler. These effects were observed in the trial of Zhang et al. [2003] during the first three weeks of age but weren't found after the 3 rd week of age due to the occurrence of infectious outbreak. Our results were in agreement with the finding of Thitaram et al. [2005]. However, Sutawee [1997] found no effects for dietary IMO supplementation of broilers diets on body weight and gain, which was attributed it to low dose of IMO supplementation. Table 3. Dressing percentages values increased with dietary supplementation od broilers diets with IMO compared to the control group. The difference was significant in the group fed high level of IMO (T2). Percentages of relative weight of breast, liver, gizzard, and heart increased significantly in treatment`s groups compared with the control group, where the largest value was for the group fed high level of IMO (T2). The difference was not significant for heart of group fed low level of IMO (T1). These improvement`s effects of IMO on carcass yield and parts might be due to its stimulant effects on growth performance. There are very few data describing the precise effects of IMO on carcass traits of broilers, although our results were in agreement with those of Ghiyasi et al. [2007] who found improved carcass and breast yields of broilers fed on low protein diets contained IMO as fermentation product of Aspergillus Orizae.

The effects of dietary treatments on carcass characteristics and some internal organs of 35 days old broilers are shown in
The mortality of each treatment was very low, and at most only one bird per control group died. Xia et al. [2001] reported that dietary IMO improved survivability of chicks. The indices of two immune organs are shown in Table 3. Differences in bursa and spleen weights due to IMO supplementation were significant compared to the control group except for bursa in groups fed low level of IMO (T1), which is numerically increased. The largest values were for the groups fed on diet contained high level of IMO (T2). There is increasing evidence that IMOs have immunomodulatory effects on systemic immune response including gut-associated lymphoid tissues system [Sung et al., 2004;Buchholz and Seibel, 2003]. Data in table (4) revealed a significant increased serum Log antibodies titer against avian flu by supplementation of broiler chicks with IMO at 21 day, 28 day, and 35 day of age compared with the control group. Difference between treatment`s groups and control one were significant except in the group fed low level of IMO only at 28 day of age. The increased weights of immune organs and serum antibody titer against avian flu along the experimental periods of broilers with IMO supplementation proved a stimulant effect of IMO on the immune system. Our results were agreed with those of Zhang et al. [2003] who found increased thymus index and weights of immune organs with dietary supplementation of broilers with IMO. Studies in mice gave strong evidence that IMO might stimulate the immune system [Watanabe et al., 2002].
As shown in table 5, although the serum levels of total protein and albumen were not affected by dietary supplementation with IMO, the serum globulin levels increased in all groups fed on diets supplemented with IMO compared with the control group. The difference was significant for group received low levels and insignificant for that received high levels of IMO. The increased levels of blood globulins confirmed the stimulant effects for dietary supplementation of broilers with IMO on the immunoglobulin's content of blood. Mizubuchi et al. [2005] observed greater levels of IgA in feces in mice fed for 4 weeks on a diet supplemented with 20% IMO. Unfortunately, our results data did not included the profile of white blood cells, which needs further investigation to assesses the finding of Li et al.

Conclusion
It was concluded that dietary supplementation of broiler chickens with 0.5 g/Kg of Isomaltooligosaccharides improved the growth performance, carcass traits and immune status but increasing dietary level of IMO to be 1 g/Kg decreased the total feed intake. O c t o b e r 05, 2 0 1 4