The diet and plane of nutrition are prominent factors that influence the blood picture and serum biochemistry of animals, which are sensitive indicators of the state of health and reflect the intensity of metabolic processes taking place in the animals (Vazquez-Anon et al., 2008; Andrews et al., 2006; Roy et al., 2013). Similarly, it has been reported that the serum biochemical parameters are the good indicators of the physiological, nutritional, and pathological status of animals/birds and can be correlated to identify the impact of nutritional factors and additives supplied in the diet (Toghyani et al., 2010). Thus, the dietary manipulations in broiler chicken are expected to reveal differential effects on the blood picture and serum biochemistry of broiler chicken. In this regard, the rice distillers dried grains with solubles (rDDGS) and rice gluten meal (RGM) are two potential economic feed ingredients of poultry ration produced by rice processing industries. India is one of the largest producers of rice in world producing approximately 105 million tons of rice in 2015-16 (Govt. of India, 2016). The rising cost of broiler chicken ration can be overcome by the use of ‘rice DDGS and rice gluten meal’, protein rich alternative feed ingredients to replace costly soybean meal. An increase in ethanol production over the last 5-10 years, due to increasing prices of conventional oil and limited underground reserves, has led to an increased supply of DDGS that may be available as livestock feed (Noll et al., 2007). On the other hand, the RGM, a by-product of wet-milling of rice, is available in appreciable amounts at lower cost compared to soybean. RGM contains 3152 kcal ME/kg, 46.45% crude protein, 3.4% ether extract and a favorable amino acid profile with relatively higher abundance of methionine (Kumar et al., 2016). It has been also designated as a source of rumen undegradable protein with the highest known metabolizable protein value among plant proteins (Heuze et al., 2015). Further, the exogenous enzyme supplementations in poultry diets, nutritionally, economically and environmentally justified, increase nutrient digestibility, reduce water content and viscosity of the excreta, and accelerate the rate of passage of digesta through the gastrointestinal tract (Sarmiento-Franco et al., 2003; Lazaro et al., 2004). In this study, it is hypothesised that the use of rDDGS and RGM in combination up to 12.5 and 15% levels respectively along with enzyme supplementation will not have any significant effect on the blood parameters and serum biochemistry of broiler chicken at 21 days of age. Thus, to test this hypothesis the objective of this study was to evaluate the effects of feeding different combinations of rDDGS and RGM levels along with enzyme supplementation on the haematology and serum biochemical parameters of broiler chicken.

Materials and Methods

Birds, Experimental Diets and Design

A total of 240 CARIBRO Vishal broiler chicks of same hatch and uniform weight were procured from the institutional hatchery and housed in specially designed battery brooder cages with standard watering and feeding facilities. A biological experiment of 42 days duration was undertaken in which day old 240 chicks were divided into six dietary treatments as per 2×3 factorial design having 5 replicates per treatment with 8 birds in each.  The broiler chicken ration were formulated by using rDDGS and RGM as replacement of soybean meal in the basal diets along with enzyme supplementation (ICAR, 2013). The levels of rDDGS and RGM and the suitable enzymes were standardised in a preliminary trail.  The rDDGS levels of 10% & 12.5%, and RGM levels of 12.5% & 15% along with either multienzyme or protease enzyme were selected to formulate the experimental diets as the pre-starter, starter and finisher diets. The feed ingredients and the nutrient composition of diets have been given in Table 1. A 2×3 factorial design resulted in six experimental diets viz T1 (no rDDGS/RGM/enzyme), T2 (no rDDGS/RGM, with multienzymes), T3 (12.5% rDDGS, 15% RGM, no enzyme), T4 (12.5% rDDGS, 15% RGM, with protease enzyme), T5 (10% rDDGS, 12.5% RGM, no enzyme), and T6 (10% rDDGS, 12.5% RGM, with protease enzyme). Each treatment was allocated 5 replicates of chicks, with 8 birds in each. The feeding trial was conducted for six weeks and the feed as well as drinking water were provided ad libitum to the birds during the entire experimental period.

Table 1: Ingredient and nutrient composition of broiler chicken diets

SBM: Soybean meal,, DORB: Deoiled rice bran, DDGS: Dried distillers grains with solubles, RGM: Rice gluten meal, LSP: Limestone powder, DCP: Di-calcium phosphate, CP: Crude protein, ME: Metabolizable energy; *Constant (0.4% salt, 0.1% trace mineral premix, 0.15% vitamin premix, 0.015% vit. B complex, 0.05% choline chloride and 0.05% Toxin binder); 1: Trace mineral premix supplied (mg/kg diet): Mg 300; Mn 55; I 0.4; Fe 56; Zn 30; Cu 4.; 2: Vitamin premix supplied (per kg diet): Vitamin A 8250 IU; Vitamin D3 1200 IU; Vitamin K 1mg; Vitamin E 40 IU.; 3: B complex: Vitamin B1 2 mg; Vitamin B2 4 mg; Vitamin B12 10 µg; niacin 60 mg; pantothenic acid 10 mg; choline 500 mg.

Sampling and Measurement

All the procedures carried out on the birds were approved by the Institutional Animal Ethics Committee of Indian Veterinary Research Institute, Izatnagar (452/01/ab/CPCSEA). For the analysis of haematological parameters 2 ml blood samples were collected from 8 birds (4 males and 4 females) per treatment at random in sterile heparin vials at 21 days of age of birds. All the blood samples were analyzed by automatic Abacus junior vet 5 haemato analyzer. The parameters studied were total leucocyte count (TLC), differential leucocyte count (DLC), Hb (%), packed cell volume (PCV), mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), mean corpuscular haemoglobin concentration (MCHC), RBC distribution width  (RDWc), platelet count, mean platelet volume (MPV) and platelet distribution width (PDWc). For evaluation of serum biochemistry of broiler chicken at 21 days of age, the blood samples were collected in the same manner as for the haematological analysis but in vials without any anticoagulant. The various serological parameters studied were serum glucose (Trinder, 1969), total protein (Doumas et al., 1971), albumin (Gustafsson, 1978), globulin, albumin: globulin ratio (A:G), cholesterol (Wybenga and Pileggi, 1970), triglyceride (Fossati and Lorenzo, 1982), SGOT (Reitman and Frankel, 1957), SGPT (Reitman and Frankel, 1957) and ALP (Kind and King, 1954).

The data collected were analysed by two way ANOVA using GLM procedure to present the results as means and standard errors (Snedecor and Cochran,1989) by using statistical package for social sciences (SPSS) 16.0 version and the comparison of significant mean differences was done by Duncan’s multiple range test (Duncan, 1955) with significance level set at P<0.05.

Results and Discussion

The results of feeding different level of rDDGS and RGM combinations with or without enzymes on haematological parameters of the broiler chicken of 21 days of age are given in Table 2. The blood profile was studied in terms of total erythrocyte count (TEC), total leukocyte count (TLC), differential leukocyte count (DLC), platelet count, Hb %, PCV, MCV, MCH, MCHC, MPV, heterophils and leukocyte ratio, RBC distribution width (RDWc) and platelet distribution width (PDWc).The results revealed that no significant (P>0.05) differences were observed in blood profile between control and other different dietary treatments having different levels of rDDGS and RGM combinations with or without enzymes and their interactions. All the blood parameters studied were well within the normal physiological range. In line with the results of the present study, Ghazalah et al. (2011) and Youssef et al. (2013) reported no significant effect of DDGS inclusion up to 15% level. Similarly, Wani et al. (2017) reported no significant (P>0.05) difference in Hb and PCV of broiler chicken by incorporating RGM up to 17.5% level with or without protease supplementation. However, in contrast to the results of the present study Gupta et al. (2017) reported inclusion of rDDGS level above 5% level significantly (P<0.01) enhance the PCV and Hb values of broiler chicken compared to control diet fed birds.

Table 2: Effect of feeding different level of rDDGS and RGM combinations haematological parameters

Values bearing different superscripts within the column differ significantly; NS: Non-significant (P>0.05); Units:WBC (x 104 ul),RBC (x106ul),Hb (g/dl),MCV (fl),MCH(pg),MCHC(g/dl) and PLT(x103ul)    

The results pertaining to feeding different level of rDDGS and RGM combinations on serological parameters on 21 days are presented in Table 3. The results revealed that no significant (P>0.05) difference was found in serum globulin (G), albumen(A): globulin (G) ratio, serum enzymes SGOT, SGPT and ALP activity between different dietary treatments and control as result of feeding 0% rDDGS +0% RGM (DR1), 12.5% rDDGS +15% RGM (DR2) and 10 % rDDGS +12.5 % RGM (DR3) combination levels. Serum glucose, total protein and albumen levels decreased significantly (P<0.01) in DR2 and DR3 as compared to DR1 level. However, serum glucose, total protein and albumen levels did not show any significant (P>0.05) difference from DR2 as compared to DR3. Serum triglyceride levels were decreased significantly (P<0.01) in DR2 level as compared to DR1 and DR3 levels. Serum triglyceride level did not show any significant (P>0.05) difference from DR3 as compared to DR1 level. Effect of with or without enzymes supplementation on feeding different level of rDDGS and RGM combinations did not show any significant (P>0.05) difference in serological parameters except serum glucose. Serum glucose level was increased significantly (P<0.01) in enzymes supplemented groups as compared to without enzymes groups. Interaction of rDDGS and RGM combinations with or without enzymes did not show any significant (P>0.05) difference on serological parameters on 21 days of broiler chickens. Our results were in agreement with Choi et al. (2014), Metwally and Farhat (2015), Kumar et al. (2016) and Wani et al. (2017). Choi et al.(2014) reported dietary inclusion of rice DDGS up to 25% did not affect the plasma content of total protein, glucose, cholesterol and triglyceride in juvenile red sea bream (Pagrus major) fish. Metwally and Farhat (2015) reported no significant (P>0.05) difference in serum biochemical parameters (serum lipid profile, glucose, total protein, albumin and globulin) up to addition of 12.5% RGM in diet of broiler chicken. Kumar et al. (2016) also reported no significant (P>0.05)  difference in the serological variables (glucose, blood urea nitrogen, plasma proteins and non-esterified fatty acids) on addition of RGM up to 21% level in the diet of growing dairy calves. Wani (2017) and Wani et al. (2017) reported no significant (P>0.05) difference on feeding different levels of RGM up to 20% level with or without protease enzyme supplementation. Moreover, serum biochemical parameters are indicators of the physiological, nutritional and pathological status of birds and can be correlated to identify the impact of nutritional factors and additives supplied in the diet.

Our results were in disagreement with Hack et al. (2015) and Gupta et al. (2017). Hack et al. (2015) reported that increasing corn DDGS level significantly (P<0.01) increased serum triglycerides, cholesterol and LDL for hens fed diet contained 22% DDGS in the diet. Gupta et al. (2017) reported 10% rice DDGS had significantly (P<0.01) higher effect on serum albumin, total protein, A:G ratio, glucose value and significantly (P<0.01) lowering effect on serum lipid profile (triglycerides, cholesterol, LDL and VLDL). However, no such study on rDDGS and RGM combination feeding were done in poultry on blood biochemical parameters till now.

Table 3: Effect of feeding different level of rDDGS and RGM combinations on serological parameters

Values bearing different superscripts within the column differ significantly; A:G ratio: Albumin: Globulin ratio, NS: Non-significant (P>0.05); M: Multienzyme, P: Protease

Conclusion

The study concludes that the feeding of rDDGS and RGM in combination up to the levels of 12.5% and 15%, respectively with or without enzyme supplementation do not have any adverse effects on the haematological parameters but adversely affecting serum biochemical parameters in terms of serum glucose, total protein and albumen levels of broiler chicken in 21 days.

Acknowledgement

Authors are highly thankful to ICAR-Central Avian Research Institute, Izatnagar, Bareilly, U.P. for providing all necessary facilities and inputs.

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