Influence of prepelleting inclusion of whole corn on performance, nutrient utilization, digestive tract measurements, and cecal microbiota of young broilers Y. Singh,* V. Ravindran,*1 T. J. Wester,* A. L. Molan,†2 and G. Ravindran† *Institute of Veterinary, Animal and Biomedical Sciences, and †Institute of Food, Nutrition and Human Health, Massey University, Palmerston North 4442, New Zealand further inclusions. Relative gizzard weight (quadratic effect, P < 0.05) increased with increasing inclusion of whole corn up to 300 g/kg and then levelled off. The AME (quadratic effect, P < 0.05) increased up to 300 g/kg of whole corn inclusion and then decreased with further inclusion. Apparent ileal digestibility of DM (P < 0.001), N (linear effect, P = 0.07), and starch (linear effect, P < 0.001) increased with increasing inclusion levels of whole corn. Based on the fluorescence in situ hybridization method, a linear (P < 0.05) effect was determined for cecal microbiota numbers. Lactobacillus spp. counts increased and counts of Clostridium spp., Campylobacterium spp., and Bacteroides spp. decreased with increasing inclusion levels of whole corn. The present data showed that, despite increased gizzard weight and nutrient utilization, weight gain of broilers was poorer with prepelleting inclusion of whole corn due to reductions in the feed intake.

Key words: whole corn, prepelleting, digestive tract, cecal microbiota, broiler 2014 Poultry Science 93:3073–3082 http://dx.doi.org/10.3382/ps.2014-04110

INTRODUCTION There is growing interest in the feeding of whole grains to broilers as a means of lowering feed costs and because of the reported positive effects on production performance and nutrient utilization (Singh et al., 2014). These positive effects are generally attributed to the effect of feed structure on gizzard development. In addition, there is some evidence that whole grain feeding may also influence intestinal microbiota ecology (Engberg et al., 2004; Santos et al., 2008). This has encouraged nutritionists to consider whole grain feeding as a natural feeding strategy to improve bird ©2014 Poultry Science Association Inc. Received April 11, 2014. Accepted September 5, 2014. 1 Corresponding author: [email protected] 2 Present address: Department of Biology, College of Sciences, Diyala University, Diyala, Iraq.

health (Mateos et al., 2002), and such an approach is becoming especially relevant in the context of increasing consumer concerns over the use of antibiotic growth promoters in poultry diets. Wheat has been the grain of choice for whole grain feeding in poultry. However, throughout the world, poultry diets are primarily based on corn. Despite this, little or no attempt has been made to use whole corn in poultry diets. The size of corn grain may physically limit feeding it whole, and this may be responsible for the lack of published data on the use of whole corn in poultry diets. Prepelleting inclusion of whole corn could be a strategy to overcome this limitation, while delivering the benefits associated with whole grain feeding. Studies examining the influence of prepelleting inclusion of whole grains in broiler diets are limited and have produced equivocal results. Wu et al. (2004) reported that prepelleting inclusion of 200 g/kg of whole wheat had no effect on gizzard development, but improved

3073

Downloaded from http://ps.oxfordjournals.org/ at Universite Laval on December 1, 2014

ABSTRACT The objective of the present study was to examine the effects of prepelleting inclusion of graded levels of whole corn on performance, digestive tract measurements, nutrient utilization, and cecal microbiota in broiler starters. Five diets, containing 600 g/ kg of ground corn or 150, 300, 450, and 600 g/kg of whole corn replacing (wt/wt) ground corn, were formulated and cold-pelleted at 65°C. Each diet was offered ad libitum to 6 replicates (8 birds per replicate cage) from d 1 to 21 posthatch. The proportion of coarse particles (>1 mm) increased with increasing prepelleting inclusion of whole corn. Pellet quality, measured as pellet durability index, increased (quadratic effect, P < 0.001) with the inclusion of whole corn to 450 g/kg and then plateaued. Weight gain and feed intake decreased (linear effect, P < 0.001) with increasing prepelleting inclusion of whole corn. Feed per gain (quadratic effect, P < 0.05) increased as the inclusion level of whole corn increased to 300 g/kg and then plateaued with

3074

Singh et al.

MATERIALS AND METHODS Hardness of Corn Corn was purchased as whole grain from a local supplier. Upon receipt, requisite samples were obtained for the measurement of grain hardness. Hardness was determined using a micro hammer mill (Stenvert Hardness Tester, Glen Creston Ltd. Stanmore, UK) as per the method described by Abdollahi et al. (2011). Corn samples (20 g) were ground using a micro hammermill (5,692 rpm) equipped with a 0.2-mm sieve, and the energy (KJ) needed to grind the sample was determined. Measurements were made on 5 samples and adjusted to 140 g/kg moisture. Corn used in the experiment was determined to require 6.2 KJ energy to mill a 20-g sample, which confirmed that it was yellow dent, a soft corn variety (Li et al., 1996).

Birds and Housing Experimental procedures were approved by the Massey University Ethics Committee. One-day-old male broiler chicks (Ross 308), obtained from a commercial hatchery, were assigned to 30 cages (8 birds per cage) in 3-tier electrically heated battery brooders in a manner so that the average bird weight per cage was similar. The birds were transferred to grower colony cages on d 11. Battery brooders and grower colony cages were housed in an environmentally controlled room. In both these cages, a floor space of 680 cm2 per bird was provided. Room temperature was maintained at 32

± 1°C during the first week and gradually reduced to 21 ± 1°C by the end of the third week. Twenty hours of fluorescent lighting was provided per day throughout the experimental period.

Diets Five diets containing 600 g/kg of ground corn or 150, 300, 450, and 600 g/kg of whole corn replacing (wt/wt) ground corn were formulated to meet Ross 308 strain recommendations for major nutrients for broiler starters (Ross, 2007; Table 1). The whole corn and ground corn used in the study were from the same lot. For the ground corn component, whole corn was ground in a single batch in a hammer mill (Bisley’s Farm Machinery, Auckland, New Zealand) to pass through a 4.0mm screen. All ingredients and titanium dioxide were mixed, the diets were conditioned at 65°C by adjusting the steam flow and pelleted. Pelleting was done in a pellet mill (Richard Size Limited Engineers, Orbit, Kingston-upon-Hull, UK) capable of manufacturing 180 kg of feed/h and equipped with a die ring of 3-mm hole size and 35-mm thickness. Smaller pellets of 3 mm length and 3 mm diameter were produced, and these pellets resembled crumbles. Titanium dioxide (3 g/kg) was included in diets as an inert marker. Each of the 5 dietary treatments was then randomly allocated to 6 replicate cages. Feed was offered ad libitum and water was available at all times throughout the 21-d trial.

Determination of Particle Size Distribution in Pelleted Diets The wet sieving method was used to determine particle size distribution of pelleted diets (Lentle et al., 2006). Briefly, a representative sample of the pelleted diet (50 g) was divided into 2 portions. One-half of the sample was oven-dried at 80°C in a forced draft oven for 24 h to determine DM content, whereas the other half was suspended in 50 mL of water for 30 min before being washed though a series of sieves (Endecott, London, UK) sized 2, 1, 0.5, 0.212, 0.106, and 0.075 mm. Contents of each sieve were subsequently washed onto preweighed filter papers, and then dried for 24 h in a forced draft oven at 80°C. The amount of diet retained by each sieve and fines that passed through all sieves were weighed and expressed as a percentage of total DM recovered.

Pellet Quality Pellet durability of pelleted diets was measured by the method described by Svihus et al. (2004b) using a Holmen Pellet Tester (New Holmen Pellet Tester, Tekpro Ltd., Norfolk, UK). Briefly, weighed samples of pellet (100 g) were circulated pneumatically through a closed chamber before being passed through a 2-mm sieve. Pellet durability index (PDI) was calculated

Downloaded from http://ps.oxfordjournals.org/ at Universite Laval on December 1, 2014

the performance of broilers. In contrast, Jones and Taylor (2001) and Taylor and Jones (2004) observed that prepelleting inclusion of 200 g/kg of whole wheat had no effect on broiler performance, but increased relative gizzard weights. Svihus et al. (2004a) increased the prepellet inclusion level of whole wheat to 500 g/kg and found no differences in performance parameters, but significant increases in gizzard weight compared with birds fed ground wheat diets. Differences in experimental methodology, inclusion level of whole grain, length of feeding, age of birds, and type and quality of grain may be responsible for these inconsistent results (Singh et al., 2014). Most previous studies have evaluated a constant dietary inclusion level of whole grain (Plavnik et al., 2002; Taylor and Jones, 2004; Wu et al., 2004; Ravindran et al., 2006; Amerah and Ravindran, 2008). Graded levels of prepelleting inclusion of whole grain in broiler diets may provide a better understanding of its effects on production performance and nutrient utilization responses. The aim of the present study was to examine the effects of prepelleting inclusion of graded levels of whole corn on performance, energy and nutrient utilization, digestive tract development, and cecal microbial counts of young broiler chickens.

PREPELLETING INCLUSION OF WHOLE CORN Table 1. Composition and analysis of the starter diet1 Item

600.0 297.4 20.7 59.4 2.0 8.0 1.0 2.5 3.0 3.0 3.0   3,035 223 12.6 9.6 9.0 2.4 10.4 6.9 4.5   880 3,940 229 359

1In the treatment diets, ground corn was replaced (wt/wt) by 0, 150, 300, 450, and 600 g/kg of whole corn before pelleting. 2Supplied per kilogram of premix: ethoxyquin, 100 mg; biotin, 0.2 mg; calcium pantothenate, 12.8 mg; cholecalciferol, 60 µg; cyanocobalamin, 0.017 mg; folic acid, 5.2 mg; menadione, 4 mg; niacin, 35 mg; pyridoxine, 10 mg; trans-retinol, 3.33 mg; riboflavin, 12 mg; thiamine, 3.0 mg; dl-αtocopheryl acetate, 60 mg; choline chloride, 638 mg; Co, 0.3 mg; Cu, 3 mg; Fe, 25 mg; I, 1 mg; Mn, 125 mg; Mo, 0.5 mg; Se, 200 µg; Zn, 60 mg. 3Analyzed values for the ground corn diet.

as the ratio between the amount pellets not passing through the sieve after the test to amount of whole pellets at the beginning. Pellet hardness was measured in a Stable Micro System Texture Analyzer (TA-XT Plus, Godalming, Surrey, UK) using the method as described by Svihus et al. (2004b). Fifteen individual pellet samples were placed between the pressure piston and bar. By increasing the pressure applied by means of the pressure piston, the force (Newtons) needed to break the pellet was determined.

Performance Parameters Body weights and feed intake were recorded on a cage basis at weekly intervals. Mortality was recorded daily. Any bird that died was weighed, and the weight was included in weekly weight gain data and used to adjust feed per gain.

AME Determination Feed intake and excreta output were measured quantitatively for 4 d between d 17 and 20 posthatch. Excreta were collected daily and stored at −20°C. Daily

collections were then pooled within a cage, mixed well in a blender, and subsampled. Subsamples were freezedried, mixed well, ground to pass through a 0.5-mm screen, and stored in airtight plastic containers at 4°C until analysis. Excreta and diet samples were analyzed for the DM and gross energy (GE).

Ileal Digestibility Determination On d 21, four birds per cage were selected randomly and euthanized by intravenous injection of sodium pentobarbitone (Provet NZ Pvt. Ltd., Auckland, New Zealand) at 0.5 mL per kg of BW. Immediately after euthanasia, portion of small intestine extending from Meckel’s diverticulum to a point 40 mm proximal to the ileo-cecal junction (ileum) was carefully excised. The contents of the lower half of ileum were collected by gently flushing with distilled water into plastic containers. Digesta samples were pooled within a cage, freezedried, ground to pass through a 0.5-mm screen size, and stored in an air-tight container at 4°C until laboratory analysis. Digesta and diet samples were analyzed for DM, N, Ti, and starch.

Determination of Cecal Microbiota Cecal contents from birds fed diets containing 0, 300, and 600 g/kg of whole corn were collected. Following ileal digesta collection (4 birds/cage), ceca were carefully excised and stored at −20°C until determination of microbiota using a fluorescence in situ hybridization (FISH) technique as per the procedure described by Dinoto et al. (2006) with some minor modifications (Molan et al. 2009). In brief, cecal contents were prepared by mixing 1 g with 9 mL of sterile-filtered phosphate buffer (PBS; pH 7.2), and the bacteria contained in the supernatant were fixed in 4% (wt/vol) of paraformaldheyde in PBS (pH 7.2). After the fixing step, the samples were applied to Teflon-coated microscopic slides (Biolab, North Shore City, New Zealand) and air dried. Bacterial cells were then dehydrated with ethanol solutions and fixed on the glass slides. Slides were hybridized by hybridization buffer, labeled with Cy3-labeled oligonucleotide specific probes (50 ng/μL), and placed in a plastic box containing wet sponges (soaked in hybridization buffer) at 46°C for 2 h. Slices were rinsed with warm hybridization buffer at 48°C and washed in prewarmed washing buffer for 20 min at 48°C. After the washing step, slides were rinsed with ice-cold distilled water and thoroughly dried before being observed using a fluorescence scanning microscope (Olympus BX51, Olympus Corporation, Tokyo, Japan) under 400× magnification. The images were captured using an Optronics MagnaFIRE SS99802 digital camera with MagnaFIRE frame-grabbing software on a Pentium IV computer. Fluorescent cells were counted automatically using ImageJ program following Abramoff et al. (2004).

Downloaded from http://ps.oxfordjournals.org/ at Universite Laval on December 1, 2014

Ingredient  Corn   Soybean meal, 480 g/kg CP   Soybean oil   Meat and bone meal   Dicalcium phosphate  Limestone   l-Lysine∙HCl   dl-Methionine   Sodium chloride   Titanium dioxide   Vitamin-trace mineral premix2 Calculated analysis   AME (kcal/kg)  CP  Lysine   Methionine + cysteine  Threonine  Tryptophan  Calcium   Total phosphorus   Available phosphorus Analyzed value3  DM   Gross energy (kcal/kg)   CP (N × 6.25)  Starch

g/kg, as fed

3075

3076

Singh et al.

Digesta pH and Digestive Tract Measurements

Chemical Analysis Dry matter was determined using standard procedures (method 930.15; AOAC International, 2005). Gross energy was determined by adiabatic bomb calorimeter (Gallenkamp Autobomb, London, UK) stan-

Calculations The AME values were calculated using the following formula with appropriate corrections made for differences in DM content: AME (kcal/kg) = [(feed intake × GEdiet) − (excreta output × GEexcreta)]/feed intake, where GEdiet is the GE of diet and GEexcreta is the GE of excreta. Similarly, the total tract DM retention was calculated based on differences in DM intake and DM output. Apparent ileal digestibility coefficients of DM, N, and starch were calculated based on Ti marker ratios in the diet and ileal digesta using the formula below: Ileal digestibility coefficient = [(nutrient/Ti)d − (nutrient/Ti)i]/(nutrient/Ti)d, where (nutrient/Ti)d is the ratio of nutrient to Ti in the diet, and (nutrient/Ti)i is the ratio of nutrient to Ti in the ileal digesta.

Figure 1. Particle size distribution of pelleted diets containing 0, 150, 300, 450, and 600 g/kg of whole corn (WC).

Downloaded from http://ps.oxfordjournals.org/ at Universite Laval on December 1, 2014

On d 21, two birds (closest to the mean cage weight) were selected from each cage. Body weights were recorded, and birds were killed by cervical dislocation. The gastrointestinal tract and organs were carefully excised, and adherent mesentery and fat were removed. Digesta pH was measured with a calibrated digital pH meter (model IQ120, 2075-E Corte Del Nogal, Carlsbad, CA). In a sequential manner, segments of the gastrointestinal tract from crop to cecum were opened in the middle by an incision and split along the length longitudinally, and the pH was quickly recorded by inserting the glass-tipped probe directly into the digesta (German and Bittong, 2009). Readings were recorded after the value stabilized. Empty weights of proventriculus and gizzard, and weights of pancreas, liver, and spleen were recorded. Different segments of small intestine were emptied by gentle pressure, and empty weight and length of duodenum (pancreatic loop), jejunum (from the pancreatic loop to Meckel’s diverticulum), ileum (from Meckel’s diverticulum to 1 cm above ileo-cecal junction), and ceca (ostium to tip) were recorded. The length of various intestinal segments was determined with a flexible tape on a wet glass surface to prevent inadvertent stretching. Relative organ weights (g/kg of BW) and relative length (cm/kg of BW) were calculated.

dardized with benzoic acid. Nitrogen was analyzed by combustion (method 968.06; AOAC International, 2005) using a CNS-200 carbon, nitrogen, and sulfur autoanalyzer (Leco Corporation, St. Joseph, MI). Titanium was determined by sulfuric acid digestion followed by colorimetric determination on a UV spectrophotometer as described by Short et al. (1996). Starch was measured using an assay kit (Megazyme, International Ireland Ltd., Wicklow, Ireland) based on conversion of starch to glucose using thermostable α-amylase and amyloglucosidase (McCleary et al., 1997).

3077

PREPELLETING INCLUSION OF WHOLE CORN

Pellet Quality

Table 2. Influence of inclusion of graded levels of whole corn on the pellet durability index (PDI) and pellet hardness of the diets

Item Whole corn inclusion (g/kg)  0  150  300  450  600  SEM3 Probability   Overall treatment effect   Linear effect   Quadratic effect

PDI1 (%)

Pellet hardness2 (N)

64.4 77.9 77.5 81.9 83.5 0.42   0.0001 0.0001 0.0001

16.4 22.2 26.0 27.9 35.5 1.44   0.001 0.0001 0.75

Influence of dietary treatments on the pellet durability and hardness is presented in Table 2. Pellet durability, measured as PDI, showed a quadratic (P < 0.001) response. Pellet durability index increased with the inclusion of whole corn to 450 g/kg and then plateaued. A linear (P < 0.001) increase in pellet hardness was observed with increasing inclusion of whole corn.

Performance

1Each

value represents the mean of 6 samples. value represents the mean of 15 samples. 3Pooled SEM. 2Each

Data Analysis Cage means served as the experimental unit for performance parameters, nutrient digestibility, and microbiota numbers. For digestive tract parameters and pH, individual birds were considered as the experimental unit. All data were subjected to orthogonal polynomial contrasts using the GLM procedure of SAS Institute Inc. (2004) to examine whether responses to increasing levels of prepelleting inclusion of whole corn were of a linear or quadratic nature. Effects were considered significant when P < 0.05.

Nutrient Utilization A quadratic effect (P < 0.05) was observed for AME and total tract retention of DM with increasing inclusion levels of whole corn (Table 4). The AME (quadratic effect, P < 0.05) increased up to 300 g/kg of whole corn inclusion and then decreased with further inclusion. Inclusion of whole corn increased the total tract DM retention up to 300 g/kg of whole corn inclusion and then levelled off with further inclusion. The apparent ileal digestibility coefficient of DM (P < 0.001) and starch (P < 0.001) increased linearly, and there was a tendency (P = 0.07) for that of N to linearly increase as the inclusion level of whole corn increased.

RESULTS Particle Size Distribution Particle size distribution of pelleted diets is shown in Figure 1. The proportion of coarse particles (>1 mm) increased with increasing prepelleting inclusion of whole corn. The percentage of particles over 1 mm were 8.9, 14.1, 16.0, 18.3, and 19.5% and those over 2 mm were 0.28, 2.1, 3.8, 4.7, and 6.0% in diets prepelleted with 0, 150, 300, 450, and 600 g/kg of whole corn, respectively.

Digestive Tract and Organ Measurements Influence of prepelleting inclusion of whole corn on the relative weights of proventriculus, gizzard, and di-

Table 3. Influence of prepelleting inclusion of graded levels of whole corn on weight gain, feed intake, and feed per gain of broilers from 1 to 21 d posthatch1 Whole corn inclusion (g/kg) Item

0

Weight gain (g/bird)   Wk 1   Wk 2   Wk 3 Feed intake (g/bird)   Wk 1   Wk 2   Wk 3 Feed per gain (g/g)   Wk 1   Wk 2   Wk 3 1Each

148 489 1,005 163 589 1,303 1.107 1.214 1.304

150  

142 470 990   162 596 1,312   1.117 1.267 1.324

300  

121 416 919   145 528 1,214   1.199 1.270 1.334

450  

120 421 933   149 536 1,226   1.253 1.297 1.341

value represents the mean of 6 replicates (8 birds per replicate). SEM.

2Pooled

Probability SEM2

600  

109 368 857   139 484 1,136   1.295 1.327 1.341

 

3.9 11.7 19.8   3.4 12.5 25.6   0.020 0.012 0.006

Treatment  

0.0001 0.0001 0.0001   0.0001 0.0001 0.0001   0.0001 0.0001 0.0001

 

Linear

Quadratic

0.0001 0.0001 0.0001   0.0001 0.0001 0.0001   0.0001 0.0001 0.0001

  0.77 0.82 0.62   0.51 0.33 0.37   0.16 0.12 0.05

Downloaded from http://ps.oxfordjournals.org/ at Universite Laval on December 1, 2014

Mortality during the experiment was negligible. Only 3 birds out of 240 died and the deaths were not related to any specific treatment. Linear (P < 0.001) decreases in weight gain and feed intake were observed throughout the trial period with increasing inclusions of whole corn (Table 3). However, a quadratic (P < 0.05) response was observed for feed per gain. Feed per gain increased with increasing inclusion levels of whole corn to 300 g/kg and then plateauing with further inclusion.

3078

Singh et al.

Table 4. Influence of prepelleting inclusion of graded levels of whole corn on the AME (kcal/kg of DM) and coefficients of total tract DM retention and apparent ileal DM, N, and starch digestibility in broilers1 Item Whole corn inclusion (g/kg)  0  150  300  450  600  SEM2 Probability   Overall treatment effect   Linear effect   Quadratic effect

AME

Total tract DM retention

Ileal DM digestibility

Ileal N digestibility

Ileal starch digestibility

3,401 3,446 3,466 3,424 3,435 11.1   0.05 0.26 0.05

0.696 0.718 0.743 0.724 0.738 0.003   0.01 0.001 0.01

0.683 0.691 0.715 0.722 0.724 0.008   0.01 0.01 0.13

0.788 0.802 0.809 0.827 0.826 0.008   0.40 0.07 0.65

0.966 0.973 0.978 0.988 0.993 0.001   0.001 0.0001 0.09

1Each value represents the mean of 6 replicates (4 or 8 birds per replicate). The AME and total tract DM retention were determined using total excreta collection between d 17 and 20 posthatch. Ileal digestibility measurements were made on d 21 posthatch. 2Pooled SEM.

of ceca and overall length of small intestine was unaffected when prepelleting inclusion of whole corn was increased to 450 g/kg, whereas an increase from 450 to 600 g/kg resulted in a significant increase in relative lengths. A linear (P < 0.001) decrease in pH of gizzard contents was observed with increasing inclusion level of whole corn (Table 7). A quadratic response (P < 0.05) was seen for pH of proventriculus contents. But no effects (P > 0.05) were observed for pH of digesta from any intestinal segment or ceca. A linear (P < 0.05) increase in pH of excreta was observed with increasing inclusion of whole corn.

Cecal Microbiota Counts Effects of prepelleting inclusion of 0, 300, and 600 g/ kg of whole corn on cecal microbial populations are presented in Table 8. With increasing inclusion of whole corn, there was a linear increase in numbers of Lactobacillus spp. (P < 0.01) and linear decreases in numbers of Clostridium spp. (P < 0.01), Campylobacter spp. (P < 0.05), and Bacteroides spp. (P < 0.05). Dietary

Table 5. Relative weights of proventriculus, gizzard, and intestinal organs (pancreas, liver, and spleen) of broilers as influenced by prepelleting inclusion of graded levels of whole corn1 Relative weight (g/kg of BW) Item Whole corn inclusion (g/kg)  0  150  300  450  600  SEM2 Probability   Overall treatment effect   Linear effect   Quadratic effect 1Each

Proventriculus

Gizzard

Pancreas

Liver

Spleen

3.94 4.33 4.27 4.57 4.70 0.141   0.001 0.01 0.77

11.9 14.4 15.0 15.7 15.5 0.56   0.0001 0.0001 0.05

2.37 2.93 2.78 2.93 2.89 0.160   0.05 0.05 0.06

31.1 29.6 28.5 28.5 29.7 0.85   0.18 0.16 0.05

0.872 0.938 0.835 0.800 0.844 0.069   0.70 0.38 0.93

value represents the mean of 6 replicates (2 birds per replicate). Measurements were made on d 21 posthatch. SEM.

2Pooled

Downloaded from http://ps.oxfordjournals.org/ at Universite Laval on December 1, 2014

gestive organs is summarized in Table 5. A quadratic effect (P < 0.05) was observed for the relative gizzard weight, with the weight increasing as inclusion of whole corn increased to 300 g/kg and then plateauing with further inclusion. Relative weights of proventriculus and pancreas showed a linear (P < 0.05) increase with increasing inclusion of whole corn. A quadratic (P < 0.05) response was observed for the relative weight of liver, with the weight decreasing at 150 g/kg inclusion, then plateauing with further inclusion and increasing from 450 to 600 g/kg inclusion. The relative weight of the spleen was unaffected (P > 0.05) by dietary treatments. Effects of prepelleting inclusion of graded levels of whole corn on the relative weight and length of different segments of the intestinal tract are shown in Table 6. A linear (P < 0.05) increase in the relative weight of jejunum with increasing inclusion of whole corn. Responses were observed for the relative length of ileum (linear effect, P < 0.01) and ceca (quadratic effect, P < 0.05). A quadratic (P < 0.05) effect was also observed for the relative length of the small intestine. As the inclusion level of whole corn was increased, a linear increase was observed for the length of the ileum. Length

151 157 155 155 170 4.6   0.06 0.01 0.05

treatments had no effect (P > 0.05) on Bifidobacteria spp. counts.

63.7 66.3 66.7 66.9 73.0 2.10   0.05 0.01 0.39

14.6 14.2 14.2 14.0 15.7 0.47   0.16 0.10 0.05

DISCUSSION

value represents the mean of 6 replicates (2 birds per replicate). Measurements were made on d 21 posthatch. intestine = duodenum + jejunum + ileum. 3Pooled SEM. 2Small

1Each

Whole corn inclusion (g/kg)  0  150  300  450  600  SEM3 Probability   Overall treatment effect  Linear  Quadratic

6.45 7.03 6.30 6.03 6.41 0.28   0.17 0.28 0.96

9.62 10.1 9.82 10.2 10.8 0.35   0.14 0.05 0.51

6.94 7.28 6.95 6.96 7.60 0.295   0.43 0.29 0.40

3.40 3.24 3.09 3.16 3.18 0.120   0.49 0.20 0.21

23.0 24.4 23.0 23.2 24.8 0.69   0.21 0.07 0.68

26.5 28.6 27.5 26.0 28.3 1.06   0.44 0.78 0.94

61.0 62.0 61.7 62.9 68.8 2.06   0.06 0.01 0.13

Ceca Ileum Jejunum Item

Duodenum

Jejunum

Ileum

Ceca

Small intestine2

Duodenum

Relative length (cm/kg of BW) Relative weight (g/kg of BW)

3079

To the authors’ knowledge, no published data are available on the effects of whole corn inclusion in poultry diets. In the present study, feeding broiler starter diets with increasing prepelleting inclusion levels of whole corn resulted in poor weight gain despite better development of the gizzard. Weight gain decreased linearly with increasing inclusion of whole corn and this paralleled the linear depression observed for feed intake. Reasons for the reductions in feed intake are unclear, especially because the pellet quality, measured as PDI, improved with increasing inclusion of whole corn. Benefits in broiler performance associated with pellet quality have been well documented (Calet, 1965; Abdollahi et al., 2013). Poor pellet quality reduces feed intake and bird performance (Proudfoot and Sefton, 1978; Nir et al., 1995). In the present study, however, feed intake was reduced despite improved PDI. This observation calls into question whether the PDI measured using the Holmen Pellet Tester is a good measure of pellet quality when pellets with whole grains or coarse ingredients are tested. This method of PDI determination is most reliable when the particle size of ingredients making up the pellet is similar. In the present study, there were large differences in the particle size among ingredients because of the relatively large size of whole corn, and as corn inclusion rate increased, proportion of larger particles making up the pellets increased. The Holmen Pellet Tester measures pellet durability by circulating a sample of pellets in a stream of air, forcing them into contact with each other and tester walls. An excluder screen allows smaller particles (e.g., from broken pellets) to leave the tester. The weight of material remaining within the tester is a measure of pellet durability. The current findings suggest that, when pellets contain particles too large to pass through the excluder screen and leave the pellet tester even if pellets break, the standard Holman Pellet Testing (Svihus et al., 2004b) is not useful to measure the true durability of pellets with whole grains. It would appear that modifications such as changing the size of the excluder screen are needed to improve the durability measurement of pellets containing whole grains. From visual observations, it appeared that the selective ingestion of larger corn particles, whose proportion progressively increased in pellets with increasing prepelleting inclusion level of whole corn, was partly responsible for the reductions in feed intake. It is also possible that the birds were attracted by the color of the corn particles. It is possible that the linear increases in pellet hardness with increasing inclusion of whole corn may also have partly contributed to the observed reductions in feed intake. Clark et al. (2009) similarly reported linear decrease in weight gain and feed intake in broilers fed diets containing 150, 300, 450, or 600

Downloaded from http://ps.oxfordjournals.org/ at Universite Laval on December 1, 2014

Table 6. Relative weight and length of the intestinal tract of broilers as influenced by prepelleting inclusion of graded levels of whole corn1

Small intestine2

PREPELLETING INCLUSION OF WHOLE CORN

3080

Singh et al.

Table 7. Influence of prepelleting inclusion of whole corn on pH of digesta from various segments of digestive tract and excreta of broilers1 Item Whole corn inclusion (g/kg)  0  150  300  450  600  SEM2 Probability   Overall treatment effect   Linear effect   Quadratic effect 1Each

Proventriculus

Gizzard

Duodenum

Jejunum

Ileum

Ceca

Excreta

3.88 3.01 3.00 3.16 3.20 0.231

4.01 3.98 3.48 3.35 3.50 0.134   0.01 0.001 0.16

5.76 6.06 5.96 5.93 6.01 0.086   0.17 0.19 0.27

5.93 6.15 6.06 6.08 6.10 0.080   0.42 0.30 0.33

6.63 6.66 6.93 6.90 6.96 0.125   0.21 0.05 0.62

6.46 6.58 6.38 6.60 6.53 0.080   0.32 0.56 0.87

7.16 7.55 7.61 7.65 7.94 0.208   0.16 0.05 0.38

0.07 0.11 0.05

value represents the mean of 6 birds. Measurements were made on d 21 posthatch. SEM.

2Pooled

hus, 2011). Grinding action of the gizzard activates cholecystokinin release (Li and Owyang, 1993; Svihus et al., 2004a), which in turn stimulates secretion of pancreatic enzymes and gastro-duodenal reflux (Duke, 1992). Increased gut motility serves to re-expose digesta to pepsin, enhancing mixing of digesta with pancreatic enzymes, and improving digestion of nutrients (Hetland et al., 2002; Ravindran et al., 2006). Increases in the relative weight of the pancreas and in the ileal digestibility of DM, N, and starch observed in the present study lend support to this thesis. Increased relative weight of the pancreas with whole wheat feeding has been reported previously (Banfield et al., 2002; Engberg et al., 2004; Wu et al., 2004; Gabriel et al., 2008). In the present study, prepelleting inclusion of whole corn increased the AME and total tract DM retention up to 300 g/kg of whole corn inclusion and then decreased or levelled off with further inclusions. These findings are in general agreement to previous studies with whole wheat that showed inclusion of whole wheat at levels of 200 to 375 g/kg improved the AME (Svihus et al., 2004a; Wu et al., 2004). It is also relevant to note that the inclusion of 500 g/kg of whole wheat had no effect on the AME (Svihus et al., 2004a). The increase in the relative weight of the proventriculus observed in the present study with increasing inclusion of whole corn is in contrast to several studies showing that birds fed whole wheat had either reduced

Table 8. Influence of prepelleting inclusion of graded levels of whole corn on cecal microbiota counts (log10 cells/g of cecal content) in broilers1 Item Whole corn inclusion (g/kg)  0  300  600  SEM2 Probability   Overall treatment effect   Linear effect   Quadratic effect 1Each

Lactobacillus spp.

Bifidobacteria spp.

Clostridium spp.

Campylobacter spp.

Bacteroides spp.

6.94 7.21 7.35 0.079

6.29 6.40 6.29 0.069   0.48 0.99 0.24

7.43 6.93 6.84 0.138   0.05 0.01 0.23

6.24 6.10 5.91 0.109   0.14 0.05 0.84

6.40 6.17 6.04 0.110   0.09 0.05 0.75

0.01 0.01 0.53

value represents the mean of 6 replicates (4 birds per replicate). Measurements were made on cecal samples collected on d 21 posthatch. SEM.

2Pooled

Downloaded from http://ps.oxfordjournals.org/ at Universite Laval on December 1, 2014

g/kg of cracked corn blended with pellets during the starter phase (0 to 18 d). These researchers also speculated that feed sorting and selection could possibly be the reason for the observed responses. It is noteworthy that such reductions in feed intake have not been reported in studies with prepelleting inclusion of whole wheat (Jones and Taylor, 2001; Svihus et al., 2004a; Wu et al., 2004). Higher relative gizzard weights observed with prepelleting inclusion of whole corn is consistent with published reports on whole wheat feeding (Jones and Taylor, 2001; Taylor and Jones, 2004; Svihus et al., 2004a). In the current study, however, gizzard weight plateaued at the corn inclusion level of 300 g/kg, suggesting gizzard growth may have a threshold beyond which development may be restricted. However, such a threshold on gizzard mass may not be necessarily associated with its grinding efficiency and functionality. This thesis is supported by the finding that whereas gizzard weight plateaued, ileal digestibilities of starch and N linearly increased with increasing whole corn inclusions. Increasing prepelleting inclusion levels of whole corn resulted in a greater proportion of coarse particles in the diets, and this would be expected to increase the contraction frequency of the gizzard to reduce the particle size before entry to the duodenum. Regardless of original feed structure size, particles need to be ground to a critical size before they can leave the gizzard (Svi-

PREPELLETING INCLUSION OF WHOLE CORN

In the present study, the relative weight of jejunum, and the relative lengths of jejunum and ileum increased linearly with increasing inclusion levels of whole corn. Published data on the effects of whole wheat feeding on development of digestive tract segments and organs, other than of the gizzard, are contradictory (Jones and Taylor, 2001; Banfield et al., 2002; Gabriel et al., 2003; Engberg et al., 2004; Wu and Ravindran, 2004; Ravindran et al., 2006; Amerah and Ravindran, 2008; Gabriel et al., 2008). In broilers fed whole wheat, Amerah and Ravindran (2008) reported an increase in the relative lengths of jejunum and ileum, but Gabriel et al. (2008) reported a decrease in the length of the jejunum. The reduction in the relative weight of the liver with inclusion of whole corn is in agreement with prepelleting inclusion of whole wheat (Wu and Ravindran, 2004), but no plausible explanation can be provided for these observations. It is not clear why the effect on relative length, but not weight, of the small intestine was markedly greater at 600 g/kg of whole corn inclusion compared with lower inclusion levels. Amerah and Ravindran (2008) similarly reported increases in the relative length of all intestinal segments in broilers fed whole wheat in a free choice feeding system compared with those fed ground wheat. In contrast, most studies where ground wheat was substituted with whole wheat showed no changes in the relative weight and length of intestinal segments (Preston et al., 2000; Jones and Taylor, 2001; Banfield et al., 2002; Engberg et al., 2004; Wu et al., 2004; Ravindran et al., 2006). In summary, it was hypothesized that prepelleting inclusion of whole corn could overcome its kernel size limitation. However, despite increases in gizzard weight and nutrient utilization, this strategy resulted in lower weight gain in broilers due largely to reductions in feed intake. Selective ingestion of whole corn particles appears to be the main reason for the observed intake reductions. Increased gizzard weight with the prepelleting inclusion of whole corn had a positive effect on the profile of cecal microbiota, highlighting its barrier function in preventing the entry of pathogenic bacteria into the distal digestive tract. Further studies are warranted to better understand the exact mechanisms driving the changes in microbiota profile with whole grain feeding.

REFERENCES Abdollahi, M. R., V. Ravindran, and B. Svihus. 2013. Pelleting of broiler diets: An overview with emphasis on pellet quality and nutritional value. Anim. Feed Sci. Technol. 179:1–23. Abdollahi, M. R., V. Ravindran, T. Wester, G. Ravindran, and D. V. Thomas. 2011. Influence of feed form and conditioning temperature on performance, apparent metabolisable energy and ileal digestibility of starch and nitrogen in broiler starters fed wheat-based diet. Anim. Feed Sci. Technol. 168:88–99. Abramoff, M. D., P. J. Magelhaes, and S. J. Ram. 2004. Image processing with ImageJ. Biophoto Int. 11:36–42. Amerah, A. M., and V. Ravindran. 2008. Influence of method of whole-wheat feeding on the performance, digestive tract development and carcass traits of broiler chickens. Anim. Feed Sci. Technol. 147:326–339.

Downloaded from http://ps.oxfordjournals.org/ at Universite Laval on December 1, 2014

weight of the proventriculus (Jones and Taylor, 2001; Taylor and Jones, 2004) or had no effect (Banfield et al., 2002; Bennett et al., 2002; Gabriel et al., 2003; Wu and Ravindran, 2004; Ravindran et al., 2006; Amerah and Ravindran, 2008) compared with those fed ground wheat. The increase in proventricular weight in birds fed whole corn diets was not associated with proventricular hypertrophy and the reasons for this finding are not clear. Gizzard pH data in the current work indicate that gastric hydrochloric acid secretion, and thus gastric function, was stimulated by the inclusion of whole corn. The observed increase in proventricular weight may be a further indication of enhanced gastric function with whole corn feeding. Similar decreases in gizzard pH have been reported in broilers fed coarsely ground compared with finely ground corn (Nir et al., 1994) and those fed whole wheat versus ground wheat diets (Gabriel et al., 2003). The more acidic environment in the gizzard may be responsible, at least in part, for the observed linear decreases in the populations of less preferred bacteria, namely Clostridium spp., Campylobacter spp., and Bacteroides spp., with an increase in the inclusion levels of whole corn. Furthermore, the increase in the numbers of beneficial Lactobacillus spp. may also have decreased the numbers of harmful bacteria through competitive exclusion. It appears that whole grain inclusion in diets encourages the colonization of commensal bacteria and discourages the colonization of less preferred bacteria through competitive exclusion, increased hydrochloric acid secretion, increased gizzard functionality, normal gut motility, enhanced digestive efficiency, and the resultant changes in substrate availability in the distal intestinal tract or a combination of these. The gizzard thus has an important function as a barrier organ that prevents pathogenic bacteria from entering the distal digestive tract (Engberg et al., 2004; Bjerrum et al., 2005; Santos et al., 2008). These results are in agreement to those of Engberg et al. (2002) who reported that feeding whole wheat resulted in a more acidic gizzard environment and greater numbers of Lactobacillus spp. and fewer less preferred bacteria such as Escherichia coli. But it must be noted that not all Lactobacillus strains are beneficial to the bird (De Smet et al., 1995). The numbers of bacteria determined in the current study are lower than those reported in previous studies. For example, using PCR, Bjerrum et al. (2006) determined cecal lactobacilli counts to be approximately 109, compared with the 107 to 108 determined in the current work. It is possible that the FISH method is not sensitive enough to detect minor bacterial groups. Moreno et al. (2001) evaluated the use of PCR and FISH techniques for the detection of thermotolerant campylobacters in naturally contaminated chicken products and reported that the FISH was less sensitive than PCR. The absence of more detailed analysis of the genus members is a limitation of the FISH method.

3081

3082

Singh et al. α-amylase method: Collaborative study. J. AOAC Int. 80:571– 579. Molan, A. L., J. Flanagan, W. Wei, and P. J. Moughan. 2009. Selenium-containing green tea has higher antioxidant and prebiotic activities than regular green tea. Food Chem. 114:829–835. Moreno, Y., M. Hernandez, M. A. Ferrus, J. L. Alonso, S. Botella, R. Montes, and J. Hernandez. 2001. Direct detection of thermotolerant campylobacters in chicken products by PCR and in situ hybridization. Res. Microbiol. 152:577–582. Nir, I., R. Hillel, I. Pitchi, and G. Shefet. 1995. Effect of particle size on performance. 3. Grinding pelleting interactions. Poult. Sci. 74:771–783. Nir, I., R. Hillel, G. Shefet, and Z. Nitsan. 1994. Effect of grain particle size on performance. 2. Grain texture interactions. Poult. Sci. 73:781–791. Plavnik, I., B. Macovsky, and D. Sklan. 2002. Effect of feeding whole wheat on performance of broiler chickens. Anim. Feed Sci. Technol. 96:229–236. Preston, C., K. McCracken, and A. McAllister. 2000. Effect of diet form and enzyme supplementation on growth, efficiency and energy utilisation of wheat-based diets for broilers. Br. Poult. Sci. 41:324–331. Proudfoot, F. G., and A. E. Sefton. 1978. Feed texture and light treatment effects on the performance of chicken broilers. Poult. Sci. 57:408–416. Ravindran, V., Y. B. Wu, D. G. Thomas, and P. C. H. Morel. 2006. Influence of whole wheat feeding on the development of gastrointestinal tract and performance of broiler chickens. Aust. J. Agric. Res. 57:21–26. Ross. 2007. Ross 308 Broiler: Nutrition specifications, June 2007. Ross Breeders Limited, Newbridge, Midlothian, Scotland, UK. Santos, F. B. O., B. W. Sheldon, A. A. Santos, and P. R. Ferket. 2008. Influence of housing system, grain type, and particle size on Salmonella colonization and shedding of broilers fed triticale or corn-soybean meal diets. Poult. Sci. 87:405–420. SAS Institute Inc. 2004. SAS® Qualification Tool User’s Guide. Version 9.12. SAS Inst. Inc., Cary, NC. Short, F. J., P. Gorton, J. Wiseman, and K. N. Boorman. 1996. Determination of titanium dioxide added as an inert marker in chicken digestibility studies. Anim. Feed Sci. Technol. 59:215– 221. Singh, Y., A. M. Amerah, and V. Ravindran. 2014. Whole grain feeding: Methodologies and effects on performance, digestive tract development and nutrient utilisation of poultry. Anim. Feed Sci. Technol. 190:1–18. Svihus, B. 2011. The gizzard: Function, influence of diet structure and effects on nutrient availability. World’s Poult. Sci. J. 67:207–224. Svihus, B., E. Juvik, H. Hetland, and Å. Krogdahl. 2004a. Causes for improvement in nutritive value of broiler chicken diets with whole wheat instead of ground wheat. Br. Poult. Sci. 45:55–60. Svihus, B., K. Kløvstad, V. Perez, O. Zimonja, S. Sahlström, R. B. Schüller, W. K. Jeksrud, and E. Prestløkken. 2004b. Physical and nutritional effects of pelleting of broiler chicken diets made from wheat ground to different coarsenesses by the use of roller mill and hammer mill. Anim. Feed Sci. Technol. 117:281–293. Taylor, R. D., and G. P. D. Jones. 2004. The incorporation of whole grain into pelleted broiler chicken diets. II. Gastrointestinal and digesta characteristics. Br. Poult. Sci. 45:237–246. Wu, Y. B., and V. Ravindran. 2004. Influence of whole wheat inclusion and xylanase supplementation on the performance, digestive tract measurements and carcass characteristics of broiler chickens. Anim. Feed Sci. Technol. 116:129–139. Wu, Y. B., V. Ravindran, D. G. Thomas, M. J. Birtles, and W. H. Hendriks. 2004. Influence of method of whole wheat inclusion and xylanase supplementation on the performance, apparent metabolisable energy, digestive tract measurements and gut morphology of broilers. Br. Poult. Sci. 45:385–394.

Downloaded from http://ps.oxfordjournals.org/ at Universite Laval on December 1, 2014

AOAC International. 2005. Official Methods of Analysis. 18th ed. Assoc. Off. Anal. Chem., Washington, DC. Banfield, M., R. Kwakkel, and J. Forbes. 2002. Effects of wheat structure and viscosity on coccidiosis in broiler chickens. Anim. Feed Sci. Technol. 98:37–48. Bennett, C. D., H. L. Classen, and C. Riddell. 2002. Feeding broiler chickens wheat and barley diets containing whole, ground and pelleted grain. Poult. Sci. 81:995–1003. Bjerrum, L., R. M. Engberg, T. D. Leser, B. B. Jensen, K. Finster, and K. Pedersen. 2006. Microbial community composition of the ileum and cecum of broiler chickens as revealed by molecular and culture-based techniques. Poult. Sci. 85:1151–1164. Bjerrum, L., K. Pedersen, and R. M. Engberg. 2005. The influence of whole wheat feeding on Salmonella infection and gut flora composition in broilers. Avian Dis. 49:9–15. Calet, C. 1965. The relative value of pellets versus mash and grain in poultry nutrition. World’s Poult. Sci. J. 21:23–52. Clark, P. M., K. C. Behnke, and A. C. Fahrenholz. 2009. Effects of feeding cracked corn and concentrate protein pellets on broiler growth performance. J. Appl. Poult. Res. 18:259–268. De Smet, I., L. Van Hoorde, M. Vande Woestyne, H. Christiaens, and W. Verstraete. 1995. Significance of bile salt hydrolytic activities of lactobacilli. J. Appl. Bacteriol. 79:292–301. Dinoto, A., A. Suksomcheep, S. Ishizuka, H. Kimura, S. Hanada, Y. Kamagata, K. Asano, F. Tomita, and A. Yokota. 2006. Modulation of rat cecal microbiota by administration of raffinose and encapsulated Bifidobacterium breve. Appl. Environ. Microbiol. 72:784–792. Duke, G. E. 1992. Recent studies on regulation of gastric motility in turkeys. Poult. Sci. 71:1–8. Engberg, R. M., M. S. Hedemann, and B. B. Jensen. 2002. The influence of grinding and pelleting of feed on the microbial composition and activity in the digestive tract of broiler chickens. Br. Poult. Sci. 43:569–579. Engberg, R. M., M. S. Hedemann, S. Steenfeldt, and B. B. Jensen. 2004. Influence of whole wheat and xylanase on broiler performance and microbial composition and activity in the digestive tract. Poult. Sci. 83:925–938. Gabriel, I., S. Mallet, and M. Leconte. 2003. Differences in the digestive tract characteristics of broiler chickens fed on complete pelleted diet or on whole wheat added to pelleted protein concentrate. Br. Poult. Sci. 44:283–290. Gabriel, I., S. Mallet, M. Leconte, A. Travel, and J. P. Lalles. 2008. Effects of whole wheat feeding on the development of the digestive tract of broiler chickens. Anim. Feed Sci. Technol. 142:144– 162. German, D. P., and R. A. Bittong. 2009. Digestive enzyme activities and gastrointestinal fermentation in wood-eating catfishes. J. Comp. Physiol. B Biochem. Syst. Environ. Physiol. 179:1025– 1042. Hetland, H., B. Svihus, and V. Olaisen. 2002. Effect of feeding whole cereals on performance, starch digestibility and duodenal particle size distribution in broiler chickens. Br. Poult. Sci. 43:416–423. Jones, G. P. D., and R. D. Taylor. 2001. The incorporation of whole grain into pelleted broiler chicken diets: Production and physiological responses. Br. Poult. Sci. 42:477–483. Lentle, R. G., V. Ravindran, G. Ravindran, and D. V. Thomas. 2006. Influence of feed particle size on the efficiency of broiler chickens fed wheat-based diets. Jpn. Poult. Sci. 43:135–142. Li, P. X. P., A. K. Hardacre, O. H. Campanella, and K. J. Kirkpatrick. 1996. Determination of endosperm characteristics of 38 corn hybrids using the Stenvert hardness test. Cereal Chem. 73:466–471. Li, Y., and C. Owyang. 1993. Vagal afferent pathway mediates physiological action of cholecystokinin on pancreatic enzyme secretion. J. Clin. Invest. 92:418–424. Mateos, G. G., R. Lazaro, and M. L. Gracia. 2002. The feasibility of using nutritional modifications to replace drugs in poultry feeds. J. Appl. Poult. Res. 11:437–452. McCleary, B. V., T. S. Gibson, and D. C. Mugford. 1997. Measurement of total starch in cereal products by amyloglucosidase vs.