Insoluble fibre and enzyme supplementation in mash or pellets diets on growth performance, apparent ileal digestibility and intestinal morphology of broiler chickens fed barley containing diets

Abstract

Background

Enzyme supplementation and the inclusion of fibre in the barley‐based diets have been some of the alternatives proposed to improve productivity in the absence of growth promoters.

Objective

This study was performed to investigate the effect of adding sunflower hulls (SFH), a multi‐enzyme carbohydrate, and feed forms (mash and pellet) on performance and some physiological parameters in broiler chickens fed barley containing diets.

Methods

Treatments were two feed forms (mash vs. pelleted), and four diets consisted of a barley‐based diet (control, CTL) or test diets which contained either SFH at 30 g/kg, enzyme (ENZ; 0.2 g/kg) or combination of SFH and enzyme (SFH + ENZ).

Results

The results showed that average daily feed intake and average daily gain were significantly increased in chickens that were fed ENZ (p < 0.05). The highest digestibility of ether extract (EE) was observed in the treatment containing SFH and SFH + ENZ (p < 0.05). The highest population of Lactobacillus spp. was observed in the treatment containing SFH (p < 0.05). The villus height and villus height to crypt depth ratios of duodenum and jejunum were significantly higher (p < 0.05) in broilers fed pellet diets compared to the mash.

Conclusion

It can be concluded that pellet diets reduce digesta viscosity and harmful microorganisms (Escherichia coli), increase growth performance, and improve intestinal morphology in barley‐based diets. Moreover, SFH and ENZ had favourable effects on EE digestibility and caecal microbial population of broilers fed with barley containing diets.

This experiment was conducted to evaluate two feed forms (mash vs. pelleted), and four diets consisted of a barley‐based diet (control, CTL) or test diets which contained either sunflower hulls at 30 g/kg (SFH), enzyme (ENZ; 0.2 g/kg) or combination of SFH and enzyme (SFH + ENZ). Pellet diets reduced digesta viscosity and harmful microorganisms (E. coli), increased growth performance, and improved intestinal morphology in barley‐based diets. Moreover, SFH and ENZ had favourable effects on EE digestibility and caecal microbial population of broilers fed with barley containing diets.

1. INTRODUCTION

It has been shown attendance of soluble nonstarch polysaccharides (NSP), such as β‐glucans and arabinoxylans, in barley, wheat and rye grains may affect nutrient use adversely and reduction growth performance of broiler chickens. The Anti‐nutritional effects of soluble NSP are mainly ascribable to an increased viscosity of digesta associated to higher levels of microbial populations in the intestinal tract. This results in decreased digestibility of nutrients, increased competition for nutrients, and alteration in the structural morphology of the intestinal wall. The use of highly digestible ingredients (Bedford, 2000; Dahiya et al., 2006), enzyme supplementation (Lázaro et al., 2003a), heat processing (HP) of the cereal (Gracia et al., 2003) and the inclusion of fibre in the diet (Jørgensen et al., 1996; Rogel et al., 1987) have been some of the alternatives proposed to improve productivity in the absence of growth promoters.

On the other hand, the use of enzyme supplementation in the diet containing non‐starch polysaccharides reduces intestinal viscosity, increases the growth rate and improves feed conversion efficiency (Lázaro et al., 2003a), as a result increasing broiler productivity. Ouhida et al. (2000) and Meng and Slominski (2005) showed in their experiments that adding appropriate exogenous enzymes such as xylanases and β‐glucanases to broiler diets containing wheat or barley, or both reduces intestinal viscosity and improves digestibility and performance.

The form of feed is considered a very important factor in improving the performance of broiler chickens (Abdollahi et al., 2013a; Mateos et al., 2002). Many researchers have reported that broilers fed pellet diets had higher weight gain and better feed conversion ratio (FCR) than birds fed meal diets (Chewning et al., 2012; Mckinney & Teeter, 2004). However, fine grinding of ingredients for pelleting has become the standard practice in feed manufacture. Therefore, the lack of structural components in pellet diets may be lead to suboptimal functionality of the foregut, followed by poor nutrient digestibility and increased consumption of litter, leading to poor intestinal health (Rodrigues & Choct, 2018; Svihus, 2011). This concern has increased the interest to inclusion of insoluble fibre to the diet (Hetland et al., 2004). Insoluble fibre can be incorporated up to a certain level because of possible nutrient dilution (Rodrigues & Choct, 2018; Singh et al., 2014). It was shown that dietary inclusion of insoluble fibre improved growth performance, increased apparent ileal digestibility of nutrients, and decreased ileal viscosity in the birds fed diets containing barley (Pourazadi et al., 2020). Moreover, Gonzalez‐Alvarado et al. (2007) reported that the use of 3% of oat hulls (OH) or soybean hulls in the feed of broiler chickens improves their performance at the age of 1–21 days and the digestibility of nutrients at 18 of age. Therefore, the present study was performed to investigate the addition of insoluble fibre and enzymes, along with the physical form of feed, on growth performance, apparent ileal digestibility and intestinal morphology.

2. MATERIALS AND METHODS

2.1. Chemical analysis of insoluble fibre and enzyme supplementation

The sunflower hulls (SFH) were obtained from Maz Maz factory and were ground using a hammer mill equipped with a 3 mm screen. The chemical composition of SFH was analysed according to Association of Official Analytical Chemist (AOAC) (2000) methods (Table 1). Crude fibre (CF) was measured by sequential extraction with diluted acid and alkali (method 978.10; AOAC, 2000). Dry matter (DM) and crude protein (CP) were determined using methods 930.15 and 990.03, respectively (AOAC, 2000). Ether extract (EE) was analysed as EE by the Soxhlet method after acid hydrolysis (method 954.02; AOAC, 2000). Neutral (NDF) and acid detergent fibre (ADF) of samples were determined sequentially as described by Van Soest et al. (1991) and expressed on an ash‐free basis. The moisture and ash contents were determined by methods reported by Debon and Tester (2001). Non‐fibre carbohydrates (NFC) was calculated from the following formula:

$$\mathrm{NFC}=100-\left(\mathrm{CP}+\mathrm{ash}+\mathrm{EE}+\mathrm{NDF}\right).$$

TABLE 1.

Chemical composition (g/kg) and particle size distribution of sunflower hulls.

Parameter	g/kg
Dry matter	923.0
Ash	30.3
Crude protein	48.0
Ether extract	46.0
Crude fibre	484.0
NDF	693.0
ADF	468.0
NFC	183.0
NFE	392.0

Screen size (μm)
3000	0.63
2000	3.24
1000	12.69
500	74.18
212	5.25
125	1.57
75	2.44
GMD ± GSD	729.72 ± 1.73

Note: NFC = 100 − (CP + Ash + EE + NDF); NFE = 100 −(CP + Ash + CF + EE).

Abbreviations: ADF; acid detergent fibre; GMD, geometric mean diameter; GSD, geometric standard deviation; NDF, neutral detergent fibre; NFC, non‐fibre‐carbohydrate; NFE, nitrogen free extract.

Nitrogen free extract (NFE) was calculated by

$$\mathrm{NFE}=100-\left(\mathrm{CP}+\mathrm{ash}+\mathrm{CF}+\mathrm{EF}\right).$$

The dry sieving method was used to determine the particle size distribution of SFH following the method described by Baker and Herrman (2002). A set of seven sieves (Endecott), sized 3000, 2000, 1000, 500, 212, 125 and 75 μm, with an Endecott testing sieve shaker, were used to separate feed particles into different size fractions. The geometric mean diameter and geometric standard deviation were determined according to the American Society of Agricultural Engineers (2003).

A multicomponent NSP‐degrading enzyme, Endo‐Power was obtained from Easy Bio. The activities of α‐galactosidase, galactomannanase, xylanase and β‐glucanase enzymes were 35, 110, 1500 and 1100 U/g, respectively. One unit of galactosidase is defined as enzyme that hydrolyzes 1 μmol of ρNPG within 1 min at 30°C and pH 4. One unit of galactomannanase is defined as enzyme activity that releases 1 μmol of reducing sugar within 1 min at 40°C and pH 4. One unit of xylanase and β‐glucanase is defined as enzyme activity that releases 1 μmol of reducing sugar within 1 min at 30°C and pH 4. The Endo‐Power was thermostable enzyme.

2.2. Birds, diets and management

A total of 320 1‐day‐old male broiler chicks (Ross 308) were obtained from a commercial hatchery in Iran. Birds were randomly allocated to 8 treatments with 5 replicate pens of 8 birds each. The experimental design was completely randomized with 8 treatments arranged as a 2 × 4 factorial. Treatments were two feed forms (mash vs. pelleted) and four diets consisted of a barley‐based diet (control, CTL) or test diets which contained either SFH (30 g/kg), enzyme (0.2 g/kg) or enzyme (0.2 g/kg) plus SFH (30 g/kg). The basal diet included 30 g/kg fine silica sand that was replaced by the same amount of SFH in the treatment diets. All the diets were formulated to meet the nutrient requirements of broilers according to National Research Council (NRC) (1994) across different growing periods and fed in mash and pellet form (Table 2). The SFH was first ground with a hammer mill provided with a 3‐mm screen and then as well as the enzyme was added to the experimental diets. The resulting batches of these 4 diets were divided into two equal portions; the first portion was fed as such, whereas the second portion was steam conditioned first and then passed through a pellet press equipped with a 2‐mm die and an effective thickness of 35 mm for starter period (1–21days) and 4‐mm die and an effective thickness of 60 mm for grower period (22–42 days). The temperature of the feed at the exit of the pellet press was 79 ± 3°C. Chicks were reared until 42 days of age in floor pens. Clean wood shavings were used as bedding material. Each pen contained one suspended drinker and one feeder. Feed and water were provided ad libitum throughout the trial.

TABLE 2.

Ingredients and chemical composition of the experimental diets (g/kg dry matter [DM]).

Ingredients	Starter	Grower
	Day 1–21	Day 22–42
Maize	248.0	295.7
Barley	300.0	300.0
Soybean meal (440 g CP/kg)	302.7	256.0
Fish meal (605 g CP/kg)	30.0	20.0
Sunflower oil	50.0	65.0
Limestone	13.5	14.0
Dicalcium phosphate	12.0	8.5
Sodium chloride	3.0	2.0
Sodium bicarbonate	1.8	1.8
Mineral and Vitamin premix a	2.5	2.5
dl‐Methionine	2.5	1.2
l‐Lysine HCl	1.5	0.8
Silica sand b	30.0	30.0
Calculated analysis
AME (MJ/kg)	12.22	12.80
Crude protein	210.2	191.5
Calcium	10.2	90.0
Available phosphorus	4.5	3.5
Sodium	2.0	1.5
Arginine	14.4	12.6
Lysine	14	11.6
Methionine	5.8	4.2
Methionine + Cysteine	9.3	7.3
Determined analysis
DM	900.0	898.0
Crude protein	211.8	192.9
Crude fibre	42.20	39.90
Ether extract	70.1	85.6

Abbreviation: CP, crude protein.

^a Provided the following (per kg of diet): Fe, 60 mg; Mn, 100 mg; Zn, 60 mg; Cu, 10 mg; I, 1 mg; Co, 0.2 mg; Se, 0.15 mg; retinyl acetate, 1.55 mg; cholecalciferol, 0.025 mg; α‐tocopherol acetate, 20 mg; menadione, 1.3 mg; thiamine, 2.2 mg; riboflavin, 10 mg; calcium pantothenate, 10 mg; choline chloride, 400 mg; nicotinamide, 50 mg; pyridoxine HCl, 4 mg; biotin, 0.04 mg; folic acid, 1 mg; vitamin B12 (cobalamin), 1.013 mg.

^b Silica sand was replaced by 30.0 g/kg sunflower hulls in starter and grower phases of experiment.

2.3. Growth performance

Average daily gain (ADG) and average daily feed intake (ADFI) for each pen were recorded. FCR was calculated by dividing ADFI with ADG for each period of the experiment (starter: 1–21 days and grower: 22–42 days) and total period (1–42 days). Birds that died during the experiment were weighed, and the data were used in the calculation of FCR.

2.4. Size of different organs and length of intestine

At the end of the experiment, two birds from each replicate were randomly selected and killed by cervical dislocation to evaluate the weight (relative to BW) of the breast, thigh, gastrointestinal tract, gizzard, proventriculus, pancreas, liver and abdominal fat. The length of intestinal segments, including the duodenum from the pylorus to the distal portion of the duodenal loop, the jejunum as the segment between the point of entry of the bile ducts and Meckel's diverticulum, the ileum from Meckel's diverticulum to the ileocaecal junction, and caecum (left and right), were measured separately (Sadeghi et al., 2015).

2.5. Apparent ileal digestibility of nutrients and gut microflora

Chromium oxide (Cr₂O₃) was added on top to the complete feed at a rate of 3 g/kg for nutrient digestibility assessment. Marker‐containing diets were given for five consecutive days, from day 23–28 of the experimental period. At day 28, two birds per replicate were killed by cervical dislocation and the ileal contents, from Meckel's diverticulum to the ileocaecal junction, were collected by gently squeezing the contents into sealed bags. The samples were kept frozen at −20°C until analysis. Feed samples and digesta were ground (using a 0.5 mm screen) prior to chemical analysis. The samples were analysed for organic matter (OM), CP and EE according to the standard procedures of AOAC (2000), as above. All values were expressed on a DM basis. Chromium oxide content in the experimental diets and digesta was measured according to the method published by Saha and Gilbreath (1991). Digestibility coefficients were calculated using the following formula (Hafeez et al., 2016):

$$\mathrm{Apparent}\mathrm{digestibility}=1-\left[\left(\frac{{\mathrm{Cr}}_2{\mathrm{O}}_3\mathrm{diet}}{{\mathrm{Cr}}_2{\mathrm{O}}_3\mathrm{digesta}}\right)\times \left(\frac{\mathrm{nutrient}\mathrm{in}\mathrm{digesta}}{\mathrm{nutrient}\mathrm{in}\mathrm{diet}}\right)\right]$$

On day 28, to measure the caecal microflora, one caecum from two birds in each replicate, which had been euthanized by cervical dislocation for ileal collection, was used. The caecal contents were aseptically collected in individual sterile culture tubes, kept on ice, and immediately transferred to the microbiological lab. Caecal microflora colonies were measured as described by Masouri et al. (2017).

2.6. pH of gizzard and ileal digesta viscosity

On day 42, two birds were randomly selected from each pen, weighed, and euthanized by cervical dislocation. Then, the pH of the contents from gizzard was measured using a digital pH meter (model 507, Crison Instruments S. A.). Gizzard opened by an incision and split longitudinally and pH was immediately recorded by inserting the pH meter directly into the digesta (Singh et al., 2014). On day 42, a further two birds per replicate (i.e. eight birds per treatment) were randomly selected and euthanized by cervical dislocation. The small intestine was removed, and the digesta from the ileum (from Meckel's diverticulum to the ileo‐caecal junction) was immediately collected. The samples were placed into clean tubes and centrifuged at 9000 × g at room temperature (4°C) for 10 min. The supernatant was withdrawn and viscosity (in centipoise, cps = 1/100 dyn s/cm²) was determined using a Brookfield digital viscometer (Model LVDVII, Brookfield Engineering Labs, Inc.; Jamroz et al., 2003).

2.7. Intestinal morphology

At the end of 42 days, the duodenum and jejunum segments were collected and analysed according to Liu et al (2022). Briefly, 1 cm sections from the middle portion of the duodenum and jejunum tissues were fixed in 10% formaldehyde phosphate buffer after washed with 0.1 M phosphate buffered saline, and the fixed sections were processed, dehydrated, embedded in paraffin wax, sectioned at 5 mm, and stained with the haematoxylin–eosin. Histological sections were examined with villus height (VH) and crypt depth (CD), which were performed on 10 well‐oriented villi chosen using a digital camera microscope (BA400 Digital, McAudi Industrial Group Co., Ltd.) and the Motic Advanced 3.2 digital image analysis system. The ratio of villus height to crypt depth (VH/CD) was calculated subsequently.

2.8. Statistical analysis

All data were analysed as a completely randomized design with eight treatments arranged as a 2 × 4 factorial with two feed forms (mash vs. pelleted) and four diets (barley‐based diet, control and CTL) or test diets which contained either SFH (30 g/kg), enzyme (0.2 g/kg) or enzyme (0.2 g/kg) plus SFH (30 g/kg) using the PROC GLM of SAS (SAS Institute, 2008). The experimental unit was the pen for all traits. Microbiological counts were subjected to base‐10 logarithm transformation before analysis. Significant differences among the treatments were determined using Tukey's test at p < 0.05.

3. RESULTS

3.1. Growth performance

The impact of dietary treatments on bird performance at each dietary phase (1–21, 22–42 and 1–42 days of age) is presented in Table 3. Mortality during the performance experiment was negligible and not related to any dietary treatment. There was no significant interaction between type of additive and feed form throughout the experiment on growth performance. The ADFI and ADG of chickens were significantly affected by the type of additives at 1–21 days of age (p < 0.05). ADFI and ADG were significantly increased in chickens fed diets containing enzymes additive (p < 0.05). Chickens fed with diet containing SFH had the lowest ADFI compared to the other treatments at 1–21 days of age (p < 0.05). The effect of feed form (pellets and mash) on ADG, ADFI and FCR ratio was significant (p < 0.05) and ADFI in chickens fed with pelleted diets compared to mash increased (p < 0.05) at 1–21 days of age and at 1–42 days of age. Chickens fed with pelleted diets gained more weight in the starter, grower and total period of experiment and improved FCR at 1–42 days of age (p < 0.05).

TABLE 3.

Effect of experimental treatments on growth performance of broiler chickens.

Additive type	Feed form	ADFI (g/day)			ADG (g/day)			FCR
		1–21 days	22–42 days	1–42 days	1–21 days	22–42 days	1–42 days	1–21 days	22–42 days	1–42 days
CTL	Mash	43.62	162	103	29.63	78.36	53.99	1.472	2.067	1.907
SFH		42.93	156	99	30.02	78.30	54.16	1.430	1.992	1.840
ENZ		45.96	157	101	32.05	82.12	57.09	1.434	1.911	1.769
SFH + ENZ		45.15	153	99	30.52	78.05	54.28	1.479	1.960	1.829
CTL	Pellet	51.68	155	103	35.65	84.13	59.89	1.449	1.842	1.719
SFH		48.27	160	104	33.79	82.20	57.99	1.428	1.946	1.793
ENZ		52.20	158	105	38.16	81.94	60.05	1.367	1.928	1.748
SFH + ENZ		46.62	159	102	33.74	85.71	59.73	1.381	1.855	1.707
SEM		1.17	3.31	1.82	0.74	2.21	1.19	0.03	0.07	0.05
Additive type
CTL		47.65ab	159	103	32.64b	81.25	56.94	1.459	1.956	1.808
SFH		45.60b	158	102	31.90b	80.25	56.08	1.429	1.968	1.818
ENZ		49.08a	157	103	35.10a	82.03	58.57	1.398	1.913	1.758
SFH + ENZ		45.89b	156	101	32.13b	81.88	57.00	1.428	1.905	1.771
SEM		0.83	2.34	1.29	0.52	1.56	0.84	0.02	0.05	0.03
Feed form
Mash		44.42b	157	100b	30.55b	79.21b	54.88b	1.454	1.982	1.822a
Pellet		49.69a	158	104a	35.33a	83.49a	59.41a	1.406	1.892	1.750b
SEM		0.58	1.65	0.91	0.37	1.10	0.59	0.01	0.03	0.02
p‐Value
Additive type		0.02	0.85	0.55	0.0008	0.84	0.24	0.40	0.73	0.57
Feed form		<0.0001	0.65	0.02	<0.0001	0.01	<0.0001	0.06	0.08	0.02
Additive type × feed form		0.06	0.28	0.73	0.134	0.35	0.58	0.49	0.37	0.45

Note: Means with different letters (a, b) within the same column differ significantly (p < 0.05).

Abbreviations: ADFI, average daily feed intake; ADG, average daily gain; ENZ, endo‐power enzyme; FCR, feed conversion ratio; SFH, sunflower hulls.

3.2. Carcass and digestive trait characteristics

The effect of experimental treatments on carcass and digestive traits is shown in Table 4. The relative weight of carcass, breast, thigh, proventriculus, liver, pancrease and abdominal fat of chickens were not affected by the type of additive and the interaction between the type of additive and the feed form (p > 0.05). On the other hand, the relative weight of gizzard and gastrointestinal tract significantly reduced in pelleted diets compared to mash (p < 0.05), and other organs were not affected by the feed form (p > 0.05). Moreover, the relative weight of the duodenum, jejunum, ileum and ceca, as well as their length, were not affected by the experimental treatments (Table 5) (p > 0.05).

TABLE 4.

Effect of experimental treatments on carcass traits of broiler chickens on d 42 (g/100 g BW).

		Carcass traits
Additive type	Feed form	Carcass	Breast	Thigh	Gastrointestinal tract	Proventriculus	Gizzard	Liver	Pancreas	Abdominal fat
CTL	Mash	73.26	25.23	21.11	11.89	0.44	1.97	2.00	0.30	1.09
SFH		72.60	24.59	20.48	13.25	0.47	2.57	2.47	0.29	1.32
ENZ		70.87	24.65	20.44	12.34	0.35	1.97	2.04	0.35	1.16
SFH + ENZ		74.40	25.03	21.21	13.02	0.54	2.18	1.95	0.37	1.32
CTL	Pellet	74.71	26.24	20.75	11.10	0.44	1.84	1.98	0.37	1.39
SFH		74.83	24.07	20.62	11.68	0.42	1.67	1.76	0.26	1.36
ENZ		72.98	21.99	20.19	11.58	0.51	1.65	1.78	0.32	1.13
SFH + ENZ			25.25	21.08	12.13	0.46	2.16	2.07	0.31	1.35
SEM		1.03	1.34	0.63	0.66	0.05	0.20	0.16	0.06	0.19
Main effects
Additive type
CTL		74.00	25.74	20.93	11.50	0.44	1.91	1.9	0.33	1.24
SFH		73.72	24.33	20.55	12.47	0.44	2.12	2.1	0.28	1.34
ENZ		71.90	23.32	20.31	11.96	0.43	1.81	1.9	0.33	1.14
SFH + ENZ		74.22	25.14	21.15	12.58	0.50	2.17	2.0	0.34	1.34
SEM		0.73	0.94	0.45	0.46	0.04	0.14	0.12	0.04	0.13
Feed form
Mash		72.80	24.87	20.81	12.63a	0.45	2.17a	2.11	0.33	1.22
Pellet		74.11	24.39	20.66	11.62b	0.46	1.83b	1.90	0.32	1.31
SEM		0.51	0.67	0.31	0.33	0.02	0.10	0.08	0.03	0.09
p‐Value
Additive type		0.12	0.32	0.56	0.35	0.58	0.27	0.68	0.68	0.72
Feed form		0.07	0.61	0.74	0.04	0.95	0.02	0.08	0.79	0.55
Additive type × feed form		0.58	0.56	0.98	0.91	0.19	0.17	0.09	0.78	0.85

Note: Means with different letters (a, b) within the same column differ significantly (p<0.05).

Abbreviations: ENZ, endo‐power enzyme; SFH, sunflower hulls.

TABLE 5.

Effect of experimental treatments on the relative weight and length of the different parts of the intestine of broiler chickens on day 42.

Additive type	Feed form	Relative weight (% of BW)				Length (cm)
		Duodenum	Jejunum	Ileum	Cecaa	Duodenum	Jejunum	Ileum	Cecab
CTL	Mash	0.81	2.39	2.27	0.76	33.00	76.50	93.87	19.25
SFH		0.80	2.14	2.07	0.80	32.00	75.75	90.75	17.75
ENZ		0.89	2.23	2.21	0.77	32.50	72.50	91.25	18.25
SFH + ENZ		0.79	2.17	2.20	0.80	30.67	72.75	85.50	19.00
CTL	Pellet	0.78	2.04	1.79	0.76	33.00	70.25	84.50	18.00
SFH		0.83	2.11	1.95	0.84	33.62	75.50	89.75	19.25
ENZ		0.86	2.36	2.11	0.82	31.75	74.25	88.50	18.25
SFH + ENZ			2.31	2.15	0.85	31.50	75.25	87.50	18.25
SEM		0.04	0.17	0.15	0.03	1.14	3.56	2.16	0.89
Main effects
Additive type
CTL		0.79	2.21	2.03	0.76	33.00	73.37	89.18	18.62
SFH		0.81	2.12	2.01	0.82	32.81	75.62	90.25	18.50
ENZ		0.87	2.30	2.16	0.80	32.12	73.37	89.87	18.25
SFH + ENZ		0.82	2.24	2.17	0.83	31.08	74.00	86.50	18.62
SEM		0.02	0.12	0.10	0.02	0.80	2.51	1.53	0.63
Feed form
Mash		0.82	2.23	2.19	0.78	32.04	74.37	90.34	18.56
Pellet		0.83	2.20	2.00	0.82	32.46	73.81	87.56	18.43
SEM		0.02	0.08	0.07	0.01	0.57	1.78	1.08	0.44
p‐Value
Additive type		0.24	0.80	0.61	0.29	0.34	0.91	0.32	0.97
Feed form		0.89	0.81	0.09	0.20	0.60	0.82	0.08	0.84
Additive type × feed form		0.62	0.49	0.49	0.88	0.75	0.60	0.08	0.45

Abbreviations: ENZ, endo‐power enzyme; SFH, sunflower hulls.

^a Weight of the 2 ceca.

^b Length of the 1 ceca.

3.3. Digestibility of nutrients

The digestibility of OM, CP and EE was not affected by the form of feed and the interaction between the type of additive and the form of feed (Table 6) (p > 0.05). By using the additives, the highest digestibility of EE was observed in the treatment containing SFH and SFH plus enzyme (p < 0.05), and the highest digestibility of OM was observed in the treatment containing enzyme. The digestibility of CP was not affected by the type of additive (p > 0.05; Table 6).

TABLE 6.

Effect of experimental treatments on apparent ileal digestibility (%) of broiler chickens on day 28.

Additive type	Feed form	Crude protein	Ether extract	Organic matter
CTL	Mash	65.44	43.86	68.14
SFH		69.81	50.71	68.41
ENZ		69.77	45.77	71.32
SFH + ENZ		69.37	47.10	66.97
CTL	Pellet	68.63	44.18	68.67
SFH		68.57	48.09	70.01
ENZ		70.48	49.52	74.89
SFH + ENZ		69.05	48.63	68.31
SEM		1.13	1.57	1.63
Main effects
Additive type
CTL		67.03	44.02b	68.40b
SFH		69.19	49.40a	69.21b
ENZ		70.13	47.65ab	73.10a
SFH + ENZ		69.21	47.86a	67.64b
SEM		0.80	1.24	1.15
Feed form
Mash		69.18	46.86	70.47
Pellet		68.60	47.61	68.71
SEM		0.56	0.87	0.81
p‐Value
Additive type		0.06	0.03	0.01
Feed form		0.47	0.55	0.14
Additive type × feed form		0.26	0.35	0.81

Note: Means with different letters (a, b) within the same column differ significantly (p<0.05).

Abbreviations: ENZ, endo‐power enzyme; SFH, sunflower hulls.

3.4. Caecal microbial population

There was no significant interaction between type of additive and feed form on caecal microbial population on d 42 (p > 0.05, Table 7). The total counts of aerobic bacteria and coliforms were not affected by the main effect (additive type and feed form) (p > 0.05). The population of Lactobacillus spp. was only affected by the type of additive and the highest population was observed in the treatment containing SFH (p < 0.05). Moreover, Escherichia coli population was affected by the main effect of type of additive and feed form. The E. coli population significantly decreased in treatments containing additives. As well as, pelleting the diets compared to the mash diets significantly decreased E. coli population (p < 0.05).

TABLE 7.

Effect of experimental treatments on digesta viscosity (cP1) of ileum and pH gizzard on day 42 and caecal microbial population (log cfu/g) of broiler chickens on day 28.

Additive type	Feed form	Ileum viscosity	Gizzard pH	Total aerobic bacteria	Lactobacilli spp.	Escherichia coli	Coliforms
CTL	Mash	1.96a	4.27	8.89	8.99	8.82	8.96
SFH		1.54bc	3.95	8.15	9.68	8.07	8.46
ENZ		1.68ab	4.20	9.09	9.16	7.60	8.94
SFH + ENZ		1.77ab	4.00	9.17	8.97	8.23	8.97
CTL	Pellet	1.97a	4.65	8.42	9.11	8.48	8.44
SFH		1.56b	3.95	9.04	9.39	7.63	9.05
ENZ		1.63b	4.30	8.42	8.91	7.79	9.04
SFH + ENZ		1.23c	4.10	8.98	9.16	7.15	8.55
SEM		0.06	0.16	0.33	0.16	0.27	0.25
Main effects
Additive type
CTL		1.96a	4.46a	8.66	9.05b	8.65a	8.70
SFH		1.55bc	3.95b	8.59	9.53a	7.85b	8.75
ENZ		1.65b	4.25ab	8.75	9.04b	7.70b	8.99
SFH + ENZ		1.50c	4.05b	9.07	9.07b	7.69b	8.76
SEM		0.03	0.11	0.23	0.11	0.19	0.18
Feed form
Mash		1.74a	4.25	8.83	9.20	8.18a	8.83
Pellet		1.60b	4.10	8.71	9.14	7.76b	8.77
SEM		0.04	0.08	0.16	0.08	0.13	0.13
p‐Value
Additive type		<0.0001	0.02	0.63	0.01	0.04	0.68
Feed form		0.008	0.23	0.49	0.63	0.005	0.73
Additive type × feed form		0.0009	0.70	0.11	0.35	0.18	0.14

Note: Means with different letters (a–c) within the same column differ significantly (p < 0.05).

Abbreviations: ENZ, endo‐power enzyme; SFH, sunflower hulls.

3.5. Ileum viscosity and pH of gizzard

There was a significant (p < 0.05) interaction between type of additive and feed form for ileum viscosity (Table 7). In pellet diets, dietary supplementation of SFH plus enzyme resulted in lower (p < 0.05) ileum viscosity compared to the enzyme, SFH and CTL, whereas, in mash diets, there was no effect (p > 0.05) of feed additives. There was no significant interaction between type of additive and feed form on pH of the gizzard (p > 0.05). Gizzard pH was modified by type of additives inclusion (p < 0.05), SFH decreased pH of the gizzard as compared to the CTL, but it does not have any difference with other additives (Table 7).

3.6. Intestinal morphology

The effect of experimental treatments on intestinal morphology in broiler chickens at the age of 42 d is presented in Table 8. No significant (p > 0.05) interaction between type of additives and feed form and also the main effect of feed additives were detected for any of the intestinal morphology, and therefore, only main effect of feed form was reported. In broilers fed with pellet diets, the VH and VH/CD ratios of duodenum and jejunum were significantly higher compared to the mash diets (p < 0.05).

TABLE 8.

Effect of experimental treatments on villus height (μm), crypt depth (μm) and villus height/crypt depth of the duodenum and jejunum of broiler chickens on day 42.

Additive type	Feed form	Villus height		Crypt depth		Villus height/crypt depth
		Duodenum	Jejunum	Duodenum	Jejunum	Duodenum	Jejunum
CTL	Mash	1078.62	1058.15	147.18	159.93	7.33	6.68
SFH		1141.34	1070.82	147.98	165.15	7.75	6.48
ENZ		1051.56	1044.86	143.39	174.28	7.32	6.04
SFH + ENZ		1089.44	1059.66	137.78	177.62	7.90	6.02
CTL	Pellet	1169.66	1145.19	149.38	158.41	7.83	7.33
SFH		1341.39	1186.66	141.11	167.20	9.31	7.09
ENZ		1431.10	1154.82	147.35	160.55	9.72	7.19
SFH + ENZ		1326.11	1170.82	146.84	148.81	9.02	7.88
SEM		71.44	46.66	3.41	7.70	0.47	0.39
Main effects
Additive type
CTL		1124.14	1101.67	148.28	159.17	7.58	7.00
SFH		1227.86	1128.74	144.54	166.18	8.53	6.79
ENZ		1241.33	1099.84	145.37	167.41	8.52	6.62
SFH + ENZ		1207.14	1115.24	142.31	163.21	8.46	6.95
SEM		50.52	32.99	2.41	5.44	0.33	0.28
Feed form
Mash		1090.24b	1058.37b	144.08	169.24	7.58b	6.30b
Pellet		1310.31a	1164.37a	146.17	158.74	8.97a	7.37a
SEM		35.72	23.33	1.70	3.85	0.23	0.19
p‐Value
Additive type		0.37	0.91	0.39	0.71	0.15	0.76
Feed form		0.0002	0.003	0.39	0.06	0.004	0.0009
Additive type × feed form		0.25	0.98	0.15	0.20	0.27	0.38

Note: Means with different letters (a, b) within the same column differ significantly (p < 0.05).

Abbreviations: ENZ, endo‐power enzyme; SFH, sunflower hulls.

4. DISCUSSION

4.1. Growth performance

In the present research, the increase in ADFI and ADG due to the use of enzyme additive at 1–21 days of age can be due to the reducing digesta viscosity and probably increasing the passage rate (Almirall et al., 1995), and also increasing digestion and absorption due to the effect of enzyme additive (Bedford & Scholz, 1998). On the other hand, the use of enzyme additive in the diet during the grower period (22–42 days of age) and throughout the experimental period (1–42 days of age) did not have significant effects on ADFI and ADG. This effect may be caused by improving the efficiency and ability of the digestive system of chickens in order to overcome the digestive problems of barley‐based diet with increasing age (Angelovicova et al., 2005; Tarkan et al., 2007). Although the FCR was not affected by additives in any of the age periods, but it was numerically improved in the treatment receiving the enzyme additives, which may be related to the optimal use of energy due to the increase in the digestibility of nutrients (Garcia et al., 2008; Lesson et al., 2000).

In the present experiment, the increase of BWG and the improvement of the FCR in chicks that are fed with pellet diet can be related to less activity during feed intake (McKinney & Teeter, 2004), and as a result, the energy will be spent on performance and BWG of the chicks (Skinner‐Nobel et al., 2003). Jiménez‐Moreno et al. (2016) showed that broiler chickens that were fed with pellet diet had a 32% higher BWG, and the FCR was 3% better than mash diet. In another experiment, the physical form of feed (pellet and mash) was investigated in wheat‐based diet. Feeding with wheat‐based pellet diets increased BWG and improved FCR in chickens at the age of 21 days than mash diet (Amerah et al., 2007). Moreover, Engberg et al. (2002) reported that the use of wheat‐based pellet diets compared to mash led to increase in BWG due to an increase in feed intake and an improvement in the FCR. Abdollahi et al. (2014) also stated that sorghum‐based pellet diets increased BWG and increased feed intake of chickens compared to mash diet at 21 days of age. It appears that HP during the pelleting process disrupts the cell structures and releases the encapsulated nutrients (Gracia et al., 2003; García et al., 2008), facilitating nutrient utilization in barley‐based diets. However, thermal processing can increase solubilization of NSP (Silversides & Bedford, 1999), leading to greater viscosity in both feed and intestinal contents particularly in diets based on viscous grains such as barley (Cowieson et al., 2005; García et al., 2008; Svihus et al., 2000). Accordingly, to achieve the desired outcome of the thermal processing, the application of optimum conditions during feed manufacture is vital.

4.2. Carcass and digestive trait characteristics

By pelleting the diet, the amount of fine particles in the feed increases, and these fine particles remain in the gizzard for a shorter period of time compared to coarse particles. As a result, these fine particles quickly pass into the small intestine and have little effect on muscle growth and gizzard function. Moreover, gastric emptying rates of fine particles are significantly higher than coarse particles, which could be the reason for the lower weight of gizzard and gastrointestinal tract in pelleted feed. In agreement with our results, Amerah et al. (2007) reported that the relative weight of gizzard and its content decreased when chickens were fed pelleted diets compared to mash. Jiménez‐Moreno et al. (2019) observed that pelleted feed caused a decrease in the relative weight of the full gastrointestinal tract, the empty weight of the gizzard, the relative length of the small intestine, ceca, and an increase in the weight of full crop and the weight of the liver. Also, in the study of Abdollahi et al. (2013b), the weight of the gizzard decreased in chickens fed with pellet diet compared to mash. In the present study, SFH had not effect on carcass components in broiler chickens. In contrast, it has been shown the presence of insoluble fibre (sugarcane bagasses, wheat bran and SFH) in barley‐based diets of broilers resulted in higher relative weight of breast and thigh and lower relative weight of the liver than the CTL group at 42 days of age (Pourazadi et al., 2020).

4.3. Digestibility of nutrients

In the present study, increases in EE digestibility in barley‐base diets supplemented by SFH were observed. These results were in agreement with the Pourazadi et al. (2020), who reported increase in EE and OM ileal digestibility in broilers fed wheat bran, sugarcane bagasses and SFH in barley‐based diets. The inclusion of fibre in broilers and laying hens’ diet is known to increase the concentration of bile acids (Hetland et al., 2003). Bile acids are strong emulsifiers and facilitate nutrient solubilisation by effective emulsification of liberated lipids (Hetland et al., 2004). This could possibly explain the improvement of the digestibility of EE in the present study by dietary inclusion of SFH. In agreement with our results, Kalmendal et al. (2011) stated that adding oat husk to corn‐based diets increases digestibility of EE. However, contrary to our results, it has been shown that the ileal digestibility of EE was reduced by the addition of OH to a wheat‐based diet compared to the control (Hetland & Svihus, 2001). Moreover, the digestibility of OM decreased by adding sugar beet pulp to corn‐based diets (Pettersson & Razdan, 1993). In the present study, the reason for increasing the digestibility of OM by adding enzymes to the barley‐based diet can be attributed to the positive effects of enzymes on the breakdown of NSP, reducing viscosity, and improving the use of nutrients. Liu and Kim (2016) reported that xylanase supplementation of wheat‐based diet increased the apparent digestibility of DM, CP and GE in chickens. In our experiment, digestibility of nutrients was not affected by the physical form of feed. In this regard, Zhang et al. (2009) observed that pelleting corn‐based diets led to improved AME and apparent digestibility of CP and OM. Jiménez‐Moreno et al. (2019) stated that pellet rations reduced the retention of EE but had no effect on the retention of nitrogen and AME _n diet.

4.4. Caecal microbial population

In our study, the use of SFH improved the count of Lactobacillus spp. It was shown that insoluble fibre increases mucin secretion and subsequently facilitating the colonization of beneficial intestinal bacteria. Dietary fibre, as the substrate for the microbiota, stimulates microbial fermentation and thereby increases the production of short‐chain fatty acids (SCFAs), such as lactic, acetic, butyric, valeric and propionic acids. These SCFA can serve as an extra source of energy to the broiler (Jamroz et al., 2003). Next to their antimicrobial and energy‐yielding effects, these SCFAs, in particular butyrate, are also known for their potential in maintaining gut integrity and strengthening of the gut mucosal barrier through, for example the provision of extra energy to the gut epithelial cells and the regulation of mucin production, respectively (Guilloteau et al., 2010).

In addition, by increasing the fermentation of insoluble fibre in the posterior part of the intestine, SCFAs are produced, and the antibacterial effects of these acids are another mechanism to explain the effect of insoluble fibre on the intestinal microbial population (Wielen et al., 2002). The development of intestinal Lactobacillus spp. population generally indicates the health of the digestive tract. The Lactobacillus spp. produces antimicrobial compounds such as bacteriocins, which have inhibitory effects against the growth of pathogenic microorganisms. In agreement with the results of this study, Pourazadi et al. (2020) stated that the caecal population of Lactobacillus spp. increased in the chickens fed with SFH with coarse particle size compared to the control in barley‐based diets. They also reported that feeding sugarcane bagasse, wheat bran and SFH with coarse particle size and SFH with small particle size significantly reduced the coliforms population compared to the control. In the present study, with the addition of enzyme to the experimental diets with or without SFH, caecal population of E. coli decreased compared to the CTL. This agrees with the findings of Mathlouthi et al. (2002), who reported that xylanase and β‐glucanase supplementation to a wheat‐ and barley‐based diet reduced the counts of E. coli in the ceca but had no effect on the number of lactobacilli.

Enberg et al. (2002) reported that feeding pellet diets increased coliforms and Enterococci bacteria in the ileum and decreased the number of Clostridium perfringens and Lactobacillus spp. in the end part of the digestive tract (cecum and rectum) of chickens. However, the decrease in the population of E. coli in the cecum of chickens fed with pellet diet in the present experiment is probably due to the reduction of digesta viscosity in these treatments compared to the mash diet. It may be due to the presence of insoluble fibre sources and enzymes in pellet rations, because the interaction effects between fibre sources and enzymes in pellet rations have reduced the amount of digesta viscosity.

4.5. Ileum viscosity and pH of gizzard

In the present study, the decrease in the pH of the gizzard in chickens fed with insoluble fibre could be due to more retention of these substances in the proventriculus and gizzard and, as a result, stimulating more acid secretion (Hetland et al., 2005; Jiménez‐Moreno et al., 2010). Jiménez‐Moreno et al. (2009) stated that the addition of 3% oats hulls to a rice‐based diet decreased the pH of the gizzard. Moreover, the use of different sources of insoluble fibre (OH, rice bran and SFH) at two levels of 2.5% and 5% in the rice‐based ration decreased the pH of the gizzard (Jiménez‐Moreno et al., 2019). Kimiaeitalab et al. (2016) stated that the use of 3% SFH increased the weight of the gizzard, decreased the pH of the gizzard, and improved the energy content in the diet.

The increase in ileum viscosity in the treatment without additives is due to the presence of β‐glucan in barley. By absorbing water, non‐starch polysaccharides (especially β‐glucan) create a sticky mass that increases the viscosity and surrounds the digestible substances, which hinders the activity of digestive enzymes and reduces digestibility (Garcia et al., 2008; Rebolé et al., 2010). Increasing the digestibility of nutrients and reducing the viscosity of digesta using exogenous enzymes such as β‐glucan and xylanase due to the destruction of NSP by using enzymes in wheat and barley‐based diets has been reported (Lázaro et al., 2003a, Garcia, Aranibar, et al., 2003; Lázaro et al., 2003b, Garcia, Medel, et al., 2003). Moreover, the reason for the decrease in the digesta viscosity of chickens fed with the diet containing SFH in the present study can be related to the increase in the passage rate of the digesta. Because it has been stated that the increase in the passage rate of the digesta along the digestive tract can reduce the viscosity of digesta in the intestine (Mateos et al., 2002). In this regard, insoluble fibre by increasing the water‐holding capacity in its structure, decreasing the solubility of NSP (Amerah et al., 2009), and increasing the intestinal movements (Hetland et al., 2005), causes an increase in the passage rate. In another research, it has been shown that the use of cellulose reduces viscosity of digesta compared to pectin (Shakouri et al., 2006). Other researchers also stated that insoluble fibre can reduce the viscosity of digesta in broilers fed with wheat and barley‐based diet (Hetland et al., 2003, Montagne et al., 2003; Mateos et al., 2002; Pourazadi et al., 2020; Taheri et al., 2016).

In the present experiment, the decrease in viscosity in pelleted diets due to their interaction effects can be due to the presence of additives (enzymes and insoluble fibre). Ankrah et al. (1999) showed that pelleting oat‐based diets decreased the digesta viscosity of broilers. However, contrary to our results, the viscosity of digesta increased in wheat‐based pellet diets compared to mash (Cowieson et al., 2005) and also stated that using xylanase enzyme in the diet decreased viscosity of digesta.

4.6. Intestinal morphology

VH and villus surface area were measured as a possible mechanism affecting the feed efficiency. Changes in intestinal morphology may affect nutrient metabolism and function. An increase in the height of the intestinal villi increases the surface of absorption, activity of digestive enzymes and transfer of nutrients on the surface of the villi (Cera et al., 1988). In the present experiment, the increase in the VH and the VH/CD ratio of duodenum and jejunum for chickens fed with pelleted diets was in agreement with the increase in growth performance (BWG and FCR), which indicates the response of the digestive and absorptive capacity of the small intestine to the absorption of pellet feed nutrients. In agreement with our results, Naderinejad et al. (2016) reported that the VH in the duodenum and jejunum of chickens that receiving pellet diet was higher than mash. Amerah et al. (2007) reported that feeding birds with wheat‐based pellet diet increased the VH and CD compared to mash diet. Moreover, Zhang et al. (2009) showed that pelleting a corn‐based diet increased VH and the ratio of VH/CD in duodenum, jejunum and ileum compared to mash diet. In the present study, the inclusion of SFH at level of 3% in barley‐based diet numerically increased the VH of duodenum and jejunum compared to the CTL. The beneficial effect of fibre on the VH was reported by Sarikhan et al. (2010). They showed that dietary inclusion of insoluble fibre at different levels (0.25, 0.5 and 0.75% inclusion) improved performance and increased the VH of ileum. The decreased transit time of the digesta in diets with insoluble fibre might reduce the proliferation and also deleterious products of bacteria into the epithelium of the gut wall (Taheri et al., 2016).

5. CONCLUSION

The current study demonstrated that when barley‐based diets are fed to broilers, dietary supplementation of SFH and ENZ could decrease ileum viscosity, with effects being more pronounced with pellets than with mash. Feeding pellets improved ADFI, ADG and FCR during total period of experiment and decreased E. coli population of cecum, regardless of the type of additives used. The dietary inclusion of SFH (3%) of as insoluble fibre source into barley‐based diets increased apparent ileal digestibility of EE and caecal microbial population of Lactobacillus, although dietary inclusion of additives decreased caecal population of E. coli. It seems that dietary supplementation of SFH and ENZ had favourable effects on broilers fed with barley containing diets.

AUTHOR CONTRIBUTIONS

Investigation; data curation: Zeinab Pourazadi. Conceptualization; supervision; writing – original draft preparation: Somayyeh Salari. Software; methodology: Mohammad Reza Tabandeh. Writing – review and editing: Mohammad Reza Abdollahi.

CONFLICT OF INTEREST STATEMENT

The authors declare no conflicts of interest.

FUNDING INFORMATION

Agricultural Sciences and Natural Resources University of Khuzestan

ETHICS STATEMENT

The authors confirm that the ethical policies of the journal, as noted on the journal's author guidelines page, have been adhered to and the appropriate ethical review committee approval has been received.

Pourazadi, Z. , Salari, S. , Tabandeh, M. R. , & Abdollahi, M. R. (2024). Insoluble fibre and enzyme supplementation in mash or pellets diets on growth performance, apparent ileal digestibility and intestinal morphology of broiler chickens fed barley containing diets. Veterinary Medicine and Science, 10, e1399. 10.1002/vms3.1399

DATA AVAILABILITY STATEMENT

Research data are not shared.