Influence of palm kernel cake on the growth performance, gut health and hematochemical indices of slow-growing broilers

ABSTRACT

Four hundred and fifty (450) 3-week-old male and female broiler chicks were randomly allocated to a control diet and four dietary treatments with six replicates in a completely randomized design (CRD) with 15 birds per replicate arranged in a 2 × 2 factorial design to evaluate the effect of palm kernel cake (PKC) diet and multi-blend enzymes on the performance of Sasso broilers. The five dietary treatments were T0 = (control, standard diet), T1 = PKC1*0% (10% PKC with no enzyme), T2 = PKC1*0.05% (10% PKC with enzyme), T3 = PKC2*0% (20% PKC with no enzyme) and T4 = PKC2*0.05% (20% PKC with enzyme). The interactive effect of PKC and enzyme showed improved (p < 0.05) FCR in T1 and T2. Higher counts (p < 0.05) of Lactobacillus spp. were found in PKC2 than in PKC1 and lower Clostridium perfringens counts in T3 compared to T2 at 8 weeks. At week 12, the PKC-enzyme effect resulted in higher (p < 0.05) platelets in the T2 and T4 groups than in the other treatments. The incorporation of 10% PKC with enzyme into the diets of Sasso broilers could maintain bacteria balance in their gut and enhance growth performance.

KEYWORDS:

• Sasso broilers
• palm kernel cake
• multi-blend enzyme
• cecal bacteria
• blood parameters

Introduction

The production and consumption of animal products will continue to increase in order to satisfy the growing demand for livestock products such as meat, egg, and milk (OECD/FAO 2017; Alshelmani et al. 2021). According to FAOSTAT (2013), poultry falls under the dominant livestock type, which is estimated to contribute 109.02 MT of meat. Therefore, a global demand exists for poultry feed ingredients, especially protein (soyabean meal) and energy (maize) sources, subsequently causing an increase in the production cost of poultry feed (Aguzey et al. 2020; Alshelmani et al. 2021). Unconventional sources of protein and energy are accessible to help reduce the cost of producing poultry feed (Koranteng et al. 2022). Such sources include the use of palm kernel cake (PKC), which, is an essential agro-industrial by-product obtained from the extraction of palm kernel (Elaeis guineensis) oil from the palm fruit. It is rich in nutrients (14–18% crude protein, 12–20% fibre, and 3–9% ether extract) to support the growth and productivity of chickens (Aguzey et al. 2020; Azizi et al. 2021). Nevertheless, PKC could contain non-starch polysaccharides (NSPs) such as β-mannans and anti-nutritional factors (Azizi et al. 2021), which have the tendency of impairing the metabolic performance of the chicken, hence PKC inclusion in the diet is limited (Alshelmani et al. 2021). The high content of fibre (Sharmila et al. 2014) present in PKC in addition to its coarse and gritty texture (Alshelmani et al. 2016), also limits its inclusion. Another reason for PKC’s minimal usage in poultry diets is that chickens have a simple digestive system and lack fibre digestive enzyme activities in their gastrointestinal tract (Montagne et al. 2003). Aside from the limits in inclusion level, it is important to limit the fibre inclusion in the diets of younger chicks as their gut is not fully developed to effectively digest it (dos Santos et al. 2019; Ravindran and Abdollahi 2021). According to Singh and Kim (2021), poultry is deficient in endogenous enzymes required for the breakdown of NSPs in PKC. It is imperative to explore means of enhancing the digestive process of chickens fed PKC-incorporated diets. One such means is the use of enzymes as recommended by Soltan (2009). McDonald et al. (2010) explained that the function of enzymes is to break down materials that inhibit the digestion, absorption, and utilization of nutrients and several authors have affirmed the inclusion of exogenous enzymes in poultry diets (Bedford and Cowieson 2012; Chen et al. 2018; Aguzey et al. 2020; Singh and Kim 2021). This has been accepted because exogenous enzymes aid in the breakdown of complex polymeric compounds, resulting in improvement in the utilization of the nutrients (Farhangi and Carter 2007), and poultry tend to benefit due to their limited digestive tract.

According to Aguzey et al. (2020), enzyme activity also helps in the aggregation of the population of beneficial microbes of the GIT of chicken, which, consequently helps in boosting their immune system. In addition, diets play a relevant role in the determination of the composition and density of the intestinal microflora with dietary fibres influencing microbial ecology (Jha and Berrocoso 2015; Yadav and Jha 2019). Azizi et al. (2021) have also opined that intestinal microflora aids the host by assisting in the digestion, absorption, and storage of nutrients.

The choice of feed ingredients should establish a balance between the environment, the host, and the microbiota, with the microbiota interrelating among themselves, with the host, and with the diet, as suggested by Yadav and Jha (2019). Bacteria present in the crop contribute to the fermentation of PKC, thereby increasing the availability of the nutrient content of the diet. PKC also acts as a substrate for microorganisms, and its degradation contributes to the breakdown of inhibitors (Aguzey et al. 2020). According to Choct (2009), manipulating the diet of animals can be very crucial to improving their performance and PKC with its relatively high fibre could be used to influence the gut performance of broiler chicken as poultry require some amount of dietary fibre for normal intestinal physiological functions (González-Alvarado et al. 2007). Dietary fibre levels have been reported to modify gut microflora population and composition and stimulate the digestive system, which may ultimately improve the health of birds (Jha and Mishra 2021). The gut of the broiler can therefore be manipulated to effectively utilize the nutrients consumed. It was hypothesized that broiler diets compounded with PKC, and supplemented with a multi-blend enzyme will improve the gut performance of Sasso broilers alongside their blood indices. This study, therefore, aimed to investigate the effect of feeding graded levels of PKC with or without multi-blend enzymes on the growth performance, gut microbes and blood profile of Sasso broilers.

Materials and methods

The study was conducted under the supervision of the research team leader following the guidelines of the Canadian Council on Animal Care (2009).

Birds husbandry and experimental design

Four hundred and fifty (450) unsexed (male and female) day-old dual-purpose broiler chicks (Sasso X44) obtained from Maison Diop Hatchery were used for this study. The chicks were brooded for 3 weeks and were randomly allocated to a control diet and four dietary treatments having six replicates with 15 birds per replicate in a completely randomized design. The chicks were weighed individually and randomly placed in their respective treatment in an open-sided poultry pen. The floor was littered with wood shavings of 4 cm thickness with a stocking density of 10 birds/m². The experiment lasted for 9 weeks (3–12 weeks of age) with treatments arranged in a 2 × 2 factorial design. Main effects: PKC1 (10% palm kernel cake), PKC2 (20% palm kernel cake diet): enzyme levels (0% and 0.05% enzyme supplement): Treatments T0 = (control, standard diet), T1 = PKC1*0% (10% palm kernel cake with no enzyme), T2 = PKC1*0.05% (10% palm kernel cake with enzyme), T3 = PKC2*0% (20% palm kernel cake with no enzyme) and T4 = PKC2*0.05% (20% palm kernel cake with enzyme). A typical PKC contains 14–18% of crude protein (CP), 12–20% crude fiber (CF), and 3–9% ether extract (EE) (Ibrahim 2013; Azizi et al. 2021). The levels of the incorporation of the PKC adopted in this study were based on earlier works of Okeudo et al. (2006), Soltan (2009), Saminathan et al. (2020) and Hakim et al. (2021) to ascertain whether the source of the test material and environment can impact birds’ performance. The birds were fed these diets in the form of mash and had access to feed and water ad libitum with 12 h of light/day with a mean temperature of 27.8°C and relative humidity of 83%. Experimental birds were vaccinated at designated stages against Gumboro disease (Zoetis, Poulvac*, Bursaplex*Live Virus, Marysville, Kansas), Newcastle and infectious bronchitis disease (Zoetis, B1 Type, B1 Strain, Mass. & Conns. Type, Live Virus Marysville, Kansas). The multi-blend (Kemzyme Plus P) used is a commercial product from Kemin Industries and the main active ingredients were xylanase (20,000,000 U/kg), β-glucanase (2,350,000 U/kg), cellulase (4,000,000 U/kg), phytase; for the degradation of NSPs and α-amylase (400, 000 U/kg) and protease for the enhancement of the action of endogenous digestive enzymes. The PKC (a residue obtained from the extraction of oil for the production of palm kernel oil) was obtained from the GVC Ave Palm processing plant and was milled before incorporation into diets.

Chemical and proximate analysis

Analysis was done on the PKC for anti-nutrients (tannin, saponin, phytate, oxalate) and non-starch polysaccharides including cellulose, lignin, acid detergent fiber (ADF), and neutral detergent fiber (NDF). Proximate analysis was conducted on treatment diets with the AOAC (2006) method. Tannins and saponins levels in the PKC were determined by the method described by Obadoni and Ochuko (2002). Cellulose, ADF, and NDF were determined based on the procedure by Van Soest (1994). The oxalate was determined by the method described by Day and Underwood (1986). Phytate was determined by the method of Reddy and Love (1999).

Experimental rations

Experimental feeds were formulated according to NRC (1994) with the specification of broiler finisher feed requirements. Diets were formulated to be both iso-caloric and iso-nitrogenous. The feed composition and proximate analysis results are presented in Table 1.

Table 1. Ingredients and nutritional composition of experimental diets.

INGREDIENTS (%)	T0	T1	T2	T3	T4
Maize	64.35	56.35	56.35	51.35	51.35
Fishmeal	2.00	2.00	2.00	2.00	2.00
Roasted soya	25.15	23.00	23.00	22.80	22.80
Wheat bran	5.00	5.00	5.00	–	–
PKC	–	10.00	10.00	20.00	20.00
Oyster shell	1.50	1.80	1.80	2.00	2.00
DCP	1.00	1.00	1.00	1.00	1.00
Vitamin Premix	0.30	0.30	0.30	0.30	0.30
Methionine	0.15	0.15	0.15	0.15	0.15
Lysine	0.20	0.20	0.20	0.20	0.20
Salt	0.35	0.20	0.20	0.20	0.20
Multi-blend Enzyme	–	–	0.05	–	0.05
Total	100.0	100.00	100.00	100.00	100.00
Nutrient analysis (%)
Moisture content	9.54	10.37	10.33	10.97	10.04
Ash content	5.65	6.33	6.42	6.55	6.50
Crude protein	19.24	19.31	19.25	19.19	19.29
Crude fat	5.12	5.39	5.34	5.41	5.49
Crude fibre	3.98	5.15	5.12	5.19	5.21
Lysine	1.41	1.49	1.52	1.61	1.63
Methionine	0.64	0.66	0.71	0.81	0.83
NFE	50.73	51.25	46.28	49.31	48.85
Metabolizable energy (Kcal/kg)	3057	3077	3049	3053	3087

PKC = palm kernel cake, DCP = Dicalcium Phosphate, NFE = Nitrogen-free Extra, T0 (control, Standard diet), T1 (PKC1*0% = 10% palm kernel cake with no enzyme), T2 (PKC1*0.05 = 10% palm kernel cake + enzyme), T3 (PKC2*0% = 20% palm kernel cake with no enzyme) and T4 (PKC2*0.05% = 20% palm kernel cake + enzyme).

Data collection

Average daily feed intake was recorded and birds were weighed individually on weekly basis for all replicates. Average daily weight gain (ADWG) and feed conversion ratio (FCR) were calculated at the end of the experimental period though single feed was served during the entire period, based on the recommendation for the Sasso breed at the study institution. At the end of the experimental period, birds from each replicate were weighed and the average body weight of the group was recorded. The ADWG and FCR were calculated using the formula: $\mathrm{ADWG}={\displaystyle \frac{\mathrm{weekly}\mathrm{weight}}{7}\mathrm{FCR}={\displaystyle \frac{\mathrm{feed}\mathrm{intake}}{\mathrm{weight}\mathrm{gain}}}}$

Intestinal sampling

At 8, 10, and 12 weeks of age, six birds (both males and females) from each treatment were randomly selected and humanely sacrificed by severing the jugular vein after being fasted (with free access to water) for 12 h. The entire intestine from each bird was removed aseptically, both ceca were removed and the contents were carefully removed into petri dishes for bacteria counts. Commercial culture media provided by OXOID Diagnostics (France) and BIOKAR (United Kingdom) were used. A 1 g sample was serially diluted (1:10) in 9 ml of tryptone salt (from 10⁻¹ to 10⁻⁵) and spread (0.1 ml of suspension) on a specific selective agar culture medium. Total number of bacteria (total plate count) was enumerated on plate count agar incubated at 30°C for 72 h, while Lactobacillus spp were counted after incubation at 37˚C for 48 h on Gelose Man, Rogosa, and Sharpe (MRS) agar medium. Total Streptococcus spp. were counted after 24 h of incubation at 37°C on Slanetz and Bartley agar medium. Clostridium perfringens were enumerated after incubation at 44°C for 24 h, anaerobically on Tryptone Sulfite Neomycin (TSN) agar medium. All bacteria were counted and expressed as total colony-forming units (CFU) per gram of the digesta, and the results were presented as log₁₀-transformed data.

Hematology and serum biochemistry

At the end of the grower (4–8 weeks) and the finisher (9–12 weeks) stages, 6 birds from each treatment were randomly selected and two sets of 5 ml of blood samples were collected from each bird from the left wing with a sterile syringe and needle for hematological and serum biochemistry analysis. For the determination of the hematological parameters, the blood samples were collected into 5 ml vacutainer tubes containing ethylene-diamine tetra-acetic acid (EDTA), and for the biochemical parameters, blood samples were collected into gel tubes and were centrifuged at 3000 rpm for 15 min to obtain the serum.

Hematology parameters

Hematoanalyzer was used to determine the values of platelets, mean corpuscular hemoglobin (MCH), white blood cells (WBC), hemoglobin, red blood cells (RBC), hematocrit, and mean corpuscular volume (MCV).

Serum biochemistry parameters

The serum obtained was used to measure the concentrations of total cholesterol, triglycerides, HDL-cholesterol, albumin, and total proteins, by the colorimetric method using an automatic device.

Statistical analysis of data

Data obtained on cecal microbial counts, hematology, and serum biochemistry parameters were subjected to a two-way analysis of variance (ANOVA) to determine the main effects of PKC and enzyme incorporation levels using the GLM procedure of Minitab (Version 19), at a 5% level of significance and significant differences between means were tested using the Tukey’s pairwise comparisons of the same statistical software.

Results

Chemical composition/analytical evaluation of PKC

The chemical composition and fibre fraction of the test material is presented in Table 2. The fibre fractions present in the PKC were 24.58%, 24.31% and 7.75% respectively for hemicellulose, cellulose, and lignin. The ADF and NDF were 35.62% and 60.20% respectively. The anti-nutrients included tannin (3500 ppm), saponin (690 ppm), phytate (109.50 ppm), and oxalate (13.4 ppm).

Table 2. Determined fibre fractions and anti-nutrients of palm kernel cake.

Parameters	Composition
Acid detergent fibre (%)	35.62
Neutral detergent fibre (%)	60.20
Hemicellulose (%)	24.58
Cellulose (%)	24.31
Lignin (%)	7.75
Tannin (ppm)	3500
Saponin (ppm)	690
Phytate (ppm)	109.50
Oxalate (ppm)	13.40

Growth performance

Table 3 shows the results recorded on the growth performance of Sasso broilers fed varying levels of PKC supplemented with or without multi-blend enzymes. The initial body weights of all treatments were similar (p > 0.05) at the onset of the experiment. The interactive effect of enzyme and 20% PKC resulted in T4 birds having a higher (p < 0.05) feed consumption compared to the control (T0) and the other treatment groups. Birds fed the 20% PKC consumed more feed (p < 0.05) compared to those fed the 10% PKC diet, in addition, those fed the enzyme-supplemented diet consumed significantly (p < 0.05) more feed than the non-enzyme group. The interactive effect of enzyme and 10% PKC (T2) increased (p < 0.05) the average daily weight gain of birds relative to T0, T1 and T3. In the main effects, birds in the 10% PKC and enzyme-supplemented groups had superior (p < 0.05) average daily weight gain. Birds fed the 10% PKC diet with or without enzymes (T1 and T2) were more efficient (p < 0.05) in feed utilization as opposed to those fed the control diet (T0) and 20% PKC diets (T3 and T4). The body weight was significantly heavier (p < 0.05) in birds fed the 10% PKC with enzyme (T2) compared to the control (T0) and the other treatment groups without enzyme supplementation (T1 and T3). The main effects showed that enzyme-supplemented diets and 10% PKC had superior (p < 0.05) body weight relative to the non-supplemented diets and 20% PKC.

Table 3. Effect of PKC diet supplemented with or without multi-blend enzymes on the overall growth performance of Sasso broilers.

	Parameters
		IBW (g)	ADFI (g)	TFI (g)	ADWG (g)	FCR (g/g)	BODYWT (g)
PKC	PKC1	246.73	78.27^b	4931.01^b	24.32^a	3.22^b	1778.66^a
	PKC2	248.31	82.67^a	5208.21^a	23.16^b	3.57^a	1707.67^b
	SEM	5.24	0.65	138.60	0.26	0.039	16.4
ENZYME LEVELS (%)	0.0	246.85	78.58^b	4950.54^b	22.89^b	3.44	1690.67^b
	0.05	245.00	82.37^a	5189.31^a	24.60^a	3.35	1795.66^a
	SEM	5.24	0.65	119.39	0.26	0.039	16.4
PKC*ENZYME	T0	242.75	80.62^b	5079.06^b	23.04^bc	3.50^a	1694.79^c
	T1	247.71	77.11^b	4857.93^c	23.48^bc	3.29^b	1726.86^bc
	T2	245.75	79.44^b	5004.72^b	25.15^a	3.16^b	1830.46^a
	T3	250.11	80.05^b	5043.15^b	22.29^c	3.60^a	1654.47^c
	T4	246.51	85.30^a	5373.90^a	24.04^ab	3.55^a	1760.86^ab
	SEM	6.06	1.34	84.37	0.36	0.055	29.95
P-VALUE	PKC	0.685	0.0001	0.001	0.006	0.0001	0.008
	ENZYME	1.000	0.001	0.023	0.0001	0.132	0.0001
	PKC*ENZYME	0.957	0.013	0.043	0.023	0.048	0.043

^abc Means with different superscripts within a row are significantly different from each other at P < 0.05, SEM = Standard error of the mean, P-value = probability value. PKC1 (10% palm kernel cake), PKC2 (20% palm kernel cake diet), Enzyme levels (0% and 0.05% enzyme supplement), T0 (control = standard diet), T1 (PKC1*0% = 10% palm kernel cake with no enzyme), T2 (PKC1*0.05 = 10% palm kernel cake + enzyme), T3 (PKC2*0% = 20% palm kernel cake with no enzyme) and T4 (PKC2*0.05% = 20% palm kernel cake + enzyme). IBW (Initial body weight), ADFI (Average daily feed intake), TFI (Total feed intake), ADWG (Average daily weight gain), FCR (Feed conversion ratio), BODYWT (Final body weight).

Composition of cecal microbiota

Table 4 presents the results for the effect of PKC supplemented with or without enzymes and their interaction effect on the population of some selected microbes in the ceca of Sasso broilers. It was observed that at 8 weeks, the interactive effect of enzyme and 20% PKC (T4) recorded lower (p < 0.05) total plate counts than the control (T0) and 10% PKC with enzyme group (T2). The main effect at 10 weeks showed that the addition of 20% PKC (PKC2) to the diet increased total plate count compared to the 10% PKC group (PKC1). However, at 12 weeks, no significant (p > 0.05) changes were observed in the interactive effect and the main effects of the total plate counts in the ceca of the birds. With regards to Lactobacillus spp., significantly higher (p < 0.05) counts were recorded for birds given the 20% PKC (PKC2) diet compared to the PKC1 group at the end of 10 weeks of age, whereas at 8 and 12 weeks, no significant (p > 0.05) interactive and main effects were seen in the population of Lactobacillus spp. in the ceca of the birds. At week 8, the interactive effect showed higher (p < 0.05) counts of total coliforms in T4 compared to T2 and T3 but during week 10, the PKC with enzyme treatment groups (T2 and T4) had lower (p < 0.05) counts of total coliforms compared to T3. Besides, at week 12, the interactive effect showed that T2 had significantly reduced (p < 0.05) counts of total coliforms compared to T0, T3 and T4. Clostridium perfringens were lower (p < 0.05) in the cecum of birds given the control diet (T0) and 20% PKC with no enzyme (T3) diet at 8 weeks compared to those fed the 10% PKC diet with enzyme (T2). The main effect at week 8 also showed lower (p < 0.05) counts of Clostridium perfringens in the PKC2 group than in the PKC1 group. Nevertheless, at 12 weeks, the main effect of enzyme levels showed significantly lower (p < 0.05) counts in the enzyme-supplemented groups than in the non-enzyme-supplemented groups. At 8 weeks, the interactive effects of PKC and multi-blend enzyme supplementation resulted in lower (p < 0.05) counts of cecal Streptococcus spp. in the T4 group compared to the control (T0) and the other treatment groups but at week 10, T0 and T4 recorded the highest (p < 0.05) count compared to T1 and T2. At 12 weeks, Streptococcus spp. counts were influenced by PKC and enzyme interaction with higher (p < 0.05) counts in the control (T0) and T4 groups than in T3.

Table 4. Effect of palm kernel cake diet supplemented with or without enzymes on the bacterial population of Sasso broilers.

		Parameters
		Total plate count			Lactobacillus spp.			Total coliforms			Clostridium perfringens			Streptococcus spp.
Age (Weeks)		8	10	12	8	10	12	8	10	12	8	10	12	8	10	12
PKC	PKC1	7.53	6.10^b	7.32	7.40	5.98^b	7.30	2.99	0.35^b	4.24^b	3.23^a	1.44	1.18	6.71	5.80^b	6.32
	PKC2	7.10	7.44^a	7.72	7.01	6.96^a	7.03	3.51	2.45^a	5.88^a	1.18^b	1.97	1.63	6.24	6.61^a	6.60
	SEM	0.18	0.33	0.17	0.16	0.20	0.25	0.48	0.53	0.24	0.50	0.47	0.43	0.19	0.12	0.15
ENZYME LEVELS (%)	0.0	7.43	7.03	7.36	7.32	6.43	6.93	3.28	2.79^a	5.01	1.80	1.50	2.23^a	6.64	6.18	6.24
	0.05	7.17	6.51	7.68	7.10	6.51	7.41	3.21	0.00^b	5.11	2.61	1.92	0.58^b	6.31	6.22	6.68
	SEM	0.18	0.33	0.17	0.16	0.20	0.25	0.48	0.53	0.24	0.50	0.47	0.43	0.19	0.12	0.15
PKC*ENZYME	T0	7.78^a	7.73	7.43	7.64	7.52	7.68	3.56^ab	0.79^b	5.67^ab	0.48^b	2.10	1.08	7.00^a	7.11^a	6.69^a
	T1	7.38^ab	6.12	7.37	7.33	5.92	7.40	3.76^ab	0.69^b	4.38^bc	2.73^ab	1.44	1.71	6.58^a	5.86^bc	6.33^ab
	T2	7.70^a	6.10	7.27	7.48	6.05	7.20	2.21^c	0.00^b	4.08^c	3.73^a	1.44	0.64	6.85^a	5.73^c	6.31^ab
	T3	7.48^ab	7.95	7.35	7.32	6.95	6.45	2.81^bc	4.89^a	5.65^ab	0.87^b	1.55	2.75	6.72^a	6.50^ab	6.14^b
	T4	6.65^b	6.94	8.10	6.71	6.97	7.61	4.20^a	0.00^b	6.11^a	1.48^ab	2.39	0.52	5.77^b	6.71^a	7.05^a
	SEM	0.25	0.39	0.23	0.23	0.28	0.36	0.67	0.75	0.33	0.71	0.19	0.41	0.22	0.26	0.16
P-VALUE	PKC	0.078	0.010	0.108	0.110	0.003	0.464	0.454	0.013	0.0001	0.010	0.435	0.464	0.098	0.0001	0.220
	ENZYME	0.311	0.275	0.197	0.340	0.780	0.198	0.918	0.002	0.770	0.270	0.536	0.016	0.229	0.829	0.052
	PKC*ENZYME	0.035	0.306	0.091	0.122	0.860	0.073	0.045	0.013	0.028	0.037	0.536	0.354	0.036	0.035	0.044

^abc Means with different superscripts within a row are significantly different from each other at P < 0.05, SEM = Standard error of the mean, P-value = probability value. PKC1 (10% palm kernel cake), PKC2 (20% palm kernel cake diet), Enzyme levels (0% and 0.05% enzyme supplement), T0 (control = standard diet), T1 (PKC1*0% = 10% palm kernel cake with no enzyme), T2 (PKC1*0.05 = 10% palm kernel cake + enzyme), T3 (PKC2*0% = 20% palm kernel cake with no enzyme) and T4 (PKC2*0.05% = 20% palm kernel cake + enzyme).

Hematological profile

The hematological profile of Sasso broilers fed PKC diets supplemented with or without multi-blend enzymes are presented in Table 5. In this study, hemoglobin, red blood cells and hematocrit were not affected (p > 0.05) by PKC inclusion and multi-blend enzyme supplementation. At 8 weeks, the PKC-enzyme interaction revealed that the platelet concentration of birds in T2 was higher (p < 0.05) than those in the control (T0) and the other treatment groups. The main effect of the enzyme levels showed a superior (p < 0.05) concentration of platelets in the enzyme-supplemented groups compared to the non-enzyme-supplemented groups at week 8. The results at week 12 on blood platelets revealed higher (p < 0.05) concentration in T2 than in T0 and T4. The mean corpuscular hemoglobin concentration was high (p < 0.05) in T4 when compared to T2 and T3 at week 8 but at week 12, T4 was superior to T1 and T3. Meanwhile, the interactive effect of PKC and enzyme showed that white blood cells count was significantly lower (p < 0.05) in T2 than in T0 and T3 at week 12. Additionally, at 12 weeks, the main effect of PKC showed that the count of white blood cells in the PKC1 group was significantly lower (p < 0.05) than that in the PKC2 group. The main effect of PKC levels at 12 weeks revealed significantly superior (p < 0.05) counts of mean corpuscular volume in the PKC2 group compared to the PKC1 group.

Table 5. Hematological characteristics of Sasso broilers fed palm kernel cake diets supplemented with or without enzymes.

		Parameters
Age (Weeks)		Plt (x10^9/L)		MCH (pg)		WBC (x10^9/µl)		Hb (g/dl)		RBC (x10^6/µl)		HTC (%)		MCV (fl)
		8	12	8	12	8	12	8	12	8	12	8	12	8	12
PKC	PKC1	34.15	45.65^a	36.90	33.02	247.87	173.51^b	10.81	10.67	2.34	2.61	29.13	32.29	124.77	124.08^b
	PKC2	28.45	35.30^b	37.40	32.98	257.29	186.72^a	11.56	11.20	2.50	2.66	30.83	33.94	123.54	127.85^a
	SEM	2.20	3.28	0.27	0.17	5.89	2.87	0.45	0.18	0.077	0.037	1.08	0.55	0.69	1.17
ENZYME LEVELS (%)	0.0	27.25^b	41.20	36.92	32.45^b	246.60	183.0	10.92	10.96	2.37	2.66	29.51	33.76	124.63	127.21
	0.05	35.35^a	39.75	37.38	33.55^a	258.57	177.23	11.45	10.91	2.46	2.61	30.45	32.47	123.69	124.71
	SEM	2.20	3.28	0.27	0.17	5.89	2.87	0.45	0.18	0.077	0.037	1.08	0.55	0.69	1.17
PKC*ENZYME	T0	22.70^b	29.80^b	37.10^ab	32.82^ab	257.90	190.45^a	11.49	11.38	2.46	2.85	30.88	34.62	125.80	121.63
	T1	25.40^b	39.20^ab	37.25^ab	32.55^bc	243.02	177.46^ab	10.88	10.54	2.35	2.62	29.13	32.37	124.25	124.11
	T2	42.90^a	52.10^a	36.55^b	33.49^ab	252.72	169.56^b	10.73	10.80	2.32	2.60	29.13	32.21	125.29	124.04
	T3	29.10^b	43.20^ab	36.59^b	32.35^c	250.17	188.54^a	10.95	11.38	2.39	2.70	29.88	35.14	125.0	130.31
	T4	27.80^b	27.40^b	38.21^a	33.60^a	264.41	184.89^ab	12.16	11.02	2.60	2.61	31.77	32.73	122.08	125.38
	SEM	3.11	4.64	0.38	0.24	8.33	4.06	0.63	0.26	0.11	0.052	1.52	0.78	0.98	1.66
P-VALUE	PKC	0.085	0.040	0.203	0.852	0.275	0.005	0.251	0.056	0.159	0.370	0.282	0.051	0.226	0.037
	ENZYME	0.019	0.759	0.239	0.0001	0.170	0.174	0.412	0.848	0.441	0.341	0.544	0.118	0.350	0.150
	PKC*ENZYME	0.008	0.007	0.007	0.023	0.789	0.036	0.296	0.245	0.280	0.522	0.544	0.168	0.060	0.161

^abc Means with different superscripts within a row are significantly different from each other at P < 0.05, SEM = Standard error of the mean, P-value = probability value. PKC1 (10% palm kernel cake), PKC2 (20% palm kernel cake diet), Enzyme levels (0% and 0.05% enzyme supplement), T0 (control = standard diet), T1 (PKC1*0% = 10% palm kernel cake with no enzyme), T2 (PKC1*0.05 = 10% palm kernel cake + enzyme), T3 (PKC2*0% = 20% palm kernel cake with no enzyme) and T4 (PKC2*0.05% = 20% palm kernel cake + enzyme). Plt (Platelets), MCH (Mean corpuscular hemoglobin), WBC (White blood cells), Hb (Hemoglobin), RBC (Red blood cells), HCT (Hematocrit), MCV (Mean corpuscular volume).

Serum biochemical profile

Table 6 indicates the results of the effect of PKC supplemented with or without enzymes on the serum biochemistry of Sasso broilers. There was no influence (p > 0.05) across the main effects and interactive effect of PKC and enzymes on total cholesterol, triglycerides and HDL-cholesterol concentrations in the serum of the birds. The interactive effect of PKC and enzyme showed a higher (p < 0.05) concentration of albumin in T2 than in T1 at week 8. The PKC-multi-blend enzyme interactive effect showed that at 8 and 12 weeks, the total protein concentration in the serum of birds in T2 was significantly higher (p < 0.05) than that of the birds in the control (T0) and the other treatment groups.

Table 6. Effect of palm kernel cake diet supplemented with or without enzymes on the serum biochemistry of Sasso broilers.

Age (Weeks)		Parameters
		TCHO (g/L)		TRIG (g/L)		HDL (g/L)		ALB (g/dL)		TP (g/dL)
		8	12	8	12	8	12	8	12	8	12
PKC	PKC1	0.78	0.91	0.35	0.37	0.84	0.86	9.36	11.56	30.78^a	36.40
	PKC2	0.76	0.87	0.34	0.52	0.77	0.89	8.90	10.58	25.62^b	32.76
	SEM	0.028	0.036	0.021	0.058	0.030	0.057	0.38	0.65	1.45	1.68
ENZYME LEVELS (%)
	0.0	0.73	0.90	0.32	0.48	0.79	0.91	8.44^b	11.20	24.76^b	33.84
	0.05	0.81	0.88	0.37	0.41	0.83	0.84	9.82^a	10.95	31.64^a	35.32
	SEM	0.028	0.036	0.021	0.058	0.030	0.057	0.38	0.65	1.45	1.68
PKC*ENZYME	T0	0.82	0.85	0.49	0.98	0.85	0.81	8.96^ab	11.76	24.41^b	38.03^b
	T1	0.72	0.92	0.31	0.39	0.80	0.84	8.24^b	10.90	24.50^b	32.68^cd
	T2	0.84	0.90	0.39	0.34	0.89	0.88	10.48^a	12.22	37.10^a	40.12^a
	T3	0.74	0.87	0.33	0.57	0.78	0.99	8.64^ab	11.49	25.02^b	35.0^c
	T4	0.78	0.87	0.37	0.47	0.77	0.80	9.16^ab	9.67	26.22^b	30.51^d
	SEM	0.040	0.051	0.030	0.081	0.042	0.080	0.53	0.92	2.05	2.38
P-VALUE	PKC	0.591	0.490	0.884	0.072	0.109	0.704	0.399	0.301	0.023	0.145
	ENZYME	0.073	0.802	0.063	0.360	0.304	0.357	0.019	0.789	0.004	0.544
	PKC*ENZYME	0.322	0.908	0.550	0.804	0.238	0.170	0.025	0.106	0.013	0.023

^abc Means with different superscripts within a row are significantly different from each other at P < 0.05, SEM = Standard error of the mean, P-value = probability value. PKC1 (10% palm kernel cake), PKC2 (20% palm kernel cake diet), Enzyme levels (0% and 0.05% enzyme supplement), T0 (control = standard diet), T1 (PKC1*0% = 10% palm kernel cake with no enzyme), T2 (PKC1*0.05 = 10% palm kernel cake + enzyme), T3 (PKC2*0% = 20% palm kernel cake with no enzyme) and T4 (PKC2*0.05% = 20% palm kernel cake + enzyme). TCHO (total cholesterol), TRIG (triglycerides), HDL (high-density lipoprotein), ALB (albumin), TP (total protein).

Discussion

The PKC utilized in this study, based on the chemical analysis, contains several anti-nutritive components and non-starch polysaccharides such as tannins, saponins, ADF, and cellulose. In contrast to the 260 ppm saponin and 310 ppm tannin reported by Hossain et al. (2016), the percent saponin and tannin of the PKC evaluated in the present study were 690 and 3500 ppm, respectively. Tannins are known to reduce the digestibility of dry matter, protein and other nutrients while oxalate interacts with the nutrients in the gastrointestinal tract (Ramteke et al. 2019), but the results revealed that the oxalate and phytate levels present in the tested PKC in this present study were outside the lethal dose of 2–5 g/kg and 50–60 mg/kg, respectively as reported by Inuwa et al. (2011). The values of ADF, NDF, hemicellulose, and cellulose observed in this study were comparable to those of Mustafa et al. (2004) who reported 36.14% ADF, 61.54% NDF, and 25.40% hemicellulose but lower than that (51.48% ADF, 82.29% NDF, 30.81% hemicellulose, and 35.55% cellulose) reported by Alshelmani et al. (2017) . The amount of lignin obtained from PKC for the present study was less than that which Cerveró et al. (2010) reported. Low quantities of lignin can absorb enzymes, promoting enzymatic hydrolysis and minimizing permanent loss of enzyme function (Eriksson et al. 2002). According to Sundu et al. (2006) and Azizi et al. (2021), the variations in the values reported for anti-nutrients and fibre composition might be explained by variations in the geographical areas and species of the palm tree, as well as in quality and the extraction and processing method employed.

The interactive effect of PKC and multi-blend enzyme benefited birds in the control, T1 and T2 treatment groups, as they consumed less feed. The addition of enzymes to the diet of the birds increased their feed intake. This might have been a result of the effect of the multi-blend enzyme improving nutrient digestibility due to the breakdown of non-starch polysaccharides by the activity of the enzymes (Viveros et al. 2000; Okukpe et al. 2019), thereby enhancing the palatability of the feed and nutrient uptake. Overall, adding enzymes to broiler feed can improve nutritional digestibility, intestinal health, and palatability, all of which can result in elevated feed consumption (Elbaz et al. 2023). The ADWG was increased in the birds fed the multi-blend enzymes of the improvement in fibre digestibility. The FCR was better for the birds that were supplemented with enzymes, suggesting a better efficiency that resulted in a better final body weight. The multi-blend enzyme enzyme-supplemented diet used in this study affected the final body weight positively, as recommended by Azizi et al. (2021), that enzyme enzyme-supplemented palm kernel meal helps with nutrient digestibility compared to non-enzyme diets.

The cecum has been described by Sun et al. (2021), as the principal place for microbial fermentation of dietary fibre in chickens, therefore it was chosen as the site for sample collection for this present study. Singh and Kim (2021) reported that dietary fibre inclusion in the diet of chicken can support cellulolytic and beneficial bacteria, such as Lactobacillus and Bifidobacterium, and augment the production of short-chain fatty acids that helps in the avoidance of digestive disorders. Apart from the high fibre and non-starch polysaccharide nature of PKC, it also contains β-mannan, the main component of palm kernel by-products, which functions as a prebiotic (Azizi et al. 2021), and β-mannan is known to improve birds’ immune function by improving non-pathogenic bacteria and decreasing pathogenic bacteria in the small intestines (Sundu et al. 2006).

Total plate count in this study was lower in the 20% PKC with enzyme relative to the control and 10% PKC with the enzyme at week 8 and the main effect of PKC at week 10 showed higher counts in the PKC2 group. Okukpe et al. (2019) reported increased total viable counts in birds fed 10, 20 and 30% PKC diets compared to their control group. In this study, the lower counts observed in the 20% PKC with enzyme were probably due to the presence of β-mannan in PKC, which is known to improve birds’ immune function by improving non-pathogenic bacteria and decreasing pathogenic bacteria in the small intestines (Sundu et al. 2006). The population of Lactobacillus spp. in the ceca only differed during week 10 as the PKC2 group had superior numbers compared to PKC1. Okukpe et al. (2019) also recorded improved Lactobacillus spp. counts at 20% PKC level over the control and the 10% PKC supplemented groups. Ohimain and Ofongo (2013) also reported higher Lactobacillus spp. counts with enzyme incorporation. This could be due to the increased production of short-chain fatty acids as a result of the high dietary fibre, which supports cellulolytic and beneficial bacteria (Singh and Kim 2021). Conversely, Alshelmani et al. (2016) found no significant impact on the lactic acid bacteria counts when birds were fed 10% PKC diets. At the end of the experiment, the interactive effect of PKC and enzyme revealed lower counts of total coliforms in the 10% PKC with enzyme group. Lower total coliform counts in bird digesta can indicate a healthier gut microbiome, which is linked to better digestive function and general health (Aruwa et al. 2021). There were lower counts of Clostridium perfringens in the control group and 20% PKC without enzyme groups than in the 10% PKC with the enzyme. Sundu et al. (2006) indicated that high fibre decreases the number of pathogenic bacteria and in this present study, the addition of enzymes to the diets improved gut functioning better than the diets with no enzyme supplementation. The increasing population of Clostridium perfringens in the gastrointestinal tract is responsible for subclinical infections including chronic damage of the intestinal mucosa causing poor feed efficiency and performance (Skinner et al. 2010). This implies that the population of Clostridium perfringens obtained in this present study was not beyond the threshold to warrant intestinal infections. These findings have been attributed to the fact that mannose-based carbohydrates, such as β-mannan and manno-oligosaccharides in the PKC, are fermented in the ceca due to their indigestible nature (Sundu et al. 2006). Furthermore, according to Gülşen et al. (2002), fermentation of non-starch polysaccharides could generate short-chain fatty acids which can cause a reduction in gastrointestinal pH and present an unfavourable acidic environment, that may decrease pathogenic microbiota growth (Kermanshahi and Rostami 2006). Hakim et al. (2021) reported a reduction in pathogenic bacteria (E. coli) when a fermented PKC-based diet at 5 and 20% with palm oil (5 and 9.5%) was fed to broilers. In this study, an interactive effect was observed in Streptococcus spp. counts and at 12 weeks, the counts were significantly higher in birds given a 20% PKC diet supplemented with enzymes and the control diet. This result affirms that of Zulkifli et al. (2009), who reported a proliferation when a 25% PKC diet was fed to finisher broilers.

Palm kernel incorporation with or without multi-blend enzymes affected some hematological parameters in this study. At 12 weeks the platelet concentration of birds in the enzyme-supplemented groups was higher suggesting the bird’s ability to cope with blood coagulation effectively. On the contrary, Aderibigbe et al. (2018) reported a non-significant interactive effect of PKC and xylanase but rather a decline in blood platelet counts in their enzyme-supplemented treated birds compared to their control counterparts. In addition, the authors found out that the bird fed the 10 and 20% PKC diets also had lower platelet counts but attributed to the presence of anti-nutritional factors present in the PKC, but in this present study, the anti-nutrient levels of the PKC used were within the acceptable limits. The mean corpuscular hemoglobin concentration was high in the 20% PKC supplemented with enzymes. Aderibigbe et al. (2018) again reported a non-significant interactive effect of PKC-xylanase, however, the 20 and 30% PKC inclusion resulted in higher concentrations of MCH compared to their control. In this study, no interactive and main effects were recorded for hemoglobin, RBC and hematocrit. This finding agrees with Aderibigbe et al. (2018), who reported no significant differences in the interactive effect of PKC and xylanase for hemoglobin, RBC, and hematocrit. The findings of Bello et al. (2011) indicated no difference in hemoglobin concentrations when they compared their control group to 10% and 20% PKC groups. Yet, the hemoglobin concentration values obtained in this study were within the normal ranges of 7–13 g/dl stated by Aderibigbe et al. (2018). Conversely, Aderibigbe et al. (2018) reported a higher hemoglobin concentration in their control group compared to the 10, 20 and 30% PKC groups. Results obtained for WBC indicated a lower value for birds given the 10% PKC supplemented with enzyme diet relative to the control and 20% PKC with no enzyme diets suggesting that 10% PKC could reduce the effect of the GIT pathogenic bacteria and may thus indicate a lower phagocytic and humoral immune activity in these birds due to the absence of harmful bacteria as observed in the main effect of the PKC levels. It is well known that a high WBC count is associated with a bacterial infection in the circulatory system. Subhadarsini and Silpa (2020) were of the view that the WBC has an important role in the immune system and that when any foreign matter like bacteria and viruses enter the body, the WBC fight against it, thus, its augmentation in the 20% PKC group. Nevertheless, Ugwu et al. (2008) did not find any difference in WBC counts when PKC was incorporated into the diet of broilers up to 20%. It is worthy of note, that, the hematological (MCH, WBC, Hb, RBC and MCV) values obtained for this study stayed within the normal range of values stated by Simaraks et al. (2004). The difference in our findings could be due to the strain of broilers used in addition to the composition of enzymes.

In this study, the level of PKC and the use of multi-blend enzymes, and their interaction affected only the albumin and protein concentrations of the serum of Sasso broilers. The concentration of albumin in the serum of birds at week 8 was high in the 10% PKC with enzyme group relative to 10% PKC with no enzyme. The total protein concentration in the serum of birds fed 10% PKC with enzyme was relatively high, suggesting that these birds tend to put on more muscles. Meanwhile, Alshelmani et al. (2016) also reported no significant impact on serum parameters upon feeding broilers with 10% PKC. Bello et al. (2011), also reported no significant differences when birds were fed 10% and 20% PKC levels. The health status of birds could be derived from their biochemical blood parameters including total cholesterol, protein, albumin, and triglycerides (Kpomasse et al. 2020). Khadijat et al. (2012) suggested that the quality and quantity of protein consumed by birds constitute the total protein content in the blood. For total cholesterol, triglycerides, and HDL-cholesterol, no significant changes were observed but values of the total cholesterol and triglycerides were numerically similar to that earlier reported for Sasso birds fed standard diets (Kpomasse et al. 2020).

Conclusion

It is inferred from the results that feeding 10% PKC with enzyme improved feed efficiency and body weight. The incorporation of PKC improved gut-beneficial bacteria and reduced pathogenic bacteria. Additionally, 10% PKC in the diet of the birds can improve the health of the birds by enhancing white blood cell counts. Alternative research approaches including bacterial and enzymatic fermentation to break down the high fibre content of PKC is critical and therefore, require much attention for its usage in the commercial poultry feed industry.

Data availability statement

All data used is available and can be provided upon reasonable request.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Additional information

Funding

The authors are much grateful to the World Bank grant International Development of Association (IDA) 5424 through the Centre d’Excellence Régional sur les Sciences Aviaires (CERSA) of the University of Lomé (Togo) for sponsoring this study.