Comparative effects of herbal additive, symbiotic and antibiotic on growth performance, blood constituents, gut microbiota, and immune response in broiler chickens

Abstract

Background:

Using medicinal plants in broiler diets has been gaining attention as an alternative to synthetic additives due to their potential health benefits and lower risk of residue accumulation.

Aims:

The present study primarily aimed to evaluate and compare the effects of herbal additives, specifically barberry (Berberis vulgaris), sumac (Rhus coriaria, L), symbiotic, and antibiotic on broiler chickens.

Methods:

A total of 384 one-day-old broiler chicks (Ross 308) were assigned to eight different dietary treatments, with six replicates per treatment. Experimental diets included control diet (CON), and other experimental groups were supplemented with 0.2 g/kg virginiamycin (VM), symbiotic (SS), 1 g/kg (B₁) and 2 g/kg (B₂) barberry seed powder, 1 g/kg (S₁) and 2 g/kg (S₂) sumac seed powder and 1 g/kg sumac seed powder + 1 g/kg barberry seed powder (B₁ + S₁).

Results:

The study results indicated that body weight increased in birds fed with VM and SS-supplemented diets (P<0.05) during 1 to 42 days of age. All dietary treatments except CON increased the count of Lactobacillus spp. and decreased the number of coliforms versus at the end of the experiment (P<0.05). Antibody titers against Gambaro disease were higher in birds fed diets B2 and B1 + S1 (P<0.05). The results also indicated that the heterophil to lymphocyte ratio was significantly lower in the SS and B1 + S1 groups as compared with the control group.

Conclusion:

A mix of sumac and barberry (1%) powdered seeds has the potential to improve performance, and disease responsiveness and intestinal microbiota in broiler.

Introduction

Over the past few decades, the importance of gastrointestinal health and its role in the health of birds has been an important subject of poultry research. Due to high-stocking densities and genetic selection for faster-growing birds, poultry farms always need a proper implementation of biosecurity programs to enhance the immune system, balance of microorganisms in the gut and resistance to pathogens. Gut microbiota impose several adverse and positive impacts on bird’s health and productivity where they compete with the host animal in obtaining nutrients from the gastrointestinal tract (GIT) and by destroying and altering bile acids, important entities in fat digestion and absorption (Gaskins et al., 2002).

Furthermore, according to Kogut et al. (2020), the relationship between the gut microbiota and the innate immune system of broiler chickens may result in an adaptive immune response and mucosal cells. Based on the findings of this study, it appears that manipulating the gut microbiota in broiler chickens through the use of feed additive supplements, either individually or in combination, could be a promising strategy. This could potentially improve growth performance, enhance immune response, and positively impact the development of mucosal cells in broiler chickens.

Following the European Union ban on antibiotics as growth promoters (AGP) in animal feed in 2006 (Phillips, 2007), several investigations have been conducted to find alternatives to antibiotics for use in poultry feed. The results of these studies have led to the introduction of a few classes of feed additives known as “non-antibiotic feed additives”, including probiotics, prebiotics, symbiotics, and organic acids. Symbiotics, specifically, refer to a synergistic combination of prebiotics and probiotics designed to improve the survival and replacement of dietary microbial supplements in the gastrointestinal tract (GIT) (Dibaji et al., 2014). Symbiotics are not only used for improving the survival of beneficial microorganisms added to food or feed, but also for stimulating the proliferation of specific native bacterial strains present in GIT.

Another class of non-antibiotic feed additive that has received high attention is phytogenic. Phytogenics are known for their ability to improve growth performance, stimulate the secretion of endogenous digestive enzymes, activate the immune response, and exhibit antibacterial, antiviral, and antioxidant actions (Alsamri et al., 2021), as well as their effectiveness against the most common poultry pathogens such as Clostridium perfringens, Campylobacter jejuni, and Escherichia coli. According to Greene et al. (2020), plant-based products can be used as feed additives to improve the performance of broilers under heat stress conditions. In the poultry industry, the perspective of health management through nutrition is one of the most significant and popular topics. Many of the beneficial properties of photogenic compounds are due to their bioactive molecules. Berberis vulgaris (family Berberidaceae) is a candidate herbal remedy that contains an isoquinoline alkaloid, exhibiting many biological activities such as boosting the immune system, antibacterial, antioxidant, anti-inflammatory, hypotensive, anti-cholesterolemic, and hepatoprotective effects (Abushouk et al., 2017).

Berberis is extensively cultivated in Iran (typically in Birjand), central and southern Europe and the northeastern United States (Tabeshpour et al., 2017). Sumac (Rhus coriaria L) is another herbal medicine belonging to the Anacardiaceous family. It is a wild-growing plant mainly found in Mediterranean bordering countries, South Europe, North Africa, Iran, and Afghanistan (Abu-Reidah et al., 2014). Sumac contains flavanols, tannins, phenolic acids, organic acids, and anthocyanin and many studies have confirmed antioxidant effects (Abu-Reidah et al., 2015), hypocholesterolemic and antibacterial effects against both gram-negative and gram-positive bacteria for this plant (Nasar-Abbas and Halkman, 2004). Azizi et al. (2020) by examining different levels (0.05, 0.1, 0.15, and 0.2%) of sumac observed significant effects in reducing cholesterol and LDL cholesterol concentration as well as the amount of E. coli in the ileum digesta in chickens fed with 0.1% of sumac powder decreased as compared with other treatments. In a study conducted by Mansoub (2011), different levels of sumac (0.75%, 1%, 1.5%, and 2%) were added to the diet of broiler chickens. The results showed that the groups fed with 1.5% and 2% sumac had higher percentages of breast and thigh, and the lowest percentage of abdominal fat was observed in the 1.5% sumac group.

In recent years, the plant compounds have been investigated due to availability, high distribution, low cost, and fewer side effects as new food additives with natural properties and prevention of liver damage and lipid peroxidation. Given the promising results of previous studies on the use of sumac and barberry fruit in broiler feed, including improved meat quality and potential health benefits for humans, this study aimed to investigate the effects of feeding powdered sumac and barberry fruit in broiler diets. Specifically, the study aimed to evaluate the impact of these dietary supplements on growth performance, organ characteristics, serum lipid profile, immunity, hematology, and Gut microbiota in broiler chickens. The results of these interventions will be compared with those of symbiotic and a well-documented antibiotic growth promoter, virginiamycin.

Materials and Methods

Ethics statement

By adhering to the guidelines approved by the Animal Health and Care Committee of Ilam Agricultural University, this experiment ensured the ethical treatment and welfare of the animals involved. Throughout the trial, the birds were handled in compliance with local laws and regulations.

Birds and management

A total of 384 Ross 308 broiler chickens (one-day male) with an average initial weight of 43 ± 0.05 and randomly allocated to one of the eight dietary treatments. There were six repetitions of each treatment, each involving eight chicks. All the chickens were raised in wire cages where the lightening regimen was 23 h light/1 h darkness. On the 28th, the temperature ranged from 33°C to 22°C and then remained constant. Experimental treatments consisted of:

1) A maize and soybean seed basal diet in mash form (Table 1) as control diet (CON)

Table 1.

Constituents and chemical compositions of basal diets

Ingredient (kg)	Starter (days 1-21)	Grower (days 22-42)
Maize	645.90	681.90
Soybean meal (44%)	291.0	270.0
Fish meal	28.70	14.10
Dicalcium phosphate	14.70	14.60
Oyster shells	10.60	10.60
Mineral premixa	2.50	2.50
vitamin premixb	2.50	2.50
Common salt	0.25	0.25
DL-Methionine	1.40	1.30
L-Lysine (HCL)	0.20	0.40
Energy (ME) (MJ. Kg-1)	12.14	12.35
Crude protein	204.30	188.5
Crude fiber (%)	3.57	3.94
Ether extract (%)	2.82	2.88
Ca (%)	9.20	9.60
Available phosphorous	4.60	4.20
Na	1.50	1.40
Methionine	5.20	4.70
Methionine + cysteine (%)	8.30	7.60
Lysine	1.16	1.02

^{a, b} Vitamin-mineral premix provided the following nutrients per kg of diet: vit A, 9000 IU; vit D3, 2000 IU; vit E, 18 IU; K3, 2.1 mg; thiamine, 1.8 mg; riboflavin, 6.6 mg; niacin, 30 mg; pantothenic acid, 10 mg; pyridoxine, 3 mg; biotin, 0.1 mg; choline chloride, 100 g; folic acid, 1 mg; cyanocobalamin, 0.015 mg; phosphorous, 1.6 mg; calcium, 2.3 mg; Zn, 100 mg; Mn, 100 mg; Fe, 50 mg; Cu, 10 mg; I, 1 mg; Se, 0.2

2) CON + 0.2 g/kg virginiamycin (VM)

3) CON + 0.5 g/kg symbiotic (SS)

4) CON + 1 g/kg barberry seed powder (B₁)

5) CON + 2 g/kg barberry seed powder (B₂)

6) CON + 1 g/kg sumac seed powder (S₁)

7) CON + 2 g/kg sumac seed powder (S₂)

8) CON + 1 g/kg sumac +1 g/kg barberry (B₁+S₁)

Virginiamycin was received form Tolide Darouhai Dami Iran Co., Tehran, Iran. The symbiotic (Biomin^®IMBO) was prepared from a commercial producer (Protexin aquatic, Nikotak, Tehran). It is made up of probiotic strain Enterococcus faecium prebiotic fructo-oligosaccharides, cell wall fragments (made of β-glucans, lipopolysaccharides, peptidoglycans and nucleotides), and phycophytic substances derived from sea algae. The sumac and barberry seeds were obtained from the nearby herbal market in Esfahan, Iran. They were dried in a dark place and ground into a powder using a mixer grinder. Chemical analysis of Rhuscoriaria L including 95.5% DM, 4.55% CP, 22% crude fiber, 18.5% ether extract and 6% ash and Berberis vulgaris 94.5% DM, 2.30% CP, 24% crude fiber, 8% ether extract and 5% ash were determined using the method of the Association of Official Analytical Chemists (AOAC International, 2000). Diet was prepared in accordance with Ross 308’s guidelines to satisfy the nutritional needs of broilers. The basal control diet was changed according to the bird’s age (grower diet 1-21 d; finisher diet 22-42 d) and did not include anticoccidials or antibiotics. Broilers had ad libitum access to experimental feed and tap water. The birds were regularly vaccinated against infectious bronchitis, Newcastle disease and Gambaro disease, but were not drugged for the entire experimental period.

Performance parameters

Feed intake (FI) and body weight (BW) were measured for all birds in a cage on days 10, 28, and 42 of age. The ratio of feed intake (FI) to body weight gain (BWG) was used to compute the feed conversion ratio (FCR) for the phases of 1 to 10 d, 11 to 21 d, 22 to 42 d, and 1 to 42 d. Mortality was recorded upon occurrence. We did not investigate the mortality rate because there were just two deaths over the entire growth period. At day 42 of age, ten birds from each treatment were randomly chosen and killed by cervical dislocation to evaluate relative weight (expressed as a percentage of the eviscerated carcass) of abdominal fat and immune organs (spleen, bursa of Fabricius, and thymus).

Blood sampling

At day 42 of age, blood sample were collected from the wing vein of 10 birds per group (2 birds/replicate) into plasma-separating tubes containing lithium heparin (100 IU/5 ml). Blood samples were centrifuged at 3000 × g for 10 min; plasma was then collected and stored at -20°C until analysis. Plasma glucose (Glu), high-density lipoprotein cholesterol (HDL-C), total cholesterol (TC), triglycerides (TG), low-density lipoprotein cholesterol (LDL-C), and total protein (TP) were measured using standard protocols of commercial laboratory kits (Pars Azmoon, Co., Tehran, Iran). For heterophils (H) and lymphocytes (L), a blood smear was prepared using Giemsa staining, and the number of H and L were counted to a total of 100 cells following a schematic diagram and using a Leica DM500 microscope with a magnification of ×100 immersion oil (Fidan et al., 2017). The H/L ratio was calculated by dividing the number of heterophil cells by lymphocyte cells.

Bacteriological analysis

Ten birds were selected for one treatment and killed by dislocation of the cervical vertebrae on day 42. The carcass was then opened and the entire gastrointestinal tract removed under aseptic conditions. The jejunum (from the most distal insertion of the duodenal mesentery to the junction with the Meckel’s diverticulum), the ileum (from the junction with the Meckels diverticulum to the ileocaecal junction), and the cecum (from the mouth to the end of each cecum) were then excised. Samples (1 g) of the contents of the jejenum, ileum and caecum were immediately collected in glass containers and were mixed with 9 ml of sterile dilution blank solution, and homogenized for three minutes. Then, serial dilutions (10⁻¹ to 10⁻⁷) were prepared for further analysis by Jin et al. (1998). All microbial analyzes were performed in duplicate, and mean values were used for statistical analysis. Lactobacillus species were grown on in MRS (de Man, Rogosa, and Sharpe) agar (Merck, Darmstadt, Germany) after incubation in anaerobic chamber at 37°C for 48 h, and coliform bacteria were grown on McConkey agar (Merck, Darmstadt, Germany). The selective agar used to enumerate coliforms was incubated under aerobic condition, at 37°C for 24 h. The number of bacteria was expressed as l g CFU/g digestion of jejunum, ileum, or caecum.

Humoral immune response

Newcastle (NDV) and Gambaro (IBD) antibody levels were measured in sera samples through inhibition (HI) test according to Cunningham (1971) and enzyme-linked immunosorbent assay (ELISA) using a commercial kit according to the instructions provided by the manufacturer (Pars Azmoon, Co., Tehran, Iran). To do so, blood samples were collected from two randomly taken birds per replication on days 10 and 20 after the related immunization. All antibody titers were recorded as log₂ of the highest dilution of serum that agglutinated an equal volume of a 0.5% suspension in PBS.

Statistical analysis

Data were subjected to one-way ANOVA in a randomized complete block design using the GLM procedure of the statistical analysis system (SAS Institute, 2001). Cages were used as the experimental unit for data analysis. Differences between means were tested using Tukey’s multiple range method, and P<0.05 was statistically significant.

Results

Growth performance and carcass composition

No significant effect of dietary experimental was observed on FI, BW and FCR in 1-10 d (P>0.05). Mean BW was greater in the birds fed on the feed additive supplementation-diets compared with those fed on the CON diet during days 11 to 21 of age (P<0.05; Table 2). In the same period, FI was lower in the birds fed with diets B₁ + S₁ and SS than that in those fed with the VM, B₁ and B₂ diets (P<0.05). Mean FCR was better in all herbal supplemented groups than that of those offered by CON and B₂ diets from days 11 to 21 (P<0.05). Broilers fed with VM showed a greater BW than other groups during 22 to 42 days of age (P<0.05). In this period, mean FCR decreased in the SS and VM-fed birds compared with those fed on the CON, B₁ and S₁ diet (P<0.05). At the end of experiment, the birds fed with diets VM and SS exhibited greater body weight gains (P<0.05; Table 2). Mean FI was lower in the birds fed with the SS, B₂, S₂ and B₁ + S₁ diets than that of those received the CON, VM, and S₁ diets during 22 to 42 and 1 to 42 days of age. Feed conversion ratio was better in the birds fed with all experimental diets compared with that in the CON group during 1 to 42 days of age (P<0.05).

Table 2.

Effects of different dietary treatments on growth performance of broilers

Diets	1 to 10 d			11 to 21 d			22 to 42 d			1 to 42 d
	BW (g)	FI (g)	FCR	BW (g)	FI (g)	FCR	BW (g)	FI (g)	FCR	BW (g)	FI (g)	FCR
CON	99	146.85	1.49	514.60b	766.37abc	1.48a	1795.30c	3656.67a	2.03a	2408.67c	4642.30a	1.92a
VM	102.14	147.85	1.45	622.60a	801.43a	1.28bc	1962.40a	3649.47a	1.86b	2687.14a	4598.76a	1.71cd
SS	102.14	144.97	1.42	594.33a	720.90bc	1.21dc	1943.00ab	3481.17bc	1.76b	2639.48a	4347.04c	1.64d
B1	100.42	146.57	1.46	609.23a	818.83a	1.34bc	1733.37c	3554.18abc	2.05a	2443.03bc	4519.58ab	1.85ab
B2	105.08	148.29	1.41	585.77a	815.70a	1.39ab	1776.17c	3440.00c	1.93ab	2467.02bc	4403.99bc	1.78bc
S1	97.85	138.20	1.41	585.87a	787.97ab	1.34bc	1779.30c	3666.83a	2.06a	2465.02bc	4593.00a	1.86ab
S2	102.57	149.42	1.46	579.87a	764.37abc	1.32bc	1802.14c	3497.33bc	1.95ab	2484.58bc	4411.13cb	1.78bc
B1+S1	102.85	147.85	1.44	623.33a	704.17c	1.12d	1841.28bc	3579.00ab	1.94ab	2567.47ab	4431.02cb	1.72cd
SEM	3.441	3.502	0.043	14.924	21.367	0.044	38.946	41.902	0.052	42.989	42.493	0.037
P-value	0.86	0.45	0.89	0.0006	0.0043	0.002	0.0020	0.0023	0.0124	0.0005	0.0001	0.0003

^{a, b} Means within same column with different superscripts differ significantly (P<0.05), the number of observations n=48 in each group. CON: Control diet, VM: 0.2 g/kg virginiamycin, SS: Synbioti, B₁: 1 and B₂: 2 g/kg barberry powdered seeds, S₁: 1 and S₂: 2 g/kg sumac powdered seeds, and B₁+S₁: Mixture of barberry 1 g/kg + sumac 1 g/kg. BW: Body weight, FI: Feed intake, and FCR: Feed conversation ratio

Blood biochemical parameters

Table 3 shows that the chicks fed with S2 diet had a greater plasma total protein level than those fed with CON or S1 diet (P<0.05). The broilers fed with the CON diet had lower plasma glucose levels (P<0.05) than the broilers fed with the SS-supplemented diet. All diets, except VM and B₁, decreased plasma TG levels as compared with the control birds. The mean blood concentration of cholesterol was larger in the birds fed on the CON diet than in the birds receiving B₁ + S₂ diet (P<0.05).

Table 3.

Effect of different dietary treatment on biochemical parameters of 42-day-old broilers

Diets	Protein (g/dl)	Glucose (mmol/l)	Triglyceride (mmol/l)	Cholesterol (mmol/l)	HDL-C (mmol/l)	LDL-C (mmol/l)
CON	3.75c	12.03c	5.17a	8.79a	3.60	4.16
VM	4.05abc	12.56abc	4.37ab	7.79ab	3.82	3.82
SS	4.04ab	12.97a	3.46bc	6.70bc	3.69	2.31
B1	3.94abc	12.27bc	4.40ab	7.98abc	3.96	3.13
B2	4.19ab	12.75ab	3.87bc	7.71abc	3.69	3.24
S1	3.85bc	12.29bc	4.18bc	8.42ab	4.14	3.46
S2	4.30a	12.76ab	3.35c	7.47abc	4.05	2.91
B1+S1	4.18ab	12.77ab	3.49bc	6.59c	3.58	2.30
SEM	0.13	0.09	0.11	0.19	0.12	0.22
P-value	0.05	0.03	0.002	0.002	0.9	0.2

^{a, b} Means within same column with different superscripts differ significantly (P<0.05), the number of observations n=48 in each group. CON: Control die, VM: 0.2 g/kg virginiamycin, SS: Synbiotic, B₁: 1 and B₂: 2 g/kg barberry powdered seeds, S₁: 1 and S₂: 2 g/kg sumac powdered seeds, and B₁+S₁: Mixture of barberry 1 g/kg + sumac 1 g/kg. BW: Body weight, FI: Feed intake, and FCR: Feed conversation ratio

Organ indices and immunological parameters

Table 4 shows that the percentage of abdominal fat in the birds fed with the SS, B2, S2, and B₁ + S₁ diets was lower than that of the birds fed with the CON diet (P<0.05). Experimental treatments exert no influence on the weight of thymus and bursa of Fabricius at 42 days of age (Table 4; P>0.05), whereas a marked increase was observed in the weight of the spleen in broilers fed with VM, SS and S₁ diets compared with those fed with S₂ and B₁ + S₁ diets (P<0.05). Heterophil (H) and lymphocyte (L) counts and H/L ratio was affected by dietary treatments (P<0.05), where the lower heterophil counts and H/L ratio were observed in the birds fed on the SS and B₁ + S₁ diets compared with the control birds. Lymphocyte count was also greater in the birds fed on the SS diet compared with the other groups (P<0.05; Table 4).

Table 4.

Effect of different dietary treatment on organ indices and immune parameters of 42-day-old broilers

Diets	Abdominal fat (g)	Spleen (g)	Thymus (g)	Bursa of fabricius (g)	Heterophil (%)	Lymphocyte (%)	H/L (%)
CON	39.10a	3.54ab	3.48	3.34	22.1a	59.3c	0.38a
VM	30.28ab	4.06a	4.34	3.21	20.2ab	67.3b	0.30cd
SS	19.82b	3.87a	4.19	3.56	15.7c	71.7a	0.22e
B1	30.92ab	3.37ab	3.65	3.32	20.6ab	59.5c	0.35ab
B2	26.82b	3.08ab	4.10	3.47	18.9b	66.8b	0.28cd
S1	29.13ab	3.97a	3.86	3.09	18.5b	58.8c	0.32bc
S2	25.28b	2.41b	3.83	2.21	19.7ab	64.5b	0.30cd
B1+S1	29.62b	2.80ab	3.85	2.04	18.1bc	67.1b	0.27d
SEM	1.35	0.15	0.16	0.12	0.84	1.09	0.013
P-value	0.032	0.049	0.937	0.5006	0.011	0.0001	0.0001

^{a, b} the number of observations n=48 in each group. CON: Control diet: VM: 0.2 g/kg virginiamycin, SS: Synbiotic, B₁: 1 and B₂: 2 g/kg barberry powdered seeds, S₁: 1 and S₂: 2 g/kg sumac powdered seeds, and B₁+S₁: Mixture of barberry 1 g/kg + sumac 1 g/kg

Intestinal Lactobacillus and Coliform bacteria communities

The numbers of Lactobacillus in the jejunum of birds fed VM and CON diets were significantly (Table 5; P<0.05) lower than in the birds fed with other experimental diets. However, the total count of Lactobacillus in the ileum decreased when the broilers were fed on VM and CON diets (P<0.05). The cecal population of Lactobacillus significantly was greater in the birds receiving the SS, B₂, S₂ and B₁ + S₁ diets compared with birds fed on the CON diet at 42 days of age (P<0.05). The numbers of coliforms in the jejunum, ileum and caecum of birds fed CON diet were significantly higher compared with the birds fed with other experimental diets (Table 5; P<0.05).

Table 5.

Effects of different dietary treatment on populations of lactobacillus and coliforms jejunum, ileum, and caecum of 42-day-old broiler (log10 CFU^* g⁻¹ fresh digesta)

Diets	Lactobacillus				Coliforms
	Jejunum	Ileum	Ceca		Jejunum	Ileum	Ceca
CON	7.85b	7.88c	8.00c		3.40a	3.96c	7.53a
VM	7.98b	8.29b	8.55ab		1.90bc	2.65b	5.35b
SS	8.64a	8.44ab	8.85a		2.00bc	2.78b	5.48b
B1	8.55a	8.65a	8.60ab		2.42b	2.88b	6.13b
B2	8.70a	8.72a	8.81a		1.69bc	2.62b	6.08b
S1	8.36a	8.68a	8.53ab		2.03c	2.82b	5.85b
S2	8.40a	8.38ab	8.72a		1.89bc	2.65b	5.40b
B1+S1	8.48a	8.70ab	8.78a		1.95bc	2.26b	5.74b
SEM	0.15	0.055	0.090		0.172	0.33	0.35
P-value	0.031	0.001	0.004		0.007	0.019	0.043

^{a, b} Means within same column with different superscripts differ significantly (P<0.05), the number of observations n=48 in each group. CON: Control diet, VM: 0.2 g/kg virginiamycin, SS: Symbiotic, B₁: 1 and B₂: 2 g/kg barberry powdered seeds, S₁: 1 and S₂: 2 g/kg sumac powdered seeds, and B₁+S₁: Mixture of barberry 1 g/kg + sumac 1 g/kg. ^* CFU: Colony forming unit

Humoral immune response against Newcastle (ND) and infectious bursal disease (IBD) disease

Primary antibody titer against ND and IBD was not affected by various experimental diets (Table 6; P>0.05). Broilers fed with the SS, B2, S2 and B₁ + S₁ diets showed greater titers for total antibody titers against ND virus (P<0.05) for the secondary response (at 32 days of age) compared with those fed on the control diet. Moreover, antibody titers against Gambaro virus were greater in the birds fed with the B₂ and B₁ + S₁ diets at 42 days of age (P<0.05).

Table 6.

Effect of different dietary treatment on the levels of humoral immune antibody titer against Newcastle and infectious bursal vaccine in broiler

Diets	ND titer			IBD titer
	Primary titer	Secondary titer		Primary titer	Secondary titer
CON	5.25	4.50b		9.92	8.59c
VM	5.50	5.50ab		10.00	9.09abc
SS	5.25	5.75a		10.61	9.09abc
B1	5.25	5.25ab		9.92	9.10abc
B2	5.00	6.00a		10.36	9.82a
S1	5.50	5.50ab		9.84	8.59c
S2	5.25	5.75a		10.34	9.64ab
B1+S1	4.75	6.00a		10.28	9.83a
SEM	0.490	0.373		0.300	0.311
P-value	0.3102	0.0021		0.2111	0.001

^{a, b} Means within same column with different superscripts differ significantly (P<0.05), the number of observations n=48 in each group. CON: Control diet, VM: 0.2 g/kg virginiamycin, SS: Symbiotic, B₁: 1 and B₂: 2 g/kg barberry powdered seeds, S₁: 1 and S₂: 2 g/kg sumac powdered seeds, and B₁+S₁: Mixture of barberry 1 g/kg + sumac 1 g/kg. ND: Newcastle disease, and IBD: Infectious bursal disease

Discussion

As our results showed, the addition of antibiotics to the diet improved weight gain and decreased FCR, while FI was like that recorded in control chickens. This confirms the findings of previous studies that showed the addition of antibiotic growth stimulants has a positive effect on the growth of broilers due to its effect on intestinal microbiota (Khodambashi Emami et al., 2015). This was highlighted by Bedford (2000), that germ-free animals were not affected by growth-promoting antibiotics. The use of growth-promoting antibiotics can increase the uptake of nutrients by the host and productivity in poultry by manipulating the community of beneficial bacteria (Baskara et al., 2020).

In the present study, supplementation of symbiotic in the diet improved body weight and FCR of birds possibly due to providing a better gut health status and microbiota function. In agreement with our results, Chayatid et al. (2019) reported that symbiotics (MOS mixed with Bacillus subtilis and Bacillus licheniformis) assist to optimize digestive system health and microbial balance in the intestine, thereby improving growth performance and reduction in the mortality of poultry. It seems that the synergistic increase the availability of indigestible nutrients and beneficial changes in food metabolism by improving intestinal microbial balance, increasing the activity of digestive enzymes, and activating digestive enzymes, thereby improving feed efficiency. Studies have been conducted recently to actively characterize the impact of phytogenic in enhancing animal health and performance.

In this study, the B₁+S₁ added diet, despite no difference with VM and SS included diets, was able to increase BWG compared with CON diet. This improvement is likely realized through synergistic effects between active plant compounds in protecting the intestinal absorption surface leading to increased use of nutrients. Medicinal plants can improve body weight by reducing harmful bacteria in GIT and increasing digestion and absorption of nutrients. Reduction of pathogenic bacteria due to reduced competition with nutrients and prevention of several intestinal diseases has a positive effect on increasing access to nutrients for animal use (Baskara et al., 2020). Consistent with the present results, previous studies have suggested an improvement in the performance of the chickens fed with a diet supplemented with sumac (Toghyani and Faghan, 2017) and barberry (Zhu et al., 2021). According to Ghasemi et al. (2014), the improved broiler’s performance with sumac-supplemented diets is due to the presence of specific biochemical components (cinnamaldehyde and eugenol) found in sumac, causing greater efficiency in the use of feed, resulting in an enhanced weight gain. In line with the findings of previous studies (Cho et al., 2014), the results of this study showed that supplementation of broiler diets with phytogenic feed additive reduced the proportion of the intestinal coliforms and increased lactobacilli populations, leading to improvement in nutrients digestibility of feed and beneficially alters material metabolism, the results which collectively improved the efficiency of feeding and increased weight gain of the birds.

In the present study, abdominal fat percentage decreased by 0.55, 0.57, 0.64 and 0.70% in the broilers fed with B₁ + S₁, S₂, B₂ and SS diets compared with those fed with CON (1.82%) diet, respectively. Ahmadian et al. (2020) reported that abdominal fat weight was primarily affected by the feed supplements, regardless of dose, being reduced by 41% for chickens fed the thyme supplements and 62% for chickens fed the sumac supplements. The presence of harmful microbes in GIT, increases the breakdown of proteins and amino acids in digestive substances, the deamination activity of proteins and amino acids consumed, and increases their rate of breakdown due to the secretion of substances such as urease by microbes. Accordingly, since medicinal plants have reduced the harmful microbial population of GIT, the rate of breakdown of proteins and amino acids in the digestive tract is reduced and more of them are absorbed and stored in the body, leading to improved carcass percentage. As a result, it reduces the conversion of protein to fat and less fat can accumulate in the body. Blood lipid-lowering effects of barberry and sumac in broiler’s chicken may be attributed to the presence of alkaloid and tannins contents, as well as an increase in dietary crude fiber. Chand et al. (2007) observed that inclusion of 20 g/kg Berberis lycium decreased serum total cholesterol, triglyceride and LDL and increased HDL. The active components of barberry, such as alkaloids are associated with the up-regulation of the hepatic expression of LDL-receptors in cells.

Plant active compounds help the growth and proliferation of lactobacilli by lowering the pH of GIT, especially the intestines, and by producing enzymes that change the chemical structure of bile acids and decongest them, which in turn reduces the amount of cholesterol. In a normal broiler, Lactobacillus spp. are the most predominant bacteria in the crop. They are released from the lining of the crop, travel through the rest of the digestive tract, and therefore play a significant role in the populations of bacteria present in the small intestine. Lactobacillus strains can decrease harmful pathogens such as E. coli in the crop and small intestine, thereby help to maintain the microbial balance in GIT. In the present study, the populations of Lactobacilli, was lower in the birds given any of the herbal treatments compared with those fed with the control diet. Similar results were reported by Dibaji et al. (2014). According to Li et al. (2019), Bacillus species controlled the makeup of the intestinal bacterial flora, preserved the equilibrium of the GIT microbiota, and enhanced the intestinal mucosa’s immune response. Dietary probiotic and prebiotic can decrease the intestinal luminal pH due to the production of lactic acid, which could affect the composition of the microbiota, especially pathogenic bacteria and reduce the development of pathogens. Bukhari et al. (2011) evaluated potential positive activities of barberry against E. coli and proteus (100-80%), pseudomonas and staphylococcus (60-70%) microorganisms compared with penicillin G. Shariatmadari and Shariatmadari (2020) also suggested that the intestinal Lactobacillus counts were increased in broiler fed with a sumac supplemented-diet than those receiving the control diet. Antimicrobial activity is one of the primary properties of plants against pathogenic intestinal microbiota.

In this regard, it has been confirmed that the phenolic compounds in phytogenics are among the main substituents responsible for disrupting bacterial cell wall by changing the Na+ and K permeability and reacting to cytoplasmic membrane causing leakage of intracellular compounds (Lopez Romero et al., 2015) and disintegration cell membrane and inhibition of ATP synthesis and eventually cell death. The H/L ratio is a stress indicator and a reduction in this ratio indicates an increase in immunoregulatory capacity of the host.

In this study, immune-stimulatory effects of symbiotic, barberry, and sumac were confirmed by lower H/L ratio in the broilers fed diets SS, B₂, S₁, S₂ and B₁ + S₁ compared with CON diet. Results of Toghyani and Faghan (2017) showed no treatment effect on antibody titers against NDV by adding 3 and 7 g/kg sumac powder in the broiler diet in comparison with an antibiotic growth promoter (AGP), but H/L ratio was decreased in response to 7 g/kg sumac and AGP. Since there was no disease challenge in this study, the ratio of heterophile to lymphocyte has not been expected to increase. Therefore, a decreased H/L ratio might show that sumac powder imposed no stress in birds. In general, the results of the present study showed that in comparison with the control group, the addition of VM in the chicken’s diet showed a positive effect on growth performance and health of broilers, but due to the potential of bacterial resistance and antibiotic residues in animal products, attempts are being made to replace them with other feed additives such as symbiotics and phytogenics.

According to our results, symbiotics stimulated selective growth and activity of beneficial microorganisms in the gut and thereby benefit health. Phylogenetics can increase the number of lactic acid-producing bacteria in the ileum and cecum of broilers and decrease the number of coliforms by lowering of the pH value. Since Lactobacillus species play an important role in creating this balance and increasing the level of local and temporal immunity of the host. By manipulating the microorganisms of GIT and creating a proper microbial balance in this organ, the immune response and performance of the bird can be optimally improved. The results of the present study demonstrated that the sumac- and barberry-supplementation diets significantly improved body weight gain, feed conversion ratio, immune system, and intestinal microbiota, with significant benefits in reducing fat deposition in broiler chickens. Plant’s effects may be due to greater efficiency in the use of feed which results in enhanced growth.

According to the study results, dietary inclusion of the sumac and barberry mixture can be applied as an alternative to antibiotic growth promoters without any adverse effects on immune responses in broiler diets. The effects of the plant may be due to greater efficiency in feed use, which leads to increased growth and modulation of gut microbiota. With better use of nutrients, it has a positive effect on nutrition, health, and growth of the host as well.

Conflict of interest

The authors declare there is no conflict of interest regarding the current study and its publication.