Citation of this paper

Prebiotic effects of agavins in poultry

Yanelys García Curbelo, R Bocourt Salabarría, Bárbara Rodríguez Sanchez, Nereyda Albelo Dorta, Alejandro Albelo González, Magali Herrera Villafranca¹ and Mercedes G López Pérez²

Departamento de Monogástricos, Instituto de Ciencia Animal (ICA), Carretera central, Km 47 ½, San José de las Lajas, Mayabeque, Cuba
ygarcia@ica.co.cu
¹ Departamento de Biomatemática, Instituto de Ciencia Animal (ICA), Carretera central, Km 47 ½, San José de las Lajas, Mayabeque, Cuba
² Departamento de Biotecnología y Bioquímica, Centro de Investigaciones y Estudios de Avanzadas del IPN-Unidad Irapuato (CINVESTAV), México

Abstract

The objective of this study was to investigate the effects of Agave fourcroydes agavins supplementation on growth performance, carcass characteristic and physiological indicators of broiler. A total of 288 one-day-old Cuban breeder hybrid EB34 male broilers were randomly allocated into three group of eight replicate pens and given a maize-soybean standard diet supplemented with 0, 0.25 and 0.50% of Agave fourcroydes agavins during the 42 days of the experiment. Growth performance, carcass characteristics, morphological and fermentation indicators in cecum and immunological indicators were determined.

The dietary level of 0.5% of agavins increased the concentrations of acetic and butyric acids in the cecum with a lowering of the cecal pH. The weight of the bursa of fabricius was increased linearly with increasing level of agavins in the diet. There was a positive effect of agavin supplementation on hemagglutination inhibition titers to Newcastle diseases. Supplementing broiler diets with agavins improved carcass characteristics but did not affect growth performance.

Key words: prebiotic, agavin, broiler

Introduction

Antibiotics as growth promoter additives (AGP) have been traditionally used in the intensive poultry industry to improve the productive performance and the prevention of diseases (Agostini et al 2012). However, since 2006 began their prohibition in animal nutrition due to the development of antimicrobial resistance. The scientific community research safe and harmless alternatives such as prebiotics in animal nutrition.

Prebiotics are defined as a substrate that is selectively utilized by host microorganisms conferring a health benefit (Gibson et al 2017). The most common prebiotics used in poultry are inulin-type fructans because studies have showed various beneficial effects on animal performance when they are used as prebiotics (Huang et al 2015; Taraz et al 2015).

However, recent research has indicated the potential of agavins oobtained froom Agave as prebiotics (Santiago-García and López 2014; García-Vieyra et al 2014; García-Curbelo et al 2015a), but little is known about the effects of these compounds in animal nutrition. The aim of this study was to investigate the effects of Agave fourcroydes agavins as prebiotic in broiler chickens.

Materials and methods

Animals, experimental design and feeding management

A total of 288, one-day-old Cuban breeder hybrid EB34 male broiler chickens (average body weight 40 ± 0.5g) were used in the trial. All experimental protocols were approved by the Committee of Institute of Animal Science (REG-PNO-PR06 (CyT-P-02-1). The chicks were weighed and randomly assigned to three treatments, each comprising eight replicate cages, with 12 birds per cage.

The treatments were a maize –soybean standard diet supplemented with 0, 0.25 and 0.50% of Agave fourcroydes agavins (STD; 0.25% AF-C; 0.50% AF-C). Agavins from Agave fourcroydes present oligosaccharides with a DP < 10, linkages of the type β (2-1), β(2-6), and branched chains of the neo type (data not shown) (García-Curbelo et al 2015b). The treatments were arranged as a completely randomized design. From 1 to 42 d of age, all the chickens were provided with water and feed ad libitum. A light period of 24 h per day was provided throughout the experiment.

The composition of the diet varied: starter 0-14d; grower 15-34 d; finisher 35-42d, according to the standards of National Research Council (1994). The diets were free of antibiotics and coccidiostats. The animal vaccination system consisted of a dose of Marek, the day after the chickens were hatched, and a dose of Gumboro at 1, 7 and 21 day of age. The Newcastle vaccine was also administered at 14 days of age.

Body weight (BW), feed intake (FI), body weight gain (BWG) and feed conversion ratio (FCR) were determied at the end of the experiment period. Viability rate was recorded on a daily basis.

Sample collection

On days 35 and 42 of the trial, one bird from each cage ( 8 birds of each experimental treatment) were selected according to average BW within the group, individually weighed, and sacrificed by bleeding he jugular vein (Sánchez 1990).

Determination of morphological indicators

Following feather removal, the organs: liver, proventriculus, gizzard, small intestine, cecum, gastrointestinal tract were separated, individually weighed, and expressed as percentage of live body weight.

Determination of fermentation indicators

At 35 d and 42 d the cecal contents were collected, and their pH contents were measured (pH metro AB150). The methodology proposed by Jounay (1982) was used for analysis of short-chain fatty acids (SCFA). Commercially available acetic acid, propionic acid, and butyric acid (Sigma) were used as the external standards.

Determination of immunological indicators

The spleen and burse of Fabricius were excised, weighed and calculated as a percentage of live body weight. Blood samples were collected from the brachial vein of 8 chicks in each treatment al 35 and 42 days of age. The serum was separated from each blood sample by centrifugation and stored at -20 ^oC until use.

The antibody titers against Newcastle in serum were measured according to Hemoglutination inhibiting (HI) test for the Newcastle vaccine by procedure of Wegmann and Smithies (1966).

Carcass characteristics

At the end of the experimental period (42 days), all broilers were weighed individually and sacrificed by bleeding of the jugular vein (Sánchez 1990). The carcass weight was determined after removing the feathers, blood, head, feet, and organs. The breast, leg (thigh+drumstick) were removed from the carcass and individually weighed. The carcass yield was expressed as a percentage of live weight.

Statistical analysis

Results are expressed as mean values with their standard errors of the mean. Statistical variations between the groups were evaluated using one-way ANOVA followed by analysis of simple classification. Duncan’s (1955), multiple-range test was employed in the necessary cases through the use of INFOSTAT (Di Rienzo et al 2012). Significant differences were considered at p < 0.05. The values of viability were analyzed through the proportions comparison with the ComparPro 1.0 program (Font et al 2007).

Results

Growth performance

The initial body weight (40 g/chick) did not different between groups prior to the assignment of the experimental diets (Table 1). At the end of the experiment (42 d), the growth performances of the chickens were not significantly modified by any treatment.

Table 1. Effect of the addition of Agave fourcroydes (AF-C) agavins on growth performance of broilers at 42 d of age
	AF-C treatments			SEM	p
	STD	0.25 %	0.50 %	SEM	p

Body weight, g/bird	2373	2334	2355	44.8	0.83
Feed intake, g/bird	3925.14	3967.00	3956.14	47.6	0.81
Feed conversion, g/g	1.83	1.81	1.85	0.04	0.78
Body weight gain, g	2333	2294	2315	40.1	0.61
Viability, %	97.7	96.0	96.6	1.61	0.02

Morpho-physiological indicators

Only the animals that received agavins showed a significant increase in cecum relative weight, by about 12% and 14% at 35 d and by13% and 17% at 42 d, with the administrations of 0.25% and 0.50% AF-C fructans (Table 2). Other gastrointestinal organs were unaffected by the administration of agavins.

Table 2. Effect of Agave fourcroydes (AF-C) agavins on some morphometric indicators of the gastrointestinal tract of broilers at 35 and 42 days of age
Relative wall weight (g·kg^-1 live weight)	Age (days)	AF-C			SEM	p
Relative wall weight (g·kg^-1 live weight)	Age (days)	STD	0.25 %	0.50 %	SEM	p

Proventriculus	35	5.04	5.32	5.44	0.63	0.83
Proventriculus	42	3.94	3.86	3.03	0.75	0.81

Gizzard	35	21.3	22.7	23.1	0.79	0.72
Gizzard	42	18.32	18.02	18.93	0.83	0.69

Small intestine	35	39.24	41.5	41.92	1.25	0.92
Small intestine	42	38.53	38.10	38.87	1.05	0.83

Two cecum	35	5.72^a	6.42^b	6.56^b	0.15	<0.05*
Two cecum	42	5.21^a	5.91^b	6.13^b	0.05	<0.05*

Liver	35	25.61	25.70	25.54	0.47	0.78
Liver	42	24.84	25.01	24.93	0.52	0.72

Gastrointestinal tract	35	101.37	102.98	103.45	1.33	0.90
Gastrointestinal tract	42	98.03	98.96	99.01	1.95	0.92
^ab Means in each row without common superscripts are different at p<0.05

Fermentation indicators

Agavins supplementation did not influence the cecal pH values at day 35 (Table 3). However, after 42 d, there was a decrease in cecal pH of broilers supplemented with 0.50% AF-C compared to the standard diet.

The different concentrations of Agave fourcroydes agavins had no effect on propionic acid concentration. Nevertheless, the acetic acid concentration was increased at 42 d for birds fed the 0.50% AF-C level. Butyric acid concentration increased with both levels of supplementation of agavins at both sampling times (Table 3 y Figure 1).

Table 3. Effect of Agave fourcroydes (AF-C) agavins on cecal pH and short-chain fatty acids (SCFA) at 35 and 42 d of age
Indicators (mmol.l^-1)	Age (days)	AF-Cs			SEM	p
Indicators (mmol.l^-1)	Age (days)	STD	0.25 %	50 %	SEM	p

pH	35	7.28	6.98	6.95	0.09	0.64
pH	42	7.15^a	6.90^ab	6.85^b	0.05	<0.01**

Acetic	35	9.94	9.88	9.92	0.28	0.70
Acetic	42	11.2^a	11.6^a	12.3^b	0.15	<0.05*

Propionic	35	2.43	2.54	2.58	0.11	0.62
Propionic	42	2.01	2.05	2.06	0.12	0.67

Butyric	35	1.37^a	1.82^b	1.93^b	0.02	<0.05*
Butyric	42	1.25^a	1.40^b	1.42^b	0.04	<0.05*
^ab Means in each row without common superscripts are different at p<0.05

Figure 1. Effect of agavins on acetic and butyric acid cooncentrations in broilers at 42d of age

Immunological indicators

The weight of the bursa of Fabricius was increased by supplementation with 0.50% of AF-C but there was no effect on the relative weight of the spleen (Table 4 and Figure 2).

The Newcastle HI titer at 42d was increased when 0.50% AF-C was added to the diet (Figure 3).

Table 4. Effect of Agave fourcroydes (AF-C) agavins addition on some immunological organs of broilers at 35 and 42 d of age
Relative weight (g·kg^-1 live weight)	Age (day)	AF-C				SEM	p
		STD	0.25 %		A0.50 %

Spleen	35	1.99		2.06	2.01	0.10	0.68
	42	2.06		2.10	2.10	0.08	0.79

Bursa of Fabricius	35	2.15^a		2.25^a	2.87^b	0.11	<0.05*
	42	2.12^a		2.14^a	2.80^b	0.07	<0.05*
^ab Means in each row without common superscripts are different at p<0.05


Figure 2. Effect of agavins on size of Bursa of Fabricius in broilers at 42 d of age	Figure 3. Hemagglutination inhibition (HI) titers to Newcastle disease in chickens at 35 and 42d of age fed a diet supplemented with agavins

Carcass characteristic

The carcass characteristics of broilers are present in Table 5. The effects of agavins diets were significant for carcass and leg weights (P <0.001). There was no significant difference in the weight of breast. Carcass yield was significantly increased in the group with diet supplemented with agavins (P <0.001).

Table 5. Effect of the addition of Agave fourcroydes (AF-C) agavins on carcass characteristic of broilers at 42 d of age
Indicators	AF-C			SEM	p
Indicators	STD	0.25 %	0.50 %	SEM	p

Carcass weight, g	1598.29^a	1647.29^b	1658.78^b	9.57	<0.001***
Breast, g	469.25	508.44	489.25	12.99	0.97
Leg (thigh+drumstick), g	442.13^a	490.38^b	520.63^c	8.44	<0.001***
Carcass yield, %	67.35^a	70.57^b	70.44^b	0.74	<0.001***
^abc Means in each row without common superscripts are different at p<0.05

Discussion

The reports of prebiotics on growth performance in chickens are not consistent (Waldroup et al 1993; Chen et al 2005). Yusrizal and Chen (2003) showed that supplemental inulin improved the growth performance of female but not of male broilers. However, Kim et al (2011) reported that supplementation of prebiotics to the diet improved broiler weight gain when compared to controls, although differences in feed intake, feed conversion and mortality were not observed. It is necessary to consider that prebiotic effects depend on different factors, including physiological and production responses as reported in the literature (Kim et al 2011; García-Curbelo et al 2012).

In our study, the agavins increased cecum weight, regardless of concentration. Agavins are an energy source of for acid lactic bacteria resulting in generation of short-chain fatty acids, which in turn may increase crypt depth and cell proliferation in the cecum (Santos et al 2006). The pH of the cecal contents was lowered for the broilers that consumed 0.50% agavins presumably because of the acetate and butyrate produced by fermentation of the agavins. (Table 3). In previous studies, it was reported that adding agavins of Agave forucroydes in mice diets led to increases in butyrate in the cecal digesta (García-Curbelo et al 2015a). Rehman et al (2008) reported that adding inulin to broiler diets increased the butyrate concentration in the cecum. Butyrate provides energy for epithelial growth (Topping and Clifton, 2001) and is involved directly or indirectly in various mechanisms such as cellular differentiation, growth and permeability, and gene expression (Mroz et al 2006).

The immunomodulatory properties of prebiotic have been described in different researches (Dubert-Ferrandon et al 2008; Madej et al 2015). Our study showed that the agavins at 0.50% of the diet increased the relative weight of the bursa of Fabricius, an organ specialized in the development of B lymphocytes in birds and in antibodies for the Newcastle vaccine. These results confirm the need for a dose-response studies of prebiotic action as suggested by the data in Figure 2. Perez (2000)reported that broilers fed β-glucans at 100 ml/g diet showed an increase of antibodies for Newcastle vaccine. The relative weight of the bursa of Fabricius was also related with the doses and their values were major in the group of high dose. These results indicate the effect of the prebiotics dose in some indicators of the animal response.

The reasons why supplements of agavins might have enhanced the immune system of broilers were not investigated in this study, but it is known that prebiotic compounds prevent the colonization of pathogens through adhering to, and blocking , of the intestinal surface, stimulating the intestinal immune-competent cells, associated to the lymphoid issue (Seifert and Watzl 2008; Vidanarachchi et al 2013).

The major effects of the agavins on the carcass were the increases in leg and carcass yield. The use of prebiotic additives guarantees a state of eubiosis that improves the digestibility of nutrients and makes possible a higher rate of absorption, contributing to the development of muscles. Our results do not coincide with the investigations of Midilli et al (2008) who found that dietary supplementation with probiotics and prebiotics in broilers did not affect carcass yield. The different responses to prebiotic in animals has relationship with genetic, feed, exploitation conditions, sex and weight (Havenstein et al 2003; Brickett et al 2007).

Conclusions

The dietary level of 0.5% of agavins increased the concentrations of acetic and butyric acids in the cecum with a lowering of the cecal pH.
The weight of the bursa of Fabricius was increased linearly with increasing level of agavins in the diet.
There was a positive effect of agavin supplementation on hemagglutination inhibition titers to Newcastle disease.
Supplementing broiler diets with agavins improved carcass characteristics but did not affect growth performance.

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