Ameliorative Effects of Mannan Oligosaccharide and Sodium Butyrate on Production Performance and Immunity During Combined Aflatoxicosis and Ochratoxicosis in Broiler Chickens

Abstract

To evaluate the ameliorative impact of mannan oligosaccharide (MOS) and sodium butyrate (NaB) during combined aflatoxicosis and ochratoxicosis on production performance and immunity, day-old broiler chicks (n=320) were divided into 10 treatment groups (T0: control; T1: T0+300 ppb aflatoxin B1 (AFB1)+250 ppb ochratoxin A (OTA); T2: T1+0.15% MOS; T3: T1+0.3% MOS; T4: T1+500 mg/kg NaB; T5: T1+1000 mg/kg NaB; T6: T1+0.15% MOS+500 mg/kg NaB; T7: T1+0.15% MOS+1000 mg/kg NaB; T8: T1+0.3% MOS+500 mg/kg NaB; T9: T1+0.3% MOS+1000 mg/kg NaB). Each diet was fed to 4 replicated groups of 8 birds each for 42 days. During overall growth phase (0-6 wk) the body weight gain (BWG) and feed intake (FI) in T0 was higher (P<0.05) than T1. The BWG in T8 was similar to T0. The BWG in all other groups was higher (P<0.05) than T1 but could not match with T0. The FI in T2, T3, T6, T7 and T8 was statistically comparable to T0. The feed conversion ratio (FCR) of group T8 was statistically (P<0.05) similar to T0. The FCR in other groups was lower (P<0.05) than T1, but did not match with T0. The CMI and HA titre in T1 was lower (P<0.05) than T0. The CMI in T3 and T8 groups was similar to T0. The HA titre of T2, T3 and T8 groups was similar to T0. It was concluded that mannan oligosaccharides at 0.15 and 0.3% levels and sodium butyrate at 500 mg/kg diet proved beneficial in alleviating both aflatoxicosis and ochratoxicosis partially, while the combination of 0.3% MOS+500 mg/kg NaB was effective in counteracting mycotoxicosis maximally, as accessed through production performance and immunity in broiler chickens.

Keywords: Aflatoxicosis, Ochratoxicosis, Production performance, Immunity, Mannan oligosaccharides, Sodium butyrate, Broiler chicken.

1. Introduction

Presence of mycotoxins in feed is one of the major constraints in maintaining feed quality, especially in tropical country like India. The toxic effects of mycotoxins i.e. mycoxicosis, is characterized by its nephrotoxic, immunosuppressive, hepatotoxic, carcinogenic, mutagenic and teratogenic effects in animals and poultry (Patil et al., 2005, 2006, 2014, 2017a, b; Katole et al., 2013; Patial et al., 2013; Patel et al., 2015; Patil and Degloorkar, 2016a, b, 2018; Singh, 2019a-f; Singh et al., 2019a, b). Aflatoxicosis in poultry causes lowered performance in terms of reduced body weight gain, feed intake and feed efficiency (Silambarasan et al., 2013; Singh et al., 2015a; Singh et al., 2016c), reduced nutrient utilisation (Silambarasan et al., 2013), increased mortality (Khatke et al., 2012b; Sharma et al., 2014), anemia (Singh et al., 2015a; Singh et al., 2016c), hepatotoxicosis and haemorrhage (Churchil et al., 2014; Singh et al., 2015a; Singh et al., 2016c; Pathak et al., 2017), altered biochemistry (Singh and Mandal, 2013; Singh et al., 2013a). Moreover, it impairs humoral and cellular immune responses in poultry and increases susceptibility to environmental and infectious agents (Khatke et al., 2012a; Singh, 2019a, b, e, f) leading to severe economic losses. Use of mannan oligosaccharide (MOS) and Saccharomyces cerevisiae (SC) (@ 0.05%, 0.1%, 0.2%) alone or their combination moderately ameliorated the adverse effects of 300 ppb AFB1 (Khatke et al., 2012a, b, c). The 0.2% level of MOS and SC was more effective than 0.05% and 0.1% level in counteracting aflatoxin in the feed. MOS appeared to be more efficacious than SC in counteracting aflatoxicosis in broiler chickens.

However, the combination of SC and MOS did not show any synergistic effect in counteracting aflatoxicosis (Khatke et al., 2012c). Ochratoxicosis causes a reduction in production performance viz. reduced growth rate, decreased feed consumption, poorer feed conversion (Singh et al., 2016b) and increased mortality (Singh et al., 2015b, 2016b). Ochratoxin contamination at 200 ppb level in broiler diet led to decreased protein, haemoglobin and creatinine, while increased uric acid, alkaline phosphatase, AST and ALT levels in blood. Moreover, ochratoxin impaired both cell mediated and humoral immunity (Singh et al., 2016a). Short chain fatty acids production in the poultry gut supports a healthy gut wall by promoting gut healing and among those short chain fatty acids butyric acid is most important one. MOS also eliminates the mycotoxins from the gut of livestock and poultry by physical adsorption (Zaghini et al., 2005). In the present study, therefore, such agents that improve the microbial and structural gut health, and improve general health and immunity such as mannan oligosaccharide and Sodium butyrate have been chosen for experimentation.

2. Materials and Methods

2.1 Production and Analysis of Mycotoxins

Aflatoxin was produced by growing Aspergillus flavus NRRL 6513 on maize. This fungus was obtained from US Department of Agriculture, Illinois, USA. Fermentations were carried out in batches as per the method described by Shotwell et al. (1966). The extraction and estimation of aflatoxin B1 was done as per the procedure of Pons et al. (1966). Aflatoxin contents were finally quantified using a UV spectrophotometer. Ochratoxin was produced as per the method described by Singh et al. (2013b) using Aspergillus westerdijkiae NRRL 3174. This fungus was also obtained from US Department of Agriculture, Illinois, USA. The extraction and estimation of ochratoxin A was done as per the procedure of AOAC (1995).

2.2 Experimental Design

Experimental design was completely randomized design. There were ten dietary treatments. Each dietary treatment had 4 replicates and each replicate had 8 chicks. The experiment was conducted in broiler chickens for 6 weeks of age. The various dietary treatments were prepared by mixing the required quantity of mycotoxins, MOS and NaB to get their desired concentration in basal diet (Table 1).

Table 1: Experimental groups and treatments

Ameliorative Effects of Mannan Oligosaccharide and Sodium Butyrate on Production Performance and Immunity During Combined Aflatoxicosis and Ochratoxicosis in Broiler Chickens - Image 1

2.3 Biological

Experiment and Analysis Day-old broiler chicks were obtained from experimental hatchery, ICAR-Central Avian Research Institute, Izatnagar. The chicks were wing banded, weighed individually and distributed randomly into ten treatment groups on equal weight basis. All birds were reared under standard management conditions and fed with broiler starter ration for 1-21 days and broiler finisher ration from 22 to 42 days. The composition of broiler starter and finisher ration was as below: The starter diet with maize 53.93, soybean meal 42.70, limestone 0.9, dicalcium phosphate 1.70, common salt 1.30, DL-methionine 0.16, TM premix 0.10, vitamin premix 0.15, B complex 0.015, choline chloride 0.05; and finisher diet with maize 63.63, soybean meal 33.40, limestone 0.95, dicalcium phosphate 1.40, common salt 0.20, DL-methionine 0.11, TM premix 0.10, vitamin premix 0.15, B complex 0.015, choline chloride 0.05 were formulated. TM premix supplied Mg, 300; Mn, 55; I, 0.4; Fe, 56; Zn, 30; Cu, 4 mg/kg diet. Vitamin premix supplied Vit A, 8250 IU; Vit. D3, 1200 ICU; Vit. K, 1mg; vitamin E 40 IU per kg diet. B complex supplied Vit. B1, 2 mg; Vit. B2, 4 mg; Vit. B12, 10 mcg; niacin, 60 mg; pantothenic acid, 10 mg; choline, 500 mg per kg diet. The starter diet contained 22.09% crude protein, 2,848 Kcal ME/kg, calcium 0.99%, available P 0.45%, lysine 1.27%, methionine 0.53% and threonine 1.01%. The corresponding values in finisher diet were 19.04%, 2,946 Kcal/kg, 0.90%, 0.38%, 1.05%, 0.44% and 0.87%. The protein as per AOAC (1995) and calcium contents as per Talapatra et al. (1940) were estimated, while the concentrations of lysine, methionine, threonine, available P and metabolizable energy values were calculated. Weekly individual body weight and feed consumption of each group were recorded and the FCR was calculated.

Humoral immune response to sheep red blood cells (SRBCs) and cellular immunity to PHA-P was observed on 30th day and 24th day of experimental trial, respectively.

2.4 Statistical Analysis

All data were statistically analyzed using SPSS software package version 20.0 following one way analysis. All the observations were recorded at 95% (P<0.05) level of significance.

3. Results and Discussion

3.1 Growth Performance

The data pertaining to effect of various dietary treatments on average body weight gain (BWG), feed intake (FI) and feed conversion ratio (FCR) of broilers in different growth phases is presented in Table 2.

3.1.1 Body Weight Gain (BWG)

During starter phase of growth (0-3 wk) the BWG of birds in control group (T0) was significantly (P<0.05) reduced due to feeding combination of AFB1 and OTA in T1. The BWG among groups T2, T6 and T7; T3 and T8; T3 and T6; T4 and T7; and T4, T5 and T9 was statistically similar. The BWG of groups T1 and T5 did not vary significantly. The BWG of T8 was statistically (P<0.05) similar to that of control group (T0). With regards to finisher phase (4-6 wk) of growth trial, the BWG of control group (T0) was highest among all other dietary treatment barring T8, which was statistically (P<0.05) similar to that of control group. The BWG of mycotoxins fed group without any dietary supplementation (T1) was statistically (P<0.05) similar to groups T4 and T5. The BWG was statistically (P<0.05) similar among dietary treatments T2, T3 and T7; T3 and T9; T4, T5 and T9; T6 and T7. During overall growth phase (0-6 wk) the BWG of broiler in control group (T0) was 1681.69 g as against 1369.66 g in toxins fed group (T1) which was significantly (P<0.05) lower. The overall BWG in groups T8 was statistically similar to that of control (T0), however the weight gain in all other groups was higher (P<0.05) than T1 but could not match with that of control (T0). The overall BWG between groups T2 and T7; T3 and T6; T4 and T9; T5 and T9 was statistically similar. The present study indicated that inclusion of mycotoxins (300 ppb AFB1+250 ppb OTA) in diet caused severe depression in BWG of birds. Reduced BWG is a common feature of aflatoxicosis and ochratoxicosis in poultry (Singh, 2019c; Silambarasan et al., 2013; Singh et al., 2015a; Singh et al., 2016c; Khatke et al., 2012b; Sharma et al., 2014; Singh et al., 2016a; Singh et al., 2015b; Singh and Mandal, 2013; Singh et al., 2013a; Singh et al., 2016b). Results of the present study revealed that dietary supplementation of both MOS and NaB were able to ameliorate the adverse effect of mycotoxins and partially improved the BWG as compared to control birds. Improvement in BWG due to supplementation of MOS on aflatoxin or ochratoxin contaminated diet was in agreement with previous investigation by different researcher.

Use of MOS and SC (@ 0.05%, 0.1%, 0.2%) alone or their combination moderately ameliorated the adverse effects of 300 ppb AFB1 on BWG of broilers (Khatke et al., 2012b). Jahanian et al. (2016) found that the retarded average daily gain (ADG) was ameliorated by inclusion of 2 g/kg of MOS into the diet of 2 ppm aflatoxin-challenged broiler chicks. Oguz and Parlat (2004) also reported significant improvement in BWG of growing Japanese quails due to MOS supplementation in diet during experimental aflatoxicosis. Singh et al. (2016b) reported that addition of SC at 0.1% level to the 200 ppb ochratoxin contaminated feed ameliorated the adverse effects of ochratoxin on body weight gain in broiler chickens. Contrary to this study, Santin et al. (2003) observed that the cell wall of SC at 0.1 levels did not improve the BWG in the presence of the 500 ppb OTA. It might be due to the higher level (500 ppb) of ochratoxin taken for dietary contamination in their experiment. In the present study, this result is because of adsorbent capacity of MOS (Jouany et al., 2005) and also improved growth performance by supplementing MOS might be attributed to their inhibitory effect on pathogen colonization in the gastrointestinal tract (Jahanian et al., 2016). The information on effect of sodium butyrate salt supplementation on BWG during mycotoxicosis is lacking in literature but it was found that dietary supplementation of NaB at the level of 500 mg/kg enhanced growth performance by stimulating the growth of duodenal mucosa in broilers (Hu and Guo, 2007). Growth promoting effect during mycotoxicosis might be possible as NaB enhanced the anti-inflammatory and anti-catabolic effects together with its role on anti-immune stress (Chamba et al., 2014).

3.1.2 Feed Intake (FI)

During 0-3 wk of age, the FI in control group (T0) was significantly (P<0.05) higher than that of mycotoxins alone fed group (T1). The FI of other treatment groups was statistically similar to that of control barring T4 and T5, however, the weight gain in these groups (T4 and T5) was higher (P<0.05) than T1. During finisher phase (4-6 wk) of growth trial feed consumption among groups T0, T2, T3, T6, T7 and T8 was statistically similar and higher than remaining treatment groups. The FI of group T1 was the lowest and statistically similar to that of groups T5 and T9. The feed consumption of group T4 was statistically higher than T1 but did not match to that of control group (T0). During overall growth period (0-6 wk), the FI in control group was higher than that of mycotoxins alone fed group (T1). The FI in groups T2, T3, T6, T7 and T8 treatment groups was statistically comparable to that of control (T0). The feed consumption of groups T4 and T9 was higher (P<0.05) than that of T1 but not similar to that of control group (T0). The cumulative FI of group T5 was statistically similar to that of toxin alone fed group (T1). In the present study, reduced FI due to aflatoxicosis and ochratoxicosis was in agreement with those of earlier reports available in the literature (Singh, 2019c; Silambarasan et al., 2013; Singh et al., 2015a; Khatke et al., 2012b; Sharma et al., 2014; Singh et al., 2016a; Singh et al., 2016c; Singh et al., 2015b; Singh and Mandal, 2013; Singh et al., 2013a; Singh et al., 2016b). 500 mg/kg NaB salt and 0.3% MOS along with 1000 mg/kg NaB salt partially ameliorated the adverse effect of mycotoxicosis on FI of birds. There was no ameliorative effect of 1000 mg/kg NaB salt supplementation on mycotoxins contaminated diet of broiler chickens. This might be due to bad or pungent odour of NaB, which inhibit the further FI along with mycotoxins. MOS was reported to have the toxin binding properties (Stanley and sefton, 2000) and detoxify aflatoxin. Devegowda et al. (1996, 1998) observed that a commercial MOS binds AFB1. Zaghini et al. (2005) also demonstrated the ability of MOS to adsorb and degrade AFB1 and reducing gastrointestinal absorption of AFB1.

Table 2: Body weight gain (g/bird), feed intake (g/bird) and FCR in different growth phases of broilers fed various dietary treatment

Addition of MOS to the AF-containing diet significantly reduced these adverse effects of AF on feed consumption of growing Japanese quails (Oguz and Parlat, 2004). In agreement with our findings, Jahanian et al. (2016) reported that MOS (2g/kg diet) alleviated the adverse effect of 0.5 ppm AFB1 on feed consumption of broilers. Khatke et al. (2012b) also reported that use of MOS and SC (@ 0.05%, 0.1%, 0.2%) alone or their combination moderately ameliorated the adverse effects of 300 ppb AFB1 on feed consumption of broilers. Singh (2015) observed that addition of SC to the ochratoxin contaminated feed ameliorated the adverse effects of ochratoxicosis on feed consumption of broiler chickens. Similarly, ElBarkouky (2008); El-Barkouky et al. (2010) also reported that addition of SC (3g/Kg feed) to the 50-200 ppb OTA contaminated diet improved the feed consumption in broiler chickens.

Further, it was also observed that sodium butyrate supplementation in the diet of broilers was beneficial, especially in the presence of immune stress during mycotoxicosis and this might be ascribed to the ability of NaB to maintain normal feed intake and decrease catabolism and damage of tissues (Zhang et al., 2011) which was generally seen during mycotoxicosis (300 ppb AFB1+250 ppb OTA).

3.1.3 Feed Conversion Ratio (FCR)

The FCR during 0-3 wk in control group (T0) was significantly (P<0.05) lower than that of mycotoxins fed group T1. The FCR in treatment groups T3 and T8 was statistically (P<0.05) similar to that of control, indicating that supplementation of 0.3% MOS alone or with 500 mg/kg NaB ameliorated the adverse effects of combine mycotoxicosis (300 ppb AFB1+250 ppb OTA) on feed efficiency of in broiler chickens.

The FCR in remaining groups was statistically higher than that of control (T0). During 4-6 wk of growth period, the FCR of group T5 was statistically similar to that of toxins fed group (T1) and higher than that of control. FCR of group T8 was statistically (P<0.05) similar to that of control. The FCR among remaining groups was statistically lower than T1, however, FCR were not comparable to that of control (T0). The results of overall FCR (0-6 weeks) showed similar trend as finisher phase (4-6 wk). The present study revealed that combined mycotoxins (300 ppb AFB1+250 ppb OTA) contamination in feed significantly (P<0.05) increased the FCR compared to that of control and resulted in poor feed efficiency in broiler chickens.

The present study revealed decreased feed efficiency due to aflatoxicosis and ochratoxicosis. Similar findings of decreased feed efficiency due to feeding of aflatoxin or ochratoxin were earlier reported by several researchers (Singh, 2019c; Silambarasan et al., 2013; Singh et al., 2015a; Khatke et al., 2012b; Sharma et al., 2014; Singh et al., 2016a; Singh et al., 2016c; Singh et al., 2015b; Singh and Mandal, 2013; Singh et al., 2013a; Singh et al., 2016b). The present study further revealed that supplementation of 0.3% MOS along with 500 mg/kg NaB to the mycotoxins contaminated feed ameliorated the adverse effects of combined mycotoxicosis on feed efficiency in broiler chickens.

Different levels of MOS (0.15% and 0.3%) and 500 mg/kg NaB and their different combinations partially ameliorated the adverse effect of mixed mycotoxicosis (AFB1 and OTA) on FCR. However, 1000 mg/kg NaB had no ameliorative effect on FCR in our study. The reason might be the intense odour of NaB salt, which reduced the feed intake by broiler chickens. Jahanian et al. (2016) reported that increasing supplemental MOS level 1 g/kg to 2 g/kg ameliorated the worsen FCR values in aflatoxin-challenged broiler chicks and it was due to dietary supplementation of MOS resulted in increases in average daily feed intake and average daily gain, consequently improved FCR values in broiler chicks exposed to aflatoxin challenge (0.5 and 2 ppm). Khatke et al. (2012b) also reported that MOS and SC (@ 0.05%, 0.1%, 0.2%) alone or their combination moderately ameliorated the adverse effects of 300 ppb AFB1 on feed efficiency of broilers. Singh et al. (2016b) found addition of SC (0.05% and 0.1%) to the 200 ppb ochratoxin contaminated feed ameliorated the adverse effects of ochratoxicosis on feed efficiency in broiler chickens. El-Barkouky (2008); El-Barkouky et al. (2010) also reported that addition of active dried yeast S. cerevisiae at 3 g per kg feed in the diet of broiler chickens ameliorated the ill effects at 50-200 ppb OTA.

Unlike to our findings, Santin et al. (2003) observed that the cell wall of SC did not ameliorate the feed intake, weight gain and feed conversion of birds which was impaired due to dietary addition of OTA (500 ppb). Aravind et al. (2003) also observed that esterified glucomannan (0.05%) effectively alleviated the growth depression caused by the naturally contaminated (aflatoxin 168 ppb, ochratoxin 8.4 ppb, zearalenone 54 ppb, and T-2 toxin 32 ppb) diet of broilers.

3.2 Immune Response

The data pertaining to CMI response to PHA-P measured as foot web index and humoral immune response measured as haemagglutination titre (HA) against SRBCs of broiler chickens fed different dietary treatments was statistically analyzed and presented in Table 3.

Table 3: Cellular and humoral immunity of broilers fed different treatments

3.2.1 Effect on Cell Mediated Immunity (CMI)

The CMI value in control group (T0) was 0.72 mm which significantly (P<0.05) decreased to 0.36 mm in the mycotoxins (300 ppb AFB1+250 ppb OTA) alone fed group (T1). The CMI value in T3 and T8 groups was statistically similar to that of control. The CMI value in groups T2, T4, T5, T7 and T9 was statistically similar to that of group T1. The CMI values of group T6 did not vary significantly (P<0.05) from group T0 and T1, indicating that supplementation of 0.15% MOS+500 mg/kg NaB salt in diet ameliorated the ill effects of mycotoxin on CMI response. In the present study, reduced CMI response in birds due to aflatoxicosis was in agreement with those of Singh (2019a, b). The result of present study indicated that combine feeding of 300 ppb AFB1 and 250 ppb OTA caused reduction in mean CMI value and supplementation of 0.3% MOS alone and along with 500 mg/kg NaB to the mycotoxins contaminated diet ameliorated the adverse effects of mycotoxins on CMI. Khatke et al. (2012c) also reported that MOS and SC (@ 0.05%, 0.1%, 0.2%) alone or their combination moderately ameliorated the adverse effects of 300 ppb AFB1 on CMI response in broiler chickens. Singh (2015) reported that inclusion of SC (0.05 and 0.1%), partially ameliorated the ill effects of OTA on CMI response. Khalil (2008) also observed that dietary inclusion of SC (2 g/kg) ameliorated the negative effects of OTA in growing quails. In addition, several workers explain the function of butyrate on immune system. Leeson et al. (2005) observed that butyrate stimulates the immune system in poultry and observed that when butyrate fed to the birds, a better withstand against the stress of coccidial challenge at 21 day of age was reported. In the lower part of the intestinal tract particularly epithelium of the ileum, caeca and colon butyrate stimulates the specific G-protein-coupled receptors, specifically GPR 41 and GPR 43 (Le Poul et al., 2003). When butyrate is attached to these receptors the production of several different peptides is stimulated (Tazoe et al., 2008; Cox et al., 2009). Some of these peptides have a positive effect on the development of the immune system and improve the functioning of the immune system in case of a health challenge (Cox et al., 2009).

3.2.2 Humoral Immune Response

The HA titre value in mycotoxins (300 ppb AFB1+250 ppb OTA) alone fed group (T1) was significantly (P<0.05) lower than that of control (T0). In the present study, reduced humoral immune response in birds due to aflatoxicosis was in agreement with those of Singh (2019a, b). The HA titre in groups T2, T3 and T8 groups was statistically similar to that of control. The HA titre of group T7 did not vary significantly (P<0.05) from group T0 and T1, indicating that supplementation of 0.15% MOS+1000 mg/kg NaB in diet partially ameliorated the adverse effect of mycotoxin on HA titre. Khatke et al. (2012c) also reported that MOS and SC (@ 0.05%, 0.1%, 0.2%) alone or their combination moderately ameliorated the adverse effects of 300 ppb AFB1 on humoral immune response in broiler chickens. The HA titre in groups T4, T5, T6 and T9 was statistically similar to that of group T1. In the present study, dietary mycotoxins (300 ppb AFB1+250 ppb OTA) significantly (P<0.05) reduced the HA titre against SRBC’s and incorporation of 0.15% MOS and 0.3% MOS alone and along with 500 mg/kg NaB salt ameliorated the ill effect of mycotoxicosis on humoral immune response in broiler chickens. El-Barkouky (2008) also found that inclusion of SC at 0.1% improved humoral immune response which was negatively affected by 50 and 100 ppb OTA. Similarly, Singh (2015) found that supplementation of SC (0.05 and 0.1%) to 200 ppb OTA contaminated feed partially ameliorated the ill effects of ochratoxicosis on humoral immune response in broiler chickens.

4. Conclusion

It was concluded that mannan oligosaccharide at 0.15 and 0.3% level and sodium butyrate at 500 mg/kg diet was proved beneficial in reducing both aflatoxicosis and ochratoxicosis partially, while the combination of 0.3% MOS+500 mg/kg NaB was effective in counteracting their toxic effect maximally, as accessed through production performance and immunity in broiler chickens.

This article was originally published in Livestock Research International | April-June, 2019 | Volume 07 | Issue 02 | Pages 107-115.

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