Growth, Nutrient Digestibility of Broilers fed Aflatoxin Contaminated Diets with Aflatoxin Binders 1 A. D. Ologhobo, 2 E. O. Ewuola, 3 U. U. Jerome, 4 U. O. Franca, 5 O. Ifarajimi 1, 2,3,4,5 Poultry Research Unit, Department of Animal Science, University of Ibadan ABSTRACT This study examined the effects of dietary Aflatoxin contamination and the efficacy binders on nutrient digestibility, feed conversion ratio, carcass yield, haematology, and serum biochemistry of broilers. 250 AborAcre strain broiler were randomly allotted into 5 dietary treatments, each comprising 50 birds of 10 birds per replicate for 8 weeks. A completely randomized design (CRD) arrangement containing 5 dietary treatments of: 1) +ve control without Aflatoxin and binders, 2) -ve control with Aflatoxin no binders, 3) diet + Montmorillonite, 4) diet + HSCAS and 5) diet + SB. With 2000ppb Aflatoxin, 2g Aflatoxin binder per kg feed, for a period of 8 weeks and a completely randomized design (CRD). The performance characteristics were significantly (p 0.05) influenced by the dietary treatments. The final live weight was highest (p<0.05) in birds fed Positive control diet (diet 1) and least (p<0.05) in birds fed diet 5 that contain SB binder compared to others. Relative liver weight was significantly (p<0.05) higher in birds fed diets 2, 3 and 5 than the control birds. The liver weight of birds fed diets 4 was not significantly different from the control. The serum proteins of birds at 56 days were not significantly different across the treatments groups. Keywords: Broilers, diets, Aflatoxin, binders 1. INTRODUCTION Poultry production in the last three decades has made a tremendous progress. However, in the recent years the growth is not as high as expected because of the challenges of many unknown diseases faced by the poultry industry [1]. Considerable importance in the aetiology of many unknown diseases with respect to mycosis and mycotoxicosis in poultry have been noticed. The problem of mycotoxicosis in the health of human and livestock has assumed a much importance, only after the outbreak of Turkey X disease in 1960 [2]. Mycotoxins refer to secondary metabolites of certain fungi which are toxic to humans, animals and plants health as well. Depending on the concentration, these may be carcinogenic, mutagenic, teratogenic or oestrogenic with the result of reduced immune response and acute disease syndrome. Mycotoxin contamination is very costly for the animal industry and is a food safety concern because of potential mycotoxin residue in meat, dairy and eggs [3, 4]. Aflatoxins are one of the most dangerous mycotoxins known due to their high toxicity to both animals and humans. These mycotoxins are known to have strong hepatotoxic and carcinogenic effects mainly in the monogastrics, which feed on high quantities of cereal grains. Aflatoxins are secondary metabolites of mycotoxins produced by the fungi Aspergillus spp. (flavus and parasiticus), These mycotoxins contaminate a wide variety of tropical and subtropical agricultural feedstuffs including oilseed meals, dried fruits, spices, and cereals, when environmental conditions are favourable. Among the various types of aflatoxins, Aflatoxin B1 (AFB1) is most commonly encountered, biologically active component and it is also considered to have higher toxicity than other aflatoxins. With the global shortage of grains and increase in International Agricultural Trade, mycotoxin contamination has become more of a reality for animal producers worldwide. Aflatoxicos is has a wide variety of effects on animals, including weight loss and poor performance, changes in the clinical biochemistry panel, and an increase in susceptibility to infectious diseases. Recent studies indicated that Aflatoxin affects hepatic gene expression [5] weakens gut morphology and function. Poor growth and feed conversion, increased mortality, leg problems and carcass condemnations are some of the economic losses associated with Aflatoxin exposure in broilers. Aflatoxin is known to cause a wide range of metabolic changes in poultry associated with reduced digestive enzyme activities, immunosuppression and liver damage [6]. The nature of crop production and feed distribution system in the poultry industry, the avoiding feed with preformed Aflatoxin contamination is difficult an economical solution would be the incorporation of a dietary additive, Aflatoxin binder [7]. The use of adsorbing agents, which can trap the Aflatoxin molecule by means of ion exchange and thereby hindering their absorption into blood from the gastrointestinal tract, has gained much attention in prevention of aflatoxins. Hydrated sodium calcium aluminosilicate (HSCAS) [8, 9], bentonite [10], zeolite ore compounds [11], activated charcoal [12], inorganic sorbents [13] and a blend of organic acids and aluminosilicates have shown considerable promise in preventing aflatoxicosis. However, the efficacy of these products and their effectiveness in binding to Aflatoxin is not always known and their model of action not well understood. Hence, study was carried out to evaluate the effects of dietary Aflatoxin contamination with binders on nutrient digestibility, feed conversion ratio, carcass yield, haematology, and serum biochemistry of broiler chickens. 257
2. MATERIALS AND METHODS 2.1 Experimental Location This research work was conducted at the Poultry Research Unit, Teaching and Research Farm, University of Ibadan, Ibadan for a period of 8 weeks. The Teaching and Research Farm is located at latitude 7 o 20 o N and longitude 3 o 50 o E, 200 meters above sea level with average day time temperature of 20 o C - 25 o C and relative humidity of 80-85%. 2.2 Experimental Animals and diets A total of 250, one-day-old Abore Acre strain broiler chicks were purchased from a reliable commercial hatchery All birds were housed and brooded together for one week, after which they were weighed and randomly allotted into 5 dietary treatments: 1) +ve control diet no Aflatoxin and binder; 2) 2000ppb Aflatoxin and No binder; 3) 2000ppb Aflatoxin and 0.2kg Montmorillonite/100kg feed; 4) 2000ppb Aflatoxin and 0.2kg HSCAS/100kg feed; 5) 2000ppb Aflatoxin and 0.2kg SB/100kg feed. Each treatment consisted of 50 birds of 5 replicates per treatment and 10 birds per replicate. Feed was supplied ad libitum and water was made available during the experimental period. A common corn-soybean meal was compounded. Weighed amounts of the Aflatoxin contaminated maize were added to the diet at a concentration to give 2000ppb Aflatoxin. Routine health and vaccination practices were strictly carried out according to recommendation. The health condition of all birds was closely monitored. Daily check was carried out on the birds for any health challenges. 2.3 Production and Quantification of Aflatoxin The fungal culture used was Aspergillus flavus (NRRL 2999) for Aflatoxin production. Aflatoxin in maize was produced by employing solid substrate fermentation methods. Sterile maize was inoculated with a spore suspension of Aspergillus flavus (NRRL 2999) and the treated maize was then incubated for 7 days with incremental increases in the incubation temperature. The maize was then autoclaved to kill the mould, washed, dried, and ground to fine particles. Aflatoxin was extracted from weighed sample of the ground maize powder with chloroform and its concentration quantified by Thin Layer Chromatography (TLC) at the Pathology unit, International Institute for Tropical Agriculture (IITA), Ibadan. A weighed amount of the treated maize was incorporated into the basal diet to produce the desired level of 2000ppb/kg in the different treatment. 3. RESULTS AND DISCUSSIONS Table 1: weight gain (g) and Feed intake (g) of broilers fed different Aflatoxin Parameters 1(+ve control) 2(-ve control) 3(Mont) 4(HSCAS) 5(SB) Heart 0.40±0.02 0.48±0.02 0.47±0.02 0.44±0.04 0.49±0.03 Liver 1.98±0.07 b 3.68±0.54 a 3.44±0.18 a 2.81±0.17 ab 3.67±0.30 a Lungs 0.49±0.02 b 0.56±0.04 ab 0.56±0.04 ab 0.52±0.03 ab 0.61±0.05 a Kidney 0.62±0.01 c 0.93±0.08 ab 0.93±0.04 ab 0.78±0.04 bc 1.03±0.08 a Adrenal gland 0.03±0.01 0.03±0.01 0.04±0.01 0.03±0.01 0.04±0.02 Pancreas 0.22±0.02 0.25±0.02 0.25±0.02 0.24±0.02 0.21±0.02 Spleen 0.11±0.01 b 0.14±0.02 ab 0.16±0.03 ab 0.15±0.01 ab 0.17±0.01 a GIT 9.51±0.41 b 13.75±0.56 ab 14.12±0.50 ab 13.22±0.96 b 21.79±5.96 a Thymus 0.11±0.01 0.07±0.01 0.09±0.02 0.11±0.03 0.12±0.02 Abdominal Fat 1.51±0.38 a 1.53±0.15 b 0.62±0.18 b 0.87±0.22 ab 0.59±0.24 b Bile 0.13±0.02 0.15±0.02 0.18±0.03 0.15±0.01 0.17±0.02 Bursa 0.06±0.01 b 0.08±0.01 ab 0.07±0.01 ab 0.10±0.01 a 0.07±0.01 ab Gizzard 2.00±0.15 b 2.46±0.12 ab 2.39±0.27 ab 2.26±0.19 ab 2.67±0.26 a abc: mean in the same row with different superscripts are significantly (P<0.05) different. Table 2: Relative Organs weight of Broilers fed different Aflatoxin Binders Parameters T1(+ve control) T2(-ve control) T3(Mont) T4(HSCAS) T5(SB) WG/WK 10.87±1.54 a 5.33±0.88 b 5.25±0.94b 4.04±0.90 b 4.51±1.05 b WGING/WK 10873.75±1454.62 a 5328.75±879.94 a 5245.00±937.75 b 4041.25±901.46 b 4508.75±1046.73 b WG/DAY/G 1553.39±207.80 a 761.25±125.71 b 749.29±133.94 b 577.32±128.78 b 644.11±149.53 b WG/BIRD/Day 31.07±4.6 a 16.28±2.61 b 21.39±2.25 b 15.95±2.46 b 14.97±3.36 b WFI 0.60±0.09 0.41±0.06 0.50±0.09 0.48±0.07 0.48±0.07 WFI/G 602.00±94.36 406.88±62.00 496.25±87.52 475.75±74.50 482.38±67.16 DFI 86.00±13.48 58.13±8.86 70.89±12.50 67.96±10.64 68.91±9.59 FCR 3.29±0.72 4.29±0.85 3.86±1.14 3.82±0.87 5.88±1.84 FCE 41.07±8.10 31.30±6.56 37.95±7.27 30.20±6.92 24.53±6.81 Mortality (%) 2 18 44 38 24 abc: mean in the same row with different superscripts are significantly (P<0.05) different. 258
Table 3: Hematological parameters and serum biochemistry of broiler chickens fed different Aflatoxin binders at 56th day Parameters 1(+ve control) 2(-ve control) 3(Mont) HSCAS) 5(SB) PCV (%) 26.70±1.01 ab 27.00±0.61 ab 24.80±1.06 b 28.90±0.96 a 27.10±1.94 ab Hemoglobin (g/dl) 8.86±0.33 ab 8.98±0.21 ab 8.23±0.35 b 9.61±0.32 a 9.00±0.64 ab Erythrocytes (x10 6 /µl) 3.38±0.19 3.55±0.11 3.66±0.09 3.62±0.21 3.74±0.14 Leukocytes (x10 3 /µl) 20.26±3.98 bc 22.15±9.77 ab 23.11±8.96 a 20.59±8.43 bc 18.74±3.88 c Heterophils (%) 21.30±1.93 26.40±2.32 25.30±1.45 27.40±2.42 25.30±1.56 Lymphocytes (%) 73.20±1.64 69.50±2.51 69.80±1.44 67.10±2.42 70.40±1.45 Monocytes (%) 2.20±0.44 1.80±0.36 2.10±0.28 2.70±0.30 2.30±0.39 Eosinophils (%) 2.30±0.42 2.40±0.56 2.60±0.40 2.40±0.45 2.20±0.20 Basophils (%) 0.20±0.13 0.10±0.10 0.00±0.0 0.40±0.22 0.20±0.13 Platelets (x10 3 /µl) 121.80±38.9 b 146.50±88.75 a 135.70±92.78 ab 125.30±32.7 b 125.60±43.04 b Total Protein (g/dl) 4.03±0.36 3.69±0.19 3.62±0.23 4.39±0.19 4.03±0.25 Albumin (g/dl) 1.23±0.07 b 1.26±0.09 b 1.24±0.13 b 1.70±0.06 a 1.54±0.13 ab Albumin/Globulin 0.49±0.06 b 0.53±0.04 ab 0.52±0.05 ab 0.65±0.03 a 0.61±0.03 ab AST (U.I/l) 99.85±6.65 ab 113.80±10.65 a 91.29±7.46 b 99.22±4.33 ab 89.18±4.00 b Cholesterol (mg/dl) 104.57±6.78 a 92.19±3.76 ab 86.86±1.82 b 97.37±4.58 ab 100.11±6.29 ab abc: mean in the same row with different superscripts are significantly (P<0.05) different. PCV: Packed Cell Volume. AST: Aspartate amino transferase. therefore the inclusion levels are good enough that they might have bound with vitamins, minerals and amino acids as well as Aflatoxin. 3.1 Performance Characteristics Table 1 The final live weight was highest (P<0.05) in birds fed 1 (+ve control) and least (P<0.05) in birds fed diet 5 (SB) compared to others. Average daily weight gain of birds fed diet 1 (+ve control) was significantly (P<0.05) higher than those fed diets 2, 3, 4 and 5 which were not significantly different from one another. Weight gain of birds fed diet 3 (Montmorillonite) was apparently higher than those fed HSCAS, SB and the diet 2 (-ve positive). Daily feed intake of birds fed Montmorillonite, HSCAS and SB were not significantly different from the control. The apparently least feed intake was recorded in birds fed diet 2 (-ve control). The feed conversion ratio was least in the control birds without Aflatoxin follow by those fed diet3 (Montmorillonite). The percentage mortality was least in the diet 1 (2%) and highest in birds fed diet 3 (44%). This study do not agree with the result of Talat et al [14] and Mussaddeq et al [15].They all reported an increase in weight gain of chicken fed Aflatoxin contaminated diet and binders. For weekly, daily feed intake, feed conversion ratio and feed conversion efficiency there is no significant difference amongst the five treatments. Massaddeq et al [15] also reported no significant different in feed intake and feed conversion ratio. Talat et al [14] reported no significant difference in both the control diet and other treatments with the toxin binders fed sodium bentonite and HSCAS. These results are in line with findings of Awan [16] who reported a decrease in dressing percentage due to the Aflatoxin in the diet. Also this result do not agree with the report of Ibrahim et al [17] who observed consistency in growth rate of chicks fed 2 5mg AF kg 1 feed with chicks receiving similar levels of dietary AF. These results could be attributed to the factor that the binders have a non-selective binding property 3.2 Relative Organ Weights Table 2, relative organ weights were significantly (P<0.05) influenced by the dietary treatment except that of heart, adrenal gland, pancreas, thymus, and bile. Relative liver weight was significantly (P<0.05) higher in birds fed diets 2, 3 and 5 than the control birds. The liver weight of birds fed diets 4 was not significantly different from that of control. Relative lung weight of the birds fed diets 5 (SB) was not significantly different from those fed diets 2, 3 and 4 but significantly (P<0.05) higher than birds fed positive control (diet 1). Relative kidney weight of birds fed diet 5 was not significantly different those fed diets 3 and 2 but not significantly (P<0.05) higher than those fed diets 4 (HSCAS) and 1 (+ve control). The kidney weight of birds fed diet 4 was not significantly different from those of birds fed diets 3, 2 and 1. Relative spleen weight of birds fed diet 5 was not significantly different from those fed diets 2, 3 and 4. However, it was significantly (P<0.05) higher than those fed diet 1 (+ve control). Relative weight of the GIT in birds fed diets 1 and 4 were statistically the same, likewise those that fed diets 2, 3 and 5. However, the GIT weight of birds fed diets 5 (SB) was significantly (P<0.05) higher than birds fed diet 4 (HSCAS) and diet 1 (+ve control). The birds fed diets 3, 4 and 5 containing toxin binders. Abdominal fat was lower in birds fed diets 3, 4 and 5 than the birds on diets 2 and 1. Abdominal fat decreased significantly (P<0.05) in treatments with Aflatoxin and binders compared to the controls. Relative weight of bursa of birds fed diet 4 was significantly (P<0.05) higher than diet 1. Bursa weight of birds fed diet 259
1 was not significantly different from those fed diets 2, 3 and 5. Relative Gizzard weight was significantly (P<0.05) highest in birds fed diet 5 while the least weight was observed in diet 1. Gizzard weight was statistically the same in birds fed diets 2, 3 and 4. This study is in line with Huff et al [18] who reported that liver is more affected in aflatoxicosis with pin-point hemorrhages and fat deposition, kidney enlargement and bursa regression in avian species. The weight of abdominal fat decreased significantly (P<0.05) in treatments with binders compared to control. This result contradicts the work of Lon-Wo et al [19] who indicated that adding zeolite to broiler diet resulted in decrease in abdominal fat compared to control. Lotfollahian et al [20] reported that using low level of zeolite (2%) resulted in a decrease in abdominal fat. 3.3 Hematological Characteristics Table 3 Hematological parameters such as PCV, hemoglobin, erythrocytes, leucocytes and platelets do not show significant difference in birds fed Aflatoxin contaminated diets, contaminated diets plus binders and diets free from aflatoxins. This may be due to the ability of the binders to ameliorate the negative effects of Aflatoxin contamination in birds. The addition of various binders (0.2kg/100kg) to an Aflatoxin contaminated diet significantly reduced the adverse effect of Aflatoxin on haematology, total protein, albumin value, globulins and AST activity. Aflatoxin contaminated feeds decreased the activities of several enzymes important for digestion of proteins, lipid and nucleic acid in broiler chicken [21]. The serum proteins of birds at 56 days were not significantly different across the treatments. This result is contrary to the report of Arab et al [22] where serum protein decreased in birds fed Aflatoxin contaminated diets. The no significant difference in the serum protein values may be attributed to the ability of the binders to reverse the negative effects of aflatoxins in birds on treatments 3, 4 and 5. 4. CONCLUSION The inclusion of Aflatoxin binders has been reported to be beneficial in the prevention of aflatoxicosis, but the dose and duration of administration play a significant role in this regard. Numerous strategies for the prevention of bioavailability of Aflatoxin in animals have been reported. Some are clearly more practical than others. The modification of toxicity and carcinogenicity in animals through diverse chemicals and adsorbent interventions including the use of HSCAS clay to diminish animal exposure to the aflatoxins, show great promise and have significant economic advantage. 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