14. Physical and chemical methods

non-exposed birds. Moreover, birds 3rd weeks of age that received 1800 μg/kg AFs were found

Aflatoxins intoxications suppress immunoglobulins (IgM, IgG and IgA) and enhance susceptibility of birds to parasitic, viral and bacterial infections. At 0.5 to 1 mg/kg Aflatoxins, these interfere with B and T-lymphocytes functioning [73], apparent alteration of splenic functioning, atrophy of bursa of Fabricius [74], suppresses cell mediated immune response, phagocytosis, and complement system as well as interferon production. Moreover, hematopoietic suppression and anemia have been observed by decrease in RBCs, packed cell volume and

AFs decrease total serum proteins due to a reduction in α, β and γ globulins, with IgG being more sensitive than IgM [79] which may cause substantial suppression of acquired immunity from vaccination programs in some disease models. The Low levels of AFB1 appears to affect the vaccinal immunity negatively and may enhance the occurrence of diseases such as Marek's disease, IBD virus, congenitally acquired salmonellosis and duodenal and cecal coccidiosis, etc. even in properly vaccinated flocks [80]. The failure of vaccines is correlated to the immunotoxic effect of toxins which compromise for immune function of birds by decreasing cellmediated immunity and inducing an inflammatory response [81]. Decrease chemotactic ability of leucocytes, impaired heterophils phagocytosis [3] and cellular and serum factors required for optimal phagocytosis can be observed in aflatoxicated chickens. Although dietary AFs depress thrombocyte counts, no effect on their phagocytic activity has been observed [82].

Due to synergistic effect of Aflatoxin B1 and hepatitis B exposure, there are no specific safe levels for aflatoxin regarding resistance/tolerance to AFs. Ideally, there should be zero level for AFs in feed [83]. The Food and Drug Administration and European Union have established 20 μg/kg and 10 μg/kg AFs as maximum level for poultry, respectively. Based on feeds available, AF contaminated feeds should be fed at lowest possible level and for the shortest period of time [84]. The production of AFs can be controlled by maintaining physical integrity of cereal grains, drying and use of anti-fungal especially propionic acid to inhibits molds growth by decreasing pH and ATP formation through electron transport pathway. UV, X-rays or microwave irradiation and dilution of contaminated feed with AF free feed is also one of the methods to dilute the concentration of AFs [9]. However, AFB1 contamination of feed is practically unavoidable universally [85]. Mycotoxins decontamination refers to methods by which these metabolites are removed or neutralized in contaminated feed, while mycotoxins detoxification refers to methods by which the toxic properties of the mycotoxins are eliminated [86]. Since early 1990s, studies on mycotoxin adsorbents have yielded success but high

to have detectable levels of AFB1 in the liver.

13. Safe level of aflatoxins and detoxification

12. Immunosuppression

134 Mycotoxins - Impact and Management Strategies

hemoglobin [75–78].

Thermal inactivation, cleaning of the kernel surface, and hence the removal of highly contaminated particulate matter, have proven effective in reducing moderate mycotoxins contamination of feed [43, 89]. However, it seems quite laborious to remove highly contaminated feedstuffs. On the other hand, a lot of chemicals e.g. acids (sulfuric acid, hydrochloric acid, phosphoric acid, benzoic acid, citric acid, acetic acid), alkaline compounds (ammonia, sodium bicarbonate, sodium hydroxide, potassium hydroxide, calcium hydroxide, caustic soda), salts (acetate ammonium, sodium bisulfite, sodium hydrosulfite, sodium chloride, sodium sulfate), oxidants (H2O2, sodium hypochlorite, ozone), reducing agents (bisulfites), chlorinated agents and formaldehyde, etc., are being used for the degradation of mycotoxins in feed [90]. These methods are inefficient but comparatively expensive. Ammoniation has been demonstrated to reduce AFs levels but not accepted in the United States [91].

High level dosages of methyl bromide, ethylene dibromide, propane/propene ethylene oxide, sulfur dioxide, phosphine propionic, acetic and isobutyric acids show fungicidal activity. However, these chemicals lower nutritional quality and are corrosive on human and animal tissues [92]. Therefore, the use of these chemicals is discouraged. Several related patents involving the use of ozone in agricultural products decontamination are found. This decontamination method involves placing the agricultural products in a treatment chamber, generating ozone in the vicinity of chamber, supplying ozone to the product through continuous flow and exposing the agricultural product to ozone, which then reacts with the toxins and/or microorganisms.

There are different types of adsorbents, which can be used for the detoxification of AFs in the feed. The use of activated carbon for the detoxification of mycotoxins can also be another option but different activated charcoals have less/no effect against mycotoxins, which show their unspecified adsorbent nature. Moreover, certain essential nutrients are also adsorbed when at higher concentration in as compared to mycotoxins [93].

The most applied method for protecting animals against mycotoxicoses is the utilization of adsorbents in the feed, aimed at binding mycotoxins efficiently in the gastrointestinal tract, thus limiting or at best preventing the toxins from being absorbed by the body thereby, preventing their toxic effects and "carry over" of the toxins to animal products [89]. Selected adsorbents added to AFs-contaminated feeds as feed additives can sequester AFs during the digestive process, allowing the mycotoxins to pass harmlessly through the gastrointestinal tract of animal [94]. This is one of the more effective and practical approaches to address the problem of AFs. The degree of adsorption capacities may vary (0–87%) among various mineral clay materials [95], and very few are actually used commercially. These considered as good absorbents include bentonites, zeolites and aluminosilicates. Studies have shown that sodium aluminosilicates, HSCAS (hydrated sodium calcium aluminosilicates) and sodium bentonites adsorb AFs [96] with adsorption potential of bentonites varying from 17 to 36%. A major advantage of these adsorbents is that they are relatively inexpensive and safe and can be easily incorporated in animal feeds [97].

showed significant (p < 0.05) binding activity against mycotoxins as compared to control with non-treated group. However Hydroxy propyl methyl cellulose, Sodium Carboxy methyl cellulose, and Microcrystalline Cellulose showed better adsorbent capacity for all mycotoxins when

Aflatoxins: Their Toxic Effect on Poultry and Recent Advances in Their Treatment

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Various bacterial, yeast and fungal species are able to degrade/remove mycotoxins and also can restrict fungal growth. This includes the use of Bacillus subtilis, NK-330 and NK-C-3 that effectively inhibit the fungus growth and AFs production [92]. The application of microorganisms e.g. Corynebacterium rubrum for bio-transformation of mycotoxins into less toxic metabolites is another option [9]. These micro-organisms act in intestinal tract of animals prior to absorption of mycotoxins but the concerned toxicity of products by enzymatic degradation and undesired effects of fermentation with non-native micro-organisms on food quality is yet

Saccharomyces cerevisiae and lactic acid bacteria (LAB) i.e. propionibacteria, bifidobacteria and lactobacillus rhamnosus strongly bind to their cell wall constituents mycotoxins without deleterious effects on animal health [9, 85, 93, 112]. Most yeast strains bind more than 15% (w/w) AFB1, which is highly strain specific by S. cerevisiae [112] and LAB for mycotoxins detoxification [113]. Generally, S. cerevisiae shows very low adhesion to the intestines [114], as opposed to LAB that show considerable adhesion to intestinal cells [115]. Coallier-Ascah and Idziak [116] and Thyagaraja and Hosono [117] found LAB to be inefficient binders of AFB1 due to the strains used, which may also depend on initial concentration of AFs [118]. Haskard et al. [119] showed that cell wall of L. rhamnosus has the ability to bind AFs predominantly to carbohydrates and to some extent, protein components that which is unaffected by pH of GI tract. The outer part of cell wall (26–32%) of S. cerevisiae contains a structure called glucomannan, which binds against mycotoxins [9]. The yeast cell wall comprises of 30–60% polysaccharides (β-glucan and mannan sugar polymers), 15–30% protein, 5–20% lipids and a small amount of chitin. Mainly, it contains 15–30% β-glucan and 15–30% MOS. Lahtinen et al. [120] found that peptidoglycans might be the most likely carbohydrate involved in the AFB1 binding process [121]. Kusumaningtyas et al. [122] used S. cerevisiae, Rhizopus oligosporus and their combination for detoxifying AFB1 in the chicken feed. The supplementation of whole yeast and only yeast cell wall rather [53, 112, 123] have shown a reduction in mycotoxins toxicities, indicating possible stability of the yeast-mycotoxins complex along the gastrointestinal tract. The cell wall represents about 30% of total weight of yeast cell [112]. Glucomannan is a bi-layered structure that consists of a network of β-1,3 glucan with β-1,6 glucan side chains. This network is in turn attached to highly glycosylated mannoproteins. The proteins and glucans provide numerous easily accessible binding sites with different binding mechanisms such as Van Der Waals bonds, hydrogen bonding, ionic or hydrophobic interactions [93, 112, 124, 125]. Yeast glucomannan showed markedly high binding ability with AFs in vitro (75–90%) and in vivo [126, 127]. The carbohydrate fractions of cell wall may represent 90% of mannoproteins. MOS constitute approximately 50% of total carbohydrates [112]. The effect of 500 g of glucomannan is comparable with that of 8 Kg of clay for mycotoxins bindings [9].

compared with Cholistan.

15. Biological methods

to be investigated completely.

Mineral adsorbents based on zeolites, silicates and phyllosilicates show different abilities to bind AFs. These possess active sites within interlayer channels at the basal planes on the surfaces or within pores, and at the edges of particles [98]. Bentonites are white, light weight and originate from volcanic ash comprising mainly of montmorillonite, the main constituent of bentonites. These are composed mostly of salts of Na, K, Ca of hydrated aluminosilicates and occasionally Fe, Mg, Zn, Ni, etc. but the composition varies from one deposit to another because of interchangeable mono and divalent ions e.g. Na+ , K+ , Ca+2, and Mg+2. So they can be classified as Ca, Mg, K or Na bentonites [86]. They have a layered microstructure, which allows AFs to bind at multiple sites including edges and basal surfaces especially at the interlayer region for adsorption [99, 100].

Zeolites possess strong colloidal properties to absorb water rapidly resulting in swelling and manifold increase in volume, giving rise to a thixotropic gelatinous substance [101, 102]. Hydration of the exchangeable cations creates a hydrophilic environment in the interlayer of montmorillonite, which influence the adsorption of different organic molecules, including mycotoxins on zeolite and montmorillonite particles [103]. The surfaces of zeolites derived HSCAS, attract polar functional groups of AFs, thus inhibit their absorption [93, 104] but is less effective against other mycotoxins. Zeolites selectively retain or release calcium during its passage through digestive system. Zeolites can absorb nitrogen of some amino acids and reduce the energy required for meat production. Zeolites suppress phosphorus utilization by forming indigestible compound with phosphorus through its aluminosilicate component [105]. Supplementation of HSCAS at the rate of 1.0% seems to diminish significantly, the adverse effects of AFs in young animals [93] as these have a high negative charge and are balanced by cations of such metals as magnesium, potassium and sodium located in the cavities, and therefore do not react with food/feed ingredients and act as inert material due to their neutral pH or slightly alkaline nature [106].

Aluminosilicates are also used at a level up to 2% as "anti-caking" agents but a several disadvantages have been observed including the impairment of minerals utilization and having a narrow range of binding efficacy [93]. Bentonites minerals can influence Ca-metabolism and bind nitrogenous cations such as NH4 + . These are found to be effective for the adsorption of AFB1 and T-2 toxin but not for zearalenone. Kececi et al. [107] determined decrease in calcium and phosphorus levels by AFs (2.5 mg/kg) for 21 days. Southern et al. [108] did not find any adverse effect on the growth and tibial mineral concentrations in chicks fed nutrient-deficient diets. Mineral clays reduce utilization of minerals including manganese, zinc, magnesium [109], chloride [95], copper and sodium [110]. Solís-Cruz et al. [111] conducted an in vitro study to evaluate the adsorption capacity of Chitosan (CHI), and three cellulosic polymers (Hydroxy propyl methyl cellulose, Sodium Carboxy methyl cellulose, and Microcrystalline Cellulose), on six mycotoxins (AFB1; FUB1; OTA; T-2; DON; and, ZEA) for poultry. All four cellulosic polymers showed significant (p < 0.05) binding activity against mycotoxins as compared to control with non-treated group. However Hydroxy propyl methyl cellulose, Sodium Carboxy methyl cellulose, and Microcrystalline Cellulose showed better adsorbent capacity for all mycotoxins when compared with Cholistan.
