Endotoxins in Swine – Effects and Strategies for Control

Endotoxins in swine – effects and strategies for control

by Josep Garcia-Sirera, Special extremely sensitive to detergents, antibiotics Nutrients Inc, 2766 Douglas Road, and bile salts. So it seems that this region is Cocunut Grove, Florida 33133, USA. essential for the maintenance of outer mem - brane functions as a biological barrier. ndotoxins are structural components Mutations altering the lipid A component of bacteria. They are part of the outer are mostly not viable, suggesting that it is Emembrane of Gram negative bacteria, important for the maintenance of outer which are released mainly when bacteria are membrane integrity as a whole. lysed, due to the use of antibiotics or because of the body’s defence mechanism. Together with phospholipids and mem - Lipid A and virulence brane bound proteins they are constituents of the outer cell membrane. The typical The physiological activities of LPS are medi - Fig. 2. Schematic view of the cellular structure of an endotoxin consists of a ated mainly by the lipid A component of endotoxin (LPS) signal transduction lypopolysaccharide (LPS). These LPS define LPS. Lipid A is a powerful biological pathway. LPS binding protein (LBP) many of the properties of host-parasite response modifier that can stimulate the mammalian immune system. During infec - solved in the plasma ligates at LPS (1). interactions. This complex binds to the cell surface tious disease caused by Gram negative bac - LPS consists of three structural elements. receptor CD14 (2). Aggregation of teria, endotoxins released from, or part of, One is a hydrophobic component, called LBP/LPS/CD14 complex with the protein multiplying cells have similar effects on ani - lipid A, which serves to anchor the molecule MD2 and the transmembrane toll like mals and significantly contribute to the into the membrane. The second is a core receptor (TLR) 4 (3) induces the signal symptoms and pathology of the disease oligosaccharide. The third component is a transduction cascade in the cell (4, 5). hydrophilic O-polysaccharide projecting into encountered. Finally, transcription factor NF κB is acti - the extracellular space. More than 150 dif - Since lipid A is embedded in the outer vated (6) and starts translation of sev - ferent variants of the third component are membrane of bacterial cells, it probably only eral genes to proteins, for example known. The O-polysaccharide portion exerts its toxic effects when the bacteria are pro-inflammatory mediators like IL-1 β seems to be relevant to host-parasite inter - lysed as a result of autolysis and the mem - (7,8). actions because its disappearance results in brane attack complex (MAC), ingestion and loss of virulence. killing by phagocytes, or killing with certain The loss of the proximal part of the core types of antibiotics. tein, MD2, which associates with toll-like oligosaccharide induces bacteria to become The injection of living or killed Gram nega - receptor-4 (TLR4). tive cells or purified LPS into experimental This triggers the signaling cascade for Fig. 1. Lypopolysaccharide structure. animals causes a wide spectrum of non-spe - macrophage/endothelial cells to secrete cific pathophysiological reactions, such as pro-inflammatory cytokines and nitric oxide fever, changes in white blood cell counts, that lead to the characteristic ‘endotoxic disseminated intravascular coagulation, shock’. hypotension, shock and death. Injection of CD14 and TLR4 are present on several fairly small doses of endotoxin results in cells of the immunological system, including death in most mammals. macrophages and dendritic cells. The sequence of events follows a regular In monocytes and macrophages, three pattern: types of events are triggered during their G Latent period. interaction with LPS: G Physiological distress (diarrhoea, prostra - G Production of cytokines, including IL-1, tion, shock). IL-6, IL-8, tumour necrosis factor (TNF) and G Death. platelet activating factor. These, in turn, How soon death occurs varies on the stimulate production of prostaglandins and dose of the endotoxin, route of administra - leukotrienes. These are powerful mediators tion, and species of animal. Animals vary in of inflammation and septic shock that their susceptibility to endotoxin and the accompanies endotoxin toxaemia. LPS acti - mechanism is complex. vates macrophages to enhanced phagocyto - In humans, LPS binds to a lipid binding pro - sis and cytotoxicity. Macrophages are tein (LBP) in the serum which transfers it to stimulated to produce and release lysosomal CD14 on the cell membrane, which in turn enzymes, IL-1 (‘endogenous pyrogen’), and transfers it to another non-anchored pro - Continued on page 14

International Pig Topics — Volume 25 Number 3 13 Continued from page 13 Treatment Final ADG Feed CI tumour necrosis factor (TNFalpha), as well weight (g) (g) intake (g/d) as other cytokines and mediators. Activation of the complement cascade. G Control 749.68±14.16 a 33.61±0.66 a 44.96±0.88 a 1.388±0.010 b C3a and C5a cause histamine release (lead - b b ab a ing to vasodilation) and affect neutrophil LPS 684.63±16.81 30.51±0.79 42.58±0.97 1.396±0.018 chemotaxis and accumulation. The result is YCL 744.40±19.12 a 33.35±0.91 a 44.28±1.04 a 1.328±0.012 b inflammation. LPS + YCL 692.93±8.25 b 30.91±0.38 b 41.56±0.58 b 1.344±0.014 b G Activation of the coagulation cascade. Initial activation of Hageman factor (blood- Means in the same column with a different letter are statistically different (P<0.05) clotting Factor XII) can activate several Table 1. Effect of inclusion of yeast cell wall in the diet on production parameters humoral systems resulting in: (1-21 days) of broilers inoculated with LPS of E. coli (Badia R. et al). – coagulation: a blood clotting cascade that leads to coagulation, thrombosis, acute dis - – activation of the complement alternative – kinin activation releases bradykinins and seminated intravascular coagulation, which pathway (which leads to inflammation). other vasoactive peptides which causes depletes platelets and various clotting fac - – plasmin activation which leads to fibrinoly - hypotension. tors resulting in internal bleeding. sis and haemorrhaging. The net effect is to induce inflammation, intravascular coagulation, haemorrhage and shock. LPS also acts as a B cell mitogen, stimulating the polyclonal differentiation and multiplication of B-cells and the secretion of immunoglobulins, especially IgG and IgM.

Control of endotoxins

In general, strategies to control endotoxin contamination in animals include all of those aimed at the reduction of bacterial contami - nation. These strategies include, but are not limited to, biosecurity, use of prebiotics, probiotics and improved nutrient digestibil - ity. Other strategies such as vaccination and use of toxin binders specifically target endo - toxin contamination. G Vaccination. As explained before, the lipid A portion is responsible for endotoxin toxicity. Also it is the more consistent por - tion of LPS structure. Currently, immunisa - tion against lipid A is being developed, but the high cost makes it a non-viable option for livestock production. Another option considered in vaccination is to immunise against LBP, in an attempt to reduce the formation of LBS-LBP complex that initiates the cascade of events leading to pathogenesis. This option is also expensive and currently only to be considered for human use. G Immunity modulators. Use of immune modulators to compensate the effects of endotoxins have been tested in animal pro - duction. In broilers, inoculation with LPS induces an activation of the immune system. Some studies show that broilers inocu - lated with LPS decrease productivity. This reduction is related to the action of inter - leukins produced during the acute inflamma - tory phase. On the other hand, some studies show the immune-modulating action of B-glucans present in yeast cell wall. Table 1 shows the results of using yeast cell wall (YCL) in diet of broilers inoculated with LPS. YCL was capable of counteracting the effect of LPS in conversion index (CI). Further tests would be needed to study the possibility of using immune modulators against endotoxins. A more practical approach to reduce absorption of endotoxins from the gastroin - testinal tract of livestock is the use of toxin

14 International Pig Topics — Volume 25 Number 3 binders. Toxin binders are widely used to against endotoxins in swine. Fig. 3 shows the control other toxins such mycotoxins. 35 Control AZ 0.05% AZ 0.10% results of using a commercially available The binder and the mycotoxin form a toxin binder to reduce free endotoxin con - complex that is too large to be absorbed 30 centration in endotoxin producing bacterial into the blood system. The complex is then cell culture. eliminated in the faeces. 25 The product was capable of binding endo - Most mycotoxin binders are hydrophilic toxins at two different inclusion rates. This 20 molecules (bentonites, aluminosilicates) effi - binding capacity was confirmed in an in vivo cient at capturing polar molecules such as test. 15 aflatoxin. The capacity of these traditional Table 2 shows free endotoxin concentra - mycotoxin binders to capture more 10 tion in blood of sows before and after feed - lipophilic-like molecules such zearalanenone ing them the commercial toxin binder or DON is questionable. There are also 5 This preliminary data suggest that the toxin organic mycotoxin binders (MOS based) binder is capable of binding endotoxin and that claim to be able to adsorb a wider 0 the gastrointestinal tract, and thus prevent - range of mycotoxins. Farm 1 Farm 2 ing endotoxins from being absorbed. Further studies are currently underway to Fig. 3. Endotoxin concentration in bac - further test this product. Table 2. Endotoxin concentration in terial cell culture of sow faeces (1-2pp) blood of sows before and after (three after in vitro supplementation of differ - weeks) addition (1kg/MT of feed) of ent MycoAD AZ concentrations (Univ- Summary MycoAD AZ (Biocheck, Leipzig, ersity of Leipzig). Germany). Endotoxins have a wide variety of effects on Sow Endotoxin ml/L When considering the possibility of using livestock affecting performance parameters. number Before After toxin binders to capture endotoxins, the Different means of control of endotoxins binder binder ideal binder candidate should target lipid A. have been tested, but most of them are too The reason being that lipid A is responsible expensive to be considered in animal pro - 123 4.55 <0.05 for the pathogenic effects of endotoxins, duction. 125 30.86 <0.05 and it is the portion of the structure that A cost effective method would be the use 126 <0.05 <0.05 remains constant across different endotox - of toxin binders to capture endotoxins in 127 >50 <0.05 ins, so by targeting lipid A, the toxin binder the gastrointestinal tract and preventing 128 22.88 <0.05 would have a wider range of action. them from entering the blood systems. 129 2.85 <0.05 Some preliminary studies have already Some studies are underway to test this 130 47.33 <0.05 tested the possibility of using toxin binders possibility. I

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