Dietary Meat and Bone Meal, and Calcium as Precursors for the Onset of Subclinical Necrotic Enteritis in Broilers Fed Diets Containing Exogenous Phytase Enzyme

Abstract

<p>One of the predisposing factors for the onset of necrotic enteritis (NE) in poultry is indigestible nutrients or nutrient imbalance. Poorly digested diets lead to the accumulation of undigested nutrients in the lower gut feeding pathogenic bacteria or create favorable conditions (high pH) that favour their proliferation. Meat and bone meal (MBM) and high dietary calcium (Ca) are examples of such dietary factors that might favor the overgrowth of enteric pathogenic bacteria.</p> <p>Three experiments were conducted in this project employing a 2 × 2 × 2 factorial arrangement of treatments. The factors were: Exp. 1, MBM (no or yes), AB (no or yes, zinc bacitracin + salinomycin) and phytase level (500 or 1500 FTU/kg; both using 500 matrix recommendations); Exp 2, NE challenge (no or yes), Ca level (0.6 or 1.0% starter, 0.5 or 0.9% grower, 0.4 or 0.8% finisher with the same level of P in each phase of the study), phytase (500 or 1500 FTU/kg both using 500 matrix recommendations) and Exp. 3, NE challenge (no or yes), (500 or 5000 FTU/kg both using 500 matrix recommendations) and MBM (as-received, AR or over-processed, OP). The diets used in all the experiments were based on wheat, SBM and canola meal. All the birds in Exp. 1 were challenged and only half of the birds in exp 2 and 3 were challenged. The challenge birds were inoculated with <i>Eimeria</i> spp. on d 9, and <i>Clostridium perfringens</i> (<i>C. perfringens</i>) strain EHE-NE18 on d 14 and 15. The effects of the factors/treatments were measured as changes in performance, nutrient digestibility, phytic acid hydrolysis, bone mineralization, haematology, intestinal morphology, jejunal gene expression and welfare (hock burns and litter quality). </p> <p>Chapter 2 presents a review of the possibility of dietary MBM and excessive dietary Ca to exacerbate the outcome of NE in broiler chickens.</p> <p>Chapter 3 examines the influence of MBM, phytase and antibiotics on broiler chickens challenged with subclinical NE as measured by growth performance, intestinal pH, apparent ileal digestibility, caecal microbiota and tibial mineralization. The results revealed that on d 21 (post-challenge), birds fed MBM had reduced weight gain (WG; P < 0.05) relative to without MBM. The WG on d 14 (P < 0.001) and d 21 (P < 0.001), and FCR on d 21 (P < 0.001) and d 42 (P < 0.01) indicated positive effects of high phytase on bird performance in the presence of AB. On d 42, high phytase increased WG with AB (P < 0.01), relative to the birds without AB in the presence of MBM. There was a notable reduction in Ca and P digestibility in birds fed MBM-free diets and a low phytase level on d 16 (P < 0.01), but with the high phytase level, Ca and P digestibility was not influenced by MBM. Thus, it was concluded that in NE challenged birds, high phytase has a beneficial effect on leg health and mineral utilization, to the extent that it can replace MBM and has beneficial effects on bird performance in the presence of AB.</p> <p>Chapter 4 examines the influence of MBM, phytase and antibiotics on broiler chickens challenged with subclinical NE as measured by intestinal permeability, organ weights, haematology, intestinal morphology and jejunal gene expression. The results indicated that lymphocyte counts were lower (P < 0.01) with MBM and AB compared to MBM without AB. Villi length was increased (P < 0.05) with high phytase and no AB compared to AB. Inclusion of MBM increased blood <i>fluorescein</i> isothiocyanate dextran (FICT-d; P < 0.05) concentration whereas AB decreased it (P < 0.001). Antibiotics increased red blood cells (RBC; P < 0.05), haemoglobin, (Hgb; P < 0.05) and packed cell volume (PCV; P < 0.05) and expression of Ca-binding protein (CALB1; P < 0.05). Overall, it was demonstrated that dietary MBM has a detrimental effect on the gut health of broilers, but the effect may be reduced by using AB.</p> <p>Chapter 5 evaluated the effect of OP MBM and phytase effects on broilers challenged with subclinical NE as measured by performance, intestinal lesions and pH, bacterial counts and apparent ileal digestibility. The results indicated that challenged birds fed OP MBM had decreased BW (P < 0.05) and FI (P < 0.05) at d 14, increased FCR (P < 0.01) at d 21 and decreased BW (P < 0.05) and FI (P < 0.05) at d 28. Birds fed low phytase had increased livability (P < 0.05) at d 42. The challenge increased the prevalence and severity of NE induced lesions in the jejunum (P < 0.001) and ileum (P < 0.01). The birds fed OP MBM had decreased pH in the jejunum (P < 0.05) and ileum (P < 0.05) at d 16. High phytase increased apparent ileal digestibility of Ca (P < 0.05) and P (P < 0.001). The challenge increased the counts of <i>Lactobacillus</i> spp. (P < 0.01) and <i>C. perfringens</i> (P < 0.001) at d 16. It was concluded that the supplementation of diets with high phytase reduces the negative impact on performance from OP MBM during NE as a result of increased nutrient digestibility.</p> <p>Chapter 6 assessed the effect of OP MBM and phytase effects on broilers challenged with subclinical NE as measured by changes in inositol phosphate esters hydrolysis, intestinal permeability, haematology, jejunal gene expression and intestinal morphology. The result revealed that birds fed low phytase had increased inositol penta-phosphate (IP5; P < 0.05) and inositol hexaphosphate (IP6; P < 0.05) in unchallenged birds only when diets contained OP MBM. Challenge with NE increased intestinal permeability as measured by serum FITC-d (P < 0.001), increased white blood cells (WBC, P < 0.001), decreased mean corpuscular volume (MCV; P < 0.001) and mean corpuscular hemoglobin (MCH; P < 0.05), and decreased crypt depth to villi length ratio (P < 0.05). The OP MBM reduced the villi: crypt ratio (P < 0.05). Only in unchallenged birds fed OP MBM did high phytase reduce MUC2 expression (P < 0.05). It was concluded that NE had a negative impact on the gut and haematology of broilers, but its effect on phytate hydrolysis was minimal. </p> <p>Chapter 7 evaluated the effect of OP MBM and phytase effects on broilers challenged with subclinical NE as measured by changes in litter quality and bone mineralization. The study showed that at d 16, challenged birds had lower toe ash (P < 0.01), femur ash (P < 0.001), tibia ash (P < 0.001) and tibial breaking strength (BS) (P < 0.001) than unchallenged birds. At d 16, only challenged birds fed high phytase and OP MBM had higher toe Mn than those fed low phytase and AR MBM. At d 16, tibial Ca (P < 0.05) and P (P < 0.05) were lower in the challenged whereas the femur K (P < 0.001), Mn (P < 0.01) and Na (P < 0.001) were higher in the challenged at d 16. At d 42, challenged birds had higher litter DM (P = 0.058) and fewer hock burns than those unchallenged (P < 0.05). It was concluded that NE impaired bone traits while high phytase and OP MBM increased bone mineral contents.</p> <p>Chapter 8 investigated the interactive effect of two dietary calcium and phytase levels on broilers challenged with subclinical NE as measured by changes in broiler performance, gut lesions and pH, bacterial counts, and apparent ileal digestibility. The results showed that gain was higher in birds fed high phytase on d 14 (P < 0.01), d 21 (P < 0.01), d 28 (P < 0.01) and d 35 (P < 0.01). Birds fed high phytase had greater livability on d 0-21 (P < 0.01). Calcium was more digestible in high Ca diets on d 16 and a NE × Ca interaction showed this effect to be more pronounced in unchallenged compared to challenged birds. A challenge × Ca interaction was observed for apparent ileal digestibility (AID) of CP (P < 0.05) indicating a lower AID of CP in challenged birds fed high Ca. The challenge decreased AID of Ca (P < 0.01). Calcium had no impact on <i>C. perfringens</i> count, but it decreased <i>Lactobacillus</i> (P < 0.05) and Bifidobacteria (P < 0.05) populations in the caeca. Thus, high Ca in the presence of phytase was beneficial for growth except that Ca created a condition in the gut where the incidence could have been worse if the challenge was clinical.</p> <p>Chapter 9 investigated the interactive effect of two dietary calcium and phytase levels on broilers challenged with subclinical NE as measured by gut permeability, phytate esters degradation, jejunal gene expression, and intestinal morphology. The results showed that the inositol triphosphate (IP3; P < 0.01) and inositol tetra-phosphate (IP4; P < 0.05) concentrations were similar for both doses of phytase in the presence of low Ca, but with high Ca both increased significantly but to a greater extent when the high dose of phytase was used. A phytase × Ca interaction was detected for vitamin D receptor (VDR; P < 0.05) expression where bird fed low phytase and low Ca recorded the highest expression of VDR, all other treatments being equivalent. Challenge birds had higher FITC-d (P < 0.05) in blood compared to unchallenged birds. Thus, high Ca and high phytase, whilst not the best for IP6 destruction, did not lead to huge reductions in indicators of gut health.</p> <p>Chapter 10 investigated the interactive effect of two dietary calcium and phytase levels on broilers challenged with subclinical NE as measured by changes in serum calcium and phosphorus, and bone mineralization. The challenge decreased serum (P < 0.05) Ca⁺ in birds regardless of dietary Ca level (d 16). Tibial BS (d 16) in challenge birds was far greater (P < 0.05) in the high Ca diet compared with low. Femur ash (d 16) in challenged birds was increased due to the phytase (P < 0.05). The challenge decreased (P < 0.05) the BS of femur and tibia at d 16 and 29. Birds on high dietary Ca had lower tibial Mg (P < 0.001), Na (P < 0.001) and Zn (P < 0.05) concentrations (d 29). Altogether, high dietary Ca and phytase improved bone mineralization.</p> <p>Chapter 11 examined volatile basic nitrogen measurement in the caecal digesta of broiler from the 3 experiments using a Berthelot reaction in automated Skalar® instrumentation. The results demonstrated that an increase (P < 0.05) in caecal NH₃ as a result of feeding MBM (Exp. 1) or high dietary Ca (Exp. 2) with a corresponding increase in pH. In Exp. 3, birds challenged with NE had lower caecal NH₃ and lower pH on d 16 but not d 29 compared to unchallenged controls. It was concluded that the Berthelot method using Skalar instrumentation equipment is suitable to measure the concentration of volatile nitrogen as NH₃ in caecal contents and other digesta of chickens. </p> <p>Overall, it was concluded that MBM is a potential predisposing factor for the onset of NE while the benefit from feeding high phytase in a MBM-based diet is only realized only in the presence of antibiotics. Also, overprocessing of MBM by way of heating beyond the existing industry practice increases the incidence of NE and impairs growth performance. Finally, high phytase is only beneficial to the growth performance of broilers only in the presence of low dietary Ca and high Ca promotes a gut environment that could increase the outcome of NE.</p>