I. INTRODUCTION
The modern broiler chicken has been genetically selected for rapid growth, increased muscle mass and heavier breast weight (Garner et al., 2002). This increase in production may also be associated with poor leg health and lameness and is often linked to skeletal abnormalities due to reduced bone mineralisation. Poor leg health and lameness affects millions of broiler chickens worldwide, with lame birds having significantly reduced performance and altered behaviour patterns (Weeks et al., 2002; Berg and Santora, 2003). Poor leg health in the broiler industry is often attributed to reduced bone quality. Studies have shown that the dimensions of the tibiotarus of the modern broiler are the appropriate size for weight bearing, however, the bone itself is weak (Sherlock et al., 2010). The cortical bones of broilers have lower mineralisation which is attributed to rapid deposition in the outer layers to increase cortical width to support the increased weight of the bird (Sherlock et al., 2010).
Calcium (Ca) and phosphorus (P) are two of the most abundant minerals in bone and form the majority of the inorganic matrix. The structure and composition of bone vary according to the age and nutritional status of the bird, with the extent of bone mineralisation influencing bone strength (Waldenstedt, 2006). Calcium, provided in the form of calcium carbonate (CaCO3) grit or flour is a major source of Ca in broiler diets. High concentrations of CaCO3 may increase the pH in the proximal gastrointestinal tract due to its high acid binding capacity leading to a decrease in P and amino acid digestibility (Selle et al., 2009). Further, due to its potent net negative charge at intestinal pH, phytate from vegetable ingredients chelates Ca in the small intestine, reducing the bioavailability of both minerals. Diets with lower CaCO3 concentration in the presence of phytase are therefore desirable to enhance P digestibility, feed conversion efficiency and weight gain. However, low levels of dietary Ca may lead to poor skeletal integrity. Displacement of limestone with a more digestible Ca source may be beneficial for reducing total dietary Ca concentration, whilst maintaining intestinal absorption of Ca. One such source is novel calcified seaweed that is mined sustainably from the ocean floor near Iceland and Ireland. Due to a unique porous architecture, this CaCO3 source is highly soluble and may meet the digestible Ca requirement at a lower total dietary Ca concentration. In addition to benefits in Ca and P retention, this creates 'space' in least cost formulation, increasing diet energy density and reducing the reliance on added fat sources. Therefore, the aim of the research presented here was to assess the efficacy of a marine Ca source on skeletal integrity and bird behaviour.
II. MATERIALS AND METHODS
A total of 1820 male Cobb-500 broilers were obtained as day olds from a commercial hatchery. Chicks were randomly allocated to 91 deep litter floor pens (1.5 m x 1.5 m) of 20 broilers. Birds received experimental diets from d 1 to d 40 (d 1 to 14 starter and d 15 to 40 grower), with temperature and lighting controlled according to Cobb broiler breeder instructions. The thirteen experimental treatments (6 replicate pens/treatment) were arranged as a 2 x 2 x 3 + 1 factorial, including calcium source (limestone or marine), two dietary calcium concentrations (0.77 or 0.60% in the starter and 0.57% or 0.40% in the grower), phytase (0, or 500 FTU/kg from either QPT2 or Quantum) and an industry standard reference diet containing 1% total Ca and 0.50% available P (avP) in the starter and 0.85% total Ca and 0.42% available P in the grower. All 12 factorially-arranged diets contained 0.35% av.P in the starter and 0.25% avP in the grower. Both QPT2 and Quantum are enhanced phytases from Escherichia coli (AB Vista Feed Ingredients, Marlborough, UK). All diets were based on corn and soybean meal, were isocaloric and isoenergetic and were formulated to meet breeder targets for all nutrients other than Ca and avP as dictated by the experimental design. All diets were steam pelleted at 80°C and provided, with water on an ad libitum basis.
On d 24 and 39 four focal birds, randomly selected from each pen, were marked for identification for use in the LTL test that was conducted on d 27 and 42. The LTL procedure was performed as described by Berg and Santora (2003). Individual focal birds were removed from the home pen and taken into the area outside the rearing compartment. Birds were individually placed into plastic tubs filled with 3 cm of tepid water (30 to 33 °C). All birds were placed into the tub in a standing position and their time spend standing was recorded. When the bird made an attempt to sit down timing was stopped. If after five minutes no attempt had been made to sit down the test was terminated. Once the test was over, birds were taken out of the tubs, their body weight was recorded and the bird was returned to their home pen.
Statistical analyses were carried out using GenStat (GenStat 14th ed). The GenStat survival analysis model was used for modelling time against the proportion of birds which remained standing for each diet. Treatment differences were considered significant at P < 0.05. The hazard risk ratio used reflects the analysis of time survived to the bird sitting when indexed against the control "industry standard" reference diet.
III. RESULTS AND DISCUSSION
As the purpose of the present paper is to explore leg integrity and bird behaviour the full performance effects will not be presented in detail. However, in order to set the context, birds fed the "industry standard" diet had mean terminal body weights (d 40) of 3.17kg and an FCR of 1.98. There was an overall mortality rate of 2.91% that was unrelated (P > 0.05) to dietary treatment. Inclusion of phytase from both sources significantly increased terminal body weight and reduced FCR (P < 0.05). Higher Ca concentrations enhanced (P<0.05) terminal body weight and reduced (P<0.05) FCR. However, there was a significant Ca level*phytase interaction for FCR where phytase enhanced FCR only in the low Ca diet. The marine Ca source tended (P = 0.08) to improve FCR compared to limestone.
Latency-to-lie results were significantly correlated (P < 0.001) with terminal body weight (Table 1). Overall, for every 100g increase in body weight the chance of the bird sitting in the water increased by 12%. As bodyweight was influenced by dietary treatment (data not shown) this factor was included in the model to remove the effect of bodyweight per se and to allow conclusions regarding the absolute diet effects. Diet C and Diet D were the only diets that were statistically different from the "industry standard" reference diet (Diet A; Table 1). Birds that were fed these diets were found to be 59% and 54% respectively less likely to sit in the water at any given point during the test, as indicated by the Hazard risk ratio. Though not statistically significant, only diet E tended to increase the likelihood of birds to sit compared with Diet A (+28%; P=0.285). Diet E was the novel marine Ca source included at a high concentration and this suggests that high dietary concentrations of very soluble Ca sources may be unwise (especially given the significant benefit at a lower level) due to precipitation with P possible exacerbation of a P deficiency.
Table 1. Dietary comparison and composition
Behavioural responses of broilers in the LTL test are highly correlated with gait scores, which is a widely used, but subjective method for assessing leg weakness and lameness in broilers (Weeks et al., 2002). The results of this study show a significant correlation between the body weight of broilers and the standing times during the LTL test. Body weight may significantly influenced lameness as indicated by a strong correlation among heavier broilers having shorter standing times (Table 1). Kestin et al. (2001) was the first study to show a clear relationship between body weight and lameness, across a wide range of genotypes including fast and slow growing birds. The previous study showed a threshold weight of 1.25 kg at 54 days of age, with birds becoming increasingly more lame in a linear relationship as body weight increased. The results of the present study are consistent with Kestin et al. (2001) where weight gain was associated with lameness and leg weakness. Leg weakness and lameness in broilers is also attributed to the rapid growth of the birds as at slaughter age skeletal maturity has not been reached resulting in softer skeletal bones. To compensate for this, there is an increased deposition in the outer layers of the cortical bones resulting in cortical bones that have high porosity and low mineral content (Venalainen et al., 2006; Sherlock et al., 2010)
Individual diet LTL analysis revealed birds fed Diets C (limestone, high calcium concentration) and Diet D (marine, low calcium concentration) were statistically superior compared with the "industry standard" reference diet, Diet A (Table). The results show that a lower total calcium concentration in general is desirable and that a low concentration of a more digestible calcium source performed comparably with a higher total calcium concentration of limestone. These findings suggest that a more digestible calcium source may be effective when used at low concentrations.
Further research is required to better understand the usefulness of a novel marine Ca source on broiler leg health, bone mineral content and bone histology. Although this research is a pilot study only it provides a starting point for understanding the effect of total Ca concentrations and Ca sources on broiler leg health. Further evaluation (economic and nutritional) of the novel marine Ca source is required before it can be considered for commercial use.
ACKNOWLEDGEMENTS
This study was funded by AB Vista Feed Ingredients. The senior author is in receipt of a scholarship from the Poultry CRC. Associate Professor Peter Thomson (University of Sydney) is thanked for his assistance with statistical analysis.
REFERENCES
Berg C and Santora GS (2003) Animal Welfare 12, 655-659.
Garner JP, Falcone C, Wakenell P, Martin M, and Mench JA (2002) British Poultry Science 43, 355-363.
Kestin SC, Gordon S and Sorensen P (2001) The Veterinary Record 148, 195-197.
Selle PH, Cowieson AJ and Ravindran V (2009) Livestock Science 124, 126-141
Sherlock L, Demmers TG, Goodship AE, McCarthy ID, and Wathes CM (2010) British Poultry Science 51 22-30.
Venalainen E, Valaja J and Jalava T (2006) British Poultry Science 47, 301-310.
Waldenstedt L (2006) Animal Feed Science and Technology 126, 291-307.
Weeks CS, Knowles TG, Gordon RG, Kerr AE, Peyton ST, and Tilbrook NT (2002) The Veterinary Record 21, 762-764.
This paper was presented at the 23rd Annual Australian Poultry Science Symposium, Sydney, New South Wales, February 19-22, 2012 organized by the Poultry Research Foundation (University of Sidney) and the World´s Poultry Science Association (Australian Branch). Engormix.com thanks the University, the WPSA and the authors for this huge contribution.
