Effect of Bacterial Phytase on Growth Performance, Nutrient Utilization, and Bone Mineralization in Broilers Fed Pelleted Diets

**Background:** Microbial phytases are widely used in poultry nutrition to hydrolyze phytate complexes and improve phosphorus (P) and calcium (Ca) bioavailability. However, the optimal dietary Ca:P ratio in the context of phytase supplementation at varying dose levels remains poorly defined. High dietary Ca can form insoluble Ca-phytate complexes that resist enzymatic digestion and reduce phytase efficacy. This study aimed to evaluate the effects of maintaining a constant Ca-to-available P (avP) ratio (resulting in reduced Ca levels) alongside bacterial 6-phytase supplementation on growth performance, nutrient digestibility, blood parameters, and bone mineralization in broilers fed pelleted diets from day 1 to 35.

**Methods:** A total of 630 one-day-old male Ross 308 broilers were allocated to 7 dietary treatments (6 pens/treatment, 15 birds/pen) in a completely randomized design. The positive control (PC) diet was formulated to meet minimum nutrient requirements. Two negative control diets were formulated: NC1 (reduced by 2.0 g/kg Ca and 1.0 g/kg avP) and NC2 (reduced by 3.0 g/kg Ca and 1.5 g/kg avP). Four additional diets comprised NC1 and NC2 supplemented with 500 or 1000 FTU/kg of bacterial 6-phytase (Endo-Phos®, from E. coli). A constant Ca:avP ratio was maintained across all diets. Diets were steam-conditioned at 85°C for 30 seconds and pelleted. Body weight and feed intake were measured on days 1, 21, and 35. On day 35, ileal digesta was collected from 3 birds/pen for CAID determination using titanium dioxide as an indigestible marker. Tibia and toe samples (2 birds/pen) were analyzed for ash, Ca, and P content. Blood samples (2 birds/pen) were analyzed for alkaline phosphatase (ALP), Ca, and P. Excreta P concentration was measured from samples collected over two consecutive days.

**Key Results:** Phytase recovery was close to expected in starter diets (621 and 1035 FTU/kg for 500 and 1000 FTU/kg targets) and finisher diets at 500 FTU/kg (537 FTU/kg), but slightly lower in the finisher 1000 FTU/kg diet (925 FTU/kg). Birds fed NC1 and NC2 diets without phytase had significantly reduced BWG and worsened FCR compared to PC (p < 0.01). Phytase supplementation at both 500 and 1000 FTU/kg restored BWG and FCR to levels equivalent to PC across all phases. Notably, birds fed NC2 + 1000 FTU/kg had 5.6% higher BWG (1940.4 g vs. 1836.6 g, p < 0.01) and 4.1% better FCR (1.452 vs. 1.514, p < 0.01) compared to PC over days 1–35. Feed intake did not differ among treatments (p = 0.47). Phytase at 1000 FTU/kg maintained CAID of P equivalent to PC (0.783 and 0.761 for NC1 and NC2, respectively, vs. 0.762 for PC). CAID of N and gross energy were lower in all NC diets compared to PC (p < 0.01). Phytase linearly increased CAID of N (p < 0.01), Ca (p < 0.05), and P (p < 0.01) in the NC1 diet. Serum Ca was lower in NC diets (8.33 and 8.5 mg/dL for NC1 and NC2) vs. PC (10.09 mg/dL, p = 0.001), and phytase supplementation restored serum Ca to PC-equivalent levels. Serum ALP and P did not differ among treatments (p > 0.05). Tibia ash tended to differ among treatments (p = 0.08), with NC2 + 500 and NC2 + 1000 FTU/kg showing numerically higher values than PC. Toe ash was significantly higher in NC2 + 500 FTU/kg (11.52%) and NC2 + 1000 FTU/kg (11.25%) compared to PC (10.09%, p = 0.006). Excreta P concentration was reduced by approximately 27% with phytase supplementation (p < 0.01), with the lowest values in NC1 + 1000 FTU/kg (0.42%) and NC2 diets (0.442–0.537%) vs. PC (0.702%).

**Clinical Implications:** This study demonstrates that bacterial 6-phytase at 500–1000 FTU/kg can effectively replace 1.5 g/kg avP and 3 g/kg Ca in broiler diets when a constant Ca:avP ratio of approximately 2 is maintained. The 1000 FTU/kg dose in moderately Ca- and P-deficient diets yielded superior growth performance and bone mineralization compared to the nutritionally adequate control, likely due to reduced formation of insoluble Ca-phytate complexes and improved phytate hydrolysis. The 27% reduction in excreta P highlights the environmental benefits of phytase supplementation. Poultry nutritionists should consider reducing dietary Ca levels in phytase-supplemented diets rather than maintaining safety margins, as excessive Ca impairs phytase efficacy and P utilization. A dose of 1000 FTU/kg is recommended for optimal digestible Ca and P supply, though 500 FTU/kg may suffice for adequate bone mineralization in moderately deficient diets.