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The Efficacy of Quantum Phytase in a Forty-Week Production Trial Using White Leghorn Laying Hens Fed Corn-Soybean Meal-Based Diets

A. L. Hughes^*, J. P. Dahiya^*, C. L. Wyatt and H. L. Classen^*,1

^* Department of Animal and Poultry Science, University of Saskatchewan, Saskatoon, Saskatchewan, Canada S7N 5A8; Syngenta Animal Nutrition Inc., Research Triangle Park, NC 27709

¹ Corresponding author: hank.classen{at}usask.ca

	ABSTRACT

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Microbial phytase is a prominent feed enzyme used in animal feeds, but there is relatively little information on its use in laying hen diets. In this experiment, an Escherichia coli 6-phytase (Quantum) was evaluated for its efficacy in a 40-wk laying hen production trial. A total of 1,080 White Leghorn hens (540 each of Shaver and Bovan strains) were fed mash corn-soybean meal-based diets containing 0.35% (positive control, PC), 0.25% (negative control, NC1), or 0.15% (NC2) nonphytate phosphorus (NPP). Six more diets were manufactured by supplementing the negative control diets with 200, 400, and 600 U/kg of exogenous phytase, resulting in a total of 9 treatments. Each dietary treatment x strain subclass was replicated 4 times with 5 adjoining cages per replicate (3 hens per cage) in a randomized complete block design. Production performance was measured from 21 to 61 wk of age. Only minor differences in production characteristics were found between the PC and NC1 treatments regardless of phytase addition, indicating that 0.25% NPP resulted in P intake that was at or above the hen’s requirement. In contrast, the hens fed 0.15% NPP diet without phytase supplementation had significantly (P < 0.05) reduced total hen housed egg production and body weight at 61 wk of age in comparison to the PC treatment, whereas the incidence of soft-shelled, cracked, and broken eggs was increased significantly (P < 0.05) in hens fed the NC2 diet. Addition of phytase to the NC2 diet improved these production characteristics to levels equal or better than the PC diet. The results indicated that Quantum phytase was efficacious in corn-soybean meal-based diets fed to White Leghorn laying hens and can be used to reduce diet supplementation with inorganic phosphorus.

Key Words: phytase • efficacy • laying hen • production performance • phosphorus

	INTRODUCTION

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Phytate is the primary storage form of phosphorus found in plant feed ingredients and accounts for approximately two-thirds of the total phosphorus in plant seeds (Cosgrove, 1966; Maenz, 2001). Phytate phosphorus in feed is poorly utilized by monogastric animals despite the presence of phytase activity in the brush border membrane of their digestive tracts (Maenz and Classen, 1998). As a consequence, inorganic phosphorus is added to feed to facilitate optimal growth and production. This practice ultimately leads to a large portion of dietary phosphorus not being utilized by the animal and being excreted in feces. Exogenous phytase can be added to diets to hydrolyze phytate within the digestive tract, making more phytate phosphorus available for use by the animal and decreasing the need for dietary inorganic phosphorus supplementation (Van Der Klis et al., 1997; Maenz, 2001).

Microbial phytase is now the prominent feed enzyme used in animal diets. Extensive research has been conducted on the use of phytase in broilers (Simons et al., 1990; Broz et al., 1994; Denbow et al., 1995; Rutherfurd et al., 2004), but research on its use in laying hen diets is less extensive.

Punna and Ronald (1999) found that incorporating 300 U/kg of phytase into a laying hen diet containing 0.1% nonphytate phosphorus (NPP) resulted in an improvement in egg production and feed intake along with a decrease in mortality. Um and Paik (1999) and Lim et al. (2003) found that supplementing a low phytate diet with phytase increased egg production in laying hens. Van Der Klis et al. (1997) found that production performance was significantly improved by dietary supplementation, whereas Jalal and Scheideler (2001) found that phytase supplementation improved feed intake, feed conversion, and egg mass. Gordon and Roland (1997) and Francesch et al. (2005) found that laying hens consuming a diet low in NPP with supplementary phytase performed as well as hens that were fed diets containing higher levels of NPP without supplementary phytase. Gordon and Roland (1997) saw an improvement in egg production, feed consumption, egg weight, and egg-specific gravity, and Francesch et al. (2005) saw an improvement in egg production, hen weight gain, and feed consumption in hens that were fed a diet low in NPP with supplementary phytase when compared with hens fed a low NPP diet without supplemental phytase. Gordon and Roland (1997) also indicated that phytase supplementation of diets containing high levels of NPP gave no further improvements in hen performance.

It should be remembered that all phytase products are not equal. Phytase enzymes differ in the source from which they are derived. They may differ in characteristics such as pH optimum, thermostability, and ability to resist hydrolysis within the digestive tract. Any difference in these characteristics will affect the ability of the phytase enzyme to function effectively and consistently within the digestive tract (Onyango et al., 2005). Therefore, all phytase enzymes produced must be tested in vivo to ensure efficacy before they are introduced to the monogastric feed market.

Previously, there has been little work done on the effect of an E. coli-derived 6-phytase (Quantum phytase) supplementation in laying hen diets. The present study was, therefore, conducted to assess the efficacy of Quantum phytase in a 40-wk production trial using White Leghorn laying hens fed corn-soybean meal-based diets and varying levels of NPP.

	MATERIALS AND METHODS

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Experimental protocol was approved by the Animal Care Committee of the University of Saskatchewan and was performed in accordance with recommendations of the Canadian Council on Animal Care (1993) as specified in the Guide to the Care and Use of Experimental Animals.

Animals and Housing

At 17 wk of age, a total of 1,080 White Leghorn pullets (540 each of Shaver White and Bovan strains) were housed in laying cages under controlled climate conditions at the Poultry Centre on the University of Saskatchewan campus. Three birds were placed in each cage (cage dimensions 30.5 x 46 cm with a height of 52 cm; floor space per hen = 468 cm²), and each experimental unit consisted of 5 adjoining cages. At 18 wk of age, the length of the light period was increased from 8L:16D to 14L:10D with a light intensity of 10 lx. The ambient temperature was maintained at a minimum of approximately 20°C through out the trial. During the preexperimental period (i.e., up to 21 wk of age) a commercial laying hen diet was offered ad libitum, whereas from 21 wk onwards, the hens were randomly assigned to 1 of the 9 dietary treatments. Each dietary treatment x strain subclass was replicated 4 times with 5 adjoining cages per replicate (3 hens per cage) in a randomized complete block design.

Experimental Diets

The ingredient and nutrient composition of the experimental diets is shown in Table 1. Three isocaloric (2,900 kcal/kg) and isonitrogenous (16.13% CP) corn-soybean meal-based diets based on National Research Council (1994) recommendations were formulated to contain 3 levels of NPP; 0.35% (positive control, PC), 0.25% (negative control 1, NC1), and 0.15% (negative control 2, NC2). The Ca supply was equal (3.8% of the diet) in all dietary treatments. The PC diet was fed without supplemental phytase. Three levels of phytase (200, 400, and 600 U/ kg) were added to each negative control diet. The phytase used was an evolved E. coli 6-phytase (Quantum phytase, Syngenta Animal Nutrition Inc., Research Triangle Park, NC) optimized for improved gastric and thermal tolerance and expressed in Pichia pastoris. Experimental diets were fed from 21 to 61 wk of age. Birds had free access to feed and water throughout the experiment. Experimental diets were analyzed for total phosphorus and calcium levels using the method of Zasoski and Barau (1977).

Egg production was recorded on a replication basis 5 d per wk and was then corrected to a 7 d per wk basis. Eggs were classified as normal, broken, cracked, soft-shelled, double-yolked, and abnormal; abnormal was defined as eggs that were misshapen or mini. The egg classification was based on all the eggs collected during the experiment. All eggs from one day were collected and weighed on a replication basis every 4 wk. Subsequently, the same eggs were used to assess specific gravity using 9 saline solutions ranging from 1.060 to 1.100 with 0.005 increments. Saline solutions were calibrated before each test. Feed intake was determined on a replication basis by weighing back all feed every 4 wk. Body weight of all birds was measured at 21, 41, and 61 wk of age. Mortality was recorded on a replication basis. Dead birds were collected, weighed, and recorded daily.

Statistical Analyses

The data were analyzed as 2 separate experiments using PC as the control group with each NC treatment. Each set of experimental data was analyzed as a 5 x 2 factorial arrangement (5 experimental diets x 2 strains) using the Proc GLM procedure of SAS (SAS Institute, 2002). Dun-can’s multiple range test was used to separate the means when ANOVA was significant, and regression analyses were used as appropriate. Differences were considered significant when P < 0.05.

	RESULTS

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The chemical analysis of the experimental diets is shown in Table 1. The calculated and analyzed total P levels were in good conformity. The analyzed Ca level was higher than the planned level, but it was consistently higher across the diets. The high Ca level was due the fact that the soybean meal contained much higher Ca (0.63%) than the expected level (0.25%).

PC and NC1 Comparisons

Only minor nonsignificant differences were observed between the 0.35% NPP (PC) and 0.25% NPP without phytase supplementation (NC1) treatments for most of the production characteristics studied (Table 2). The hens fed NC1 diet without exogenous phytase supplementation had comparatively high incidence of abnormal eggs compared with the PC treatment, but the differences were not significant. The addition of phytase to the NC1 diet resulted in production performance values that were similar to those observed for the PC treatment. There were no significant (P > 0.05) differences observed for total hen housed egg production (THHP), mortality, feed to egg mass ratio, feed intake, egg weight, body weight at trial end, incidence of soft-shelled, cracked, and broken eggs, and incidence of double-yolked eggs when comparing the PC and NC1 treatments, regardless of phytase supplementation. Total hen day egg production (THDP), body weight at the end of trial, and egg-specific gravity were significantly (P < 0.05) higher for Shaver hens, whereas incidence of soft-shelled, cracked, and broken eggs, double eggs, abnormal eggs, feed to egg mass ratio, feed intake, and egg weight were significantly lower for the Shaver hens compared with Bovan hens. There was no dietary treatment x strain interaction observed for any of the variables studied.

Hens consuming 0.15% NPP diet (NC2) without phytase supplementation had significantly (P < 0.05) reduced THHP and body weight at the end of trial in comparison to the PC treatment, whereas the incidence of soft-shelled, cracked, and broken eggs was increased significantly in hens fed the NC2 diet (Table 3). Although the mortality rate was around 70% higher in the NC2 treatment, the difference was not statistically significant. The addition of microbial phytase to the 0.15% NPP diet resulted in an improvement in the production characteristics (mainly THDP, THHP, body weight at the trial end, feed to egg mass ratio, and the incidence of soft-shelled, cracked, and broken eggs) to levels that were equal to or greater than those observed for the PC treatment. There was a linear improvement in THDP, THHP, feed to egg mass ratio, and mortality with the addition of exogenous phytase (200, 400, and 600 U/kg diet) to the NC2 diet.

	DISCUSSION

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The results of the current experiment showed that the production performance of the laying hens fed diet containing 0.25% NPP (NC1) was not significantly different from those that were fed diet containing 0.35% NPP (PC), regardless of phytase addition. The data demonstrated that the 0.25% NPP (NC1) diets provided sufficient phosphorus intake to support maximum hen performance and therefore were at or above the hen’s requirement. Our results showed that laying hens can maintain optimal health and production when fed a diet containing 0.25% NPP, provided feed intake is normal. The National Research Council (1994) recommends that white-egg layers require 250 mg of NPP per day per hen if the feed intake is 100 g per day to maintain optimal health and production. Previous research conducted by Um and Paik (1999) also found that the supplementation of phytase to diets containing 0.24% NPP gave no further improvements in hen performance.

In contrast to the PC and NC1 comparisons, the results showed that feeding 0.15% NPP diet (NC2) caused significant reductions in hen performance when compared with the PC treatment. It demonstrates that 0.15% NPP was insufficient at providing hens with their daily P requirement. Phytase supplementation completely corrected the significant reductions in hen performance that was caused by a diet deficient in available P. Our results on the effects of phytase supplementation to a 0.15% NPP diet on hen production performance were similar to those found by other workers.

Previous research has found that supplementing a diet containing 0.10 to 0.15% NPP with phytase resulted in a significant improvement in laying hen egg production when compared with the same diet that was not supplemented with exogenous phytase (Punna and Roland, 1999; Keshavarz, 2000; Lim et al., 2003; Wu et al., 2006). We observed that there was a linear increase in both hen day egg production and hen housed egg production with the addition of phytase to the 0.15% NPP diet. Hens consuming a diet containing reduced levels of NPP without phytase supplementation were least efficient in terms of feed conversion (Jalal and Scheideler, 2001). The addition of phytase to 0.1% (Jalal and Scheideler, 2001) and 0.2% NPP (Scott et al., 2001) diets has the ability to significantly improve feed efficiency. We found that the diet containing 0.15% NPP (NC2) without enzyme addition was the least efficient in terms of feed conversion (feed to egg mass ratio). Lim et al. (2003) documented that phytase supplementation decreased the percentage of broken and soft-shelled eggs. There was a quadratic decrease in the percentage of soft-shelled, broken, and cracked eggs with the addition of phytase to the NC2 diet in the current study.

It has been previously found that the body weight of hens fed a diet containing 0.1% NPP was significantly less than that of hens fed the same diet with the addition of phytase (Gordon and Roland, 1997; Van Der Klis et al., 1997). The results of our study are in agreement with these previous studies. The body weight of the hens fed the 0.15% NPP diet was significantly lower than that of the hens fed the same diet with phytase supplementation. Higher mortality in hens consuming diets deficient in NPP have been found by Punna and Roland (1999) and Jalal and Scheideler (2001). Our data showed that there was a linear decrease in mortality with the addition of phytase to the NC2 diet.

A few production characteristics were unaffected by the NPP deficiency or exogenous phytase supplementation, this includes egg weight, egg-specific gravity, feed intake, and the incidence of double-yolked and abnormal eggs. Punna and Roland (1999) found that phytase supplementation increased egg weight in hens fed a diet containing 0.1% available phosphorus, but had no effect with 0.2, 0.3, and 0.4% available phosphorus diets. We may have seen a difference in egg weight if the dietary NPP level in this study would have been lower than 0.15%. Boling et al. (2000) also found that there were no significant differences in egg-specific gravity, regardless of NPP level or phytase inclusion. It is unexplainable why we did not see a significant effect of NPP level on egg-specific gravity in our study. There was a clear decrease in egg shell quality as shown by the significant increase in soft-shelled, cracked, and broken eggs when P was removed from the diet. The significant increase in soft-shelled, cracked, and broken eggs is an indication that egg shell quality has diminished, and therefore one might also expect a response in egg-specific gravity. Contrary to the results of present study, Gordon and Roland (1997) and Jalal and Scheideler (2001) documented that the feed consumption increased in hens fed diets containing 0.1% NPP supplemented with phytase. An increase in feed intake with phytase supplementation may have been seen because of the more severe P deficiency in a 0.1% NPP diet.

The results indicate that the addition of phytase to diets deficient in NPP improved hen production characteristics up to levels seen for hens fed a diet that is adequate in NPP. This shows that Quantum phytase is efficacious in corn-soybean meal diets fed to White Leghorn laying hens and can be used to reduce diet supplementation with inorganic P. Therefore, Quantum phytase can be added to diets deficient in NPP to ensure that birds consuming these diets are provided with adequate P for optimal health and production throughout the laying period. Many producers are concerned about production losses caused by diets deficient in NPP. If producers are hesitant to feed 0.15% NPP with phytase supplementation, we recommend that phytase be fed with a 0.25% NPP diet. National Research Council (1994) recommends that a hen’s daily P requirement is 250 mg. Most producers feed diets high in inorganic P containing approximately 0.35% NPP to ensure that P requirements are met. In place of inorganic P, we suggest that Quantum phytase be fed on top of a 0.25% NPP diet as a safety margin. As a bonus, replacing inorganic P with exogenous phytase has some benefits—it is more environmentally friendly due to reduced P excretion, and in the long run, may be more cost effective.

	ACKNOWLEDGMENTS

 
The authors would like to acknowledge the Syngenta Animal Nutrition Inc. for providing Quantum phytase and funding support for this project, and the staff at the University of Saskatchewan Poultry Centre for their assistance. The technical assistance of Dawn Abbott (University of Saskatchewan, Saskatoon, Canada) is greatly appreciated.

Received for publication December 14, 2007. Accepted for publication March 2, 2008.

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This Article Free Via Open Access Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in Web of Science Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Hughes, A. L. Articles by Classen, H. L. Search for Related Content PubMed PubMed Citation Articles by Hughes, A. L. Articles by Classen, H. L.