A comparison of long bone development in historical and ...

MOLECULAR, CELLULAR, AND DEVELOPMENTAL BIOLOGY A comparison of long bone development in historical and contemporary ducks R. C. Van Wyhe,* T. J. Applegate,† M. S. Lilburn,‡ and D. M. Karcher*1 *Department of Animal Sciences, Michigan State University, East Lansing 48824; †Department of Animal Sciences, Purdue University, West Lafayette, IN 47907; and ‡Department of Animal Sciences, The Ohio State University, Ohio Agricultural Research and Development Center, Wooster 44691 ABSTRACT The selection for growth and carcass traits in poultry meat species has contributed to increased interest in understanding and characterizing skeletal growth as the birds struggle to balance skeletal development with increased BW and muscle mass. The objective of this study was to compare the physical characteristics and mineralization of the tibia and femur from commercial Pekin ducks representing circa 1993 and 2010 commercial strains. In 1993, the femur and tibia were collected from 8 ducks at 11 ages between 11 and 53 d. A similar study was done in 2010 in which the femur and tibia were collected from 8 ducks at 12 sample ages between 10 and 49 d. All bones were weighed and the length and width at 50% of length were measured. Each bone was subsequently cut into epiphyseal (top 25% of length) and diaphyseal (midregion at 50% of length) sections. Each bone segment

was extracted with ether, hot weighed, and ashed. The 2010 contemporary ducks reached market weight faster than the 1993 ducks. Therefore, statistical comparisons were made at common BW as well as at common ages. The mean tibia length of the 2010 duck was 0.75 cm greater (P < 0.05) at similar ages and similar BW. The percentage ash in the diaphyseal region of the tibia was 3% greater (P < 0.05) in the 2010 versus 1993 ducks. The percentage epiphyseal ash in the femur was 10% lower (P < 0.01) at 10 d and 14 d in the 2010 ducks but there were no significant differences by 18 d of age. The lower epiphyseal ash values at both younger ages and smaller BW in the 2010 contemporary ducks suggests that it is critical to monitor those factors that influence bone mineralization in contemporary ducklings that can achieve market BW at earlier chronological ages.

Key words: bone, duck, mineralization, tibia, femur 2012 Poultry Science 91:2858–2865 http://dx.doi.org/10.3382/ps.2012-02385

INTRODUCTION The selection for increased BW at a common age and proportional increases in muscle mass in commercial meat strains of poultry has not necessarily resulted in concomitant increases in skeletal integrity. Tibial dyschondroplasia, femoral fractures, and long bone distortions are some of the skeletal issues that are most often associated with genetic increases in growth (Sullivan, 1994; Cook, 2000; Petek et al., 2005). These structural anomalies may contribute to increased mortality and morbidity that will reduce the overall well being of commercial meat genotypes. There are numerous factors that influence bone development, including nutrition, genetics, and management practices (Leeson, 1988; Bond et al., 1991; Edwards, 2000; Oviedo-Rondón et al., 2006; Talaty and Katanbaf, 2009). Examining the changes in bone characteristics and bone composition ©2012 Poultry Science Association Inc. Received April 4, 2012. Accepted July 17, 2012. 1 Corresponding author: [email protected]

over time may help identify those ages or BW at which the potential for skeletal problems to occur is the greatest. There are several traditional bone traits that are most often studied when one is trying to characterize skeletal development. Bone mineralization or ash is often determined as it is assumed to be highly correlated with the structural strength of a bone (Martin and Ishida, 1989; Martin et al., 1996). In laying hens, Mazzuco and Hester (2005) demonstrated that bone mineralization was negatively correlated with bone breakage strength in laying hen bones with reduced medullary bone content. A change in mineralization rates will influence bone strength by varying the structural properties of the bone. Given that not every bone contributes equally to the structural support of the bird, most studies have focused on long bone development in poultry as being representative of skeletal health. Commercial poultry species have several key long bones that contribute to their overall skeletal stability and a single bone does not necessarily represent the growth and development status of the entire skeletal

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system. The tibia has been used extensively as an indicator of optimal skeletal growth in poultry (Buckner et al., 1950; Norman and Hurwitz, 1993; Lilburn, 1994; Jendral et al., 2008). The femur, however, may be a better index of overall skeletal development given that it is more closely associated with overall bone mineral density than the tibia (Melton et al., 1999). In broiler chicks, Applegate and Lilburn (2002) reported that the femur and tibia have different mineralization rates. The humerus is part of the upper or thoracic portion of the body (nonweight bearing). It has been primarily studied in laying hens due to its different mineralization properties when compared with the tibia (Hester et al., 2004). The extent to which each region of a long bone is mineralized is not the same at a given age. The epiphyseal or proximal region of a long bone is primarily involved in longitudinal bone growth and has a lower percentage of bone ash than the diaphyseal region (Applegate and Lilburn, 2002; Dibner et al., 2007). The diaphyseal or middle region of the bone has a higher and more consistent percentage ash content throughout the life of the bird. Comparisons of the ash content in these different regions of the bone can thus be used as a descriptive index of skeletal development differences across ages and between genotypes. It has been documented that genetic differences exist in the strength and mineralization of growing long bones. Williams et al. (2000) reported that overall bone strength and mineralization were decreased in high weight versus low weight selected broiler lines. Talaty and Katanbaf (2009) subsequently reported that variability in bone mineral density among purebred broiler lines might prove useful as a selection tool. Chickens have been the primary poultry species studied with respect to skeletal development although similar genetic improvements in BW and the associated challenges with optimizing skeletal health have been observed in commercial turkeys and ducks (Applegate et al., 1999; C. M. Turk, Maple Leaf Farms, personal communication). Previous research on duck bone development is limited, but duck time to market weight is similar among ducks and chickens (Applegate and Lilburn, 2002). Ducks also suffer from similar skeletal problems that plague other species, such as tibial dyschondroplasia. Therefore, the primary objective of the study reported herein was to compare the physical and mineralization characteristics of the femur and tibia from contemporary and historical commercial Pekin ducks. A secondary objective was to compare the developmental status of the humerus to that of the femur and tibia in the contemporary 2010 ducks.

MATERIALS AND METHODS Experimental Design In 2010, 8 commercial Pekin drakes were randomly sampled from the same commercial flock at 10, 14, 18,

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21, 25, 29, 32, 35, 39, 42, 46, and 49 d of age. The right humerus, tibia, and femur were harvested from each bird. The data obtained from these bones were compared with similar data generated in 1993 from commercial Maple Leaf ducks. The 1993 bones were collected at 14, 18, 21, 25, 28, 32, 35, 39, 42, 46, 49, and 53 d of age. Body weight was obtained for each duck at the time of sampling. Ducks were reared following commercial conditions that were present for 1993 (litter flooring) and 2010 (raised plastic flooring). Besides flooring types, nutritional differences existed with ducks fed proprietary commercial diets during both sampling years. Bone parameters from the 1993 ducks were measured in a similar fashion to what was described by Applegate and Lilburn (2002). The adhering tissue from the 2010 bones was removed and the length and width (at 50% of length) of each bone was subsequently measured. All bones were cut with a bone saw into 2 sections. The epiphyseal section was defined as the proximal and distal 25% of the determined individual bone length and the diaphyseal was defined as the middle 50% of the bone. Bones were placed in a soxhlet and the fat removed via diethyl ether extraction for 48 h. Following fat extraction, bones were placed in crucibles and placed in a static air drying oven set at 105°C for at least 2 h. Dried bones were removed, hot weighed in the crucible, and placed in an ashing oven which was set at 600°C for 24 to 48 h. The remaining ash was weighed for final ash determination.

Statistics All data were analyzed using the GLM procedure of SAS version 9.2. Differences between means were tested using Tukey’s adjustment with significance accepted at P < 0.05. Body weight differences were analyzed using the estimates statement of PROC GLM. Contemporary (2010) ducks reached market weight faster than the 1993 ducks; therefore, statistical comparisons were made at similar BW (800 g, 1,600 g, 2,400 g, 3,150 g, 4,000 g, and 4,300 g) as well as at common ages (10, 14, 18, 25, 28/29, 32, 35, 39, 42, and 46 d). Age-based comparisons were conducted using the following model: the observation (yijk) was equal to the sum of the mean (μ), the main effect of strain (τi), the main effect of age (βj), the interaction between age and strain (τβij) and the error term (εijk), (yijk = μ + τi + τβij + εijk). The subscripts i and j represent the replicates for strain and age, respectively. The subscript k is the experimental unit representing a unique combination of i and j. All similar BW-based comparisons were conducted using the following model: the observation (yijk) is equal to the sum of the mean (μ), the main effect of strain (τi), the main effect of age (βj(i)), and the error term (εijk), (yijk = μ + τi + βj(i)+ εijk). All subscripts were the same as the age-based comparisons with the exception that j stands for replicates for BW in this model. The BW was unique to each duck and thus considered

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nested within strain, whereas age was considered crossclassified.

RESULTS In the 17 yr that passed between the 1993 and 2010 data sets used for this report, the Maple Leaf Farms genetics program significantly improved the performance of their commercial ducks. The BW selected for analysis, 0.8, 1.6, 2.4, 3.15, 4.0, and 4.3 kg, approximately correspond to 14, 21, 28, 35, 42, and 49 d of age for the 2010 ducks. The current-generation ducks are 200 g heavier than the 1993 duck at d 14 and reach market weight (3,100 g) approximately 7 d earlier.

Tibia Length and Width The tibia growth curve was curvilinear in both 1993 and 2010 (Figure 1A–D). Even though the tibia growth curves were similar, the 2010 duck tibias were approximately 0.75 cm longer at all common ages and common BW (Figure 1A,B). There was also a main effect of strain as the overall average for 2010 bones was significantly longer (P < 0.001). There was an age-by-strain interaction (P = 0.03) due to the 2010 ducks reaching mature length at a younger age (Figure 1A). The width

of the tibia remained relatively unchanged over the 17yr period between studies (Figure 1C,D). There was a main effect of age (P < 0.001), but no changes due to strain (year) or a strain-by-age interaction. Similarly, there was a BW main effect (P < 0.001) but no interaction of BW and strain on tibia width.

Tibia Ash The main effects of age and BW on tibia epiphyseal ash were both significant (P < 0.001) as the ash percentage increased with both duck age and BW (Figure 2A,B). There were no overall differences in tibia epiphyseal ash due to strain (P = 0.12). However, there was a strain by age (P < 0.001) and strain by BW (P = 0.03) interaction. At 800 g, 4,000 g, and 4,300 g of BW, there was an approximate 4% increase in epiphyseal ash in the 2010 ducks and no differences at 1,600 g and 3,150 g. The diaphyseal region is more mature physiologically and the percentage ash content is relatively constant throughout the production period. There was less than a 10% change in tibial diaphyseal bone ash over the ages studied in this report. While the overall change in percentage diaphyseal tibia ash was small, there was still an age effect (Figure 2C). Polynomial regression

Figure 1. A comparison of 1993 and 2010 duck tibial length and width. A) Regression of 1993 duck and 2010 duck tibial lengths by age. For 1993: Y = −0.0048x2 + 0.4315x + 2.4619, r2 = 0.99. For 2010, Y = −0.0046x2 + 0.397x + 4.101, r2 = 0.99. Means represent 8 birds per age. B) Regression of 1993 duck and 2010 duck tibial lengths on BW. For 1993, Y = −6E-07x2 + 0.0042x + 4.8845, r2 = 0.97. For 2010, Y = −4E-07x2 + 0.0031x + 6.5083, r2 = 0.93. C) Regression of 1993 duck and 2010 duck tibial lengths by age. For 1993, Y = −0.003x2 + 0.2869x + 0.9209, r2 = 0.98. For 2010, Y = −0.0019x2 + 0.2219x + 1.6485, r2 = 0.94. Means represent 8 birds per age. D) Regression of 1993 duck and 2010 duck tibial widths on BW. For 1993, Y = −3E-07x2 + 0.0026x + 2.7179, r2 = 0.90. For 2010, Y = −2E-07x2 + 0.0019x + 2.9369, r2 = 0.75. *Indicates significant difference between 1993 and 2010 duck strains at common age or BW (P < 0.05). Poly. = polynomial regression.


equations for a regression based on BW and diaphyseal ash could not be formulated with an acceptable R2 to be used for statistical analysis in the study (Figure 2D). There was a strain effect on diaphyseal ash, as the overall mean for tibial diaphyseal ash was higher (P < 0.001) in the 2010 (58%) compared with the 1993 ducks (55%). The tibia diaphyseal ash in the 2010 duck was already increased at d 10 and this difference was consistent through market age (42 d). There was also an interaction between age and strain (P < 0.001), as the differences in ash were not significant after d 42.

Femur Length and Width The femur had a curvilinear growth pattern with both age and BW main effects (Figure 3A,B). There was no significant effect due to strain or an interaction between age and strain although there was an interaction between BW and strain. Similar to what was observed for the tibia, femur length in the 2010 ducks was greater through 28 d but there were no age effects thereafter (Figure 3A). When comparing ducks at a common BW (Figure 3B), the 2010 ducks had a longer femur when BW was 800 g, whereas the 1993

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ducks had a longer femur at 2,400 g and 3,150 g of BW, which would have corresponded to older ages as well. Although, there was no effect of strain on the femur length, there was a significant effect of strain on femur width (Figure 3C,D). There were also main effects due to BW and age as well as interactions between both age and BW with strain. Although, no evident regression line of the BW-strain interaction was seen because the R2 value was low (Figure 3D).

Femur Ash The ash content in the epiphyseal region of the femur was highly variable throughout the ages sampled (P < 0.05). There were no significant strain effects on mean femur epiphyseal ash (Figure 4A). Although there was no main effect of strain, there was a significant interaction between age and strain. The biggest difference in ash between the 1993 and 2010 ducks compared at similar ages occurred in the epiphyseal region of the femur (Figure 4A). Mean femoral epiphyseal ash was 21% at 14 d in 2010 versus 32% in 1993. In the 2010 ducks, femoral epiphyseal ash increased rapidly and was higher than 1993 ducks by 29 d.

Figure 2. A comparison of 1993 and 2010 duck tibial epiphyseal and diaphyseal ash. A) Regression of 1993 duck and 2010 duck tibial epiphyseal ash by age. For 1993, Y = 4E-05x2 + 0.0006x + 0.3162, r2 = 0.93. For 2010, Y = −6E-05x2 + 0.0075x + 0.2043, r2 = 0.97. Means represent 8 birds per age. B) Regression of 1993 duck and 2010 duck tibial epiphyseal ash on BW. For 1993, Y = 9E-09x2 − 4E-06x + 0.3352, r2 = 0.71. For 2010, Y = −5E-09x2 + 7E-05x + 0.2472, r2 = 0.84. C) Regression of 1993 duck and 2010 duck tibial diaphyseal ash by age. For 1993: y = 1E-05x2 + 0.0002x + 0.5317, r2 = 0.66. For 2010, y = −9E-05x2 + 0.0048x + 0.5304, r2 = 0.51.. Means represent 8 birds per age. D) Regression of 1993 duck and 2010 duck tibial diaphyseal ash on BW. Overall means not significantly different. R-squared value too low for analysis using regression equations. *Indicates significant difference between 1993 and 2010 duck strains at common age or BW (P < 0.05). Poly. = polynomial regression; epi = epiphyseal; dia = diaphyseal.

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The ash content in the diaphyseal portion of the femur remained fairly constant over all the ages sampled, similar to what was observed for the tibia. There was a main effect of age as the ash content increased slightly with age (Figure 4C). For femoral diaphyseal ash the relationship between BW and diaphyseal bone ash was not analyzed using the regression equation as the R2 values were deemed too low (Figure 4D). The overall mean femoral diaphyseal ash was higher (P < 0.05) in the 2010 ducks (57%) compared with the 1993 strain (55%). There was a significant strain-by-age interaction (P < 0.05) due to increased ash in the 2010 ducks at d 18 and d 25.

Humerus Length and Width The 2010 humerus data was distinctly different from the femur and tibia. The humerus and tibia were the longest bones (Figure 5A). Figure 5B illustrates the humerus length was less than 30% of its 35 d length at 10 d but increased rapidly and was approximately the same length as the tibia by 49 d. The humerus did not reach 80% of its market size until 29 d or 95% of mature size until 32 d, whereas the tibia and femur

reached 80% of their market size by 21 d and 95% of their mature size by 29 d.

Humerus Ash The humerus ash responded differently over the course of the sample ages compared with the tibia and femur. The epiphyseal ash in the humerus did not increase with length. Instead, bone ash decreased in the humerus from hatch through d 25 and thereafter increased from d 29 until the end of the study. The humerus diaphyseal ash was slightly lower than the tibia and femur, but all bones had an overall mean of 55% (Figure 5D) or greater and there was a limited range of fluctuation (under 5%) in mineralization throughout life in all 3 bones. This indicates that the mineralization of the diaphyseal portion is relatively static throughout life.

DISCUSSION The curvilinear bone growth curves support the data with broilers reported by Bond et al. (1991) and Applegate and Lilburn (2002). That the tibia and femur were

Figure 3. A comparison of 1993 and 2010 duck femoral lengths and widths. A) Regression of 1993 duck and 2010 duck femur lengths by age. For 1993, Y = −0.0033x2 + 0.294x + 1.3257, r2 = 0.99. For 2010, Y = −0.0029x2 + 0.253x + 2.3543, r2 = 0.99. Means represent 8 birds per age. B) Regression of 1993 duck and 2010 duck femur lengths on BW. For 1993, Y = −4E-07x2 + 0.0028x + 2.9961, r2 = 0.96. For 2010, Y = −3E07x2 + 0.002x + 3.8412, r2 = 0.93. C) Regression of 1993 duck and 2010 duck femur widths by age. For 1993, Y = −0.002x2 + 0.2016x + 2.6151, r2 = 0.97. For 2010, Y = −0.00153x2 + 0.1458x + 2.3127, r2 = 0.92. Means represent 8 birds per age. D) Regression of 1993 duck and 2010 duck femur widths on BW. Overall means for 1993, 6.86 mm, for 2010, 5.12 mm (P < 0.001). R-squared value too low for analysis using regression equations. *Indicates significant difference between 1993 and 2010 duck strains at common age or BW (P < 0.05). Poly. = polynomial regression.


longer at the same age in the faster-growing 2010 ducks has also been shown in different lines of turkeys (Lilburn and Nestor, 1991) and broiler chickens (Reddish and Lilburn, 2004). However, it was not anticipated that the 2010 ducks would have longer bones at the same BW. A longer bone at the same BW suggests that selection for increased growth rate in ducks may also exert indirect selection pressure for increased skeletal support in the heavier, 2010 ducks. We do not have a rationale for why these changes were only observed in the tibia. The 17 yr between experiments and concomitant changes in growth rate did, however, result in other skeletal changes that were only observed in the femur. Femur length in the 2010 ducks was greater through 28 d but there were no age effects thereafter. The decrease in days to market weight could have contributed to the observed differences in femur length. The 2010 ducks are obviously growing more rapidly and the data suggests that at younger ages and heavier BW, the femur may be adapting to these changes in growth. These differences were only observed at younger ages, however, suggesting that the mature size of the femur has not changed in 17 yr. The femur connects the upper body with the lower skeleton and is responsive to both changes in BW as well as conformation of the ani-

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mal. Due to the load-bearing nature of the bone, it may be more resistant to longer-term changes in architecture (length, width). Nestor et al. (1987) reported that direct selection for one aspect of skeletal development (shank width) in a fast-growing turkey line increased that particular trait without changing other components of the lower skeletal axis. The humerus, femur, and tibia demonstrated different growth patterns, which supports previous research from Bruno et al. (2007). This is different than what was observed by Church and Johnson (1964), who reported that the humerus has a similar growth curve as the tibia and femur. Humerus data were not collected in 1993, so it is unclear what changes may have occurred in the years between experiments. The tibia and femur responded differently with respect to changes in width. The increase in tibia length would be expected to correlate positively with increased tibia width indicating an overall larger bone. Conversely, femur width was expected to be similar in both contemporary and modern ducks later in life. The 1993 ducks having a shorter bone early in life would be expected to have a narrower width early in life. This again would correlate with an overall decrease in bone size early in life. Bone width is strongly correlated with

Figure 4. A comparison of 1993 and 2010 duck femoral epiphyseal and diaphyseal ash. A) Regression of 1993 duck and 2010 duck femur epiphyseal ash by age. For 1993, Y = 7E-05x2 − 0.0027x + 0.3523, r2 = 0.82. For 2010, Y = −0.0002x2 + 0.0169x + 0.0418, r2 = 0.81. Means represent 8 birds per age. B) Regression of 1993 duck and 2010 duck femur epiphyseal ash on BW. Overall means not significantly different. Rsquared value too low for analysis using regression equations. C) Regression of 1993 duck and 2010 duck femur diaphyseal ash by age. R-squared value too low for analysis using regression equations. Means represent 8 birds per age. D) Regression of 1993 duck and 2010 duck femur diaphyseal ash on BW. Overall means, for 1993, 54.8%, and for 2010, 57.1%. R-squared value too low for analysis using regression equations. *Indicates significant difference (P < 0.05). Poly. = polynomial regression; epi = epiphyseal; dia = diaphyseal.

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bone strength and narrow bones lead to increased fractures in human males (Tommasini and Nasser, 2005). A hypothesis could be developed that if bone length continues to increase without proportionate changes in bone width, this could predispose birds to increased skeletal problems. The epiphyseal region of the bone is responsible for the linear increase in bone growth and the process involves chondrocyte replication in the growth plate, maturational development, and ultimately mineralization. The percentage ash in this region of bone would be expected to vary considerably with age because of the physiological changes that are occurring, particularly during the early stages of growth (Applegate and Lilburn, 2002). If the low epiphyseal bone ash values in the 2010 ducks at the early ages are correlated with total bone strength, as suggested by Rath et al. (2000) and Seeman (2008), this certainly emphasizes the importance of early diet and management strategies geared to maximizing optimal skeletal development. The tibia and humeral epiphyseal ash did not exhibit the same trends that were observed in the femur. These

small differences in the tibia may not be critical to overall skeletal development but certainly suggest that the tibia has not been affected by selection over time nearly to the extent as the femur. The observations of epiphyseal ash run counter to the bone length data where the tibia showed the greatest response to selection effects on BW. Few studies have looked at epiphyseal ash in the humerus but total ash or mineral content in the humerus bone has been shown to increase throughout life in varying purebred chicken lines (Talaty and Katanbaf, 2009). The cause of the bone ash decrease in the current study is unclear; it may simply be a reflection of the normal variability in humeral bone ash in ducks. The diaphyseal portion of the bone is the more mature portion of the bone and the bone ash percentage will be more consistent than the epiphyseal end. The femur, humerus, and tibia had less than a 10% change in diaphyseal bone ash throughout life. Previous studies have shown similar results (Lilburn, 1994; Applegate and Lilburn, 2002). Though diaphyseal and epiphyseal ash were examined, total bone ash was not examined in the 1993 duck, so it is not pos-

Figure 5. A comparison of femoral, humoral, and tibial bone parameters in 2010 ducks. A) Regression of 2010 duck femur, humerus, and tibia length by age. Femur, −0.0029x2 + 0.253x + 2.3543, r2 = 0.99. Humerus, −0.0059x2 + 0.5989x − 2.6755, r2 = 0.99. Tibia, −0.0046x2 + 0.397x + 4.101, r2 = 0.99. Means represent 8 birds per age. Tibia and femur significantly different at all ages (P < 0.05). B) Fractional tibia, femur, and humoral length (after fat extraction) as a proportion of the measure at d 35, approximately market size. Means represent 8 birds per age. C) Regression of 2010 duck epiphyseal ash by age. For femur, Y = −0.0002x2 + 0.0169x + 0.0418, r2 = 0.81. For humerus, Y = 0.0003x2 − 0.0122x + 0.4053, r2 = 0.81. For tibia, Y = −6E-05x2 + 0.0075x + 0.2043, r2 = 0.97. Means represent 8 birds per age. D) Femur, humerus, and tibia diaphyseal ash by age. R-squared value too low for analysis using regression equations. *Indicates all bones are significantly different from one another (P < 0.05). ◊Indicates significant difference for humerus and tibia (P < 0.05). ‡Indicates the femur is significantly different from the humerus and tibia, but the tibia is not significantly different from the humerus (P < 0.05). +Indicates the humerus is significantly different from the tibia and femur, but the tibia is not significantly different from the femur (P < 0.05). ∞Indicates the tibia is significantly different from the humerus and femur, but the femur is not significantly different from the humerus (P < 0.05). Poly. = polynomial regression; Epi= epiphyseal; Dia = diaphyseal.


sible to compare total ash. It is unclear from previous studies how ducks compare with chickens as total ash results have varied (Bond et al., 1991; Rath et al., 2000; Mutuş et al., 2006). The results of the study show how important it is to examine multiple bones when looking at skeletal development. Each bone exhibits different changes over time (years) and suggests that a single bone is not necessarily representative of overall changes in skeletal development, particularly in species that are continually under selection pressure for traits of commercial importance. There were several changes in skeletal development reported herein that occurred during the 17-yr period between sample collection. There could be several factors that contributed to the observed changes, such as nutrition, flooring system, or genetics. However, the data does not lend itself to a single, simple reason for why the changes occurred. Future research may want to examine the female ducks as well, due to sex differences in skeletal development.

ACKNOWLEDGMENTS The authors kindly thank Maple Leaf Farms (Leesburg, IN) for their cooperation and partial research support.

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