DOI: 10.25952/bnf9-3b78
title: Sex Effects and the Use of the ANCOVA Model

description: <p><b>Chapter 4 dataset</b><br />
With known variation in performance between male and female broilers and the fact that sourcing single-sex birds for use in research is becoming increasingly difficult, it becomes important to determine the effect of rearing method with male and female broilers on between-pen variation and body weight (BW) uniformity. We evaluated the performance response of broilers reared as single or mixed-sex to standard and reduced crude protein (CP) diets. The study was designed as a 2 × 3 factorial arrangement of treatments consisting of 672 Cobb-500 broilers assigned to 48 floor pens with 6 treatments, 8 replicates, and 14 birds per pen. The factors were: rearing method (male single-sex, female single-sex or equally mixed-sex) and dietary CP level (standard or reduced). For the overall period of the trial (d 0-35) there was a significant effect (P < 0.001) of rearing method and CP level on feed intake (FI) and feed conversion ratio (FCR). There was also a significant interaction between rearing method and CP level for BWG during d 0-35 (P < 0.01).  There was a significant interaction between CP level and sex on d 34 BW (P < 0.01) where the reduced CP diet decreased the BW of both males and females, but to a greater extent the BW of the female birds. Dietary CP level had a significant effect on relative breast and drumstick weights with birds fed the reduced CP diet having significantly lower breast weights (P < 0.001) and higher drumstick weights (P < 0.01). This study suggests that male and female broilers have different CP requirements, and rearing birds as equally mixed-sex results in the lowest CV% for performance parameters and best BW uniformity compared to single-sex birds. Furthermore, when low CP diets are fed to broilers, they will prioritize the growth of more important body parts such as the legs.
</p><p><b>Chapter 5 dataset</b><br />
The availability of sexed day-old broiler chicks is becoming an issue as feather sexing is no longer possible. This has great implications for broiler researchers as the use of randomly distributed mixed-sex birds may result in a greater between-pen variation and thus less statistical power than the use of single-sex birds. The objective of this study was to evaluate the effect of including sex proportion as a covariate in an analysis of covariance (ANCOVA) on the statistical power compared to analysis of variance (ANOVA) where sex was not considered. The statistical parameters examined include mean square error (MSE), the F-statistic, model fit, model significance and observed power. A total of 4 separate experiments that used mixed-sex broilers with unequal numbers of male and female birds per pen were conducted during which performance of the birds was measured. The male % in each pen was recorded during each experiment and corrected for mortality. The performance results were analysed by ANOVA and the statistical parameters were then compared to ANCOVA where sex proportion was included as a covariate. The results showed that a set of assumptions first needed to be met to run ANCOVA. In addition, if the ANOVA results show a high level of model significance and power, then ANCOVA may not be necessary. In other circumstances where the assumptions are met and model significance and observed power are low, the inclusion of sex proportion as a covariate in the analysis will help to reduce MSE, increase the F-statistic value and improve the model significance, model fit and observed power. Therefore, it is suggested that sex proportion should be considered as a covariate in ANCOVA to improve statistical power in nutritional experiments when male and female broilers are unequally and randomly distributed in pens.
</p><p><b>Chapter 6 dataset</b><br />
It is well known that male and female broilers differ in their growth performance and that many physiological factors contribute to this difference. The aim of this experiment is to investigate if there are differences between males and female broilers in caecal microbiota and nutrient transporter gene expression and if these differences play a role in the growth performance of broilers. The possible effect of protein level and its interaction with sex on microbiota and expression of the nutrient transporters were also investigated. Samples were collected from male and female birds fed either standard crude protein (SCP) or reduced crude protein diets (RCP) at the age of d 35. The experiment was designed as a 2 × 2 factorial arrangement of treatments consisting of 448 Cobb 500 broilers assigned to 32 floor pens with 4 treatments, 8 replicates, and 14 birds per pen for performance measurements. The factors were sex (male or female) and dietary crude protein (CP) level (standard or reduced). Body weight gain, feed intake and feed conversion ratio were recorded for each pen. Sex had a significant effect on BWG and FCR (<i>P</i> < 0.001) where males had a significantly higher BWG and better FCR compared to females. There was a significant interaction between sex and protein level on FI (<i>P</i> < 0.05), where male birds had a higher FI compared to female birds only when the birds were fed SCP but not RCP diets. There was a significant interaction between CP level and sex on the expression of CAT2 (<i>P</i> = 0.02) and PEPT2 (<i>P</i> = 0.026) where the genes were significantly upregulated in females but only when the RCP diet was fed. The RCP diet upregulated the expression of B<sup>o</sup>AT (<i>P</i> = 0.03) as a main effect. Female birds had significantly higher expression of the PepT-2 gene compared to the males. The alpha diversity of caecal microbiota showed differences among the treatments. The Shannon diversity index was statistically higher (<i>P</i> = 0.036) for males fed the SCP diet and the Chao1 index for evenness was statistically higher (<i>P</i> = 0.027) in females fed the SCP diet. There was also a difference in the relative abundance of the 15 most common genera found in the caecal content of the broilers in this experiment and lastly the differential composition of microbiota between the different treatments was also significantly different.</p><p>
This study suggests that chickens are able to compensate for a reduction in AA substrates when fed a low CP diet through the upregulation of certain AA transporters, females may adapt to low CP diets better by such upregulation compared to males, and lastly, sex has an effect on the caecal microbial population and these differences contribute towards the performance differences between male and female broilers.</p>
description: <p><b>Chapter 4 dataset</b><br />
With known variation in performance between male and female broilers and the fact that sourcing single-sex birds for use in research is becoming increasingly difficult, it becomes important to determine the effect of rearing method with male and female broilers on between-pen variation and body weight (BW) uniformity. We evaluated the performance response of broilers reared as single or mixed-sex to standard and reduced crude protein (CP) diets. The study was designed as a 2 × 3 factorial arrangement of treatments consisting of 672 Cobb-500 broilers assigned to 48 floor pens with 6 treatments, 8 replicates, and 14 birds per pen. The factors were: rearing method (male single-sex, female single-sex or equally mixed-sex) and dietary CP level (standard or reduced). For the overall period of the trial (d 0-35) there was a significant effect (P < 0.001) of rearing method and CP level on feed intake (FI) and feed conversion ratio (FCR). There was also a significant interaction between rearing method and CP level for BWG during d 0-35 (P < 0.01).  There was a significant interaction between CP level and sex on d 34 BW (P < 0.01) where the reduced CP diet decreased the BW of both males and females, but to a greater extent the BW of the female birds. Dietary CP level had a significant effect on relative breast and drumstick weights with birds fed the reduced CP diet having significantly lower breast weights (P < 0.001) and higher drumstick weights (P < 0.01). This study suggests that male and female broilers have different CP requirements, and rearing birds as equally mixed-sex results in the lowest CV% for performance parameters and best BW uniformity compared to single-sex birds. Furthermore, when low CP diets are fed to broilers, they will prioritize the growth of more important body parts such as the legs.
</p><p><b>Chapter 5 dataset</b><br />
The availability of sexed day-old broiler chicks is becoming an issue as feather sexing is no longer possible. This has great implications for broiler researchers as the use of randomly distributed mixed-sex birds may result in a greater between-pen variation and thus less statistical power than the use of single-sex birds. The objective of this study was to evaluate the effect of including sex proportion as a covariate in an analysis of covariance (ANCOVA) on the statistical power compared to analysis of variance (ANOVA) where sex was not considered. The statistical parameters examined include mean square error (MSE), the F-statistic, model fit, model significance and observed power. A total of 4 separate experiments that used mixed-sex broilers with unequal numbers of male and female birds per pen were conducted during which performance of the birds was measured. The male % in each pen was recorded during each experiment and corrected for mortality. The performance results were analysed by ANOVA and the statistical parameters were then compared to ANCOVA where sex proportion was included as a covariate. The results showed that a set of assumptions first needed to be met to run ANCOVA. In addition, if the ANOVA results show a high level of model significance and power, then ANCOVA may not be necessary. In other circumstances where the assumptions are met and model significance and observed power are low, the inclusion of sex proportion as a covariate in the analysis will help to reduce MSE, increase the F-statistic value and improve the model significance, model fit and observed power. Therefore, it is suggested that sex proportion should be considered as a covariate in ANCOVA to improve statistical power in nutritional experiments when male and female broilers are unequally and randomly distributed in pens.
</p><p><b>Chapter 6 dataset</b><br />
It is well known that male and female broilers differ in their growth performance and that many physiological factors contribute to this difference. The aim of this experiment is to investigate if there are differences between males and female broilers in caecal microbiota and nutrient transporter gene expression and if these differences play a role in the growth performance of broilers. The possible effect of protein level and its interaction with sex on microbiota and expression of the nutrient transporters were also investigated. Samples were collected from male and female birds fed either standard crude protein (SCP) or reduced crude protein diets (RCP) at the age of d 35. The experiment was designed as a 2 × 2 factorial arrangement of treatments consisting of 448 Cobb 500 broilers assigned to 32 floor pens with 4 treatments, 8 replicates, and 14 birds per pen for performance measurements. The factors were sex (male or female) and dietary crude protein (CP) level (standard or reduced). Body weight gain, feed intake and feed conversion ratio were recorded for each pen. Sex had a significant effect on BWG and FCR (<i>P</i> < 0.001) where males had a significantly higher BWG and better FCR compared to females. There was a significant interaction between sex and protein level on FI (<i>P</i> < 0.05), where male birds had a higher FI compared to female birds only when the birds were fed SCP but not RCP diets. There was a significant interaction between CP level and sex on the expression of CAT2 (<i>P</i> = 0.02) and PEPT2 (<i>P</i> = 0.026) where the genes were significantly upregulated in females but only when the RCP diet was fed. The RCP diet upregulated the expression of B<sup>o</sup>AT (<i>P</i> = 0.03) as a main effect. Female birds had significantly higher expression of the PepT-2 gene compared to the males. The alpha diversity of caecal microbiota showed differences among the treatments. The Shannon diversity index was statistically higher (<i>P</i> = 0.036) for males fed the SCP diet and the Chao1 index for evenness was statistically higher (<i>P</i> = 0.027) in females fed the SCP diet. There was also a difference in the relative abundance of the 15 most common genera found in the caecal content of the broilers in this experiment and lastly the differential composition of microbiota between the different treatments was also significantly different.</p><p>
This study suggests that chickens are able to compensate for a reduction in AA substrates when fed a low CP diet through the upregulation of certain AA transporters, females may adapt to low CP diets better by such upregulation compared to males, and lastly, sex has an effect on the caecal microbial population and these differences contribute towards the performance differences between male and female broilers.</p>