Breeders for the first parent generation were selected from REL breeder stock in July of 2018. After bleeding and genotyping, two P0 crosses were then created: one consisting of birds with all non-reference SNPs (compared to galGal6, ) for each gene region, designated P0-1, and the other consisting of birds with non-reference SNPs for CPQ and heterozygous SNPs for LRRTM4, designated P0-2. P0-1 consisted of 10 males and 13 females; P0-2 consisted of 12 males and 24 females. Separately, ungenotyped REL birds were used to breed generation 1 of the control population. Semen was collected from all the males of each P0 group, pooled, and used to artificially inseminate each of the females from the same P0 group.
After hatching, the F1 progeny produced from P0-1 and P0-2 were kept in floor pens and managed as breeders until they reached 18 weeks of age, at which point they were then bled and genotyped. Breeders for each F1 population were placed in individual breeder cages and photo stimulated. In May of 2019, breeders from the F1 were selected based on being homozygous for the non-reference SNPs for both genes; though all breeder SNP genotypes from this point forward were the same, the F1 populations were kept separate in order to complete reciprocal matings between them. Breeders from F1-1 (from P0-1) consisted of 12 males and 31 females; F1-2 (from P0-2) consisted of 12 males and 37 females. To produce the control group, 24 males and 72 females from the REL line were also utilized. Insemination began at the same time, when the MAS breeders were 22 weeks of age and the REL control breeders were 20 weeks of age. For the REL, pooled semen from all 24 males were used to artificially inseminate all 72 REL females. For the MAS, pooled semen from the F1-1 males was used to artificially inseminate the F1-2 females and similarly the F1-2 males were used to inseminate the F1-1 females. This reciprocal mating scheme produced the F2 generation of birds which would possess only the non-reference SNPs for the CPQ and LRRTM4 genes. Sets of eggs for hatching included at least 250 eggs each from REL and the F2 of the MAS. At transfer, all eggs were candled and infertile or eggs with embryonic mortality were removed and stored for breakout on hatch day along with eggs that did not hatch; no significant difference (P>0.05) in hatchery breakout was found between the two lines. After hatch, birds received a wing band that represented their genetic line (either MAS or REL). Sets of eggs were produced every two weeks for four total hatches. The first and fourth hatches were subjected to 5-week hypobaric challenges. The second and third hatches were placed for 8-week floor pen trials to evaluate change in production traits associated with MAS.
F2 chicks for challenge in the hypobaric (Hypo) chamber were placed on November 28, 2019 (Hypo1) and January 8, 2020 (Hypo2). Hypo1 was maintained at 9,000 ft simulated altitude while Hypo2 was initially set at 9,000 ft simulated altitude, then after 2 weeks increased to 11,000 ft simulated elevation to induce a higher incidence of ascites. In Hypo1, all hatched birds were placed in the chamber (n=578) whereas in Hypo2, similar numbers of birds were placed from each line and fewer total birds were placed (n=433) which reduced the number of birds that would need to be culled for compliance with bird density regulations.
The hypobaric challenge results indicate a sex- and elevation-dependent reduction in ascites incidence in both cohorts. Hypo1 saw an overall decrease (P=0.041) in ascites mortality between the MAS and REL birds, with a 27.3% reduction for ascites in MAS males and a 39.8% in MAS females [Table 2]. For Hypo2, there was an overall numerical, although not statistically significant (P=0.162), decrease in ascites mortality between the lines, with reductions of 23.4% in males (P=0.126) and only 5.2% reduction in females [Table 1]. There was no significant difference (P>0.05) between the lines for the right ventricle to total ventricle (RVTV) ratio or body weight. The Kaplan-Meier survival model curve visually echoes these trends, however the analyzed P-values from this model are only numerically different (P>0.05) [Figure 1].
Birds reared for the floor trials were placed on December 12, 2019 (Floor1) and December 28, 2019 (Floor2). On the day of placement, the total number of hatched birds was counted for each line and the number of birds placed per pen determined based on the smallest group; for Floor1 this was 14 birds per pen (0.133 m2/bird) and for Floor2 13 birds per pen (0.143 m2/bird) with all excess birds placed in spare pens for mortality replacement up until the beginning of the FCR measurement period from d 49 to d 55. As both floor trials were run concurrently in the same barn, though offset by two weeks, we considered whether the live performance and processing data should be analyzed as completely separate trials with separate analyses or together by adding the main effect of trial. After consultation with a professional agricultural statistician and colleagues at the University of Arkansas, the latter option was chosen. However, the interaction between line and trial was assessed for each measurement, and any traits identified as having an interaction between line and trial were noted and are discussed. Conclusions about the overall affect of MAS on that particular measurement were not drawn in those cases.
Live performance data from both cohorts is shown in Table 3. Significant differences were found between trials: d0 body weight (BW) (P<0.001), d42 BW (P=0.038), d0-42 body weight gain (BWG) (P=0.045), d49 BW (P=0.006), d0-49 BWG (P=0.007), d54 BW (P=0.017), and full-trial BWG (P=0.019). Additionally, the genetic lines were significantly different in d49-54 BWG (P=0.036) and FCR (P<0.001), both of which were improved in the MAS. Significant differences were found between trial and genetic line for d14 BW (P=0.019) and d0-14 BWG (P=0.015).
Total sample sizes and tabular representations of the data are as follows: Table 4, live weight and carcass characteristics (n=868); Table 5, deboned parts (n=868); Table 6, organ weights (n=295); Table 7, heart characteristics (n=295); Table 8, meat quality characteristics (n=395). A significant improvement was seen in the MAS birds for absolute and relative tender weights (P<0.001, P<0.001, respectively), relative drumstick weight (P=0.016), and significant differences were found for 24-hour L* color measurement (P<0.001), 24-hour a* color measurement (P=0.037), and 24-hour pH (P=0.003). Significant differences were found between sexes in relative hot carcass weight (P<0.001), wing weight (P<0.001), relative thigh weight (P<0.001), relative drumstick weight (P<0.001), absolute and relative heart weight (P<0.001, P=0.018, respectively), liver weight (P<0.001), lung weight (P<0.001), spleen weight (P<0.001), RV weight (P<0.001), TV weight (P<0.001), drip loss (P<0.001), 24-hour L* color measurement (P<0.001), 24-hour pH (P<0.001), and shearing peaks (P=0.016).
Significant differences were also found between the two floor trials for many characteristics, including wing weight (P<0.001), relative breast weight (P<0.001), relative thigh weight (P<0.001), relative drumstick weight (P=0.002), relative heart weight (P<0.001), liver weight (P=0.009), relative spleen weight (P=0.014), 24-hour a* color measurement (P=0.003), 24-hour b* color measurement (P=0.017), 24-hour pH (P<0.001), and shearing peaks (P=0.002). Due to this, there were also several significant interactions. Between trial and genetic line, significant differences were found for RV weight (P=0.046), RVTV (P=0.005), shear force requirement (P<0.001), and 1:3 shear area (P<0.001). While RV was found to be significantly different, Tukey’s HSD test was unable to separate the means. RVTV was found to be the largest in MAS birds from both trials and the smallest in REL birds from both trials. The largest shear force measurements were found in the Floor2 REL breasts, with intermediate force requirements in the Floor1 MAS group, and the lowest requirements in the Floor1 REL and Floor2 MAS groups. The largest 1:3 shear area measurements were found in the Floor2 REL group, with all other groups having comparable lower measurements.
A number of significant differences were found between trial and sex. These were live weight (P=0.035), hot carcass weight (P=0.028), fat pad weight (P=0.019), chilled carcass weight (P=0.036), absolute and relative tender weight (P=0.010, P=0.014, respectively), thigh weight (P=0.025), drumstick weight (P=0.032), relative liver weight (P=0.032), shear force requirement (P=0.010), and 1:3 shear area (P=0.040). The live weight and weights of hot carcass, chilled carcass, and tenders were the largest for the Floor1 males, moderate for Floor2 males, and smallest for females in both trials. The largest fat pads were found in the Floor1 males, moderate for Floor1 females, small intermediate for Floor2 females, and smallest in Floor2 males. Relative tender weight was found to be the greatest in Floor2 females, moderate in Floor1 females, and the lowest in males from both trials. The mean weights of thighs and drumsticks were separated into four distinct groups, from largest to smallest being Floor1 males, Floor2 males, Floor1 females, and Floor2 females. The relative weight of liver was found to be the largest in females from both trials and the smallest in males from both trials. For both the shear force requirement and the 1:3 shear area, the measurements from Floor1 females were found to be the largest, the Floor1 males were found to be the smallest, and both sexes in Floor2 were intermediate.
Finally, there were interactions found between genetic line and sex. These were live weight (P=0.011) hot carcass weight (P=0.012), chilled carcass weight (P=0.019), breast weight (P=0.050), thigh weight (P=0.006), drumstick weight (P=0.032), and 24-hour b* color measurement (P=0.025). For live weight and the weights of hot carcass, chilled carcass, thighs, and drumsticks, MAS males were found to be the largest, REL males were intermediate, and females from both the MAS and REL lines were found to be the smallest. Breast weight was the largest for males from both the MAS and REL lines, with the smallest weight in the females from both lines. The 24-hour b* color measurement was found to be the highest in MAS females, with the three other groups having comparable lower b* measurements.
There was also a single three-way interaction (P=0.039) found for the processing measurements, which was fat pad. In order from largest to smallest, the means were separated as follows: Floor1 MAS females, Floor1 REL females, Floor2 females from both lines, Floor1 REL males, Floor1 MAS males, Floor2 MAS males, and Floor2 REL males.