Environmental and Biological Determinants of Semen Quality in AI Bulls: Breed, Age, and THI Thresholds over 5 years

Abstract: Heat stress is a major environmental factor affecting bull fertility due to its impact on semen quality. However, the temperature-humidity index (THI) at which semen quality begins to decline under commercial artificial insemination (AI) centre conditions remains unclear. This study aimed to identify environmental and biological factors associated with sperm motility and to explore a THI threshold under commercial production conditions. Semen records were analysed from 20,067 ejaculates collected from 473 Dutch bulls representing seven breeds between January 2021 and September 2025. Due to commercial sensitivity, breed identities were masked and reported using breed codes. Semen quality was evaluated as the percentage of non-motile sperm (mean 12.9 ± 7.6%) and values were log transformed to stabilize variance. Daily mean THI was calculated on the day of semen collection using meteorological data published by Royal Netherlands Meteorological Institute. THI values ranged from 23.3 to 82.9 with most ejaculates (74.4%) collected under THI<60 and only 0.26% under THI≥80. Statistical models identified THI, bull age, and breed as significant predictors of non‑motile sperm (p<0.001). THI was first analysed as a continuous variable and a significant association with non-motile sperm was observed (p<0.001), corresponding to an estimated 1.5% relative increase in non-motile sperm per 10-unit increase in THI. When analysed categorically, THI<60 was used as the reference. Mean non‑motile sperm proportions were 12.31% for THI<60, 12.68% for THI 60-69, 12.35% for THI 70-79, and 13.14% for THI≥80. A significant increase was observed at THI 60-69 (p<0.001) compared to the reference, while higher THI categories showed numerically greater proportions, however statistically non‑significant (THI 70-79: p=0.830; THI≥80: p=0.183), likely due to the limited number of observations. Segmented regression analysis further indicated that non‑motile sperm increased when THI was higher than 60 to 62 (p<0.001). A weaker breakpoint appeared at THI 47.1 ± 7.4 (p=0.047). However, due to substantial variability in semen quality across THI classes, no single definitive heat-stress threshold could be identified. Breed and age were strong predictors of semen quality (p<0.01). Breed A showed the highest non‑motile sperm (~13.3%), and values increased with age (p<0.001), from 11.37% in younger bulls (14-20 months) to 14.10% at 40 months. Overall, bulls generally tolerate mild to moderate heat, but semen quality declines once THI exceeds approximately 60-62, with breed and age influencing resilience. These findings provide practical THI benchmarks to support heat‑stress management in AI centres and help maintain semen quality as environmental temperatures continue to rise.