Improved Training and Semen Collection Outcomes Using the Closed Box Chair for Macaques

Abstract

Collaborative semen collection in monkeys is a valuable tool in research, animal collection management, and conservation efforts. To obtain samples, monkeys are often restrained in open restraint chairs (ORC) with the “pole and collar” technique. While commonly used, this restraint is not tolerated by all individuals; some become anxious or aggressive towards the poles and people. In an effort to refine this procedure and improve welfare of the monkeys, we examined the use of a “closed box chair” (CBC), a clear, plexiglass box in which the monkey is trained to sit for sperm collection. The CBC does not require pole and collar, and although legs are secured, the arms and neck are not restrained. The use of CBCs has increased in recent years; however, there are few studies demonstrating its effects on scientific outcomes. We used positive reinforcement techniques to train 34 adult male rhesus macaques (Macaca mulatta) to provide semen samples using either the ORC or the CBC. While all CBC monkeys (n = 14) were reliably trained for this procedure, only 75% of ORC (n = 20) males completed the training (p = 0.04). It took significantly less time to train animals in the CBC than the ORC (201.0 vs. 412.4 min; p <0.001). In a controlled subset, males restrained with ORC (n = 7) produced a significantly lower ejaculatory volume than those collected by CBC (n = 10) (297.6 µL vs 522.1 µL respectively; p = 0.04) and had a lower concentration of sperm (186.0 × 106/mL vs 367.5 × 106/mL respectively; p = 0.017), although there were no differences with respect to sperm motility (p = 0.15). Our data suggest the closed box chair technique reduces stress on the animals while enhancing semen quality, supporting the use of the CBC as an important refinement.

Short Communication: Characteristics of the endangered Javan banteng (Bos javanicus) spermatozoa

“…the time was not reported (Das and Sarkar 2004). The habituation process reported by Houser et al (2021) showed mixed results among individuals from the same species regarding the time needed to progress from each stage. These findings imply the presence of the influence of species or individuals in their response to the habituation process.…”

“…This process is necessary, especially for wild animals that are unfamiliar with human encounters for the animals and human safety. Animal habituation for semen collection was reported in rhinos, camels, macaques, and even manatees (Agil et al 2008;El-Hassanein 2017;Cowart et al 2020;Houser et al 2021). The habituation time needed for semen collection in rhinos was for two months (Agil et al 2008), whereas, in the yak (Poephagus grunniens).…”

Rapid, accurate mapping of transgene integration in viable rhesus macaque embryos using enhanced-specificity tagmentation-assisted PCR

“…Rhesus macaque semen was collected from a male at the same day of oocyte collection and the sperm washed in warmed TALP-HEPES to a final concentration of 20 million sperm/mL. 32 A standard in vitro fertilization protocol was followed with sperm activated by 1 mM caffeine + 1 mM dibutyryl-cAMP for 15 min before adding 10 μL to each culture drops containing ova. 31 …”

Establishing the normal range of sperm DNA fragmentation index (% DFI) for rhesus macaques

“…All semen collections were performed by penile electroejaculation without sedation using a PTE 110 Volt AC electroejaculator (P-T Electronics, Model 303, Boring, OR) as previously described 13 . Ejaculate was collected directly into a wide mouth container and sat at 37 °C for 30 min to allow for liquefaction.…”

“…Samples were washed twice by centrifugation at 300×g for 7 min at 37 °C. After removal of the supernatant (11 mL), the sperm pellet was resuspended in the remaining 1 mL of TALP-HEPES and analyzed by a computer assisted sperm analysis (CASA) system (IVOS II-Animal Motility software, version 1.11, Hamilton Thorne, Beverly, MA) for sperm concentration and motility 13 . After analysis, aliquots containing 250,000 spermatozoa were placed in separate 0.6 ml centrifuge tubes and snap frozen in liquid nitrogen for storage in a − 80 °C freezer until analysis.…”