Theriogenology Wild (Jan 2025)
Comparison of staining protocols to assess sperm viability in cockatiels (Nymphycus hollandicus)
Abstract
Abstract/Summary: The assessment of sperm viability belongs to the core elements of semen analysis in human and veterinary andrology. In avian medicine various live/dead stains and protocols have been evaluated in a number of commercial poultry and companion bird species. However, viability results are known to differ largely between avian species and the used stains and protocols. Therefore, species specific comparisons of live/dead stains are required, which are absent in numerous bird orders. The aim of this study was to compare the conventional live/dead stains eosin blue 2% (EB), eosin yellow-nigrosin 10% (EYN), eosin blue-nigrosin 5% (EBN), bromphenol blue nigrosin (BBN) and the fluorescence stain SYBR green - propidium iodide (SYBR-PI) in spermatozoa of cockatiels as model for psittacine species. The latter comparison was performed conventionally via cell count as well as using a computer-assisted semen analyzer (CASA). Moreover, all viability stains were correlated to sperm motility values of the same sample, as this has been suggested as suitable method to validate sperm viability. Pairwise comparison between the motility values and the viability values of EB, EYN, EBN, BBN, SYBR-PI were performed immediately, 24 hours and 48 hours after semen collection. Progressive motility values (PMOT) correlated significantly (p < 0.05) with all viability results immediately after semen collection, but PMOT after 24 hours and general motility values only with EB and SYBR/PI. In microscopy EB and SYBR were judged as best suited, because alive and damaged spermatozoa could be effectively distinguished. Conventional and computer-assisted analysis using SYBR/PI demonstrated significant and strong correlations (rs = 0.939; p < 0.0001). The results of this study, suggest EB and SYBR/PI as suitable stains for viability assessment in cockatiels, while EYN and EBN is not recommendable and BBN seems not suitable at all in psittacine spermatozoa.
