Viability Assessment of Spermatozoa in Large Falcons (falco Spp.) Using Various Staining Protocols

Viability assessment is an important part of semen analysis and various live/dead staining protocols have been used in semen of avian species. Results of live/dead count differed between dyes, staining protocols and bird species, impeding comparability between studies and requiring species specific comparisons of viability stains. In raptor semen similar comparisons are absent. Thus, the aim of the present study was to compare eight conventional viability stains. Eosin blue 2 % [EB], eosin blue 2 % with the addition of 3 % sodium citrate [EB2], eosin blue nigrosin 5 % [EBN5], eosin yellow nigrosin 5 % [EYN5], eosin yellow nigrosin 10 % [EYN10], eosin blue aniline blue [EBA], eosin yellow aniline blue [EYA], and bromphenol blue nigrosin [BBN] were evaluated in comparison to the fluorescence stain SYBR® Green - propidium iodide [SYBR-PI] in spermatozoa of falcons. The comparison was performed using conventional light microscopy which is applicable in breeding centers, veterinary practices and field studies. Additionally, live/dead stains were correlated to motility values of the same samples to validate sperm viability. Light microscopy using EB and using SYBR-PI enabled an effective and clear differentiation between alive and dead spermatozoa of falcons. Motility values correlated significantly and strong with EB only (r = 0.629; p < 0.001), but not with any other stain used in the study. Therefore, our results suggest EB as the most suitable stain for viability assessment in the semen of large falcons.