Descriptive analysis of sperm head morphometry in Iberian ibex (Capra pyrenaica): Optimum sampling procedure and staining methods using Sperm-Class Analyzer®

Sperm morphology has been identified as one characteristic which can be useful in prediction of fertility in a species. The development of computer automated sperm morphometry analysis allows for objective analysis of sperm head dimensions. The aim of the current study was to develop an optimum sampling procedure to characterize the Iberian ibex (Capra pyrenaica) sperm head morphometrically. Fresh semen from 11 males was collected using transrectal ultrasonic-guided massage of accessory sex glands and electroejaculation and prepared on slides for morphometric analysis to evaluate technical variation and standardize automated sperm morphometry analysis procedures by Sperm-Class Analyzer®. Three staining methods (Diff-Quik®, Hemacolor®, Spermblue®), number of sperm cells necessary to sample and repeatability of the staining technique were assessed. There were significant differences in size of sperm head depending on stain used. Hemacolor® was stain most suitable for sperm head morphometry evaluation (length = 8.42 μm; width = 4.21 μm; area = 29.37 μm2; perimeter = 21.93 μm; elongation = 0.33; elipticity = 2.01; regularity = 0.95; rugosity = 0.77). Morphometric values obtained from samples of 50, 100, 150, 175 and 200 sperm heads were compared. The most efficient method of analyzing sperm morphometry was to evaluate 100 sperm cells at 60× objective magnification. Thus, this study has allowed for description of optimal sample processing to determine morphometric parameters of sperm heads (size and shape) in Iberian ibex by Sperm-Class Analyzer® and provides a basis for future studies on the relationship with freezability and fertility in this species.