Comparison of sperm subpopulation structures in first and second ejaculated semen from Japanese black bulls by a cluster analysis of sperm motility evaluated by a CASA system

Abstract

In the present study, bull sperm in the first and second ejaculates were divided into subpopulations based on their motility characteristics using a cluster analysis of data from computer-assisted sperm motility analysis (CASA). Semen samples were collected from 4 Japanese black bulls. Data from 9,228 motile sperm were classified into 4 clusters; 1) very rapid and progressively motile sperm, 2) rapid and circularly motile sperm with widely moving heads, 3) moderately motile sperm with heads moving frequently in a short length, and 4) poorly motile sperm. The percentage of cluster 1 varied between bulls. The first ejaculates had a higher proportion of cluster 2 and lower proportion of cluster 3 than the second ejaculates.

Effects of Media and Frame Rates on Hyperactivated Bovine Sperm Motility and an Analysis of Sperm Motility Subpopulation Structures in Sex-Sorted and Non-Sorted Semen

“…Therefore, sperm kinetic parameters other than VSL were affected by the frame rate. The number of bovine sperm incubated in BO for clustering in the present study (44,570 motile sperm) was markedly higher than in previous studies (16,,585 motile sperm) [24][25][26][27], and sperm in BO clearly exhibited hyperactivation-like characteristics; therefore, data on sperm captured at 150 fps were used as reference data for the discriminant analysis. The cluster analysis categorized motile sperm into 6 clusters in the present study.…”

“…Sperm motility was evaluated as described in a previous study [16]. Briefly, semen samples were introduced into a 20-µm-deep chamber (SC20-01-04-B, Leja, GN Nieuw-Vennep, Netherlands) preliminary warmed at 37°C on a hot plate (Kitazato Corporation, Shizuoka, Japan), and sperm motility was evaluated using a CASA system (SMAS, DITECT, Tokyo, Japan) based on the digitalized images obtained by a ×10 negative-phase contrast microscope (E200, Nikon, Tokyo, Japan).…”

“…A cluster analysis of sperm motility was performed as described in a previous study [16]. Briefly, sperm kinetic parameters were obtained at 150 fps by the CASA system, and VSL, VCL, VAP, ALH, and BCF were used as parameters in the cluster analysis.…”

Epidermal growth factor alleviates the negative impact of urea on frozen-thawed bovine sperm, but the subsequent developmental competence is compromised

“…After this step, sperm cells were washed using Tyrode's medium containing lactate, pyruvate, and HEPES (TL-HEPES) and subsequently used for in vitro fertilization (IVF). At this point, progressive sperm motility was subjectively assessed by a computer-assisted semen analyzer (CASA) visual estimation as described by others 59 . In brief, samples of semen were deposited in a 20-µm-deep chamber (SC20-01-04-B, Leja, GN Nieuw-Vennep, Netherlands) which was pre-heated to 37 °C on a hot plate.…”

Current perspectives of CASA applications in diverse mammalian spermatozoa

“…However, a well-grounded statistical model to determine the number of clusters (subpopulations) to be considered is not yet available for CASA data (Martínez-Pastor et al 2011). Most of these studies considered variations in the frequencies of determined motile subpopulations to assess differences among individual or repeated ejaculates (Vázquez et al 2015;Kanno et al 2017), possible fertility of ejaculates (Quintero-Moreno et al 2007;García-Á lvarez et al 2014) and the effect of short-term storage and cryopreservation on sperm quality (Martinez-Pastor et al 2005;Núñez-Martínez et al 2006;Dorado et al 2011).…”