100 The evolution and premature hyperactivation of kinetic sperm subpopulations are affected by inbreeding level in Retinta cattle
Authors: Teran E. , Molina A. , Rodriguez Z. P. , Dorado J. , Peyrás S. D. 
Published: January 2021

Computer-assisted sperm analysis (CASA) has become a powerful tool to study sperm heterogeneity because it allows to cluster individual sperm in subpopulations (Sp) according to their kinetic parameters, which are associated with fertility and sperm quality. In cattle, inbreeding (mating between close relatives) affects sperm quality. Inbreeding estimation has become more accurate with the increasing availability of genomic methodologies, such as the detection of runs of homozygosity (ROH). Additionally, genomic inbreeding values (FROH) allow us to determine which metabolic pathways are differentially affected by this genetic trait. The aim of this study was to assess the effect of inbreeding on evolution of sperm Sp over time in cattle. Sperm samples (n=100) from 50 Retinta bulls (two replicates) were analysed in a sperm longevity experiment at time 0 (T0, after thawing), 1 (T1) and 2 (T2) h. At each time point, eight parameters were measured using a CASA system (Sperm Class Analyzer 5.4), including curvilinear velocity (VCL, μm/s), straight-line velocity (VSL, μm/s); average path velocity (VAP, μm/s), percentage of linearity (LIN,%: VSL/VCL), percentage of straightness (STR,%: VSL/VAP), wobble coefficient (WOB,%: VAP/VCL), mean amplitude of lateral head displacement (ALH, μm) and beat-cross frequency (BCF, Hz). The presence of Sp was determined by a two-step multivariate analysis including non-hierarchical followed by hierarchical analysis in 80,154 motile sperm. Four sperm Sp were identified: Sp1 (rapid and highly progressive sperm); Sp2 (progressive sperm with intermediate speed); Sp3 (slow and non-progressive sperm); and Sp4 (fastest, hyperactive-like, non-progressive sperm). To determine the effect of inbreeding, individuals were clustered into lowly (FROH &lt; 0.125, n=27) and highly (FROH &gt; 0.125, n=23) inbred individuals. After thawing (T0), the percentage of sperm Sp4 was higher and showed premature hyperactivation in highly inbred animals, which was previously associated with reduced fertility (Table 1). However, highly inbred individuals showed an increased percentage of rapid (Sp1) and intermediate (Sp2) progressive sperm after T1 and T2 incubation, respectively, compared with lowly inbred bulls (P&lt;0.001). These results suggest that sperm velocity and progression persist for longer in highly inbred bulls, at least after a short incubation. In conclusion, our results demonstrate that inbreeding affects sperm motility pattern and premature hyperactivation in cattle.

Table 1.
Percentage of subpopulations (Sp) by inbreeding group and time

Time
Sp1
Sp2
Sp3
Sp4

Highly inbred
Lowly inbred
Highly inbred
Lowly inbred
Highly inbred
Lowly inbred
Highly inbred
Lowly inbred

0
33.17a
34a
32.39a
32.56a
28.07a
27.93a
6.37b
5.51a

1
31.73b
26.13a
36.75a
41.65b
23.80a
24.37a
7.72a
7.85a

2
11.73b
7.67a
53.75b
48.78a
29.68a
39.27b
4.83a
4.28a

a,bDifferent superscripts by subpopulation in the same row denote significant difference (P&lt;0.05).