Cryopreservation of Danube barbel Barbus balcanicus sperm and its effects on sperm subpopulation structure

Abstract

The objective of this study was to develop a successful freezing protocol for cryopreservation of Danube barbel sperm, and to identify the presence of different spermatozoa subpopulations. By testing different concentrations of different cryoprotectants, we determined that the use of 5% dimethyl sulfoxide (DMSO) yielded the highest total motility of ~25%. Cooling rates influenced by frame height and cooling time in liquid nitrogen vapor showed that a frame height of 3 cm and cooling time of 2 min yielded the highest post-thaw motility. Supplementation of cryomedia with 0.1 M of sugars led to an increase in the total post-thaw motility by ~50%, while protein supplementation lowered post-thaw motility. Motile spermatozoa hierarchically clustered according to their motility parameters, displaying a four-subpopulation (SP1-SP4) structure. SP1 was defined by low values of velocity but high overall linearity; SP2 was comprised of fast non-linear spermatozoa, that had high velocity values but low linearity; SP3 was characterized by fast linear spermatozoa, and SP4 by slow non-linear spermatozoa. Protocols developed in this study will lead to the creation of new and enhanced conservation strategies for this species.

Sperm Quality and Cryopreservation in Teleost: Effect of Seminal Plasma Component and Climate Change

“…For other fish species such as Barbus balcacinus and Solea senegalensis, four sperm subpopulations have been described (Kojadinović et al, 2020;Martínez-Pastor et al, 2008). Although the multivariate analysis method used to identify Barbus balcacinus subpopulations did not follow the same two-step clustering procedure, this method is the most widely used for fish.…”