The use of antioxidant supplements in broodstock diets proved to be beneficial for the reproductive performance of some marine fish. Protection against oxidative stress is important during spermatogenesis, because of the high levels of reactive oxygen species (ROS) inherent to this extremely active replicative process. Spermatogonia and spermatocytes have a natural mechanism of protection against ROS associated to enzymes and cytoplasmic molecules that act as scavengers. However, this protection diminishes in spermatozoa, in which the cytoplasmic content is reduced. As a biological response to oxidative stress and to protect spermatozoa, seminal plasma can be a powerful source of antioxidants, where diet composition has a pivotal role. In fact, not only seminal or plasma membrane composition may be modulated through the incorporation of certain compounds in the feeds, such as trace elements or vitamins, but also other sperm traits can be affected. Studies have been conducted to search for novel feed formulations with incorporation of micro and macroalgae or directly by the increase of compounds such as zinc, selenium and vitamin C and E, known by their antioxidant properties. Having algae a diversity of bioactive compounds with considerable antioxidant capacity, they are clearly good candidates. However, it is not always clear the effect of different antioxidant sources and most of the research conducted had positive and negative effects depending on the fish species.
In the present study gilthead seabream (Sparus aurata), European seabass (Dicentrarchus labrax) and Senegalese sole (Solea senegalensis) breeder diets were supplemented with macroalgae (Plocamium or Sargassum), as natural antioxidants sources, or directly by administrating zinc, selenium and vitamins C and E into the feeds. Experiments were conducted during gametogenesis, spermiation and post-spermiation in some species to identify putative effects on gonadal antioxidant system and sperm traits. Sperm samples were collected and several quality tests were determined: spermatozoa motility, viability, ROS, DNA fragmentation and lipid peroxidation. Expression of apoptotic, anti-apoptotic, antioxidant genes and enzymes as well as total antioxidant capacity (TAS) were determined in reproductive tissues and cells.
Results demonstrated a clear effect of antioxidants in sperm traits, very dependent on the species and antioxidant used. In Senegalese sole the supplementation of 5% Plocamium cartilagineum increased spermatozoa motility and viability when compared with control or with Sargassum vulgare, although in terms of cells protection against ROS, Sargassum supplementation proved to be more efficient, probably due to a high phenolic content present in this algae. In seabream and seabass, there was higher total antioxidant activity during gametogenesis than spermiation, with an increase of TAS in fish fed on vitamin supplementation. In seminal plasma, higher levels of vitamins and selenium were detected when compared with control. Sperm GSR and GPx activity was positively affected by treatments. In seabream, bax expression (pro-apoptosis) was downregulated in Se and Zn treatments, where bclx (anti-apoptosis) was upregulated. Funding: Projects SPERMANTIOX and GERMROS (PTDC/CVT-CVT/4109/2020 and EXPL/CVT/0305/2021, FCT), BREEDFLAT (PT-INNOVATION-0080-EEA grants), and CCMAR and LA Strategic Projects (UIDB/04326/2020, LA/P/0101/2020).