The cryopreservation potential and ultrastructure of Agulhas sole Austroglossus pectoralis spermatozoa

by Markovina, M. Z.

Abstract (Summary)
As the estimated market demand for the Agulhas sole Austroglossus pectoralis exceeds the annual catch from trawlers, this species is a potential aquaculture candidate. Broodstock conditioning and gamete preservation is part of research and development aiming at establishing a breeding protocol for a new aquaculture species. Based on a literature review of the morphology of pleuronectiform spermatozoa, this study was designed firstly, to contribute to the field of spermatozoan morphology by describing the ultrastructure of A. pectoralis spermatozoa. This was followed by an experiment to cryopreserve mature spermatozoa to provide baseline data for future studies on this and related species. The testis of A. pectoralis was a paired structure encased in a membrane, the tunica albuginea. The primary testis was located on the dorsal surface of the rib cage and the secondary testis on the ventral side. The testis was of an unrestricted spermatogonial type, based upon observations of spermatogonia along the entire length of the lobule. Mature spermatozoa of A. pectoralis had an acrosome-free ovoid head 1.68 ± 1.6?m in length and 1.7 ± 1.6?m in diameter, a short mid-piece of 0.5 ± 0.1?m in length, containing 7 irregularly shaped mitochondria forming a ring-like structure at the base of the nucleus. The flagellae were 47.4 ± 4.8?m in length, most with two plasma membrane lateral fin-like projections. However, some flagellae had either zero or three lateral fin projections. Cross-sections of the flagellae showed an axenome with a 9+2 microtubule configuration. The proximal and distal centriols were coaxal, situated deep within the nuclear fossa. The structure of A. pectoralis spermatozoa conformed to the type 1 ect-aquasperm, also found in externally fertilizing species. This type has been suggested to be the plesiomorphic form in Neopterigians. Finally, this study contributed to a cryopreservation protocol for A. pectoralis spermatozoa by testing the two cryoprotectants dimethyl sulphoxide (DMSO) and glycerol. Glycerol, at a concentration of 10%, offered better cryoprotection than DMSO. This was established using flow cytometry analysis of post-thaw nuclear membrane integrity after 64 days of storage in liquid nitrogen. The toxicity of DMSO to isolated cellular proteins may have resulted in DMSO-treated sperm having the highest percent (35.2% ± 3.2%) of non-viable cells compared with 23.0% ± 2.5% and 27.8% ± 3.4% for glycerol and the control, respectively. The presence of sucrose in the Modified Mounib Medium xtender solution may explain why 45.5% ± 5% of the sperm cells were potentially viable in the control treatment. Initially, the white margined sole Dagatichthys marginatus (Soleidae) was selected as the most suitable candidate for flatfish aquaculture in South Africa. Thus, the aim of this study was to investigate the cryogenic potential and ultrastructure of D. marginatus spermatozoa. However, due to a skewed sex ratio, there were not enough males available to study this species. A skewed sex ratio is common amongst soleids, thus, the need to develop effective cryopreservation methods and to develop an understanding of sperm morphology so that the best time for cryopreservation can be chosen. In conclusion, this first description of spermatozan morphology of A. pectoralis contributed to our understanding of soleid sperm ultrastructure. In addition, a comparison of testis appearance between fish sampled just prior to spawning season and fish with mature sperm provided information on the spawning season of this species. The findings from the cryopreservation experiment suggested that glycerol was a feasible cryoprotectant for this species when sperm was prepared under field conditions.
Bibliographical Information:


School:Rhodes University

School Location:South Africa

Source Type:Master's Thesis

Keywords:ichthyology fisheries science


Date of Publication:01/01/2008

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