Sperm chromatin dispersion test (SCDt) for the assessment of sperm DNA fragmentation in black tiger prawn, Penaeus monodon

Abstract

The decapod sperm cell possesses no flagellum so that sperm evaluation has been primarily limited to the integrity of the plasma membrane and acrosome morphology. More recently, andrologists are turning to methods that are capable of assessing sperm DNA quality in an attempt to help understand infertility that cannot be explained by traditional sperm parameters. This study has successfully validated and applied a sperm chromatin dispersion test (SCDt) for the assessment of sperm DNA fragmentation of Penaeus monodon. After selecting an appropriate lysing solution, prawn sperm nuclear morphotypes were confirmed with respect to their DNA status using the two-tail comet assay. Once the SCDt methodology was established, we investigated and compared baseline levels of sperm DNA fragmentation (SDF) present in domesticated and wild male broodstock. Additionally, given that mature prawn sperm possess relatively uncondensed chromatin, the susceptibility of the nucleus to mechanical stress (vortex shearing forces) was investigated in an attempt to artificially induce sperm DNA damage. Results revealed that (1) the percentages of SDF indicated by the SCDt were strongly correlated (Pearson R2 = 0.989; P = .01) with those identified using the two-tailed comet assay, (2) mechanical stress associated with vortexing did not increase SDF level (P = .76) and (3) domesticated male broodstock had a higher (P < .001) SDF level (6.8 ± 4.5%) than that of the wild male broodstock (3.3 ± 1.5%). This study has provided a technique that allows for the assessment of sperm DNA integrity and with further development it could be incorporated as a potential index for male prawn fertility evaluation. We propose that the SCDt can be considered as one of the male fertility parameters in the routine management of male broodstock in prawn aquaculture.