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. 2015 Spring;16(2):161-6.

Analysis of chromatin integrity and DNA damage of buffalo spermatozoa

K Gh M Mahmoud  1 , A A E El-Sokary  2 , A E Abdel-Ghaffar  3 , M E A Abou El-Roos  3 , Y F Ahmed  1
Affiliations

Analysis of chromatin integrity and DNA damage of buffalo spermatozoa

K Gh M Mahmoud et al. Iran J Vet Res. 2015 Spring.

Abstract

This study was conducted to determine chromatin integrity and DNA damage by DNA electrophoresis and comet assays of buffalo fresh and frozen semen. Semen samples were collected from four buffalo bulls and evaluated after freezing for semen motility, viability, sperm abnormalities, chromatin integrity and DNA damage. A significant variation was found in semen parameters after thawing. Highly significant differences (P<0.001) in chromatin integrity were observed between fresh and frozen semen. For the fresh semen, there was no significant difference between the bulls for chromatin integrity; however, a significant variation (P<0.05) was detected in their frozen semen. No DNA fragmentation was observed by agarose gel electrophoresis. The percentage of sperm with damaged DNA detected by comet assay differed significantly between fresh and frozen semen. A significant negative correlation was recorded between motility and DNA damage (r=-0.68, P<0.05). Sperm abnormalities and DNA fragmentation were significantly positively correlated (r=0.59, P<0.05). In conclusion, DNA damage evaluation can provide reassurance about genomic normalcy and guide the development of improved methods of selecting spermatozoa with intact DNA to be used in artificial insemination.

Keywords: Buffalo bull; Chromatin integrity; DNA damage; Semen quality.

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Figures

Fig. 1
Fig. 1
Fluorescence microscopy displaying buffalo spermatozoa with damaged chromatin as shown by yellow or red colours of the acridine orange stain, and normal chromatin with green colour (×400)
Fig. 2
Fig. 2
DNA profile of frozen semen after electrophoresis. M is λ DNA hind III molecular weight marker (23130, 9416, 6557, 4361, 2322, 2027 and 564 bp). 1, 2, 3 and 4 represent frozen semen samples of four buffalo bulls
Fig. 3
Fig. 3
Fluorescence photomicrograph of buffalo spermatozoa (×400) following neutral microgel electrophoresis showing different forms of fragmentation (A and B). Spermatozoa with non-fragmented DNA do not have a comet (without arrows) and spermatozoa with fragmented DNA exhibit the characteristic formation of comet (arrows
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