Morphometric comparison by the ISAS^® CASA-DNAf system of two techniques for the evaluation of DNA fragmentation in human spermatozoa

Abstract

DNA fragmentation has been shown to be one of the causes of male infertility, particularly related to repeated abortions, and different methods have been developed to analyze it. In the present study, two commercial kits based on the SCD technique (Halosperm® and SDFA) were evaluated by the use of the DNA fragmentation module of the ISAS® v1 CASA system. Seven semen samples from volunteers were analyzed. To compare the results between techniques, the Kruskal–Wallis test was used. Data were used for calculation of Principal Components (two PCs were obtained), and subsequent subpopulations were identified using the Halo, Halo/Core Ratio, and PC data. Results from both kits were significantly different (P < 0.001). In each case, four subpopulations were obtained, independently of the classification method used. The distribution of subpopulations differed depending on the kit used. From the PC data, a discriminant analysis matrix was obtained and a good a posteriori classification was obtained (97.1% for Halosperm and 96.6% for SDFA). The present results are the first approach on morphometric evaluation of DNA fragmentation from the SCD technique. This approach could be used for the future definition of a classification matrix surpassing the current subjective evaluation of this important sperm factor.

Afterword to Sperm morphometrics today and tomorrow special issue in Asian Journal of Andrology

“…By integrating the original variables according to their coherence in a database into a new complex mathematical variable, clearly defined homogeneous subpopulations of spermatozoa can be defined. In support of the theory above, the papers presented here showed that most of the variance from up to 13 morphometric variables could be explained by only two or three PCs: two in bulls,16 adolescent humans,8 adult human sperm head DNA,9 domestic cats,10 puma,11 roosters, and guinea fowls12 and three PCs in adult human split ejaculate samples 7…”

“…Here, the number of subpopulations ranged from two to five: two (for the X-/Y-bearing bovine sperm heads,15 for large+elongated/small+elongated sperm heads in human adolescents8), three (for large+round/elongated/small spermatozoa in human sperm heads in split ejaculate fractions,7 for elongated+intermediate/large+high acrosome/short+small sperm heads in the puma,11 for small, wide and slightly elliptical/average size, long, narrow and very elliptical/very large, wide and elliptical sperm heads in the rooster12), four (for large/high medium/low medium/small in human sperm head DNA,9 for small/short/large/narrow sperm heads in the bull,14 for shape-related sperm heads in both normo- and terato-zoospermic cats10) to five (for very small, wide, very short and slightly elliptical/small, very short, very wide and slightly elliptical/very large, very wide, short and slightly elliptical/average size, very long, very narrow and very elliptical/average size, long, narrow and elliptical sperm heads in the guinea fowl12). …”

“…For example, if the changes in sperm populations upon maturation in the cat epididymis10 are indicative of the epididymal maturation process, aberrant populations in the ejaculate could be indicative of epididymal malfunction in endangered feline species; the rapid and automated assessment of human sperm DNA damage9 could be useful in optimizing selection methods that enrich populations in the less damaged cells required for ART; likewise, it would be interesting to find out whether differences in bovine sperm subpopulations between bulls, ejaculates and thawed straws16 are present in the native semen or introduced by the cryopreservation protocol, and to use these subpopulations to monitor the development of methods to select the sperm subpopulation (of elongated and tapered spermatozoa) previously associated with fertility for AI.…”

“…This Special Issue on computer-aided sperm morphology assessment comprises four reviews (on sperm transport in mammals,3 the current status of sperm morphometry in mammals4 and birds,5 and the relevant statistical methods to assess the morphometric results6); three clinical research papers (on sperm subpopulations in split ejaculates from adult men with normozoospermia7 and in ejaculates from adolescents with or without varicocoele,8 including the use of computer-assisted sperm analysis in assessing sperm nuclear DNA fragmentation9); and six veterinary research papers on sperm populations in the epididymis (in normozoospermic and teratozoospermic domestic cats10) and in ejaculated spermatozoa (from an endangered puma species,11 roosters and guinea fowls,12 rams, bulls and boars,13 and on cryopreserved bovine semen14). …”

Effect of different oxidative stress degrees generated by hydrogen peroxide on motility and DNA fragmentation of zebrafish (Danio rerio) spermatozoa

“…Several works that used SCDT to determine SDF have classified four levels of DNA damage according to halo size (absence, small, medium, and large), but not halo area [Human: (Absalan & Ghannadi, ; Tandara et al, ); Pig: (Alkmin et al, ); Llama: (Carretero, Lombardo, Arraztoa, Giuliano, & Gambarotta, )]. In the recent past, Sadeghi et al () evaluated halo areas with two SCD techniques (Halosperm ® vs. SDFA) in human sperm using the DNA fragmentation module of the ISAS ® . Sebastian and Raghavan () found significant differences in the halo area in spermatozoa treated with endosulfan.…”