ORIGINAL ARTICLE A human morphologically normal spermatozoon may have noncondensed chromatin F. Boitrelle 1,2 , M. Pagnier 2 , Y. Athiel 2 , N. Swierkowski-Blanchard 1,2 , A. Torre 1,2 , L. Alter 1,2 , C. Muratorio 1 , F. Vialard 1,2 , M. Albert 1,2 & J. Selva 1,2 1 Department of Reproductive Biology and Cytogenetics, Poissy General Hospital, Poissy, France; 2 EA 2493, University of Saint-Quentin-en-Yvelines, France Keywords Aniline blue—chromatin condensation— human spermatozoon—sperm morphology Correspondence Florence Boitrelle, Department of Reproduc- tive Biology and Cytogenetics, Poissy General Hospital, F-78303 Poissy, France. Tel.: +33 139 275155; Fax: +33 139 274425; E-mail: florenceboitrelle@yahoo.fr Accepted: July 29, 2014 doi: 10.1111/and.12341 Summary According to numerous assisted reproductive medicine practitioners, semen with normal characteristics might not require further investigation. However, on the scale of the individual spermatozoon, it is well known that normal mor- phology does not guarantee optimal nuclear quality. Here, for 20 patients with normal sperm characteristics and a high proportion of spermatozoa with non- condensed chromatin, we subsequently assessed chromatin condensation status (aniline blue staining) and morphology (Papanicolaou staining) of the same 3749 spermatozoa. Although the overall proportion of morphologically normal spermatozoa was not correlated with the overall proportion of spermatozoa with noncondensed chromatin, an individual spermatozoon’s morphology appeared to be closely related to its chromatin condensation status. Morpho- logically normal spermatozoa with noncondensed chromatin were seen in all patients; the proportion averaged 23.3% [min 10.9%–max 44.4%]. Morpholog- ically abnormal spermatozoa were more likely to have noncondensed chromatin than morphologically normal ones (P < 0.0001). Small-, large- or multiple-headed spermatozoa presented the highest degree of noncondensation (>80% for each type), and more than half the vacuolated spermatozoa also presented noncondensed chromatin. However, a morphologically normal sper- matozoon may also have a noncondensed chromatin. Introduction The ‘World Health Organization Laboratory Manual for the Examination and Processing of Human Semen’ sug- gests that human male fertility can be predicted by the application of low cut-off values to the microscopic assessment of standard parameters (i.e. sperm count, motility, viability and morphology in particular) in native semen (WHO, 2010). Indeed, the cut-off values in the latest edition of the manual (particularly those concerning sperm morphology) are lower than in previous editions. For example, a semen with >4% of normal sperm forms (and otherwise normal features) is considered Depart- ment of Reproductive Biology and Cytogenetics, Poissy General Hospital, considered normal. However, many reports have shown that semen samples are heterogeneous and that the nuclear quality can vary greatly from one spermatozoon to another. Furthermore, normal morphol- ogy does not guarantee normal sperm nuclear quality, and a morphologically normal spermatozoon could present noncondensed chromatin, DNA damage and/or an abnormal chromosome content (Avenda~ no et al., 2010; Abu et al., 2012). In the present study, we focused on the chromatin con- densation status of individual spermatozoa. In correctly condensed sperm chromatin, 85% of the DNA is bound to protamines and only 15% is bound to histones (for a review, see Oliva & de Mateo, 2011). This histone–prot- amine replacement results from complex epigenetic events that also appear to influence embryo development after fertilisation (Jenkins & Carrell, 2012; Gannon et al., 2014). Indeed, it was recently reported that histones retained within the sperm nucleus are localised at particular chro- mosomal regions (e.g. the promoter regions of genes involved in embryo development), rather than being ran- domly distributed (Hammoud et al., 2011). Hence, the degree of sperm chromatin condensation could be a mar- ker of a spermatozoon’s potential for yielding an embryo © 2014 Blackwell Verlag GmbH 1 Andrologia 2014, xx, 1–8