DNA methylation marks, a key modification of imprinting, are erased in 
primordial germ cells and sex specifically re-established during gametogenesis. 
Abnormal epigenetic programming has been proposed as a possible mechanism 
compromising male fertility. We analysed by pyrosequencing the DNA methylation 
status of 47 CpGs located in differentially methylated regions (DMRs), the DMR0 
and DMR2 of the IGF2 gene and in the 3rd and 6th CTCF-binding sites of the H19 
DMR in human sperm from men with normal semen and patients with teratozoospermia 
(T) and/or oligo-astheno-teratozoospermia (OAT). All normal semen samples 
presented the expected high global methylation level for all CpGs analysed. In 
the teratozoospermia group, 11 of 19 patients presented a loss of methylation at 
variable CpG positions either in the IGF2 DMR2 or in both the IGF2 DMR2 and the 
6th CTCF of the H19 DMR. In the OAT group, 16 of 22 patients presented a severe 
loss of methylation of the 6th CTCF, closely correlated with sperm 
concentration. The methylation state of DMR0 and of the 3rd CTCF was never 
affected by the pathological status of sperm samples. This study demonstrates 
that epigenetic perturbations of the 6th CTCF site of the H19 DMR might be a 
relevant biomarker for quantitative defects of spermatogenesis in humans. 
Moreover, we defined a methylation threshold sustaining the classification of 
patients in two groups, unmethylated and methylated. Using this new 
classification of patients, the observed intrinsic imprinting defects of 
spermatozoa appear not to impair significantly the outcome of assisted 
reproductive technologies.