The purpose of this study was to detect the effects of bacterial infection on human sperm nuclear protamines and DNA fragmentation. In this study, 120 semen samples were collected from unselected male partners of couples consulting for infertility in infertility and obstetrics clinic. All the samples were screened bacteriologically according to World Health Organization guidelines, and also sperm parameters and DNA fragmentation were evaluated. The concentrations of protamines P1 and P2 were quantified using acid urea acrylamide gel electrophoresis. Of a total number of 120 sample, 36 (30%) of them were infected with bacteria. Nine species of bacteria belonging to five genera, Staphylococcus, Escherichia, Streptococcus, Enterococcus and Klebsiella, were identified. The comparison between infected (36) and noninfected (84) samples appeared the negative impact of bacterial infection on sperm parameters and P1/P2 ratios. The percentages of P1/P2 ratio abnormality were significantly higher in infected patients. Sperm concentration, motility, progression and chromatin condensation were significantly lower in infected patients (p < .010). Depending on these results, we concluded that the bacterial infections have significant negative effects on sperm chromatin condensation and protamine P1/P2 ratio. Moreover, the negative relationship between the bacterial infections and sperm parameters, such as concentration, motility and progressive motility, has been shown.