Background: Tramadol abuse is a major health problem in Egypt. It promotes oxidative stress via lowering the antioxidant levels. Nigella sativa (NS) possesses strong antioxidant properties. Objectives: the aim of the study was to assess the testicular toxicity after oral administration of tramadol, testicular recovery after tramadol withdrawal and the potential ameliorative effect of NS on tramadol-induced testicular impairment. Material and Methods: Sixty-six adult male rats were allocated into 4 groups: control group (divided into 3 subgroups orally received normal saline, corn oil, and NS dissolved in corn oil respectively), tramadol-treated group (40mg/kg/day for 30 days), recovery group left to recover for 2 weeks after 30 days of tramadol administration, and NS-cotreated group administered NS (1ml/kg/day orally) along with tramadol. At the time of sacrifice, venous blood samples were obtained for assaying serum testosterone levels and the testes were processed for histological, morphometric and immunohistochemical examination. Results: Tramadol treatment resulted in deterioration of the testicular microarchitecture, intense expression of iNOS immunoreactivity, and statistically significant decreases in the tubular diameters, seminiferous epithelial heights, and serum testosterone levels. The recovery group revealed partial improvement of all the parameters tested. Coadministration of NS significantly improved the detrimental testicular effects of tramadol; this improvement was clearer than that resulted from tramadol withdrawal, proving the protective role of NS. Conclusion: NS has a significant role in protection against tramadol-induced testicular damage. Recommendations: Tramadol should be administered only when indicated with appropriate dose monitoring. NS supplementation is also advisable in patients on tramadol therapy.