Background:The study sought to determine whether hepatitis B infection in females and males plays a role in the outcome of the IVF/ICSI cycle. Methods: We performed a retrospective analysis of data collected from IVF/ICSI cycles carried out from January 2011 to December 2019 at the University Medical Centre Ljubljana. The data was analyzed using Pearson's chi-square test, Fisher's exact test, and Kruskal-Wallis test in order to determine the differences between the groups. Results: This study included 117 cycles with a past female hepatitis B infection (HF group), 91 cycles with couples with a past male hepatitis B infection (HM group), and 10,216 cycles with no past male or female hepatitis B infection (control group). There was no difference in sperm concentration, but total sperm motility was significantly higher in the HF and control groups compared to the HM group (p = 0.008 and p < 0.001). Also, sperm morphology was significantly higher in the control group, compared to both, the HM group (p < 0.001) and the HF group (p = 0.004). Furthermore, the rate of fertilized oocytes per number of retrieved oocytes was significantly higher in the HF (p < 0.001) and control groups (p = 0.003) compared to the HM group, but on the contrary, the rate of immature oocytes was lower in the HM (p = 0.009) and control groups (p = 0.001) compared to the HF group. The number of obtained embryos, blastocyst, and embryo utilization rates were similar between the groups, although the rate of cryopreserved embryos was higher in the HF (p = 0.007) and the HM groups (p = 0.027) compared to the control group. No significant difference was observed in the pregnancy and live birth rate per embryo transfer, while a trend towards a lower birth rate per aspiration was observed in the HM and in HF groups, which can be explained with a significantly higher miscarriage rate observed in the HM (p < 0.001) and HF groups (p = 0.042) compared to the control group. Gestational age was similar for all groups, although we observed a strong trend towards a higher birth weight of singletons in the HF group (p = 0.043) compared to the control group. Conclusions: The results indicate that hepatitis B infection could have a detrimental influence on sperm motility and morphology and cycle outcome, especially in terms of a higher miscarriage rate.