Background/Objectives: Hepatitis B virus (HBV) remains a persistent challenge for transfusion safety. Although testing for hepatitis B surface antigen (HBsAg) and nucleic acid testing (NAT) reduces transmission risk, antibodies to hepatitis B core antigen (anti-HBc) and antibodies to hepatitis B surface antigen (anti-HBs) provide additional insight into past infection and vaccine-induced immunity. We aimed to determine their seroprevalence among blood donors in southern Croatia and assess associations with age, occupation, and time since vaccination. Methods: This cross-sectional study was conducted between February and November 2024 at two regional transfusion centers in southern Croatia. A total of 1008 voluntary blood donors, all HBsAg- and NAT-negative, were tested for anti-HBc and anti-HBs using chemiluminescent microparticle immunoassay. Demographic and vaccination data were collected through verified medical records. Results: Anti-HBc was detected in 0.5% of donors, exclusively among the unvaccinated. Protective anti-HBs levels were found in 38.1% overall and 70.6% of vaccinated donors, with significant declines by age and more than 15 years post-vaccination (p = 0.024). Healthcare workers showed higher seroprotection than non-healthcare donors (67.0% vs. 35.1%; p < 0.001), although one-third still lacked protective levels. Conclusions: HBV exposure was rare, but waning vaccine-induced immunity was evident, with protective anti-HBs levels in 70.6% of vaccinated donors, declining with age and time since vaccination. These findings highlight the need for periodic monitoring of anti-HBs and targeted booster strategies, especially in older and occupationally exposed groups. HBsAg and NAT provide a high level of transfusion safety, while the role of routine anti-HBc testing in this low-endemic context should be carefully evaluated in view of its potential benefits and drawbacks. Donor-based surveillance is a valuable tool for evaluating long-term vaccine effectiveness and guiding public health policy.

Introduction: Cytomegalovirus (CMV) seroprevalence varies globally. This study aimed to detect CMV seroprevalence and reactivation among patients who had been exposed to hepatitis B virus (HBV) or hepatitis C virus (HCV). Methodology: The study was conducted between 2017 and 2022 and included adult participants who were serologically diagnosed with hepatitis B (63 participants) or hepatitis C (69 participants), as well as 132 control respondents. CMV IgG and IgM levels were measured for all 264 respondents; CMV IgG avidity was further determined for those who tested positive for CMV IgG and IgM. Results: The total CMV IgG seroprevalence observed in the study was 95.4%, with 98.5% in HCV-positive, 96.8% in HBV-positive, and 95.2% in control respondents; and with a slightly higher prevalence in women (97.1%) compared to men (94.9%). The overall CMV reactivation rate among anti-HCV positive respondents was 2.9%. Although no significant difference was found in the reactivation rate of CMV between anti-HCV positive and negative respondents, the reactivation rate of CMV within the subgroup of HCV RNA positive individuals was higher at 4.4%. In contrast, no CMV reactivation was observed in the respondents without detectable HCV RNA. No CMV reactivation was detected in the HBV group. Conclusions: This study did not confirm a higher reactivation rate of CMV in HBV- or HCV-positive respondents compared to the control cases. Given the high CMV seroprevalence among adults in Bosnia and Herzegovina, future research should include CMV DNA testing for more accurate assessment.