Objective: To determine if there is a link between prolonged COVID symptoms and the reactivation of EBV. Methods: This study was an observational (case-control) analysis involving 140 patients who tested positive for COVID-19 and are experiencing persistent symptoms such as fatigue and post-exertional malaise. Additionally, a control group of 80 individuals, matched for age and gender, who have fully recovered from SARS-CoV-2 infection without these symptoms, was included. The research took place between December 2023 and March 2024 at Benha University Hospitals in Benha, Egypt. The reactivation of the Epstein–Barr virus (EBV) was identified by detecting EBV genetic material using TaqMan probes, along with at least one set of primers (BamHI and LMP2). Results: Initial hospitalization during acute COVID-19 infection is significantly associated with post-COVID fatigue (p-value 0.007*). No significant associations were found for risk factors like diabetes and hypertension. EBV replication observed was due to EBV reactivation rather than primary infection. EBV-specific antibody titers EBNA-1 IgG (p-value 0.004*) and EA-D IgG (p-value 0.008*). 40/140 (28.6 %) patients with COVID-19 with persistent fatigue showed EBV reactivation in contrast to 9/80 (11.3 %) of controls (P-value 0.003*) using the same detection methods. Conclusion: EBV reactivation plays a role in Long-COVID syndrome following COVID-19 infection supporting the usage of EBV inhibitors for long-term COVID-19 treatment.