Epstein–Barr virus (EBV) and Kaposi’s sarcoma-associated herpesvirus (KSHV) are oncogenic gammaherpesviruses with biphasic lifecycles. Both can reactivate from latency in response to various stimuli. In sub-Saharan Africa, EBV and KSHV prevalence is high, often obscured by co-infections such as HIV, tuberculosis, and, more recently, SARS-CoV-2.

SARS-CoV-2, the causative agent of COVID-19, has caused substantial morbidity in SA. Its dysregulated immune responses, including cytokine storms, may trigger the reactivation of latent viruses. Both EBV and KSHV have been reported to reactivate in severe COVID-19 cases, exacerbating outcomes.

Given their reactivation potential in immunocompromised individuals, we investigated EBV/KSHV dynamics in 407 non-hospitalized people living with HIV during the COVID-19 pandemic. To further study the viruses’ interaction, a tissue-culture-based system was established using EBV latently infected cell lines (Raji, Namalwa, IARC-171, AGS-EBV) and EBV-KSHV dually infected BC-1 cells. Cultures were then exposed to chemical stimulants, SARS-CoV-2 pseudovirus and a COVID-19 “cytokine cocktail”. Reactivation was quantified via real-time qPCR.

Clinically, EBV viral load (VL) was detectable in 97.0% of patients, with 12.4% exhibiting elevated VL (≥1×10^5 copies/10^6 cells). In contrast, KSHV VL was detectable in only 21% of patients. SARS-CoV-2 exposure and COVID-19 vaccination did not significantly influence EBV reactivation; however, KSHV reactivation increased post-SARS-CoV-2 exposure in unvaccinated individuals. Logistic regression confirmed higher odds of KSHV reactivation in previously SARS-CoV-2-exposed patients and in patients with elevated EBV VL.

In vitro, SARS-CoV-2 infection did not induce EBV or KSHV reactivation in the tested cell lines. These findings warrant further investigation into whether the SARS-CoV-2-induced cytokine storm contributes to EBV or KSHV reactivation.

Our in vitro studies support the clinical observations that EBV and KSHV reactivation is unlikely to be driven by SARS-CoV-2 itself, but rather results from the dysregulated inflammatory response. This may have long-term percussions for gammaherpesvirus-associated tumorigenesis outlasting the pandemic.