Crimean–Congo hemorrhagic fever virus (CCHFV) causes severe or fatal infections in humans and is geographically widespread. The virus has coevolved with its tick vectors, establishing persistent infections critical to its transmission. This study explored the mechanisms underpinning these persistent infections, using tick cell lines and the Hazara virus (HAZV) as a biosafety level 2 (BSL-2) model for CCHFV. Initially, an RT-qPCR protocol was developed to detect HAZV in tick cells. The study then focused on the production of virus-derived DNA (vDNAs) by tick cells as a defensive response to infection. These vDNAs regulate viral particle production, enabling tick cells to maintain viability and establish persistent infections. The experiments characterized vDNAs production, viral titers, and subcellular localization, and they examined the effect of the reverse transcriptase inhibitor azidothymidine triphosphate (AZT). The results showed that all tested tick cell lines supported HAZV replication, achieving persistent infections without cytopathic effects. vDNAs was detected in both the cytoplasm and nucleus, and its formation was dependent on HAZV infection. Importantly, vDNAs presence was linked to infection persistence; cells treated with AZT exhibited a marked reduction in vDNAs production and an associated increase in viral particle production, which correlated with higher cell death. These findings underscore the critical role of vDNAs in balancing viral replication and promoting long-term cell survival in tick cells, highlighting their importance in the coevolution of tick-borne viruses and their vectors.

Jingmen tick virus (JMTV) is a novel flavi-like virus first identified in 2010 in Rhipicephalus microplus in the Jingmen region of Hubei Province, China and has been reported in different Asian countries, Central and South America, Africa, and Europe. Beyond ticks, JMTV has been detected in a range of other arthropods and in vertebrate hosts. In humans, JMTV has been found in patients with Crimean-Congo hemorrhagic fever in Kosovo and Turkey, and in febrile patients with a history of tick bites in China, suggesting it may be a novel human pathogen. To investigate the presence of JMTV in Italy, we developed a One-step real-time RT-PCR assay and applied it to individually screen 1150 ticks collected from northeastern, central, and southern Italy. JMTV RNA was detected in multiple tick species, including Ixodes ricinus, Rhipicephalus bursa, Rhipicephalus sanguineus s.l., Dermacentor marginatus, and Hyalomma marginatum with a prevalence ranging from 0.52% to 18.42% in questing ticks. The detection of JMTV in ticks from all surveyed areas, indicates that the virus is geographically widespread in Italy. These findings highlight the need for comprehensive surveillance strategies to identify new areas of active virus circulation and to investigate the potential impact of JMTV on public health.

Crimean-Congo hemorrhagic fever virus (CCHFV) causes severe or fatal infections in humans and is geographically widespread. The virus has coevolved with its tick vectors, establishing persistent infections critical to its transmission. This study explores the mechanisms underpinning these persistent infections, using tick cell lines and Hazara virus (HAZV) as a biosafety level 2 (BSL-2) model for CCHFV. Initially, an RT-qPCR protocol was developed to detect HAZV in tick cells. The study then focused on the production of virus-derived DNA (vDNAs) by tick cells as a defensive response to infection. These vDNAs regulate viral particle production, enabling tick cells to maintain viability and establish persistent infections. Experiments characterized vDNAs production, viral titers, and subcellular localization, and examined the effect of the reverse transcriptase inhibitor azidothymidine triphosphate (AZT). Results showed that all tested tick cell lines supported HAZV replication, achieving persistent infections without cytopathic effects. vDNAs was detected in both the cytoplasm and nucleus, and its formation was dependent on HAZV infection. Importantly, vDNAs presence was linked to infection persistence; cells treated with AZT exhibited a marked reduction in vDNAs production and an associated increase in viral particle production, which correlated with higher cell death. These findings underscore the critical role of vDNAs in balancing viral replication and promoting long-term cell survival in tick cells, highlighting their importance in the coevolution of tick-borne viruses and their vectors.