Molecular screening for tick-borne encephalitis virus in urban tick populations

Climate change is reshaping tick ecology by prolonging their seasonal activity and enabling expansion into previously unsuitable habitats [1]. Consequently, the incidence of tick-borne diseases is increasing in regions once considered low risk, creating growing public health challenges. Tick-borne encephalitis (TBE) remains one of the most important vector-borne diseases in Europe, with the Baltic countries accounting for nearly one-third of reported cases [2, 3]. This study investigated the prevalence of tick-borne encephalitis virus (TBEV) in questing ticks in Lithuania and evaluated key transmission risk factors. During 2023–2024, 544 ticks (Ixodes ricinus, n = 525; Dermacentor reticulatus, n = 19) were collected by flagging method in urban habitats and pooled into 47 groups by species and developmental stage. TBEV detection was carried out using real-time RT-PCR and virus isolation in two cell lines (Neuro-2a and MARC-145). TBEV RNA was detected in 15.26% (83/544) of individual tick samples. Virus isolation proved more sensitive than direct RT-PCR, yielding a total minimum infection rate (MIR) of 2.8% across both cell lines, compared with 1.47% by direct RT-PCR. MIR was 1.3% (7/525) for I. ricinus and 5.3% (1/19) for D. reticulatus. Positive samples included two nymphs, three females, and three males, indicating active TBEV circulation across developmental stages and sexes. These findings highlight a potentially greater role of D. reticulatus in TBEV transmission than previously recognised and emphasise the need for continued tick surveillance, robust public health interventions, and the inclusion of virus isolation techniques to enhance detection accuracy.