Tick-borne Encephalitis (TBE) is an important zoonosis in many parts of eastern and central Europe. The most efficient transmission route for tick-borne encephalitis virus (TBEv) seems to be the saliva-activated non-viraemic transmission between co-feeding Ixodes ricinus ticks taking place on rodent hosts. During the period 2001-2014, a population of yellow-necked mouse, Apodemus flavicollis, was intensively live-trapped in a known TBE focus in Trentino (northern Italy). Ticks feeding on individual rodents, distinguishing between larval and nymphal life-stages, and counts of co-feeding groups on rodents, were recorded. In addition, serological test for the detection of antibodies to TBEv was carried out on trapped rodents. First, a combined effect of climatic conditions and rodent density on the number of co-feeding groups was observed. Specifically, co-feeding occurrence on rodents during the season was affected by autumnal cooling of the previous season. On the other hand, co-feeding occurrence was associated with rodent density during the sampling season showing a humped (concave-down) relationship. Individual features of rodents such as weight and gender also affected co-feeding occurrence with heaviest males carrying more co-feeding ticks. In addition, we found that the overall number of co-feeding ticks on rodents positively affected TBEv infection in rodents the following season. This is due to a higher probability of larvae to acquire TBEv infection during the season through co-feeding transmission; these larvae would moult and feed on rodents as infected nymphs the following season, thus increasing TBEv circulation and transmission risk. In conclusion, specific climatic conditions in conjunction with suitable rodent density are the principal drivers of co-feeding ticks on rodents and they can be used to provide a hazard early warning system for TBE. This potentially allows targeted public health actions, with implications for prevention and control within TBEv circulation areas.