Our study tested whether two European bird-specialized ticks, Ixodes arboricola and I. frontalis, can act as vectors in the transmission cycles of Borrelia burgdorferi s.l. The ticks have contrasting ecologies but share songbird hosts (such as the great tit, Parus major) with the generalist I. ricinus which may therefore act as a bridging vector. In the first phase of the experiment, we obtained Borrelia-infected ornithophilic nymphs by exposing larvae to great tits that had previously been exposed to I. ricinus nymphs carrying a community of genospecies (Borrelia garinii, valaisiana, afzelii, burgdorferi s.s., spielmanii). Skin samples showed that birds selectively amplified B. garinii and B. valaisiana. The spirochetes were transmitted to the ornithophilic ticks and survived moulting, leading to infection rates of 16% and 27% in nymphs of I. arboricola and I. frontalis respectively. In the second phase, pathogen-free great tits were exposed to the Borrelia-infected ornithophilic nymphs. None of these ticks were able to infect the birds, as indicated by the tissue samples. Analysis of xenodiagnostic I. ricinus larvae found no evidence for co-feeding or systemic transmission of B. burgdorferi s.l. These outcomes do not support the occurrence of enzootic cycles of Borrelia burgdorferi s.l. involving songbirds and their specialized ornithophilic ticks.