Roads fragment vertebrate populations and affect the dynamics and dispersal patterns of vertebrate parasites. We evaluated how vertebrate ticks distribute near roads and road underpasses in human-caused road-fragmented landscapes in Seville, SW Spain. We sampled 49 stations with 93 individualized sampling points and assessed tick abundance. We explored the relationship between tick presence and abundance and distance to the nearest road and underpass (drainage culverts and other passages used by vertebrates), and landscape features through categorical regression and nonparametric statistics. The presence of the tick-borne pathogens Borrelia sp. and Bartonella sp. was also analyzed by PCR. We found preliminary evidence of high relative tick abundance next to roads and in the vicinity of road underpasses. Plant cover type was related to tick presence in this road context. Implications of road permeability and edge effect in patterns of vertebrate–tick relationships in road fragmentation contexts are discussed. Both Borrelia sp. and Bartonella sp. were detected in the ticks analyzed. This is the first report of these bacteria in ticks from Seville. The results confirm the potential risk of acquiring Lyme disease and bartonellosis in this area.