Vector-borne pathogens often consist of genetically distinct strains that can establish co-infections in the vertebrate host and the arthropod vector. Co-infections (or mixed infections) can result in competitive interactions between strains with important consequences for strain abundance and transmission. Here we used the spirochete bacterium, Borrelia afzelii, as a model system to investigate the interactions between strains inside its tick vector, Ixodes ricinus. Larvae were fed on mice infected with either one or two strains of B. afzelii. Engorged larvae were allowed to molt into nymphs that were subsequently exposed to three seasonal treatments (artificial summer, artificial winter, and natural winter), which differed in temperature and light conditions. We used strain-specific qPCRs to quantify the presence and abundance of each strain in the immature ticks. Co-infection in the mice reduced host-to-tick transmission to larval ticks and this effect was maintained in the resultant nymphs at 1 and 4 months after the larva-to-nymph molt. Competition between strains in co-infected ticks reduced the abundance of both strains. This inter-strain competition occurred in the three life stages that we investigated: engorged larvae, recently molted nymphs, and overwintered nymphs. The abundance of B. afzelii in the nymphs declined by 40.5% over a period of 3 months, but this phenomenon was not influenced by the seasonal treatment. Future studies should investigate whether inter-strain competition in the tick influences the subsequent strain-specific transmission success from the tick to the vertebrate host.