Ixodes ricinus serves as vector for a range of microorganisms capable of causing clinical illness in humans. The microorganisms occur in the same vector populations and are generally affected by the same tick-host interactions. Still, the instars have different host preferences which should manifest in different transmission patterns for various microorganisms in the tick populations, i.e., most microorganisms increase in prevalence rate from larvae to nymphs because their reservoirs are among small mammals and birds that serve as blood hosts for larvae. Other microorganisms, like Anaplasma phagocytophilum, mainly increase in prevalence rates from nymphs to adults, because their reservoirs are larger ungulates that serve as primary blood hosts for nymphs and adults. We sampled a representative sample of ticks from 12 locations on Zealand and Funen, Denmark, and investigated the differences in prevalence rate of infection in larvae, nymphs and adults for multiple pathogens. Prevalence of infection for larvae, nymphs and adults, respectively, was: 0, 1.5 and 4.5% for Borrelia burgdorferi; 0, 4.2 and 3.9% for Borrelia garinii; 0, 6.6 and 6.1% for Borrelia afzelii; 0, 0 and 0.6% for Borrelia valaisiana; 0, 3.7 and 0.6% for Borrelia spielmanii; 0, 0.7 and 1.2% for Babesia divergens; 0, 0, 0.6% for Babesia venatorum; 0, 1.5 and 6.1% for A. phagocytophilum. The results were in general compatible with the hypothesis i.e., that differences in blood host for larvae and nymphs define differences in transmission of infectious agents, but other factors than differences in blood hosts between larvae and nymphs may also be important to consider.