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Trypanosoma rhipicephalis Marotta et al. (2018) is a species isolated from Rhipicephalus microplus ticks collected from native bovine Seropédica, in the state of Rio de Janeiro. This study aimed to investigate the interaction of the tick Rhipicephalus sanguineus s. l. with T. rhipicephalis by in vitro artificial feeding. Eight females of R. sanguineus s. l. partially fed on rabbits. Tick infection was performed by an artificial feeding system using plastic tips for 12 hours. Canine blood used for feeding the ticks test group was previously infected with T. rhipicephalis 108/mL. The hemolymph smear test was performed in all females after experimental infection. The daily posture was collected and organized in pools of each female per posture day. The eggs were divided into three groups, the first group for eggs PCR, the second for hatching and larval PCR, and the third group of eggs for isolation in cell culture. The evaluation of the presence of DNA in the macerated eggs of experimentally infected ticks showed two positive PCR samples. The evaluation of the presence of DNA in experimentally infected females showed all samples tested positive. For the hemolymph test, tick number 7 presented epimastigote developmental forms and amastigotes of T. rhipicephalis. Experimental infection by artificial feeding proved to be a suitable tool to study the interaction of T. rhipicephalis in R. sanguineus s. l. ticks. The results show the transovarial transmission of T. rhipicephalis by R. sanguineus s. l., as well as the interaction of the protozoan in the organism of this tick species.