Canine visceral leishmaniasis (CVL) can present as a severe debilitating or subclinical form of the disease; progression is dependent on several factors, including the host immunological status. We performed a previous cohort study in an endemic area of visceral leishmaniasis (VL) that identified and classified dogs as resistant or susceptible to CVL. The bone marrow (BM) is one of the tissues of preference for Leishmania infantum infection and myeloid cells are the main host cells of the parasite. However, little is known about the impact of immune cellular alterations in the BM on disease severity. Recently, it was demonstrated in a murine model that L. donovani infection induces the expansion of hematopoietic stem cells that reside in the BM of infected mice. In the present study, we focused on the response characterization of BM cells from susceptible dogs naturally infected with L. infantum. Longitudinal analysis of the hematological profile and peripheral blood counts revealed significantly decreased red blood cell counts, hemoglobin and hematocrit levels. Reassessment and characterization of BM cellular profile compared to uninfected dogs as controls revealed erythroid cell hypoplasia in susceptible animals. In the characterization by flow cytometry, the T lymphocyte count was significantly higher in the BM of susceptible dogs, when compared to the bone marrow of control dogs. For further BM cell characterization, gene expression profiling by RNA sequencing was performed. Transcriptomic analysis identified 327 differentially expressed genes (DEGs) in susceptible animals versus controls. Enrichment analysis revealed that pathways related to DNA repair were negatively regulated, while pathways related to cell migration were positively and negatively modulated in susceptible dogs. A machine learning algorithm identified a set of four genes (EGR2, FOS, TINAGL1 and ADCY9) with high power to detect susceptible dogs. Currently, these DEGs and their associated pathways are undergoing validation by RT-qPCR, immunofluorescence and functional assays, such as neutral comet and transwell cell migration assays. To perform validation, new BM samples were collected. RT-qPCR will be used to validate the set of four DEGs identified by bioinformatic analysis and those involved in DNA double-break repair and cell migration pathways. Assays evaluating the migratory behavior of cell subsets in susceptible and healthy dogs were conducted; at first, in a two-dimensional environment, no significant differences were observed in migration profiles between the groups. In addition, assays using a 3D environment will be performed to characterize the mesenchymal migration performed by cells that migrate from the BM to the periphery. Immunofluorescence was also performed to validate proteins involved in cell migration processes and revealed higher RAC expression in BM cells from susceptible dogs when compared to cells from dogs not infected with L. infantum. Neutral comet assays will be later performed to identify the double breakage of DNA in the BM cells. In conclusion, the deep transcriptomic analysis of the cells from the BM, one of the central tissues of VL infection, showed profound alterations in the gene expression profile of susceptible dogs to L. infantum infection that in the end would explain the fatal outcome of VL in susceptible animals.