<p>Human parvovirus B19 (B19V) infection is a significant but underrecognized complication, commonly linked not only to aplastic anemia but also pancytopenia, especially in hematopoietic stem cell transplantation (HSCT) recipients. While B19V’s tropism for erythroid progenitor cells (EPCs) is well-documented, its restriction to EPCs fails to fully explain the pathogenesis of pancytopenia. In this study, we used PrimeFlow RNA assay, and single-cell full-length transcriptome sequencing (scFAST-seq) to show that B19V could infect hematopoietic stem cells (HSCs) and initiate viral transcription, resulting in increased apoptosis, impaired self-renewal and multilineage differentiation of HSCs, which may contribute directly to pancytopenia. Further analysis revealed that B19V could activate the JAK2/STAT5 signaling pathway in HSCs to promote viral persistence. Pharmacological inhibition with baricitinib markedly reduced the viral load and partially restored hematopoietic differentiation capacity in vitro. Taken together, our findings reveal B19V as a previously unrecognized HSC-tropic virus that disrupts function of HSCs and may drive pancytopenia. Targeting the JAK2/STAT5 signaling by baricitinib shows promising therapeutic potential for reversing virus-induced bone marrow failure. This work not only deepens our understanding of viral tropism and pathogenesis in the hematopoietic niche, but also opens up new possibilities for treating bone marrow damage after transplantation and in other hematological diseases.</p>