Abstract

Vaccinia virus (VACV) and monkeypox virus (MPXV) are closely related members of the family Poxviridae, genus Orthopoxvirus, both capable of causing systemic infections with potential neurological complications. Although live, replication-competent VACV strains were historically used in smallpox vaccination, their clinical use was associated with rare but severe central nervous system (CNS)-related adverse events. Despite this, the mechanisms underlying VACV-induced CNS pathology, particularly olfactory dysfunction, remain poorly characterized. In this study, we found VACV-VR1354, a tissue culture-adapted derivative of the neurovirulent Western Reserve strain, can invade the CNS via the olfactory route and induce olfactory impairment. By using an intranasal infection model in two inbred mouse strains—C57BL/6N and BALB/c, we demonstrate that VACV-VR1354 efficiently disseminates from the nasal mucosa to the brain, as evidenced by a spatiotemporal gradient of viral DNA load (nasal mucosa > olfactory bulb > cerebrum > cerebellum). Evans blue extravasation assays indicated a transient increase in blood–brain barrier (BBB) permeability in the olfactory bulb, peaking at 7 days post-infection (dpi) and resolving by 14 dpi, with more pronounced effects in C57BL/6N mice. Neuroinvasion was accompanied by robust microglial and astrocytic activation, as well as injury to mature olfactory sensory neurons, particularly at 7 dpi. Transcriptomic profiling of the olfactory bulb revealed significant downregulation of olfactory receptor (OR) genes, with the downregulated genes significantly enriched in olfactory transduction pathways. Concurrently, strong upregulation of proinflammatory cytokines, chemokines, and interferon-stimulated genes (ISGs) was detected in the olfactory bulb tissue, indicative of intense neuroinflammation. Behaviorally, infected C57BL/6N mice exhibited impaired aversion to camphor odor between 14 and 49 dpi, with full functional recovery observed by 56 dpi. Coll