Attenuation of the 2022 global outbreak monkeypox virus relative to its clade IIb ancestor

Monkeypox virus (MPXV) is a zoonotic virus endemic to Africa that has recently re-emerged, becoming the first orthopoxvirus with human-to-human transmission since smallpox eradication. The 2022 global epidemic of MPXV was caused by Clade IIb lineage B.1 virus that derives from the Clade IIb lineage A.1, which is endemic to West Africa1-3. Here we compared an early lineage B.1 virus with its closest earlier A.1 ancestor and show that despite only 46 nucleotide differences, lineage B.1 has attenuating phenotypic changes. Both B.1 and A.1 viruses replicated equivalently in cell culture, but B.1 had defects in long-range spread. MPXV B.1 also displayed defects in innate immune control, inducing higher IFN{beta}, innate immune signalling and MAPK gene expression, and a reduced capacity to suppress interleukin-1{beta}-mediated immune responses. Finally, MPXV B.1 had reduced virulence in a mouse model of MPXV infection relative to the ancestral A.1 endemic strain. Our work demonstrates biological differences between human circulating MPXV lineages that correlate with clinical observations documenting reduced in-host viral dissemination and disease severity for lineage B. Our data also reveal that the APOBEC3-like mutational footprint in MPXV is not only a signature of sustained human transmission but can also drive phenotypic changes.