Borrelia isolates associated with Lyme borreliosis were previously divided into 3 genospecies, B. burgdorferi sensu stricto, B. garinii and group VS461, on the basis of DNA homology. B. burgdorferi sensu stricto and B. garinii were identified by monoclonal antibodies (MAbs), H3TS and D6 respectively, but no MAbs were available to identify group VS461. Two MAbs were produced, I 17.3 and J 8.3 which reacted with OspB and OspA proteins, respectively, of strains belonging to group VS461, which should be named B. afzelii sp. nov. 24 strains were assigned to B. afzelii sp. nov., 11 of them being isolated from skin lesions, 6 from acrodermatitis chronica atrophicans (ACA) and 5 from erythema chronicum migrans (ECM). Although quite unknown in the USA, ACA has frequently been reported in northern Europe where B. afzelii sp. nov. is commonly isolated. This study documents the involvement of B. afzelii sp. nov. as a specific aetiological agent of ACA.