BackgroundCarbapenemase-producing makes a great contribution to carbapenem resistance in Gram-negative bacilli.BlaAFM-1gene was first discovered by us inAlcaligenes faecalisAN70 strain isolated in Guangzhou of China and, was submitted to NCBI on 16 November 2018.MethodsAntimicrobial susceptibility testing was performed by broth microdilution assay using BD Phoenix 100. The phylogenetic tree of AFM and other B1 metallo-β-lactamases was visualized by MEGA7.0. Whole-genome sequencing technology was used to sequence carbapenem-resistant strains including theblaAFM-1gene. Cloning and expressing ofblaAFM-1were designed to verify the function of AFM-1 to hydrolyze carbapenems and common β-lactamase substrates. Carba NP and Etest experiments were conducted to evaluate the activity of carbapenemase. Homology modeling was applied to predict the spatial structure of AFM-1. A conjugation assay was performed to test the ability of horizontal transfer of AFM-1 enzyme. The genetic context ofblaAFM-1was performed by Blast alignment.ResultsAlcaligenes faecalisstrain AN70,Comamonas testosteronistrain NFYY023,Bordetella trematumstrain E202, andStenotrophomonas maltophiliastrain NCTC10498 were identified as carrying theblaAFM-1gene. All of these four strains were carbapenem-resistant strains. Phylogenetic analysis revealed that AFM-1 shares little nucleotide and amino acid identity with other class B carbapenemases (the highest identity (86%) with NDM-1 at the amino acid sequence level). The spatial structure of the AFM-1 enzyme was predicted to be αβ/βα sandwich structure, with two zinc atoms at its active site structure. Cloning and expressing ofblaAFM-1verified AFM-1 could hydrolyze carbapenems and common β-lactamase substrates. Carba NP test presented that the AFM-1 enzyme possesses carbapenemase activity. The successful transfer of pAN70-1(plasmid of AN70) toE.coliJ53 suggested that theblaAFM-1gene could be disseminated by the plasmid. The genetic context ofblaAFMindicated that the downstream of theblaAFMgene was always adjacent totrpFandbleMBL. Comparative genome analysis revealed thatblaAFMappeared to have been mobilized by an ISCR27-related mediated event.ConclusionsTheblaAFM-1gene is derived from chromosome and plasmid, and theblaAFM-1gene derived from the pAN70-1 plasmid can transfer carbapenem resistance to susceptible strains through horizontal transfer. SeveralblaAFM-1-positive species have been isolated from feces in Guangzhou, China.Supplementary InformationThe online version contains supplementary material available at 10.1186/s12941-023-00592-0.