An uncharacterized Rickettsia species was previously identified by molecular detection in Amblyomma americanum ticks from Oklahoma, a state reported to have high Rickettsia seroprevalence. Amblyomma americanum ticks are aggressive feeders capable of transmitting viral, protozoal and bacterial species that cause diseases in humans and animals. Discovering and characterizing novel microorganisms in this tick species is crucial for identifying potential new pathogens. Using A. americanum ticks collected from Oklahoma, we isolated, cultured and sequenced the entire genome of a previously detected, but uncharacterized, novel Rickettsia species. Triturated A. americanum ticks were used as inoculum to culture the novel Rickettsia species in Vero E6 cells, and qPCR testing confirmed the presence of the new Rickettsia species while ruling out the presence of other tick-borne organisms. The total genome size was 1.17 Mbp consisting of a complete chromosome with a 30.7 % G+C content (GenBank accession CP157197). We predicted 1037 genes, 997 coding gene open reading frames, along with 33 tRNAs, 4 ncRNAs and 3 rRNAs. This genome was most similar to Rickettsia canadensis strain CA410 at 91.1 % identity, based on average nucleotide identity analysis. A maximum-likelihood phylogeny tree, constructed using 636 concatenated core proteins, placed the novel Rickettsia species in a clade with Rickettsia canadensis. We propose the name Rickettsia oklahomensis sp. nov., strain Oklahoma 10, which is available from the Centers for Disease Control and Prevention Rickettsial Isolate Reference Collection (WDCM 1093), Atlanta, GA, USA (CRIRC accession number ROK001).