Comparative genomics analysis of three conserved plasmid families in the Western Hemisphere soft tick-borne relapsing fever borreliae provides insight into variation in genome structure and antigenic variation systems.

spirochetes, causative agents of Lyme disease and relapsing fever (RF), have uniquely complex genomes, consisting of a linear chromosome and both circular and linear plasmids. The plasmids harbor genes important for the vector-host life cycle of these tick-borne bacteria. The role of plasmids from Lyme disease causing spirochetes is more refined compared to RF because of limited plasmid-resolved genome assemblies for the latter. We recently addressed this limitation and found that three linear plasmid families (F6, F27, and F28) were syntenic across all the RF species that we examined. Given this conservation, we further investigated the three plasmid families. The F6 family, also known as the megaplasmid, contained regions of repetitive DNA. The F27 was the smallest, encoding genes with unknown function. The F28 family encoded the putative expression locus for antigenic variation in all species except and . Taken together, this work provides a foundation for future investigations to identify essential plasmid-localized genes that drive the vector-host life cycle of RF . IMPORTANCE spp. spirochetes are arthropod-borne bacteria found globally that infect humans and other vertebrates. RF borreliae are understudied and misdiagnosed pathogens because of the vague clinical presentation of disease and the elusive feeding behavior of argasid ticks. Consequently, genomics resources for RF spirochetes have been limited. Analyses of plasmids have been challenging because they are often highly fragmented and unassembled in most available genome assemblies. By utilizing Oxford Nanopore Technologies, we recently generated plasmid-resolved genome assemblies for seven spp. found in the Western Hemisphere. This current study is an in-depth investigation into the linear plasmids that were conserved and syntenic across species. We identified differences in genome structure and, importantly, in antigenic variation systems between species. This work is an important step in identifying crucial plasmid-localized genetic elements essential for the life cycle of RF spirochetes.