Strict Conservation yet Non-Essential Nature of Plasmid Gene bba40 in the Lyme Disease Spirochete Borrelia burgdorferi

The highly segmented genome of Borrelia burgdorferi, the tick-borne bacterium that causes Lyme disease, is composed of a linear chromosome and more than 20 co-existing endogenous plasmids. Many plasmid-borne genes are unique to B. burgdorferi and some have been shown to provide essential functions at discrete points of the infectious cycle between a tick vector and rodent host. In this study, we investigated the role of bba40, a highly conserved and differentially expressed gene on a ubiquitous linear plasmid of B. burgdorferi. In a prior genome-wide analysis, inactivation of bba40 by transposon insertion was linked with a noninfectious phenotype in mice, suggesting that conservation of the gene in the Lyme disease spirochete reflected a critical function of the encoded protein. To address this hypothesis, we moved the bba40::Tn allele into a similar wild-type background and compared the phenotypes of isogenic wild-type, mutant and complemented strains in vitro and throughout the in vivo mouse/tick infectious cycle. In contrast to the previous study, we identified no defect in the ability of the bba40 mutant to colonize the tick vector or murine host, or to be efficiently transmitted between them. We conclude that bba40 joins a growing list of unique, highly conserved, yet fully dispensable plasmid-borne genes of the Lyme disease spirochete. We infer that the experimental infectious cycle, while including the tick vector and murine host, lacks key selective forces imposed during the natural enzootic cycle. IMPORTANCE The key finding of this study contradicts our premise that the ubiquitous presence and strict sequence conservation of a unique gene in the Lyme disease spirochete, Borrelia burgdorferi, reflect a critical role in either the murine host or tick vector in which these bacteria are maintained in nature. Instead, the outcome of this investigation illustrates the inadequate nature of the experimental infectious cycle currently employed in the laboratory to fully model the enzootic cycle of the Lyme disease spirochete. This study also highlights the importance of complementation for accurate interpretation of mutant phenotypes in genetic studies of Borrelia burgdorferi.