Regulatory Protein BBD18 of the Lyme Disease Spirochete: Essential Role During Tick Acquisition?

Abstract

The Lyme disease spirochete Borrelia burgdorferi senses and responds to environmental cues as it transits between the tick vector and vertebrate host. Failure to properly adapt can block transmission of the spirochete and persistence in either vector or host. We previously identified BBD18, a novel plasmid-encoded protein of B. burgdorferi, as a putative repressor of the host-essential factor OspC. In this study, we investigate the in vivo role of BBD18 as a regulatory protein, using an experimental mouse-tick model system that closely resembles the natural infectious cycle of B. burgdorferi. We show that spirochetes that have been engineered to constitutively produce BBD18 can colonize and persist in ticks but do not infect mice when introduced by either tick bite or needle inoculation. Conversely, spirochetes lacking BBD18 can persistently infect mice but are not acquired by feeding ticks. Through site-directed mutagenesis, we have demonstrated that abrogation of spirochete infection in mice by overexpression of BBD18 occurs only with bbd18 alleles that can suppress OspC synthesis. Finally, we demonstrate that BBD18-mediated regulation does not utilize a previously described ospC operator sequence required by B. burgdorferi for persistence in immunocompetent mice. These data lead us to conclude that BBD18 does not represent the putative repressor utilized by B. burgdorferi for the specific downregulation of OspC in the mammalian host. Rather, we suggest that BBD18 exhibits features more consistent with those of a global regulatory protein whose critical role occurs during spirochete acquisition by feeding ticks.

A small intergenic region of lp17 is required for evasion of adaptive immunity and induction of pathology by the Lyme disease spirochete

“…The results reported in this study are part of a growing body of work implicating lp17 as an important player in the infectious cycle of B. burgdorferi. This highly conserved, yet relatively uncharacterised plasmid has now been suggested to encode at least two mammalian virulence determinants, in addition to a regulator protein that is essential for acquisition by larval ticks (Casselli et al, 2012;Hayes et al, 2014). Lp17 remains an attractive target for future studies involving the Lyme disease spirochete, and investigation into the mechanism of action of the bbd07 locus described herein could lead to improved strategies for combating this important animal and human pathogen.…”

“…Clonal isolates of B. burgdorferi lacking various combinations of non‐essential plasmids have provided insights into their respective requirements during infection in mice (Dulebohn, Bestor, Rego, Stewart, & Rosa, , Grimm et al, , Jewett et al, , Kawabata, Norris, & Watanabe, , Labandeira‐Rey, Seshu, & Skare, , Labandeira‐Rey & Skare, , Purser & Norris, ). Investigations into the infectious requirement of other plasmids that are maintained with higher fidelity or that are essential for viability have proven more difficult (Bestor et al, , Casselli, Tourand, & Bankhead, , Hayes et al, , Jewett et al, ). One such plasmid is the ~17 kb linear plasmid, lp17.…”

“…Another study has implicated the mechanism of BBD18 as a post‐transcriptional negative regulator of the alternative sigma factor RpoS (Dulebohn, Hayes, & Rosa, ). Additionally, unidentified factors encoded on the extreme right‐end of lp17 may be required for maximum tissue colonisation in mice (Casselli et al, ; Hayes et al, ). Furthermore, previous studies have implicated several lp17‐encoded genes located near the telomere ends as being transcriptionally activated by the mammalian host‐specific BosR/Rrp2‐RpoN‐RpoS regulon(s) (Boardman et al, ; Caimano et al, ; Ojaimi et al, ; Ouyang et al, ; Ouyang, Blevins, & Norgard, ), although the involvement of these genes during infection is not well understood.…”

“…Thus far, studies on lp17 have been conducted primarily in vitro using non-infectious B. burgdorferi clones, as this plasmid has been demonstrated as nonessential and easily manipulatable in these clones (Beaurepaire & Chaconas, 2005;Chaconas, Stewart, Tilly, Bono, & Rosa, 2001;Sadziene, Wilske, Ferdows, & Barbour, 1993;Sarkar, Hayes, Dulebohn, & Rosa, 2011;Tourand et al, 2006). As spontaneous loss of lp17 in cultured spirochetes has been reported only rarely, and genetic manipulation of this plasmid in infectious clones is challenging, the role for lp17 during infection of mammals and ticks has only begun to be elucidated (Casselli et al, 2012;Grimm, Elias, Tilly, & Rosa, 2003;Hayes et al, 2014;Norris et al, 2011;Sadziene et al, 1993). Recent works have demonstrated that bbd18, a gene located on the right-end of lp17, is required for suppression of the known surface immunogen OspC, and that this likely has biological significance during colonisation of the tick vector (Casselli et al, 2012;Hayes et al, 2014;Sarkar et al, 2011).…”

The need to unravel the twisted nature of the Borrelia burgdorferi sensu lato complex across Europe

“…Since then, the development of diverse molecular tools has enabled investigators to probe and uncover some of the underlying pathogenic mechanisms of the spirochete on a molecular genetic level [1]. These include the use of targeted mutagenesis studies for investigating the role of genes in a tick-animal model in a high-throughput manner [49][50][51][52], the use of the cre-lox recombination system for the deletion of a set of genes on a plasmid [53] and the use of reporter genes including cat gene [54], lacZ [55,56], luc [57,58] and GFP [33,59]. Also published on and in use are experiments with an inducible gene expression system when the gene of interest is essential for viability and cannot be deleted.…”