Development of Genetic System to Inactivate a Borrelia turicatae Surface Protein Selectively Produced within the Salivary Glands of the Arthropod Vector

Job E Lopez,Paul A Beare,Daniel E Sturdevant,Sandra J Raffel,Karin E Peterson,Tom G Schwan,Reid Hargrove,Christopher P Brooks,Vijayaraj Nagarajan,Hannah K Wilder,Janet Foley

doi:10.1371/journal.pntd.0002514

Abstract

Background Borrelia turicatae, an agent of tick-borne relapsing fever, is an example of a pathogen that can adapt to disparate conditions found when colonizing the mammalian host and arthropod vector. However, little is known about the genetic factors necessary during the tick-mammalian infectious cycle, therefore we developed a genetic system to transform this species of spirochete. We also identified a plasmid gene that was up-regulated in vitro when B. turicatae was grown in conditions mimicking the tick environment. This 40 kilodalton protein was predicted to be surface localized and designated the Borrelia repeat protein A (brpA) due to the redundancy of the amino acid motif Gln-Gly-Asn-Val-Glu.Methodology/Principal FindingsQuantitative reverse-transcriptase polymerase chain reaction using RNA from B. turicatae infected ticks and mice indicated differential regulation of brpA during the tick-mammalian infectious cycle. The surface localization was determined, and production of the protein within the salivary glands of the tick was demonstrated. We then applied a novel genetic system for B. turicatae to inactivate brpA and examined the role of the gene product for vector colonization and the ability to establish murine infection.Conclusions/SignificanceThese results demonstrate the complexity of protein production in a population of spirochetes within the tick. Additionally, the development of a genetic system is important for future studies to evaluate the requirement of specific B. turicatae genes for vector colonization and transmission.

Highlights

Tick-borne pathogens must efficiently adapt to the arthropod vector and mammalian host to ensure survival of the microorganism
Most species of tick-borne relapsing fever spirochetes are maintained in enzootic cycles, and given an approximately 20 year life span, the arthropod vector for Borrelia turicatae represents a reservoir for the pathogens
We have identified a gene that was designated the Borrelia repeat protein A. brpA was up-regulated in a portion of the spirochetes colonizing Ornithodoros turicata, the vector for B. turicatae

Summary

Introduction

Tick-borne pathogens must efficiently adapt to the arthropod vector and mammalian host to ensure survival of the microorganism. The surface localization and production of the protein was determined, and a system to inactivate the brpA was developed to evaluate the necessity of the gene during the tick-mammalian infectious cycle These results demonstrate the up-regulation of a B. turicatae gene within the vector, differential protein production during midgut and salivary gland colonization, and the ability to inactivate B. turicatae genes by allelic exchange. An agent of tick-borne relapsing fever, is an example of a pathogen that can adapt to disparate conditions found when colonizing the mammalian host and arthropod vector. We identified a plasmid gene that was up-regulated in vitro when B. turicatae was grown in conditions mimicking the tick environment This 40 kilodalton protein was predicted to be surface localized and designated the Borrelia repeat protein A (brpA) due to the redundancy of the amino acid motif Gln-Gly-Asn-Val-Glu

Methods

Results

Discussion

Conclusion