Patricia Rosa, Ph.D.

Chief, Molecular Genetics Section
Senior Investigator

Description of Research Program

Research in this laboratory focuses on the spirochete Borrelia burgdorferi, the cause of Lyme disease, the most common arthropod-borne disorder in the United States. B. burgdorferi is maintained in nature through an infectious cycle between wild mammals and ticks. Occasionally, infected ticks feed upon humans and transmit Lyme disease.

Although human infection is not relevant to the transmission cycle, it has medical significance as a multisystemic, potentially chronic illness. The tick vector and the mammalian host represent two very different environments, and there is good evidence for differential gene expression by borreliae in these locations.

The Infectious Cycle of B. burgdorferi

• B. burgdorferi spirochetes persist in a latent state in midguts of infected ticks for many months.
• After a tick attaches to a mammalian host and ingests a bloodmeal, spirochetes multiply and efficiently move to the salivary glands.
• B. burgdorferi is then transmitted via tick saliva and remains in the mammalian skin for several days before dissemination via the bloodstream.
• Spirochetes persist in low numbers in infected mammals, yet are efficiently acquired by feeding ticks following attachment.

This scenario suggests that B. burgdorferi responds to environmental cues in order to adapt to and move between the arthropod (tick) vector and mammalian host. Recent experiments document modulation of spirochetal outer surface proteins in response to environmental conditions and reinforce this hypothesis.

Our broad objective is to use a molecular genetic approach to elucidate the mechanisms of adaptation and variation in B. burgdorferi and their roles in the infectious cycle. The specific aims of our research are as follows:

1. Develop basic genetic tools to manipulate borrelial genes of interest. The availability of the complete genomic sequence of B. burgdorferi represents a wealth of information that can be effectively utilized through a genetic approach. However, studies of the biology of B. burgdorferi and the pathogenesis of Lyme disease have been severely limited by a lack of genetic tools, because most methods that have been developed for other bacteria can not be directly applied to borreliae. To address our scientific goals, we aim to develop a set of basic genetic tools for B. burgdorferi. The ability to perform routine genetic manipulations in B. burgdorferi will greatly facilitate our research objectives as well as those of other investigators in the field.
2. Understand the structure and function of plasmids in B. burgdorferi. A distinguishing feature of the B. burgdorferi genome is the presence of a linear chromosome and multiple linear and circular plasmids. The genomic sequence of B. burgdorferi identified 21 different plasmids, representing the largest known complement of plasmids of all bacteria and constituting one third of the spirochete's DNA. More than 90% of the plasmid-encoded genes are unique to B. burgdorferi, without homologs in any other organisms, suggesting they encode functions pertinent to the distinctive lifestyle of the spirochete. Consistent with this hypothesis is growing evidence that B. burgdorferi plasmids carry genes critical for survival in or transmission between the tick and mammal host.
   
   Despite the apparent significance of B. burgdorferi plasmids to its life cycle, relatively little is known about the mechanisms of plasmid replication and partitioning, or the functions of the proteins they encode. We have begun a series of experiments to test our hypothesis that the multiplicity and unusual structure of the DNA molecules that form the B. burgdorferi genome are intrinsic to the ability of the spirochete to adapt to, persist in, and be transmitted between the alternating mouse/tick environments of the infectious cycle. These experiments aim to
   • Define the minimal plasmid elements required for replication, partitioning, and incompatibility of linear and circular replicons
   • Assess the roles of individual plasmids for survival in, or transmission between, the tick vector, and mammalian host
3. Determine how B. burgdorferi responds to particular environmental cues in order to persist in and be transmitted between the tick vector and mammalian host. We hypothesize that discrete environmental signals induce appropriate bacterial responses that are critical for survival and transmission of the spirochete during the infectious cycle. We plan to
   • Determine which proteins are made in different sites or at different stages of the infection in ticks and mammals
   • Determine what these proteins do and how the genes encoding them are regulated
   • Decipher the signals that mediate the adaptive responses

   Knowing which bacterial proteins are synthesized in the mammal versus the tick and gaining insight into their functions will contribute to a better understanding of the pathogenesis of Lyme disease. This knowledge is relevant to the diagnosis and prevention of Lyme disease.

Conclusion

The transmission of B. burgdorferi between ticks and mammals represents an ideal system in which to study the adaptive responses of a bacterial pathogen to its vector and host environments. All steps of this infectious cycle can be reproduced in the laboratory, making it accessible to scientific investigation. Molecular genetics represents a powerful method with which to address this system.

Previous studies have identified spirochetal components that should be important in the adaptation of B. burgdorferi to its environment. Ongoing and future studies are designed to test the roles of these genes and their products in the infectious cycle and to identify additional genes that allow the spirochetes to adapt, persist, and be transmitted between ticks and mammals. This research should elucidate the biological basis of these bacterium-host-vector relationships and the factors that contribute to the pathogenesis of disease in an incidental human host. 

Research Group Members

Back row, left to right: Aaron Bestor, Patti Rosa, Mollie Jewett, Kit Tilly, Philip Stewart. Front row, left to right: Ryan Rego, Melissa Jessop, Claire Checroun, Amit Sarkar

Selected Publications

(View list in PubMed.)

Jewett MW, Byram R, Bestor A, Tilly K, Lawrence K, Burtnick MN, Gherardini F, Rosa PA. Genetic basis for retention of a critical virulence plasmid of Borrelia burgdorferi. Mol Microbiol. 2007 Nov;66(4):975-90.

Jewett MW, Lawrence K, Bestor AC, Tilly K, Grimm D, Shaw P, VanRaden M, Gherardini F, Rosa PA. The critical role of the linear plasmid lp36 in the infectious cycle of Borrelia burgdorferi. Mol Microbiol. 2007 Jun;64(5):1358-74.

Tilly K, Bestor A, Jewett MW, Rosa P. Rapid clearance of Lyme disease spirochetes lacking OspC from skin. Infect Immun. 2007 Mar;75(3):1517-9.

Tilly K, Krum JG, Bestor A, Jewett MW, Grimm D, Bueschel D, Byram R, Dorward D, Vanraden MJ, Stewart P, Rosa P. Borrelia burgdorferi OspC protein required exclusively in a crucial early stage of mammalian infection. Infect Immun. 2006 Jun;74(6):3554-64.

Stewart PE, Wang X, Bueschel DM, Clifton DR, Grimm D, Tilly K, Carroll JA, Weis JJ, Rosa PA. Delineating the requirement for the Borrelia burgdorferi virulence factor OspC in the mammalian host. Infect Immun. 2006 Jun;74(6):3547-53.

Rosa PA, Tilly K, Stewart PE. The burgeoning molecular genetics of the Lyme disease spirochaete. Nat Rev Microbiol. 2005 Feb;3(2):129-43. Review.

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