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Laboratory of Zoonotic Pathogens

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Frank Gherardini, Ph.D.

Photo of Frank Gherardini, Ph.D.

Chief, Gene Regulation Section, LZP

Major Areas of Research

  • Physiology, biochemistry, gene regulation, and pathogenesis of Borrelia burgdorferi
  • Treponema pallidum
  • Burkholderia mallei
  • Identification of genes required for intracellular survival of Burkholderia pseudomallei
 

Program Description

Borrelia Projects: This research focuses on the physiology, biochemistry, gene regulation, and pathogenesis of Borrelia burgdorferi, the causative agent of Lyme disease in humans. B. burgdorferi faces several environmental and immunological challenges during its infective cycle and must alter (regulate) gene expression to successfully adapt to these conditions.

Analysis of the B. burgdorferi genome sequence has revealed that there are very few known regulatory proteins in this bacterium. Conspicuously absent are global regulatory proteins such as CRP, LexA, Fnr, IHF, Lrp, and the sigma factors involved in the heat shock response, ┐σ32 and σ24.

This suggests that, compared to other well-characterized pathogenic bacterial systems, the global regulatory systems operating in B. burgdorferi are relatively simple. Clearly, these systems are required for B. burgdorferi to adapt as it encounters very different environments during transfer from an animal reservoir to the tick and then to a human host.

Research efforts in this group have focused on three important regulatory proteins: 1)BosR, a Zn-dependent transcriptional activator that regulates key antioxidant enzymes; 2) σ54, an alternate sigma factor that also regulates certain parts of the oxidative stress response and regulates the osmotic stress response; and 3) vs., which controls the stationary phase of growth and the expression of genes that are critical to the pathogenesis of Lyme disease.

Illustration of the transmission of Lyme disease, reactive oxygen (ROS) and reactive nitrogen (NOS) present a challenge to the survival of B. burgdorferi. As the bacteria move from the midgut to the feeding site, they encounter a dramatic increase in ROS and respond by increasing the expression of key protective enzymes (NapA, NpX, TrxR, and Trx).
During the transmission of Lyme disease, reactive oxygen (ROS) and reactive nitrogen (NOS) present a challenge to the survival of B. burgdorferi. As the bacteria move from the midgut to the feeding site, they encounter a dramatic increase in ROS and respond by increasing the expression of key protective enzymes (NapA, NpX, TrxR, and Trx).

Burkholderia Projects: Burkholderia pseudomallei, the etiological agent of melioidosis, is a Gram-negative, facultatively anaerobic, motile bacillus that is responsible for a broad spectrum of illnesses observed in both humans and animals.

While epidemiological surveys have demonstrated that B. pseudomallei is endemic to regions that typically border the equator, the incidence of disease is particularly high in Southeast Asia and northern Australia. In northeastern Thailand alone, an estimated 20 percent of community-acquired septicemias and approximately 40 percent of deaths due to complications associated with bacterial sepsis can be attributed to this organism. The manifestations of melioidosis are commonly represented by acute, sub-acute, and chronic illnesses, with the clinical manifestations often being mistaken for malaria, plague, pneumonia, and miliary tuberculosis. Infections are typically acquired via inhalation or aspiration, ingestion, or via the direct contact of damaged surface tissues with contaminated water or soil.

Burkholderia mallei, the etiological agent of glanders, is a Gram-negative bacterium that is responsible for disease in donkeys, mules, horses, and occasionally humans. Unlike the environmental saprophyte B. pseudomallei, however, B. mallei does not persist in nature outside of its soliped hosts. While B. mallei and B. pseudomallei are genotypically similar, significant phenotypic differences do exist between the two pathogenic species. Although glanders is one of the oldest diseases known to man, relatively little is known about the pathogenesis of disease caused by B. mallei. This phenomenon is due primarily to the lack of the disease in North America, along with the fact that B. mallei can be a particularly dangerous organism to study even in a controlled laboratory environment.

B. pseudomallei is known to resist the bactericidal activity of both reactive oxygen and reactive nitrogen intermediates as well as to survive and multiply within several mouse and human macrophage cell lines (see electron micrograph). In order to identify genetic loci associated with these virulence phenotypes, specifically those genes that are up- or down-regulated in an intracellular environment, we use a standard macrophage uptake assay, recover the intracellular bacteria, and isolate bacterial mRNA to probe DNA micro-arrays. The data sets obtained from these studies are used to identify genes required for intracellular survival. Genes of interest are disrupted by allelic exchange and their functions are defined using both in vivo and in vitro model systems (mouse macrophage invasion, infectivity in hamsters and mice).

Photo of B. pseudomallei surviving inside a human macrophage.
B. pseudomallei (designated by arrow) surviving inside a human macrophage.

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Biography

Dr. Gherardini received his doctorate in 1987 from the University of Illinois, studying enzymes involved in the utilization of galactomannans by Bacteroides ovatus. From 1991 to 2001, he was a tenured professor in the Department of Microbiology at the University of Georgia. In 2001, Dr. Gherardini joined the Rocky Mountain Laboratories, where he is chief of the Gene Regulation Section and a senior investigator in the Laboratory of Zoonotic Pathogens.

Research Group

Research Group Members
Left to right: Travis Bourret, Crystal Richards, David Revelli, Natalie Moran, Kevin Lawrence, Julie Boylan, Hua Su, Frank Gherardini; front: Dexter

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Selected Publications

Warawa JM, Long D, Rosenke R, Gardner D, Gherardini FC. Bioluminescent diagnostic imaging to characterize altered respiratory tract colonization by the Burkholderia pseudomallei capsule mutant. Front Microbiol. 2011;2:133.

Bourret TJ, Boylan JA, Lawrence KA, Gherardini FC. Nitrosative damage to free and zinc-bound cysteine thiols underlies nitric oxide toxicity in wild-type Borrelia burgdorferi. Mol Microbiol. 2011 Jul;81(1):259-73.

Brett PJ, Burtnick MN, Heiss C, Azadi P, DeShazer D, Woods DE, Gherardini FC.  Burkholderia thailandensis oacA mutants facilitate the expression of Burkholderia mallei-like O polysaccharides.Infect Immun. 2011 Feb;79(2):961-9.

Sun YC, Koumoutsi A, Jarrett C, Lawrence K, Gherardini FC, Darby C, Hinnebusch BJ. Differential control of Yersinia pestis biofilm formation in vitro and in the flea vector by two c-di-GMP diguanylate cyclases. PLoS One. 2011 Apr 29;6(4):e19267

Burtnick MN, DeShazer D, Nair V, Gherardini FC, Brett PJ. Burkholderia mallei cluster 1 type VI secretion mutants exhibit growth and actin polymerization defects in RAW 264.7 murine macrophages. Infect Immun. 2010 Jan;78(1):88-99.

Xu H, Caimano MJ, Lin T, He M, Radolf JD, Norris SJ, Gherardini FC, Wolfe AJ, Yang X.  Role of acetyl-phosphate in activation of the Rrp2-RpoN-RpoS pathway in Borrelia burgdorferi.  PLoS Pathog. 2010. Sep 16;6(9).

Visit PubMed for a complete publications listing.

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Last Updated March 25, 2013