CD8+T cells are significant because they clear intracellular infections;however, subverting these immune cells has implications to long-term infections. We will study how Brucella melitensis, a facultative intracellular bacterium, chronically persists in animals in the presence of an immune response. We hypothesize that low numbers and ineffectual CD8+T cells permit continuing infection. We have found BALB/c mice infected with B. melitensis results in chronic infection lasting >12 months using bioluminescent Brucella. Evaluation of the CD8+T cell memory pool from these mice reveals a failure to maintain the CD8+T cell memory phenotype (LFA1hi, KLRG1lo, and CD127hi), or polyfunctional cytokine expression (IL-2, IFN-? and TNF-?). These findings indicate CD8+T cells express an "exhausted" phenotype suggesting that Brucella evades this known effector mechanism for removing intracellular pathogens. Further, a Brucella protein, TcpB, can inhibit CD8+T cell killing of Brucella peptide expressing target cells in vivo. Our long-term goal is to understand how the bacteria remain in a chronic state in the presence of an immune response, by investigating the following Aims:
Aim 1 : To determine the CD8+T cell response in BALB/c mice during acute and chronic infection. We will compare the magnitude and effectiveness of CD8+Tcell responses in mice during acute and chronic infection using CD8+Teffector and memory markers, transcription factor expression, cytokine production, and in vivo killing. Impact: We will identify CD8+T cell phenotypic differences between acute and chronic infection and hypothesize an "exhausted" phenotype and ineffectual CD8+Tcells contribute to chronic brucellosis.
Aim 2 : To determine the protective capacity of Brucella-induced memory CD8+T cells. We will examine the capacity of adoptively transferred CD8+T cells from acute versus chronic infections to protect naive animals from a first infection. Impact: We expect CD8+Tcells from acute infected mice will protect naive mice better than cells from chronically infected mice supporting a loss of functional CD8+Tcells with chronic infection.
Aim 3 : To determine the inhibitory ability of Brucella TcpB on cytotoxic CD8+T cells. We will determine the ability of the TcpB protein to inhibit cytotoxic CD8+T cell killing of infected cells and the kinetics of this inhibition in vivo during infection. Impact: We expect that TcpB protein modulates the adaptive immune response by inhibiting CD8+T cell killing permitting long-term survival of Brucella-infected cells. Our studies represent a critical first step in elucidating how Brucella infection shapes CD8+T cell effector and memory responses. This work will fill a serious void in understanding the role of CD8+T cells in brucellosis that likely participate in the resolution of tis disease.

Public Health Relevance

Brucellosis is a chronic debilitating bacterial disease and a prevalent zoonosis worldwide. CD8+ T cells likely play a critical role in controlling the intracellular infection, but little is known regarding effector and memory responses of these CD8+T cells during infection. We will study how effector and memory CD8+T cells respond in acute versus chronic Brucella infection, and we hypothesize that these studies will identify a decline in functional CD8+T cells as brucellosis transitions from early to late infection. Our findings will provide future opportunities to enhance development and persistence of functional CD8+T cells leading to improved bacterial clearance and designing vaccines that can potentate CD8+T memory cells.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI073558-06
Application #
8462526
Study Section
Vaccines Against Microbial Diseases (VMD)
Program Officer
Mukhopadhyay, Suman
Project Start
2007-04-01
Project End
2017-04-30
Budget Start
2013-05-01
Budget End
2014-04-30
Support Year
6
Fiscal Year
2013
Total Cost
$353,675
Indirect Cost
$118,675
Name
University of Wisconsin Madison
Department
Pathology
Type
Schools of Veterinary Medicine
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715
Kassem, Issmat I; Splitter, Gary A; Miller, Sally et al. (2016) Let There Be Light! Bioluminescent Imaging to Study Bacterial Pathogenesis in Live Animals and Plants. Adv Biochem Eng Biotechnol 154:119-45
Khan, Mike; Harms, Jerome S; Marim, Fernanda M et al. (2016) The Bacterial Second Messenger Cyclic di-GMP Regulates Brucella Pathogenesis and Leads to Altered Host Immune Response. Infect Immun 84:3458-3470
Gourley, Christopher R; Petersen, Erik; Harms, Jerome et al. (2015) Decreased in vivo virulence and altered gene expression by a Brucella melitensis light-sensing histidine kinase mutant. Pathog Dis 73:1-8
Durward-Diioia, Marina; Harms, Jerome; Khan, Mike et al. (2015) CD8+ T cell exhaustion, suppressed gamma interferon production, and delayed memory response induced by chronic Brucella melitensis infection. Infect Immun 83:4759-71
Splitter, Gary; Harms, Jerome; Petersen, Erik et al. (2014) Studying host-pathogen interaction events in living mice visualized in real time using biophotonic imaging. Methods Mol Biol 1197:67-85
Smith, Judith A; Khan, Mike; Magnani, Diogo D et al. (2013) Brucella induces an unfolded protein response via TcpB that supports intracellular replication in macrophages. PLoS Pathog 9:e1003785
Petersen, Erik; Rajashekara, Gireesh; Sanakkayala, Neelima et al. (2013) Erythritol triggers expression of virulence traits in Brucella melitensis. Microbes Infect 15:440-9
Magnani, Diogo M; Lyons, Elizabeth T; Forde, Toni S et al. (2013) Osteoarticular tissue infection and development of skeletal pathology in murine brucellosis. Dis Model Mech 6:811-8
Durward, Marina; Radhakrishnan, Girish; Harms, Jerome et al. (2012) Active evasion of CTL mediated killing and low quality responding CD8+ T cells contribute to persistence of brucellosis. PLoS One 7:e34925
Gupta, V K; Radhakrishnan, G; Harms, J et al. (2012) Invasive Escherichia coli vaccines expressing Brucella melitensis outer membrane proteins 31 or 16 or periplasmic protein BP26 confer protection in mice challenged with B. melitensis. Vaccine 30:4017-22

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