Research in the Division currently centers on three diseases: tuberculosis, Lyme disease, and relapsing fever.
Tuberculosis
Tuberculosis (TB), along with AIDS and malaria, is one of the three major infectious disease killers. One third of the world's population is infected with Mycobacterium tuberculosis (Mtb). Each year an estimated ten million people develop, and about two million people die of, TB. The TNPRC has developed a highly visible research program in TB using NHPs, specifically macaques. Key aspects of our model system are the natural route of infection with Mtb and the ability to model TB/AIDS comorbidity using SIV as a surrogate for HIV. Using this model we are currently studying the following various aspects of disease and attempting to develop therapeutics and vaccines against the infection.
1. Understanding the response of Mtb to in vivo stress: Understanding bacterial gene expression in vivo is central to unraveling mechanisms of disease caused by intracellular bacteria such as Mtb. We are specifically interested in the intersection of the pathways governed by stress-response transcription factors SigH, DosR and ClgR and their role in potentiating the survival and persistence of Mtb in vivo. It is hypothesized that these regulatory genes play a crucial role in the process of adaptation of M. tuberculosis to the intracellular milieu, and hold the secret of this bacterium's virulence. This work is funded by the NIH (R01AI089323) and the Wetmore Foundation. Suggested reading: Veatch AV and Kaushal D. Opening Pandora’s box: Mechanisms of Mycobacterium tuberculosis resuscitation. Trends in Microbiology. pii: S0966-842X(17)30191-9. doi: 10.1016/j.tim.2017.08.001.
2. Understanding the role of inducible bronchus associated lymphoid tissue in mediating granuloma level protection from TB: iBALT contains spatially organized T cells, B cells and macrophages and its presence is associated with better protective outcomes during TB. Additionally, a dominant feature of the granulomatous inflammation is the accumulation of neutrophils that produce inflammatory molecules such as S100A8/A9 proteins. Incidentally, increased neutrophil accumulation has also been associated with increased Mtb and viral burden in TB/HIV co-infected patients. Based on these new data, we are testing a paradigm-shifting hypothesis that a protective TB granuloma is one that contains iBALT and contributes to Mtb containment during LTBI. In contrast, progression to a more neutrophilic, inflammatory granuloma causes TB reactivation, loss of Mtb control and progression to PTB. We are mechanistically identifying Mtb genes and pathways that modulate iBALT formation, providing crucial new information about the mechanism(s) employed by Mtb in interfering with the formation of protective iBALTs. We are also addressing the function of persistent iBALT in limiting reactivation and dissemination in the NHP model of TB/SIV co-infection. Finally, we are addressing whether iBALT structures can be enhanced, or neutrophilic granulomas reversed, to decrease TB reactivation and disease severity during latency and SIV co-infection. This work is primarily funded by the NIH (R01AI134240 and R01AI111914). Suggested reading: Kaushal D, Foreman TW, Gautam US, Alvarez X, Adekambi T, Rangel-Moreno J, Golden NA, Kissee R, Johnson-May AF, Phillips BL, Ahsan MH, Roy CJ, Didier PJ, Doyle LA, Russell-Lodrigue KE, Blanchard JL, Rengarajan J, Lackner AA, Khader SA, Mehra S. 2015. Mucosal vaccination with attenuated Mycobacterium tuberculosis induces strong central memory responses and protects against tuberculosis. Nature Communications. 6:8533. PMID: 26460802. PMCID: PMC4608260 (accompanied by NIH, Tulane and Nature press releases, along with other press coverage available on the web).
3. Identifying classically activated T cell based antigen-specific responses associated with protection vs. disease. This aim encompasses three major NIH grants (R01AI111943, R01AI123047 and U19AI111211). We are focusing on the mechanisms by which Mtb-specific T cell responses maintain LTBI, confer protective immunity or result in HIV-induced reactivation of LTBI. We hypothesize that co-infection with HIV depletes and/or impairs in the functional capacities of Mtb-specific CD4 and CD8 T cells to drive reactivation of LTBI and that antiretroviral therapy (ART) only partially restores these functions. We propose to test this hypothesis using mechanistic experiments in the nonhuman primate model of inhalation TB. We are also focusing on the identity of specific T cell responses that direct the control of Mtb in the LTBI state, or are responsible for reactivation. We propose to identify the immune signatures associated with control of Mtb infection as LTBI either spontaneously or upon therapy, in macaques. Suggested reading: Foreman TW*, Mehra S*, LoBato DN, Malek A, Alvarez X, Golden NA, Bucsan AN, Didier PJ, Doyle-Meyers LA, Russell-Lodrigue KE, Roy CJ, Blanchard JL, Kuroda MJ, Lackner AA, Chan J, Khader SA, Jacobs WR, Jr., Kaushal D. CD4+ T-cell-independent mechanisms suppress reactivation of latent tuberculosis in a macaque model of HIV co-infection. Proceedings of the National Academy of Sciences USA. 113:E5636-44 PMID: 27601645. PMCID: PMC5 035858. (accompanied by Tulane and NAS press releases, along with other press coverage available on the web).
4. CD4 T cell independent mechanisms of protection from TB and HIV-induced reactivation of TB. Here we are using the macaque model of TB and of Mtb/SIV co-infection to study a variety of immune components, such as CD8 T cells, macrophages, NK cells, and innate lymphoid cells, especially ILC3 cells, and mucosal associated invariant T (MAIT) cells. This work is supported by R01AI134236, and the Wetmore Foundation.
5. Identification of biomarkers of the various stages of disease and infection. We are using transcriptomic, miRNA, epigenetic and proteomic approaches to define active vs. latent TB as well as reactivation of latent TB due to SIV (as a surrogate for HIV) co-infection. We are also interested in better defining the immunological mechanism of SIV-mediated reactivation of latent TB in primate lungs. This work is supported by R01HL106790, R01AI123780 and the Wetmore Foundation.
6. Host-directed approaches to better treat TB. We are testing several host-directed approaches. Of these, one (Imatinib) has shown promising preclinical results adjunctive to chemotherapy and is now included in an NIH-supported clinical trial. Another has shown promise as a stand-alone reagent and is now being tested by Dr. Mehra adjunctive to chemotherapy. Agents targeting multiple pathways such as AMPK, NAD+ and SIRT are under investigation. These studies are supported by UH2AI122320 and UH3AI122320. Suggested reading: Gautam US, Foreman TW, Bucsan AN, Veatch AV, Gentry KM, Golden NA, Alvarez X, Adekambi T, Doyle-Myers LA, Russell-Lodrigue KE, Didier PJ, Kousoulas KG, Kalman D, Lackner AA, Rengarajan J, Khader SA, Kaushal D, Mehra S. Inhibition of tryptophan catabolism reorganizes the tuberculoma and augments immune-mediated control of Mycobacterium tuberculosis. Proceedings of the National Academy of Sciences USA. 115(1):E62-E71. PMID: 29255022.
7. Development of attenuated vaccines against TB. We are focused on the development of an extremely safe vaccine based on stress-response attenuation. Our focus is to develop vaccines that can be mucosally delivered, elicit potent lung T and B cell responses and would be safe in the HIV+ population. Additional vaccination approaches being applied include MAPS, and viral vectors. This work is supported by the Bill and Melinda Gates Foundation, AERAS, and the NIH grant R01AI135729. Suggested reading: Foreman TW, Veatch AV, LoBato DN, Didier PJ, Doyle-Meyers LA, Russell-Lodrigue KE, Lackner AA, K. Gus Kousoulas, Khader SA, Kaushal D, Mehra S. Non-pathological infection of macaques by an attenuated mycobacterial vaccine is not reactivated in the setting of HIV coinfection. American Journal of an attenuated mycobacterial vaccine is not reactivated in the setting of HIV coinfection. American Journal of Pathology. pii: S0002-9440(17)30363-2.
Lyme Disease
Lyme disease, or Lyme borreliosis is an emerging infectious disease that affects people in North America, Europe, and Asia. Lyme borreliosis is caused by the spirochete Borrelia burgdorferi and is transmitted by ticks. The disease may manifest in numerous ways, e.g. as an inflammation of the skin (erythema migrans), as arthritis, and in the peripheral and central nervous systems as, for example, facial paralysis and through neurocognitive symptoms. Lyme disease that affects the nervous system is also called Lyme neuroborreliosis. After developing a nonhuman primate model of Lyme disease, divisional faculty are currently using this model to try to understand the pathogenesis of neuroborreliosis of the central nervous system.
1) One major finding was that inflammation caused by B. burgdorferi is the key factor in neuroborreliosis pathogenesis. Suggested reading. Ramesh G, Didier PJ, England JD, Santana-Gould L, Doyle-Meyers LA, Martin DS, Jacobs MB, Philipp MT. Inflammation in the pathogenesis of Lyme neuroborreliosis. Am J Pathol. 2015 May;185(5):1344-60. doi: 10.1016/j.ajpath.2015.01.024. Epub 2015 Apr 16. PMCID: PMC4419210. Supported by NIH grant R01-NS048952.
Delving deeper into the mechanisms mediating inflammation by B. burgdorferi we determined that MAP kinase pathways were crucial to effecting inflammation and apoptosis in cells of the CNS, and that the neurokinin Substance P accentuates the inflammatory response caused by B. burgdorferi in the CNS in vitro, ex vivo, and in vivo. Suggested reading. Parthasarathy G, Philipp MT. The MEK/ERK pathway is the primary conduit for Borrelia burgdorferi-induced inflammation and P53-mediated apoptosis in oligodendrocytes. Apoptosis. 2014 Jan;19(1):76-89. doi:10.1007/s10495-013-0913-8. PubMed Central PMCID: PMC3947362. Martinez AN, Ramesh G, Jacobs MB, Philipp MT. Antagonist of the neurokinin-1 receptor curbs neuroinflammation in ex vivo and in vitro models of Lyme neuroborreliosis. J Neuroinflammation. 2015 Dec 30;12:243. doi: 10.1186/s12974-015-0453-y; PubMed Central PMCID: PMC4696197. Martinez AN, Burmeister AR, Ramesh G, Doyle-Meyers L, Marriott I, Philipp MT. Aprepitant limits in vivo neuroinflammatory responses in a rhesus model of Lyme neuroborreliosis. J Neuroinflammation. 2017 Feb 15;14(1):37. doi: 10.1186/s12974-017-0813-x; PubMed Central PMCID. PMC5312540. Supported by NIH grant R01 NS 050325.
Improvement of methods for the serological diagnosis of Lyme disease is also of major interest to divisional faculty. The significance of this line of research stems from the fact that Lyme disease that is accurately diagnosed early in the course of infection is more efficaciously treated with antibiotics than when the disease remains undiagnosed for extended periods. The C6 test, an antibody detection assay that is more sensitive and specific than the tests available heretofore, was developed by investigators in the Division. An additional advantage is that the test does not yield false-positive results with serum specimens from humans or dogs that have received Lyme disease vaccination. The test was approved by the FDA and the USDA for human and animal use, respectively, and licensed by Tulane University to Immunetics, Inc., of Boston, MA, for human use, and to IDEXX Laboratories, Inc. of Westbrook, ME, for veterinary purposes. Currently, research on diagnosis is focused on the use of a multiplex platform for Lyme disease diagnosis at all phases of disease and for gauging treatment response. In a related project, collaborative research is aimed at identification of Borrelia burgdorferi diagnostic biomarkers in humans and nonhuman primates, focused on early detection of B. burgdorferi proteins in blood and/or body fluids, prior to the generation of specific antibodies by infected individuals. Divisional faculty and collaborators are also engaged in the identification of the molecule(s) present in tick saliva that attract B. burgdorferi to the feeding tick. Suggested reading: M. E. Embers, N.R. Hasenkampf, M.B. Barnes, E. S. Didier, M. T. Philipp and A.C.Tardo. A Five-Antigen Fluorescent Bead-based Assay for Diagnosis of Lyme Disease. Clinical and Vaccine Immunology. 2016 Apr 4;23(4):294-303. doi: 10.1128/CVI.00685-15. Embers ME, Hasenkampf NR, Jacobs MB, and Philipp MT. Dynamic Longitudinal Antibody Responses during Borrelia burgdorferi Infection and Antibiotic Treatment of Rhesus Macaques. Clinical and Vaccine Immunology 2012, 19(8):1218.
This work is supported by grants R33AI100166, R41AI114049 and R21AI121894 from NIH/NIAID
With respect to treatment of Lyme disease, definitive experiments on the persistence of B. burgdorferi post-antibiotic treatment in the NHP have been conducted by researchers in the Division. Current projects aimed at improving treatment include:(1) In vivo testing of antibiotic efficacy against B. burgdorferi spirochetes that have been adapted to the host; (2) evaluation of combination antibiotic therapy against B. burgdorferi infection in animal models; (3) resuscitation of viable, non-cultivable B. burgdorferi treated with antibiotics to evaluate the viability of spirochetes that persist in treated animals; and (4) the discovery and evaluation of novel small molecule inhibitors targeting Borrelia and Bartonella for antimicrobial efficacy. Suggested reading: M. E. Embers, N.R. Hasenkampf, M.B. Jacobs, A.C. Tardo, N.A. Crossland, L.A. Doyle-Meyers, M.T. Philipp and E. Hodzic. Variable Manifestations, Diverse Seroreactivity and Post-treatment Persistence in Nonhuman Primates Exposed to Borrelia burgdorferi by Tick Feeding. 2017. PLOS ONE. December 13, 2017 https://doi.org/10.1371/journal.pone.0189071 N.A. Crossland, X. Alvarez, and M.E. Embers. Late Disseminated Lyme Disease: Associated Pathology and Spirochete Persistence Post-Treatment in Rhesus Macaques. 2017. American Journal of Pathology. http://dx.doi.org/10.1016/j.ajpath.2017.11.005 Caskey JR and ME Embers. Persister Development by B. burgdorferi Populations In Vitro. Antimicrobial Agents and Chemotherapy, 2015 Oct;59(10):6288-95. doi: 10.1128/AAC.00883-15. Epub 2015 Jul 27. Embers ME, Barthold SW, Borda JT, Bowers L, Doyle LA, Hodzic E, Jacobs MB, Hasenkampf NR, Martin DS, Narasimhan S, Phillippi-Falkenstein KM, Purcell JE, Ratterree MS, and Philipp MT. Persistence of Borrelia burgdorferi in rhesus macaques following antibiotic treatment of disseminated infection. PLoS One. 2012;7(1):e29914. Epub 2012 Jan 11. PMID: 22253822.
This work is supported by Global Lyme Alliance, the Bay Area Lyme Foundation and the Steven and Alexandra Cohen Foundation.
Relapsing Fever
Relapsing Fever (RF) borreliosis is a neglected disease that is present in 5 of 7 continents, and infection with the spirochete pathogen is particularly devastating for fetal and neonatal health. Key elements of human disease, such as multiple febrile episodes, high levels of spirochetemia, and cardiac impairment are not recapitulated in mice. With a collaborative research project, Division faculty developed a nonhuman primate model of RF borreliosis that closely resembles human disease. The aim for subsequent study is to use this model to evaluate diagnostic and vaccine candidates, with the goal of improving maternal and fetal health worldwide. Suggested reading: Lopez JE, Oliphant HV, Wilder HK, Grasperge BJ, Morgan TW, Stuckey KJ and ME Embers. Real-Time Monitoring of Disease in Rhesus Macaques Infected by Tick Bite with Borrelia turicatae. 2014. Journal of Infectious Diseases. Nov 15;210(10):1639-48
