UC Berkeley

Mycobacterium tuberculosis (Mtb) is an intracellular bacterial pathogen and causative agent of the disease tuberculosis. Mtb establishes infection in the lungs via inhalation of aerosol droplets containing the bacteria. Approximately a quarter of the world's population is latently infected with Mtb. Control of Mtb infection is dependent on the adaptive immune response to Mtb infection, however which cell types and immune pathways are necessary for controlling Mtb infection remains unclear. The scientific community has identified type 1 CD4+ T helper cells (Th1) and their key cytokine interferon gamma (IFN-γ) as important for controlling infection, yet infected humans will eventually lose control of Mtb infection and develop active tuberculosis disease while Th1s are present in the lungs. There is growing evidence in human studies, non-human primate infection models, and murine tuberculosis vaccine models that type 17 CD4+ T helper cells (Th17s) can promote control of Mtb infection, but Th17s are not always induced in Mtb infected humans and never induced in unvaccinated mice. The mechanism(s) by which Mtb-infected hosts induce Th1 and/or Th17 responses remain unclear. This highlights the necessity for understanding what are the controlling factors directing Th1 and Th17 development during Mtb infection. A more complete understanding of Th1 and Th17 development and their roles during Mtb infection will improve Mtb vaccine development which is required for eliminating the tuberculosis pandemic worldwide. Here, we demonstrate that mice forced into a Th17 adaptive immune response are protected against Mtb infection mediated by the key cytokine produced by Th17s, IL-17a. We found that Mtb uses the type VII secretion system ESX-1 and lipid virulence factor phthiocerol dimycocerosate (PDIM) to promote infection by preventing the development of Th17s. ESX-1 and PDIM are known to be essential for the detrimental type I interferon response during Mtb infection, although the type I interferon response is not responsible for the inhibition of Th17 development. ESX-1 and PDIM alter conventional dendritic cells in the draining mediastinal lymph node to promote expression of Th1-polarizing cytokine IL-12 and hinder expression of Th17-polarizing cytokine IL-23. These results indicate that Mtb uses the type VII secretion system ESX-1 and lipid virulence factor PDIM to force the host into a limiting but permitting Th1 adaptive immune response by preventing the development of protective Th17 cells. Separately, Staphylococcus aureus (S. aureus) is an extracellular bacterial pathogen known to cause skin and soft tissue infections. S. aureus is a member of the ESKAPE pathogens known for their increasing frequency of antibiotic resistance and occurrence of infection in hospitals. The more pathogenic and antibiotic-resistant version of S. aureus, methicillin-resistant S. aureus (MRSA) causes approximately 300,000 hospitalizations and 10,000 deaths every year in the United States of America alone. MRSA contains an arsenal of virulence factors used to cause disease, however only about two-thirds of the pangenome of S. aureus remains consistent making virulence factor-targeted therapy difficult to design as the frequency of S. aureus infections being MRSA increases and the ability to use antibiotics to treat MRSA infections declines. A more comprehensive understanding of all the virulence factors used by MRSA to promote infection is required for designing novel therapeutics to treat this dangerous pathogen. Herein, we performed a surface and secreted proteome mass spectrometry experiment and identified 21 potential hits with unknown protein function. Utilization of a genome-wide arrayed transposon library of MRSA to infect WT C57BL/6 bone marrow-derived macrophages revealed 6 out of the 21 hits with some difference in virulency compared to WT MRSA. 2 of the 6 potential virulence factors (SAUSA300_1739 and SAUSA300_1740) with unknown protein functions were located next to each other in the MRSA genome. These 2 genes are located in an operon of 6 genes (SAUSA300_1739 to SAUSA300_1744), which we show are transcribed together as an operon of 6 genes. Chromosomal knockouts of any of these 6 genes did not impair growth in broth, however each knockout was attenuated in a murine subcutaneous infection model. Genetic analysis of the SAUSA300_1739 to SAUSA300_1744 operon in several S. aureus strains indicate that this operon is highly conserved amongst S. aureus strains. And finally, we identified the protein function of SAUSA300_1739 and SAUSA300_1740 as DNases. These results reveal that MRSA uses this operon to promote infection and expands our knowledge of the arsenal of virulence factors used by MRSA.