Mucosal Immunization with Iron Receptor Antigens Protects against Urinary Tract Infection

Uncomplicated infections of the urinary tract, caused by uropathogenic Escherichia coli, are among the most common diseases requiring medical intervention. A preventive vaccine to reduce the morbidity and fiscal burden these infections have upon the healthcare system would be beneficial. Here, we describe the results of a large-scale selection process that incorporates bioinformatic, genomic, transcriptomic, and proteomic screens to identify six vaccine candidates from the 5379 predicted proteins encoded by uropathogenic E. coli strain CFT073. The vaccine candidates, ChuA, Hma, Iha, IreA, IroN, and IutA, all belong to a functional class of molecules that is involved in iron acquisition, a process critical for pathogenesis in all microbes. Intranasal immunization of CBA/J mice with these outer membrane iron receptors elicited a systemic and mucosal immune response that included the production of antigen-specific IgM, IgG, and IgA antibodies. The cellular response to vaccination was characterized by the induction and secretion of IFN-γ and IL-17. Of the six potential vaccine candidates, IreA, Hma, and IutA provided significant protection from experimental infection. In immunized animals, class-switching from IgM to IgG and production of antigen-specific IgA in the urine represent immunological correlates of protection from E. coli bladder colonization. These findings are an important first step toward the development of a subunit vaccine to prevent urinary tract infections and demonstrate how targeting an entire class of molecules that are collectively required for pathogenesis may represent a fundamental strategy to combat infections

Direct Activation of STING in the Tumor Microenvironment Leads to Potent and Systemic Tumor Regression and Immunity

SummarySpontaneous tumor-initiated T cell priming is dependent on IFN-β production by tumor-resident dendritic cells. On the basis of recent observations indicating that IFN-β expression was dependent upon activation of the host STING pathway, we hypothesized that direct engagement of STING through intratumoral (IT) administration of specific agonists would result in effective anti-tumor therapy. After proof-of-principle studies using the mouse STING agonist DMXAA showed a potent therapeutic effect, we generated synthetic cyclic dinucleotide (CDN) derivatives that activated all human STING alleles as well as murine STING. IT injection of STING agonists induced profound regression of established tumors in mice and generated substantial systemic immune responses capable of rejecting distant metastases and providing long-lived immunologic memory. Synthetic CDNs have high translational potential as a cancer therapeutic

Waging War against Uropathogenic Escherichia coli: Winning Back the Urinary Tract▿

Urinary tract infection (UTI) caused by uropathogenic Escherichia coli (UPEC) is a substantial economic and societal burden—a formidable public health issue. Symptomatic UTI causes significant discomfort in infected patients, results in lost productivity, predisposes individuals to more serious infections, and usually necessitates antibiotic therapy. There is no licensed vaccine available for prevention of UTI in humans in the United States, likely due to the challenge of targeting a relatively heterogeneous group of pathogenic strains in a unique physiological niche. Despite significant advances in the understanding of UPEC biology, mechanistic details regarding the host response to UTI and full comprehension of genetic loci that influence susceptibility require additional work. Currently, there is an appreciation for the role of classic innate immune responses—from pattern receptor recognition to recruitment of phagocytic cells—that occur during UPEC-mediated UTI. There is, however, a clear disconnect regarding how factors involved in the innate immune response to UPEC stimulate acquired immunity that facilitates enhanced clearance upon reinfection. Unraveling the molecular details of this process is vital in the development of a successful vaccine for prevention of human UTI. Here, we survey the current understanding of host responses to UPEC-mediated UTI with an eye on molecular and cellular factors whose activity may be harnessed by a vaccine that stimulates lasting and sterilizing immunity

Protease Activity, Secretion, Cell Entry, Cytotoxicity, and Cellular Targets of Secreted Autotransporter Toxin of Uropathogenic Escherichia coli

The secreted autotransporter toxin (Sat), found predominantly in uropathogenic Escherichia coli, is a member of the SPATE (serine protease autotransporters of Enterobacteriaceae) family and, as such, has serine protease activity and causes cytopathic effects on various cell types. To assess the contribution of the serine protease active site to the mechanism of action of Sat, mutations were made in the first (S256I), in the second (S258A), or in both (S256I/S258A) serine residues within the active site motif. Mutations in the first or both serines reduced protease activity to background levels (P < 0.001); a single mutation in the second serine reduced activity by 60% compared to wild type (P < 0.001). After reversion of the S256I mutation to wild type (I256S), we confirmed S(256) as the catalytically active serine. None of these mutations affected secretion of the mature passenger domain or release into the supernatant. The S256I mutation, however, abrogated the cytotoxicity of Sat on human bladder (UM-UC-3) and kidney (HEK 293) epithelial cells, characterized by rounding and elongation, respectively, and a high level of cell detachment. Moreover, S(256) is essential for Sat to mediate cytoskeletal contraction and actin loss in host cells as well as to degrade specific membrane/cytoskeletal (fodrin and leukocyte function-associated molecule 1) and nuclear [microtubule-associated proteins, LIM domain-only protein 7, Rap GTPase-activating protein, poly(ADP-ribose) polymerase] proteins in vitro. Lastly, Sat was internalized by host cells and localized to the cytoskeletal fraction where membrane/cytoskeletal target proteins reside

Toll-like Receptor-Deficient Mice Reveal How Innate Immune Signaling Influences Salmonella Virulence Strategies

Pathogens utilize features of the host response as cues to regulate virulence gene expression. Salmonella enterica serovar Typhimurium (ST) sense Toll-like receptor (TLR)-dependent signals to induce Salmonella Pathogenicity Island 2 (SPI2), a locus required for intracellular replication. To examine pathogenicity in the absence of such cues, we evaluated ST virulence in mice lacking all TLR function (Tlr2(-/-)xTlr4(-/-)xUnc93b1(3d/3d)). When delivered systemically to TLR-deficient mice, ST do not require SPI2 and maintain virulence by replicating extracellularly. In contrast, SPI2 mutant ST are highly attenuated after oral infection of the same mice, revealing a role for SPI2 in the earliest stages&nbsp;of infection, even when intracellular replication is not required. This early requirement for SPI2 is abolished in MyD88(-/-)xTRIF(-/-) mice lacking both TLR- and other MyD88-dependent signaling pathways, a potential consequence of compromised intestinal permeability. These results demonstrate how pathogens use plasticity in virulence strategies to respond to different host immune environments

Toll-like Receptor-Deficient Mice Reveal How Innate Immune Signaling Influences Salmonella Virulence Strategies

Pathogens utilize features of the host response as cues to regulate virulence gene expression. Salmonella enterica serovar Typhimurium (ST) sense Toll-like receptor (TLR)-dependent signals to induce Salmonella Pathogenicity Island 2 (SPI2), a locus required for intracellular replication. To examine pathogenicity in the absence of such cues, we evaluated ST virulence in mice lacking all TLR function (Tlr2(-/-)xTlr4(-/-)xUnc93b1(3d/3d)). When delivered systemically to TLR-deficient mice, ST do not require SPI2 and maintain virulence by replicating extracellularly. In contrast, SPI2 mutant ST are highly attenuated after oral infection of the same mice, revealing a role for SPI2 in the earliest stages&nbsp;of infection, even when intracellular replication is not required. This early requirement for SPI2 is abolished in MyD88(-/-)xTRIF(-/-) mice lacking both TLR- and other MyD88-dependent signaling pathways, a potential consequence of compromised intestinal permeability. These results demonstrate how pathogens use plasticity in virulence strategies to respond to different host immune environments