Amanda Robison

Flagella overexpression attenuates Salmonella pathogenesis.

Flagella are cell surface appendages involved in a number of bacterial behaviors, such as motility, biofilm formation, and chemotaxis. Despite these important functions, flagella can pose a liability to a bacterium when serving as potent immunogens resulting in the stimulation of the innate and adaptive immune systems. Previous work showing appendage overexpression, referred to as attenuating gene expression (AGE), was found to enfeeble wild-type Salmonella. Thus, this approach was adapted to discern whether flagella overexpression could induce similar attenuation. To test its feasibility, flagellar filament subunit FliC and flagellar regulon master regulator FlhDC were overexpressed in Salmonella enterica serovar Typhimurium wild-type strain H71. The results show that the expression of either FliC or FlhDC alone, and co-expression of the two, significantly attenuates Salmonella. The flagellated bacilli were unable to replicate within macrophages and thus were not lethal to mice. In-depth investigation suggests that flagellum-mediated AGE was due to the disruptive effects of flagella on the bacterial membrane, resulting in heightened susceptibilities to hydrogen peroxide and bile. Furthermore, flagellum-attenuated Salmonella elicited elevated immune responses to Salmonella presumably via FliC’s adjuvant effect and conferred robust protection against wild-type Salmonella challenge.

Recombinant Salmonella vaccination technology and its application to human bacterial pathogens.

Salmonella enterica is a Gram-negative intracellular bacterial pathogen which causes salmonellosis in humans and animals. During the past several decades, extensive studies have shown that the attenuated Salmonella vaccine vector is an optimal vehicle for delivering passenger antigens to mucosal sites to induce humoral, cellular, and mucosal immunity. This immunity leads to protection against challenges with the wild-type pathogens from which the passenger antigens were derived. A myriad of studies have demonstrated that using attenuated Salmonella vaccines for recombinant multivalent vaccine construction has multiple advantages. In this review, we summarize these advantages and further evaluate the Salmonella-based vaccines against five bacterial diseases. Four of these are Gram-negative pathogens- Escherichia coli, Helicobacter pylori, Shigella dysenteriae, and Yersinia pestis-and one is a mycobacterial pathogen, Mycobacterium tuberculosis. Apart from H. pylori, the Salmonella-based vaccines against the other four pathogens exhibit excellent performance in safety, immunogenicity, and protection. These properties qualify them to be as a new generation of vaccines for preventing infections from bacterial pathogens.

Roles of Toll-Like Receptor 2 (TLR2), TLR4, and MyD88 during Pulmonary Coxiella burnetii Infection

ABSTRACT Coxiella burnetii, the causative agent of Q fever, is an obligate intracellular, primarily pulmonary, bacterial pathogen. Although much is known about adaptive immune responses against this bacterium, our understanding of innate immune responses against C. burnetii is not well defined, particularly within the target tissue for infection, the lung. Previous studies examined the roles of the innate immune system receptors Toll-like receptor 2 (TLR2) and TLR4 in peripheral infection models and described minimal phenotypes in specific gene deletion animals compared to those of their wild-type controls (S. Meghari et al., Ann N Y Acad Sci 1063:161–166, 2005, http://dx.doi.org/10.1196/annals.1355.025; A. Honstettre et al., J Immunol 172:3695–3703, 2004, http://dx.doi.org/10.4049/jimmunol.172.6.3695) . Here, we assessed the roles for TLR2, TLR4, and MyD88 in pulmonary C. burnetii infection and compared responses to those that occurred in TLR2- and TLR4-deficient animals following peripheral infection. As observed previously, neither TLR2 nor TLR4 was needed for limiting bacterial growth after peripheral infection. In contrast, TLR2 and, to a lesser extent, TLR4 limited growth (or dissemination) of the bacterium in the lung and spleen after pulmonary infection. TLR2, TLR4, and MyD88 were not required for the general inflammatory response in the lungs after pulmonary infection. However, MyD88 signaling was important for infection-induced morbidity. Finally, TLR2 expression on hematopoietic cells was most important for limiting bacterial growth in the lung. These results expand on our knowledge of the roles for TLR2 and TLR4 in C. burnetii infection and suggest various roles for these receptors that are dictated by the site of infection.

Type I Interferon Counters or Promotes Coxiella burnetii Replication Dependent on Tissue

ABSTRACT Coxiella burnetii is an intracellular pathogen and the cause of Q fever. Gamma interferon (IFN-γ) is critical for host protection from infection, but a role for type I IFN in C. burnetii infection has not been determined. Type I IFN supports host protection from a related pathogen, Legionella pneumophila, and we hypothesized that it would be similarly protective in C. burnetii infection. In contrast to our prediction, IFN-α receptor-deficient (IFNAR−/−) mice were protected from C. burnetii-induced infection. Therefore, the role of type I IFN in C. burnetii infection was distinct from that in L. pneumophila. Mice treated with a double-stranded-RNA mimetic were protected from C. burnetii-induced weight loss through an IFNAR-independent pathway. We next treated mice with recombinant IFN-α (rIFN-α). When rIFN-α was injected by the intraperitoneal route during infection, disease-induced weight loss was exacerbated. Mice that received rIFN-α by this route had dampened interleukin 1β (IL-1β) expression in bronchoalveolar lavage fluids. However, when rIFN-α was delivered to the lung, bacterial replication was decreased in all tissues. Thus, the presence of type I IFN in the lung protected from infection, but when delivered to the periphery, type I IFN enhanced disease, potentially by dampening inflammatory cytokines. To better characterize the capacity for type I IFN induction by C. burnetii, we assessed expression of IFN-β transcripts by human macrophages following stimulation with lipopolysaccharide (LPS) from C. burnetii. Understanding innate responses in C. burnetii infection will support the discovery of novel therapies that may be alternative or complementary to the current antibiotic treatment.

Oral delivery of oligomeric procyanidins in Apple Poly® enhances type I IFN responses in vivo

Type I IFN signaling is a central pathway that provides critical innate protection from viral and bacterial infection and can have regulatory outcomes in inflammatory settings. We determined previously that OPCs contained in the dietary supplement APP enhanced responses to type I IFN in vitro. Here, we confirm that OPCs from two different sources significantly increased pSTAT1, whereas a monomeric form of procyanidin did not. We hypothesized that similar responses could be induced in vivo following ingestion of APP. Ingestion of APP before injection of polyI:C enhanced in vivo responses to type I IFNs in mice. When human subjects ingested APP, enhanced responses to type I IFN and enhanced pSTAT1 ex vivo were detected, whereas ingestion of RES, a monomeric polyphenol, induced minimal such changes. Polyphenols are best known for induction of anti‐inflammatory and antioxidant responses; however, our findings suggest a unique, nonantioxidant aspect of OPCs that is broadly applicable to many disease settings. The capacity of oral OPCs to enhance type I IFN signaling in vivo can augment innate protection and may, in part, contribute to the noted anti‐inflammatory outcome of ingestion of OPCs from many sources.