Infection and Immunity最新文献

Chlamydia trachomatis is the leading cause of bacterial sexually transmitted infection in the United States. The high rate of asymptomatic cases and absence of a vaccine often leave infections untreated, increasing the risk of serious complications in women, like pelvic inflammatory disease, ectopic pregnancy, and infertility. The generation of C. trachomatis mutants is crucial for studying C. trachomatis gene function, identifying potential vaccine candidates, and understanding host-pathogen interactions. However, the obligate intracellular nature of the bacteria hinders the development of genetic tools for mutagenesis. Counterselectable markers are effective systems for selecting bacterial mutants; however, such systems have yet to be optimized for use in C. trachomatis. In this study, we created a toxin-antitoxin (TA) system-based counterselection marker. Two TA systems were tested, toxin CcdB and its antitoxin CcdA (CcdAB), and toxin MvpT and its antitoxin MvpA (MvpAT). For each system, the antitoxin was expressed from a constitutive promoter, while the toxin was controlled by an inducible promoter. We first showed that, in Escherichia coli, toxin induction in both TA systems overcame the protective effect of the antitoxin, resulting in growth inhibition. The two systems were subsequently tested in C. trachomatis. While the CcdAB system did not significantly inhibit the growth of C. trachomatis, the MvpAT system did. Altogether, we have developed an MvpAT-based counterselection system for use in C. trachomatis. Implementation of this system will enable more efficient genetic manipulation, facilitating the identification of bacterial virulence factors and advancing translational research toward improved treatment and prevention.

Bacteroides fragilis is an important member of the human gut microbiota, where it contributes to immune modulation, intestinal barrier integrity, and colonization resistance. Despite its beneficial roles as a symbiont in the gut, B. fragilis is also the most commonly isolated anaerobe in clinical infections, implicated in intra-abdominal abscesses, bloodstream infections, and soft tissue infections. Antimicrobial resistance (AMR) is increasingly recognized as a major factor in its transition from symbiont to opportunist; however, the relationship between resistance and anatomical site of isolation remains poorly defined. Here, we compared AMR phenotypes and genotypes between intestinal and extra-intestinal B. fragilis isolates to assess whether clinical strains are enriched for resistance determinants. Surprisingly, we found comparable susceptibility profiles and AMR gene content between the two groups. Minimal inhibitory concentrations (MICs) were broadly similar, and β-lactamase activity was detected in ~70% of the isolates regardless of the isolation site. We found that resistance genes were similarly distributed across both intestinal and clinical strains. A microbial genome-wide association study (mGWAS) confirmed the known resistance markers, such as ermF, aadS, and tetQ, and identified novel associations with conjugative transposons, efflux transporters, regulatory genes, and previously uncharacterized loci. These findings suggest that intestinal strains serve as a reservoir of clinically relevant resistance determinants that may be mobilized under selective pressure. Although prior work has largely focused on clinical isolates, our findings highlight the need to surveil AMR within the gut microbiota, where widespread resistance in commensal bacteria has the potential to complicate treatment of extra-intestinal infections.

 Chlamydia trachomatis (C.t.), the leading bacterial cause of sexually transmitted infections, replicates within a unique intracellular compartment called the inclusion, which is modified by secreted proteins known as inclusion membrane (Inc) proteins. Here, we further characterize CpoS, an Inc protein previously shown to be critical for bacterial replication and inclusion development. We demonstrate that CpoS directly binds multiple coiled-coil region-containing Incs and engages Rab GTPases at a separate site. Notably, CpoS-InaC interactions facilitate the recruitment of select Arf GTPases to the inclusion membrane, while Rab recruitment occurs independently of these interactions. Biochemical and biophysical analyses revealed that Incs self-oligomerize to form higher-ordered structures, with CpoS adopting a tetrameric conformation resembling that of eukaryotic SNARE proteins. We propose that these assemblies serve as scaffolds to orchestrate vesicle docking, tethering, and fusion. Our findings highlight the intricate interplay between bacterial and host factors, revealing how C.t. leverages both Inc-Inc interactions and host protein engagement to manipulate vesicular trafficking and sustain infection.

A key challenge in immunology is understanding how the innate immune system achieves specificity against diverse pathogens. Our previous work in Caenorhabditis elegans identified NMUR-1, a neuronal G protein-coupled receptor homologous to mammalian neuromedin U receptors, as a regulator of pathogen-specific innate immune responses. Here, we used quantitative proteomics and functional analyses to show that NMUR-1 modulates the expression of mitochondrial F₁F_O ATP synthase subunits and regulates ATP levels during infection, linking neuronal signaling to host energy metabolism. Loss of NMUR-1 leads to reduced ATP and reactive oxygen species (ROS) concentrations in infected animals, altering survival outcomes in a pathogen-specific manner. We further demonstrate that ATP availability and its contribution to host defense are neurally controlled by the NMUR-1 ligand CAPA-1 and its source neurons, ASG. These findings uncover a neuroimmune mechanism whereby NMUR-1 regulates energy homeostasis as a determinant of innate immune specificity. Our study also provides mechanistic insights into the emerging roles of conserved NMU signaling in neuroimmune regulation across animal phyla.

Candida auris is an emerging fungal pathogen recognized among the Centers for Disease Control and Prevention's urgent threats and designated a critical priority by the World Health Organization due to its global spread, high mortality rates, potential for pan-drug resistance, and its persistent transmission within healthcare settings. The clinical management of C. auris infections is further hindered by the lack of both rapid/specific diagnosis and effective antifungals. These challenges emphasize the urgent need for alternative therapeutic strategies. In this study, we investigated the antifungal, immunomodulatory, and protective effects of engineered Lectin-Fc(IgG) fusion proteins against a fluconazole-resistant C. auris strain. Specifically, Dectin-1-Fc(IgG2a), Dectin-1-Fc(IgG2b), and wheat germ agglutinin (WGA)-Fc(IgG2a) demonstrated dose-dependent binding to key fungal cell wall components, β-1,3-glucan and chitin, with Dectin-1-Fc(IgG2b) exhibiting the highest reactivity, followed by Dectin-1-Fc(IgG2a) and WGA-Fc(IgG2a). In vitro, all constructs exhibited fungistatic activity and reduced the biofilm biomass and metabolism, with the Dectin-1-Fc(IgG) variants displaying the most potent effects. As opsonins, Lectin-Fc(IgG)s significantly enhanced the macrophage-yeast association and macrophage-mediated killing of C. auris. In a systemic murine C. auris infection model, a single therapeutic administration of Dectin-1-Fc(IgG2b) or WGA-Fc(IgG2a) conferred 100% protection, while Dectin-1-Fc(IgG2a) achieved >80% protection, with all treated mice manifesting clinical improvement. Quantification of fungal burden at day 7 post-infection revealed at least a ~1 log reduction in colony-forming units in the spleen, kidney, and liver of Lectin-Fc(IgG)-treated animals. Cytokine profiling indicated a Th1-type-skewed immune response in Lectin-Fc(IgG)-treated mice. Collectively, these findings support the antifungal and immunotherapeutic potential of Lectin-Fc(IgG)s against C. auris, offering a novel broad-spectrum strategy to overcome current therapeutic limitations.

Current vaccine development efforts encompass diverse and innovative approaches; however, studies with basic vaccines (e.g., whole cell inactivated) continue to inform these efforts. Perhaps the world's most effective and durable human vaccine, the query (Q) fever vaccine known as Q-VAX, can trigger a severe post-vaccination hypersensitivity reaction, precluding its widespread deployment. The history of Q fever vaccine development serves as a rich example of the power of scientific collaboration, elegant experimentation, and the study of adverse post-vaccination events. Here, we review the history of Q fever vaccine development, profiling seminal studies while also relating this information to modern vaccine development efforts.

Rocky Mountain spotted fever (RMSF) caused by Rickettsia rickettsii is the most fatal tick-borne disease in people and dogs in the Americas. This pathogen is transmitted by several hard ticks: Dermacentor species, Rhipicephalus sanguineus, and Amblyomma americanum. RMSF can quickly progress to a life-threatening illness with fatalities ranging from 30% to 80%. Doxycycline is the only treatment option, and currently, no methods are available to prevent RMSF. We previously reported that vaccination with R. rickettsii whole cell antigen vaccine (WCAV) with Montanide gel adjuvant offers protection against virulent R. rickettsii infection in dogs. Here, we compared three adjuvants to optimize the safety and immunogenicity of WCAV: Alhydrogel, Montanide, and Quil A. Independent of adjuvants, all vaccinees were protected, whereas unvaccinated dogs developed the clinical disease. Vaccination reduced the pathogen to undetectable levels in blood and various tissues. An R. rickettsii-specific IgG response was observed following primary vaccination in all vaccinated groups, which augmented after booster. The vaccine with Quil A had the highest IgG response with a significant rise in CD4⁺ and CD8⁺ T cell numbers, while Montanide and Alhydrogel resulted in a balanced IgG response. IgG2 was the primary antibody detected in unvaccinated infection controls. Several systemic proinflammatory cytokines varied after infection in both vaccinees and controls. Plasma concentration of intercellular adhesion molecule 1 was higher in unvaccinated compared to vaccinees in the first 9 days after infection. This study demonstrates that the WCAV efficacy is independent of the adjuvant, although Quil A induced a higher IgG response and expansion of CD4 and CD8 T cells.

Klebsiella pneumoniae (K. pneumoniae), a common nosocomial pathogen causing severe pulmonary infections, is often complicated by coinfections. Bacterial ghosts, which are empty bacterial cell envelopes, hold significant promise as vaccine adjuvants. This study aims to develop and evaluate a novel combination vaccine platform utilizing K. pneumoniae ghosts (KP ghosts) to explore their intrinsic immunogenic properties as both vaccine and natural adjuvants. In this study, we showed that KP ghosts enhanced maturation and activation of bone marrow-derived dendritic cells, increasing surface markers (CD40, CD80, CD86, and MHC II) and cytokine secretion (IL-1β, TNF-α, and IL-12p70). The KP ghost-based vaccine provided strong immune protection in mice, significantly improving survival rates and reducing bacterial loads in organs after bacterial challenge. Additionally, to assess the adjuvant potential of KP ghosts, C57BL/6 mice were co-immunized with KP ghosts and a model antigen, ovalbumin (OVA). In comparison to OVA alone, the combination of OVA and KP ghosts elicited higher levels of specific IgG antibodies. Furthermore, OVA combined with KP ghosts increased the expression of the early activation marker CD69 on T cells after in vitro antigen stimulation and raised the frequencies of central memory T cells (Tcm) and CD4⁺ IFN-γ⁺ T cells. In conclusion, KP ghosts are effective as both vaccine and adjuvant components, enhancing the innate immune response of dendritic cells and the antigen-specific response of T cells. These findings highlight KP ghosts as a dual-purpose vaccine/adjuvant platform for broader antibacterial vaccine development.

Placental malaria (PM) causes mortality and severe morbidity in areas with stable Plasmodium falciparum transmission. The selective placental accumulation of P. falciparum-infected erythrocytes (IEs) is mediated by VAR2CSA, a PfEMP1-type parasite ligand that binds exclusively to placenta-restricted CSA. VAR2CSA-specific IgG is therefore generally restricted to women exposed to P. falciparum infection during pregnancy. However, widespread acquisition of VAR2CSA-reactive IgG outside pregnancy among Colombian and Brazilian individuals has been reported, supposedly due to cross-reactivity between VAR2CSA and the P. vivax-specific antigen PvDBP. Here, we measured levels and Fc-afucosylation of VAR2CSA-reactive IgG in plasma from pregnant Brazilian women at delivery, using full-length VAR2CSA (FV2) expressed in baculovirus-transfected insect cells (FV2_BIC) or Chinese hamster ovary cells (FV2_CHO) as well as the corresponding native antigen (IT4VAR04) on the IE surface. We also measured levels of IgG specific for GLURP (P. falciparum-specific) and PvDBP (P. vivax-specific). FV2_CHO-specific IgG levels were lower than FV2_BIC-reactive IgG levels. Furthermore, only FV2_CHO-specific IgG was restricted to women exposed to P. falciparum during pregnancy. Levels of PvDBP-specific IgG were significantly higher among P. vivax-exposed pregnant women but did not correlate with FV2-specific IgG levels. Finally, FV2_CHO-specific IgG was markedly Fc-afucosylated in contrast to FV2_BIC-reactive IgG. Our findings caution against using levels of IgG reacting with recombinant proteins expressed in insect cells as a measure of exposure to VAR2CSA during pregnancy, at least in South America. Furthermore, our data do not support the hypothesis that exposure to PvDBP induces IgG that cross-reacts with VAR2CSA and contributes to protection against PM.

Adverse pregnancy outcomes represent a global health burden. Bacterial infection and subsequent inflammation in gestational membranes lead to immunological and physiological changes that contribute to adverse pregnancy outcomes. Although animal models of infection during pregnancy are useful to interrogate tissue and cellular level changes in host responses, these models also have numerous drawbacks, including cost, complexity, and ethical considerations. The advent of organ-on-a-chip models provides cutting-edge new approaches to model host-pathogen interactions in multicellular organ and tissue environments. In this work, we employ an organ-on-a-chip model of the maternal-fetal interface as a tool to study immunological responses to infection with the perinatal pathogen, Group B Streptococcus (GBS). Furthermore, we validate the organ-on-a-chip assays using an ex vivo culture model of primary human gestational membranes. GBS infection leads to enhanced production of EGF, FGF-2, G-CSF, GRO-α, IL-6, IL-8, MCP-1, MIP-1α, TNF-β, IL-10, IL-17F in gestational membranes and both the maternal and fetal chambers of the organ-on-a-chip model. Additionally, GBS infection is associated with enhanced TNF-α, RANTES, IL-12p70, IP-10, MIG, FLT3L, GM-CSF, IL-1β, IL-2, PDGF-AB/BB, and IL-17E/IL-25 cytokine production in gestational membranes and the maternal compartment of the organ-on-a-chip model. Gestational membranes challenged with GBS produce IL-15, IL-27, M-CSF, MCP-3, MDC, and MIP-1β, a result that was not seen in the organ-on-a-chip model. GBS infection leads to enhanced production of eotaxin, IFN-γ, IL-1α, IL-4, IL-12p40, IL-13, and SCD40L in the maternal and fetal chambers of the organ-on-a-chip model, but not the gestational membranes ex vivo. Together, these results indicate that GBS infection induces comparable production of a repertoire of cytokines and chemokines in both models, with some salient differences, underscoring the utility of these complementary approaches to study immunological responses to infection at the maternal-fetal interface.