Infection and Immunity最新文献

Previous research demonstrated that Campylobacter jejuni encodes a heme utilization system that facilitates heme-dependent growth under iron-limiting conditions, and that transcription of this system is induced during human infection. Despite these observations, it remained unknown whether the heme transport system is required for colonization and disease in a susceptible host. To address this, we created individual non-polar deletion mutants of each component of the heme transport system and examined their ability to promote heme-dependent growth and iron uptake. From this work, we found that only the heme receptor, ChuA, was required for heme-dependent growth and iron acquisition, which supports the earlier work of another group. Furthermore, we examined whether intestinal colonization, immune activation, and pathology were altered during infection with these mutants. After establishing that elevated heme and chuABCD expression occur during C. jejuni infection of IL-10^-/- mice, we found that a mutant of the heme receptor, ChuA, exhibited significantly reduced colonization of the colon. In addition, we found that neutrophil and circulating monocyte recruitment were significantly reduced in the colon during infection with the ChuA mutant, but that populations of self-maintaining tissue macrophages remained high. Loss of ChuA reduced colonic colonization and was accompanied by diminished innate immune cell recruitment and intestinal pathology. Together, these findings identify ChuA-dependent heme acquisition as a key determinant of efficient colonic colonization and the associated inflammatory disease.

Clostridioides difficile infection (CDI) is a common cause of antibiotic-associated diarrhea and arises from disruption of the resident microbiota in the gastrointestinal tract. Despite being a major cause of morbidity and mortality worldwide, CDI presents as a wide spectrum of clinical manifestations, ranging from completely asymptomatic infection to severe and fulminant colitis. Recent research has revealed that CDI disease severity is influenced by the complex ecosystem of the gut, involving interactions between C. difficile, the host, and the resident gut microbiota. This review will summarize the current state of the field in understanding how the balance of interactions in the gut alters CDI disease severity using clinical data and mouse models of infection. In addition, external factors influencing these interactions, such as diet and pharmaceutical drugs, will be discussed. Ultimately, a comprehensive understanding of the ecological factors influencing patient outcome will be critical for the design of novel therapeutics for primary and recurrent CDI.

The commensal bacteria of the intestinal microbiota inhabit one of the most densely populated and dynamic microbial ecosystems known, where fluctuations in nutrient availability, host-derived stresses, and microbial competition require highly coordinated regulatory responses to maintain colonization and fitness. As a stable, homeostatic microbiota performs essential functions for the host, understanding how commensals adapt to the fluctuating intestinal environment is important for maintaining intestinal homeostasis and informing the development of new microbiota-targeted therapeutics. Throughout diverse bacterial species, rapid adaptation to environmental perturbations is mediated in part by small regulatory RNAs (sRNAs). While most sRNA research has focused on a limited number of model organisms, particularly bacterial pathogens, recent studies have uncovered important roles for these noncoding RNAs in promoting the fitness and resilience of gut commensals. Advances in differential RNA sequencing, comparative genomics, and functional genetics have revealed diverse repertoires of sRNAs in prominent gut symbionts, with regulatory roles in nutrient acquisition, stress responses, and host colonization. This minireview highlights recent advances and outlines future directions to further elucidate sRNA-mediated regulation as a pillar of commensal resilience in the mammalian gut.

Providencia alcalifaciens is a gut commensal bacterium and also an emerging enteric pathogen associated with sporadic infections and outbreak cases in humans. The most notable outbreak caused by this bacterium occurred in 1996 in Fukui Prefecture, Japan, affecting 270 individuals. However, the pathogenic mechanisms responsible for this outbreak remain unknown. In this study, we identified the key virulence determinants of the Fukui outbreak strains through genomic and functional analyses. These strains uniquely carry a ~162 kb large plasmid encoding a type III secretion system (T3SS) closely homologous to the Salmonella SPI-1 T3SS. We also show that the plasmid-encoded T3SS (T3SSp) constitutes a functional secretion system and is essential for the pathogenicity of the Fukui outbreak strain, including the invasion of cultured epithelial cells and the induction of diarrhea in a rabbit model. Secretome analysis identified effectors secreted in a T3SSp-dependent manner, among which PipA-sharing limited sequence similarity with SipA, a SPI-1 T3SS effector-plays a crucial role in inducing diarrhea. Ectopic expression of PipA in HeLa cells caused focal accumulation of F-actin, indicating its cytoskeleton-modulating activity. Comparative genomics with other Providencia species revealed the dissemination of the large plasmid, with structural variations among enteropathogenic strains of P. alcalifaciens and Providencia rustigianii associated with clinical cases in humans and animals. Thus, our findings underscore the molecular basis of P. alcalifaciens pathogenicity in the Fukui outbreak and highlight the significance of the large plasmids encoding T3SS in driving pathogenic evolution among Providencia species.

Pseudomonas aeruginosa is a key bacterial pathogen causing lethal infections in the lungs of cystic fibrosis (CF) patients. These infections involve complex interactions between P. aeruginosa and the airway mucosal epithelium, which are regulated by multiple factors including bacterial adhesion. Calcium (Ca²⁺), a potent cellular signal, the levels of which increase in the airway fluids during CF, was shown to alter the expression of multiple virulence factors in P. aeruginosa. However, its role in regulating the host-pathogen interactions is not well understood. This study investigates the impact of Ca²⁺ on the initial adherence of P. aeruginosa to lung epithelial cells and its role in regulating the expression of several adhesins, including flagellin (fliC), type IV fimbrial precursor (pilA), and galactophilic lectin (lecA). Two epithelial cell lines, A549 and CuFi-5 cells, were infected with two strains of P. aeruginosa, PAO1 and FRD1. According to quantitative adherence assays, immunofluorescence microscopy, and scanning electron microscopy, adherence of P. aeruginosa to both cell lines was enhanced at elevated Ca²⁺. RNA-seq and RT-qPCR showed the transcription of adhesins (fliC, pilA, and lecA) was differentially regulated by Ca²⁺ across bacterial strains and host cell models. Deleting lecA either completely or partially abolished Ca²⁺-dependent increase in adherence of FRD1. This impact was reversed by gene complementation. In agreement with the Ca²⁺-induced expression of fliC, the proportion of flagellated PAO1 cells increased within the bacterial populations grown in the presence of Ca²⁺. The observed Ca²⁺-dependent increase in P. aeruginosa adherence to lung epithelial cells may promote the initiation of the pathogen's infections in CF airways.

Streptococcus pneumoniae (Spn) asymptomatically colonizes the upper respiratory tract (URT), a niche from which it can transmit to another host or cause invasive disease in the same host. The in vivo transcriptional adaptations that Spn undergoes during nasopharyngeal colonization, particularly during influenza A virus (IAV) co-infection, are poorly understood. Here, we leveraged an established infant mouse model of colonization, shedding, and transmission to perform genome-wide transcriptomic profiling of Spn during mono- and during IAV co-infection. Compared with broth-grown controls, pneumococci isolated from the URT exhibited distinct transcriptional programs, with over 200 genes differentially expressed across time points. Genes involved in carbohydrate uptake and metabolism, glycan degradation, amino sugar and nucleotide sugar metabolism, and amino acid biosynthesis were consistently enriched during colonization, highlighting metabolic adaptation to the nasopharyngeal niche. In contrast, IAV co-infection induced a markedly distinct transcriptional signature, including upregulation of branched-chain amino acid biosynthesis, bacteriocin production, and phosphate acquisition systems. Notably, the pilus islet-1 locus was upregulated during Spn-IAV co-infection. Functional studies demonstrated that while the pilus was dispensable for colonization under mono- and co-infection conditions, it promoted high-shedding events and enhanced inflammatory responses during IAV co-infection. However, reduced inflammation and reduced high-shedding events from pups inoculated with a pilus-deficient mutant did not alter transmission frequency in the infant mouse model. Collectively, our findings define the in vivo transcriptional landscape of Spn during URT colonization and reveal distinct bacterial adaptations during viral co-infection, providing insight into mechanisms that influence pneumococcal persistence, inflammation, and transmission.

Francisella tularensis is a gram-negative bacterium that causes tularemia, a fatal zoonotic disease. F. tularensis has been used in the bioweapon programs of several countries. Its potential use as a bioterrorism agent led the CDC to classify F. tularensis as a Tier 1 Select Agent. The cytosolic sensor absent in melanoma 2 (Aim2) detects double-stranded DNA in the cytosol of infected cells and subsequently assembles a multiprotein complex known as the inflammasome. Inflammasome activation drives the secretion of IL-1β and IL-18, key pro-inflammatory cytokines required for controlling F. tularensis infection. Prior studies have shown that F. tularensis actively suppresses Aim2 inflammasome activation; however, the underlying mechanism remains unknown. We hypothesized that F. tularensis suppresses Aim2-mediated responses by modulating the intracellular redox environment. We utilized an F. tularensis live vaccine strain (LVS) mutant lacking OxyR (ΔoxyR), a transcriptional regulator that controls the expression of major antioxidant enzymes. Our results show that macrophages infected with the ΔoxyR mutant exhibit significantly higher levels of Aim2-dependent caspase-1 and IL-1β than those infected with wild-type bacteria. The expression of interferon regulatory factor 1 and the guanylate-binding proteins GBP2 and GBP5, upstream signaling components of the Aim2 inflammasome, is markedly higher in ΔoxyR-infected macrophages than in controls. These changes were absent in ΔoxyR-infected NADPH oxidase-deficient macrophages, which are unable to generate reactive oxygen species. Collectively, these findings demonstrate that the macrophage redox environment plays a key role in activating the Aim2 inflammasome. This work advances understanding of how F. tularensis-encoded factors subvert host innate immune defenses.

Enterotoxigenic Escherichia coli (ETEC) remains a leading cause of diarrheal disease in low- and middle-income countries, yet vaccine efforts have struggled to achieve broad and durable protection. To better understand early steps in host-pathogen interactions, we utilized human stem cell-derived jejunal organoid monolayers to compare epithelial responses to ETEC H10407 and E24377A, prototypical strains frequently used in controlled human infection challenges. Transcriptomic analysis revealed that H10407 uniquely stimulated a robust innate immune response, characterized by upregulation of TLR5-dependent genes, including IL-17C and CXCL6. This response was found to be driven by sustained expression of flagella, the result of a naturally occurring fliZ gene disruption in H10407, which allows flagellin production despite ample nutrient availability. Complementation with fliZ restored repression of flagellar genes, confirming its regulatory role. In epithelial-macrophage co-cultures, H10407-expressed flagella enhanced macrophage sampling of bacteria as well as IL-6 production, effects that were not replicated by recombinant flagellin alone. However, exogenous flagellin did induce epithelial IL-17C release, a cytokine implicated in adaptive immune priming. These findings identify constitutive flagellar expression as a key factor in the heightened innate immune response and virulence of H10407, and highlight its potential utility in optimizing mucosal vaccine design against ETEC.

Giardiasis caused by the intestinal protozoan Giardia lamblia is a major global cause of diarrheal disease and malabsorption particularly affecting young children and populations in low- and middle-income countries. Despite evidence of immunity after infection, no human vaccine is available. Induction of effective mucosal immunity is critical for protection against this lumen-dwelling pathogen, but has been hampered by the lack of suitable adjuvants. We show here in a murine model that cyclic di-(3',5')-guanosine monophosphate (CDG) is a potent mucosal adjuvant for immunization against G. lamblia with a defined antigen, α1-giardin. Based on this observation, we developed a novel nanovaccine strategy that employs metal-organic framework (MOF) nanoparticles loaded with CDG and cloaked with native G. lamblia membranes. These particles enable simultaneous delivery of multiple antigens in their native form together with a powerful mucosal adjuvant. CDG-loaded MOFs exhibited efficient cargo encapsulation and controlled release and strong immune-stimulatory activity in dendritic cells. Intranasal immunization of mice with G. lamblia membrane-coated CDG-MOFs elicited robust parasite-specific IgG responses and conferred profound protection against G. lamblia infection that was superior to that achieved with a single defined antigen. Immunization-induced protection was dependent on the CDG receptor STING and required CD4⁺ T cells but was independent of secretory IgA. These findings demonstrate that MOF-based nanovaccines incorporating a cyclic dinucleotide represent an effective platform for inducing protective mucosal immunity against giardiasis. More broadly, this approach offers a versatile strategy for delivering hydrophilic adjuvants and multivalent antigens for vaccination against mucosal pathogens of the gastrointestinal tract.