Infection and Immunity最新文献

Klebsiella pneumoniae infections are sharply on the rise among at-risk populations. K. pneumoniae has nine serogroups of O-antigens. Recently, additional O-antigen subtypes within these serogroups have been identified; the contributions of these subtypes to pathogenic fitness and their immunogenicity, functional antibody responses, and cross-reactivity are unknown. We investigated how the addition of the single-branched galactose in O-antigen subtype O2b compared to O2a alters its virulence and host immune responses. We deleted the gmlABC region of an O2b strain of K. pneumoniae, converting it to an otherwise isogenic O2a strain. Complementation of this mutant allowed us to identify the specific genes responsible for the addition of the single branched galactose of O2b. Experiments using the O2a mutant and its parent O2b strain confirmed similar phenotypic expression of virulence factors beyond the O-antigen. Well-established murine models of pneumonia were used to determine the pulmonary fitness of the strains and assess the host innate immune responses. Complement-mediated killing assays suggested differences in susceptibility to innate immune defenses, with the O2a mutant being more susceptible to serum killing. Lastly, using polysaccharide-protein bioconjugate vaccines against these specific O-antigen subtypes, we determined that only partial cross-reactivity and protection are elicited. These studies advance our understanding of the immune response to K. pneumoniae O-antigens by defining a fitness advantage of O2b compared to O2a and informing vaccine design to combat this drug-resistant pathogen.

Chronic wound infection is a major global public health issue, with Enterococcus faecalis among the most commonly isolated pathogens from such wounds. Neutrophils are short-lived immune cells critical for host defense, yet E. faecalis-neutrophil interactions are poorly understood. Here, we show that instead of eliminating E. faecalis, neutrophils provide a niche for intracellular persistence and replication, potentially prolonging infection and inflammation at the wound site. In murine wound beds and ex vivo wound cells, intracellular E. faecalis was detected in recruited neutrophils at 24 h post-infection (h p.i). Unexpectedly, extended infection did not induce neutrophil death. Rather, E. faecalis infection significantly prolonged the life spans of both murine and human neutrophils in vitro compared to uninfected controls. Quantification of intracellular CFU revealed that E. faecalis were phagocytosed regardless of opsonization and persisted intracellularly up to 24 h p.i. This finding was confirmed via transmission electron microscopy and confocal microscopy. Blinded quantification and fluorescent D-amino acid staining, which marks newly synthesized bacterial peptidoglycan, revealed active replication within murine neutrophils between 6 and 18 h p.i., followed by a predominately persistent phase between 18 and 24 h p.i. Infected murine neutrophils remained immunologically active, secreting pro-inflammatory and chemoattractant cytokines. These findings highlight an underappreciated intracellular lifestyle for E. faecalis that may contribute to its ability to persist in chronic wounds and contribute to biofilm-associated infections.

Avian Pathogenic Escherichia coli (APEC) is a major cause of economic loss in poultry, exacerbated by the rising prevalence of antibiotic resistance. While sulfur metabolism is essential for bacterial growth, its specific role and regulation in APEC virulence remain poorly understood. This study identifies the LsrR-cysN axis as a novel regulatory pathway that critically governs APEC virulence. We demonstrate that the quorum-sensing regulator LsrR directly binds to the cysN promoter, activating its transcription. Functional analysis revealed that cysN deletion drastically attenuated virulence, significantly reducing biofilm formation, serum resistance, adhesion, invasion, and motility. The APEC94∆cysN also exhibited altered antibiotic resistance profiles, which were linked to the upregulation of efflux pumps acrA and tolC. Crucially, in a murine model, the APEC94∆cysN showed a 75% reduction in mortality and severe impairment in colonization of blood, lungs, liver, spleen, and kidneys. This attenuation was associated with a skewed host immune response, characterized by reduced levels of IL-2 and IL-6 and elevated levels of IL-4 and TNF-α. Our findings establish the LsrR-cysN axis as a central regulator connecting quorum sensing to virulence in APEC, revealing a promising target for novel anti-virulence strategies.

The type VI secretion system (T6SS) is a major virulence factor in Vibrio parahaemolyticus, but its pathogenic mechanisms are poorly understood or still not fully understood. This study investigates how two critical T6SS1 structural components, VipA1 and Hcp1, contribute to bacterial virulence and host inflammatory responses. Comparative proteomics revealed 149 secreted proteins dependent on T6SS1, including 28 core proteins requiring both VipA1 and Hcp1 for secretion. These proteins were functionally linked to metabolic pathways such as folate-mediated one-carbon metabolism and lysine degradation, as well as structural processes like flagellar assembly. Phenotypic analysis revealed that the ΔvipA1-hcp1 double mutant showed markedly attenuated virulence: 52.7% reduction in antibacterial activity compared to the wild-type strain. Biofilm formation increased 2.1-fold at 30°C and 2.8-fold at 37°C in ΔvipA1-hcp1, while swimming and swarming motility decreased by 30.9% and 35.5%. In vivo, ΔvipA1-hcp1 caused only 50% mortality in mice, compared to 91.7% for the wild-type strain, and exhibited 3- to 15-fold lower bacterial loads in the blood, liver, and spleen. Histopathological analysis confirmed that the ΔvipA1-hcp1 failed to induce tissue damage, unlike the wild-type strain. At the host interface, deletion of vipA1 and hcp1 led to significantly elevated inflammatory cytokine (IL-1β, IL-8, and IL-6) mRNA levels. Mechanistically, T6SS1 inhibited NF-κB activation by stabilizing IκBα and reducing p65 nuclear translocation (40.0% in wild-type-infected cells vs 85.8% in double mutant-infected cells). These findings establish VipA1 and Hcp1 as critical regulators of T6SS1-mediated coordinating effector secretion, virulence, immune evasion, and lethality, providing novel mechanistic insights into V. parahaemolyticus pathogenesis.

Antibiotic-induced microbiome injury, defined as a reduction of ecological diversity and obligate anaerobe taxa, is associated with negative health outcomes in hospitalized patients, and healthy individuals who received antibiotics in the past are at higher risk for autoimmune diseases. Postbiotics contain mixtures of bacterial fermentation metabolites and bacterial cell wall components that have the potential to modulate microbial communities. Yet, it is unknown if a fermentation-derived postbiotic can reduce antibiotic-induced microbiome injury. Here, we present the results from a single-center, randomized placebo-controlled trial involving 32 patients who received an oral, fermentation-derived postbiotic alongside oral antibiotic and probiotic therapy for non-gastrointestinal (GI) infections. At the end of the antibiotic course, patients receiving the postbiotic (n = 16) had significantly higher fecal bacterial alpha diversity (+40%, inverse Simpson index) compared to the placebo group (n = 16), and the treatment was well-tolerated. Analysis of 157 longitudinal fecal samples revealed that this increased diversity was driven by enrichment of health-associated taxa, notably obligate anaerobic Firmicutes, particularly Lachnospiraceae. In contrast, Escherichia/Shigella species, often linked to pathogenicity and antibiotic resistance, were reduced in postbiotic-treated patients at the end of antibiotic treatment and remained lower up to 10 days later. Our findings suggest that postbiotic co-administration during antibiotic therapy may augment health-associated gut microbiome composition and mitigate antibiotic-induced microbiome injury.Trial registration ISRCTN30327931 retrospectively registered.

Streptococcus agalactiae (Group B Streptococcus or GBS), a Gram-positive bacterium, and Candida albicans, a polymorphic fungus, are commensal microbes in most of the population they colonize. However, for certain patients, they can cause severe and sometimes fatal infections. Previous research has indicated that GBS and C. albicans can synergize to enhance the colonization of GBS in the bladders of mice, but not much was known prior to this study about how interactions between GBS and C. albicans alter treatment effectiveness and infection outcome in vivo. Results showed that interactions between the two opportunistic pathogens were influenced by media nutrient availability and that the presence of C. albicans in a culture reduces the effectiveness of certain antibiotics against GBS in vitro. This study also utilized a larval zebrafish model to investigate differences in virulence in solo infections vs co-infections with both pathogens in vivo. Co-infections of GBS and C. albicans into the otic vesicle were found to have increased virulence compared to solo infections of either pathogen. Co-infection also led to an increased GBS burden compared to solo GBS infections. Co-infections of GBS and C. albicans by yolk sac injection were not more virulent than solo infections with either pathogen. However, the antibiotic clindamycin was less effective in preventing mortality in co-infections compared to solo GBS infections. Overall, these findings highlight how interactions between GBS and C. albicans can influence treatment effectiveness and virulence during infection.

Neutrophils are dominant cells during acute immune response to Clostridioides difficile infection (CDI). A higher number of infiltrating colonic neutrophils is clearly linked to greater tissue damage and severe CDI (3, 4). However, the mechanism(s) by which neutrophils exacerbate tissue damage in CDI remain unknown. We investigated the role of a neutrophil subset marked by Olfactomedin-4 expression (OLFM4⁺ neutrophils) during CDI. Single-cell transcriptomics reveal that Olfm4 is increased in blood neutrophils of infected mice, and these cells exhibit gene signatures characterized by high expression of degranulation genes. In C. difficile-infected mice, OLFM4⁺ neutrophils aggregate to areas of severe intestinal epithelial cell (IEC) damage, and plasma OLFM4 was significantly increased in both C. difficile-infected mice and patients. In vitro, OLFM4⁺ neutrophils and recombinant OLFM4 protein exacerbated C. difficile toxin-induced IEC damage. In sum, our studies provide novel insights into neutrophil-mediated pathology and highlight the role of OLFM4⁺ neutrophils in worsening CDI-induced IEC damage.

Salmonella enterica infections are a major cause of morbidity and mortality worldwide, especially in sub-Saharan Africa and in the Asian continent, and are increasingly associated with antimicrobial resistance. Salmonella enterica serovars Typhi and Paratyphi A, B, and C cause enteric fever, while non-typhoidal Salmonella serovars (usually Typhimurium and Enteritidis) cause mainly gastroenteritis which can lead to systemic infections. Vaccines are only licensed against S. Typhi, but different combinations are in clinical development to prevent S. Typhi and S. Paratyphi A or S. Typhi and non-typhoidal Salmonella. Here, we describe elements of the pathogenesis of and immunity to Salmonella that are critical to guide the rational design of vaccines. We highlight how the choice of appropriate immunogenic and protective antigens would be essential to achieve the maximum coverage of serovars in a multivalent Salmonella vaccine. The principal vaccines under development at the preclinical and clinical stages are described, together with considerations on the technical and clinical feasibility of moving combination vaccines toward licensure.

Avoiding lysosomal degradation is vital to the success of intracellular pathogens. The Gram-negative bacterium Coxiella burnetii and protozoan parasites of the Leishmania genus are unique in being able to replicate within the mature phagolysosomal compartment of host cells, though the exact mechanisms utilized to withstand this hostile environment are not clearly defined. We recently reported that C. burnetii removes the lysosomal protease cathepsin B during infection of mammalian cells. Here, we aimed to determine if this virulence strategy was also employed by the intralysosomal pathogen, Leishmania mexicana. In contrast to C. burnetii, decreases in the activity of specific cathepsins were not detected in L. mexicana-infected host cells as determined using immunoblotting and protease activity-based probes. Co-infection of THP-1 macrophage-like cells with both pathogens resulted in a proteolytic and secretory phenotype consistent with C. burnetii infection, suggesting that C. burnetii-induced remodeling of the lysosome is not influenced by L. mexicana. The host cell proteome and secretome of L. mexicana-infected cells were defined using mass spectrometry. This confirmed that, unlike C. burnetii, L. mexicana does not induce increased abundance of lysosomal proteins either intracellularly or in the extracellular milieu. Collectively, this study reveals that although C. burnetii and L. mexicana reside in a phagolysosomal intracellular niche, they employ divergent mechanisms to survive within this hostile compartment.

As a bacterial pathogen enters a human host, it immediately encounters a temperature upshift to 37°C. Mimicking the early hours of infection, we analyzed the transcriptome and proteome of uropathogenic Escherichia coli CFT073 initially grown at 23°C, then shifted to 37°C for 4 h. Temperature caused a change in mRNA expression for 9% of the genome (1% false discovery rate, ≥2-fold); similar impacts were observed for the proteome with a good concordance amongst the most highly temperature-regulated genes. Comparison to E. coli K-12 MC4100 shows temperature to be a more broadly used regulatory cue in the uropathogen. Multiple operons associated with fimbrial adhesion, biofilm formation, immune evasion, and competitor defense show temperature regulation. Multiple fimbrial adhesins (pap, pap-2, foc) are increased in expression at 37°C, while others (ecp) are favored at 23°C. Decreased motility gene expression at 37°C and 23°C is correlated with the thermoregulation of multiple motility repressors (papX, focX, pdeL, and rpoS). Several biofilm formation and c-di-GMP signaling genes showed preferential expression at 37°C, suggesting human body temperature modulates this process. Growth at 37°C promotes a broad set of immune evasion genes (complement evasion, antimicrobial peptide cleavage, phagocyte killing/iron acquisition, copper export) along with genes associated with competitor bacterial and phage defense. RpoS protein expression and the genes it controls show minimal changes during this time course, indicating bacteria enter the host ready to counter diverse stresses in various niches. Together, our studies demonstrate that temperature cues a suite of genes whose expression benefits host colonization and survival.