Microbiology and Immunology最新文献

The emergence of bacteria with both antimicrobial resistance and high virulence has become a global health concern, underscoring the urgent need to elucidate the molecular basis underlying these traits. Here, we employed the silkworm (Bombyx mori) infection model, which is suitable for high-throughput screening, together with an Escherichia coli library containing plasmid clones of all genes from strain W3110, to identify genes whose overexpression enhances virulence. We found that overexpression of the uncharacterized protein YaiX promoted bacterial proliferation in silkworms and increased host lethality. Compared with the empty-vector control, the YaiX-overexpressing strain exhibited resistance to multiple antimicrobial agents with diverse mechanisms of action, including β-lactams, tetracyclines, fluoroquinolones, aminoglycosides, cationic surfactants, and hydrogen peroxide. Sequence analysis revealed that amino acids 18-52 of YaiX contain a transferase hexapeptide domain predicted to form a left-handed parallel β-helix. Overexpression of YaiX mutants lacking regions outside this domain conferred ampicillin resistance, whereas deletion of the hexapeptide domain abolished this phenotype. RNA sequencing and GO enrichment analyses further indicated that YaiX overexpression altered the expression of genes encoding RNA-binding proteins and porins. These findings suggest that YaiX overexpression, through its hexapeptide domain, modulates gene expression and contributes to both multidrug resistance and enhanced virulence in E. coli.

Dengue Virus Serotype 4 (DENV4) continues to contribute substantially to global dengue morbidity, yet current tetravalent vaccines provide inconsistent protection and may pose risks of antibody-dependent enhancement (ADE). This study aimed to design and evaluate a serotype-specific multi-epitope vaccine (MEV-DV4) targeting the conserved capsid protein of DENV4 using integrated reverse vaccinology and immunoinformatics approaches. Conserved, antigenic, non-allergenic, and non-toxic B-cell, cytotoxic T lymphocyte (CTL), and helper T lymphocyte (HTL) epitopes were identified and assembled with a β-defensin adjuvant and PADRE sequence using optimized linkers. The final construct demonstrated high antigenicity (0.8559), structural stability, favorable physicochemical properties, and excellent solubility. Structural validation confirmed 97.6% of residues in favored Ramachandran regions. Molecular docking revealed strong interactions with TLR4 and TLR8, particularly MEV-TLR8, and molecular dynamics simulations supported the overall stability of the complex. Immune simulations predicted robust humoral and cellular responses with strong memory formation. Experimental validation in an albino mouse model using an alum-adjuvanted MEV-DV4 formulation showed early and potent antibody responses, with peak HI titers at day 21. Notably, antibody titers induced by MEV-DV4 were statistically comparable to those produced by a commercial inactivated dengue vaccine at all tested time points (p > 0.05), while no adverse reactions were observed. These computational and experimental findings demonstrate that MEV-DV4 is a safe, immunogenic, and promising serotype-specific vaccine candidate against DENV4. Further neutralization and challenge studies are warranted to advance its preclinical development.

Macrophages are key components of the innate immune system, recognizing pathogen-associated molecular patterns (PAMPs) via Toll-like receptors (TLRs) to initiate immune responses. This study investigated the individual and combinatorial effects of TLR3 Poly(I:C), TLR5 (Flagellin), and TLR7 (Imiquimod) ligands on nitric oxide (NO) production and pro-inflammatory cytokine expression in RAW 264.7 mouse macrophage cells. Our results demonstrate that all three individual TLR agonists induced NO production and cytokine expression. Notably, co-stimulation with Poly(I:C) and imiquimod led to a significant synergistic enhancement of NO production, particularly at lower concentrations. A robust upregulation was observed in key Th1-type (IL-12p40, IFN-γ, TNF-α) and Th2-type (IL-6) cytokines. The optimal synergistic response for cytokine induction was observed with a 0.1 µg/mL Poly(I:C) and 1 µg/mL imiquimod combination. These findings highlight a potent crosstalk between TRIF-dependent (TLR3) and MyD88-dependent (TLR7, TLR5) signaling pathways, leading to amplified immune activation. Our study highlights the potential of synergistic TLR ligand combinations as powerful immunomodulators, offering promising avenues for the rational design of more effective vaccine adjuvants and innovative strategies in cancer immunotherapy.

The extensive and indiscriminate use of antibiotics, which are intended to halt bacterial growth or eliminate them, has unintentionally given rise to antimicrobial resistance (AMR), which is a critical threat to global health. In the pursuit of alternative solutions, researchers have focused on type II toxin-antitoxin systems prevalent within the bacterial domain. One such promising example is the MazF toxin, which has been demonstrated to cleave RNA at specific sequences. MazF toxins are now recognized as present in a diverse range of bacterial species and exhibit varying cleavage sequence specificities. This diversity offers the potential for controlling bacterial growth by silencing the expression of hundreds of essential genes. In this study, a preliminary screening of five MazF homologs from distinct taxonomic groups was conducted to identify a MazF candidate that can effectively suppress bacterial proliferation. MazFne1, an endoribonuclease toxin originating from Nitrosomonas europaea that specifically targets UGG sequences, emerged as a promising candidate. Building upon this discovery, a phagemid system was engineered to deliver mazFne1 and express it in two distinct Escherichia coli strains. This approach successfully inhibited E. coli growth, suggesting a potential strategy for developing therapeutic interventions to combat antibiotic-resistant bacterial infections.

Cover photograph: Photograph of a 96-well plate onto which each strain was cultured with rabbit red blood cells for 1 h and the supernatant was transferred after centrifugation. Microbiol Immunol: 70:57-67. Article link here

Severe acute pancreatitis (SAP) is a critical abdominal emergency, with inflammatory response being the core pathogenic mechanism. This study aims to explore the role of miR-711 in SAP and its underlying mechanisms. This study enrolled 121 MAP patients and 109 SAP patients, collecting their clinical data. qRT-PCR measured miR-711 expression in serum and cells. Cell viability was tested with CCK-8, inflammatory factors via ELISA kits, and oxidative stress using SOD and MDA kits. The dual-luciferase assay confirmed miR-711 and KRT8 binding. Compared to the mild acute pancreatitis (MAP) group, the expression level of miR-711 in the serum of patients in the SAP group was significantly upregulated and positively correlated with relevant indicators. The ROC curve demonstrated its potential diagnostic value for SAP. In vitro, the miR-711 inhibitor effectively enhances the viability of Caerulein-induced AR42J cells and alleviates their inflammation and oxidative damage. Furthermore, the study confirmed that KRT8 is a downstream target gene of miR-711, and there is a negative correlation between their expressions. KRT8 may reverse the protective effect of miR-711 inhibitor on Cerulein-induced AR42J cells by regulating the NF-κB and p38 MAPK signaling pathways. The serum level of miR-711 in SAP patients is significantly upregulated and negatively correlated with SAP disease-related indicators. In vitro models demonstrate that inhibition of miR-711 effectively alleviates cerulein-induced cellular damage. Overexpression of KRT8 can reverse the cellular damage caused by miR-711 overexpression.

Parahenipaviruses (genus Parahenipavirus) are nonsegmented negative-strand RNA viruses belonging to the family Paramyxoviridae of the order Mononegavirales. Parahenipaviruses have been detected in shrews and rodents, some of which have been reported to be associated with human diseases. Although many parahenipaviruses have been identified thus far, there still exist large phylogenetic gaps in parahenipaviruses, suggesting the existence of yet-to-be-identified parahenipaviruses. In this study, we analyzed public RNA-seq data and identified a novel parahenipavirus from northern short-tailed shrews (Blarina brevicauda [Say, 1823]), tentatively named Blarina brevicauda virus (BbV). Based on sequence comparisons between BbV and its most closely related viruses, BbV meets the International Committee on Taxonomy of Viruses species demarcation criteria, and therefore we propose that BbV is a novel species of virus in the genus Parahenipavirus. Furthermore, mapping analysis using RNA-seq data derived from multiple tissues of northern short-tailed shrews suggested a possible kidney tropism of BbV. This study provides novel insights into the diversity of parahenipaviruses, which would contribute to a deeper understanding of their evolution and control of infectious diseases.

Cover photograph: Gene organization of the assembly and insertion machinery (Aim) operon in Porphyromonas gingivalis. Schematic representation of the chromosomal arrangement of genes PGN_0296 to PGN_0301. Arrows indicate the direction of transcription. T9SS: type IX secretion system. Microbiol Immunol: 70:9-14. Article link here

HEV causes chronic infections that are detrimental to immunocompromised patients. Previous studies showed alterations of γδ T cell subsets at the acute phase of HEV infection. To assess a possible role of CMV, we have examined the frequencies and responses to CMV and HEV of blood γδ T cell subsets from control donors and acute-phase HEV patients with or without CMV. CMV DNA was mostly undetectable in the blood of CMV-seropositive HEV patients, and anti-CMV antibodies were only slightly elevated. However, Vγ9^negVδ1^pos cells were enriched in vivo, suggesting an increased CMV burden. In contrast, γ9δ1 cells were depleted in most HEV patients, regardless of CMV status. Culturing with IL-2 and IL-15 led to strong γ9δ1 T cell enrichment in samples from HEV patients. After IL-2/IL-15 sensitization, analysis of IFN-γ responses to CMV-infected fibroblasts or hepatocarcinoma cells (with IL-18) showed innate responsiveness to CMV in γ9δ1, γ9δ2, and γ9^negδ1^pos cells in some control subjects and CMV-seronegative patients. However, these responses were selectively exacerbated in CMV-seropositive HEV patients, who also showed significant αβ T cell responses to CMV. This indicates reactivation of anti-CMV immunity. Responses to HEV-infected HepG2 cells remained undetected. IFN-γ responses were not associated with TCR downmodulation in γ9δ2 or γ9^negδ1^pos cells. However, IFN-γ-producing γ9δ1 from HEV patients were characterized by high TCR expression, which was downmodulated after stimulation with CMV-infected fibroblasts or CMV/HEV-coinfected HepG2 cells. We conclude that γ9δ1 T cells are selectively mobilized in HEV patients and can be activated by CMV.