A c-di-GMP signaling module controls responses to iron in Pseudomonas aeruginosa.

Cyclic dimeric guanosine monophosphate (c-di-GMP) serves as a bacterial second messenger that modulates various processes including biofilm formation, motility, and host-microbe symbiosis. Numerous studies have conducted comprehensive analysis of c-di-GMP. However, the mechanisms by which certain environmental signals such as iron control intracellular c-di-GMP levels are unclear. Here, we show that iron regulates c-di-GMP levels in Pseudomonas aeruginosa by modulating the interaction between an iron-sensing protein, IsmP, and a diguanylate cyclase, ImcA. Binding of iron to the CHASE4 domain of IsmP inhibits the IsmP-ImcA interaction, which leads to increased c-di-GMP synthesis by ImcA, thus promoting biofilm formation and reducing bacterial motility. Structural characterization of the apo-CHASE4 domain and its binding to iron allows us to pinpoint residues defining its specificity. In addition, the cryo-electron microscopy structure of ImcA in complex with a c-di-GMP analog (GMPCPP) suggests a unique conformation in which the compound binds to the catalytic pockets and to the membrane-proximal side located at the cytoplasm. Thus, our results indicate that a CHASE4 domain directly senses iron and modulates the crosstalk between c-di-GMP metabolic enzymes.

Molecular mechanisms of DNase inhibition of early biofilm formation Pseudomonas aeruginosa or Staphylococcus aureus: A transcriptome analysis.

In vitro studies show that DNase can inhibit Pseudomonas aeruginosa and Staphylococcus aureus biofilm formation. However, the underlying molecular mechanisms remain poorly understood. This study used an RNA-sequencing transcriptomic approach to investigate the mechanism by which DNase I inhibits early P. aeruginosa and S. aureus biofilm formation on a transcriptional level, respectively. A total of 1171 differentially expressed genes (DEGs) in P. aeruginosa and 1016 DEGs in S. aureus enriched in a variety of biological processes and pathways were identified, respectively. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses revealed that the DEGs were primarily involved in P. aeruginosa two-component system, biofilm formation, and flagellar assembly and in S. aureus biosynthesis of secondary metabolites, microbial metabolism in diverse environments, and biosynthesis of amino acids, respectively. The transcriptional data were validated using quantitative real-time polymerase chain reaction (RT-qPCR), and the expression profiles of 22 major genes remained consistent. These findings suggested that DNase I may inhibit early biofilm formation by downregulating the expression of P. aeruginosa genes associated with flagellar assembly and the type VI secretion system, and by downregulating S. aureus capsular polysaccharide and amino acids metabolism gene expression, respectively. This study offers insights into the mechanisms of DNase treatment-based inhibition of early P. aeruginosa and S. aureus biofilm formation.

Metagenomic Next-Generation Sequencing Assists in the Diagnosis of Mediastinal Aspergillus fumigatus Abscess in an Immunocompetent Patient: A Case Report and Literature Review.

Background: Aspergillus fumigatus is an opportunistic fungal pathogen, which is commonly found in lungs and rarely causes infections in mediastinum. Mediastinal Aspergillus abscess is a serious infectious condition, and is characterized by difficult diagnosis due to its clinical manifestations being nonspecific. Case Presentation: Here, we report a case of a mediastinal Aspergillus fumigatus abscess in an immunocompetent patient. The patient was a 45-year-old woman who presented with a 20-day history of sore throat without any underlying diseases. Chest computed tomography (CT) showed a mass in the anterior superior mediastinum. Metagenomic next-generation sequencing (mNGS) identified Aspergillus fumigatus sequences in endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) tissue, indicating the mediastinal Aspergillus fumigatus infection of this patient. The following mediastinal biopsy histological analysis and tissue fungi culture also suggested Aspergillus fumigatus infection, confirming the mNGS detection. The patient was diagnosed with mediastinal aspergillosis caused by Aspergillus fumigatus. After timely voriconazole treatment, the patient was discharged with good condition. Conclusion: Our study presented a rare case with mediastinal Aspergillus fumigatus abscess in an immunocompetent patient. As a new clinical diagnostic method, mNGS could assist timely diagnosis and precise treatment of Aspergillus infection.

DNase inhibits early biofilm formation in Pseudomonas aeruginosa- or Staphylococcus aureus-induced empyema models.

Anti-infection strategies against pleural empyema include the use of antibiotics and drainage treatments, but bacterial eradication rates remain low. A major challenge is the formation of biofilms in the pleural cavity. DNase has antibiofilm efficacy in vitro, and intrapleural therapy with DNase is recommended to treat pleural empyema, but the relevant mechanisms remain limited. Our aim was to investigate whether DNase I inhibit the early biofilm formation in Pseudomonas aeruginosa- or Staphylococcus aureus-induced empyema models. We used various assays, such as crystal violet staining, confocal laser scanning microscopy (CLSM) analysis, peptide nucleic acid-fluorescence in situ hybridization (PNA-FISH), and scanning electron microscopy (SEM) analysis. Our results suggested that DNase I significantly inhibited early biofilm formation in a dose-dependent manner, without affecting the growth of P. aeruginosa or S. aureus in vitro. CLSM analysis confirmed that DNase I decreased the biomass and thickness of both bacterial biofilms. The PNA-FISH and SEM analyses also revealed that DNase I inhibited early (24h) biofilm formation in two empyema models. Thus, the results indicated that DNase inhibited early (24h) biofilm formation in P. aeruginosa- or S. aureus-induced rabbit empyema models and showed its therapeutic potential against empyema biofilms.