Aim: Investigation of the lateral flagellar (Laf) genes transcription by the quorum sensing (QS) regulators AphA and OpaR in Vibrio parahaemolyticus. Materials & methods: Regulation mechanisms were...

Vibrio parahaemolyticus possesses two types of flagella: a single polar flagellum (Pof) for swimming and the peritrichous lateral flagella (Laf) for swarming. Expression of Laf genes has previously been reported to be regulated by the quorum sensing (QS) regulators AphA and OpaR. In the present study, we showed that OpaR, the QS regulator at high cell density (HCD), acted as a negative regulator of swimming motility and the transcription of Pof genes in V. parahaemolyticus. OpaR bound to the promoter-proximal DNA regions of flgAMN, flgMN, and flgBCDEFGHIJ within the Pof gene loci to repress their transcription, whereas it negatively regulates the transcription of flgKL-flaC in an indirect manner. Thus, this work investigated how QS regulated the swimming motility via direct action of its master regulator OpaR on the transcription of Pof genes in V. parahaemolyticus.

Diffuse alveolar damage (DAD), a lethal manifestation of acute lung injury, remains a critical public health concern due to the absence of targeted therapies. However, the underlying cellular and molecular mechanisms responsible for immunopathology during DAD progression are largely undefined. Here, by integrating single cell RNA sequencing, functional assays, and genetic/pharmacological interventions in a mouse model of ricin-induced DAD, we revealed a significant accumulation of neutrophil with an activated phenotype that plays a critical role in immunopathology. We observed the formation of neutrophil extracellular traps (NETs) during DAD, which further intensified inflammation and tissue injury. IL-17A signaling activity was upregulated in DAD-affected lungs, while IL-17A deficiency or functional blockade significantly attenuated neutrophil recruitment, NET generation, and tissue damage. Mechanically, IL-17A stimulates lung resident fibroblasts to produce the neutrophil chemoattractant CXCL1. Notably, type 3 innate lymphoid cells (ILC3) emerged as the dominant source of IL-17A, highlighting a triad of interactions among ILC3, fibroblast, and neutrophil in DAD pathogenesis. This finding delineates a pathogenic IL-17A-neutrophil-NET axis that amplifies lung immunopathology after ricin-induced DAD, a deeper understanding of these relationships may pave the way for mitigate DAD immunopathology and other lung inflammatory disorders.

Pseudomonas aeruginosa (PA) is a key pathogen in hospital-acquired pneumonia (HAP) and ventilator-associated pneumonia (VAP), challenging clinical medicine. This study aims to elucidate the characteristics of the host's innate immune response following inhalational PA infection. We developed a mouse model by aerosolized intratracheal inoculation with PA and conducted a comprehensive analysis at the protein, cellular, and gene expression levels. Protein analysis revealed a substantial increase in inflammatory proteins in the bronchoalveolar lavage fluid and serum, indicating a robust inflammatory response in the lungs and systemic circulation. Cellular investigations showed an increase in neutrophils, monocytes, and alveolar macrophages during infection, whereas NK cells showed a marked reduction from 5.88% pre-infection to 2.41% at 24 h (p = 0.0102) and 1.55% by 48 h (p = 0.0023). To assess gene expression changes, RNA-sequencing technology was employed to map the temporal shifts in the transcriptional profile of the host lung post-infection. We analysed the expression patterns and dynamic transcriptional characteristics of differentially expressed genes (DEGs), describing the inflammation progression. Importantly, Through the analysis of single-cell RNA sequencing (scRNA-seq) datasets in public repositories, we observed the reduction in conventional natural killer (cNK) cells, rather than tissue-resident natural killer (trNK) cells in the early stages of PA infection. Sequential scRNA-seq analysis resolved NK-subset heterogeneity, revealing that cNK dominance (77.8% of total NK cells) under homeostasis collapsed to 9.2% within 24 h post-infection. Our findings establish cNK attrition as the earliest immune checkpoint failure in PA pneumonia and provide proof-of-concept for cNK-targeted immunotherapies to counteract lethal pulmonary infections. Keywords: Pseudomonas aeruginosa, aerosolized intratracheal inoculation, conventional NK cells, innate immunity, RNA-sequencing.