Balázs Rada

Pseudomonas aeruginosa remains a significant challenge to human medicine as it causes acute and chronic infections at several locations in the body, including the lung and eyes. P. aeruginosa is a very versatile, highly adaptable, opportunistic pathogen due to its large genome, ability to form protective biofilms and metabolic adaptation. Targeted, susceptibility-guided antibiotic therapy represents the best clinical approach to treat P. aeruginosa infections. However, due to the emergence of antibiotic resistance and the lack of a vaccine, it is critical to accelerate the development of novel, antipseudomonal therapies. Hypothiocyanous acid (HOSCN) occurs naturally in the human body and serves as an antimicrobial reactive oxidant in mucosal secretions, including the airway surface liquid, saliva, tears and milk. Mucosal peroxidase enzymes generate HOSCN from its substrates, thiocyanate (SCN ^- ) and H ₂ O ₂ . HOSCN has been reported to kill P. aeruginosa in vitro which indicates its potential to be used as a new mechanism to target this bacterium. This review article provides a brief overview of the human clinical relevance of P. aeruginosa and summarizes current knowledge on the in vitro antibacterial efficacy and mechanism of action of HOSCN against this pathogen. This work also reviews the mechanisms by which P. aeruginosa attempts to adapt to the antibacterial actions of HOSCN.

Chronic bacterial respiratory infections by Staphylococcus aureus are a hallmark of cystic fibrosis (CF) lung disease, affecting up to 80% of all people with CF by their mid-teens. S. aureus is able to survive and persist in the CF lung despite robust neutrophilic inflammation. As neutrophils are the immune system’s front line of defense against bacteria, the persistent nature of S. aureus infections in CF indicates both a defect in the ability of neutrophils to kill S. aureus and an enhanced ability of the bacteria to survive. S. aureus persistence in the CF lung is driven by both microbial and host, genetic and microenvironmental, factors. There doesn’t seem to be one unifying feature that makes S. aureus more virulent in the CF lung as several factors have been proposed to aide its survival. These include increased resistance to antibiotics, the ability to form small colony variants, biofilm formation, co-infection with Pseudomonas aeruginosa , and several virulence factors such as the accessory gene regulatory system, leukocidins, and staphylococcal protein A. A variety of host factors also affect the ability of neutrophils to kill S. aureus in the CF lung. Defects in the function of the cystic fibrosis transmembrane conductance regulator, the genetic cause of CF, affect phagolysosomal killing and lead to increased formation of neutrophil extracellular traps, which are less effective at killing S. aureus . Additionally, data suggest that factors within the CF lung microenvironment also affect neutrophilic killing of S. aureus . However, more research is needed to clearly identify what these environmental factors may be. This review article summarizes the current knowledge on the clinical relevance of S. aureus lung infections in CF, on microbial and host mechanisms promoting S. aureus survival in the CF lung, and on details of neutrophil- S. aureus interactions in CF. By understanding how S. aureus is able to survive in the CF lung and why neutrophils are unable to kill this bacterium, it could be possible to identify potential therapeutic targets to alleviate the consequences of S. aureus respiratory infection in CF.

Infections caused by Streptococcus pneumoniae ( Spn , pneumococcus), manifesting as pneumonia, meningitis, and otitis media, represent a leading cause of severe morbidity and mortality globally. Escalating antibiotic resistance and serotype replacement by conjugate vaccines underscore the need for serotype-independent therapies. Hypothiocyanous acid (HOSCN), a reactive oxidant naturally produced by the host innate immune system, is formed through the interaction of hydrogen peroxide, thiocyanate anions (SCN ⁻ ), and peroxidase enzymes. HOSCN is produced at mucosal surfaces, including the respiratory epithelium, where it functions as a broad-spectrum oxidative agent. HOSCN has been reported to kill Spn in vitro , but tolerance mechanisms of Spn against HOSCN have also been described. This review synthesizes recent advances in understanding HOSCN’s molecular mechanisms of action, tolerance pathways of Spn, and the effects of HOSCN on host-Spn interactions. The article also discusses current limitations and key challenges that must be addressed to translate HOSCN from preclinical promise to clinical application in the management of Spn lung infections. By leveraging an innate microbicidal pathway already active in the healthy lung, HOSCN-based therapeutics could offer rapid, resistance-refractory control of pneumococcal disease.

Cystic fibrosis (CF) lung disease manifests through abnormally thick mucus, persistent bacterial infections and a dysregulated innate immune system that involves significant neutrophilic inflammation. Neutrophils, immune cells essential to fight infections, accumulate in large numbers in CF airways and release neutrophil extracellular traps (NETs) into the airway lumen that deliver extracellular DNA, granule content and cytokines including IL-1β. Interleukin-1β, a powerful, proinflammatory cytokine, represents another, significant component of the innate immune system that is dysregulated in CF. Both defense mechanisms become problematic as NETs and IL-1β are present at elevated levels in CF airways, potentially creating a destructive cycle that exacerbates lung damage rather than protects against infections. Therefore, understanding the interplay between IL-1β and NETs is crucial for addressing CF lung disease progression. This review examines the general mechanisms of IL-1β release and NET formation, with particular focus on their role in CF lung disease, and proposes that a self-perpetuating, positive feedback loop between these two innate immune processes represents a major driving force in disease progression. This understanding suggests potential therapeutic targets for interrupting the cycle of inflammation and tissue damage in CF airways.

Loss of the flagellum marks the pathoadaptation of Pseudomonas aeruginosa to the cystic fibrosis (CF) airway environment during lung disease. Losing the flagellum is advantageous to the bacterium as the flagellum can be recognized by immune cells. The primary purpose of the flagellum is, however, to provide motility to the bacterium. Our goal was to determine whether the loss of flagellar motility or the loss of flagellum expression contributes to P. aeruginosa lung infection in CF. To address this, wild-type and gut-corrected FABP-human cystic fibrosis transmembrane conductance regulator (hCFTR) mice deficient in the murine Cftr gene were infected intratracheally with lethal doses of wild-type or flagellum-deficient P. aeruginosa . While there was no significant difference in the survival of wild-type mice after infection with either of the bacterial strains, a significantly higher mortality was observed in FABP-hCFTR mice infected with flagellum-deficient P. aeruginosa , compared to mice infected with their flagellated counterparts. When FABP-hCFTR mice were infected with isogenic, motility-deficient flagellated mutants, animal survival and lung bacterial titers were similar to those observed in mice infected with the wild-type bacterium. Airway levels of neutrophils and the amount neutrophil elastase were similar in mice infected with either the wild-type bacteria or the flagellum-deficient P. aeruginosa . Our results show that FABP-hCFTR mice have a different response to flagellum loss in P. aeruginosa compared to wild-type animals. The loss of flagellum expression, rather than the loss of motility, is the main driver behind the increased virulence of flagellum-deficient P. aeruginosa in CF. These observations provide new insight into P. aeruginosa virulence in CF. IMPORTANCE: Pseudomonas aeruginosa , a major respiratory pathogen in cystic fibrosis, is known to lose its flagellum during the course of infection in the airways. Here, we show that the loss of flagellum leads to a more enhanced virulence in Cftr-deficient cystic fibrosis mice than in control animals. Loss of flagellum expression, rather than the loss of flagellar swimming motility, represents the main driver behind this increased virulence suggesting that this appendage plays a specific role in P. aeruginosa virulence in cystic fibrosis airways.

Bone marrow stromal cells (BMSCs) have immunomodulatory activities in numerous species and have been used in clinical trials. BMSCs also make antibacterial agents. Since hepcidin is known to have antimicrobial effects in fish, we wondered if it might also be used as an antimicrobial agent by mammalian BMSCs. In the present study, we show hepcidin expression in both mouse (mBMSC) and human BMSCs (hBMSC). We observed a hBMSC hepcidin-dependent degradation of ferroportin in HEK-293 reporter cells in vitro. In human and mouse bone marrows (BM) we detected hepcidin-positive BMSCs in close proximity to hematopoietic progenitors. The conditioned culture medium of hBMSCs significantly reduced bacterial proliferation that was partially blocked by a hepcidin-neutralizing antibody. Similarly, medium in which hepcidin-deficient (Hamp^−/−) mouse BMSCs had been grown was significantly less effective in reducing bacterial counts than the medium of wild-type cells. In a zymosan-induced peritonitis mouse model we found that mBMSC-derived hepcidin reduced the number of invading polymorphonuclear (PMN) cells in the peritoneal cavity. Our results show that BMSC-derived hepcidin has antimicrobial properties in vitro and also reduces inflammation in vivo. We conclude that hepcidin should be added to the expanding arsenal of agents available to BMSCs to fight infections and inflammation.

Introduction: Severe forms of COVID-19, the disease caused by SARS-CoV-2, are characterized by acute respiratory distress syndrome, robust lung inflammation and death in some patients. Strong evidence has been accumulating that polymorphonuclear neutrophilic granulocytes (PMN) play an important role in the pathophysiology of severe COVID-19. SARS-CoV-2 directly induces in vitro PMN activation, mainly the release of neutrophil extracellular traps (NETs). However, the viral components inducing this PMN response remain unclear. Methods: In this work human PMN responses were assessed in vitro in response to the spike (S) protein of two different SARS-CoV-2 variants, anti-S IgG1 antibodies or immune complexes formed by them. Production of reactive oxygen species (ROS) was measured by Diogenes-based chemiluminescence. Release of myeloperoxidase (MPO) was assessed by ELISA while secretion of a list of cytokines and growth factors was determined by high-performance multiplex cytokine assay. Results and discussion: We show that the SARS-CoV-2 Omicron variant S protein and anti-spike IgG1, either alone or together, stimulate ROS production in human PMNs. We also observed that the SARS-CoV-2 Wuhan S protein and anti-S IgG1 antibody together trigger MPO release from PMNs. Based on the relevance of SARS-CoV-2 and influenza co-infections, we have also investigated the impact of influenza virus infection on the previous PMN responses to S proteins or anti-S antibodies. We did not detect any significant effect of influenza co-infection on ROS generation in PMNs. Our data also show that PMN stimulation by S proteins induced the release of different chemokines, growth factors, regulatory and proinflammatory cytokines. Overall, our findings show that the SARS-CoV-2 S protein, an anti-spike IgG1 antibody or their immune complex, promote oxidative responses of PMNs in a variant-dependent manner, contributing to a better understanding of the role of PMN responses during SARS-CoV-2 infection.