Biology subjects

Kirienko, N.

Publications and source records attributed to Kirienko, N..

6 recordsLinked to original sources

bioRxiv2024

Role of R5 Pyocin in the Predominance of High-Risk Pseudomonas aeruginosa Isolates

Infections with antimicrobial resistant pathogens, such as Pseudomonas aeruginosa, are a frequent occurrence in healthcare settings. Human P. aeruginosa infections are predominantly caused by a small number of sequence types (ST), such as ST235, ST111, and ST175. Although ST111 is recognized as one of the most prevalent high-risk P. aeruginosa clones worldwide and frequently exhibits multidrug-resistant or extensively drug-resistant phenotypes, the basis for this dominance remains unclear. In this study, we used a genome-wide transposon insertion library screen to discover that the competitive advantage of ST111 strains over certain non-ST111 strains is through production of R pyocins. We confirmed this finding by showing that competitive dominance was lost by ST111 mutants with R pyocin gene deletions. Further investigation showed that sensitivity to ST111 R pyocin (specifically R5 pyocin) is caused by deficiency in the O-antigen ligase waaL, which leaves lipopolysaccharide (LPS) bereft of O antigen, enabling pyocins to bind the LPS core. In contrast, sensitivity of waaL mutants to R1 or R2 pyocins depended on additional genomic changes. In addition, we found the PA14 mutants in lipopolysaccharide biosynthesis (waaL, wbpL, wbpM) that cause high susceptibility to R pyocins also exhibit poor swimming motility. Analysis of 5,135 typed P. aeruginosa strains revealed that several international, high-risk sequence types (including ST235, ST111, and ST175) are enriched for R5 pyocin production, indicating a correlation between these phenotypes and suggesting a novel approach for evaluating risk from emerging prevalent P. aeruginosa strains. Overall, our study sheds light on the mechanisms underlying the dominance of ST111 strains and highlighting the role of waaL in extending spectrum of R pyocin susceptibility.

microbiology↗

bioRxiv2023

Cytotoxic rhamnolipid micelles drive acute virulence in Pseudomonas aeruginosa

Pseudomonas aeruginosa is an opportunistic human pathogen that has developed multi- or even pan-drug resistance towards most frontline and last resort antibiotics, leading to increasing infections and deaths among hospitalized patients, especially those with compromised immune systems. Further complicating treatment, P. aeruginosa produces numerous virulence factors that contribute to host tissue damage and immune evasion, promoting bacterial colonization and pathogenesis. In this study, we demonstrate the importance of rhamnolipid production in host-pathogen interactions. Secreted rhamnolipids form micelles that exhibited highly acute toxicity towards murine macrophages, rupturing the plasma membrane and causing organellar membrane damage within minutes of exposure. While rhamnolipid micelles (RMs) were particularly toxic to macrophages, they also caused membrane damage in human lung epithelial cells, red blood cells, Gram-positive bacteria, and even non-cellular models like giant plasma membrane vesicles. Most importantly, rhamnolipid production strongly correlated to P. aeruginosa virulence against murine macrophages in various panels of clinical isolates. Altogether, our findings suggest that rhamnolipid micelles are highly cytotoxic virulence factors that drive acute cellular damage and immune evasion during P. aeruginosa infections.

microbiology↗

bioRxiv2023

Pseudomonas aeruginosa Siderophores Damage Lung Epithelial Cells and Promote Inflammation

Multidrug-resistant Pseudomonas aeruginosa is a common nosocomial respiratory pathogen that continues to threaten the lives of patients with mechanical ventilation in intensive care units and those with underlying comorbidities such as cystic fibrosis or chronic obstructive pulmonary disease. For over 20 years, studies have repeatedly demonstrated that the major siderophore pyoverdine is an important virulence factor for P. aeruginosa in invertebrate and mammalian hosts in vivo. Despite its physiological significance, an in vitro, mammalian cell culture model to characterize the impact and molecular mechanism of pyoverdine-mediated virulence has only been developed very recently. In this study, we adapt a previously-established, murine macrophage-based model for human bronchial epithelial cells (16HBE). We demonstrate that conditioned medium from P. aeruginosa induced rapid 16HBE cell death through the pyoverdine-dependent secretion of cytotoxic rhamnolipids. Genetic or chemical disruption of pyoverdine biosynthesis decreased rhamnolipid production and mitigated cell death. Consistent with these observations, chemical depletion of lipid factors or genetic disruption of rhamnolipid biosynthesis was sufficient to abrogate conditioned medium toxicity. Furthermore, we also examine the effects of purified pyoverdine exposure on 16HBE cells. While pyoverdine accumulated within cells, the siderophore was largely sequestered within early endosomes, showing minimal cytotoxicity. More membrane-permeable iron chelators, such as the siderophore pyochelin, decreased epithelial cell viability and upregulated several proinflammatory genes. However, pyoverdine potentiated these iron chelators in activating proinflammatory pathways. Altogether, these findings suggest that the siderophores pyoverdine and pyochelin play distinct roles in virulence during acute P. aeruginosa lung infection.

cell biology↗

bioRxiv2021

Box C/D Small Nucleolar Ribonucleoproteins Regulate Mitochondrial Surveillance and Innate Immunity

Monitoring of mitochondrial functions is crucial for organismal survival. This task is performed by mitochondrial surveillance or quality control pathways, which are activated by signals originating from mitochondria and relayed to the nucleus (retrograde response) to start the transcription of protective genes. In Caenorhabditis elegans, several systems exist, including the UPRmt, MAPKmt, and the ESRE pathway. These pathways are highly conserved and their loss results in compromised survival following mitochondrial stress. In this study, we found a novel interaction between the box C/D snoRNA core proteins (snoRNPs) and mitochondrial surveillance and innate immunity pathways. We showed that C/D snoRNPs are required for the full expressions of UPRmt and ESRE upon stress. Meanwhile, we found that the loss of C/D snoRNPs increased immune responses. Understanding the "molecular switch" mechanisms of interplay between these pathways may be important for understanding of multifactorial processes, including response to infection or aging.

cell biology↗

bioRxiv2020

Novel Immune Modulators Enhance Caenorhabditis elegans Resistance to Multiple Pathogens

Traditionally, treatments for bacterial infection have focused on killing the microbe or preventing its growth. As antimicrobial resistance becomes more ubiquitous, the feasibility of this approach is beginning to wane and attention has begun to shift toward disrupting the host-pathogen interaction by improving the host defense. Using a high-throughput, fragment-based screen to identify compounds that alleviate Pseudomonas aeruginosa-mediated killing of Caenorhabditis elegans, we identified over 20 compounds that stimulated host defense gene expression. Five of these molecules were selected for further characterization. Four of five compounds showed little toxicity against mammalian cells or worms, consistent with their identification in a phenotypic, high-content screen. Each of the compounds activated several host defense pathways, but the pathways were generally dispensable for compound-mediated rescue in Liquid Killing, suggesting redundancy or that the activation of one or more unknown pathways may be driving compound effects. A genetic mechanism was identified for LK56, which required the Mediator subunit MDT-15/MED15 and NHR-49/HNF4 for its function. Interestingly, LK32, LK34, LK38, and LK56 also rescue C. elegans from P. aeruginosa in an agar-based assay, which uses different virulence factors and defense mechanisms. Rescue in an agar-based assay for LK38 entirely depended upon the PMK-1/p38 MAPK pathway. Three compounds, LK32, LK34, and LK56 also conferred resistance to Enterococcus faecalis, and the two lattermost, LK34 and LK56, also reduced pathogenesis from Staphylococcus aureus. This study supports a growing role for MDT-15 and NHR-49 in immune response and identifies 5 molecules that with significant potential for use as tools in the investigation of innate immunity. Author SummaryTwo trends moving in opposite directions (the increase in antimicrobial resistance and the decline of commercial interest in the discovery and development of novel antimicrobials) have precipitated a looming crisis: a nearly complete inability to safely and effectively treat bacterial infections. To avert this, new approaches in healthcare are needed. One approach that is receiving increasing attention is to stimulate host defense pathways, to improve the clearance of bacterial infections. We describe five small molecules that promote host resistance to infectious bacteria, at least partially by activating C. elegans innate immune pathways. Several are effective against both Gram-positive and Gram-negative pathogens. Three molecules, LK34, LK35, and LK38 have highly overlapping downstream target genes, suggesting that they act on common pathways, despite having distinct chemical structures. One of the compounds was mapped to the action of MDT-15/MED15 and NHR-49/HNF4, a pair of transcriptional regulators more generally associated with fatty acid metabolism, potentially highlighting a new link between these biological functions. These studies pave the way for future characterization of the anti-infective activity of the molecules in higher organisms and highlight the compounds potential utility for further investigation of immune modulation as a novel therapeutic approach.

genetics↗

bioRxiv2017

Interplay between mitochondria and diet mediates pathogen and stress resistance in C. elegans

Diet is a crucial determinant of organismal biology. Here we demonstrate the dramatic impact of a subtle shift in diet on the ability of Caenorhabditis elegans to survive pathogenic or abiotic stress. Interestingly, this shift occurs independently of canonical host defense pathways, arising instead from improvements in mitochondrial health. Using a variety of assays, we reveal that the most common C. elegans food source (E. coli OP50) results in a vitamin B12 deficiency that compromises mitochondrial homeostasis. Increasing B12 supply by feeding on E. coli HT115 or by supplementing bacterial media with methylcobalamin restored mitochondrial function, even if the bacteria were dead. B12 supplementation also efficiently increased host health without adversely affecting lifespan. Our study forges a molecular link between a dietary deficiency (nutrition/microbiota) and a physiological consequence (host sensitivity), using the host-microbiota-diet framework. The ubiquity of B12 deficiency (~10-40% of US adults) highlights the importance of our findings.

molecular biology↗