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Chembiochem : a European Journal of... Nov 2023Legionella pneumophila is the causative agent of Legionnaires' disease, a serious form of pneumonia. Its macrophage infectivity potentiator (Mip), a member of a highly...
Legionella pneumophila is the causative agent of Legionnaires' disease, a serious form of pneumonia. Its macrophage infectivity potentiator (Mip), a member of a highly conserved family of FK506-binding proteins (FKBPs), plays a major role in the proliferation of the gram-negative bacterium in host organisms. In this work, we test our library of >1000 FKBP-focused ligands for inhibition of LpMip. The [4.3.1]-bicyclic sulfonamide turned out as a highly preferred scaffold and provided the most potent LpMip inhibitors known so far. Selected compounds were non-toxic to human cells, displayed antibacterial activity and block bacterial proliferation in cellular infection-assays as well as infectivity in human lung tissue explants. The results confirm [4.3.1]-bicyclic sulfonamides as anti-legionellal agents, although their anti-infective properties cannot be explained by inhibition of LpMip alone.
Topics: Humans; Legionnaires' Disease; Tacrolimus Binding Proteins; Peptidylprolyl Isomerase; Bacterial Proteins; Legionella pneumophila; Legionella
PubMed: 37489700
DOI: 10.1002/cbic.202300442 -
International Journal of Biological... Dec 2023Macrophage infectivity potentiator (MIP) proteins are widespread in human pathogens including Legionella pneumophila, the causative agent of Legionnaires' disease and...
Macrophage infectivity potentiator (MIP) proteins are widespread in human pathogens including Legionella pneumophila, the causative agent of Legionnaires' disease and protozoans such as Trypanosoma cruzi. All MIP proteins contain a FKBP (FK506 binding protein)-like prolyl-cis/trans-isomerase domain that hence presents an attractive drug target. Some MIPs such as the Legionella pneumophila protein (LpMIP) have additional appendage domains of mostly unknown function. In full-length, homodimeric LpMIP, the N-terminal dimerization domain is linked to the FKBP-like domain via a long, free-standing stalk helix. Combining X-ray crystallography, NMR and EPR spectroscopy and SAXS, we elucidated the importance of the stalk helix for protein dynamics and inhibitor binding to the FKBP-like domain and bidirectional crosstalk between the different protein regions. The first comparison of a microbial MIP and a human FKBP in complex with the same synthetic inhibitor was made possible by high-resolution structures of LpMIP with a [4.3.1]-aza-bicyclic sulfonamide and provides a basis for designing pathogen-selective inhibitors. Through stereospecific methylation, the affinity of inhibitors to L. pneumophila and T. cruzi MIP was greatly improved. The resulting X-ray inhibitor-complex structures of LpMIP and TcMIP at 1.49 and 1.34 Å, respectively, provide a starting point for developing potent inhibitors against MIPs from multiple pathogenic microorganisms.
Topics: Humans; Legionella pneumophila; Scattering, Small Angle; X-Ray Diffraction; Bacterial Proteins; Tacrolimus Binding Proteins; Legionnaires' Disease; Macrophages
PubMed: 37633566
DOI: 10.1016/j.ijbiomac.2023.126366 -
Infection and Immunity Aug 2023Previously, we showed that Legionella pneumophila secretes rhizoferrin, a polycarboxylate siderophore that promotes bacterial growth in iron-deplete media and the murine...
Previously, we showed that Legionella pneumophila secretes rhizoferrin, a polycarboxylate siderophore that promotes bacterial growth in iron-deplete media and the murine lung. Yet, past studies failed to identify a role for the rhizoferrin biosynthetic gene () in L. pneumophila infection of host cells, suggesting the siderophore's importance was solely linked to extracellular survival. To test the possibility that rhizoferrin's relevance to intracellular infection was missed due to functional redundancy with the ferrous iron transport (FeoB) pathway, we characterized a new mutant lacking both and . This mutant was highly impaired for growth on bacteriological media that were only modestly depleted of iron, confirming that rhizoferrin-mediated ferric iron uptake and FeoB-mediated ferrous iron uptake are critical for iron acquisition. The mutant, but not its -containing complement, was also highly defective for biofilm formation on plastic surfaces, demonstrating a new role for the L. pneumophila siderophore in extracellular survival. Finally, the mutant, but not its complement containing , proved to be greatly impaired for growth in Acanthamoeba castellanii, , and human U937 cell macrophages, revealing that rhizoferrin does promote intracellular infection by L. pneumophila. Moreover, the application of purified rhizoferrin triggered cytokine production from the U937 cells. Rhizoferrin-associated genes were fully conserved across the many sequenced strains of L. pneumophila examined but were variably present among strains from the other species of . Outside of , the closest match to the L. pneumophila rhizoferrin genes was in Aquicella siphonis, another facultative intracellular parasite of amoebae.
Topics: Animals; Mice; Humans; Legionella pneumophila; Siderophores; Amoeba; U937 Cells; Bacterial Proteins; Iron; Macrophages; Biofilms
PubMed: 37428036
DOI: 10.1128/iai.00072-23 -
Analytical and Bioanalytical Chemistry Sep 2023Legionella pneumophila are pathogenic bacteria that can be found in high concentrations in artificial water systems like evaporative cooling towers, which have been the...
Legionella pneumophila are pathogenic bacteria that can be found in high concentrations in artificial water systems like evaporative cooling towers, which have been the source of frequent outbreaks in recent years. Since inhaled L. pneumophila can lead to Legionnaires' disease, the development of suitable sampling and rapid analysis strategies for these bacteria in aerosols is therefore of great relevance. In this work, different concentrations of viable L. pneumophila Sg 1 were nebulized and sampled by the cyclone sampler Coriolis® µ under defined conditions in a bioaerosol chamber. To quantify intact Legionella cells, the collected bioaerosols were subsequently analyzed by immunomagnetic separation coupled with flow cytometry (IMS-FCM) on the platform rqmicro.COUNT. For analytical comparison, measurements with qPCR and cultivation were performed. Limits of detection (LOD) of 2.9 × 10 intact cells m for IMS-FCM and 7.8 × 10 intact cells m for qPCR indicating a comparable sensitivity as in culture (LOD = 1.5 × 10 culturable cells m). Over a working range of 10 - 10 cells mL, the analysis of nebulized and collected aerosol samples with IMS-FCM and qPCR provides higher recovery rates and more consistent results than by cultivation. Overall, IMS-FCM is a suitable culture-independent method for quantification of L. pneumophila in bioaerosols and is promising for field application due to its simplicity in sample preparation.
Topics: Humans; Legionella pneumophila; Immunomagnetic Separation; Flow Cytometry; Respiratory Aerosols and Droplets; Legionnaires' Disease; Water Microbiology
PubMed: 37204446
DOI: 10.1007/s00216-023-04738-z -
Molecular Microbiology Mar 2024The phenomenon of host cell escape exhibited by intracellular pathogens is a remarkably versatile occurrence, capable of unfolding through lytic or non-lytic pathways.... (Review)
Review
The phenomenon of host cell escape exhibited by intracellular pathogens is a remarkably versatile occurrence, capable of unfolding through lytic or non-lytic pathways. Among these pathogens, the bacterium Legionella pneumophila stands out, having adopted a diverse spectrum of strategies to disengage from their host cells. A pivotal juncture that predates most of these host cell escape modalities is the initial escape from the intracellular compartment. This critical step is increasingly supported by evidence suggesting the involvement of several secreted pathogen effectors, including lytic proteins. In this intricate landscape, L. pneumophila emerges as a focal point for research, particularly concerning secreted phospholipases. While nestled within its replicative vacuole, the bacterium deftly employs both its type II (Lsp) and type IVB (Dot/Icm) secretion systems to convey phospholipases into either the phagosomal lumen or the host cell cytoplasm. Its repertoire encompasses numerous phospholipases A (PLA), including three enzymes-PlaA, PlaC, and PlaD-bearing the GDSL motif. Additionally, there are 11 patatin-like phospholipases A as well as PlaB. Furthermore, the bacterium harbors three extracellular phospholipases C (PLCs) and one phospholipase D. Within this comprehensive review, we undertake an exploration of the pivotal role played by phospholipases in the broader context of phagosomal and host cell egress. Moreover, we embark on a detailed journey to unravel the established and potential functions of the secreted phospholipases of L. pneumophila in orchestrating this indispensable process.
Topics: Humans; Phospholipases; Legionnaires' Disease; Vacuoles; Bacterial Proteins; Legionella pneumophila; Phospholipases A
PubMed: 37891705
DOI: 10.1111/mmi.15183 -
Travel Medicine and Infectious Disease 2023
Topics: Humans; Legionnaires' Disease; Bacteria; Legionella; Pneumonia; Anti-Bacterial Agents; Community-Acquired Infections
PubMed: 37858711
DOI: 10.1016/j.tmaid.2023.102654 -
Journal of Water and Health Sep 2023Legionnaires' disease (LD) is a severe pneumonia mainly caused by the bacterium Legionella pneumophila. Although many environmental sources of LD have been described,...
Legionnaires' disease (LD) is a severe pneumonia mainly caused by the bacterium Legionella pneumophila. Although many environmental sources of LD have been described, the sources of the majority of non-outbreak LD cases have not been identified. In several outbreaks in the Netherlands, wastewater treatment plants (WWTPs) were identified as the most likely source of infection. In this study, four criteria for Legionella growth and emission to air and surface waters were selected based on the literature and a risk matrix was drafted. An inventory was made of all WWTPs and their characteristics in the Netherlands. The risk matrix was applied to identify WWTPs at risk for Legionella growth and emission. Wastewater was collected at WWTPs with moderate to high risk for Legionella growth and emission. In 18% of the sampled WWTPs, Legionella spp. was detected using culture methods. The presented risk matrix can be used to assess the risks of Legionella growth and emission for WWTPs and support surveillance by prioritizing WWTPs. When Legionella is detected in the wastewater, it is recommended to take action to prevent emission to air or discharge on surface waters and, if possible, reduce the Legionella concentration.
Topics: Humans; Legionella; Wastewater; Legionnaires' Disease; Legionella pneumophila; Disease Outbreaks
PubMed: 37756196
DOI: 10.2166/wh.2023.164 -
BioRxiv : the Preprint Server For... Dec 2023Bacterial adhesion is a fundamental process which enables colonisation of niche environments and is key for infection. However, in , the causative agent of Legionnaires'...
Bacterial adhesion is a fundamental process which enables colonisation of niche environments and is key for infection. However, in , the causative agent of Legionnaires' disease, these processes are not well understood. The collagen-like protein (Lcl) is an extracellular peripheral membrane protein that recognises sulphated glycosaminoglycans (GAGs) on the surface of eukaryotic cells, but also stimulates bacterial aggregation in response to divalent cations. Here we report the crystal structure of the Lcl C-terminal domain (Lcl-CTD) and present a model for intact Lcl. Our data reveal that Lcl-CTD forms an unusual dynamic trimer arrangement with a positively charged external surface and a negatively charged solvent exposed internal cavity. Through Molecular Dynamics (MD) simulations, we show how the GAG chondroitin-4-sulphate associates with the Lcl-CTD surface via unique binding modes. Our findings show that Lcl homologs are present across both the Pseudomonadota and Fibrobacterota-Chlorobiota-Bacteroidota phyla and suggest that Lcl may represent a versatile carbohydrate binding mechanism.
PubMed: 38106198
DOI: 10.1101/2023.12.10.570962 -
Puerto Rico Health Sciences Journal Dec 2023Case of a 44-year-old woman with past medical history of dermatomyositis who had been on Methrotexate therapy who became infected with Legionella pneumophila after...
Case of a 44-year-old woman with past medical history of dermatomyositis who had been on Methrotexate therapy who became infected with Legionella pneumophila after staying at a home rental, known commercially as an Airbnb. The patient presented to the ER with complaints of general malaise and subsequently developed sepsis with respiratory failure requiring intubation. CT scan confirmed the diagnosis of pneumonia and through extensive research L. pneumophila was identified using a pneumonia panel that works by identifying nucleic acids of fastidious organisms that are difficult and take long to grow by culture. As the patient's pneumonia progressed to ARDS and her clinical impression worsened, pronation technique was begun in addition to her course of antibiotics that were already being administered. The patient began showing significant improvement in her clinical picture and was extubated with progressive recovery.
Topics: Humans; Female; Adult; Legionnaires' Disease; Legionella pneumophila; Pneumonia; Anti-Bacterial Agents; Tomography, X-Ray Computed
PubMed: 38104291
DOI: No ID Found -
MBio Jun 2024The causative agent of Legionnaires' disease, , is an environmental bacterium, that replicates in macrophages, parasitizes amoeba, and forms biofilms. employs the...
UNLABELLED
The causative agent of Legionnaires' disease, , is an environmental bacterium, that replicates in macrophages, parasitizes amoeba, and forms biofilms. employs the quorum sensing (Lqs) system and the transcription factor LvbR to control various bacterial traits, including virulence and biofilm architecture. LvbR negatively regulates the nitric oxide (NO) receptor Hnox1, linking quorum sensing to NO signaling. Here, we assessed the response of to NO and investigated bacterial receptors underlying this process. Chemical NO donors, such as dipropylenetriamine (DPTA) NONOate and sodium nitroprusside (SNP), delayed and reduced the expression of the promoters for flagellin (P) and the 6S small regulatory RNA (P). Marker-less mutant strains lacking individual (Hnox1, Hnox2, or NosP) or all three NO receptors (triple knockout, TKO) grew like the parental strain in media. However, in the TKO strain, the reduction of P expression by DPTA NONOate was less pronounced, suggesting that the NO receptors are implicated in NO signaling. In the Δ mutant, the promoter was upregulated, indicating that NosP negatively regulates LvbR. The single and triple NO receptor mutant strains were impaired for growth in phagocytes, and phenotypic heterogeneity of non-growing/growing bacteria in amoebae was regulated by the NO receptors. The single NO receptor and TKO mutant strains showed altered biofilm architecture and lack of response of biofilms to NO. In summary, we provide evidence that regulates virulence, intracellular phenotypic heterogeneity, and biofilm formation through NO and three functionally non-redundant NO receptors, Hnox1, Hnox2, and NosP.
IMPORTANCE
The highly reactive diatomic gas molecule nitric oxide (NO) is produced by eukaryotes and bacteria to promote short-range and transient signaling within and between neighboring cells. Despite its importance as an inter-kingdom and intra-bacterial signaling molecule, the bacterial response and the underlying components of the signaling pathways are poorly characterized. The environmental bacterium forms biofilms and replicates in protozoan and mammalian phagocytes. harbors three putative NO receptors, one of which crosstalks with the quorum sensing (Lqs)-LvbR network to regulate various bacterial traits, including virulence and biofilm architecture. In this study, we used pharmacological, genetic, and cell biological approaches to assess the response of to NO and to demonstrate that the putative NO receptors are implicated in NO detection, bacterial replication in phagocytes, intracellular phenotypic heterogeneity, and biofilm formation.
Topics: Biofilms; Legionella pneumophila; Nitric Oxide; Signal Transduction; Virulence; Gene Expression Regulation, Bacterial; Bacterial Proteins; Phenotype; Macrophages; Quorum Sensing
PubMed: 38682908
DOI: 10.1128/mbio.00710-24