Refine
Has Fulltext
- yes (19) (remove)
Is part of the Bibliography
- yes (19)
Year of publication
Document Type
- Journal article (19) (remove)
Language
- English (19) (remove)
Keywords
- Staphylococcus aureus (5)
- synthesis (4)
- vancomycin (4)
- antibacterial activity (3)
- antibiotics (3)
- epithelial cells (3)
- inhibition (3)
- substituent (3)
- NSG (2)
- cytotoxicity (2)
Institute
- Institut für Molekulare Infektionsbiologie (19) (remove)
EU-Project number / Contract (GA) number
- ERC335568 (1)
\(Enterococcus\) species cause increasing numbers of infections in hospitals. They contribute to the increasing mortality rates, mostly in patients with comorbidities, who suffer from severe diseases. \(Enterococcus\) resistances against most antibiotics have been described, including novel antibiotics. Therefore, there is an ongoing demand for novel types of antibiotics that may overcome bacterial resistances. We discovered a novel class of antibiotics resulting from a simple one-pot reaction of indole and \(o\)-phthaldialdehyde. Differently substituted indolyl benzocarbazoles were yielded. Both the indole substitution and the positioning at the molecular scaffold influence the antibacterial activity towards the various strains of \(Enterococcus\) species with the highest relevance to nosocomial infections. Structure-activity relationships are discussed, and the first lead compounds were identified as also being effective in the case of a vancomycin resistance.
Community-acquired (CA) Staphylococcus aureus cause various diseases even in healthy individuals. Enhanced virulence of CA-strains is partly attributed to increased production of toxins such as phenol-soluble modulins (PSM). The pathogen is internalized efficiently by mammalian host cells and intracellular S. aureus has recently been shown to contribute to disease. Upon internalization, cytotoxic S. aureus strains can disrupt phagosomal membranes and kill host cells in a PSM-dependent manner. However, PSM are not sufficient for these processes. Here we screened for factors required for intracellular S. aureus virulence. We infected escape reporter host cells with strains from an established transposon mutant library and detected phagosomal escape rates using automated microscopy. We thereby, among other factors, identified a non-ribosomal peptide synthetase (NRPS) to be required for efficient phagosomal escape and intracellular survival of S. aureus as well as induction of host cell death. By genetic complementation as well as supplementation with the synthetic NRPS product, the cyclic dipeptide phevalin, wild-type phenotypes were restored. We further demonstrate that the NRPS is contributing to virulence in a mouse pneumonia model. Together, our data illustrate a hitherto unrecognized function of the S. aureus NRPS and its dipeptide product during S. aureus infection.
Metabolic conversion of CI-1040 turns a cellular MEK-inhibitor into an antibacterial compound
(2018)
Influenza virus (IV) infections cause severe respiratory illnesses that can be complicated by bacterial super-infections. Previously, we identified the cellular Raf-MEK-ERK cascade as a promising antiviral target. Inhibitors of MEK, such as CI-1040, showed potent antiviral activity. However, it remained unclear if this inhibitor and its active form, ATR-002, might sensitize host cells to either IV or secondary bacterial infections. To address these questions, we studied the anti-pathogen activity of ATR-002 in comparison to CI-1040, particularly, its impact on Staphylococcus aureus (S. aureus), which is a major cause of IV super-infections. We analysed IV and S. aureus titres in vitro during super-infection in the presence and absence of the drugs and characterized the direct impact of ATR-002 on bacterial growth and phenotypic changes. Importantly, neither CI-1040 nor ATR-002 treatment led to increased bacterial titres during super-infection, indicating that the drug does not sensitize cells for bacterial infection. In contrast, we rather observed reduced bacterial titres in presence of ATR-002. Surprisingly, ATR-002 also led to reduced bacterial growth in suspension cultures, reduced stress- and antibiotic tolerance without resistance induction. Our data identified for the first time that a particular MEK-inhibitor metabolite exhibits direct antibacterial activity, which is likely due to interference with the bacterial PknB kinase/Stp phosphatase signalling system.
Ongoing resistance developments against antibiotics that also affect last-resort antibiotics require novel antibacterial compounds. Strategies to discover such novel structures have been dimerization or hybridization of known antibacterial agents. We found novel antibacterial agents by dimerization of indols and hybridization with carbazoles. They were obtained in a simple one-pot reaction as bisindole tetrahydrocarbazoles. Further oxidation led to bisindole carbazoles with varied substitutions of both the indole and the carbazole scaffold. Both the tetrahydrocarbazoles and the carbazoles have been evaluated in various S. aureus strains, including MRSA strains. Those 5-cyano substituted derivatives showed best activities as determined by MIC values. The tetrahydrocarbazoles partly exceed the activity of the carbazole compounds and thus the activity of the used standard antibiotics. Thus, promising lead compounds could be identified for further studies.
Meningococcal meningitis is a severe central nervous system infection that occurs when Neisseria meningitidis (Nm) penetrates brain endothelial cells (BECs) of the meningeal blood-cerebrospinal fluid barrier. As a human-specific pathogen, in vivo models are greatly limited and pose a significant challenge. In vitro cell models have been developed, however, most lack critical BEC phenotypes limiting their usefulness. Human BECs generated from induced pluripotent stem cells (iPSCs) retain BEC properties and offer the prospect of modeling the human-specific Nm interaction with BECs. Here, we exploit iPSC-BECs as a novel cellular model to study Nm host-pathogen interactions, and provide an overview of host responses to Nm infection. Using iPSC-BECs, we first confirmed that multiple Nm strains and mutants follow similar phenotypes to previously described models. The recruitment of the recently published pilus adhesin receptor CD147 underneath meningococcal microcolonies could be verified in iPSC-BECs. Nm was also observed to significantly increase the expression of pro-inflammatory and neutrophil-specific chemokines IL6, CXCL1, CXCL2, CXCL8, and CCL20, and the secretion of IFN-γ and RANTES. For the first time, we directly observe that Nm disrupts the three tight junction proteins ZO-1, Occludin, and Claudin-5, which become frayed and/or discontinuous in BECs upon Nm challenge. In accordance with tight junction loss, a sharp loss in trans-endothelial electrical resistance, and an increase in sodium fluorescein permeability and in bacterial transmigration, was observed. Finally, we established RNA-Seq of sorted, infected iPSC-BECs, providing expression data of Nm-responsive host genes. Altogether, this model provides novel insights into Nm pathogenesis, including an impact of Nm on barrier properties and tight junction complexes, and suggests that the paracellular route may contribute to Nm traversal of BECs.
Sepsis caused by Gram-positive bacterial pathogens is a major fatal disease but its molecular basis remains elusive. Toll-like receptor 2 (TLR2) has been implicated in the orchestration of inflammation and sepsis but its role appears to vary for different pathogen species and clones. Accordingly, Staphylococcus aureus clinical isolates differ substantially in their capacity to activate TLR2. Here we show that strong TLR2 stimulation depends on high-level production of phenol-soluble modulin (PSM) peptides in response to the global virulence activator Agr. PSMs are required for mobilizing lipoproteins, the TLR2 agonists, from the staphylococcal cytoplasmic membrane. Notably, the course of sepsis caused by PSM-deficient S. aureus is similar in wild-type and TLR2-deficient mice, but TLR2 is required for protection of mice against PSM-producing S. aureus. Thus, a crucial role of TLR2 depends on agonist release by bacterial surfactants. Modulation of this process may lead to new therapeutic strategies against Gram-positive infections.
Background
The emergence of antibiotic resistant bacteria in recent decades has highlighted the importance of developing new drugs to treat infections. However, in addition to the design of new drugs, the development of accurate preclinical testing methods is essential. In vivo imaging technologies such as bioluminescence imaging (BLI) or magnetic resonance imaging (MRI) are promising approaches. In a previous study, we showed the effectiveness of \(^{19}\)F MRI using perfluorocarbon (PFC) emulsions for detecting the site of Staphylococcus aureus infection. In the present follow-up study, we investigated the use of this method for in vivo visualization of the effects of antibiotic therapy.
Methods/Principal findings
Mice were infected with S. aureus Xen29 and treated with 0.9% NaCl solution, vancomycin or linezolid. Mock treatment led to the highest bioluminescence values during infection followed by vancomycin treatment. Counting the number of colony-forming units (cfu) at 7 days post-infection (p.i.) showed the highest bacterial burden for the mock group and the lowest for the linezolid group. Administration of PFCs at day 2 p.i. led to the accumulation of \(^{19}\)F at the rim of the abscess in all mice (in the shape of a hollow sphere), and antibiotic treatment decreased the \(^{19}\)F signal intensity and volume. Linezolid showed the strongest effect. The BLI, cfu, and MRI results were comparable.
Conclusions
\(^{19}\)F-MRI with PFCs is an effective non-invasive method for assessing the effects of antibiotic therapy in vivo. This method does not depend on pathogen specific markers and can therefore be used to estimate the efficacy of antibacterial therapy against a broad range of clinically relevant pathogens, and to localize sites of infection.
Background:
During the last years, (19)F-MRI and perfluorocarbon nanoemulsion (PFC) emerged as a powerful contrast agent methodology to track cells and to visualize inflammation. We applied this new modality to visualize deep tissue abscesses during acute and chronic phase of inflammation caused by Staphylococcus aureus infection.
Methodology and Principal Findings:
In this study, a murine thigh infection model was used to induce abscess formation and PFC or CLIO (cross linked ironoxides) was administered during acute or chronic phase of inflammation. 24 h after inoculation, the contrast agent accumulation was imaged at the site of infection by MRI. Measurements revealed a strong accumulation of PFC at the abscess rim at acute and chronic phase of infection. The pattern was similar to CLIO accumulation at chronic phase and formed a hollow sphere around the edema area. Histology revealed strong influx of neutrophils at the site of infection and to a smaller extend macrophages during acute phase and strong influx of macrophages at chronic phase of inflammation.
Conclusion and Significance:
We introduce (19)F-MRI in combination with PFC nanoemulsions as a new platform to visualize abscess formation in a murine thigh infection model of S. aureus. The possibility to track immune cells in vivo by this modality offers new opportunities to investigate host immune response, the efficacy of antibacterial therapies and the influence of virulence factors for pathogenesis.
MRSA (Methicillin-resistant Staphylococcus aureus) is the second-leading cause of deaths by antibiotic-resistant bacteria globally, with more than 100,000 attributable deaths annually. Despite the high urgency to develop a vaccine to control this pathogen, all clinical trials with pre-clinically effective candidates failed so far. The recent development of “humanized” mice might help to edge the pre-clinical evaluation closer to the clinical situation and thus close this gap. We infected humanized NSG mice (huNSG: (NOD)-scid IL2R\(_γ\)\(^{null}\) mice engrafted with human CD34+ hematopoietic stem cells) locally with S. aureus USA300 LAC* lux into the thigh muscle in order to investigate the human immune response to acute and chronic infection. These mice proved not only to be more susceptible to MRSA infection than wild-type or “murinized” mice, but displayed furthermore inferior survival and signs of systemic infection in an otherwise localized infection model. The rate of humanization correlated directly with the severity of disease and survival of the mice. Human and murine cytokine levels in blood and at the primary site of infection were strongly elevated in huNSG mice compared to all control groups. And importantly, differences in human and murine immune cell lineages surfaced during the infection, with human monocyte and B cell numbers in blood and bone marrow being significantly reduced at the later time point of infection. Murine monocytes in contrast behaved conversely by increasing cell numbers. This study demonstrates significant differences in the in vivo behavior of human and murine cells towards S. aureus infection, which might help to sharpen the translational potential of pre-clinical models for future therapeutic approaches.
Next-generation humanized NSG-SGM3 mice are highly susceptible to Staphylococcus aureus infection
(2023)
Humanized hemato-lymphoid system mice, or humanized mice, emerged in recent years as a promising model to study the course of infection of human-adapted or human-specific pathogens. Though Staphylococcus aureus infects and colonizes a variety of species, it has nonetheless become one of the most successful human pathogens of our time with a wide armory of human-adapted virulence factors. Humanized mice showed increased vulnerability to S. aureus compared to wild type mice in a variety of clinically relevant disease models. Most of these studies employed humanized NSG (NOD-scid IL2Rgnull) mice which are widely used in the scientific community, but show poor human myeloid cell reconstitution. Since this immune cell compartment plays a decisive role in the defense of the human immune system against S. aureus, we asked whether next-generation humanized mice, like NSG-SGM3 (NOD-scid IL2Rgnull-3/GM/SF) with improved myeloid reconstitution, would prove to be more resistant to infection. To our surprise, we found the contrary when we infected humanized NSG-SGM3 (huSGM3) mice with S. aureus: although they had stronger human immune cell engraftment than humanized NSG mice, particularly in the myeloid compartment, they displayed even more pronounced vulnerability to S. aureus infection. HuSGM3 mice had overall higher numbers of human T cells, B cells, neutrophils and monocytes in the blood and the spleen. This was accompanied by elevated levels of pro-inflammatory human cytokines in the blood of huSGM3 mice. We further identified that the impaired survival of huSGM3 mice was not linked to higher bacterial burden nor to differences in the murine immune cell repertoire. Conversely, we could demonstrate a correlation of the rate of humanization and the severity of infection. Collectively, this study suggests a detrimental effect of the human immune system in humanized mice upon encounter with S. aureus which might help to guide future therapy approaches and analysis of virulence mechanisms.