scholarly journals Klebsiella pneumoniae Expressing VIM-1 Metallo-β-Lactamase Is Resensitized to Cefotaxime via Thiol-Mediated Zinc Chelation

2019 ◽  
Vol 88 (1) ◽  
Author(s):  
Harpa Karadottir ◽  
Maarten Coorens ◽  
Zhihai Liu ◽  
Yang Wang ◽  
Birgitta Agerberth ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Ex Vivo ◽  
Reductase Activity ◽  
Urine Samples ◽  
Dependent Manner ◽  
Content Type ◽  
Endogenous Factors ◽  
Reducing Ability ◽  
Zinc Chelation ◽  
Ht 29

ABSTRACT Antibiotic-resistant Klebsiella pneumoniae isolates constitute a great clinical challenge. One important resistance mechanism in K. pneumoniae is the metallo-β-lactamases (MBLs), which require zinc for their function. Thus, zinc chelation could be a strategy to resensitize K. pneumoniae to β-lactams. However, the potential role for endogenous zinc chelators for this purpose remains to be explored. The aim was to search for endogenous factors that could resensitize MBL-expressing K. pneumoniae to cefotaxime (CTX). Clinical K. pneumoniae isolates expressing different MBLs were screened for sensitivity to CTX in supernatants from human HT-29 colonic epithelial cells. Factors influencing CTX susceptibility were isolated and identified with chromatographic and biochemical methods. Free zinc was measured with a Zinquin assay, the thiol content was assessed with a fluorometric thiol assay, and the reducing ability of the supernatant was measured with a fluorescent l-cystine probe. Urine samples from healthy volunteers were used to validate findings ex vivo. VIM-1-expressing K. pneumoniae regained susceptibility to CTX when grown in supernatants from HT-29 cells. This effect was mediated via free thiols in the supernatant, including l-cysteine, and could be prevented by inhibiting thioredoxin reductase activity in the supernatant. Free thiols in urine samples appeared to have a similar function in restoring CTX activity against VIM-1-expressing K. pneumoniae in a zinc-dependent manner. We have identified l-cysteine as an endogenous zinc chelator resulting in the resensitization of VIM-1-expressing K. pneumoniae to CTX. These results suggest that natural zinc chelators in combination with conventional antibiotics could be used to treat infections caused by VIM-1-expressing pathogens.

scholarly journals Allergic Airway Inflammation Decreases Lung Bacterial Burden following Acute Klebsiella pneumoniae Infection in a Neutrophil- and CCL8-Dependent Manner

2014 ◽  
Vol 82 (9) ◽  
pp. 3723-3739 ◽  
Author(s):  
Daniel E. Dulek ◽  
Dawn C. Newcomb ◽  
Kasia Goleniewska ◽  
Jaqueline Cephus ◽  
Weisong Zhou ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Airway Inflammation ◽  
Ex Vivo ◽  
Allergic Airway Inflammation ◽  
Dependent Manner ◽  
Content Type ◽  
Th2 Cytokine ◽  
Interleukin 17A ◽  
Th17 Cytokines

ABSTRACTThe Th17 cytokines interleukin-17A (IL-17A), IL-17F, and IL-22 are critical for the lung immune response to a variety of bacterial pathogens, includingKlebsiella pneumoniae. Th2 cytokine expression in the airways is a characteristic feature of asthma and allergic airway inflammation. The Th2 cytokines IL-4 and IL-13 diminishex vivoandin vivoIL-17A protein expression by Th17 cells. To determine the effect of IL-4 and IL-13 on IL-17-dependent lung immune responses to acute bacterial infection, we developed a combined model in which allergic airway inflammation and lung IL-4 and IL-13 expression were induced by ovalbumin sensitization and challenge prior to acute lung infection withK. pneumoniae. We hypothesized that preexisting allergic airway inflammation decreases lung IL-17A expression and airway neutrophil recruitment in response to acuteK. pneumoniaeinfection and thereby increases the lungK. pneumoniaeburden. As hypothesized, we found that allergic airway inflammation decreased the number ofK. pneumoniae-induced airway neutrophils and lung IL-17A, IL-17F, and IL-22 expression. Despite the marked reduction in postinfection airway neutrophilia and lung expression of Th17 cytokines, allergic airway inflammation significantly decreased the lungK. pneumoniaeburden and postinfection mortality. We showed that the decreased lungK. pneumoniaeburden was independent of IL-4, IL-5, and IL-17A and partially dependent on IL-13 and STAT6. Additionally, we demonstrated that the decreased lungK. pneumoniaeburden associated with allergic airway inflammation was both neutrophil and CCL8 dependent. These findings suggest a novel role for CCL8 in lung antibacterial immunity againstK. pneumoniaeand suggest new mechanisms of orchestrating lung antibacterial immunity.

scholarly journals Highly Tigecycline-Resistant Klebsiella pneumoniae Sequence Type 11 (ST11) and ST147 Isolates from Companion Animals

2017 ◽  
Vol 61 (6) ◽  
Author(s):  
Cristina M. Ovejero ◽  
Jose Antonio Escudero ◽  
Daniel Thomas-Lopez ◽  
Andreas Hoefer ◽  
Gabriel Moyano ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Companion Animals ◽  
Sequence Type ◽  
Urine Samples ◽  
Content Type

ABSTRACT In this study, we characterized two tigecycline-resistant Klebsiella pneumoniae isolates from dog urine samples. The isolates were genetically unrelated, belonging to sequence type 11 (ST11) and ST147, both classically related to human isolates. To the best of our knowledge, this is the first identification of tigecycline-resistant isolates from animals. We unveil here the worrisome circulation among animals of bacterial clones resistant to this last-resort antibiotic.

scholarly journals Chloroquine Potentiates Primaquine Activity against Active and Latent Hepatic Plasmodia Ex Vivo: Potentials and Pitfalls

2020 ◽  
Vol 65 (1) ◽  
pp. e01416-20
Author(s):  
Laurent Dembélé ◽  
Jean-François Franetich ◽  
Valérie Soulard ◽  
Nadia Amanzougaghene ◽  
Shahin Tajeri ◽  
...  
Keyword(s):  
Cell Line ◽  
Ex Vivo ◽  
Plasmodium Yoelii ◽  
Human Hepatocytes ◽  
Dependent Manner ◽  
Primary Hepatocytes ◽  
Content Type ◽  
Primary Human Hepatocytes ◽  
Hepatocarcinoma Cell ◽  
Hepatocarcinoma Cell Line

ABSTRACTFor a long while, 8-aminoquinoline compounds have been the only therapeutic agents against latent hepatic malaria parasites. These have poor activity against the blood-stage plasmodia causing acute malaria and must be used in conjunction with partner blood schizontocidal agents. We examined the impacts of one such agent, chloroquine, upon the activity of primaquine, an 8-aminoquinoline, against hepatic stages of Plasmodium cynomolgi, Plasmodium yoelii, Plasmodium berghei, and Plasmodium falciparum within several ex vivo systems—primary hepatocytes of Macaca fascicularis, primary human hepatocytes, and stably transformed human hepatocarcinoma cell line HepG2. Primaquine exposures to formed hepatic schizonts and hypnozoites of P. cynomolgi in primary simian hepatocytes exhibited similar 50% inhibitory concentration (IC50) values near 0.4 μM, whereas chloroquine in the same system exhibited no inhibitory activities. Combining chloroquine and primaquine in this system decreased the observed primaquine IC50 for all parasite forms in a chloroquine dose-dependent manner by an average of 18-fold. Chloroquine also decreased the primaquine IC50 against hepatic P. falciparum in primary human hepatocytes, P. berghei in simian primary hepatocytes, and P. yoelii in primary human hepatocytes. Chloroquine had no impact on primaquine IC50 against P. yoelii in HepG2 cells and, likewise, had no impact on the IC50 of atovaquone (hepatic schizontocide) against P. falciparum in human hepatocytes. We describe important sources of variability in the potentiation of primaquine activity by chloroquine in these systems. Chloroquine potentiated primaquine activity against hepatic forms of several plasmodia. We conclude that chloroquine specifically potentiated 8-aminoquinoline activities against active and dormant hepatic-stage plasmodia in normal primary hepatocytes but not in a hepatocarcinoma cell line.

scholarly journals Protection against Klebsiella pneumoniae Using Lithium Chloride in an Intragastric Infection Model

2014 ◽  
Vol 59 (3) ◽  
pp. 1525-1533 ◽  
Author(s):  
Nina Tsao ◽  
Chih-Feng Kuo ◽  
Ching-Chen Chiu ◽  
Wei-Chen Lin ◽  
Wan-Hui Huang ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Lithium Chloride ◽  
Blood Levels ◽  
Infection Model ◽  
Dependent Manner ◽  
Toxic Dose ◽  
Content Type ◽  
Bipolar Mood Disorder ◽  
Liver Tissues

ABSTRACTIntragastricKlebsiella pneumoniaeinfections of mice can cause liver abscesses, necrosis of liver tissues, and bacteremia. Lithium chloride, a widely prescribed drug for bipolar mood disorder, has been reported to possess anti-inflammatory properties. Using an intragastric infection model, the effects of LiCl onK. pneumoniaeinfections were examined. Providing mice with drinking water containing LiCl immediately after infection protected them fromK. pneumoniae-induced death and liver injuries, such as necrosis of liver tissues, as well as increasing blood levels of aspartate aminotransferase and alanine aminotransferase, in a dose-dependent manner. LiCl administered as late as 24 h postinfection still provided protection. Monitoring of the LiCl concentrations in the sera ofK. pneumoniae-infected mice showed that approximately 0.33 mM LiCl was the most effective dose for protecting mice against infections, which is lower than the clinically toxic dose of LiCl. Surveys of bacterial counts and cytokine expression levels in LiCl-treated mice revealed that both were effectively inhibited in blood and liver tissues. Usingin vitroassays, we found that LiCl (5 μM to 1 mM) did not directly interfere with the growth ofK. pneumoniaebut madeK. pneumoniaecells lose the mucoid phenotype and become more susceptible to macrophage killing. Furthermore, low doses of LiCl also partially enhanced the bactericidal activity of macrophages. Taken together, these data suggest that LiCl is an alternative therapeutic agent forK. pneumoniae-induced liver infections.

scholarly journals Analysis of Endothelial Adherence of Bartonella henselae and Acinetobacter baumannii Using a Dynamic HumanEx VivoInfection Model

2015 ◽  
Vol 84 (3) ◽  
pp. 711-722 ◽  
Author(s):  
Marko Weidensdorfer ◽  
Ju Ik Chae ◽  
Celestine Makobe ◽  
Julia Stahl ◽  
Beate Averhoff ◽  
...  
Keyword(s):  
Acinetobacter Baumannii ◽  
Pathogenic Bacteria ◽  
Ex Vivo ◽  
Bartonella Henselae ◽  
Dependent Manner ◽  
Content Type ◽  
Human Pathogenic Bacteria ◽  
Infection Models ◽  
Animal Infection

Bacterial adherence determines the virulence of many human-pathogenic bacteria. Experimental approaches elucidating this early infection event in greater detail have been performed using mainly methods of cellular microbiology. However,in vitroinfections of cell monolayers reflect thein vivosituation only partially, and animal infection models are not available for many human-pathogenic bacteria. Therefore,ex vivoinfection of human organs might represent an attractive method to overcome these limitations. We infected whole human umbilical cordsex vivowithBartonella henselaeorAcinetobacter baumanniiunder dynamic flow conditions mimicking thein vivoinfection situation of human endothelium. For this purpose, methods for quantifying endothelium-adherent wild-type and trimeric autotransporter adhesin (TAA)-deficient bacteria were set up. Data revealed that (i)A. baumanniibinds in a TAA-dependent manner to endothelial cells, (ii) this organ infection model led to highly reproducible adherence rates, and furthermore, (iii) this model allowed to dissect the biological function of TAAs in the natural course of human infections. These findings indicate that infection models usingex vivohuman tissue samples (“organ microbiology”) might be a valuable tool in analyzing bacterial pathogenicity with the capacity to replace animal infection models at least partially.

scholarly journals Metabolite Transporter PEG344 Is Required for Full Virulence of Hypervirulent Klebsiella pneumoniae Strain hvKP1 after Pulmonary but Not Subcutaneous Challenge

2017 ◽  
Vol 85 (10) ◽  
Author(s):  
Jeffrey Bulger ◽  
Ulrike MacDonald ◽  
Ruth Olson ◽  
Janet Beanan ◽  
Thomas A. Russo
Keyword(s):  
Klebsiella Pneumoniae ◽  
Virulence Factors ◽  
Ex Vivo ◽  
Infection Model ◽  
Virulence Plasmid ◽  
Content Type ◽  
Model Studies ◽  
Life Threatening ◽  
Cd1 Mice

ABSTRACT Hypervirulent Klebsiella pneumoniae (hvKP) is an emerging pathotype that is capable of causing tissue-invasive and organ- and life-threatening infections in healthy individuals from the community. Knowledge on the virulence factors specific to hvKP is limited. In this report, we describe a new factor (PEG344) that increases the virulence of hvKP strain hvKP1. peg-344 is present on the hvKP1 virulence plasmid, is broadly prevalent among hvKP strains, and has increased RNA abundance when grown in human ascites. An isogenic derivative of hvKP1 (hvKP1Δpeg-344) was constructed and compared with its wild-type parent strain in in vitro, ex vivo, and infection model studies. Both survival and competition experiments with outbred CD1 mice demonstrated that PEG344 was required for full virulence after pulmonary challenge but, interestingly, not after subcutaneous challenge. In silico analysis suggested that PEG344 serves as an inner membrane transporter. Compared to hvKP1, a small but significant decrease in the growth/survival of hvKP1Δpeg-344 was observed in human ascites, but resistance to the bactericidal activity of complement was similar. These data suggested that PEG344 may transport an unidentified growth factor present in ascites. The data presented are important since they expand our limited knowledge base on virulence factors unique to hvKP, which is needed to lay the groundwork for translational approaches to prevent or treat these devastating infections.

scholarly journals Influence of Cranberry Extract on Tamm-Horsfall Protein in Human Urine and its Antiadhesive Activity Against Uropathogenic Escherichia coli

Planta Medica ◽  
2018 ◽  
Vol 85 (02) ◽  
pp. 126-138 ◽  
Author(s):  
Birte Scharf ◽  
Jandirk Sendker ◽  
Ulrich Dobrindt ◽  
Andreas Hensel
Keyword(s):  
Escherichia Coli ◽  
Bacterial Adhesion ◽  
Ex Vivo ◽  
Transmembrane Protein ◽  
Bacterial Attachment ◽  
Urine Samples ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Upec Strain ◽  
Bladder Cells

AbstractLC-MS characterized cranberry extract from the fruits of Vaccinium macrocarpon inhibited under in vitro conditions the bacterial adhesion of Escherichia coli strain 2980 uropathogenic E. coli (UPEC strains UTI89, NU14) to T24 bladder cells and adhesion of UPEC strain CFT073 to A498 kidney cells in a concentration-dependent manner. Within a biomedical study, urine samples from 16 volunteers (8 male, 8 female) consuming cranberry extract for 7 d (900 mg/d) were analyzed for potential antiadhesive activity against UPEC by ex vivo experiments. Results indicated inhibition of adhesion of UPEC strain UTI89 to human T24 bladder cells. Subgroup analysis proved significant inhibition of bacterial adhesion in case of urine samples obtained from male volunteers while female urine did not influence the bacterial attachment. Differences between antiadhesive capacity of urine samples from male/female volunteers were significant. Protein analysis of the urine samples indicated increased amounts of Tamm-Horsfall protein (THP, syn. uromodulin) in the active samples. Inhibition of bacterial adhesion by the urine samples was correlated to the respective amount of THP. As it is known that THP, a highly mannosylated glycoprotein, strongly interacts with FimH of UPEC, this will lead to a decreased interaction with uroplakin, a FimH-binding transmembrane protein of urothelial lining cells. From these data it can be concluded that the antiadhesive effect of cranberry after oral intake is not only related to the direct inhibition of bacterial adhesins by extract compounds but is additionally due to an induction of antiadhesive THP in the kidney.

scholarly journals Klebsiella pneumoniae Yersiniabactin Promotes Respiratory Tract Infection through Evasion of Lipocalin 2

2011 ◽  
Vol 79 (8) ◽  
pp. 3309-3316 ◽  
Author(s):  
Michael A. Bachman ◽  
Jennifer E. Oyler ◽  
Samuel H. Burns ◽  
Mélissa Caza ◽  
François Lépine ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Respiratory Tract ◽  
Ex Vivo ◽  
Iron Acquisition ◽  
Opportunistic Pathogen ◽  
Respiratory Pathogen ◽  
Lipocalin 2 ◽  
Content Type ◽  
Tract Infections ◽  
Disseminated Disease

ABSTRACTKlebsiella pneumoniaeis a pathogen of increasing concern because of multidrug resistance, especially due toK. pneumoniaecarbapenemases (KPCs).K. pneumoniaemust acquire iron to replicate, and it utilizes iron-scavenging siderophores, such as enterobactin (Ent). The innate immune protein lipocalin 2 (Lcn2) is able to specifically bind Ent and disrupt iron acquisition. To determine whetherK. pneumoniaemust produce Lcn2-resistant siderophores to cause disease, we examined siderophore production by clinical isolates (n= 129) from respiratory, urine, blood, and stool samples and by defined siderophore mutants through genotyping and liquid chromatography-mass spectrometry. Three categories ofK. pneumoniaeisolates were identified: enterobactin positive (Ent+) (81%), enterobactin and yersiniabactin positive (Ent+Ybt+) (17%), and enterobactin and salmochelin (glycosylated Ent) positive (Ent+gly-Ent+) with or without Ybt (2%). Ent+Ybt+strains were significantly overrepresented among respiratory tract isolates (P= 0.0068) and β-lactam-resistant isolates (P= 0.0019), including the epidemic KPC-producing clone multilocus sequence type 258 (ST258). Inex vivogrowth assays, gly-Ent but not Ybt allowed evasion of Lcn2 in human serum, whereas siderophores were dispensable for growth in human urine. In a murine pneumonia model, an Ent+strain was an opportunistic pathogen that was completely inhibited by Lcn2 but caused severe, disseminated disease inLcn2−/−mice. In contrast, an Ent+Ybt+strain was a frank respiratory pathogen, causing pneumonia despite Lcn2. However, Lcn2 retained partial protection against disseminated disease. In summary, Ybt is a virulence factor that is prevalent among KPC-producingK. pneumoniaeisolates and promotes respiratory tract infections through evasion of Lcn2.

scholarly journals Klebsiella pneumoniae Reduces SUMOylation To Limit Host Defense Responses

mBio ◽  
2020 ◽  
Vol 11 (5) ◽  
Author(s):  
Joana Sá-Pessoa ◽  
Kornelia Przybyszewska ◽  
Filipe Nuno Vasconcelos ◽  
Amy Dumigan ◽  
Christian G. Frank ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Klebsiella Pneumoniae ◽  
Glycogen Synthase ◽  
Defense Responses ◽  
Multidrug Resistant ◽  
Lung Epithelial Cells ◽  
Type I ◽  
Dependent Manner ◽  
Content Type ◽  
Medical Needs

ABSTRACT Klebsiella pneumoniae is an important cause of multidrug-resistant infections worldwide. Understanding the virulence mechanisms of K. pneumoniae is a priority and timely to design new therapeutics. Here, we demonstrate that K. pneumoniae limits the SUMOylation of host proteins in epithelial cells and macrophages (mouse and human) to subvert cell innate immunity. Mechanistically, in lung epithelial cells, Klebsiella increases the levels of the deSUMOylase SENP2 in the cytosol by affecting its K48 ubiquitylation and its subsequent degradation by the ubiquitin proteasome. This is dependent on Klebsiella preventing the NEDDylation of the Cullin-1 subunit of the ubiquitin ligase complex E3-SCF-βTrCP by exploiting the CSN5 deNEDDylase. Klebsiella induces the expression of CSN5 in an epidermal growth factor receptor (EGFR)-phosphatidylinositol 3-kinase (PI3K)-protein kinase B (AKT)-extracellular signal-regulated kinase (ERK)-glycogen synthase kinase 3 beta (GSK3β) signaling pathway-dependent manner. In macrophages, Toll-like receptor 4 (TLR4)-TRAM-TRIF-induced type I interferon (IFN) via IFN receptor 1 (IFNAR1)-controlled signaling mediates Klebsiella-triggered decrease in the levels of SUMOylation via let-7 microRNAs (miRNAs). Our results revealed the crucial role played by Klebsiella polysaccharides, the capsule, and the lipopolysaccharide (LPS) O-polysaccharide, to decrease the levels of SUMO-conjugated proteins in epithelial cells and macrophages. A Klebsiella-induced decrease in SUMOylation promotes infection by limiting the activation of inflammatory responses and increasing intracellular survival in macrophages. IMPORTANCE Klebsiella pneumoniae has been singled out as an urgent threat to human health due to the increasing isolation of strains resistant to “last-line” antimicrobials, narrowing the treatment options against Klebsiella infections. Unfortunately, at present, we cannot identify candidate compounds in late-stage development for treatment of multidrug-resistant Klebsiella infections; this pathogen is exemplary of the mismatch between unmet medical needs and the current antimicrobial research and development pipeline. Furthermore, there is still limited evidence on K. pneumoniae pathogenesis at the molecular and cellular levels in the context of the interactions between bacterial pathogens and their hosts. In this research, we have uncovered a sophisticated strategy employed by Klebsiella to subvert the activation of immune defenses by controlling the modification of proteins. Our research may open opportunities to develop new therapeutics based on counteracting this Klebsiella-controlled immune evasion strategy.

scholarly journals Immunomodulatory Role of Clarithromycin in Acinetobacter baumannii Infection via Formation of Neutrophil Extracellular Traps

2015 ◽  
Vol 60 (2) ◽  
pp. 1040-1048 ◽  
Author(s):  
Theocharis Konstantinidis ◽  
Konstantinos Kambas ◽  
Alexandros Mitsios ◽  
Maria Panopoulou ◽  
Victoria Tsironidou ◽  
...  
Keyword(s):  
Acinetobacter Baumannii ◽  
Ex Vivo ◽  
Neutrophil Extracellular Traps ◽  
Macrolide Antibiotics ◽  
Dependent Manner ◽  
Biofilm Inhibition ◽  
Content Type ◽  
Extracellular Traps

ABSTRACTMacrolide antibiotics have been shown to act as immunomodulatory molecules in various immune cells. However, their effect on neutrophils has not been extensively investigated. In this study, we investigated the role of macrolide antibiotics in the generation of neutrophil extracellular traps (NETs). By assessingex vivoandin vivoNET formation, we demonstrated that clarithromycin is able to induce NET generation bothin vitroandin vivo. Clarithromycin utilizes autophagy in order to form NETs, and these NETs are decorated with antimicrobial peptide LL-37. Clarithromycin-induced NETs are able to inhibitAcinetobacter baumanniigrowth and biofilm formation in an LL-37-dependent manner. Additionally, LL-37 antimicrobial function depends on NET scaffold integrity. Collectively, these data expand the knowledge on the immunomodulatory role of macrolide antibiotics via the generation of LL-37-bearing NETs, which demonstrate LL-37-dependent antimicrobial activity and biofilm inhibition againstA. baumannii.

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