scholarly journals Studying the surfaces of bacteria using neutron scattering: finding new openings for antibiotics

2020 ◽  
Vol 48 (5) ◽  
pp. 2139-2149
Author(s):  
Nicolò Paracini ◽  
Luke A. Clifton ◽  
Jeremy H. Lakey
Keyword(s):  
Antibiotic Resistance ◽  
Neutron Scattering ◽  
Structural Information ◽  
Cell Envelope ◽  
In Vitro Models ◽  
Differential Sensitivity ◽  
Asymmetric Membranes ◽  
Gram Negative ◽  
Biological Barrier

The use of neutrons as a scattering probe to investigate biological membranes has steadily grown in the past three decades, shedding light on the structure and behaviour of this ubiquitous and fundamental biological barrier. Meanwhile, the rise of antibiotic resistance has catalysed a renewed interest in understanding the mechanisms underlying the dynamics of antibiotics interaction with the bacterial cell envelope. It is widely recognised that the key reason behind the remarkable success of Gram-negative pathogens in developing antibiotic resistance lies in the effectiveness of their outer membrane (OM) in defending the cell from antibacterial compounds. Critical to its function, the highly asymmetric lipid distribution between the inner and outer bilayer leaflets of the OM, adds an extra level of complexity to the study of this crucial defence barrier. Here we review the opportunities offered by neutron scattering techniques, in particular reflectometry, to provide structural information on the interactions of antimicrobials with in vitro models of the OM. The differential sensitivity of neutrons towards hydrogen and deuterium makes them a unique probe to study the structure and behaviour of asymmetric membranes. Molecular-level understanding of the interactions between antimicrobials and the Gram-negative OM provides valuable insights that can aid drug development and broaden our knowledge of this critically important biological barrier.

Core oligosaccharide portion of lipopolysaccharide plays important roles on multiple antibiotic resistance in Escherichia coli

2021 ◽  
Author(s):  
Jianli Wang ◽  
Wenjian Ma ◽  
Yu Fang ◽  
Hao Liang ◽  
Huiting Yang ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Antibiotic Resistance ◽  
Cell Envelope ◽  
Gene Clusters ◽  
Gram Negative Bacteria ◽  
Multiple Antibiotic Resistance ◽  
Core Oligosaccharide ◽  
Gram Negative ◽  
The Core ◽  
K 12

Gram-negative bacteria are intrinsically resistant to antibiotics due to the presence of the cell envelope, but mechanisms are still not fully understood. In this study, a series of mutants that lack one or more major components associated with the cell envelope were constructed from Escherichia coli K-12 W3110. WJW02 can only synthesize Kdo 2 -lipid A which lacks the core oligosaccharide portion of lipopolysaccharide. WJW04, WJW07 and WJW08 were constructed from WJW02 by deleting the gene clusters relevant to the biosynthesis of exopolysaccharide, flagella and fimbria, respectively. WJW09, WJW010 and WJW011 cells cannot synthesize exopolysaccharide, flagella and fimbria, respectively. Comparing to the wild type W3110, mutants WJW02, WJW04, WJW07 and WJW08 cells showed decreased resistance to more than 10 different antibacterial drugs, but not the mutants WJW09, WJW010 and WJW011. This indicates that the core oligosaccharide portion of lipopolysaccharide plays important roles on multiple antibiotic resistance in E. coli and the 1 st heptose in core oligosaccharide portion is critical. Furthermore, the removal of the core oligosaccharide of LPS leads to influences on cell wall morphology, cell phenotypes, porins, efflux systems, and the respond behaviors to antibiotic stimulation. The results demonstrated the important role of lipopolysaccharide on the antibiotic resistance of Gram-negative bacteria.

scholarly journals Facile synthesis of silver nanoparticles using Averrhoa bilimbi L and Plum extracts and investigation on the synergistic bioactivity using in vitro models

2019 ◽  
Vol 8 (1) ◽  
pp. 873-884 ◽  
Author(s):  
Suresh Sagadevan ◽  
Selvaraj Vennila ◽  
Preeti Singh ◽  
Jayasingh Anita Lett ◽  
Mohd Rafie Johan ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Silver Nanoparticles ◽  
Salmonella Typhi ◽  
In Vitro Models ◽  
Diffusion Method ◽  
Gram Positive ◽  
Gram Negative ◽  
Vis Spectroscopy ◽  
Test Cultures

Abstract The bacterial communities (Gram-negative and Gram-positive) form the biofilms which oppose the mode of action of antibiotics and affecting the immune system of the human. These chronic infections related to biofilm are always hard to be cured because of their inherent resistance to both antimicrobial agents and host defense. The present study is devoted to the synthesis of silver nanoparticles (AgNPs) using aqueous extracts of Averrhoa bilimbi leaf and Plum fruit (Prunus bokharensis) and its inhibitory effect on Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, and Salmonella typhi. Here, AgNPs) were successfully prepared by green synthesis method obtained by the reduction of silver nitrate into silver ions. The synthesized AgNPs were characterized by UV-Vis Spectroscopy, XRD, FTIR, FESEM-EDAX and DLS analysis. The antimicrobial effects of the two extract were evaluated by the disk diffusion method. The green synthesized AgNPs exhibited good antibacterial activity against both Gram-negative and Gram-positive. They were also analyzed for their significant antibacterial activities against all the test cultures exposed whereas Escherichia coli and Salmonella typhi topped among other test cultures. The anti-proliferative activity of phytochemical mediated synthesis of Ag NPs was investigated for their cytotoxicity in Vero and Human epidermoid larynx carcinoma cell lines (HEp-2) based on their viability using MTT assay. The present study also represents the synergistic bioactivity of silver nanoparticles using in vitro models.

scholarly journals MICROBIOLOGICAL SPECTRE OF TERTIARY PERITONITIS AS A COMPONENT OF ITS DIAGNOSTICS AND TREATMENT

2017 ◽  
Vol 6 ◽  
pp. 10-18
Author(s):  
Petro Fomin ◽  
Oleh Matviychuk ◽  
Olena Korniychuk
Keyword(s):  
Antibiotic Resistance ◽  
Abdominal Sepsis ◽  
Peritoneal Exudate ◽  
Secondary Peritonitis ◽  
Blood Sampling ◽  
Candida Spp ◽  
Gram Negative ◽  
Tertiary Peritonitis

The aim of the research was to investigate the microbial spectre of tertiary peritonits (TP) and its antibiotic resistance as the way to improve the diagnostics and treatment of TP. Materials and methods. Prospective research enrolled 109 patients with secondary peritonitis. Tertiary peritonitis developed in 18,3 % of cases. Samples of peritoneal exudate had been drawn upon index operation, relaparotomy and on the day of diagnosis of TP. Blood sampling was performed in patients with persistent fever, impaired consciousness, prolonged (>4 days) discharge from drainage tubes and on the 1st day of diagnosis of TP. Antibacterial susceptibility was evaluated using Hinton-Müller media. Results and discussion. Patients were divided into 2 groups: with secondary peritonitis (89) and with TP (20). In TP group, cultivation of 76,2 % of primary specimens resulted in replantable and identifiable growth, presenting a shift towards Gram-negative flora and higher incidence of Candida albicans. Following the development of TP, hemocultures were positive in 15,1 %, presented mainly by Proteus spp. and non-albicans Candida spp. Second-group carbapenems, tigecycline and piperacillin-tazobactam had shown the highest activity in pathogens of TP. Caspofungin proved to be the most potent antifungal agent, especially towards non-albicans Candida spp. Antibiotic resistance in TP group was marked in 63,8 %. Conclusions. Tertiary peritonitis is one of the most severe forms of abdominal sepsis with highest mortality. Causing pathogenic flora in case of TP is mainly Gram-negative and coccal with high rates of antibiotic resistance both in vitro and in vivo. Fungi, presented predominantly by Candida non-albicans substrains, show an increasing content in peritoneal exudate and major effect upon mortality in TP. In case of TP, a significant percent of peritoneal specimens do not provide any culture growth despite of observing stringent sampling, transportation and cultivation rules. Antimicrobial therapy of TP can never be standardized and should always be thoroughly based upon regular and proper peritoneal and blood sampling.

scholarly journals Defining the function of OmpA in the Rcs stress response

2020 ◽  
Author(s):  
Juliette Létoquart ◽  
Kilian Dekoninck ◽  
Cédric Laguri ◽  
Pascal Demange ◽  
Robin Bevernaegie ◽  
...  
Keyword(s):  
Stress Response ◽  
Outer Membrane ◽  
Cell Envelope ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Bacterial Surface ◽  
Periplasmic Domain ◽  
Inner Membrane Protein

AbstractOmpA, a protein commonly found in the outer membrane of Gram-negative bacteria, has served as a paradigm for the study of β-barrel proteins for several decades. In Escherichia coli, OmpA was previously reported to form complexes with RcsF, a surface-exposed lipoprotein that triggers the Rcs stress response when damage occurs in the outer membrane and the peptidoglycan. How OmpA interacts with RcsF and whether this interaction allows RcsF to reach the surface has remained unclear. Here, we integrated in vivo and in vitro approaches to establish that RcsF interacts with the C-terminal, periplasmic domain of OmpA, not with the N-terminal β-barrel, thus implying that RcsF does not reach the bacterial surface via OmpA. Our results reveal a novel function for OmpA in the cell envelope: OmpA competes with the inner membrane protein IgaA, the downstream Rcs component, for RcsF binding across the periplasm, thereby regulating the Rcs response.

scholarly journals Reciprocal Regulation of PASTA Kinase Signaling by Differential Modification

2019 ◽  
Vol 201 (10) ◽  
Author(s):  
Benjamin D. Labbe ◽  
Cherisse L. Hall ◽  
Stephanie L. Kellogg ◽  
Yao Chen ◽  
Olivia Koehn ◽  
...  
Keyword(s):  
Cell Wall ◽  
Antibiotic Resistance ◽  
Kinase Activity ◽  
Cell Envelope ◽  
Activation Loop ◽  
Intrinsic Resistance ◽  
Content Type

ABSTRACTTransmembrane Ser/Thr kinases containing extracellular PASTA (penicillin-binding protein [PBP]andSer/Thr-associated) domains are ubiquitous amongActinobacteriaandFirmicutesspecies. Such PASTA kinases regulate critical bacterial processes, including antibiotic resistance, cell division, cell envelope homeostasis, and virulence, and are sometimes essential for viability. Previous studies of purified PASTA kinase fragments revealed they are capable of autophosphorylationin vitro, typically at multiple sites on the kinase domain. Autophosphorylation of a specific structural element of the kinase known as the activation loop is thought to enhance kinase activity in response to stimuli. However, the role of kinase phosphorylation at other sites is largely unknown. Moreover, the mechanisms by which PASTA kinases are deactivated once their stimulus has diminished are poorly understood.Enterococcus faecalisis a Gram-positive intestinal bacterium and a major antibiotic-resistant opportunistic pathogen. InE. faecalis, the PASTA kinase IreK drives intrinsic resistance to cell wall-active antimicrobials, and such antimicrobials trigger enhanced phosphorylation of IreKin vivo. Here we identify multiple sites of phosphorylation on IreK and evaluate their functionin vivoandin vitro. While phosphorylation of the IreK activation loop is required for kinase activity, we found that phosphorylation at a site distinct from the activation loop reciprocally modulates IreK activityin vivo, leading to diminished activity (and diminished antimicrobial resistance). Moreover, this site is important for deactivation of IreKin vivoupon removal of an activating stimulus. Our results are consistent with a model in which phosphorylation of IreK at distinct sites reciprocally regulates IreK activityin vivoto promote adaptation to cell wall stresses.IMPORTANCETransmembrane Ser/Thr kinases containing extracellular PASTA domains are ubiquitous amongActinobacteriaandFirmicutesspecies and regulate critical processes, including antibiotic resistance, cell division, and cell envelope homeostasis. Previous studies of PASTA kinase fragments revealed autophosphorylation at multiple sites. However, the functional role of autophosphorylation and the relative impacts of phosphorylation at distinct sites are poorly understood. The PASTA kinase ofEnterococcus faecalis, IreK, regulates intrinsic resistance to antimicrobials. Here we identify multiple sites of phosphorylation on IreK and show that modification of IreK at distinct sites reciprocally regulates IreK activity and antimicrobial resistancein vivo. Thus, these results provide new insights into the mechanisms by which PASTA kinases can regulate critical physiological processes in a wide variety of bacterial species.

scholarly journals Caprylic Acid and Glyceryl Trinitrate Combination for Eradication of Biofilm

2014 ◽  
Vol 59 (3) ◽  
pp. 1786-1788 ◽  
Author(s):  
Joel Rosenblatt ◽  
Ruth A. Reitzel ◽  
Issam Raad
Keyword(s):  
Antibiotic Resistance ◽  
Glyceryl Trinitrate ◽  
Broad Spectrum ◽  
Caprylic Acid ◽  
Gram Positive ◽  
Gram Negative ◽  
Antimicrobial Treatments ◽  
Synergistic Combinations

ABSTRACTThere is a growing need for biocompatible, broad-spectrum, nonantibiotic, antimicrobial treatments because of the frequent ineffectiveness of antibiotics against biofilms and the increasing incidence of antibiotic resistance. In this study, we demonstrate rapid and complete biofilm eradication in anin vitromodel with synergistic combinations of glyceryl trinitrate and caprylic acid against resistant Gram-positive, Gram-negative, and fungal biofilms.

scholarly journals Mutant Alleles oflptDIncrease the Permeability of Pseudomonas aeruginosa and Define Determinants of Intrinsic Resistance to Antibiotics

2015 ◽  
Vol 60 (2) ◽  
pp. 845-854 ◽  
Author(s):  
Carl J. Balibar ◽  
Marcin Grabowicz
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Cell Envelope ◽  
Antibacterial Drug ◽  
Multidrug Efflux ◽  
Intrinsic Resistance ◽  
Gram Negative ◽  
Content Type ◽  
Design And Optimization ◽  
Multidrug Efflux Pumps

ABSTRACTGram-negative bacteria provide a particular challenge to antibacterial drug discovery due to their cell envelope structure. Compound entry is impeded by the lipopolysaccharide (LPS) of the outer membrane (OM), and those molecules that overcome this barrier are often expelled by multidrug efflux pumps. Understanding how efflux and permeability affect the ability of a compound to reach its target is paramount to translatingin vitrobiochemical potency to cellular bioactivity. Herein, a suite ofPseudomonas aeruginosastrains were constructed in either a wild-type or efflux-null background in which mutations were engineered in LptD, the final protein involved in LPS transport to the OM. These mutants were demonstrated to be defective in LPS transport, resulting in compromised barrier function. Using isogenic strain sets harboring these newly created alleles, we were able to define the contributions of permeability and efflux to the intrinsic resistance ofP. aeruginosato a variety of antibiotics. These strains will be useful in the design and optimization of future antibiotics against Gram-negative pathogens.

scholarly journals Adjunctive transferrin to reduce the emergence of antibiotic resistance in Gram-negative bacteria

2019 ◽  
Vol 74 (9) ◽  
pp. 2631-2639 ◽  
Author(s):  
Brian M Luna ◽  
Ksenia Ershova ◽  
Jun Yan ◽  
Amber Ulhaq ◽  
Travis B Nielsen ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Bacterial Pathogens ◽  
Microbial Pathogens ◽  
Gram Negative Bacteria ◽  
Antibiotic Resistant ◽  
Gram Negative ◽  
Resistant Mutants ◽  
Time Kill ◽  
Emergence Of Resistance

AbstractBackgroundNew strategies are needed to slow the emergence of antibiotic resistance among bacterial pathogens. In particular, society is experiencing a crisis of antibiotic-resistant infections caused by Gram-negative bacterial pathogens and novel therapeutics are desperately needed to combat such diseases. Acquisition of iron from the host is a nearly universal requirement for microbial pathogens—including Gram-negative bacteria—to cause infection. We have previously reported that apo-transferrin (lacking iron) can inhibit the growth of Staphylococcus aureus in culture and diminish emergence of resistance to rifampicin.ObjectivesTo define the potential of apo-transferrin to inhibit in vitro growth of Klebsiella pneumoniae and Acinetobacter baumannii, key Gram-negative pathogens, and to reduce emergence of resistance to antibiotics.MethodsThe efficacy of apo-transferrin alone or in combination with meropenem or ciprofloxacin against K. pneumoniae and A. baumannii clinical isolates was tested by MIC assay, time–kill assay and assays for the selection of resistant mutants.ResultsWe confirmed that apo-transferrin had detectable MICs for all strains tested of both pathogens. Apo-transferrin mediated an additive antimicrobial effect for both antibiotics against multiple strains in time–kill assays. Finally, adding apo-transferrin to ciprofloxacin or meropenem reduced the emergence of resistant mutants during 20 day serial passaging of both species.ConclusionsThese results suggest that apo-transferrin may have promise to suppress the emergence of antibiotic-resistant mutants when treating infections caused by Gram-negative bacteria.

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