Publications by authors named "Kenneth W Bayles"

93 Publications

CyDisCo production of functional recombinant SARS-CoV-2 spike receptor binding domain.

Protein Sci 2021 09 16;30(9):1983-1990. Epub 2021 Jul 16.

Eppley Institute for Research in Cancer and Allied Diseases, University of Nebraska Medical Center, Omaha, Nebraska, USA.

The COVID-19 pandemic caused by SARS-CoV-2 has applied significant pressure on overtaxed healthcare around the world, underscoring the urgent need for rapid diagnosis and treatment. We have developed a bacterial strategy for the expression and purification of a SARS-CoV-2 spike protein receptor binding domain (RBD) that includes the SD1 domain. Bacterial cytoplasm is a reductive environment, which is problematic when the recombinant protein of interest requires complicated folding and/or processing. The use of the CyDisCo system (cytoplasmic disulfide bond formation in E. coli) bypasses this issue by pre-expressing a sulfhydryl oxidase and a disulfide isomerase, allowing the recombinant protein to be correctly folded with disulfide bonds for protein integrity and functionality. We show that it is possible to quickly and inexpensively produce an active RBD in bacteria that is capable of recognizing and binding to the ACE2 (angiotensin-converting enzyme) receptor as well as antibodies in COVID-19 patient sera.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/pro.4152DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8376421PMC
September 2021

Accumulation of Succinyl Coenzyme A Perturbs the Methicillin-Resistant (MRSA) Succinylome and Is Associated with Increased Susceptibility to Beta-Lactam Antibiotics.

mBio 2021 06 29;12(3):e0053021. Epub 2021 Jun 29.

Microbiology, School of Natural Sciences, National University of Ireland, Galway, Ireland.

Penicillin binding protein 2a (PBP2a)-dependent resistance to β-lactam antibiotics in methicillin-resistant Staphylococcus aureus (MRSA) is regulated by the activity of the tricarboxylic acid (TCA) cycle via a poorly understood mechanism. We report that mutations in and , but not other TCA cycle enzymes, negatively impact β-lactam resistance without changing PBP2a expression. Increased intracellular levels of succinyl coenzyme A (succinyl-CoA) in the mutant significantly perturbed lysine succinylation in the MRSA proteome. Suppressor mutations in or , responsible for succinyl-CoA biosynthesis, reversed mutant phenotypes. The major autolysin (Atl) was the most succinylated protein in the proteome, and increased Atl succinylation in the mutant was associated with loss of autolytic activity. Although PBP2a and PBP2 were also among the most succinylated proteins in the MRSA proteome, peptidoglycan architecture and cross-linking were unchanged in the mutant. These data reveal that perturbation of the MRSA succinylome impacts two interconnected cell wall phenotypes, leading to repression of autolytic activity and increased susceptibility to β-lactam antibiotics. -dependent methicillin resistance in MRSA is subject to regulation by numerous accessory factors involved in cell wall biosynthesis, nucleotide signaling, and central metabolism. Here, we report that mutations in the TCA cycle gene, , increased susceptibility to β-lactam antibiotics and was accompanied by significant accumulation of succinyl-CoA, which in turn perturbed lysine succinylation in the proteome. Although cell wall structure and cross-linking were unchanged, significantly increased succinylation of the major autolysin Atl, which was the most succinylated protein in the proteome, was accompanied by near complete repression of autolytic activity. These findings link central metabolism and levels of succinyl-CoA to the regulation of β-lactam antibiotic resistance in MRSA through succinylome-mediated control of two interlinked cell wall phenotypes. Drug-mediated interference of the SucCD-controlled succinylome may help overcome β-lactam resistance.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/mBio.00530-21DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8437408PMC
June 2021

Integrative network analyses of transcriptomics data reveal potential drug targets for acute radiation syndrome.

Sci Rep 2021 Mar 10;11(1):5585. Epub 2021 Mar 10.

Department of Biochemistry, University of Nebraska-Lincoln, Lincoln, NE, USA.

Recent political unrest has highlighted the importance of understanding the short- and long-term effects of gamma-radiation exposure on human health and survivability. In this regard, effective treatment for acute radiation syndrome (ARS) is a necessity in cases of nuclear disasters. Here, we propose 20 therapeutic targets for ARS identified using a systematic approach that integrates gene coexpression networks obtained under radiation treatment in humans and mice, drug databases, disease-gene association, radiation-induced differential gene expression, and literature mining. By selecting gene targets with existing drugs, we identified potential candidates for drug repurposing. Eight of these genes (BRD4, NFKBIA, CDKN1A, TFPI, MMP9, CBR1, ZAP70, IDH3B) were confirmed through literature to have shown radioprotective effect upon perturbation. This study provided a new perspective for the treatment of ARS using systems-level gene associations integrated with multiple biological information. The identified genes might provide high confidence drug target candidates for potential drug repurposing for ARS.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-021-85044-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7946886PMC
March 2021

Inactivation of the Pta-AckA pathway impairs fitness of during overflow metabolism.

J Bacteriol 2021 Feb 16. Epub 2021 Feb 16.

Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, NE 68198, USA

Under conditions of glucose excess, aerobically growing bacteria predominantly direct carbon flux towards acetate fermentation, a phenomenon known as overflow metabolism or the bacterial 'Crabtree effect'. Numerous studies of the major acetate-generating pathway, the Pta-AckA, revealed its important role in bacterial fitness through the control of central metabolism to sustain balanced growth and cellular homeostasis. In this work, we highlight the contribution of the Pta-AckA pathway to fitness of the spore-forming bacterium, We demonstrate that disruption of the Pta-AckA pathway causes a drastic growth reduction in the mutants and alters the metabolic and energy status of the cells. Our results revealed that inactivation of the Pta-AckA pathway increases the glucose consumption rate, affects intracellular ATP, NAD and NADH levels and leads to a metabolic block at the pyruvate and acetyl-CoA nodes. Consequently, accumulation of intracellular acetyl-CoA and pyruvate forces bacteria to direct carbon into the TCA and/or glyoxylate cycles as well as fatty acid and poly(3-hydroxybutyrate) (PHB) biosynthesis pathways. Notably, the presence of phosphate butyryltransferase in partially compensates for the loss of phosphotransacetylase activity. Furthermore, overexpression of the gene not only eliminates the negative impact of the mutation on fitness, but also restores normal growth in the mutant of the non-butyrate-producing bacterium, Taken together, the results of this study demonstrate the importance of the Pta-AckA pathway for fitness by revealing its critical contribution to the maintenance of metabolic homeostasis during aerobic growth under conditions of carbon overflow. , the etiologic agent of anthrax, is a highly pathogenic, spore-forming bacterium that causes acute, life-threatening disease in both humans and livestock. A greater understanding of the metabolic determinants governing fitness of is essential for the development of successful therapeutic and vaccination strategies aimed at lessening the potential impact of this important biodefense pathogen. This study is the first to demonstrate the vital role of the Pta-AckA pathway in preserving energy and metabolic homeostasis in under conditions of carbon overflow, therefore, highlighting this pathway as a potential therapeutic target for drug discovery. Overall, the results of this study provide important insight into understanding the metabolic processes and requirements driving rapid proliferation during vegetative growth.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/JB.00660-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8092162PMC
February 2021

Bromelain Inhibits SARS-CoV-2 Infection in VeroE6 Cells.

bioRxiv 2020 Sep 16. Epub 2020 Sep 16.

Eppley Institute for Research in Cancer and Allied Diseases, University of Nebraska Medical Center, Omaha, Nebraska, USA 68198.

Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). The initial interaction between Transmembrane Serine Protease 2 (TMPRSS2) primed SARS-CoV-2 spike (S) protein and host cell receptor angiotensin-converting enzyme 2 (ACE-2) is a pre-requisite step for this novel coronavirus pathogenesis. Here, we expressed a GFP-tagged SARS-CoV-2 S-Ectodomain in Tni insect cells. That contained sialic acid-enriched N- and O-glycans. Surface resonance plasmon (SPR) and Luminex assay showed that the purified S-Ectodomain binding to human ACE-2 and immunoreactivity with COVID-19 positive samples. We demonstrate that bromelain (isolated from pineapple stem and used as a dietary supplement) treatment diminishes the expression of ACE-2 and TMPRSS2 in VeroE6 cells and dramatically lowers the expression of S-Ectodomain. Importantly, bromelain treatment reduced the interaction between S-Ectodomain and VeroE6 cells. Most importantly, bromelain treatment significantly diminished the SARS-CoV-2 infection in VeroE6 cells. Altogether, our results suggest that bromelain or bromelain rich pineapple stem may be used as an antiviral against COVID-19.

Highlights: Bromelain inhibits / cleaves the expression of ACE-2 and TMPRSS2Bromelain cleaves / degrades SARS-CoV-2 spike proteinBromelain inhibits S-Ectodomain binding and SARS-CoV-2 infection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1101/2020.09.16.297366DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7523097PMC
September 2020

Genetic and Biochemical Analysis of CodY-Mediated Cell Aggregation in Staphylococcus aureus Reveals an Interaction between Extracellular DNA and Polysaccharide in the Extracellular Matrix.

J Bacteriol 2020 03 26;202(8). Epub 2020 Mar 26.

Department of Biology, Georgetown University, Washington, DC, USA

The global regulator CodY links nutrient availability to the regulation of virulence factor gene expression in , including many genes whose products affect biofilm formation. Antithetical phenotypes of both biofilm deficiency and accumulation have been reported for -null mutants; thus, the role of CodY in biofilm development remains unclear. mutant cells of a strain producing a robust biofilm elaborate proaggregation surface-associated features not present on mutant cells that do not produce a robust biofilm. Biochemical analysis of the clinical isolate SA564, which aggregates when deficient for CodY, revealed that these features are sensitive to nuclease treatment and are resistant to protease exposure. Genetic analyses revealed that disrupting (the diacylglycerol transferase gene) in mutant cells severely weakened aggregation, indicating a role for lipoproteins in the attachment of the biofilm matrix to the cell surface. An additional and critical role of IcaB in producing functional poly--acetylglucosamine (PIA) polysaccharide in extracellular DNA (eDNA)-dependent biofilm formation was shown. Moreover, overproducing PIA is sufficient to promote aggregation in a DNA-dependent manner regardless of source of nucleic acids. Taken together, our results point to PIA synthesis as the primary determinant of biofilm formation when CodY activity is reduced and suggest a modified electrostatic net model for matrix attachment whereby PIA associates with eDNA, which interacts with the cell surface via covalently attached membrane lipoproteins. This work counters the prevailing view that polysaccharide- and eDNA/protein-based biofilms are mutually exclusive. Rather, we demonstrate that eDNA and PIA can work synergistically to form a biofilm. remains a global health concern and exemplifies the ability of an opportunistic pathogen to adapt and persist within multiple environments, including host tissue. Not only does biofilm contribute to persistence and immune evasion in the host environment, it also may aid in the transition to invasive disease. Thus, understanding how biofilms form is critical for developing strategies for dispersing biofilms and improving biofilm disease-related outcomes. Using biochemical, genetic, and cell biology approaches, we reveal a synergistic interaction between PIA and eDNA that promotes cell aggregation and biofilm formation in a CodY-dependent manner in We also reveal that envelope-associated lipoproteins mediate attachment of the biofilm matrix to the cell surface.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/JB.00593-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7099133PMC
March 2020

Fluorescent Sensor Arrays Can Predict and Quantify the Composition of Multicomponent Bacterial Samples.

Front Chem 2019 15;7:916. Epub 2020 Jan 15.

Department of Pharmaceutical Sciences, University of Nebraska Medical Center, Omaha, NE, United States.

Fast and reliable identification of infectious disease agents is among the most important challenges for the healthcare system. The discrimination of individual components of mixed infections represents a particularly difficult task. In the current study we further expand the functionality of a ratiometric sensor array technology based on small-molecule environmentally-sensitive organic dyes, which can be successfully applied for the analysis of mixed bacterial samples. Using pattern recognition methods and data from pure bacterial species, we demonstrate that this approach can be used to quantify the composition of mixtures, as well as to predict their components with the accuracy of ~80% without the need to acquire additional reference data. The described approach significantly expands the functionality of sensor arrays and provides important insights into data processing for the analysis of other complex samples.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fchem.2019.00916DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6974461PMC
January 2020

An integrated computational and experimental study to investigate Staphylococcus aureus metabolism.

NPJ Syst Biol Appl 2020 01 30;6(1). Epub 2020 Jan 30.

Department of Chemical and Biomolecular Engineering, University of Nebraska-Lincoln, Lincoln, NE, USA.

Staphylococcus aureus is a metabolically versatile pathogen that colonizes nearly all organs of the human body. A detailed and comprehensive knowledge of staphylococcal metabolism is essential to understand its pathogenesis. To this end, we have reconstructed and experimentally validated an updated and enhanced genome-scale metabolic model of S. aureus USA300_FPR3757. The model combined genome annotation data, reaction stoichiometry, and regulation information from biochemical databases and previous strain-specific models. Reactions in the model were checked and fixed to ensure chemical balance and thermodynamic consistency. To further refine the model, growth assessment of 1920 nonessential mutants from the Nebraska Transposon Mutant Library was performed, and metabolite excretion profiles of important mutants in carbon and nitrogen metabolism were determined. The growth and no-growth inconsistencies between the model predictions and in vivo essentiality data were resolved using extensive manual curation based on optimization-based reconciliation algorithms. Upon intensive curation and refinements, the model contains 863 metabolic genes, 1379 metabolites (including 1159 unique metabolites), and 1545 reactions including transport and exchange reactions. To improve the accuracy and predictability of the model to environmental changes, condition-specific regulation information curated from the existing knowledgebase was incorporated. These critical additions improved the model performance significantly in capturing gene essentiality, substrate utilization, and metabolite production capabilities and increased the ability to generate model-based discoveries of therapeutic significance. Use of this highly curated model will enhance the functional utility of omics data, and therefore, serve as a resource to support future investigations of S. aureus and to augment staphylococcal research worldwide.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41540-019-0122-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6992624PMC
January 2020

Stochastic Expression of Sae-Dependent Virulence Genes during Staphylococcus aureus Biofilm Development Is Dependent on SaeS.

mBio 2020 01 14;11(1). Epub 2020 Jan 14.

Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, Nebraska, USA

The intricate process of biofilm formation in the human pathogen involves distinct stages during which a complex mixture of matrix molecules is produced and modified throughout the developmental cycle. Early in biofilm development, a subpopulation of cells detaches from its substrate in an event termed "exodus" that is mediated by SaePQRS-dependent stochastic expression of a secreted staphylococcal nuclease, which degrades extracellular DNA within the matrix, causing the release of cells and subsequently allowing for the formation of metabolically heterogenous microcolonies. Since the SaePQRS regulatory system is involved in the transcriptional control of multiple virulence factors, the expression of several additional virulence genes was examined within a developing biofilm by introducing fluorescent gene reporter plasmids into wild-type and isogenic regulatory mutants and growing these strains in a microfluidic system that supplies the bacteria with a constant flow of media while simultaneously imaging developing biofilms in 5-min intervals. This study demonstrated that multiple virulence genes, including , were expressed stochastically within a specialized subpopulation of cells in nascent biofilms. We demonstrated that virulence genes regulated by SaePQRS were stochastically expressed in nearly all strains examined whereas Agr-regulated genes were expressed more homogenously within maturing microcolonies. The commonly used Newman strain contains a variant of SaeS (SaeS) that confers constitutive kinase activity to the protein and caused this strain to lack the stochastic expression pattern observed in other strain backgrounds. Importantly, repair of the SaeS allele resulting in reversion to the well-conserved SaeS allele found in other strains restored stochastic expression in this strain. is an important human pathogen capable of colonizing diverse tissue types and inducing severe disease in both immunocompromised and otherwise healthy individuals. Biofilm infections caused by this bacterial species are of particular concern because of their persistence, even in the face of intensive therapeutic intervention. The results of the current study demonstrate the stochastic nature of Sae-mediated virulence gene expression in and indicate that this regulatory system may function as a "bistable switch" in a manner similar to that seen with regulators controlling competence gene expression in and persister cell formation in The results of this study provide a new perspective on the complex mechanisms utilized by during the establishment of infections.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/mBio.03081-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6960292PMC
January 2020

CidR and CcpA Synergistically Regulate Staphylococcus aureus Expression.

J Bacteriol 2019 12 5;201(23). Epub 2019 Nov 5.

Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, Nebraska, USA

The death and lysis of a subpopulation of cells during biofilm development benefit the whole bacterial population through the release of an important component of the biofilm matrix, extracellular DNA. Previously, we have demonstrated that these processes are affected by the gene products of the operon, the expression of which is controlled by the LysR-type transcriptional regulator, CidR. In this study, we characterized - and -acting elements essential for the induction of the operon. In addition to a CidR-binding site located within the promoter region, sequence analysis revealed the presence of a putative catabolite responsive element ( box), suggestive of the involvement of the catabolite control protein A (CcpA) in the regulation of expression. This was confirmed using electrophoretic mobility shift assays and real-time reverse transcriptase PCR analysis demonstrating the direct positive control of transcription by the master regulator of carbon metabolism. Furthermore, the importance of CcpA and the identified site for the induction of the operon was demonstrated by examining the expression of P reporter fusions in various mutant strains in which the genes involved in carbon metabolism and carbon catabolite repression were disrupted. Together the results of this study demonstrate the necessity of both transcriptional regulators, CidR and CcpA, for the induction of the operon and reveal the complexity of molecular interactions controlling its expression. This work focuses on the characterization of - and -acting elements essential for the induction of the operon in The results of this study are the first to demonstrate the synergistic control of expression by transcriptional regulators CidR and CcpA during carbohydrate metabolism. We established that the full induction of expression depends on the metabolic state of bacteria and requires both CidR and CcpA. Together, these findings delineate regulatory control of expression under different metabolic conditions and provide important new insights into our understanding of cell death mechanisms during biofilm development in .
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/JB.00371-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6832070PMC
December 2019

Identification of Extracellular DNA-Binding Proteins in the Biofilm Matrix.

mBio 2019 06 25;10(3). Epub 2019 Jun 25.

Department of Immunology and Microbiology, University of Colorado School of Medicine, Aurora, Colorado, USA

We developed a new approach that couples Southwestern blotting and mass spectrometry to discover proteins that bind extracellular DNA (eDNA) in bacterial biofilms. Using as a model pathogen, we identified proteins with known DNA-binding activity and uncovered a series of lipoproteins with previously unrecognized DNA-binding activity. We demonstrated that expression of these lipoproteins results in an eDNA-dependent biofilm enhancement. Additionally, we found that while deletion of lipoproteins had a minimal impact on biofilm accumulation, these lipoprotein mutations increased biofilm porosity, suggesting that lipoproteins and their associated interactions contribute to biofilm structure. For one of the lipoproteins, SaeP, we showed that the biofilm phenotype requires the lipoprotein to be anchored to the outside of the cellular membrane, and we further showed that increased SaeP expression correlates with more retention of high-molecular-weight DNA on the bacterial cell surface. SaeP is a known auxiliary protein of the SaeRS system, and we also demonstrated that the levels of SaeP correlate with nuclease production, which can further impact biofilm development. It has been reported that biofilms are stabilized by positively charged cytoplasmic proteins that are released into the extracellular environment, where they make favorable electrostatic interactions with the negatively charged cell surface and eDNA. In this work we extend this electrostatic net model to include secreted eDNA-binding proteins and membrane-attached lipoproteins that can function as anchor points between eDNA in the biofilm matrix and the bacterial cell surface. Many bacteria are capable of forming biofilms encased in a matrix of self-produced extracellular polymeric substances (EPS) that protects them from chemotherapies and the host defenses. As a result of these inherent resistance mechanisms, bacterial biofilms are extremely difficult to eradicate and are associated with chronic wounds, orthopedic and surgical wound infections, and invasive infections, such as infective endocarditis and osteomyelitis. It is therefore important to understand the nature of the interactions between the bacterial cell surface and EPS that stabilize biofilms. Extracellular DNA (eDNA) has been recognized as an EPS constituent for many bacterial species and has been shown to be important in promoting biofilm formation. Using biofilms, we show that membrane-attached lipoproteins can interact with the eDNA in the biofilm matrix and promote biofilm formation, which suggests that lipoproteins are potential targets for novel therapies aimed at disrupting bacterial biofilms.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/mBio.01137-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6593408PMC
June 2019

Construction of a Sequence-Defined Transposon Mutant Library in Staphylococcus aureus.

Methods Mol Biol 2019 ;2016:29-37

Department of Pathology and Microbiology, Center for Staphylococcal Research, University of Nebraska Medical Center, Omaha, NE, USA.

Transposon mutagenesis is one of the most widely used strategies to generate a large number of random mutations within a bacterial genome and then to precisely identify the mutated sites. The generation of sequence-defined transposon mutant libraries that are composed of a collection of different mutants, each containing a single transposon insertion mutation within nearly all of the nonessential genes within the genome, is a rapid and reliable way to enhance the study of gene function. In this chapter, we describe the process to generate a sequence-defined transposon mutant library in Staphylococcus aureus utilizing the mariner-based bursa aurealis transposon.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/978-1-4939-9570-7_3DOI Listing
March 2020

Guanine Limitation Results in CodY-Dependent and -Independent Alteration of Staphylococcus aureus Physiology and Gene Expression.

J Bacteriol 2018 07 25;200(14). Epub 2018 Jun 25.

Department of Biology, Georgetown University, Washington, DC, USA

In , the global transcriptional regulator CodY modulates the expression of hundreds of genes in response to the availability of GTP and the branched-chain amino acids isoleucine, leucine, and valine (ILV). CodY DNA-binding activity is high when GTP and ILV are abundant. When GTP and ILV are limited, CodY's affinity for DNA drops, altering expression of CodY-regulated targets. In this work, we investigated the impact of guanine nucleotides (GNs) on physiology and CodY activity by constructing a null mutant (Δ strain). biosynthesis of guanine monophosphate is abolished due to the mutation; thus, the mutant cells require exogenous guanosine for growth. We also found that CodY activity was reduced when we knocked out , activating the Agr two-component system and increasing secreted protease activity. Notably, in a rich, complex medium, we detected an increase in alternative sigma factor B activity in the Δ mutant, which results in a 5-fold increase in production of the antioxidant pigment staphyloxanthin. Under biologically relevant flow conditions, Δ cells failed to form robust biofilms when limited for guanine or guanosine. Transcriptome sequencing (RNA-Seq) analysis of the transcriptome during growth in guanosine-limited chemostats revealed substantial CodY-dependent and -independent alterations of gene expression profiles. Importantly, these changes increase production of proteases and δ-toxin, suggesting that exhibits a more invasive lifestyle when limited for guanosine. Further, gene products upregulated under GN limitation, including those necessary for lipoic acid biosynthesis and sugar transport, may prove to be useful drug targets for treating Gram-positive infections. infections impose a serious economic burden on health care facilities and patients because of the emergence of strains resistant to last-line antibiotics. Understanding the physiological processes governing fitness and virulence of in response to environmental cues is critical for developing efficient diagnostics and treatments. purine biosynthesis is essential for both fitness and virulence in since inhibiting production cripples 's ability to cause infection. Here, we corroborate these findings and show that blocking guanine nucleotide synthesis severely affects fitness by altering metabolic and virulence gene expression. Characterizing pathways and gene products upregulated in response to guanine limitation can aid in the development of novel adjuvant strategies to combat infections.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/JB.00136-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6018361PMC
July 2018

Ratiometric Fluorescent Sensor Array as a Versatile Tool for Bacterial Pathogen Identification and Analysis.

ACS Sens 2018 03 9;3(3):700-708. Epub 2018 Mar 9.

Department of Pharmaceutical Sciences , University of Nebraska Medical Center , Omaha , Nebraska 68198-6858 , United States.

Rapid and reliable identification of pathogenic microorganisms is of great importance for human and animal health. Most conventional approaches are time-consuming and require expensive reagents, sophisticated equipment, trained personnel, and special storage and handling conditions. Sensor arrays based on small molecules offer a chemically stable and cost-effective alternative. Here we present a ratiometric fluorescent sensor array based on the derivatives of 2-(4'- N, N-dimethylamino)-3-hydroxyflavone and investigate its ability to provide a dual-channel ratiometric response. We demonstrate that, by using discriminant analysis of the sensor array responses, it is possible to effectively distinguish between eight bacterial species and recognize their Gram status. Thus, multiple parameters can be derived from the same data set. Moreover, the predictive potential of this sensor array is discussed, and its ability to analyze unknown samples beyond the list of species used for the training matrix is demonstrated. The proposed sensor array and analysis strategies open new avenues for the development of advanced ratiometric sensors for multiparametric analysis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acssensors.8b00025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5938749PMC
March 2018

Nutritional Regulation of the Sae Two-Component System by CodY in Staphylococcus aureus.

J Bacteriol 2018 04 26;200(8). Epub 2018 Mar 26.

Department of Biology, Georgetown University, Washington, DC, USA

subverts innate defenses during infection in part by killing host immune cells to exacerbate disease. This human pathogen intercepts host cues and activates a transcriptional response via the exoprotein expression (SaeR/SaeS [SaeR/S]) two-component system to secrete virulence factors critical for pathogenesis. We recently showed that the transcriptional repressor CodY adjusts nuclease () gene expression via SaeR/S, but the mechanism remained unknown. Here, we identified two CodY binding motifs upstream of the P1 promoter, which suggested direct regulation by this global regulator. We show that CodY shares a binding site with the positive activator SaeR and that alleviating direct CodY repression at this site is sufficient to abrogate stochastic expression, suggesting that CodY represses expression by blocking SaeR binding. Epistasis experiments support a model that CodY also controls indirectly through Agr and Rot-mediated repression of the P1 promoter. We also demonstrate that CodY repression of restrains production of secreted cytotoxins that kill human neutrophils. We conclude that CodY plays a previously unrecognized role in controlling virulence gene expression via SaeR/S and suggest a mechanism by which CodY acts as a master regulator of pathogenesis by tying nutrient availability to virulence gene expression. Bacterial mechanisms that mediate the switch from a commensal to pathogenic lifestyle are among the biggest unanswered questions in infectious disease research. Since the expression of most virulence genes is often correlated with nutrient depletion, this implies that virulence is a response to the lack of nourishment in host tissues and that pathogens like produce virulence factors in order to gain access to nutrients in the host. Here, we show that specific nutrient depletion signals appear to be funneled to the SaeR/S system through the global regulator CodY. Our findings reveal a strategy by which delays the production of immune evasion and immune-cell-killing proteins until key nutrients are depleted.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/JB.00012-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5869476PMC
April 2018

Staphylococcus aureus CidC Is a Pyruvate:Menaquinone Oxidoreductase.

Biochemistry 2017 09 25;56(36):4819-4829. Epub 2017 Aug 25.

Department of Pharmaceutical Sciences and ‡Department of Pathology & Microbiology, University of Nebraska Medical Center , Omaha, Nebraska 68198-5900, United States.

Recent studies have revealed an important role for the Staphylococcus aureus CidC enzyme in cell death during the stationary phase and in biofilm development and have contributed to our understanding of the metabolic processes that are important in the induction of bacterial programmed cell death (PCD). To gain more insight into the characteristics of this enzyme, we performed an in-depth biochemical and biophysical analysis of its catalytic properties. In vitro experiments show that this flavoprotein catalyzes the oxidative decarboxylation of pyruvate to acetate and carbon dioxide. CidC efficiently reduces menadione, but not CoenzymeQ, suggesting a specific role in the S. aureus respiratory chain. CidC exists as a monomer under neutral-pH conditions but tends to aggregate and bind to artificial lipid membranes at acidic pH, resulting in enhanced enzymatic activity. Unlike its Escherichia coli counterpart, PoxB, CidC does not appear to be activated by other amphiphiles like Triton X-100 or octyl β-d-glucopyranoside. In addition, only reduced CidC is protected from proteolytic cleavage by chymotrypsin, and unlike its homologues in other bacteria, protease treatment does not increase CidC enzymatic activity. Finally, CidC exhibits maximal activity at pH 5.5-5.8 and negligible activity at pH 7-8. The results of this study are consistent with a model in which CidC functions as a pyruvate:menaquinone oxidoreductase whose activity is induced at the cellular membrane during cytoplasmic acidification, a process previously shown to be important for the induction of bacterial PCD.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acs.biochem.7b00570DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5853648PMC
September 2017

Poly(3-hydroxybutyrate) fuels the tricarboxylic acid cycle and de novo lipid biosynthesis during Bacillus anthracis sporulation.

Mol Microbiol 2017 Jun 30;104(5):793-803. Epub 2017 Mar 30.

Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, NE, 68198, USA.

Numerous bacteria accumulate poly(3-hydroxybutyrate) (PHB) as an intracellular reservoir of carbon and energy in response to imbalanced nutritional conditions. In Bacillus spp., where PHB biosynthesis precedes the formation of the dormant cell type called the spore (sporulation), the direct link between PHB accumulation and efficiency of sporulation was observed in multiple studies. Although the idea of PHB as an intracellular carbon and energy source fueling sporulation was proposed several decades ago, the mechanisms underlying PHB contribution to sporulation have not been defined. Here, we demonstrate that PHB deficiency impairs Bacillus anthracis sporulation through diminishing the energy status of the cells and by reducing carbon flux into the tricarboxylic acid (TCA) cycle and de novo lipid biosynthesis. Consequently, this metabolic imbalance decreased biosynthesis of the critical components required for spore integrity and resistance, such as dipicolinic acid (DPA) and the spore's inner membrane. Supplementation of the PHB deficient mutant with exogenous fatty acids overcame these sporulation defects, highlighting the importance of the TCA cycle and lipid biosynthesis during sporulation. Combined, the results of this work reveal the molecular mechanisms of PHB contribution to B. anthracis sporulation and provide valuable insight into the metabolic requirements for this developmental process in Bacillus species.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/mmi.13665DOI Listing
June 2017

Staphylococcus aureus biofilm: a complex developmental organism.

Mol Microbiol 2017 May 8;104(3):365-376. Epub 2017 Mar 8.

Center for Staphylococcal Research, Department of Pathology & Microbiology, University of Nebraska Medical Center, Omaha, NE, USA.

Chronic biofilm-associated infections caused by Staphylococcus aureus often lead to significant increases in morbidity and mortality, particularly when associated with indwelling medical devices. This has triggered a great deal of research attempting to understand the molecular mechanisms that control S. aureus biofilm formation and the basis for the recalcitrance of these multicellular structures to antibiotic therapy. The purpose of this review is to summarize our current understanding of S. aureus biofilm development, focusing on the description of a newly-defined, five-stage model of biofilm development and the mechanisms required for each stage. Importantly, this model includes an alternate view of the processes involved in microcolony formation in S. aureus and suggests that these structures originate as a result of stochastically regulated metabolic heterogeneity and proliferation within a maturing biofilm population, rather than a subtractive process involving the release of cell clusters from a thick, unstructured biofilm. Importantly, it is proposed that this new model of biofilm development involves the genetically programmed generation of metabolically distinct subpopulations of cells, resulting in an overall population that is better able to adapt to rapidly changing environmental conditions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/mmi.13634DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5397344PMC
May 2017

Simple synthesis of endophenazine G and other phenazines and their evaluation as anti-methicillin-resistant Staphylococcus aureus agents.

Eur J Med Chem 2017 Jan 28;125:710-721. Epub 2016 Sep 28.

Department of Pharmaceutical Sciences, College of Pharmacy, University of Nebraska Medical Center, Omaha, NE 68198, United States. Electronic address:

Community-associated methicillin resistant Staphylococcus aureus (CA-MRSA) has become a severe health concern because of its treatment difficulties. Herein, we report the synthesis and biological evaluation of two phenazine natural products and a series of phenazines that show promising activities against MRSA with MIC values in the low micromolar range. Basic studies revealed that these compounds are bacteriostatic agents. The most active compound also displayed promising IC50 values against HaCat cells. Finally, a QSAR model was developed to understand the key structural features of the molecules.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ejmech.2016.09.079DOI Listing
January 2017

Identification of inhibitors for single-stranded DNA-binding proteins in eubacteria.

J Antimicrob Chemother 2016 12 8;71(12):3432-3440. Epub 2016 Sep 8.

Department of Oral Biology, University of Nebraska Medical Center, Omaha, NE 68583, USA

Objectives: The increasing threat of drug-resistant bacteria establishes a continuing need for the development of new strategies to fight infection. We examine the inhibition of the essential single-stranded DNA-binding proteins (SSBs) SSBA and SSBB as a potential antimicrobial therapy due to their importance in DNA replication, activating the SOS response and promoting competence-based mechanisms of resistance by incorporating new DNA.

Methods: Purified recombinant SSBs from Gram-positive (Staphylococcus aureus and Bacillus anthracis) and Gram-negative (Escherichia coli and Francisella tularensis) bacteria were assessed in a high-throughput screen for inhibition of duplex DNA unwinding by small molecule inhibitors. Secondary electrophoretic mobility shift assays further validated the top hits that were then tested for MICs using in vitro assays.

Results: We have identified compounds that show cross-reactivity in vitro, as well as inhibition of both F. tularensis and B. anthracis SSBA. Five compounds were moderately toxic to at least two of the four bacterial strains in vivo, including two compounds that were selectively non-toxic to human cells, 9-hydroxyphenylfluoron and purpurogallin. Three of the SSBA inhibitors also inhibited S. aureus SSBB in Gram-positive bacteria.

Conclusions: Results from our study support the potential for SSB inhibitors as broad-spectrum antibacterial agents, with dual targeting capabilities against Gram-positive bacteria.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/jac/dkw340DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5181397PMC
December 2016

Novel fluorinated pyrrolomycins as potent anti-staphylococcal biofilm agents: Design, synthesis, pharmacokinetics and antibacterial activities.

Eur J Med Chem 2016 Nov 12;124:129-137. Epub 2016 Aug 12.

UNMC Center for Drug Discovery and Department of Pharmaceutical Sciences, United States; Center for Staphylococcal Research, United States; Fred and Pamela Buffett Cancer Center, University of Nebraska Medical Center, 986125 Nebraska Medical Center, Omaha, NE 68198, United States. Electronic address:

Staphylococcus aureus (SA) is a major cause of hospital- and community-associated bacterial infections in the U.S. and around the world. These infections have become increasingly difficult to treat due to the propensity to develop antibiotic resistance and form biofilm. To date, no antibiofilm agents are available for clinical use. To add to the repertoire of antibiotics for clinical use and to provide novel agents for combating both SA and biofilm infections, we previously reported marinopyrroles as potent anti-SA agents. In this study, we used fragment-based and bioisostere approaches to design and synthesize a series of novel fluorinated pyrrolomycins for the first time, performed analyses of their physicochemical and drug-like properties, and investigated structure activity relationships and pharmacokinetics. These promising fluorinated pyrrolomycins demonstrate potent antibacterial activity against SA with favorable drug-like properties and pharmacokinetic profiles. Importantly, these compounds kill staphylococcal biofilm-associated cells with a lack of mammalian cell cytotoxicity and no occurrence of bacterial resistance. Our novel fluorinated pyrrolomycin 4 has a clogP value of 4.1, an MIC of 73 ng/mL, MBC of 4 μg/mL, kill staphylococcal-associated biofilm at 8 μg/mL, bioavailability of 35%, and the elimination half-life of 6.04 h and 6.75 h by intravenous and oral administration, respectively. This is the first report of comprehensive drug discovery studies on pyrrolomycin-based antibiotics.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ejmech.2016.08.017DOI Listing
November 2016

Potassium Uptake Modulates Staphylococcus aureus Metabolism.

mSphere 2016 May-Jun;1(3). Epub 2016 Jun 15.

Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, Nebraska, USA.

As a leading cause of community-associated and nosocomial infections, Staphylococcus aureus requires sophisticated mechanisms that function to maintain cellular homeostasis in response to its exposure to changing environmental conditions. The adaptation to stress and maintenance of homeostasis depend largely on membrane activity, including supporting electrochemical gradients and synthesis of ATP. This is largely achieved through potassium (K(+)) transport, which plays an essential role in maintaining chemiosmotic homeostasis, affects antimicrobial resistance, and contributes to fitness in vivo. Here, we report that S. aureus Ktr-mediated K(+) uptake is necessary for maintaining cytoplasmic pH and the establishment of a proton motive force. Metabolite analyses revealed that K(+) deficiency affects both metabolic and energy states of S. aureus by impairing oxidative phosphorylation and directing carbon flux toward substrate-level phosphorylation. Taken together, these results underline the importance of K(+) uptake in maintaining essential components of S. aureus metabolism. IMPORTANCE Previous studies describing mechanisms for K(+) uptake in S. aureus revealed that the Ktr-mediated K(+) transport system was required for normal growth under alkaline conditions but not under neutral or acidic conditions. This work focuses on the effect of K(+) uptake on S. aureus metabolism, including intracellular pH and carbon flux, and is the first to utilize a pH-dependent green fluorescent protein (GFP) to measure S. aureus cytoplasmic pH. These studies highlight the role of K(+) uptake in supporting proton efflux under alkaline conditions and uncover a critical role for K(+) uptake in establishing efficient carbon utilization.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/mSphere.00125-16DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4911797PMC
June 2016

The LysR-type transcriptional regulator, CidR, regulates stationary phase cell death in Staphylococcus aureus.

Mol Microbiol 2016 09 4;101(6):942-53. Epub 2016 Jul 4.

Center for Staphylococcal Research, Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, Nebraska, 68198-5900, USA.

The Staphylococcus aureus LysR-type transcriptional regulator, CidR, activates the expression of two operons including cidABC and alsSD that display pro- and anti-death functions, respectively. Although several investigations have focused on the functions of different genes associated with these operons, the collective role of the CidR regulon in staphylococcal physiology is not clearly understood. Here we reveal that the primary role of this regulon is to limit acetate-dependent potentiation of cell death in staphylococcal populations. Although both CidB and CidC promote acetate generation and cell death, the CidR-dependent co-activation of CidA and AlsSD counters the effects of CidBC by redirecting intracellular carbon flux towards acetoin formation. From a mechanistic standpoint, we demonstrate that CidB is necessary for full activation of CidC, whereas CidA limits the abundance of CidC in the cell.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/mmi.13433DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5014633PMC
September 2016

Electron Paramagnetic Resonance (EPR) Spectroscopy to Detect Reactive Oxygen Species in .

Bio Protoc 2015 Sep;5(17)

Center for Staphylococcal Research, Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, USA.

Under aerobic conditions, () primarily metabolizes glucose to acetic acid. Although normally is able to re-utilize acetate as a carbon source following glucose exhaustion, significantly high levels of acetate in the culture media may not only be growth inhibitory but also potentiates cell death in stationary phase cultures by a mechanism dependent on cytoplasmic acidification. One consequence of acetic acid toxicity is the production of reactive oxygen species (ROS). The present protocol describes the detection of ROS in undergoing cell death by electron paramagnetic resonance (EPR) spectroscopy. Using 1-hydroxy-3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine (CMH) as a cell permeable spin probe, we demonstrate the detection of various oxygen radicals generated by bacteria. Although standardized for , the methods described here should be easily adapted for other bacterial species. This protocol is adapted from Thomas (2014) and Thomas (2010).
View Article and Find Full Text PDF

Download full-text PDF

Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4863951PMC
http://dx.doi.org/10.21769/bioprotoc.1586DOI Listing
September 2015

Resistance to Acute Macrophage Killing Promotes Airway Fitness of Prevalent Community-Acquired Staphylococcus aureus Strains.

J Immunol 2016 05 6;196(10):4196-203. Epub 2016 Apr 6.

Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, NE 68198

The incidence of methicillin-resistant Staphylococcus aureus (MRSA) pneumonia in otherwise healthy individuals is increasing. To investigate the mechanism underlying the epidemiological success of predominant community-associated (CA)-MRSA strains, we examined their fitness traits during the initial interaction between bacteria and the host occurring in the lower airway. Using a mouse respiratory infection model, we show that clinical isolates often responsible for CA infections are highly resistant to clearance from healthy airways, whereas S. aureus strains not as prevalent or traditionally associated with hospital-associated infections are relatively susceptible. Mechanistically, the competitive fitness of S. aureus is a result of both agr-dependent and -independent resistance to innate bacterial killing. Furthermore, we show that rather than evasion from neutrophil-dependent bactericidal process, the observed S. aureus fitness in the lower airways is due to its intrinsic resistance to resident alveolar macrophage-mediated intracellular killing. Importantly, we demonstrate that the virulence determinants responsible for bacterial persistence in immune-competent mice are dispensable in mice with predisposing conditions such as influenza infection. Taken together, these novel findings of the improved competence of predominant CA-MRSA strains to survive innate killing in healthy hosts, particularly at the very beginning stage of infection, provide a unique insight into their epidemiological success.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.4049/jimmunol.1600081DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4868659PMC
May 2016

The major autolysin is redundant for Staphylococcus aureus USA300 LAC JE2 virulence in a murine device-related infection model.

FEMS Microbiol Lett 2016 05 3;363(9). Epub 2016 Apr 3.

Department of Microbiology, School of Natural Sciences, National University of Ireland, Galway, Ireland

The major Staphylococcus aureus autolysin, Atl, has been implicated in attachment to surfaces and release of extracellular DNA during biofilm formation under laboratory conditions. Consistent with this, polyclonal antibodies to the amidase and glucosaminidase domains of Atl inhibited in vitro biofilm formation. However, in a murine model of device-related infection the community-associated S. aureus strain USA300 LAC JE2 established a successful infection in the absence of atl These data indicate that Atl activity is not required for biofilm production in this infection model and reveal the importance of characterizing the contribution of biofilm phenotypes to virulence under in vivo conditions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/femsle/fnw087DOI Listing
May 2016

Redox Imbalance Underlies the Fitness Defect Associated with Inactivation of the Pta-AckA Pathway in Staphylococcus aureus.

J Proteome Res 2016 Apr 24;15(4):1205-12. Epub 2016 Mar 24.

Department of Chemistry, University of Nebraska-Lincoln , Lincoln, Nebraska 68588, United States.

The phosphotransacetylase-acetate kinase (Pta-AckA) pathway is thought to be a vital ATP generating pathway for Staphylococcus aureus. Disruption of the Pta-AckA pathway during overflow metabolism causes significant reduction in growth rate and viability, albeit not due to intracellular ATP depletion. Here, we demonstrate that toxicity associated with inactivation of the Pta-AckA pathway resulted from an altered intracellular redox environment. Growth of the pta and ackA mutants under anaerobic conditions partially restored cell viability. NMR metabolomics analyses and (13)C6-glucose metabolism tracing experiments revealed the activity of multiple pathways that promote redox (NADH/NAD(+)) turnover to be enhanced in the pta and ackA mutants during anaerobic growth. Restoration of redox homeostasis in the pta mutant by overexpressing l- lactate dehydrogenase partially restored its viability under aerobic conditions. Together, our findings suggest that during overflow metabolism, the Pta-AckA pathway plays a critical role in preventing cell viability defects by promoting intracellular redox homeostasis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acs.jproteome.5b01089DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4875753PMC
April 2016

Effects of Low-Dose Amoxicillin on Staphylococcus aureus USA300 Biofilms.

Antimicrob Agents Chemother 2016 05 22;60(5):2639-51. Epub 2016 Apr 22.

Department of Biology, American University, Washington, DC, USA

Previous studies showed that sub-MIC levels of β-lactam antibiotics stimulate biofilm formation in most methicillin-resistant Staphylococcus aureus (MRSA) strains. Here, we investigated this process by measuring the effects of sub-MIC amoxicillin on biofilm formation by the epidemic community-associated MRSA strain USA300. We found that sub-MIC amoxicillin increased the ability of USA300 cells to attach to surfaces and form biofilms under both static and flow conditions. We also found that USA300 biofilms cultured in sub-MIC amoxicillin were thicker, contained more pillar and channel structures, and were less porous than biofilms cultured without antibiotic. Biofilm formation in sub-MIC amoxicillin correlated with the production of extracellular DNA (eDNA). However, eDNA released by amoxicillin-induced cell lysis alone was evidently not sufficient to stimulate biofilm. Sub-MIC levels of two other cell wall-active agents with different mechanisms of action-d-cycloserine and fosfomycin-also stimulated eDNA-dependent biofilm, suggesting that biofilm formation may be a mechanistic adaptation to cell wall stress. Screening a USA300 mariner transposon library for mutants deficient in biofilm formation in sub-MIC amoxicillin identified numerous known mediators of S. aureus β-lactam resistance and biofilm formation, as well as novel genes not previously associated with these phenotypes. Our results link cell wall stress and biofilm formation in MRSA and suggest that eDNA-dependent biofilm formation by strain USA300 in low-dose amoxicillin is an inducible phenotype that can be used to identify novel genes impacting MRSA β-lactam resistance and biofilm formation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/AAC.02070-15DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4862544PMC
May 2016
-->