scholarly journals Staphylococcus aureus Preferentially Liberates Inorganic Phosphate from Organophosphates in Environments where This Nutrient Is Limiting

2020 ◽  
Vol 202 (22) ◽  
Author(s):  
Jessica L. Kelliher ◽  
Aleeza J. Leder Macek ◽  
Kevin M. Grudzinski ◽  
Jana N. Radin ◽  
Thomas E. Kehl-Fie
Keyword(s):  
Staphylococcus Aureus ◽  
Inorganic Phosphate ◽  
Organic Molecules ◽  
Model Organism ◽  
Mammalian Host ◽  
Content Type ◽  
Phosphoamino Acids ◽  
Phosphorus Containing ◽  
Essential Nutrient ◽  
Phosphate Source

ABSTRACT Phosphate is an essential nutrient that Staphylococcus aureus and other pathogens must acquire from the host during infection. While inorganic monophosphate (Pi) is the preferred source of this nutrient, bacteria can also obtain it from phosphate-containing organic molecules. The Pi-responsive regulator PhoPR is necessary for S. aureus to cause infection, suggesting that Pi is not freely available during infection and that this nutrient must be obtained from other sources. However, the organophosphates from which S. aureus can obtain phosphate are unknown. We evaluated the ability of 58 phosphorus-containing molecules to serve as phosphate sources for S. aureus. Forty-six of these compounds, including phosphorylated amino acids, sugars, and nucleotides, supported growth. Among the organophosphate sources was glycerol-3-phosphate (G3P), which is commonly found in the mammalian host. Differing from the model organism Escherichia coli, S. aureus does not import G3P intact to obtain Pi. Instead, S. aureus relies on the phosphatase PhoB to release Pi from G3P, which is subsequently imported by Pi transporters. To determine if this strategy is used by S. aureus to extract phosphate from other phosphate sources, we assessed the ability of PhoB- and Pi transporter-deficient strains to grow on the same library of phosphorus-containing molecules. Sixty percent of the substrates (28/46) relied on the PhoB/Pi transporter pathway, and an additional 10/46 (22%) were PhoB independent but still required Pi transport through the Pi transporters. Cumulatively, these results suggest that in Pi-limited environments, S. aureus preferentially generates Pi from organophosphates and then relies on Pi transporters to import this nutrient. IMPORTANCE For bacteria, the preferred form of the essential nutrient phosphate is inorganic monophosphate (Pi), but phosphate can also be extracted from a variety of phosphocompounds. Pathogens, including Staphylococcus aureus, experience Pi limitation within the host, suggesting that the use of alternative phosphate sources is important during infection. However, the alternative phosphate sources that can be used by S. aureus and others remain largely unexplored. We screened a library of phosphorus-containing compounds for the ability to support growth as a phosphate source. S. aureus could use a variety of phosphocompounds, including nucleotides, phosphosugars, and phosphoamino acids. Subsequent genetic analysis determined that a majority of these alternative phosphate sources are first processed extracellularly to liberate Pi, which is then imported through Pi transporters.

scholarly journals Staphylococcus aureusCoproporphyrinogen III Oxidase Is Required for Aerobic and Anaerobic Heme Synthesis

mSphere ◽  
2019 ◽  
Vol 4 (4) ◽  
Author(s):  
Jacob E. Choby ◽  
Eric P. Skaar
Keyword(s):  
Staphylococcus Aureus ◽  
Anaerobic Respiration ◽  
Model Organism ◽  
Anaerobic Growth ◽  
Cellular Respiration ◽  
Nitrate Respiration ◽  
Human Pathogen ◽  
Content Type ◽  
Heme Synthesis ◽  
Aerobic And Anaerobic

ABSTRACTThe virulence of the human pathogenStaphylococcus aureusis supported by many heme-dependent proteins, including key enzymes of cellular respiration. Therefore, synthesis of heme is a critical component of staphylococcal physiology.S. aureusgenerates heme via the coproporphyrin-dependent pathway, conserved across members of theFirmicutesandActinobacteria. In this work, we genetically investigate the oxidation of coproporphyrinogen to coproporphyrin in this heme synthesis pathway. The coproporphyrinogen III oxidase CgoX has previously been identified as the oxygen-dependent enzyme responsible for this conversion under aerobic conditions. However, becauseS. aureususes heme during anaerobic nitrate respiration, we hypothesized that coproporphyrin production is able to proceed in the absence of oxygen. Therefore, we tested the contribution to anaerobic heme synthesis of CgoX and two other proteins previously identified as potential oxygen-independent coproporphyrinogen dehydrogenases, NWMN_1486 and NWMN_1636. We have found that CgoX alone is responsible for aerobic and anaerobic coproporphyrin synthesis from coproporphyrinogen and is required for aerobic and anaerobic heme-dependent growth. This work provides an explanation for howS. aureusheme synthesis proceeds under both aerobic and anaerobic conditions.IMPORTANCEHeme is a critical molecule required for aerobic and anaerobic respiration by organisms across kingdoms. The human pathogenStaphylococcus aureushas served as a model organism for the study of heme synthesis and heme-dependent physiology and, like many species of the phylaFirmicutesandActinobacteria, generates heme through a coproporphyrin intermediate. A critical step in terminal heme synthesis is the production of coproporphyrin by the CgoX enzyme, which was presumed to be oxygen dependent. However,S. aureusalso requires heme during anaerobic growth; therefore, the synthesis of coproporphyrin by an oxygen-independent mechanism is required. Here, we identify CgoX as the enzyme performing the oxygen-dependent and -independent synthesis of coproporphyrin from coproporphyrinogen, resolving a key outstanding question in the coproporphyrin-dependent heme synthesis pathway.

scholarly journals De Novo Purine Biosynthesis Is Required for Intracellular Growth of Staphylococcus aureus and for the Hypervirulence Phenotype of a purR Mutant

2020 ◽  
Vol 88 (5) ◽  
Author(s):  
Mariya I. Goncheva ◽  
Ronald S. Flannagan ◽  
David E. Heinrichs
Keyword(s):  
Staphylococcus Aureus ◽  
Virulence Factors ◽  
Metabolic Regulation ◽  
De Novo ◽  
Purine Biosynthesis ◽  
Mammalian Host ◽  
Cell Binding ◽  
Intracellular Growth ◽  
Content Type ◽  
De Novo Purine Biosynthesis

ABSTRACT Staphylococcus aureus is a noted human and animal pathogen. Despite decades of research on this important bacterium, there are still many unanswered questions regarding the pathogenic mechanisms it uses to infect the mammalian host. This can be attributed to it possessing a plethora of virulence factors and complex virulence factor and metabolic regulation. PurR, the purine biosynthesis regulator, was recently also shown to regulate virulence factors in S. aureus, and mutations in purR result in derepression of fibronectin binding proteins (FnBPs) and extracellular toxins, required for a so-called hypervirulent phenotype. Here, we show that hypervirulent strains containing purR mutations can be attenuated with the addition of purine biosynthesis mutations, implicating the necessity for de novo purine biosynthesis in this phenotype and indicating that S. aureus in the mammalian host experiences purine limitation. Using cell culture, we showed that while purR mutants are not altered in epithelial cell binding, compared to that of wild-type (WT) S. aureus, purR mutants have enhanced invasion of these nonprofessional phagocytes, consistent with the requirement of FnBPs for invasion of these cells. This correlates with purR mutants having increased transcription of fnb genes, resulting in higher levels of surface-exposed FnBPs to promote invasion. These data provide important contributions to our understanding of how the pathogenesis of S. aureus is affected by sensing of purine levels during infection of the mammalian host.

scholarly journals Daptomycin Tolerance in the Staphylococcus aureus pitA6 Mutant Is Due to Upregulation of thedltOperon

2016 ◽  
Vol 60 (5) ◽  
pp. 2684-2691 ◽  
Author(s):  
Lukas Mechler ◽  
Eve-Julie Bonetti ◽  
Sebastian Reichert ◽  
Matthias Flötenmeyer ◽  
Jacques Schrenzel ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Inorganic Phosphate ◽  
Protein Biosynthesis ◽  
Resistance Mechanisms ◽  
Important Change ◽  
Intracellular Accumulation ◽  
Content Type ◽  
Carbohydrate And Lipid Metabolism ◽  
Important Object

ABSTRACTUnderstanding the mechanisms of how bacteria become tolerant toward antibiotics during clinical therapy is a very important object. In a previous study, we showed that increased daptomycin (DAP) tolerance ofStaphylococcus aureuswas due to a point mutation inpitA(inorganic phosphate transporter) that led to intracellular accumulation of both inorganic phosphate (Pi) and polyphosphate (polyP). DAP tolerance in thepitA6mutant differs from classical resistance mechanisms since there is no increase in the MIC. In this follow-up study, we demonstrate that DAP tolerance in thepitA6mutant is not triggered by the accumulation of polyP. Transcriptome analysis revealed that 234 genes were at least 2.0-fold differentially expressed in the mutant. Particularly, genes involved in protein biosynthesis, carbohydrate and lipid metabolism, and replication and maintenance of DNA were downregulated. However, the most important change was the upregulation of thedltoperon, which is induced by the accumulation of intracellular Pi. The GraXRS system, known as an activator of thedltoperon (d-alanylation of teichoic acids) and of themprFgene (multiple peptide resistance factor), is not involved in DAP tolerance of thepitA6mutant. In conclusion, DAP tolerance of thepitA6mutant is due to an upregulation of thedltoperon, triggered directly or indirectly by the accumulation of Pi.

scholarly journals ϕSa3mw Prophage as a Molecular Regulatory Switch ofStaphylococcus aureusβ-Toxin Production

2019 ◽  
Vol 201 (14) ◽  
Author(s):  
Phuong M. Tran ◽  
Michael Feiss ◽  
Kyle J. Kinney ◽  
Wilmara Salgado-Pabón
Keyword(s):  
Staphylococcus Aureus ◽  
Plaque Assay ◽  
Growth Conditions ◽  
Lytic Cycle ◽  
Host Cells ◽  
Mammalian Host ◽  
Toxin Gene ◽  
Biofilm Growth ◽  
Content Type ◽  
Conditional Expression

ABSTRACTPhage regulatory switches (phage-RSs) are a newly described form of active lysogeny where prophages function as regulatory mechanisms for expression of chromosomal bacterial genes. InStaphylococcus aureus, ϕSa3int is a widely distributed family of prophages that integrate into the β-toxin structural genehlb, effectively inactivating it. However, β-toxin-producing strains often arise during infections and are more virulent in experimental infective endocarditis and pneumonia infections. We present evidence that inS. aureusMW2, ϕSa3mw excision is temporally and differentially responsive to growth conditions relevant toS. aureuspathogenesis. PCR analyses of ϕSa3mw (integrated and excised) and of intacthlbshowed that ϕSa3mw preferentially excises in response to hydrogen peroxide-induced oxidative stress and during biofilm growth. ϕSa3mw remains as a prophage when in contact with human aortic endothelial cells in culture. A criterion for a prophage to be considered a phage-RS is the inability to lyse host cells. MW2 grown under phage-inducing conditions did not release infectious phage particles by plaque assay or transmission electron microscopy, indicating that ϕSa3mw does not carry out a productive lytic cycle. These studies highlight a dynamic, and perhaps more sophisticated,S. aureus-prophage interaction where ϕSa3int prophages provide a novel regulatory mechanism for the conditional expression of virulence factors.IMPORTANCEβ-Toxin is a sphingomyelinase hemolysin that significantly contributes toStaphylococcus aureuspathogenesis. In mostS. aureusisolates the prophage ϕSa3int inserts into the β-toxin genehlb, inactivating it, but human and experimental infections give rise to β-toxin-producing variants. However, it remained to be established whether ϕSa3mw excises in response to specific environmental cues, restoring the β-toxin gene sequence. This is not only of fundamental interest but also critical when designing intervention strategies and therapeutics. We provide evidence that ϕSa3mw actively excises, allowing the conditional expression of β-toxin. ϕSa3int prophages may play a novel and largely uncharacterized role inS. aureuspathogenesis as molecular regulatory switches that promote bacterial fitness and adaptation to the challenges presented by the mammalian host.

scholarly journals Acquisition of the Phosphate Transporter NptA Enhances Staphylococcus aureus Pathogenesis by Improving Phosphate Uptake in Divergent Environments

2017 ◽  
Vol 86 (1) ◽  
Author(s):  
Jessica L. Kelliher ◽  
Jana N. Radin ◽  
Kyle P. Grim ◽  
Paola K. Párraga Solórzano ◽  
Patrick H. Degnan ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Mouse Model ◽  
Inorganic Phosphate ◽  
Phosphate Uptake ◽  
Genomic Analysis ◽  
Phosphate Transporter ◽  
Phosphate Limitation ◽  
Content Type ◽  
Staphylococcal Species

ABSTRACTDuring infection, pathogens must obtain all inorganic nutrients, such as phosphate, from the host. Despite the essentiality of phosphate for all forms of life, howStaphylococcus aureusobtains this nutrient during infection is unknown. Differing fromEscherichia coli, the paradigm for bacterial phosphate acquisition, which has two inorganic phosphate (Pi) importers, genomic analysis suggested thatS. aureuspossesses three distinct Pitransporters: PstSCAB, PitA, and NptA. WhilepitAandnptAare expressed in phosphate-replete media, expression of all three transporters is induced by phosphate limitation. The loss of a single transporter did not affectS. aureus. However, disruption of any two systems significantly reduced Piaccumulation and growth in divergent environments. These findings indicate that PstSCAB, PitA, and NptA have overlapping but nonredundant functions, thus expanding the environments in whichS. aureuscan successfully obtain Pi. Consistent with this idea, in a systemic mouse model of disease, loss of any one transporter did not decrease staphylococcal virulence. However, loss of NptA in conjunction with either PstSCAB or PitA significantly reduced the ability ofS. aureusto cause infection. These observations suggest that Piacquisition via NptA is particularly important for the pathogenesis ofS. aureus. While our analysis suggests that NptA homologs are widely distributed among bacteria, closely related less pathogenic staphylococcal species do not possess this importer. Altogether, these observations indicate that Piuptake byS. aureusdiffers from established models and that acquisition of a third transporter enhances the ability of the bacterium to cause infection.

scholarly journals Cigarette Smoke Increases Staphylococcus aureus Biofilm Formation via Oxidative Stress

2012 ◽  
Vol 80 (11) ◽  
pp. 3804-3811 ◽  
Author(s):  
Ritwij Kulkarni ◽  
Swati Antala ◽  
Alice Wang ◽  
Fábio E. Amaral ◽  
Ryan Rampersaud ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Cigarette Smoke ◽  
Biofilm Formation ◽  
Pathogenic Bacteria ◽  
Protein A ◽  
Model Organism ◽  
Human Cells ◽  
Dependent Manner ◽  
Content Type ◽  
Tract Infections

ABSTRACTThe strong epidemiological association between cigarette smoke (CS) exposure and respiratory tract infections is conventionally attributed to immunosuppressive and irritant effects of CS on human cells. Since pathogenic bacteria such asStaphylococcus aureusare members of the normal microbiota and reside in close proximity to human nasopharyngeal cells, we hypothesized that bioactive components of CS might affect these organisms and potentiate their virulence. UsingStaphylococcus aureusas a model organism, we observed that the presence of CS increased both biofilm formation and host cell adherence. Analysis of putative molecular pathways revealed that CS exposure decreased expression of the quorum-sensingagrsystem, which is involved in biofilm dispersal, and increased transcription of biofilm inducers such assarAandrbf. CS contains bioactive compounds, including free radicals and reactive oxygen species, and we observed transcriptional induction of bacterial oxidoreductases, including superoxide dismutase, following exposure. Moreover, pretreatment of CS with an antioxidant abrogated CS-mediated enhancement of biofilms. Exposure of bacteria to hydrogen peroxide alone increased biofilm formation. These observations are consistent with the hypothesis that CS induces staphylococcal biofilm formation in an oxidant-dependent manner. CS treatment induced transcription offnbA(encoding fibronectin binding protein A), leading to increased binding of CS-treated staphylococci to immobilized fibronectin and increased adherence to human cells. These observations indicate that the bioactive effects of CS may extend to the resident microbiota of the nasopharynx, with implications for the pathogenesis of respiratory infection in CS-exposed humans.

scholarly journals Metabolic reprogramming during the Trypanosoma brucei life cycle

F1000Research ◽  
2017 ◽  
Vol 6 ◽  
pp. 683 ◽  
Author(s):  
Terry K. Smith ◽  
Frédéric Bringaud ◽  
Derek P. Nolan ◽  
Luisa M. Figueiredo
Keyword(s):  
Life Cycle ◽  
Trypanosoma Brucei ◽  
Model Organism ◽  
Protozoan Parasite ◽  
Tsetse Fly ◽  
Metabolic Reprogramming ◽  
Mammalian Host ◽  
Insect Vector ◽  
Environmental Signals ◽  
Cellular Metabolic Activity

Cellular metabolic activity is a highly complex, dynamic, regulated process that is influenced by numerous factors, including extracellular environmental signals, nutrient availability and the physiological and developmental status of the cell. The causative agent of sleeping sickness, Trypanosoma brucei, is an exclusively extracellular protozoan parasite that encounters very different extracellular environments during its life cycle within the mammalian host and tsetse fly insect vector. In order to meet these challenges, there are significant alterations in the major energetic and metabolic pathways of these highly adaptable parasites. This review highlights some of these metabolic changes in this early divergent eukaryotic model organism.

scholarly journals Pseudomonas aeruginosa PA14 Enhances the Efficacy of Norfloxacin against Staphylococcus aureus Newman Biofilms

2020 ◽  
Vol 202 (18) ◽  
Author(s):  
Giulia Orazi ◽  
Fabrice Jean-Pierre ◽  
George A. O’Toole
Keyword(s):  
Cystic Fibrosis ◽  
Staphylococcus Aureus ◽  
Pseudomonas Aeruginosa ◽  
Antimicrobial Agents ◽  
Respiratory Infections ◽  
Multiple Infection ◽  
Bacterial Biofilms ◽  
Interspecies Interactions ◽  
Chronic Infections ◽  
Content Type

ABSTRACT The thick mucus within the airways of individuals with cystic fibrosis (CF) promotes frequent respiratory infections that are often polymicrobial. Pseudomonas aeruginosa and Staphylococcus aureus are two of the most prevalent pathogens that cause CF pulmonary infections, and both are among the most common etiologic agents of chronic wound infections. Furthermore, the ability of P. aeruginosa and S. aureus to form biofilms promotes the establishment of chronic infections that are often difficult to eradicate using antimicrobial agents. In this study, we found that multiple LasR-regulated exoproducts of P. aeruginosa, including 2-heptyl-4-hydroxyquinoline N-oxide (HQNO), siderophores, phenazines, and rhamnolipids, likely contribute to the ability of P. aeruginosa PA14 to shift S. aureus Newman norfloxacin susceptibility profiles. Here, we observe that exposure to P. aeruginosa exoproducts leads to an increase in intracellular norfloxacin accumulation by S. aureus. We previously showed that P. aeruginosa supernatant dissipates the S. aureus membrane potential, and furthermore, depletion of the S. aureus proton motive force recapitulates the effect of the P. aeruginosa PA14 supernatant on shifting norfloxacin sensitivity profiles of biofilm-grown S. aureus Newman. From these results, we hypothesize that exposure to P. aeruginosa PA14 exoproducts leads to increased uptake of the drug and/or an impaired ability of S. aureus Newman to efflux norfloxacin. Surprisingly, the effect observed here of P. aeruginosa PA14 exoproducts on S. aureus Newman susceptibility to norfloxacin seemed to be specific to these strains and this antibiotic. Our results illustrate that microbially derived products can alter the ability of antimicrobial agents to kill bacterial biofilms. IMPORTANCE Pseudomonas aeruginosa and Staphylococcus aureus are frequently coisolated from multiple infection sites, including the lungs of individuals with cystic fibrosis (CF) and nonhealing diabetic foot ulcers. Coinfection with P. aeruginosa and S. aureus has been shown to produce worse outcomes compared to infection with either organism alone. Furthermore, the ability of these pathogens to form biofilms enables them to cause persistent infection and withstand antimicrobial therapy. In this study, we found that P. aeruginosa-secreted products dramatically increase the ability of the antibiotic norfloxacin to kill S. aureus biofilms. Understanding how interspecies interactions alter the antibiotic susceptibility of bacterial biofilms may inform treatment decisions and inspire the development of new therapeutic strategies.

scholarly journals The Serratia marcescens Siderophore Serratiochelin Is Necessary for Full Virulence during Bloodstream Infection

2020 ◽  
Vol 88 (8) ◽  
Author(s):  
Danelle R. Weakland ◽  
Sara N. Smith ◽  
Bailey Bell ◽  
Ashootosh Tripathi ◽  
Harry L. T. Mobley
Keyword(s):  
Serratia Marcescens ◽  
Bloodstream Infection ◽  
Iron Acquisition ◽  
Gene Clusters ◽  
Clinical Isolates ◽  
Wild Type ◽  
Serratia Plymuthica ◽  
Content Type ◽  
Cas Assay ◽  
Essential Nutrient

ABSTRACT Serratia marcescens is a bacterium frequently found in the environment, but over the last several decades it has evolved into a concerning clinical pathogen, causing fatal bacteremia. To establish such infections, pathogens require specific nutrients; one very limited but essential nutrient is iron. We sought to characterize the iron acquisition systems in S. marcescens isolate UMH9, which was recovered from a clinical bloodstream infection. Using RNA sequencing (RNA-seq), we identified two predicted siderophore gene clusters (cbs and sch) that were regulated by iron. Mutants were constructed to delete each iron acquisition locus individually and in conjunction, generating both single and double mutants for the putative siderophore systems. Mutants lacking the sch gene cluster lost their iron-chelating ability as quantified by the chrome azurol S (CAS) assay, whereas the cbs mutant retained wild-type activity. Mass spectrometry-based analysis identified the chelating siderophore to be serratiochelin, a siderophore previously identified in Serratia plymuthica. Serratiochelin-producing mutants also displayed a decreased growth rate under iron-limited conditions created by dipyridyl added to LB medium. Additionally, mutants lacking serratiochelin were significantly outcompeted during cochallenge with wild-type UMH9 in the kidneys and spleen after inoculation via the tail vein in a bacteremia mouse model. This result was further confirmed by an independent challenge, suggesting that serratiochelin is required for full S. marcescens pathogenesis in the bloodstream. Nine other clinical isolates have at least 90% protein identity to the UMH9 serratiochelin system; therefore, our results are broadly applicable to emerging clinical isolates of S. marcescens causing bacteremia.

scholarly journals Genome Sequences of a Staphylococcus aureus Clinical Isolate, Strain SMA0034-04 (UGA22), from Siaya County Referral Hospital in Siaya, Kenya

2019 ◽  
Vol 8 (17) ◽  
Author(s):  
Gary Xie ◽  
Qiuying Cheng ◽  
Hajnalka Daligault ◽  
Karen Davenport ◽  
Cheryl Gleasner ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Clinical Isolate ◽  
Referral Hospital ◽  
Genome Sequences ◽  
Content Type

Here, we report the genome sequences of a Staphylococcus aureus clinical isolate, strain SMA0034-04 (UGA22), which contains one chromosome and one plasmid. We also reveal that isolate SMA0034-04 (UGA22) contains loci in the genome that encode multiple exotoxins.

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