Alanyl-Phosphatidylglycerol and Lysyl-Phosphatidylglycerol Are Translocated by the Same MprF Flippases and Have Similar Capacities To Protect against the Antibiotic Daptomycin in Staphylococcus aureus

ABSTRACTThe lysinylation of negatively charged phosphatidylglycerol by MprF proteins reduces the affinity of cationic antimicrobial peptides (CAMPs) for bacterial cytoplasmic membranes and reduces the susceptibility of several Gram-positive bacterial pathogens to CAMPs. MprF ofStaphylococcus aureusencompasses a lysyl-phosphatidylglycerol (Lys-PG) synthase and a Lys-PG flippase domain. In contrast,Clostridium perfringensencodes two MprF homologs which specifically synthesize alanyl-phosphatidylglycerol (Ala-PG) or Lys-PG, while only the Lys-PG synthase is fused to a putative flippase domain. It remains unknown whether cationic Lys-PG and zwitterionic Ala-PG differ in their capacities to be translocated by MprF flippases and if both can reduce CAMP susceptibility in Gram-positive bacteria. By expressing the MprF proteins ofC. perfringensin anS. aureus mprFdeletion mutant, we found that both lipids can be efficiently produced inS. aureus. Simultaneous expression of the Lys-PG and Ala-PG synthases led to the production of both lipids and slightly increased the overall amounts of aminoacyl phospholipids. Ala-PG production by the correspondingC. perfringensenzyme did not affect susceptibility to CAMPs such as nisin and gallidermin or to the CAMP-like antibiotic daptomycin. However, coexpression of the Ala-PG synthase with flippase domains of Lys-PG synthesizing MprF proteins led to a wild-type level of daptomycin susceptibility, indicating that Ala-PG can also protect bacterial membranes against daptomycin and suggesting that Lys-PG flippases can also translocate the related lipid Ala-PG. Thus, bacterial aminoacyl phospholipid flippases exhibit more relaxed substrate specificity and Ala-PG and Lys-PG are more similar in their capacities to modulate membrane functions than anticipated.

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Activities of Oxadiazole Antibacterials against Staphylococcus aureus and Other Gram-Positive Bacteria

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00453-18 ◽

2018 ◽

Vol 62 (8) ◽

Cited By ~ 2

Author(s):

Sara Ceballos ◽

Choon Kim ◽

Derong Ding ◽

Shahriar Mobashery ◽

Mayland Chang ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Gram Positive ◽

Methicillin Resistant ◽

Content Type ◽

Gram Positive Bacteria ◽

Vancomycin Resistant Enterococci ◽

Vancomycin Resistant ◽

Mrsa Strains

ABSTRACT The activities of four oxadiazoles were investigated with 210 methicillin-resistant Staphylococcus aureus (MRSA) strains. MIC50 and MIC90 values of 1 to 2 and 4 μg/ml, respectively, were observed. We also evaluated the activity of oxadiazole ND-421 against other staphylococci and enterococci and in the presence of oxacillin for selected MRSA strains. The MIC for ND-421 is lowered severalfold in combination with oxacillin, as they synergize. The MIC90 of ND-421 against vancomycin-resistant enterococci is ≤1 μg/ml.

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Inhibition of Methionyl-tRNA Synthetase by REP8839 and Effects of Resistance Mutations on Enzyme Activity

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00275-08 ◽

2008 ◽

Vol 53 (1) ◽

pp. 86-94 ◽

Cited By ~ 30

Author(s):

Louis S. Green ◽

James M. Bullard ◽

Wendy Ribble ◽

Frank Dean ◽

David F. Ayers ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Mechanistic Explanation ◽

Trna Synthetase ◽

Turnover Rates ◽

Wild Type ◽

Resistance Mutations ◽

Gram Positive ◽

Gram Positive Bacteria ◽

Human Orthologs ◽

Methionyl Trna Synthetase

ABSTRACT REP8839 is a selective inhibitor of methionyl-tRNA synthetase (MetRS) with antibacterial activity against a variety of gram-positive organisms. We determined REP8839 potency against Staphylococcus aureus MetRS and assessed its selectivity for bacterial versus human orthologs of MetRS. The inhibition constant (Ki ) of REP8839 was 10 pM for Staphylococcus aureus MetRS. Inhibition of MetRS by REP8839 was competitive with methionine and uncompetitive with ATP. Thus, high physiological ATP levels would actually facilitate optimal binding of the inhibitor. While many gram-positive bacteria, such as Staphylococcus aureus, express exclusively the MetRS1 subtype, many gram-negative bacteria express an alternative homolog called MetRS2. Some gram-positive bacteria, such as Streptococcus pneumoniae and Bacillus anthracis, express both MetRS1 and MetRS2. MetRS2 orthologs were considerably less susceptible to REP8839 inhibition. REP8839 inhibition of human mitochondrial MetRS was 1,000-fold weaker than inhibition of Staphylococcus aureus MetRS; inhibition of human cytoplasmic MetRS was not detectable, corresponding to >1,000,000-fold selectivity for the bacterial target relative to its cytoplasmic counterpart. Mutations in MetRS that confer reduced susceptibility to REP8839 were examined. The mutant MetRS enzymes generally exhibited substantially impaired catalytic activity, particularly in aminoacylation turnover rates. REP8839 Ki values ranged from 4- to 190,000-fold higher for the mutant enzymes than for wild-type MetRS. These observations provide a potential mechanistic explanation for the reduced growth fitness observed with MetRS mutant strains relative to that with wild-type Staphylococcus aureus.

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Daptomycin versus Vancomycin in a Methicillin-Resistant Staphylococcus aureus Endophthalmitis Rabbit Model: Bactericidal Effect, Safety, and Ocular Pharmacokinetics

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.05745-11 ◽

2012 ◽

Vol 56 (5) ◽

pp. 2485-2492 ◽

Cited By ~ 9

Author(s):

Sophie Lefèvre ◽

Maher Saleh ◽

Luc Marcellin ◽

Audrey Subilia ◽

Tristan Bourcier ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Rabbit Model ◽

Bacterial Count ◽

Ocular Inflammation ◽

Gram Positive ◽

Methicillin Resistant ◽

Content Type ◽

Gram Positive Bacteria ◽

Bactericidal Efficacy ◽

The Mean

ABSTRACTStaphylococcus aureusis a frequent cause of acute endophthalmitis, and infection with this virulent bacterium is often associated with a poor visual outcome. In this study, we investigated the bactericidal efficacy and the safety of intravitreal daptomycin (DAP), a lipopeptide antibiotic with broad-spectrum activity against Gram-positive bacteria, compared with those of intravitreal vancomycin (VAN) in a methicillin-resistantS. aureusendophthalmitis rabbit model. The pharmacokinetics and pharmacodynamics of daptomycin in the infected eyes were also studied. Rabbits were randomly divided into three treatment groups (n= 8) and one untreated group (n= 4), to compare the effect of single intravitreal injections of 0.2 mg and 1 mg of daptomycin (DAP 0.2 and DAP 1 groups, respectively) with that of 1 mg of intravitreal vancomycin (VAN 1 group). Vitreal aspirates were regularly collected and grading of ocular inflammation was regularly performed until euthanasia on day 7. In the DAP 0.2 group, 62.5% of the eyes were sterilized and the mean bacterial count presented a reduction of 1 log unit. In the DAP 1 and VAN 1 groups, the infection was eradicated (100% and 87.5% of eyes sterilized, respectively), with a 4-log-unit reduction of the mean bacterial count. The bactericidal efficacy in the DAP 1 group was not inferior to that in the VAN 1 group and was superior to that of the other regimens in limiting the ocular inflammation and preserving the architecture of the ocular structures (P< 0.05). The elimination half-life (t1/2β) of daptomycin was independent of the administered dose (38.8 ± 16.5 h and 40.9 ± 6.7 h, respectively, for the DAP 0.2 and DAP 1 groups) and was significantly longer than thet1/2βof vancomycin (20.5 ± 2.0 h for the VAN 1 group) (P< 0.05). This antibiotic could therefore be considered for the treatment of intraocular infections caused by Gram-positive bacteria.

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Alanine Esters of Enterococcal Lipoteichoic Acid Play a Role in Biofilm Formation and Resistance to Antimicrobial Peptides

Infection and Immunity ◽

10.1128/iai.00111-06 ◽

2006 ◽

Vol 74 (7) ◽

pp. 4164-4171 ◽

Cited By ~ 130

Author(s):

Francesca Fabretti ◽

Christian Theilacker ◽

Lucilla Baldassarri ◽

Zbigniew Kaczynski ◽

Andrea Kropec ◽

...

Keyword(s):

Antimicrobial Peptides ◽

Deletion Mutant ◽

Lipoteichoic Acid ◽

Polymyxin B ◽

Resonance Spectroscopy ◽

Wild Type ◽

Gram Positive ◽

Gram Positive Bacteria ◽

Surface Molecules ◽

Environmental Interactions

ABSTRACT Enterococcus faecalis is among the predominant causes of nosocomial infections. Surface molecules like d-alanine lipoteichoic acid (LTA) perform several functions in gram-positive bacteria, such as maintenance of cationic homeostasis and modulation of autolytic activities. The aim of the present study was to evaluate the effect of d-alanine esters of teichoic acids on biofilm production and adhesion, autolysis, antimicrobial peptide sensitivity, and opsonic killing. A deletion mutant of the dltA gene was created in a clinical E. faecalis isolate. The absence of d-alanine in the LTA of the dltA deletion mutant was confirmed by nuclear magnetic resonance spectroscopy. The wild-type strain and the deletion mutant did not show any significant differences in growth curve, morphology, or autolysis. However, the mutant produced significantly less biofilm when grown in the presence of 1% glucose (51.1% compared to that of the wild type); adhesion to eukaryotic cells was diminished. The mutant absorbed 71.1% of the opsonic antibodies, while absorption with the wild type resulted in a 93.2% reduction in killing. Sensitivity to several cationic antimicrobial peptides (polymyxin B, colistin, and nisin) was considerably increased in the mutant strain, confirming similar results from other studies of gram-positive bacteria. Our data suggest that the absence of d-alanine in LTA plays a role in environmental interactions, probably by modulating the net negative charge of the bacterial cell surface, and therefore it may be involved in the pathogenesis of this organism.

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Staphylococcus aureus Extracellular Vesicles Carry Biologically Active β-Lactamase

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00522-12 ◽

2013 ◽

Vol 57 (6) ◽

pp. 2589-2595 ◽

Cited By ~ 75

Author(s):

Jaewook Lee ◽

Eun-Young Lee ◽

Si-Hyun Kim ◽

Dae-Kyum Kim ◽

Kyong-Su Park ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Extracellular Vesicles ◽

Extracellular Vesicle ◽

Biologically Active ◽

Gram Positive ◽

Content Type ◽

Gram Positive Bacteria ◽

Heat Treated ◽

Protease Digestion

ABSTRACTGram-positive bacteria naturally produce extracellular vesicles. However, little is known regarding the functions of Gram-positive bacterial extracellular vesicles, especially in the bacterial community. Here, we investigated the role ofStaphylococcus aureusextracellular vesicles in interbacterial communication to cope with antibiotic stress. We found thatS. aureusliberated BlaZ, a β-lactamase protein, via extracellular vesicles. These extracellular vesicles enabled other ampicillin-susceptible Gram-negative and Gram-positive bacteria to survive in the presence of ampicillin. However,S. aureusextracellular vesicles did not mediate the survival of tetracycline-, chloramphenicol-, or kanamycin-susceptible bacteria. Moreover,S. aureusextracellular vesicles did not contain theblaZgene. In addition, the heat-treatedS. aureusextracellular vesicles did not mediate the survival of ampicillin-susceptible bacteria. The β-lactamase activities ofS. aureussoluble and extracellular vesicle-associated BlaZ were similar, but only the extracellular vesicle-associated BlaZ was resistant to protease digestion, which suggests that the enzymatic activity of BlaZ in extracellular vesicles is largely protected by the vesicle structure. Our observations provide evidence of the important role ofS. aureusextracellular vesicles in antibiotic resistance, which allows the polymicrobial community to continue to evolve and prosper against antibiotics.

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Ruthenium based antimicrobial theranostics – using nanoscopy to identify therapeutic targets and resistance mechanisms in Staphylococcus aureus

Chemical Science ◽

10.1039/c9sc04710g ◽

2020 ◽

Vol 11 (1) ◽

pp. 70-79 ◽

Cited By ~ 7

Author(s):

Kirsty L. Smitten ◽

Simon D. Fairbanks ◽

Craig C. Robertson ◽

Jorge Bernardino de la Serna ◽

Simon J. Foster ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Antimicrobial Activity ◽

Specific Resistance ◽

Therapeutic Targets ◽

Resistance Mechanisms ◽

Wild Type ◽

Gram Positive ◽

Gram Positive Bacteria

A detailed study on the uptake and antimicrobial activity of a RuII theranostic complex with wild-type S. aureus, MRSA, and other mutants has identified the specific resistance mechanisms that Gram-positive bacteria display against this lead.

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Penile implant infection resulting in Staphylococcus aureus bacteraemia and infective endocarditis

Access Microbiology ◽

10.1099/acmi.0.000295 ◽

2021 ◽

Vol 3 (12) ◽

Author(s):

Joseph P. Creel ◽

David Triplett ◽

Mannu Nayyar ◽

Nathan A. Summers

Keyword(s):

Staphylococcus Aureus ◽

Infective Endocarditis ◽

Type Species ◽

Clinical Suspicion ◽

Gram Positive ◽

Scrotal Pain ◽

Content Type ◽

Gram Positive Bacteria ◽

Link Type ◽

Penile Implant

Introduction. Penile implant infections are a possible surgical complication that has historically been most commonly associated with Gram-positive bacteria. Staphylococcus aureus is a Gram-positive bacteria and is the most common cause of endocarditis. Case Presentation. A male patient in his 50s with a past medical history of hypertension, diabetes, end-stage renal disease (ESRD) on peritoneal dialysis (PD) and erectile dysfunction with a penile implant placed 6 years prior to the admission date presented with complaints of scrotal pain. The pump for his implant had eroded through his scrotum and was draining pus. Blood cultures returned positive for Gram-positive cocci in clusters in 4/4 bottles, which was eventually identified as methicillin-sensitive Staphylococcus aureus (MSSA). A transthoracic echocardiogram (TTE) was performed due to concern for infective endocarditis (IE) but did not show any valvular abnormalities. Due to high clinical suspicion, a transesophageal echocardiogram (TEE) was performed and revealed a vegetation on the native mitral valve. His penile implant was removed by urology and intraoperative cultures grew MSSA. Surgical valve replacement was not recommended, and the patient was sent home with IV antibiotics for 6 weeks. Discussion. Post-operative site infections are a quite uncommon point of entry for infective endocarditis, with penile implant infections being an even rarer cause. While a TTE is often used initially to attempt to diagnose infective endocarditis, it has lower sensitivity than a TEE. If clinical suspicion for infective endocarditis remains high after negative imaging with TTE, then TEE should be performed for better visualization of the heart valves.

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Linearized esculentin-2EM shows pH dependent antibacterial activity with an alkaline optimum

Molecular and Cellular Biochemistry ◽

10.1007/s11010-021-04181-7 ◽

2021 ◽

Author(s):

Erum Malik ◽

David A. Phoenix ◽

Timothy J. Snape ◽

Frederick Harris ◽

Jaipaul Singh ◽

...

Keyword(s):

Helical Structure ◽

Food Preservation ◽

Forming Process ◽

Gram Positive ◽

Gram Negative ◽

Bacterial Membranes ◽

Gram Positive Bacteria ◽

Alkaline Conditions ◽

Ph Dependent ◽

Gram Negative Organisms

AbstractHere the hypothesis that linearized esculentin 2EM (E2EM-lin) from Glandirana emeljanovi possesses pH dependent activity is investigated. The peptide showed weak activity against Gram-negative bacteria (MLCs ≥ 75.0 μM) but potent efficacy towards Gram-positive bacteria (MLCs ≤ 6.25 μM). E2EM-lin adopted an α-helical structure in the presence of bacterial membranes that increased as pH was increased from 6 to 8 (↑ 15.5–26.9%), whilst similar increases in pH enhanced the ability of the peptide to penetrate (↑ 2.3–5.1 mN m−1) and lyse (↑ 15.1–32.5%) these membranes. Theoretical analysis predicted that this membranolytic mechanism involved a tilted segment, that increased along the α-helical long axis of E2EM-lin (1–23) in the N → C direction, with − < µH > increasing overall from circa − 0.8 to − 0.3. In combination, these data showed that E2EM-lin killed bacteria via novel mechanisms that were enhanced by alkaline conditions and involved the formation of tilted and membranolytic, α-helical structure. The preference of E2EM-lin for Gram-positive bacteria over Gram-negative organisms was primarily driven by the superior ability of phosphatidylglycerol to induce α-helical structure in the peptide as compared to phosphatidylethanolamine. These data were used to generate a novel pore-forming model for the membranolytic activity of E2EM-lin, which would appear to be the first, major reported instance of pH dependent AMPs with alkaline optima using tilted structure to drive a pore-forming process. It is proposed that E2EM-lin has the potential for development to serve purposes ranging from therapeutic usage, such as chronic wound disinfection, to food preservation by killing food spoilage organisms.

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Influence of the cell wall on ciprofloxacin susceptibility in selected wild-type Gram-negative and Gram-positive bacteria

International Journal of Antimicrobial Agents ◽

10.1016/j.ijantimicag.2003.12.015 ◽

2004 ◽

Vol 23 (6) ◽

pp. 627-630 ◽

Cited By ~ 18

Author(s):

Mercedes Berlanga ◽

M.Teresa Montero ◽

Jordi Hernández-Borrell ◽

Miquel Viñas

Keyword(s):

Cell Wall ◽

Wild Type ◽

Gram Positive ◽

Gram Negative ◽

Gram Positive Bacteria

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CodY Is a Global Regulator of Virulence-Associated Properties for Clostridium perfringens Type D Strain CN3718

mBio ◽

10.1128/mbio.00770-13 ◽

2013 ◽

Vol 4 (5) ◽

Cited By ~ 27

Author(s):

Jihong Li ◽

Menglin Ma ◽

Mahfuzur R. Sarker ◽

Bruce A. McClane

Keyword(s):

Clostridium Perfringens ◽

Null Mutant ◽

Intestinal Infection ◽

Wild Type ◽

Global Regulator ◽

Content Type ◽

Gram Positive Bacteria ◽

Type D ◽

Epsilon Toxin ◽

Virulence Properties

ABSTRACT CodY is known to regulate various virulence properties in several Gram-positive bacteria but has not yet been studied in the important histotoxic and intestinal pathogen Clostridium perfringens. The present study prepared an isogenic codY-null mutant in C. perfringens type D strain CN3718 by insertional mutagenesis using the Targetron system. Western blot analysis indicated that, relative to wild-type CN3718 or a complementing strain, this isogenic codY mutant produces reduced levels of epsilon toxin (ETX). Using supernatants from cultures of the wild-type, codY-null mutant, and complementing strains, CodY regulation of ETX production was shown to have cytotoxic consequences for MDCK cells. The CodY regulatory effect on ETX production was specific, since the codY-null mutant still made wild-type levels of alpha-toxin and perfringolysin O. Sialidase activity measurements and sialidase Western blot analysis of supernatants from CN3718 and its isogenic derivatives showed that CodY represses overall exosialidase activity due to a reduced presence of NanH in culture supernatants. Inactivation of the codY gene significantly decreased the adherence of CN3718 vegetative cells or spores to host Caco-2 cells. Finally, the codY mutant showed increased spore formation under vegetative growth conditions, although germination of these spores was impaired. Overall, these results identify CodY as a global regulator of many C. perfringens virulence-associated properties. Furthermore, they establish that, via CodY, CN3718 coordinately regulates many virulence-associated properties likely needed for intestinal infection. IMPORTANCE Clostridium perfringens is a major human and livestock pathogen because it produces many potent toxins. C. perfringens type D strains cause intestinal infections by producing toxins, especially epsilon toxin (ETX). Previous studies identified CodY as a regulator of certain virulence properties in other Gram-positive bacteria. Our study now demonstrates that CodY is a global regulator of virulence-associated properties for type D strain CN3718. It promotes production of ETX, attachment of CN3718 vegetative cells or spores to host enterocyte-like Caco-2 cells, and spore germination; the last two effects may assist intestinal colonization. In contrast, CodY represses sporulation. These results provide the first evidence that CodY can function as a global regulator of C. perfringens virulence-associated properties and that this strain coordinately regulates its virulence-associated properties using CodY to increase ETX production, host cell attachment, and spore germination but to repress sporulation, as would be optimal during type D intestinal infection.

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