Antibiotic susceptibility of Staphylococcus aureus with different degrees of biofilm formation

AbstractStaphylococcus aureus is one of the most common pathogens in biofilm-associated chronic infections. S. aureus living within biofilms evades the host immune response and is more resistant to antibiotics than planktonic bacteria. In this study, we generated S. aureus with low and high levels of biofilm formation using the rbf (regulator of biofilm formation) gene and performed a BioTimer assay to determine the minimum inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) of various types of antibiotics. We showed that biofilm formation by S. aureus had a greater effect on MBC than MIC, probably due to the different growth modes between planktonic and biofilm bacteria. Importantly, we found that the MBC for biofilm S. aureus was much higher than that for planktonic cells, but there was little difference in MBC between low and high levels of biofilm formation. These results suggest that once the biofilm is formed, the bactericidal activity of antibiotics is significantly reduced, regardless of the degree of S. aureus biofilm formation. We propose that S. aureus strains with varying degrees of biofilm formation may be useful for evaluating the anti-biofilm activity of antimicrobial agents and understanding antibiotic resistance mechanisms by biofilm development.

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Antimicrobial and Antibiofilm Peptides

Biomolecules ◽

10.3390/biom10040652 ◽

2020 ◽

Vol 10 (4) ◽

pp. 652 ◽

Cited By ~ 12

Author(s):

Angela Di Somma ◽

Antonio Moretta ◽

Carolina Canè ◽

Arianna Cirillo ◽

Angela Duilio

Keyword(s):

Antimicrobial Peptides ◽

Biofilm Formation ◽

Bacterial Resistance ◽

Antimicrobial Agents ◽

Multidrug Resistant ◽

Resistant Bacteria ◽

Antibiofilm Activity ◽

Chronic Infections ◽

Planktonic Bacteria ◽

Hostile Environments

The increasing onset of multidrug-resistant bacteria has propelled microbiology research towards antimicrobial peptides as new possible antibiotics from natural sources. Antimicrobial peptides are short peptides endowed with a broad range of activity against both Gram-positive and Gram-negative bacteria and are less prone to trigger resistance. Besides their activity against planktonic bacteria, many antimicrobial peptides also show antibiofilm activity. Biofilms are ubiquitous in nature, having the ability to adhere to virtually any surface, either biotic or abiotic, including medical devices, causing chronic infections that are difficult to eradicate. The biofilm matrix protects bacteria from hostile environments, thus contributing to the bacterial resistance to antimicrobial agents. Biofilms are very difficult to treat, with options restricted to the use of large doses of antibiotics or the removal of the infected device. Antimicrobial peptides could represent good candidates to develop new antibiofilm drugs as they can act at different stages of biofilm formation, on disparate molecular targets and with various mechanisms of action. These include inhibition of biofilm formation and adhesion, downregulation of quorum sensing factors, and disruption of the pre-formed biofilm. This review focuses on the proprieties of antimicrobial and antibiofilm peptides, with a particular emphasis on their mechanism of action, reporting several examples of peptides that over time have been shown to have activity against biofilm.

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Protein A-Mediated Multicellular Behavior in Staphylococcus aureus

Journal of Bacteriology ◽

10.1128/jb.01222-08 ◽

2008 ◽

Vol 191 (3) ◽

pp. 832-843 ◽

Cited By ~ 170

Author(s):

Nekane Merino ◽

Alejandro Toledo-Arana ◽

Marta Vergara-Irigaray ◽

Jaione Valle ◽

Cristina Solano ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Biofilm Formation ◽

Protein A ◽

Surface Proteins ◽

Growth Media ◽

Dependent Manner ◽

Chronic Infections ◽

The Matrix ◽

Implanted Medical Devices ◽

Biofilm Development

ABSTRACT The capacity of Staphylococcus aureus to form biofilms on host tissues and implanted medical devices is one of the major virulence traits underlying persistent and chronic infections. The matrix in which S. aureus cells are encased in a biofilm often consists of the polysaccharide intercellular adhesin (PIA) or poly-N-acetyl glucosamine (PNAG). However, surface proteins capable of promoting biofilm development in the absence of PIA/PNAG exopolysaccharide have been described. Here, we used two-dimensional nano-liquid chromatography and mass spectrometry to investigate the composition of a proteinaceous biofilm matrix and identified protein A (spa) as an essential component of the biofilm; protein A induced bacterial aggregation in liquid medium and biofilm formation under standing and flow conditions. Exogenous addition of synthetic protein A or supernatants containing secreted protein A to growth media induced biofilm development, indicating that protein A can promote biofilm development without being covalently anchored to the cell wall. Protein A-mediated biofilm formation was completely inhibited in a dose-dependent manner by addition of serum, purified immunoglobulin G, or anti-protein A-specific antibodies. A murine model of subcutaneous catheter infection unveiled a significant role for protein A in the development of biofilm-associated infections, as the amount of protein A-deficient bacteria recovered from the catheter was significantly lower than that of wild-type bacteria when both strains were used to coinfect the implanted medical device. Our results suggest a novel role for protein A complementary to its known capacity to interact with multiple immunologically important eukaryotic receptors.

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Use of Confocal Microscopy To Analyze the Rate of Vancomycin Penetration through Staphylococcus aureus Biofilms

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.49.6.2467-2473.2005 ◽

2005 ◽

Vol 49 (6) ◽

pp. 2467-2473 ◽

Cited By ~ 177

Author(s):

Kimberly K. Jefferson ◽

Donald A. Goldmann ◽

Gerald B. Pier

Keyword(s):

Staphylococcus Aureus ◽

Antimicrobial Agents ◽

Confocal Scanning Laser Microscopy ◽

Planktonic Bacteria ◽

Transcriptional Changes ◽

Confocal Scanning ◽

Biofilm Bacteria ◽

Confocal Scanning Laser ◽

Reduced Rate

ABSTRACT When bacteria assume the biofilm mode of growth, they can tolerate levels of antimicrobial agents 10 to 1,000 times higher than the MICs of genetically equivalent planktonic bacteria. The properties of biofilms that give rise to antibiotic resistance are only partially understood. Inhibition of antibiotic penetration into the biofilm may play a role, but this has not been proven directly. In this report, penetration of the glycopeptide antibiotic vancomycin into viable Staphylococcus aureus biofilms was analyzed by confocal scanning laser microscopy using a fluorescently labeled derivative of the drug. We found that while vancomycin bound to free-floating bacteria in water within 5 min, it took more than 1 h to bind to cells within the deepest layers of a biofilm. These results indicate that the antibiotic is transported through the depth of the biofilm but that the rate is significantly reduced with respect to its transport through flowing water. This suggests that, whereas planktonic bacteria were rapidly exposed to a full bolus of vancomycin, the bacteria in the deeper layers of the biofilm were exposed to a gradually increasing dose of the drug due to its reduced rate of penetration. This gradual exposure may allow the biofilm bacteria to undergo stress-induced metabolic or transcriptional changes that increase resistance to the antibiotic. We also investigated the role of poly-N-acetylglucosamine, an important component of the S. aureus biofilm matrix, and found that its production was not involved in the observed decrease in the rate of vancomycin penetration.

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Staphylococcus aureus Develops an Alternative, ica-Independent Biofilm in the Absence of the arlRS Two-Component System

Journal of Bacteriology ◽

10.1128/jb.187.15.5318-5329.2005 ◽

2005 ◽

Vol 187 (15) ◽

pp. 5318-5329 ◽

Cited By ~ 117

Author(s):

Alejandro Toledo-Arana ◽

Nekane Merino ◽

Marta Vergara-Irigaray ◽

Michel Débarbouillé ◽

José R. Penadés ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Biofilm Formation ◽

Defined Medium ◽

Chronic Infections ◽

Negative Regulators ◽

Global Regulators ◽

Two Component ◽

Two Component Systems ◽

Important Virulence Factor ◽

Biofilm Development

ABSTRACT The biofilm formation capacity of Staphylococcus aureus clinical isolates is considered an important virulence factor for the establishment of chronic infections. Environmental conditions affect the biofilm formation capacity of S. aureus, indicating the existence of positive and negative regulators of the process. The majority of the screening procedures for identifying genes involved in biofilm development have been focused on genes whose presence is essential for the process. In this report, we have used random transposon mutagenesis and systematic disruption of all S. aureus two-component systems to identify negative regulators of S. aureus biofilm development in a chemically defined medium (Hussain-Hastings-White modified medium [HHWm]). The results of both approaches coincided in that they identified arlRS as a repressor of biofilm development under both steady-state and flow conditions. The arlRS mutant exhibited an increased initial attachment as well as increased accumulation of poly-N-acetylglucosamine (PNAG). However, the biofilm formation of the arlRS mutant was not affected when the icaADBC operon was deleted, indicating that PNAG is not an essential compound of the biofilm matrix produced in HHWm. Disruption of the major autolysin gene, atl, did not produce any effect on the biofilm phenotype of an arlRS mutant. Epistatic experiments with global regulators involved in staphylococcal-biofilm formation indicated that sarA deletion abolished, whereas agr deletion reinforced, the biofilm development promoted by the arlRS mutation.

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The antimicrobial peptide TAT-RasGAP317-326 inhibits the formation and the expansion of bacterial biofilms in vitro

10.1101/2020.09.29.318378 ◽

2020 ◽

Author(s):

Tytti Heinonen ◽

Simone Hargraves ◽

Maria Georgieva ◽

Christian Widmann ◽

Nicolas Jacquier

Keyword(s):

Staphylococcus Aureus ◽

Biofilm Formation ◽

Pathogenic Bacteria ◽

Inhibitory Concentration ◽

Antimicrobial Treatment ◽

Bacterial Biofilms ◽

Chronic Infections ◽

Planktonic Bacteria ◽

Further Development

AbstractBiofilms are structured aggregates of bacteria embedded in a self-produced matrix. Pathogenic bacteria can form biofilms on surfaces and in tissues leading to nosocomial and chronic infections. While antibiotics are largely inefficient in limiting biofilm formation and expansion, antimicrobial peptides (AMPs) are emerging as alternative anti-biofilm treatments. In this study, we explore the effect of the newly described AMP TAT-RasGAP317-326 on Acinetobacter baumannii, Pseudomonas aeruginosa and Staphylococcus aureus biofilms. We observe that TAT-RasGAP317-326 inhibits the formation of biofilms at concentrations equivalent or two times superior to the minimal inhibitory concentration (MIC) of the corresponding planktonic bacteria. Moreover, TAT-RasGAP317-326 limits the expansion of A. baumannii and P. aeruginosa established biofilms at concentrations 2-4 times superior to the MIC. These results further confirm the potential of AMPs against biofilms, expand the antimicrobial potential of TAT-RasGAP317-326 and support further development of this peptide as an alternative antimicrobial treatment.

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Antibiofilm Activity of Phorbaketals from the Marine Sponge Phorbas sp. against Staphylococcus aureus

Marine Drugs ◽

10.3390/md19060301 ◽

2021 ◽

Vol 19 (6) ◽

pp. 301

Author(s):

Yong-Guy Kim ◽

Jin-Hyung Lee ◽

Sangbum Lee ◽

Young-Kyung Lee ◽

Buyng Su Hwang ◽

...

Keyword(s):

Reactive Oxygen Species ◽

Staphylococcus Aureus ◽

Marine Sponge ◽

Biofilm Formation ◽

Antimicrobial Agents ◽

Critical Role ◽

Antibiofilm Activity ◽

Oxygen Species ◽

Chronic Infections ◽

Hemolysin Gene

Biofilm formation by Staphylococcus aureus plays a critical role in the persistence of chronic infections due to its tolerance against antimicrobial agents. Here, we investigated the antibiofilm efficacy of six phorbaketals: phorbaketal A (1), phorbaketal A acetate (2), phorbaketal B (3), phorbaketal B acetate (4), phorbaketal C (5), and phorbaketal C acetate (6), isolated from the Korean marine sponge Phorbas sp. Of these six compounds, 3 and 5 were found to be effective inhibitors of biofilm formation by two S. aureus strains, which included a methicillin-resistant S. aureus. In addition, 3 also inhibited the production of staphyloxanthin, which protects microbes from reactive oxygen species generated by neutrophils and macrophages. Transcriptional analyses showed that 3 and 5 inhibited the expression of the biofilm-related hemolysin gene hla and the nuclease gene nuc1.

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Biological Evaluation of Selected 1,2,3-triazole Derivatives as Antibacterial and Antibiofilm Agents

Current Topics in Medicinal Chemistry ◽

10.2174/1568026620666200710104737 ◽

2020 ◽

Vol 20 (24) ◽

pp. 2186-2191

Author(s):

Lialyz Soares Pereira André ◽

Renata Freire Alves Pereira ◽

Felipe Ramos Pinheiro ◽

Aislan Cristina Rheder Fagundes Pascoal ◽

Vitor Francisco Ferreira ◽

...

Keyword(s):

Electron Microscopy ◽

Scanning Electron Microscopy ◽

Staphylococcus Aureus ◽

Biofilm Formation ◽

Antimicrobial Agents ◽

Public Health Problem ◽

Biological Evaluation ◽

Major Public Health Problem ◽

Community Environments ◽

Scanning Electron

Background: Resistance to antimicrobial agents is a major public health problem, being Staphylococcus aureus prevalent in infections in hospital and community environments and, admittedly, related to biofilm formation in biotic and abiotic surfaces. Biofilms form a complex and structured community of microorganisms surrounded by an extracellular matrix adhering to each other and to a surface that gives them even more protection from and resistance against the action of antimicrobial agents, as well as against host defenses. Methods: Aiming to control and solve these problems, our study sought to evaluate the action of 1,2,3- triazoles against a Staphylococcus aureus isolate in planktonic and in the biofilm form, evaluating the activity of this triazole through Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) tests. We have also performed cytotoxic evaluation and Scanning Electron Microscopy (SEM) of the biofilms under the treatment of the compound. The 1,2,3-triazole DAN 49 showed bacteriostatic and bactericidal activity (MIC and MBC 128 μg/mL). In addition, its presence interfered with the biofilm formation stage (1/2 MIC, p <0.000001) and demonstrated an effect on young preformed biofilm (2 MICs, p <0.05). Results: Scanning Electron Microscopy images showed a reduction in the cell population and the appearance of deformations on the surface of some bacteria in the biofilm under treatment with the compound. Conclusion: Therefore, it was possible to conclude the promising anti-biofilm potential of 1,2,3-triazole, demonstrating the importance of the synthesis of new compounds with biological activity.

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

Journal of Bacteriology ◽

10.1128/jb.00159-20 ◽

2020 ◽

Vol 202 (18) ◽

Cited By ~ 1

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.

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Putrescine and its metabolic precursor arginine promote biofilm and c-di-GMP synthesis in Pseudomonas aeruginosa

Journal of Bacteriology ◽

10.1128/jb.00297-21 ◽

2021 ◽

Author(s):

Zhexian Liu ◽

Sarzana S. Hossain ◽

Zayda Morales Moreira ◽

Cara H. Haney

Keyword(s):

Pseudomonas Aeruginosa ◽

Immune Responses ◽

Biofilm Formation ◽

Antimicrobial Agents ◽

Bacterial Pathogen ◽

Guanosine Monophosphate ◽

Genetic Approach ◽

Chronic Infections ◽

Host Immune Responses ◽

Primary Mechanism

Pseudomonas aeruginosa , an opportunistic bacterial pathogen can synthesize and catabolize a number of small cationic molecules known as polyamines. In several clades of bacteria polyamines regulate biofilm formation, a lifestyle-switching process that confers resistance to environmental stress. The polyamine putrescine and its biosynthetic precursors, L-arginine and agmatine, promote biofilm formation in Pseudomonas spp. However, it remains unclear whether the effect is a direct effect of polyamines or through a metabolic derivative. Here we used a genetic approach to demonstrate that putrescine accumulation, either through disruption of the spermidine biosynthesis pathway or the catabolic putrescine aminotransferase pathway, promoted biofilm formation in P. aeruginosa . Consistent with this observation, exogenous putrescine robustly induced biofilm formation in P. aeruginosa that was dependent on putrescine uptake and biosynthesis pathways. Additionally, we show that L-arginine, the biosynthetic precursor of putrescine, also promoted biofilm formation, but via a mechanism independent of putrescine or agmatine conversion. We found that both putrescine and L-arginine induced a significant increase in the intracellular level of bis-(3′-5′)-cyclic dimeric guanosine monophosphate (c-di-GMP) (c-di-GMP), a bacterial second messenger widely found in Proteobacteria that upregulates biofilm formation. Collectively these data show that putrescine and its metabolic precursor arginine promote biofilm and c-di-GMP synthesis in P. aeruginosa . Importance: Biofilm formation allows bacteria to physically attach to a surface, confers tolerance to antimicrobial agents, and promotes resistance to host immune responses. As a result, regulation of biofilm is often crucial for bacterial pathogens to establish chronic infections. A primary mechanism of biofilm promotion in bacteria is the molecule c-di-GMP, which promotes biofilm formation. The level of c-di-GMP is tightly regulated by bacterial enzymes. In this study, we found that putrescine, a small molecule ubiquitously found in eukaryotic cells, robustly enhances P. aeruginosa biofilm and c-di-GMP. We propose that P. aeruginosa may sense putrescine as a host-associated signal that triggers a lifestyle switching that favors chronic infection.

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Antibiotic Susceptibility Study Of Metal-Gentamicin Complexes Against Staphylococcus Aureus Biofilms

IIUM Medical Journal Malaysia ◽

10.31436/imjm.v16i2.1050 ◽

2017 ◽

Vol 16 (2) ◽

Author(s):

Intan Azura Shahdan ◽

Fatimah Zahrah Mohd Sobri ◽

Mohammad Faiz Hizzuan Hanapi ◽

Hanani Ahmad Yusof ◽

Fiona N.-F. How

Keyword(s):

Staphylococcus Aureus ◽

Antibiotic Susceptibility ◽

Bacterial Species ◽

Dental Biofilm ◽

Planktonic Bacteria ◽

Metal Contents ◽

Susceptibility Study ◽

Biofilm Bacteria ◽

Dental Plaques ◽

Conventional Antibiotics

Introduction: Dental plaque is a structurally and functionally organized biofilm. Modern molecular biological techniques have identified about 1000 different bacterial species in the dental biofilm, twice as many as can be cultured. Inherent resistance of biofilm bacteria to conventional antibiotics is alarming. It induces antibiotic resistance to an order of three or more in magnitude greater than those displayed by planktonic bacteria. Staphylococcus aureus is the most dominant bacterial species isolated from the saliva and dental plaques. One of the reasons for its pathogenicity is its ability to form biofilms. In this study, the resistance of S. aureus biofilms against a eries of metal-antibiotics, an alternative to the conventional antibiotics, was investigated. Materials and Methods: A series of metal-antibiotic complexes derived from gentamicin was synthesized to give metal-gentamicin complexes. The metal contents of all the compounds were determined using Atomic Absorption Spectroscopy (AAS). Antibiotic susceptibility testing of the gentamicin-antibiotic complexes against several strains of S. aureus biofilms was conducted using broth microdilution assay. Results: The results showed that S. aureus is susceptible against Co(II) and Fe(II) gentamicin complexes; all were tested at 0.25 to 1 mmol concentrations. Conclusion(s): Co(II) and Fe(II)-gentamicin complexes demonstrated antimicrobial activity.

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