Interaction with teichoic acids contributes to highly effective antibacterial activity of graphene oxide on Gram-positive bacteria
Graphical Abstract
Introduction
Graphene, owing to its unique sp2-bonded 2D atomic monolayer carbons arranged in hexagonal network structure and excellent electronic, mechanical, and thermal properties, facilitates in addressing universal concerns in water security, energy production and biomedicine (Dziewięcka et al., 2020, Lu et al., 2017, Zhao et al., 2014). Graphene oxide (GO), as the most concerned derivative of hydrophilic graphene, has great potentials in environmental applications such as photocatalysis, contaminant removal, membrane-based separation and electrochemical sensing (Huang et al., 2019a, Malina et al., 2020, Mu et al., 2019, Tang et al., 2020, Ye et al., 2020). Meanwhile, its applications in water disinfection and pathogen control are subjects of current interests (Huang et al., 2020, Panda et al., 2018, Zhao et al., 2018). Prior reports have demonstrated that the oxygenated and hydrophilic GO usually had a broad antibacterial spectrum against various kinds of bacteria (e.g. Staphylococcus aureus (S. aureus) and Pseudomonas aeruginosa (P. aeruginosa)), regardless of its diverse sources and characteristics (Anand et al., 2019, Chen et al., 2014, He et al., 2015, Keshvardoostchokami et al., 2020). Most of researches showed that the antibacterial activities of GO against Gram-positive (Gram+) bacteria (e.g. S. aureus and Streptococcus mutans) were higher than that against Gram-negative (Gram−) bacteria (e.g. P. aeruginosa and Escherichia coli (E. coli)) in in vitro culture experiments (Anand et al., 2019, He et al., 2015, Pulingam et al., 2019). Akhavan and Ghaderi (2010) also showed that GO nanosheets exhibited stronger antibacterial activity against S. aureus than E. coli, with an inactivation efficiency of 74% and 59%, respectively, after 1 h exposure at 37 °C.
A series of mechanisms have been proposed for the antibacterial activity of GO. (1) The physical interaction between GO and bacterial cells, including “wrapping” by GO, cutting into bacterial membrane and photothermal ablation by absorbing infrared light (Dallavalle et al., 2018, He et al., 2015, Perreault et al., 2015); (2) The inductions of reactive oxygen species (ROS) or carbon radical (·C), which further caused lipid peroxidation (Li et al., 2016, Panda et al., 2018, Yao et al., 2020); (3) Interactions between GO and biological macromolecules such as proteins, lipids or DNA (Liu et al., 2011, Qi et al., 2019). The antibacterial ability of GO against different bacteria was divergent, following the order S. aureus > Enterococcus faecalis > E. coli > P. aeruginosa (Pulingam et al., 2019). However, the known mechanisms could not explain why the antibacterial ability of GO against Gram+ bacteria is superior to Gram− ones.
It is well-known that Gram+ and Gram− bacteria are distinct in the cell envelope architectures (Akhavan and Ghaderi, 2010, Pulingam et al., 2019). The cell wall of Gram+ bacteria is mainly composed of peptidoglycan, teichoic acids and some proteins. By contrast, the cell wall of Gram− bacteria contains peptidoglycan, lipopolysaccharide and some protein but not teichoic acid (Caudill et al., 2020). The formation of pores in the bacterial cell wall, initiating osmotic imbalance and cell death, was reported to govern the antibacterial activity of graphene (Pham et al., 2015). A previous study illuminated that the interactions between different bacterial components and antibacterial substance contributed to divergent antibacterial activities, which means that the interaction of antibacterial peptides with teichoic acid rather than peptidoglycan could reduce the antibacterial activity (Malanovic and Lohner, 2016). Pulingam et al. (2019) elucidated that the different morphology changes of bacterial cells after exposure of GO contributed to divergent antibacterial activity of GO, which cell entrapment and membrane disruption were observed for Gram+ and Gram− bacteria upon GO treatments, respectively. A proposed explanation for the greater antibacterial activity of GO towards Gram+ than Gram− bacteria was the differences in components of their cell walls, that is the Gram+ bacteria lacking the outer membrane were demonstrated to be more vulnerable to the cell membrane damage than Gram− bacteria with an outer membrane (Akhavan and Ghaderi, 2010, Vi et al., 2020). However, the systematic molecular-level understanding of GO interactions with different components of cell walls is still limited, which would mislead its ideal applications in environment and biomedicine. In particular, teichoic acids play critical roles in the regulation of cell morphology and division, protecting Gram+ bacteria from autolysis and sustaining cation homeostasis for the cell (Brown et al., 2013, Caudill et al., 2020, Schlag et al., 2010). The quantitative comparisons of interaction affinities between GO and diverse cell walls together with membrane components, especially the unique components of teichoic acid existing in Gram+ bacteria, at molecular level may provide an insight in mechanism elucidation.
In this study, the antibacterial activities of GO against two types of Gram+ bacteria, S. aureus, Staphylococcus epidermidis (S. epidermidis), and two types of Gram− bacteria, P. aeruginosa and E. coli were investigated. Interaction mechanisms between GO and bacterial cell walls were further characterized using scanning electron microscopy (SEM), transmission electron micrographs (TEM), atomic force microscopy (AFM) and Fourier transform infrared spectroscopy (FTIR) spectra analyses. To better elucidate the underlying antibacterial mechanisms, for the first time, the adsorption affinities of teichoic acid and peptidoglycan to GO were compared in vitro, and the relative expression of autolysin of S. aureus (AtlA) after exposure to GO was explored. The superior antibacterial ability of GO against Gram+ versus Gram− bacteria is hypothesized to be attributed to its higher adsorption affinity for teichoic acid, a distinctive component of Gram+ bacterial cell wall which regulates the activity of autolysin. Our findings would explain the enhanced antibacterial activity of GO against Gram+ bacteria and provide guidance for practical water disinfection and pathogen control applications of graphene-based materials.
Section snippets
Graphene oxide and bacterial strains
GO (~99.8%) was purchased from XFNANO Materials Tech Inc. (Nanjing, China, product no. XF224). TEM (Hitachi H-7650, Japan), AFM (Alpha300R-2018, Germany), FTIR (Nicolet AVA TAR370, USA) and Raman spectroscopy (LabRAM HR Evolution, France) were employed to characterize the morphology and property of GO.
S. aureus (ATCC29213) and S. epidermidis (KB-3) were selected as model Gram+ bacteria; P. aeruginosa (PAO1) and E. coli (DH5α) were chosen as representative Gram− bacteria. S. aureus (ATCC29213),
Characterization of graphene oxide
The TEM, AFM, FTIR and Raman characterizations of GO are exhibited in Fig. S1. The TEM image of GO presented a typical two-dimensional flake with obvious wrinkles (Fig. S1a). AFM image (Fig. S1b) showed that the lateral dimensions of GO ranged from nanometers to micrometers and the thickness of GO was ~ 2.0 nm, indicating multi-layer distribution of GO nanosheets (Yao et al., 2020). Detailed functional groups of GO were explored via the FTIR spectrum. As shown in Fig. S1c, the intense peak at
Conclusion
GO exhibited a higher antibacterial activity on Gram+ than Gram− bacteria, due to its higher adsorption efficiency with teichoic acid, a unique component of Gram+ bacteria cell walls, than peptidoglycan, in which π−π interaction is the main driving force. GO adsorbed teichoic acid preferentially, which promoted the wrap of GO on Gram+ bacterial cell surfaces. Whereas, the adhesion did not occur with Gram− bacteria. The role of teichoic acid in antibacterial mechanisms to Gram+ bacteria was
CRediT authorship contribution statement
Meizhen Wang: Funding acquisition, Resources, Visualization, Writing - review & editing. Zhangqiang Li: Methodology, Formal analysis, Investigation, Software. Yunyun Zhang: Investigation, Formal analysis, Writing - original draft. Yue Li: Investigation. Na Li: Software. Dan Huang: Visualization, Supervision, Writing - review & editing. Baile Xu: Writing - review & editing.
Declaration of Competing Interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
This work was funded by the National Science Foundation of China (Grant no. 22076167, 21906145 and 21876155) and National Key R&D Program of China (Grant no. 2018YFE0110500). B. Xu acknowledges the scholarship from China Scholarship Council and Deutscher Akademischer Austauschdienst.
References (65)
- et al.
Development of a novel three-dimensional magnetic polymer aerogel as an efficient adsorbent for malachite green removal
J. Hazard. Mater.
(2020) - et al.
Wall teichoic acids govern cationic gold nanoparticle interaction with Gram-positive bacterial cell walls
Chem. Sci.
(2020) - et al.
Effects of foliar application of graphene oxide on cadmium uptake by lettuce
J. Hazard. Mater.
(2020) - et al.
Novel insight into adsorption and co-adsorption of heavy metal ions and an organic pollutant by magnetic graphene nanomaterials in water
Chem. Eng. J.
(2019) - et al.
Adsorption and desorption of phenanthrene by magnetic graphene nanomaterials from water: roles of pH, heavy metal ions and natural organic matter
Chem. Eng. J.
(2019) - et al.
Investigations of conformational structure and enzymatic activity of trypsin after its binding interaction with graphene oxide
J. Hazard. Mater.
(2020) - et al.
FT-IR study of plant cell wall model compounds: pectic polysaccharides and hemicelluloses
Carbohydr. Polym.
(2000) - et al.
Adsorption and desorption of phthalic acid esters on graphene oxide and reduced graphene oxide as affected by humic acid
Environ. Pollut.
(2018) - et al.
Development of a novel fluorescent substrate for Autolysin E, a bacterial type II amidase
Biochem. Biophys. Res. Commun.
(2009) - et al.
Gram-positive bacterial cell envelopes: the impact on the activity of antimicrobial peptides
Biochim. Biophys. Acta Biomembr.
(2016)
The environmental fate of graphene oxide in aquatic environment—complete mitigation of its acute toxicity to planktonic and benthic crustaceans by algae
J. Hazard. Mater.
Graphene oxide quantum dots stimulate indigenous bacteria to remove oil contamination
J. Hazard. Mater.
Graphene oxide exhibits differential mechanistic action towards Gram-positive and Gram-negative bacteria
Colloids Surf. B Biointerfaces
Atomic force microscopic study on morphological alterations induced by photodynamic action of Toluidine Blue O in Staphylococcus aureus and Escherichia coli
J. Photochem. Photobiol. B Biol.
pH-Dependent adsorption of aromatic compounds on graphene oxide: an experimental, molecular dynamics simulation and density functional theory investigation
J. Hazard. Mater.
Graphene oxide and graphene oxide functionalized with silver nanoparticles as adsorbents of phosphates in waters. A comparative study
Sci. Total Environ.
Adsorption and coadsorption of organic pollutants and a heavy metal by graphene oxide and reduced graphene materials
Chem. Eng. J.
Graphene oxide enhanced ozonation of 5-chloro-2-methyl-4-isothiazolin-3-one: kinetics, degradation pathway, and toxicity
J. Hazard. Mater.
Naturally occurring layered-mineral magnesium as a bactericidal against Escherichia coli
Appl. Clay Sci.
Antibacterial clay against Gram-negative antibiotic resistant bacteria
J. Hazard. Mater.
Toxicity of graphene and graphene oxide nanowalls against bacteria
ACS Nano
Graphene oxide and carbon dots as broad-spectrum antimicrobial agents-a minireview
Nanoscale Horiz.
Proton-binding capacity of Staphylococcus aureus wall teichoic acid and its role in controlling autolysin activity
PLoS One
Wall teichoic acids of Gram-positive bacteria
Annu. Rev. Microbiol.
Antimicrobial agents targeting bacterial cell walls and cell membranes
OIE Rev. Sci. Tech.
Synthetic lethal compound combinations reveal a fundamental connection between wall teichoic acid and peptidoglycan biosyntheses in Staphylococcus aureus
ACS Chem. Biol.
A new function of graphene oxide emerges: inactivating phytopathogenic bacterium Xanthomonas oryzae pv. Oryzae
J. Nanopart. Res.
Graphene oxide exhibits broad-spectrum antimicrobial activity against bacterial phytopathogens and fungal conidia by intertwining and membrane perturbation
Nanoscale
Light-enhanced antibacterial activity of graphene oxide, mainly via accelerated electron transfer
Environ. Sci. Technol.
Functionalization pattern of graphene oxide sheets controls entry or produces lipid turmoil in phospholipid membranes
ACS Appl. Mater. Interfaces
Graphene oxide as a new anthropogenic stress factor - multigenerational study at the molecular, cellular, individual and population level of Acheta domesticus
J. Hazard. Mater.
A Staphylococcus aureus ypfP mutant with strongly reduced lipoteichoic acid (LTA) content: LTA governs bacterial surface properties and autolysin activity
Mol. Microbiol.
Cited by (33)
Chirality-influenced antibacterial behavior of gold nanoclusters
2024, Colloids and Surfaces A: Physicochemical and Engineering AspectsAntibacterial mechanism of lignin and lignin-based antimicrobial materials in different fields
2023, International Journal of Biological MacromoleculesControlling biofilm and virulence properties of Gram-positive bacteria by targeting wall teichoic acid and lipoteichoic acid
2023, International Journal of Antimicrobial AgentsTime-dependent plastic behavior of bacteria leading to rupture
2023, Journal of the Mechanical Behavior of Biomedical Materials
- 1
These authors contributed equally to this work.