Laure Giraud

Audrey Tourrette

Emmanuel Flahaut
email: flabaur@cbimie.ups-rlse.fr

Carbon nanomaterials-based polymer-matrix nanocomposites for antimicrobial applications: A review

Carbon Nanomaterials present many exceptional properties, including antimicrobial ones. The transfer of the latter through thei r incorporation into materials has generated a growing interest with a signi ficant increase in number of publications since 2011. The use of polymeric matrices presents many advantages, because polymers are efficient nanoparticles holders and also exhibit intrinsic antimicrobial properties in some cases. The state of the art of fabrication methods and presumed antimicrobial mechanism is presented, focusing on antibacterial activity as the latter is, by far, the most discussed in the literature. Even if many examples of such nanocomposites are available, most of them correspond to carbon nanomaterials incorporated in the volume, white only the surface is of interest in terms of intrinsic (and thus expected long term) activity, and there is still a lack of information concerning the occurring antimicrobial mechanisms. We have identified the good stability of carbon nanomaterials in biological environment as an interesting option in terms of durability of the antimicrobial effect and will discuss this question in detail, as well as the fact that there are currently no clear conclusions concerning toxicity aspects due to the possible release of such nanoparticles in use conditions.

42.

Carbon nanomaterials antimicrobial mechanisms . . . .

Introduction

Microbial proliferation on materials has become a major issue in various fields. Whether used in transports, biomedicine or envi ronmental conditions, materials are confronted to microbe's pro liferation leading to global health and economic issues. Microbes include bacteria, fungi, algae, protozoa, and amoebas, and are generally described as sources or vectors of diseases. For example, materials placed in environmentally challenging conditions are indeed faced to the issue of biofilm formation. A biofilm is a structured agglomeration of microorganisms embedded in a self produced matrix that adheres to a surface. Once formed, microor ganisms in a biofilm are very difficult to remove compared to in dividual entities, as this structure behaves as a reservoir of microbes and offers them protection from antimicrobial agents [START_REF] O'toole | biofilm formation as microbial development[END_REF]2]. Fighting against biofilm formation is a great issue for mate rials immersed in aquatic media for instance, as their functionality and life expectancy become limited with the appearance of a bio film [3]. It is also a problem for implantable medical devices, the development of resistant bacterial biofilm leading to an increase in mortality by nosocomial infection and being as such an important sanitary concern [4,[START_REF] Veerachamy | Bacterial adherence and biofilm formation on medical implants: a review[END_REF].

Antimicrobial materials represent an alternative to limit or eradicate microbial proliferation, either because they can directly kill or damage microbes, or because they can prevent cell adhesion onto their surface, such as passive antimicrobial polymers based materials for example [6e8]. Antimicrobial compounds able to inhibit bacterial proliferation are important because numerous bacteria have become resistant to antibiotics [9e11]. Anti infective agents with other action mechanism have been developed, such as metal ions [START_REF] Vaidya | Antimicrobial efficacy and synergy of metal ions against Enterococcus faecium, Klebsiella pneumoniae and acinetobacter baumannii in planktonic and biofilm phenotypes[END_REF], quaternary ammonium salts [13e16], and nano particles [START_REF] Fernando | Antimicrobial Nanoparticles: applications and mechanisms of action[END_REF][START_REF] Wang | The antimicrobial activity of nanoparticles: present situation and prospects for the future[END_REF]. This is why the use of silver (Ag) nanoparticles is widespread as it is efficient and works differently from antibiotics [START_REF] Matai | Antibacterial activity and mechanism of Ag ZnO nanocomposite on S. aureus and GFP-expressing antibiotic resistant E. coli[END_REF][START_REF] Li | Antibacterial activity and mechanism of silver nanoparticles on Escherichia coli[END_REF]. Among all the investigated nanoparticles, carbon based nanomaterials (CNMs) represent a specific category. Albeit known and exploited in many fields due to their exceptional properties, they have recently gained further attention thanks to their anti microbial activity [START_REF] Xin | Antibacterial carbon-based nanomaterials[END_REF][START_REF] Al-Jumaili | Review on the antimicrobial properties of carbon nanostructures[END_REF]. Most studies to date have highlighted their action as free particles in suspension [23e25], but not included within a material. CNMs may also be combined with other antimicrobial agents such as Ag [START_REF] Pant | Characterization and antibacterial properties of aminophenol grafted and Ag NPs decorated graphene nanocomposites[END_REF][START_REF] He | Antibacterial effect and proteomic analysis of graphene-based silver nanoparticles on a pathogenic bacterium Pseudomonas aeruginosa[END_REF] or ZnO [START_REF] Zanni | Evaluation of the antibacterial power and biocompatibility of zinc oxide nanorods decorated graphene nanoplatelets: new perspectives for antibiodeteriorative approaches[END_REF][START_REF] Wang | Superior antibacterial activity of zinc oxide/graphene oxide composites originating from high zinc concentration localized around bacteria[END_REF] for enhanced efficacy.

However, the use of nanoparticles in general for such applica tions raises potential issues, including the durability of their action and their potential own toxicity. The durability issue is directly related to the number of nanoparticles present in the material, which could be time limited as nanoparticles may be progressively released or dissolved in the medium to be treated. The main advantage of carbon nanomaterials (CNMs) is that they are very chemically resistant and thus not soluble, and will as such be kept at the same amount in antibacterial materials with no expectable loss of activity with time. Regarding the release of the nanoparticles themselves (related to ageing of the material due to abrasion, weathering, etc.), their potential intrinsic toxicity towards both humans and the environment should also be considered [30e33]. Therefore, composite materials containing embedded nano particles are now investigated. As polymers proved to be both efficient nanoparticle holders and to demonstrate intrinsic anti microbial properties in some cases, we have decided to focus this review on polymer based nanocomposites [START_REF] Francolini | Antimicrobial Polymers for Anti-biofilm Medical Devices: State-Of-Art and Perspectives[END_REF].

In a first part, the different microorganisms which have been investigated so far in the literature in this context will briefly be presented, as well as the tests used to assess the antimicrobial ac tivity of materials, which are usually different from the ones used for particles in suspension. Focusing on polymer based materials, the different antimicrobial strategies which have been proposed will then be compared: after describing the specific case of poly mers exhibiting an intrinsic antimicrobial activity, the antimicro bial properties of carbon nanomaterials will be listed before finally reviewing polymer based antimicrobial nanocomposites contain ing CNMs. In each case, attention will be paid to fabrication methods, antimicrobial tests used (along with the tested microor ganisms) and presumed antimicrobial mechanism. Some examples of potential applications will also be discussed, and general con clusions will be proposed.

Microorganisms and antimicrobial tests

Microorganisms are micro living entities that are part of our daily live. Composing about half of the human body [START_REF] Sender | Revised estimates for the number of human and bacteria cells in the body[END_REF] they are mostly essential for our proper functioning, although some of them have pathogenic effects on human beings. Microbes (the science of which is called microbiology [36]) include several types of micro organisms exhibiting different forms, among which fungi, bacteria or protozoa may be cited. This term does not include viruses that are usually considered on the borderline of living organisms, or even as non living, infectious entities. Fungi are eukaryotic com ponents, usually filamentous with cell walls containing chitin, that include entities with a lot of variation in size and structure. Although associated to plants for a long time, they miss many of their main characteristics (absence of chloroplasts, chlorophyll and starch) [START_REF] Hawksworth | Microorganisms[END_REF]. Bacteria are unicellular microorganisms with a size ranging from 1 to 5 mm. A least composed of cell walls, plasmic membrane, cytoplasm and genetic material, they are capable of reproduction via cellular division (contrary to viruses). They are classified through their cell walls nature into Gram negative or Gram positive categories. Gram negative bacteria have a thin peptidoglycan layer of 6e15 nm and an outer lipid membrane whereas Gram positive bacteria have a thicker peptidoglycan layer of (20e80 nm) and miss the outer lipid membrane [START_REF] Silhavy | The bacterial cell envelope[END_REF]. Contrary to bacteria, a virus is not an autonomous entity that can reproduce independently. Smaller than bacteria (20e500 nm), it is a least composed of genetic material surrounded by a protein shell known as a capsid [START_REF] Hawksworth | Microorganisms[END_REF]. Thus, the effect of antimicrobial agents varies depending on the type of microorganism because of their different structures. There are different kinds of antimicrobial activity assays available in the literature [START_REF] Balouiri | Methods for in vitro evaluating antimicrobial activity_ A review[END_REF], and they can generally be used with most antimicrobial agents to test their activity against pathogens, whatever their form (polymers, composites, or suspensions of nanoparticles).

Literature mostly reports the use of assays such as inhibition zone, colony counting or optical density (OD) measurements to quantify the antimicrobial activity of materials. These global methods, giving information at the level of the whole population of microorganisms, may roughly be summarized in three steps: contact between microorganisms and the sample, retrieval of microorgan isms, and finally investigation of microorganisms' proliferation with comparison with a control condition [START_REF] Song | Electrospun Nanofibers with Antimicrobial Properties[END_REF].

The inhibition zone assay or disc diffusion test (Fig. 1a) is a qualitative diffusion method frequently used and consists in filling petri dishes with a microbial solution of known concentration mixed with a nutritive medium (Luria Bertani agar medium; Mueller Hinton Agar medium, etc.). The tested materials are then added onto the bacterial lawn and the plate is incubated. After wards, the radius of inhibition zone (mm) is measured and reflects the antimicrobial activity of the sample. An antibiotic disc may be used as a positive control [START_REF] Khandelwal | Antimicrobial activities of graphene polymer nanocomposites[END_REF][START_REF] Yang | Electrospun chitosan/poly (vinyl alcohol)/graphene oxide nanofibrous membrane with ciprofloxacin antibiotic drug for potential WoundDressing application[END_REF]. Further variations of this diffusion method exist, such as the agar plug diffusion method [START_REF] Elleuch | Bioactive secondary metabolites from a new terrestrial streptomyces sp. TN262[END_REF] or the agar well diffusion method, that allows to test the antimicrobial activity of plants [START_REF] Valgas | Screening methods to determine antibacterial activity of natural products[END_REF]. It is also possible to quantitatively measure the antimicrobial activity of bacteria or fungi via dilution methods. The same type of tests previously presented are applied to micro organisms that are confronted to various concentrations of anti microbial substance and the minimum inhibitory concentration (MIC) may be determined, as in the agar dilution or broth dilution methods [START_REF] Jorgensen | Antimicrobial susceptibility testing: a review of general principles and contemporary practices[END_REF].

The colony counting assay (Fig. 1b) is widespread. After the in cubation of the tested samples with a microbial solution of known concentration, microorganisms are retrieved and then incubated into a culture medium before processing to colony counting to measure their viability [START_REF] Thampi | Differential adhesive and bioactive properties of the polymeric surface coated with graphene oxide thin film[END_REF]. Microbial growth following exposure to an antimicrobial agent is mainly measured by assessing the optical density after the microorganisms have been resuspended in water [START_REF] Santos | Antimicrobial graphene polymer (PVK-GO) nanocomposite films[END_REF]. Such tests thus allow to deduce the antimicrobial properties of samples through the control of their impact on the proliferation of microbes upon contact. However, they don't give any indication about the mechanisms of action. Another way to assess the effect of antimicrobial agents is to evaluate their impact on the microbes at the individual scale. Other reported tests such as RNA/DNA measurements, instead of measuring optical density, aim to detect RNA (or DNA) via spectrophotometry (or chromatog raphy) to allow the estimation of impacted cells.

Fluorescence assays (Fig. 1ced) are described in numerous pa pers and allow to gain information on damaged cells. However, they usually cannot be applied directly to the surface of a material (unless it is transparent), because the cells need to be placed on a transparent glass slide. After incubation, bacteria adhering to the tested surface sample are retrieved and put into a culture medium. Stain is then added to determine the live/dead cell ratio. SYTO9 (green fluorescence emission at 503 nm) is among the most used stains. This is a green fluorescent nuclear and chromosome coun terstain that is permeant to both prokaryotic and eukaryotic cell membranes. SYTO 9 stain has a high affinity for DNA and exhibits enhanced fluorescence upon binding to DNA. It stains both live and dead Gram positive and Gram negative bacteria. Another example is 4 0 ,6' Di Amidino 2 Phenyl Indole (DAPI, blue fluorescence emission at 461 nm) which also stains mainly DNA, but passes only through intact cell membranes (live, or fixed) and can thus be used to assess cell viability. It can also bind to RNA, but in this case the fluorescence emission maximum is up shifted to ca. 500 nm. Finally, propidium iodide (PI, red fluorescence emission at 617 nm) easily penetrates the damaged, permeable membranes of non viable cells but is excluded by viable cells. It binds to double stranded DNA by intercalating between base pairs. Fluorescence imaging finally reveals stained cells, and as such the antimicrobial activity [START_REF] Santos | Antimicrobial graphene polymer (PVK-GO) nanocomposite films[END_REF][START_REF] Santos | Graphene nanocomposite for biomedical applications: fabrication, antimicrobial and cytotoxic investigations[END_REF]. Thereby, these methods confirm impacts on cell membrane integrity (with the possible release of cell components), but they do not indicate the specific process leading to this dete rioration. Other assays take advantage of fluorescence detection after addition of a reagent that allows the identification of certain specific components (Metabolic assay, Reactive oxygen species assay, assessment of membrane lipid peroxidation, etc.) and are thus very relevant to investigate mechanistic aspects [48e54].

Microscopy imaging may be used to observe cell adhesion using a simple optical microscope. Much more details may be obtained using electron microscopy, either in scanning (SEM) or trans mission mode (TEM) to detect morphological changes of cells after incubation with the tested sample, or to detect biofilm formation. Atomic Force microscopy (AFM) may also be used to measure sur face topography and indicate the formation of biofilm or not [START_REF] Santos | Graphene nanocomposite for biomedical applications: fabrication, antimicrobial and cytotoxic investigations[END_REF][START_REF] Mejías Carpio | Toxicity of a polymer graphene oxide composite against bacterial planktonic cells, biofilms, and mammalian cells[END_REF]. It may also be used to investigate the mechanical proper ties of cell membranes [START_REF] Li | Advances in atomic force microscopy for singlecell analysis[END_REF][START_REF] Camesano | Observation of changes in bacterial cell morphology using tapping mode atomic force microscopy[END_REF]. Of course, microscopy imaging by AFM, SEM or TEM are not high throughput and require long sample preparation steps so these techniques are not used routinely to assess the antimicrobial activity (they also miss the quantitative aspect), but only to investigate mechanistic issues.

Most publications on antimicrobial activity of carbon based nanomaterials focus particularly on the antibacterial activity. As may be seen in section 5.2, the colony counting, zone inhibition or the fluorescence tests are frequently used to assess the antibacterial activity of the tested materials, with in particular the main use of gram negative bacteria E. coli. Authors also regularly report the use of gram negative P. aeruginosa and gram positive S. aureus and B. subtilis in antibacterial tests [48,58e60]. These models of bacteria are frequently identified as nosocomial infections. They may develop resistance to antibiotics and as such are of important sanitary concern [START_REF] Weinstein | Nosocomial infection update[END_REF]. It is thus very relevant to use these types of bacteria to assess the antibacterial activity of materials. Other studies also present, to a lesser extent, the evaluation of antimi crobial activity of carbon based nanomaterials against certain types of fungi or viruses [START_REF] Hegab | Fine-tuning the surface of forward osmosis membranes via grafting graphene oxide: performance patterns and biofouling propensity[END_REF][START_REF] Hegab | The controversial antibacterial activity of graphene-based materials[END_REF][START_REF] Mohamed | Novel aminohydrazide cross-linked chitosan filled with multi-walled carbon nanotubes as antimicrobial agents[END_REF].

Antimicrobial polymers

Antimicrobial polymers are briefly described here because they may be used in association with CNMs to enhance their intrinsic activity and thus need to be described to understand the role played by each different part in a nanocomposite material. They fight against pathogens through active or passive pathways. Active polymers kill microbes upon contact via electrostatic or biocidal interactions. On the contrary, antimicrobial passive polymers fight against pathogens mainly by preventing their adhesion through repelling them from the surface. As microbes are mainly hydro phobic or negatively charged, a repulsive mechanism may be ob tained by controlling the hydrophilicity, charge properties and surface energy of the polymer [START_REF] Huang | Recent advances in antimicrobial polymers: a mini-review[END_REF]. For example, Poly(ethylene glycol) (PEG), is used to prevent protein and cell adhesion due to its hydrophilic surface [START_REF] Krishnan | Advances in polymers for antibiofouling surfaces[END_REF]. According to their structure, antimicrobial polymers may be sorted in 3 categories: polymeric biocides, biocidal polymers and biocide releasing polymers [START_REF] Huang | Recent advances in antimicrobial polymers: a mini-review[END_REF]. Polymeric biocides are polymers formed by a succession of bioactive units. In biocidal polymers, on the contrary, the activity is provided by the whole macromolecule. Most biocidal polymers actively damage cells membrane through electrostatic interactions. There are indeed lots of examples of biocidal polymers containing cationic groups such as quaternary ammonium [START_REF] Xue | Antimicrobial polymeric materials with quaternary ammonium and phosphonium salts[END_REF][START_REF] Kawabata | Antibacterial activity of soluble pyridinium-type polymers[END_REF], phosphonium [START_REF] Kanazawa | Polymeric phosphonium salts as a novel class of cationic biocides. IV. Synthesis and antibacterial activity of polymers with phosphonium salts in the main chain[END_REF][START_REF] Kanazawa | Novel polycationic biocides: synthesis and antibacterial activity of polymeric phosphonium salts[END_REF], tertiary sulfonium [START_REF] Kanazawa | Antibacterial activity of polymeric sulfonium salts[END_REF] or guanidine [START_REF] Locock | Guanylated polymethacrylates: a class of potent antimicrobial polymers with low hemolytic activity[END_REF]. In contrast, biocide releasing polymers usually have no intrinsic activity but only act as a container releasing antimicrobial substances such as for example gentamicin, triclosan or silver among the most commonly cited [START_REF] Siedenbiedel | Antimicrobial polymers in solution and on surfaces: overview and functional principles[END_REF][START_REF] Yu | Engineering biomaterials surfaces to modulate the host response[END_REF]. For further information on these polymers, the reader may refer to detailed reviews on this topic [START_REF] Huang | Recent advances in antimicrobial polymers: a mini-review[END_REF][START_REF] Liu | Recent development of antifouling polymers: structure, evaluation, and biomedical applications in nano/micro-structures[END_REF][START_REF] Timofeeva | Antimicrobial polymers: mechanism action, factors of activity, and applications[END_REF]. In this document, the focus is specifically on polymers including car bon nanoparticles as antimicrobial agent.

Carbon nanomaterials

Different kinds of carbon nanomaterials

CNMs (Fig. 2) exhibit a large variety of shapes from 0D (nano diamonds, fullerenes), to 1D (nanotubes) and 2D graphene and related materials (graphene, graphene oxide, reduced graphene oxide). One common property of all carbon nanomaterials is the possibility to functionalise them by grafting different surface functional groups with important consequences on their hydro phobicity/hydrophilicity, surface charge, electronic, optical and often also mechanical properties. The most common way of doing so is through simple oxidation, leading to the grafting of oxygen containing functional groups, and is often a consequence of puri fication treatments [START_REF] Bortolamiol | Double-walled carbon nanotubes: quantitative purification assessment, balance between purification and degradation and solution filling as an evidence of opening[END_REF]. Many much more sophisticated proced ures have already been extensively reviewed [START_REF] Georgakilas | Functionalization of graphene: covalent and non-covalent approaches, derivatives and applications[END_REF]. Nanodiamonds are 0D nanostructures composed of sp 3 hybridized carbon atoms at the core, often surrounded by sp 2 carbon atoms and functionals groups [START_REF] Shenderova | Ultrananocrystalline Diamond: Synthesis, Properties and Applications[END_REF]. They exhibit exceptional mechanical and optical properties, along with a specific surface area commonly between 200 and 600 m 2 /g [75e78]. Their properties depend upon the sp 3 / sp 2 ratio [START_REF] Xin | Antibacterial carbon-based nanomaterials[END_REF][START_REF] Mochalin | The properties and applications of nanodiamonds[END_REF]. Graphene is a 2D material forming a planar structure of one atomic layer. It is composed of sp 2 hybridized carbon atoms arranged in hexagons, and represents the basic structure found in other nanomaterials such as fullerene and carbon nanotubes (CNTs) [START_REF] Xin | Antibacterial carbon-based nanomaterials[END_REF]. The classification of graphene and related materials (GRM) may be defined through the number of layers, the average lateral size, and the carbon to oxygen (C/O) atomic ratio (Fig. 3) [START_REF] Wick | Classification framework for graphene-based materials[END_REF], this last parameter having the highest impact on their physico chemical properties. The superposition of 2e10 layers leads to Few Layer Graphene (FLG), whereas the presence of 10 sheets of graphene or more is described as graphite nanoplates. Graphene Oxide (GO) corresponds to a single layer of graphene bearing many kinds of oxygenated functions, and its reduction leads to reduced Graphene Oxide (rGO). While graphene has exceptional physico chemical properties and is highly hydro phobic, with a specific surface area theoretically reaching 2600 m 2 g À1 , good electronic mobility and mechanical properties [81e83], GO is on the opposite an insulating but highly hydrophilic material and it is more easily dispersed than its non functionalized counterpart [START_REF] Shearer | Accurate thickness measurement of graphene[END_REF].

Fullerenes are spherical 0D closed nanostructures containing a majority of sp 2 hybridized carbon atoms (C 60 [START_REF] Xin | Antibacterial carbon-based nanomaterials[END_REF][START_REF] Al-Jumaili | Review on the antimicrobial properties of carbon nanostructures[END_REF], C 70 , C 76 , C 78 , C 84 etc.). Fullerenes also present size and structure dependant properties (optical, mechanical, chemical, electrical) [START_REF] Kang | Single-walled carbon nanotubes exhibit strong antimicrobial activity[END_REF]. Finally, a Carbon Nanotube (CNT) can be described as a rolled graphene sheet generally closed by hemifullerenes. Single Wall Carbon Nanotubes (SWCNTs) have a small diameter typically of only a few nanometres or less; they can behave as semiconductors or metals depending on their electronic structure. Multi Wall Carbon Nano tubes (MWCNTs) composed of concentric SWCNTs may reach up to 100 nm outer diameter [START_REF] Xin | Antibacterial carbon-based nanomaterials[END_REF]. Similarly, intrinsic parameters such as their diameter, length, number of walls and the presence of func tional groups play a significant role on their properties [86].

For all these reasons, CNMs are used for numerous applications, from sensors, optical devices, superconductors, to drug delivery systems and biomedical applications in general, including their antimicrobial properties [START_REF] Shi | The antibacterial applications of graphene and its derivatives[END_REF][START_REF] Kang | Single-walled carbon nanotubes exhibit strong antimicrobial activity[END_REF][START_REF] Akhavan | Toxicity of graphene and graphene oxide nanowalls against bacteria[END_REF]. It is important to mention here that an abundant literature has been dedicated to the issues of the potential toxicity of carbon nanomaterials towards humans [START_REF] Shvedova | Exposure to carbon nanotube material: assessment of nanotube cytotoxicity using human keratinocyte cells[END_REF][START_REF] Lam | A review of carbon nanotube toxicity and assessment of potential occupational and environmental health risks[END_REF] and the environment [START_REF] Cheng | Effect of carbon nanotubes on developing zebrafish (danio rerio) embryos[END_REF] since the early beginnings of this research field. Antimicrobial activity of carbon nanomaterials is related to the same kind of mechanisms but is considered in a positive way because it is targeted toward the elimination of pathogens. Most of the work related to CNMs has focused on the prevention of biofilm formation and/or elimination of bacteria.

Carbon nanomaterials antimicrobial mechanisms

Antimicrobial behaviour of CNMs in suspension in a liquid has been the main focus in the literature to date because this is the most common way to expose cells in culture. Authors have reported so far two main antibacterial mechanisms: physical damage and oxidative stress.

CNMs may indeed impart mechanical damage to microorgan isms through perforation of the outer membrane upon contact, causing the loss of cell components and in consequence cell death. The exact mechanism may slightly change upon the type of nano material. For example, graphene (or more likely FLG) may be considered as a "nano knife" (2D) cutting the membrane, whereas it would be more accurate to describe a CNT as a "nano dart" (1D) (Fig. 4a) while these 2 approaches certainly depend on the me chanical rigidity of the nanoparticles: indeed, it does not seem very realistic to consider actual graphene, a very flexible nanomaterial, as a knife while this may be more likely in the case of FLG, which is much more rigid. The same applies to CNTs, where few walled CNTs such as SWCNTs and DWCNTs are more likely to be progres sively included within the cell membrane while more rigid MWCNTs may indeed penetrate like needles (Fig. 4a). Destabilization of Bacterial membrane may also be obtained via electrostatic interaction with the nanomaterials. This is expected with functionalized carbon nanomaterials, especially when they have been oxidised and bear carboxylate ( COO -) groups on their surface. Furthermore, the separation of microorganism from their microenvironment via isolation is another possible mechanism. For example, a graphene sheet may wrap itself around a microor ganism and block its access to nutrients, this action is illustrated in thus increase the antimicrobial activity [113]. Therefore, it is diffi cult to predict the particles action and their efficiency [START_REF] Nowack | Potential scenarios for nanomaterial release and subsequent alteration in the environment: nanoparticle release[END_REF]. Although most earlier reviews have tried to highlight relationships between the physico chemical parameters of CNMs and their antimicrobial activity, no clear conclusions have been established so far. Authors remain cautious with the mention of phenomena that remain difficult to clearly evidence or even controversial [START_REF] Hegab | The controversial antibacterial activity of graphene-based materials[END_REF][START_REF] Zou | Mechanisms of the antimicrobial activities of graphene materials[END_REF]. The explanation is that there are too many parameters to consider, from the particles themselves, to the biological environ ment, the type of microorganism tested and the type of assay. It is thus very challenging to compare existing studies and to draw general conclusions. New studies should be conducted using standardized experimental procedures in order to reach this goal. The grouping and read across concepts may help to progress to wards this goal [START_REF] Wigger | Material-specific properties applied to an environmental risk assessment of engineered nanomaterials implications on grouping and read-across concepts[END_REF].

After having reviewed the intrinsic antimicrobial properties of CNMs, the situation of their combination with other known antimicrobial agents will now be discussed. To date, the following associations have been described in the literature: CNMs func tionalization with Nano Ag, metals, photocatalytic agents, and antibiotics. This description of the state of the art is important because part of the mechanisms proposed in the case of the CNMs dispersed in a liquid medium may not apply when they are immobilized in the case of an antimicrobial surface.

Carbon nanomaterials functionalization

Functionalization of CNMs (Fig. 5) has already been extensively described and this is not the goal here to review this topic, but only to present and discuss relevant functionalization approaches for antimicrobial applications. In the vast majority of examples found in the literature, the functionalization was obtained by non covalent adsorption of nanoparticles and/or (antibiotic) mole cules, sometimes involving pi stacking interactions.

Combinations between CNMs and Ag nanoparticles (AgNP) have been widely studied to enhance their antimicrobial activity (Table S1) [27,61,97,117e119]. The use of AgNPs is popular due to their intrinsic antibacterial properties. The antimicrobial activity of silver comes from Ag þ ions and the reason why Ag nanoparticles are more potent than direct exposure to the ions is that while the entry of the latter into the cells may be prevented by some defence mechanisms, nothing would block the penetration of the nano particles which can later dissolve directly in the cytoplasm and release Ag þ ions where nothing can block their diffusion. Ag þ ions may then react with DNA or proteins and cause cell death [START_REF] Pant | Characterization and antibacterial properties of aminophenol grafted and Ag NPs decorated graphene nanocomposites[END_REF]. The adherence of AgNPs on microbes is likely to induce ROS generation [START_REF] Nie | Bioinspired and biocompatible carbon nanotube-Ag nanohybrid coatings for robust antibacterial applications[END_REF][START_REF] Kovvuru | Oral ingestion of silver nanoparticles induces genomic instability and DNA damage in multiple tissues[END_REF]. The antimicrobial activity of AgNPs depends greatly on their size and dispersibility [START_REF] Zhou | Ecofriendly and environment-friendly synthesis of size-controlled silver nanoparticles/graphene composites for antimicrobial and SERS actions[END_REF][START_REF] Lukowiak | Antimicrobial graphene family materials: progress, advances, hopes and fears[END_REF]: the large specific surface area of CNMs ensures a good dispersion of the AgNPs upon grafting [START_REF] Pant | Characterization and antibacterial properties of aminophenol grafted and Ag NPs decorated graphene nanocomposites[END_REF][START_REF] Han | Graphene family nanomaterials (GFNs) promising materials for antimicrobial coating and film: a review[END_REF][START_REF] Nie | Bioinspired and biocompatible carbon nanotube-Ag nanohybrid coatings for robust antibacterial applications[END_REF]. Furthermore, synthesis processes on CNMs have been developed to control AgNPs size, ensuring an enhanced antimi crobial activity [START_REF] Nie | Bioinspired and biocompatible carbon nanotube-Ag nanohybrid coatings for robust antibacterial applications[END_REF].

Literature also reports CNMs functionalization with other metals or their corresponding oxide to enhance their antimicrobial properties (Table S1). Adsorption of Fe on graphene was mentioned as a good combination for enhanced adsorption of contaminants in the water such as Cr, As and Pb [START_REF] Gollavelli | Facile synthesis of smart magnetic graphene for safe drinking water: heavy metal removal and disinfection control[END_REF]. It is also possible to modify the electrical properties of CNMs upon their functionalization with metals [START_REF] Panda | Electron transfer directed antibacterial properties of graphene oxide on metals[END_REF][START_REF] Li | Antibacterial activity of large-area monolayer graphene film manipulated by charge transfer[END_REF]. For instance, deposition of graphene on p doped Ge was evidenced to attract electrons from the respiratory chain of a microorganism [START_REF] Geng | Antibacterial ability and hemocompatibility of graphene functionalized germanium[END_REF]. This action combined to the ROS production of the nanomateria leads to the death of microorganisms. The antimicrobial effect of metal ions combined to CNMs was also identified [START_REF] Cheong | Synergistic antifungal study of PEGylated graphene oxides and copper nanoparticles against Candida albicans[END_REF]. The enhanced antimicrobial action of a graphene sponge decorated with Cu ions was reported [START_REF] Deng | Graphene sponge decorated with copper nanoparticles as a novel bactericidal filter for inactivation of Escherichia coli[END_REF]. Cu ions are able Fig. 4b. The proliferation is then stopped and the microorganism may be destroyed [START_REF] Shi | The antibacterial applications of graphene and its derivatives[END_REF][START_REF] Liu | Fabrication of polylactic acid/carbon nanotubes/chitosan composite fibers by electrospinning for strawberry preservation[END_REF]. CNMs have also been reported as being able to inhibit cellular division of cancer cells. Biomimetic proper ties of MWCNTs that allow them to interfere with microtubules were reported [START_REF] García-Hevia | Inhibition of cancer cell migration by multiwalled carbon nanotubes[END_REF]. These intracellular proteins are nanotubes presenting similarities with MWCNTs, and play a central role in cellular motion and division. MWCNTs interaction with microtu bules filaments leads to interferences and finally inhibits cells proliferation. Although examples of this mechanism were not described so far in the literature, it is supposed that it may also be active against microbial pathogens.

Oxidative stress mechanisms also must be considered to un derstand antimicrobial activity of CNMs. Chemical interactions between the latter and microorganisms' surface may lead to in flammatory reactions with the production of Reactive Oxygen species (ROS) (Fig. 4c). These species may damage lipids, proteins and DNA of bacteria and cause membrane peroxidation, leading to the cell's death. It is worth mentioning that this may strongly depend on the nature of the carbon nanomaterial because fuller enes and carbon nanotubes may simultaneously exhibit intrinsic antioxidant properties [93e95]. In addition, redox reactions may take place with a charge transfer between nanomaterials and bacteria. CNMs may attract cells electrons and cause biological mechanisms such as respiration to stop [START_REF] Xin | Antibacterial carbon-based nanomaterials[END_REF][START_REF] Al-Jumaili | Review on the antimicrobial properties of carbon nanostructures[END_REF][START_REF] Kang | Single-walled carbon nanotubes exhibit strong antimicrobial activity[END_REF][START_REF] Zheng | Antibacterial applications of graphene oxides: structure-activity relationships, molecular initiating events and biosafety[END_REF][START_REF] Han | Graphene family nanomaterials (GFNs) promising materials for antimicrobial coating and film: a review[END_REF]. Bacteria, contrary to eukaryote cells, have no mitochondria and reactions associated to the respiratory chain are localised in the cell mem brane. CNMs may accept an electron from the respiratory chain and inhibit the transport reaction leading to a bactericide effect [START_REF] Mashino | Antibacterial and antiproliferative activity of cationic fullerene derivatives[END_REF][START_REF] Parise | Activity inhibition on municipal activated sludge by single-walled carbon nanotubes[END_REF]. Unlike viruses, fungi have mitochondria, however studies on CNMs impact on fungal mitochondria were not found in the literature. To date, examples of mitochondrial injuries induced by CNMs were only described in mammal cells [START_REF] Li | The triggering of apoptosis in macrophages by pristine graphene through the MAPK and TGF-beta signaling pathways[END_REF] or in plant cells [START_REF] Begum | Induction of cell death by graphene in Arabidopsis thaliana (Columbia ecotype) T87 cell suspensions[END_REF].

It is likely that the intrinsic antibacterial activity of CNMs is related to a combination of physical and chemical mechanisms. Other properties of CNMs may be combined to enhance such ac tivity [START_REF] Han | Graphene family nanomaterials (GFNs) promising materials for antimicrobial coating and film: a review[END_REF]. Among different possibilities, CNMs may decrease bac terial viability through photothermal effect with the absorption and conversion of NIR laser irradiation into local heating (Fig. 4d) [81,102 e105 ]. Local heating may also be achieved under microwave irradiation, where stimulated CNTs may induce bacteria thermal ablation [START_REF] Ibelli | Progress on utilizing hyperthermia for mitigating bacterial infections[END_REF][START_REF] Al-Hakami | Fast disinfection of Escherichia coli bacteria using carbon nanotubes interaction with microwave radiation, 458943[END_REF], or under acoustic waves where CNMs may adsorb acoustic energy to induce hyperthermia of cancer cells [108 e110]. Hyperthermia mechanisms are frequently used in can cer therapy and might also be also applied against bacterial path ogens [START_REF] Ibelli | Progress on utilizing hyperthermia for mitigating bacterial infections[END_REF].

Although CNMs antibacterial activity can be obtained through several mechanisms, it is likely that the latter depends on many parameters such as the environment and the nature of the micro organisms. Our survey of the current literature highlights that most of the work published so far has focused on CNTs and GRMs, and especially GO. Furthermore, important differences for a given nanostructure were also identified, depending on its intrinsic pa rameters, including the particles size, thickness, and surface chemistry [START_REF] Hegab | The controversial antibacterial activity of graphene-based materials[END_REF][START_REF] Raphey | Advanced biomedical applications of carbon nanotube[END_REF]. For instance, GO is more efficient than graphene because the oxygen groups facilitate its dispersion, thus allowing better cell/particle contacts. This may also be related to the surface charge effect described earlier (at biological pH, carboxylic func tions should be present as negatively charged carboxylates). In addition, size is an important parameter as a small graphene sheet may penetrate and diffuse into a cell without seriously damaging the membrane, while this would be very different for larger sheets. CNTs also have different action mechanisms upon their size. Shorter tubes seem to have more opportunities for cap/cell inter action and thus should be more efficient [START_REF] Kang | Antibacterial effects of carbon nanotubes: size does matter![END_REF]. However, longer nanotubes may cause damages inside the cell while transiting and to interact with DNA and to stop its replication [START_REF] Li | Graphene oxide loaded with copper oxide nanoparticles as an antibacterial agent against Pseudomonas syringae pv. tomato[END_REF], leading to the microorganism death.

The combination of CNMs with metal oxides is also effective to fight against microorganisms [START_REF] Hegab | The controversial antibacterial activity of graphene-based materials[END_REF] and graphene ZnO nano composites [START_REF] Oves | Graphene decorated zinc oxide and curcumin to disinfect the methicillin-resistant Staphylococcus aureus[END_REF][START_REF] Zhong | Graphene oxide-modified ZnO particles: synthesis, characterization, and antibacterial properties[END_REF] or CNT functionalization with ZnO have been reported [START_REF] Sui | Synthesis of ZnO coated multi-walled carbon nanotubes and their antibacterial activities[END_REF]. ZnO graphene has an enhanced antimicrobial ac tivity compared with pristine graphene [START_REF] Kavitha | Glucose sensing, photocatalytic and antibacterial properties of graphene ZnO nanoparticle hybrids[END_REF], the composite inducing death by interacting with its proteins [START_REF] Zanni | Evaluation of the antibacterial power and biocompatibility of zinc oxide nanorods decorated graphene nanoplatelets: new perspectives for antibiodeteriorative approaches[END_REF] and presenting a long lasting activity due to graphene slowing the dissolution of ZnO [START_REF] Wang | Superior antibacterial activity of zinc oxide/graphene oxide composites originating from high zinc concentration localized around bacteria[END_REF][START_REF] Chen | Characterizations of zinc oxide nanorods incorporating a graphene layer as antibacterial nanocomposites on silicon substrates[END_REF]. Many other examples may also be found, such asso ciations of CNMs with as CuO [START_REF] Li | Graphene oxide loaded with copper oxide nanoparticles as an antibacterial agent against Pseudomonas syringae pv. tomato[END_REF][START_REF] Palza | Antibacterial carbon nanotubes by impregnation with copper nanostructures[END_REF], SnO 2 [START_REF] Wu | Synthesis and antibacterial activities of graphene decorated with stannous dioxide[END_REF][START_REF] Mohan | Surface modified graphene/SnO 2 nanocomposite from carbon black as an efficient disinfectant against Pseudomonas aeruginosa[END_REF], Co 3 O 4 [START_REF] Alsharaeh | Novel route for the preparation of cobalt oxide nanoparticles/reduced graphene oxide nanocomposites and their antibacterial activities[END_REF][START_REF] Liu | Transgenerational effects of reduced graphene oxide modified by Au, Ag, Pd, Fe 3 O 4 , Co 3 O 4 and SnO 2 on two generations of Daphnia magna[END_REF], WO 3 [START_REF] Ahmed | Well-controlled in-situ growth of 2D WO 3 rectangular sheets on reduced graphene oxide with strong photocatalytic and antibacterial properties[END_REF], and PbO [START_REF] Fakhri | Degradation photocatalysis of tetrodotoxin as a poison by gold doped PdO nanoparticles supported on reduced graphene oxide nanocomposites and evaluation of its antibacterial activity[END_REF].

It is possible to confer photocatalytic properties to CNMs by association with a photocatalytic agent (Table S2). A photocatalytic agent can produce ROS under light irradiation, and as such they may deteriorate microorganisms via oxidative stress. As already discussed, CMNs exhibit photothermal activity, good electrical conductivity and easily produce photo induced electron; thus CNMs are good base materials for photocatalytic functionalization [START_REF] Lukowiak | Antimicrobial graphene family materials: progress, advances, hopes and fears[END_REF][START_REF] Yi | Selective prepared carbon nanomaterials for advanced photocatalytic application in environmental pollutant treatment and hydrogen production[END_REF]. TiO 2 is a well known photocatalytic agent that has been used to form CNMs based composites exhibiting antimicrobial ac tion under solar light irradiation [144e150]. Literature also reports CNMs functionalized with other photocatalysts such as ZnO [151e153], CdS [START_REF] Gao | Graphene oxide CdS composite with high photocatalytic degradation and disinfection activities under visible light irradiation[END_REF][START_REF] Deng | Graphene CdS nanocomposite inactivation performance toward Escherichia coli in the presence of humic acid under visible light irradiation[END_REF], MnS 2 [START_REF] Fakhri | Synthesis and characterization of MnS 2 /reduced graphene oxide nanohybrids for with photocatalytic and antibacterial activity[END_REF], WO 2 ; WO 3 [START_REF] Akhavan | Protein degradation and RNA efflux of viruses photocatalyzed by graphene tungsten oxide composite under visible light irradiation[END_REF], Cu 2 O [START_REF] Hou | Facile synthesis of water-dispersible Cu 2 O nanocrystal reduced graphene oxide hybrid as a promising cancer therapeutic agent[END_REF], Ag 3 PO 4 [START_REF] Liu | Graphene oxide enwrapped Ag 3 PO 4 composite: towards a highly efficient and stable visible-light-induced photocatalyst for water purification[END_REF], g C 3 N 4 [START_REF] Sun | Antibacterial activity of graphene oxide/g-C 3 N 4 composite through photocatalytic disinfection under visible light[END_REF].

Authors also explored the association of CNMs with antibiotics (Table S3). The utilization of CNMs exhibiting different mechanisms of action than antibiotics allows them to overcome the issue of the development of resistance from some microorganisms such as E. coli or S. aureus. The synergistic effects of CNMs and antibiotics produces a composite with an increased antibacterial activity compared to its individual counterparts [161e163]. Association of GO and Tetracycline (TET) was reported to enhance the interaction between the antibiotic substance and microorganisms [START_REF] Mishra | New life for an old antibiotic[END_REF]. Two modes of contact influenced by GO were observed such as the wrapping of the nanocomposite on the resistant bacteria. The important concentration of Tetracycline on GO and the direct contact with the cell does not allow the efflux pump gene responsible of the antibiotic resistance of the bacteria to eject TET from the cell. A CNT lysine nanocomposite also presented a better antimicrobial activity than its pristine counterparts with the direct contact mechanism of CNT allowing the antibiotic to diffuse into the cell and avoid rejection from the microorganism [START_REF] Amiri | Efficient method for functionalization of carbon nanotubes by lysine and improved antimicrobial activity and water-dispersion[END_REF].

The use of CNMs as a substrate was also proposed to increase the dispersion and stability of antibiotics. SWCNT ciprofloxacin exhibited good dispersion and stability for over 6 months [START_REF] Assali | Single-walled carbon nanotubesciprofloxacin nanoantibiotic: strategy to improve ciprofloxacin antibacterial activity[END_REF].

Moreover, the functionalization of CNMs with antibiotics allows more control over the release of the substance [START_REF] Yuan | Self-assembly of multilayered functional films based on graphene oxide sheets for controlled release[END_REF] and thus the requirement of lower quantities.

Finally, there are many examples of functionalization of CNMs simultaneously by different compounds to further enhance the properties of the nanocomposite (Table S4). Multiple metals or oxides may be used, such as bimetal functionalization of CNTs exhibiting enhanced activity compared to single metal based nanocomposites [168e170]. The functionalization of graphene with Cu and Ag nanoparticles where the synergistic action of the metals on the graphene support increased the composite antimi crobial activity was reported [START_REF] Perdikaki | Ag and Cu monometallic and Ag/Cu bimetallic nanoparticle graphene composites with enhanced antibacterial performance[END_REF]. It is also possible to optimize the photocatalytic action of CNMs [172e174]. The photocatalytic degradation activity of SWCNTs functionalized with Cu and TiO 2 for pharmaceutical wastewater decontamination was mentioned in literature [START_REF] Payan | Catalytic decomposition of sulfamethazine antibiotic and pharmaceutical wastewater using Cu-TiO 2 @functionalized SWCNT ternary porous nanocomposite: influential factors, mechanism, and pathway studies[END_REF]. The combined effects of TiO 2 as a photocatalytic agent and Cu (antimicrobial) enhanced the global photocatalytic activity. Combination with a polymer is another way to increase CNMs antimicrobial activity [176e178]. A zinc oxide multiwalled carbon nanotube poly(vinyl chloride) ternary nanocomposite with increased antimicrobial and biocompatibility properties due to the synergistic effects of the compounds was reported [START_REF] Manikandan | Direct glucose sensing and biocompatible properties of a zinc oxide multiwalled carbon nanotube poly(vinyl chloride) ternary composite[END_REF].

These different examples of CNMs functionalization for anti microbial application highlight the synergistic effects of such nanocomposites. Unfortunately, this approach has only been vali dated in suspension, and not after immobilisation on a surface. Even if the improvement of the dispersion of antimicrobial com pounds offered by CNMs may be transposed to the surface of ma terials, the issue of the durability of the action is generally not solved because of the need for the release of the antimicrobial substance from CNMs (dissolution of metal ions, release of anti biotic molecules). The photocatalytic properties of CNMs associated to a photocatalytic agent is one exception.

Finally, CNMs antimicrobial properties and the associated mechanisms could also occur in case of contact with human or animal cells, which raises the issue of the balance between benefits (antimicrobial activity) and risks (toxicity/environmental impact) [30e32]. Numerous studies have been conducted to determine the potential toxicity of CNMs, nonetheless it is difficult to draw general conclusions. Indeed, the comparison of studies is delicate because, as previously discussed, nanoparticles effects depend on many parameters that may even vary upon the environment [START_REF] Nowack | Potential scenarios for nanomaterial release and subsequent alteration in the environment: nanoparticle release[END_REF]. The same phenomenon dealing with the establishment of relationships between the physico chemical parameters of CNMs and their biocidal activity also exists for their toxicity assessment. Even if hazard is not always demonstrated, solutions to avoid this issue graphene, oxidised or not) are also mostly chosen because of their established antimicrobial properties in literature [START_REF] Han | Graphene family nanomaterials (GFNs) promising materials for antimicrobial coating and film: a review[END_REF][START_REF] Azizi-Lalabadi | Carbon nanomaterials against pathogens; the antimicrobial activity of carbon nanotubes, graphene/graphene oxide, fullerenes, and their nanocomposites[END_REF]. Overall, GO appears as the most investigated CNM among all antimicrobial CNM based nanocomposites, followed by CNTs.

The different types of polymer based composites containing CNMs will now be presented through their elaboration method and antimicrobial action. Fig. 6 illustrates the different possible situa tions that we have identified.

Active and passive routes refer to their main mechanism of ac tion. In the passive route, the surface of the material mainly pre vents microbial adhesion, without necessarily damaging the pathogen. In the active route, there is on the contrary some anti microbial action leading to cytotoxic effects, ideally ending up with cell death. Most of the processes described to prepare bulk com posites or coating composites for the active route may also be used for the passive one. We focus here on active materials not releasing nanoparticles, in a purpose of avoiding issues related to toxicity and loss of activity with time. Different situations may occur among antimicrobial CNM based nanocomposites. Most matrices usually reported in the literature do not exhibit intrinsic antimicrobial properties. The purpose of their use is to disperse them in order to enhance their activity, while also trapping them and preventing unwanted release. In some other cases, the matrix may also exhibit antimicrobial activity as described earlier, that combine with those of the CNMs. Finally, sometimes the addition of CNMs is not related to their antimicrobial properties but only aiming at mechanically reinforcing a matrix already exhibiting its own antimicrobial ac tivity. These different situations will now be described and commented.

Biocompatible matrices will be used for biomedical applica tions, and the question of their stability in use conditions is a very important criterion because their biodegradation would lead to the release of the CNMs. For instance agarose is a good binder [START_REF] Wang | A facile and green method to fabricate graphene-based multifunctional hydrogels for miniature-scale water purification[END_REF] and the biocompatible properties and low toxicity of PVA (Poly(vinyl alcohol)) makes it an ideal candidate for biomedical applications, but their biodegradation properties must be considered [START_REF] Yang | Preparation and characterization of antibacterial electrospun chitosan/poly (vinyl alcohol)/graphene oxide composite nanofibrous membrane[END_REF].

Polyvinyl N carbazole (PVK) was used to prepare nano composite materials with enhanced dispersion of CNMs, allowing a better contact between nanoparticles and pathogen agents, thus improving the antibacterial activity [START_REF] Santos | Antimicrobial graphene polymer (PVK-GO) nanocomposite films[END_REF][START_REF] Ahmed | Antimicrobial PVK:SWNT nanocomposite coated membrane for water purification: performance and toxicity testing[END_REF]. A better antibacterial activity from PVK GO modified surfaces compared to GO modified ones was reported [START_REF] Mejías Carpio | Toxicity of a polymer graphene oxide composite against bacterial planktonic cells, biofilms, and mammalian cells[END_REF].

On the contrary, some works mentioned the use of polymers presenting intrinsic antimicrobial activity. Examples include PVA/ Chitosan [START_REF] Liu | Facile preparation and characterization of poly(vinyl alcohol)/chitosan/graphene oxide biocomposite nanofibers[END_REF]: chitosan, with its cationic nature, is indeed known for its antibacterial activity as a biocidal polymer with direct action [START_REF] Kassem | Antibacterial activity of chitosan nanocomposites and carbon nanotubes: a review[END_REF]. The incorporation of GO particles increased the antibacterial effect of the material, leading to the conclusion that a synergistic effect occurred. Furthermore, the incorporation of CNMs may improve polymers general properties as well: such nanocomposites may thus exhibit enhanced mechanical and hydrophilic properties, in addition to their antibacterial ones [START_REF] He | Promoted water transport across graphene oxide poly(amide) thin film composite membranes and their antibacterial activity[END_REF][START_REF] Cha | Carbon-based nanomaterials: multifunctional materials for biomedical engineering[END_REF]. Some articles even reported antimicrobial composites containing CNMs only added to improve the physical properties. The addition of MWNTs into a CS/ PLA (PLA: polylactic acid) matrix was indeed mentioned for impacting only its mechanical (tensile and bending) properties [START_REF] Liu | Fabrication of polylactic acid/carbon nanotubes/chitosan composite fibers by electrospinning for strawberry preservation[END_REF].

Methods of elaboration of nanocomposites

In this section, the different methods of elaboration of nano composites are described and antimicrobial properties as well as proposed mechanisms are discussed later, but are gathered in Table 1. Two different types of nanocomposite materials may be found depending on the presence of the CNMs in the bulk or only at 1 planarTECH Graphene Facemask, https://planartech.co.uk/.

have been investigated, the most obvious one being their inclusion in composite nanomaterials to limit the release and associated toxicity. This approach is in line with the safer by design strategy which aims at proposing engineering solutions to make possible the responsible use of nanomaterials.

Composites

Carbon nanomaterial based composites

Since the last 10 years, the development of carbon nanomaterial based composites for antimicrobial applications has received increased scientific attention as illustrated in Fig. S5.

In 10 years, the number of publications concerning antimicro bial composites containing carbon nanomaterials has gradually increased. We found that the activity in this field started in 2011 and did not evolve much until 2014, when it started to grow regularly to reach 87 publications in 2020. This may be explained by the increasingly easier access to carbon nanomaterials during the last decade, and the growing need for new strategies to fight against microorganisms.

The inclusion of nanoparticles within composite materials al lows for a whole material to benefit from the advantageous prop erties of the nanoparticles without the potential toxicity issue because of their trapping in the matrix, preventing their release. It appears interesting and promising to study the transfer of these particles' properties into a bulk material following their incorpo ration into the matrix as such materials find various applications, especially in the medical area with the fabrication of implantable medical devices or antimicrobial textiles. Because the interaction between pathogens and the material occurs only at the surface, it seems unnecessary to disperse CNMs in the bulk, and this topic will be further discussed later. In the current context of worldwide sanitary crisis due to the spread of COVID 19, graphene has been highlighted with for example the development of graphene based respiratory masks, commercialised by several companies. Among them, IDEATI 1 proposes a cotton mask coated with graphene and other carbon nanomaterials that is reusable and displays antibac terial properties, although antiviral properties of graphene have not been clearly demonstrated so far. Bioserenity 2 proposes a reusable mask containing 4 layers, one containing graphene and said to display antiviral properties. Dezeen 3 developed a system where graphene contained into the mask is able to conduct electricity in order to produce charges and to repel trapped particles. Thus, graphene has become very interesting for medical research and industry as a way to limit COVID 19 proliferation among the pop ulation. Uses were mostly found in the medical area [START_REF] Aslan | Antimicrobial biomaterials based on carbon nanotubes dispersed in poly(lactic-co-glycolic acid)[END_REF][START_REF] Surud Zi C | The effect of graphene loading on mechanical, thermal and biological properties of poly(vinyl alcohol)/graphene nanocomposites[END_REF][START_REF] Mohamed | Novel aminohydrazide cross-linked chitosan filled with multi-walled carbon nanotubes as antimicrobial agents[END_REF][START_REF] Liu | Antibacterial graphene oxide coatings on polymer substrate[END_REF], but also for environmental applications with the new filters for water purification [START_REF] Musico | Surface modification of membrane filters using graphene and graphene oxide-based nanomaterials for bacterial inactivation and removal[END_REF][START_REF] Ahmed | Antimicrobial PVK:SWNT nanocomposite coated membrane for water purification: performance and toxicity testing[END_REF][START_REF] Wang | A facile and green method to fabricate graphene-based multifunctional hydrogels for miniature-scale water purification[END_REF] or in the food industry with food packaging [START_REF] Grande | Chitosan cross-linked graphene oxide References nanocomposite films with antimicrobial activity for application in food industry[END_REF][START_REF] Liu | Fabrication of polylactic acid/carbon nanotubes/chitosan composite fibers by electrospinning for strawberry preservation[END_REF][START_REF] Mahmoudi | Physicochemical and antibacterial properties of chitosan-polyvinylpyrrolidone films containing self-organized graphene oxide nanolayers[END_REF] for example.

Certain CNMs are preferred for the elaboration of antimicrobial nanocomposite materials. This concerns CNTs and graphene, along with their oxidised forms. This may be related to the availability of these materials, CNTs becoming of easier access with their decreasing cost. This selection may also result from their different intrinsic properties. The literature describes GO as the easiest to manipulate among all other CNMs because of its higher dis persibility, especially in water [184 ,185 ], while graphene (FLG) is rather difficult to disperse properly. This is due to the presence of oxygenated functions at the surface of GO which induce a better hydrophilicity and increased antimicrobial activity, possibly also related to the enhanced dispersibility. These CNMs (CNTs and the surface of the material. The interface between microorganisms and CNMs is at the surface of the composite. Thus, it seems un necessary to include CNMs in the whole material. Different ap proaches are used to create these composites, some specific methods are used to develop bulk nanocomposites whereas other techniques are designed to simply coat a surface. Fig. 7 and Fig. 8 offer a simple classification of these elaboration methods accord ing to their "bulk" or "surface" nature.

Bulk nanocomposites may be prepared in a very simple way by the solvent casting method (Fig. 7a). Basically, this involves the mixing of a polymer solution with a dispersion of CNMs. The resulting liquid is then poured onto a plate to form the final ma terial after evaporation of the solvent. Depending on the quantity casted onto the plate this method allows to prepare bulk nano composites [START_REF] Surud Zi C | The effect of graphene loading on mechanical, thermal and biological properties of poly(vinyl alcohol)/graphene nanocomposites[END_REF][START_REF] Mohamed | Novel aminohydrazide cross-linked chitosan filled with multi-walled carbon nanotubes as antimicrobial agents[END_REF][START_REF] Wang | A facile and green method to fabricate graphene-based multifunctional hydrogels for miniature-scale water purification[END_REF][START_REF] Mahmoudi | Physicochemical and antibacterial properties of chitosan-polyvinylpyrrolidone films containing self-organized graphene oxide nanolayers[END_REF] or films [START_REF] Grande | Chitosan cross-linked graphene oxide References nanocomposite films with antimicrobial activity for application in food industry[END_REF]. A variation of this tech nique was used to create a nanocomposite film from a GO/DODA solution (DODA: dimethyldioctadecylammonium) on water. Such a nanocomposite material may find applications in water treatment or solar energy conversion [START_REF] Yin | Functional free-standing graphene honeycomb films[END_REF].

Another way to fabric a bulk nanocomposite is through the electrospinning method (Fig. 7b). This technology consists in applying an electrical tension on the nozzle of a syringe containing a polymer solution. An opposite voltage is applied onto a collector placed in front. The created electric field causes drops of the polymer solution to stretch and form solid fibres as they travel to the target. This is a simple process allowing the production of few nanometre diameter fibres with large surface area and superior mechanical properties, which may be easily functionalized [START_REF] Agarwal | Use of electrospinning technique for biomedical applications[END_REF]. This technique was reported in the literature for the production of composite fibres containing CNMs [187,188,193e195]. It was used for example to produce polylactic acid/CNTs/chitosan nano composite fibres in one step because the CNMs can be included directly in the polymer solution [START_REF] Liu | Fabrication of polylactic acid/carbon nanotubes/chitosan composite fibers by electrospinning for strawberry preservation[END_REF][START_REF] Wang | A facile and green method to fabricate graphene-based multifunctional hydrogels for miniature-scale water purification[END_REF]. Polymers are selected according to their inherent properties in view of the application. For example, Polyethylene oxide (PEO), Polylactic acid (PLA) or Poly(vinyl alcohol) PVA are easy to process by electrospinning and are thus among the most used polymers to develop electrospun nanofibers [START_REF] Yang | Preparation and characterization of antibacterial electrospun chitosan/poly (vinyl alcohol)/graphene oxide composite nanofibrous membrane[END_REF]. Another method recently developed called 3D printing or additive manufacturing (Fig. 7c) may be considered to prepare bulk nanocomposites [START_REF] Chia | Recent advances in 3D printing of biomaterials[END_REF][START_REF] Sayyar | Fabrication of 3D structures from graphene-based biocomposites[END_REF]. In most cases, this fast and accurate technique allows to fabricate objects with any wanted shape. The 3D printer creates the final structure by heating a polymeric mixture and extruding it layer by layer through a nozzle [START_REF] Hoque | Extrusion based rapid prototyping technique: an advanced platform for tissue engineering scaffold fabrication[END_REF]. Different works reported 3D printed composites containing graphene or CNTs in a polymer matrix for biomedical application [199e202]. In most cases tests involved human cells to confirm the biocompatibility, however only a few studies investigated the antimicrobial properties. This was the case in a 2020 study on 3D printed GO/poly(e caprolactone) fibrous scaffolds that evaluated the antibacterial effect of such a material against S. epidermidis and E. coli [START_REF] Melo | Incorporation of graphene oxide into poly(e-caprolactone) 3D printed fibrous scaffolds improves their antimicrobial properties[END_REF].

Instead of creating bulk materials, nanocomposites may also be prepared by coating a substrate. This is especially relevant to limit the amount of CNMs, while obtaining the same surface properties. For instance, the spin coating technique (Fig. 8a) consists in drop ping the coating solution/suspension onto a rotating substrate. The centrifuge force allows the liquid to form a uniform layer on the substrate, control of rotation speed rate allows an easy control of the film thickness. This technique was described in the literature for the production of CNMs based nanocomposites [START_REF] Li | Identification and optimization of carbon radicals on hydrated graphene oxide for ubiquitous antibacterial coatings[END_REF][START_REF] Mejías Carpio | Toxicity of a polymer graphene oxide composite against bacterial planktonic cells, biofilms, and mammalian cells[END_REF]. It was used to cast a drop of a PLGA SWNT solution (PLGA: Poly(lactic co gly colic acid)) on a rotating cleaned glass substrate to coat it with the corresponding nanocomposite film [START_REF] Aslan | Antimicrobial biomaterials based on carbon nanotubes dispersed in poly(lactic-co-glycolic acid)[END_REF]. Spin coating is a very simple, fast, and effective technique when only one side of a ma terial needs to be modified. Dip coating (Fig. 8b) is another common coating method used for the fabrication of CNMs based nanocomposites [START_REF] Li | Identification and optimization of carbon radicals on hydrated graphene oxide for ubiquitous antibacterial coatings[END_REF][START_REF] Liu | Antibacterial graphene oxide coatings on polymer substrate[END_REF], where the substrate is immersed into a coating material solution/sus pension and retrieved at controlled speed. PVK SWNT coated nitrocellulose membranes were prepared using this strategy [START_REF] Ahmed | Antimicrobial PVK:SWNT nanocomposite coated membrane for water purification: performance and toxicity testing[END_REF]. Contrary to spin coating, this process is relatively slow, with only limited control of the film thickness. However, it makes possible to cover the whole surface of a substrate in a single step.

Vacuum filtration (Fig. 8c) was also mentioned in the literature as a membrane coating method. The substrate is fixed as a filter on a vacuum filtration setup, then the coating suspension is poured onto the substrate and the coating process finally takes place during its filtration. SWNT/PVDF (PVDF: Poly(vinylidene fluoride)) micropo rous membranes were successfully obtained by this technique [START_REF] Brady-Est Evez | A single-walled-carbon-nanotube filter for removal of viral and bacterial pathogens[END_REF]. Furthermore, the vacuum filtration also allows to prepare nano composite films, for example to filter a GO PEI/BC (PEI: Polyethylen imine; BC: bacterial cellulose) dispersion in order to obtain free standing disc like films instead of considering the filter as the final substrate [START_REF] Yang | Improvement of antimicrobial activity of graphene oxide/bacterial cellulose nanocomposites through the electrostatic modification[END_REF]. This method was also used to prepare a free standing paper composed of Tween 20 (Tween 20: Polyoxy ethylene(20) sorbitan monolaurate) and rGO. Similarly to dip coating, vacuum filtration is rather slow and does not offer a very accurate control of the homogeneity and film thickness [START_REF] Park | Biocompatible, robust free-standing paper composed of a TWEEN/graphene composite[END_REF]. A nanopaint designed to coat a material may be prepared by simply mixing components together. For instance a GO nanopaint exhibiting both antibacterial and antifouling properties was fabri cated via the ball milling technology [START_REF] Krishnamoorthy | Graphene oxide nanopaint[END_REF]. This method involves a bowl containing the components and grinding balls. It is an easy way to mix materials together, however a major disadvantage is the possible incorporation of impurities from the bowl and/or the balls [START_REF] Chua | Ball-milled sulfur-doped graphene materials contain metallic impurities originating from ball-milling apparatus: their influence on the catalytic properties[END_REF].

Another method to coat a substrate is through electrophoretic deposition (EDP) (Fig. 8d). This technique involves the attachment of suspended charged particles onto an opposite charged surface through the influence of an electric field. This presents several advantages such as good surface homogeneity and easy control of the film thickness. This technique was used to immobilize a PVK GO composite onto an indium tin oxide (ITO) substrate [START_REF] Santos | Antimicrobial graphene polymer (PVK-GO) nanocomposite films[END_REF]. This technique may also be considered to simply deposit particles onto materials surface. It was used to deposit graphene onto stainless steel [START_REF] Akhavan | Toxicity of graphene and graphene oxide nanowalls against bacteria[END_REF].

The Electrospraying method (Fig. 8e) is another technique which can be considered as a variation of the electrospinning Furthermore, techniques able to specifically remove the top polymer layer on the surface of a material may be used to make particles available at the surface (even if they were initially embedded in a polymer matrix) are very interesting. This area of research is not very developed yet, but surface treatments such as laser ablation, plasma or UV/O 3 oxidation processes are already available and may be used to achieve this goal. The industrial development of Carbon nanomaterials based polymer matrix nanocomposites for antimicrobial applications is thus entirely conceivable. In conclusion, there is a wide choice of different techniques and the most appropriate one may be selected depending on technical limitations. The question of the strength of the interaction between CNMs and the surface is very important both in terms of durability of the antimicrobial effect and the limitation of the release of the CNMs that would raise issues of potential toxicity.

Interaction between the components

We discuss here the different possible interactions between CNMs and a polymer matrix, in the context of the safety (no release) but also durability (longer efficacy) for antimicrobial ap plications. In a nanocomposite, different interactions occur be tween CNMs and the polymeric matrix, depending on the physio chemical properties of both components, such as the carbon nanoparticles oxidation state, or the polarity of the polymer for instance. Chemical interactions between components can be created, however physical ones are often enough to prepare a composite and many studies report only physical interactions be tween particles and the matrix to explain the formation of the nanocomposite. GOepoly(amide) thin film composite membranes were produced and the FTIR analysis suggested that the particles were only embedded in the matrix because no strong chemical binding was evidenced between the components [START_REF] He | Promoted water transport across graphene oxide poly(amide) thin film composite membranes and their antibacterial activity[END_REF]. Aslan et al. also reported the incorporation of SWNTs into a PLGA matrix, where their FTIR characterization suggested that not all SWNT were embedded in the polymer [START_REF] Aslan | Antimicrobial biomaterials based on carbon nanotubes dispersed in poly(lactic-co-glycolic acid)[END_REF]. Other works also described weak interactions between CNMs and a polymeric matrix. For example, interactions between the pi bonds of multi walled carbon nano tubes sidewalls and chemically modified chitosan derivatives were reported to ensure a homogeneous dispersion of the particles in side the matrix [START_REF] Mohamed | Novel aminohydrazide cross-linked chitosan filled with multi-walled carbon nanotubes as antimicrobial agents[END_REF]. Literature also reported interactions such as hydrogen bonding between the particles and the matrix [START_REF] Surud Zi C | The effect of graphene loading on mechanical, thermal and biological properties of poly(vinyl alcohol)/graphene nanocomposites[END_REF][START_REF] Wang | A facile and green method to fabricate graphene-based multifunctional hydrogels for miniature-scale water purification[END_REF]. Hydrogen bonds between oxygen containing groups on the surface of GO and the hydroxyl groups of an epoxy matrix were identified through FTIR characterization [START_REF] Wang | Effect of oxygen-containing functional groups in epoxy/reduced graphene oxide composite coatings on corrosion protection and antimicrobial properties[END_REF]. It is also possible to find hydrogen bonding between two polymers forming the matrix [START_REF] Liu | Fabrication of polylactic acid/carbon nanotubes/chitosan composite fibers by electrospinning for strawberry preservation[END_REF]. For instance hydrogen bonds between the chitosan and PLA (Pol ylactic acid) chains in a GO PVA CS composite were reported [START_REF] Yang | Preparation and characterization of antibacterial electrospun chitosan/poly (vinyl alcohol)/graphene oxide composite nanofibrous membrane[END_REF]. Furthermore the reported electrostatic interactions in the case of rGO DODA allowed to avoid any possible leak from the nano composite [START_REF] Yin | Functional free-standing graphene honeycomb films[END_REF]. Multiple interactions were also reported, such as the combination of electrostatic and hydrogen bonding interactions between GO sheets and lipid molecules present in alkyd resin, allowing the formation of GO nanopaint [START_REF] Krishnamoorthy | Graphene oxide nanopaint[END_REF]. Covalent bonding was also mentioned [START_REF] Li | Identification and optimization of carbon radicals on hydrated graphene oxide for ubiquitous antibacterial coatings[END_REF]. The formation of covalent bonds between GO and chitosan were identified, a crosslinking reaction through nucleophilic addition taking place with the reaction between the epoxy groups in GO and the amino and primary hydroxyl groups in chitosan. This reaction is rather easy and leads to strong binding, improving the mechanical properties of the GO chitosan films [START_REF] Grande | Chitosan cross-linked graphene oxide References nanocomposite films with antimicrobial activity for application in food industry[END_REF].

The type of interaction occurring between the components is of great importance as it may cause stability differences between nanocomposites. As introduced previously, the control of nano particles toxicity is a major issue. Numerous studies have been method, but the electrical tension applied on the liquid contained in the syringe results in the emission of small droplets on the target instead of fibres. This method allowed the preparation of GO thin films wrapped onto electrospun fibroporous polycarbonate ure thane to create a membrane that may be used for future biomedical application [START_REF] Thampi | Differential adhesive and bioactive properties of the polymeric surface coated with graphene oxide thin film[END_REF].

Other strategies have also been described, such as layer by layer assembly (LbL) which involves electrostatic interactions [START_REF] Khanna | Fabrication of long-ranged close-packed monolayer of silica nanospheres by spin coating[END_REF][START_REF] Puetz | Dip coating technique[END_REF] and atom transfer radical polymerisation ATRP [START_REF] Joo | Preparation of antibacterial PDMAEMAfunctionalized multiwalled carbon nanotube via atom transfer radical polymerization[END_REF] to graft CNMs onto a material and form a stable modified surface.

Surface coating methods are very relevant as they allow the presence of nanoparticles only on the surface of the material, which is determinant to enable a direct interaction between particles and microorganisms in term of antibacterial activity. This is generally difficult to achieve with bulk fabrication techniques as particles are often enclosed into the polymeric matrix thus not directly available at the surface. Table 1 presents different examples of composites containing CNMs from articles published between 2008 and 2019.

For each example, it details the fabrication technique, if the matrix has an intrinsic antimicrobial effect, if the antimicrobial mechanism of the composite is known, the antimicrobial assay used and microorganism tested, and finally the proposed application. It may be noticed that most studies related the biocidal activity to direct contact between carbon nanoparticles and microorganisms. Therefore, it is especially important to prepare composites with particles at the surface. It would then be advantageous to find techniques able to bring the particles to the surface even if they are initially embedded within a polymer matrix. A UV/O 3 oxidation treatment on GO nanosheets that allowed to etch away the surface polymer (2 hydroxyethyl methacrylate (HEMA)) and make possible a direct access to the nanoparticles was reported [START_REF] Lu | Enhanced antibacterial activity through the controlled alignment of graphene oxide nanosheets[END_REF]. Further more, laser ablation was chosen in order to resurface GO particles embedded in an agarose matrix [START_REF] Papi | Biomimetic antimicrobial cloak by graphene-oxide agar hydrogel[END_REF]. The laser impulse induced liquefaction and matter transport that allowed the cleaning of the nanoparticles. However, there is no extended research on this area yet and it must be emphasized that the actual demonstration of the absence of a very thin layer of polymer at the surface of the carbon nanomaterials is challenging to perform. The general ideas intro duced here, with a classification of these different methods, are summarized on Fig. 6. Methods that may be used to prepare composites with nanoparticles on the surface are regrouped and framed.

The classification of the nanocomposite's elaboration methods available in the literature and provided in this section may allow the reader to select the most suitable method for a particular application. Composites may be elaborated on a large scale via the techniques presented in this section as they are already used in the industry. This is the case for spin coating [START_REF] Balzarotti | Spin coating deposition on complex geometry substrates_ Influence of operative parameters[END_REF][START_REF] Khanna | Fabrication of long-ranged close-packed monolayer of silica nanospheres by spin coating[END_REF]; dip coating [START_REF] Puetz | Dip coating technique[END_REF]; vacuum filtration [START_REF] Zhang | Graphene based supercapacitor fabricated by vacuum filtration deposition[END_REF]; electrophoretic deposition [START_REF] Verma | Simulating industrial electrophoretic deposition on distributed memory architectures[END_REF] and electro spinning [START_REF] Jaworek | Micro-and nanoparticle production by electrospraying[END_REF]. Furthermore, access to large amounts of CNMs has become easier thanks to enhanced production perfor mances. This is especially the case of CNTs, with the development of an easier production and prices that keep lowering [START_REF] Zhang | Carbon nanotube mass production: principles and processes[END_REF]. Graphene (or more realistically few or multi layer graphene) is still harder to obtain but we may expect a similar evolution in terms of availability within the next years.

As already mentioned, the interface between microorganisms and CNMs is at the surface of the composite. It thus seems un necessary to include CNMs in the bulk, and materials with CNMs only at the surface seem more promising for antimicrobial appli cations with an enhanced activity and a lower cost. Surface coating methods are thus very relevant as they allow the presence of nanoparticles only at the material surface, which is determinant to enable a direct interaction between particles and microorganisms in term of antibacterial activity. conducted on the potential toxicity of CNMs and, if it is still difficult to draw general conclusions, avoiding their release has become a priority. The safer by design strategy aims to benefit from the antimicrobial properties of CNMs into a nanocomposite while avoiding issues related to their release and potential toxicity. Literature reports different stabilities of composites (absence of release of CNMs) depending on the interactions between their components. For instance, differences in terms of stability between a hydrated GO spin coated film and a hydrated GO covalently bonded film were observed. The strong covalent binding appeared to be more durable than the spin coated film with the ability to withstand 30 min under sonication, whereas the spin coated film was only able to resist a few scratches or washings [START_REF] Li | Identification and optimization of carbon radicals on hydrated graphene oxide for ubiquitous antibacterial coatings[END_REF].

Strong chemical interactions are thus preferred. In addition, works presenting physical embedment of the CNMs also exhibiting good stability with a durable composite without release of the nanoparticles are also available [START_REF] He | Promoted water transport across graphene oxide poly(amide) thin film composite membranes and their antibacterial activity[END_REF]. It may then be concluded that a combination between strong chemical and physical in teractions is the safest way to produce a durable and efficient nanocomposite for antimicrobial applications.

However, it is very difficult to assess with certainty the release of carbon nanoparticles from a composite as carbon is present everywhere and the issue concerns the detection of very small quantities of CNMs [START_REF] Mottier | Environmental impact of engineered carbon nanoparticles: from releases to effects on the aquatic biota[END_REF]. This makes it difficult to compare carbon nanoparticles based antimicrobial composite durability in terms of particles release in expected conditions of use. It is essential to extend research on these topics, as even without environmental and toxicity issues, the loss of carbon nanoparticles would question the durability of the device activity as in most cases the antimi crobial activity of carbon nanoparticles based composites is directly linked to the action of the nanoparticles themselves. It is essential to ensure the proper holding of the nanoparticles by the matrix to achieve the safer by design strategy, where devices pre sent durable antimicrobial activity with low risks concerning toxicity and environmental impact. This also mean that the type of matrix used must be selected carefully, because for example a biodegradable matrix will inevitably lead to the release of the particles with time. Due to the technical challenge related to the identification of negligible losses of CNMs in potentially complex environments during the expected conditions of use, it is recom mended to challenge the nanocomposites in harsh conditions (ultrasonication, temperature well above the maximum operation one, low or high pH, etc.) which may allow to certify that the nanocomposites should be used safely in normal conditions.

Antimicrobial properties of composites

Antimicrobial activity of the materials is assessed through qualitative or quantitative tests based on the analysis of materials impact on the proliferation of microorganisms upon contact the internalisation of CNMs should also be ruled out. Furthermore, local heating is likely not to occur in the case of thermal insulating polymers. However, physical action is still possible (membrane damaging, oxidative stress but also antifouling activity), and especially if the CNMs are easily accessible at the surface while being safely entrapped, thus making possible to provide unlimited antimicrobial activity because no CNM is released at any time. Action through oxidative stress without physical contact should also be considered, as nanomaterials may themselves produce oxidative species. However, this situation cannot be generalised because this effect strongly depends on the nature of the carbon nanomaterial (as discussed in part 2, fullerenes and carbon nano tubes may also exhibit intrinsic antioxidant properties).

Conclusion

Composites incorporating carbon based nanomaterials are promising for antimicrobial applications. This review focused on the activity of carbon nanomaterials (carbon nanotubes and gra phene derivatives), incorporated in or at the surface of a polymeric matrix. These types of carbon nanomaterials are still the most frequently investigated in the literature, and we have presented many examples evidencing that their incorporation in a polymer matrix allows to transfer the activity of the particles to a whole material. Most reported polymers are efficient nanoparticle holders, while also demonstrating additional intrinsic antimicro bial properties in some cases. When the antimicrobial properties of the composites are conferred by the carbon nanomaterials, most works link the biocidal activity to direct contact between particles and microorganisms. It is thus important to focus on the surface of materials, which represents the interface between CNMs and mi croorganisms. This simple observation leads to the conclusion that CNMs present in the bulk are of no use and that producing nano composites with nanoparticles at the surface seems to be much more relevant in terms of antimicrobial properties, maximizing the efficacy by localizing the nanoparticles where they are really needed. When blocked at the surface, antimicrobial mechanisms such as the creation of physical damages and subsequent oxidative stress should still be performing. The challenge is then managing to prepare polymeric materials containing carbon nanoparticles exposed at the surface (without being covered by a thin polymeric layer) while making sure that they are safely entrapped, to avoid potential release and related toxicity issues. Processes leading to strong interactions (physical, chemical, or both) with the polymer at the surface should thus be preferred. Among the different CNMs investigated in the literature, GO obviously seems to stand apart, the presence of many oxygen containing functional groups being beneficial for both improving the interface with the polymer matrix and for their intrinsic biocidal activity. However, a possible loss of activity may occur due to the ability of some bacteria to reduce it. There is no doubt that other CNMs such as CNTs, FLG and rGO may find here a favourable field of application not only in the biomedical field, but also in all situations where surfaces should remain free of microbial contamination, such as in the food in dustry with food packaging or for environmental applications with new filters for air or water purification for example. this is espe cially motivated by their expected unique unlimited action, because they should neither dissolve nor be released from the surface, thus also complying with the safer by design strategy.

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.

fluorescence assay and microscopy imaging. For instance, the in hibition zone assay presented in part 1 was used to demonstrate the antimicrobial activity of CS/PVA/GO composites (Fig. 9) [START_REF] Yang | Preparation and characterization of antibacterial electrospun chitosan/poly (vinyl alcohol)/graphene oxide composite nanofibrous membrane[END_REF].

The colony counting method was also adapted to assess the antimicrobial activity of GO poly(amide) membranes with the measurement of cell viability after incubation with the samples (Fig. 10) [START_REF] He | Promoted water transport across graphene oxide poly(amide) thin film composite membranes and their antibacterial activity[END_REF].

Literature also describes the adaptation of the fluorescence assay to measure the antimicrobial activity of a nanocomposite, this may be illustrated with Fig. 11 showing fluorescence images of PVK GO films [START_REF] Santos | Antimicrobial graphene polymer (PVK-GO) nanocomposite films[END_REF]. This technique can only be applied in the case of transparent thin films.

Scanning electron microscopy (SEM) is a common method of characterization that allows the visualisation of the surface morphology of materials in contact with microorganisms. SEM imaging was used to visualise the interaction between a PVK GO composite and B. subtilis bacteria (Fig. 12) [START_REF] Mejías Carpio | Toxicity of a polymer graphene oxide composite against bacterial planktonic cells, biofilms, and mammalian cells[END_REF].

As listed in Table 1, other assays may be used with CNM containing nanocomposites, such as the DNA release assay, Reac tive oxygen species assay, etc. Furthermore, these tools may even be used to identify the component of the nanocomposite which is responsible for the antimicrobial activity.

By testing separately the polymer (matrix), the CNMs and finally the nanocomposites and comparing the results, it is possible to attribute the antimicrobial activity to a particular component. For instance, an inert polypropylene membrane with an increased antimicrobial activity upon the incorporation of Graphene mate rials (GMs) was reported and it was demonstrated that the activity could be attributed to direct contact between graphene and mi croorganisms [209].

The antimicrobial activity of most CNM containing composites may be attributed to the carbon nanomaterials themselves. They may play either an active role by directly killing/damaging the pathogens, or a passive role if they prevent the adhesion of path ogens at the surface (without killing them). In most cases, the matrix alone does not exhibit antimicrobial properties while the nanocomposite does [182 ,190 ]. Indeed, even if the matrix does not have an intrinsic antimicrobial effect, it may enhance the activity of the CNMs by improving their dispersion within the matrix [START_REF] Musico | Surface modification of membrane filters using graphene and graphene oxide-based nanomaterials for bacterial inactivation and removal[END_REF]. Literature often concludes to a transfer of antimicrobial properties from isolated CNMs to the whole material upon their incorporation [209]. The combination of the antimicrobial properties of CNMs and the polymer was also reported, such as in the case of CS PVP GO nanocomposites where both GO and chitosan exhibited an anti microbial activity [START_REF] Mahmoudi | Physicochemical and antibacterial properties of chitosan-polyvinylpyrrolidone films containing self-organized graphene oxide nanolayers[END_REF].

Different scenarios are thus possible, yet literature usually concludes that CNMs are responsible for the antimicrobial activity of the nanocomposite and that their properties are transferred to the whole material. The situation of GO which may be reduced by bacteria [61,115,224e227] requires some specific attention, espe cially if this leads to a loss in terms of antimicrobial activity, thus questioning the durability of the action with time.

As seen in part 2, it is possible to assess the global antimicrobial efficiency of nanocomposite materials thanks to assays that may also be used for CNMs in suspension, however this does not mean that the exact antimicrobial mechanisms can be easily determined. Some antimicrobial mechanisms presented in section 2 (Fig. 4) that are specific to CNMs in suspension may be ruled out to describe what occurs in the case of nanocomposites. As the nanoparticles are immobilized/entrapped within a matrix, the wrapping mechanism where a particle wraps itself around a microorganism and blocks its access to nutrients cannot be considered. Mechanisms involving

Fig. 1 .

 1 Fig. 1. Schematic illustration of the antimicrobial assays, a) Inhibition zone assay and b) colony counting assay (representation of these assays in petri dishes filled with nutritive medium), and fluorescence assay, cells stained with c) DAPI or d) PI (on glass slides). (A colour version of this figure can be viewed online.)

Fig. 2 .

 2 Fig. 2. Carbon nanomaterials: a) Fullerene (C 60 ); b) Carbone Nanotube (CNT); c) Nanodiamond; d) Graphene; e) Graphene Oxide (GO, oxygen atoms shown in red); f) reduced Graphene Oxide (rGO). Adapted from Ref. [30].

Fig. 3 .Fig. 4 .

 34 Fig. 3. Classification grid for the categorization of different graphene types according to three fundamental graphene-based materials properties: number of graphene layers, average lateral dimension, and atomic carbon/oxygen ratio. Reprinted with permission from Ref. [80]. (A colour version of this figure can be viewed online.)

Fig. 5 .

 5 Fig. 5. CNMs possible functionalization described in literature. Adapted from Ref. [30]. (A colour version of this figure can be viewed online.)

Fig. 6 .

 6 Fig. 6. Scheme illustrating the antimicrobial action of composites containing CNMs depending on their design. The passive route corresponds to the prevention of bacterial adhesion (without killing the bacteria), while the active route leads to bacterial killing. The frame highlights different techniques that allow the creation of composites containing CNMs directly accessible on their surface.

Fig. 8 .Fig. 7 .

 87 Fig. 8. Schematic illustration of surface nanocomposite elaboration a) Spin coating; b) Dip coating; c) Vacuum filtration; d) Electrophoretic deposition; e) Electrospraying. Adapted from Ref. [207]. (A colour version of this figure can be viewed online.)

Fig. 9 .

 9 Fig. 9. Picture of inhibition zones of composite nanofibrous membranes in various ratios of CS/PVA/GO; a) CS:PVA:GO 1:9:0; b) CS:PVA:GO 2:8:0; c) CS:PVA:GO 3:7:0; d CS:PVA:GO 1:9:0.01; e) CS:PVA:GO 2:8:0.02; f) CS:PVA:GO 3:7:0.03; g) CS:PVA:GO 1:9:0.03; h) CS:PVA:GO 1:9:0.05. against E. coli (A) and S.aureus (B). Reprinted with permission from Ref. [187]. (A colour version of this figure can be viewed online.)

Fig. 11 .

 11 Fig. 11. a d: Fluorescence images of the E. coli on a) unmodified ITO; b) electrodeposited PVK; c) spin-coated GO; and d) electrodeposited PVK-GO films. Images were obtained with a 100Â oil immersion objective using a FITC filter for green fluorescence from SYTO 9 in all bacteria presented on the left column and a TRITC filter for red fluorescence from PI in membrane-compromised bacteria presented on the right column. e) Plot of the total number of bacteria (green) and among them the number of PI-positive bacteria (red) on the field of view (90 Â 70 mm 21 ), which were adsorbed on the modified surfaces. The blank is the unmodified ITO substrate as the control. f) Correlation of the % non-viable E. coli (% PIpositive) attached on each of the surfaces. Adapted from Ref. [47]. (A colour version of this figure can be viewed online.)

Table 1

 1 Examples of composites containing CNMs from articles published between 2008 and 2019. ~: noqualitative test (deduction from literature); /: no antimicrobial properties; X: no information.

	Bulk composite-Mixing-Direct casting
	Composite	Polymer antimicrobial	Mechanism	Assay	Application	Microorganisms Ref
		property
	G-PVA (Poly(vinyl alcohol))	/	~Physical damage and/or	Colony counting	Biomedical	S. aureus	[60]
			Oxidative Stress	Absorbance (MTT: 3-(4,5-		E. coli	2016
				dimethylthiazol-2-yl)-2,5-
				diphenyltetrazolium
				bromide)
	GO-DODA	antibacterial	Not described	Fluorescence (SYTO 9 PI	Environment (solar	P. aeruginosa	[81]
	(dimethyldioctadecylammonium)			(Propidium iodide))	harvesting devices) and	E. coli	2016
				SEM	biomedical		[59]
							2013
	GO-AG (agarose)	/	~Physical damage and	Fluorescence (DAPI (4',6-	Environment (water	E. coli	[81]
			Oxidative Stress	diamidino-2-ph enylindole)	purification)	S. aureus	2016
				( PI) SEM			[182]
							2013
	GO-CS (Chitosan)	/	Not described	Fluorescence	Food (packaging)	E. coli	[58]
				(SYTO 9 PI)		B. subtillis	2017
	GO-PVP-CS	CS: antibacterial (cationic)	~Physical damage and	Colony counting	Biomedical (wound	S. aureus	[81]
	(PVP: Polyvinyl	PVP:/	Oxidative Stress		dressing)	E. coli	2016
	pyrrolidone)				Food (packaging)		[183]
							2016
	rGO-epoxy	/	Radical Oxidative Stress	Colony counting	Environment (corrosion	E. coli	[213]
					protection)		2018
					Biomedical
	MWCNT-CS-derivatives	Improved performance of	Oxidative Stress	Inhibition zone	Biomedical	E. faecalis; E.	[62]
		CS/MWCNT composite				coli;	2018
		compared to CS				S. epidermidis
						Fungi: A. niger
						C. neoformans
						C. tropicalis
	Bulk composite-Electrospinning
	Composite	Polymer antimicrobial	Mechanism	Assay	Application	Microorganisms Ref
		property
	GO-CS-PVA	CS: antibacterial	Not described	Inhibition zone	Biomedical (wound	E. coli	[81]
		PVA:/			healing and		2016
					drug delivery system)		[188]
							2014
	GO-PVA-CS	CS: antibacterial	Not described	Inhibition zone	Biomedical (wound	E. coli	[187]
		PVA:/			dressing)	S. aureus	2018
	SWNTs-Psf	Psf:/	Physical damage	Fluorescence (DAPI PI)	Antibacterial coating	E. coli	[194]
	Psf: Polysulfone			SEM			2011
	t-MWNT-lignin-PVA	PVA:/	~Physical damage	Serial dilution method	Biomedical (Wound	S. aureus	[195]
		Lignin:/		(Minimum inhibitory	dressings, scaffolds, and		2018
				concentration; Minimum	antimicrobial textile).
				bactericidal concentration)
	CNTs-CS-PLA	CS: antibacterial	PLA/CNTs:/	Colony counting	Food: (Strawberry	E. coli	[91]
		PLA:/	CS/PLA/CNTs:		preservation)	S. aureus	2019
			On bacteria: electrostatic			Fungi:
			effects			B. cinerea and
			On Fungi: aggregation of			Rhizopus
			fungal spores and
			morphological
			abnormalities.
	Coating-Spin coating
	Composite	Polymer antimicrobial	Mechanism	Assay	Application	Microorganisms Ref
		property
	GO-glass	Glass: /	C radical transfer	Colony counting	Biomedical (Coatings for	E. coli	[54]
	hGO-glass (hydrated GO)	Silicon:/	Physical damage	SEM	medical devices, wound	L. crispatus	2016
	GO-Silicon			Fluorescence	healing, antibacterial
				(Bacterial Membrane Lipid	additives in dental
				Peroxidation)	devices)
				Luminescence	Environment (water
				(b-Galactosidase Release	filtration membranes)
				from Bacteria)
	GO-PVK (polyvinyl-N-carbazole) PVK:/	~Physical damage and	PVK-GO suspension:	Biomedical	E. coli, C.	[214]
			Oxidative Stress	Fluorescence (SYTO-9 PI		metallidurans;	2015
			PVK-GO solution:	(propidium iodide);		B. subtilis, R.	[55]
			Wrapping isolation	Metabolic activity)		opacus
				Colony counting

(continued on next page)

Table 1

 1

	(continued )
	Bulk composite-Mixing-Direct casting
	Composite	Polymer antimicrobial	Mechanism	Assay	Application	Microorganisms Ref
		property
		of PVK enhances
		performances)
	Film-Vacuum filtration
	Composite	Polymer antimicrobial	Mechanism	Assay	Application	Microorganisms Ref
		property
	GO PEI/BC (PEI: Polyethylen-	/	~Physical damage;	Colony counting	X	S. aureus	[81]
	imine; BC: bacterial cellulose)		Wrapping isolation and	Fluorescence (PI)		S. cerevisiae	2016
			electrostatic interaction	SEM			[204]
							2016
	rGO-TWEEN 20	TWEEN:/	Non-binding	Optical microscope	Biomedical	B. cereus	[214]
	(Polyoxyethylene(20) sorbitan		Not described	Fluorescence (Calcein PI)			2015
	monolaurate)						[82]
							2010
	Surface functionalization-Electrodeposition
	Composite	Polymer antimicrobial	Mechanism	Assay	Application	Microorganisms Ref
		property
	GO-Steel	Steel:/	~Physical damage	Colony	X	E. coli	[215]
	rGO-Steel			Counting		S. aureus	2018
				Spectrophotometry (RNA			[87]
				efflux Measurement)			2010
	Surface functionalization-Electrospraying
	Composite	Polymer antimicrobial	Mechanism	Assay	Application	Microorganisms Ref
		property
	GO-PCU (Polycarbonate	PCU:/	~Physical damage and	Inhibition zone	Biomedical	S. aureus	[46]
	urethane)		Oxidative Stress	Absorbance (MTT assay)		P. aeruginosa	2017
				Colony counting
	Surface functionalization-Layer by layer
	Composite	Polymer antimicrobial	Mechanism	Assay	Application	Microorganisms Ref
		property
	GO-MPPM (modified	PP (Polypropylene):/	~Physical damage	Colony counting	Environment	E. coli	[81]
	macroporous polypropylene						2016
	Membrane)						[209]
							2015
	GO-PLL hybrid thin film	PLL:/	PLL electrostatic	Luminescence (ATP	Environment (Filtration)	Water from a	[81]
	composite		interaction	(Adenosine Tri Phosphate)	.	local Australian	2016
	PLL: Poly(L-lysine)		GO Physical damage and	test)		lake	[53]
			Radical Oxidative Stress				2015
	SWNT-PLL-PGA	PLL-PGA:/	SWNT	Fluorescence (SYTO 9 PI) Biomedical	E. coli	[214]
	PGA: Poly(L-glutamic acid)		~Physical damage and			S. epidermidis	2015
	SWNT polyelectrolyte films		Oxidative Stress				[208]
							2012
	Surface functionalization-ATRP
	Composite	Polymer antimicrobial	Mechanism	Assay	Application	Microorganisms Ref
		property
	CNTs-PDMAEMA	PDMAEMA: antibacterial	CNTs allow mechanical	SEM	X	E. coli	[210]
	PDMAEMA: Poly[2-	(Electrostatic interactions)	reinforcement, electrical	Colony counting		S. aureus
	(dimethylamino)ethyl		conductivity and
	methacrylate];		antibacterial effects
			Polymer diffusion

Dezeen, https://www.dezeen.com/tag/graphene/.

BioSerenity, https://www.bioserenity.com/.

Acknowledgments

No project grant was used to support this research. We may however aknowledge the Ministry of Higher Education, Research and Innovation for funding the PhD grant of Mrs Laure Giraud.

Appendix A. Supplementary data

Supplementary data to this article can be found online at https://doi.org/10.1016/j.carbon.2021.06.002.