Nanomaterial-Based Antimicrobial Coating for Biomedical Implants: New Age Solution for Biofilm-Associated Infections

Abstract

Recently, the upsurge in hospital-acquired diseases has put global health at risk. Biomedical implants being the primary source of contamination, the development of biomedical implants with antimicrobial coatings has attracted the attention of a large group of researchers from around the globe. Bacteria develops biofilms on the surface of implants, making it challenging to eradicate them with the standard approach of administering antibiotics. A further issue of current concern is the fast resurgence of resistance to conventional antibiotics. As nanotechnology continues to advance, various types of nanomaterials have been created, including 2D nanoparticles and metal and metal oxide nanoparticles with antimicrobial properties. Researchers from all over the world are using these materials as a coating agent for biomedical implants to create an antimicrobial environment. This comprehensive and contemporary review summarizes various metals, metal oxide nanoparticles, 2D nanomaterials, and their composites that have been used or may be used in the future as an antimicrobial coating agent for biomedical implants, as well as their succinct mode of action to combat biofilm-associated infection and diseases.

1. Introduction

All around the world nosocomial (or clinically procured) infections have become a common health risk with a high death rate of over 15% in developing countries.¹ The increased rates of insertion of medical devices and implants have become a common source of such hospital-acquired infections or healthcare-associated infections (HAIs). These are defined as the infections acquired by patients nosocomially while receiving any treatment at a healthcare facility, and such infections include catheter-associated urinary tract infections (CAUTIs), surgical site infections (SSIs), central line associated bloodstream infections (CLABIs), ventilator-associated pneumonia (VAP), hospital-acquired pneumonia, and infections derived from orthopedic implants, dental implants, and so on.² Reed et al. in their report stated that as per the Centers for Disease Control and Prevention 2 million patients estimated annually suffer from HAIs, and 10 000 of them die from the same. Annually, up to $4.5 billion of additional healthcare expenses due to HAIs has been estimated.³ In developed countries like the United States and Europe, the financial losses due to HAIs were estimated at approximately $6.5 billion and €7 billion, respectively.⁴ It is estimated that more than 10% of patients hospitalized have urinary catheters, and more than 30% have vascular catheters. Nosocomial contaminations are thus a significant health challenge, in which 30–40% of the urinary tract infections caused are identified as catheter-related urinary parcel diseases.⁵ This has resulted in increased mental distress, suffering, and a significant financial burden on society. Data acquired in Nosocomial Infections Surveillance carried out in the United States suggested that urinary tract infections (UTIs) are the most common HAIs which were followed by pneumonia and bloodstream infections. Approximately 95% of UTIs are associated with catheter and urinary stents, and around 87% of bacteremia or blood infections are caused by implantable cardioverter defibrillators, intravascular devices, pacemakers, and prosthetic vascular grafts; moreover, approximately 86% of pneumonia was caused by ventilation.⁶ Among the pool of pathogenic microorganisms, Staphylococcus aureus (MRSA) and Escherichia coli are the major causative agents of HAIs, particularly in immunocompromised patients (like those with cancer, HIV, or an autoimmune disease), which are facilitated by implant-related systemic infections and are the main causes of mortality.⁷ Evaluation reports suggest that the prevalent pathogens in patients with stent-associated respiratory infection include 50% of S. aureus, 35.7% of P. aeruginosa, and 14.3% of Candida albicans. On the other hand, for cardiac implants like permanent pacemakers and defibrillators, the most frequent pathogens involved include methicillin-resistant S. aureus (MRSA), Pseudomonas sp., E. coli, Klebsiella sp., and S. epidermidis.⁸

With the progression of science and technology, microorganisms have also developed new complex mechanisms of interactions with several implant surfaces and extensive colonization.⁹ A majority of host-associated pathogenic microorganisms reside and form a highly ordered, complex, and spatially structured network called a biofilm.¹⁰ Biofilms are bacterial populations packed within an extracellular matrix (ECM) comprised of polymers secreted by those bacteria such as exopolysaccharides (EPSs), proteins, extracellular DNA, and certain quorum-sensing proteins required for cell-to-cell communication. The generated complex exopolysaccharide layer in the biofilm shields the residing microbes from the outside assault of a host-resistant framework and antitoxins. Microbes which are involved in biofilm formation and implant-associated infections include Staphylococcus sp., Streptococcus sp., Escherichia coli, Pseudomonas sp., Corynebacterium sp., Propionibacterium acnes, and Klebsiella pneumoniae, some fungi including Candida sp. and Mycobacterium sp.⁸ Biofilm development is a complex sequential cycle with essentially three stages: the first stage begins with attachment to a biotic or abiotic surface, in which planktonic bacterium (free-gliding bacterium) adhere to the surface and proliferate to form microcolonies; the second stage is accumulation or maturation, in which microbes in microcolonies produce an extracellular matrix that serves as a scaffold to establish a three-dimensional (3D) structure; and finally, the third stage of detachment or dispersal occurs when the microbial cells are released and dispersed upon ECM degradation after reaching a desired cell density (Figure 1).¹¹

Figure 1.

Schematic representation of biofilm formation. Created with BioRender.com.

The characteristics of the implant surface that are important for the initiation of biofilm formation by these microorganisms are roughness, hydrophobicity, charge, and physio-chemical properties. Furthermore, when compared to the free-floating planktonic form, biofilm on medical devices or implants is more resistant to antibiotics and host defense systems.¹² A few examinations showed that following good clinical practices can decrease embed-related clinically procured diseases by as much as 50–70%.¹³ However, as a result of the distressing conditions within the biofilm, the metabolic pathway of the microorganism changes, resulting in changes at the multicellular level. This reduces the antimicrobial activity of antibiotics, further protecting the biofilms.¹⁴ Bacteria can develop resistance to various broad-spectrum antibiotics through a variety of mechanisms, including limited antibiotic diffusion, enzyme neutralization, slow cell growth, efflux pumps, and so on.¹⁴

To combat the problem of antibiotic resistance in biofilm-forming microbes, nanomaterials have emerged as one of the most effective means of eradicating drug-resistant biofilm-associated nosocomial infections of implants, medical devices, and other biomaterials. These are materials with one or more dimensions in the nanoscale range of 1–100 nm due to which they exhibit unique physiochemical properties compared to bulk materials. Nanoparticles have a high surface-to-volume ratio, which contributes to their chemical reactivity and biological activities, and their small size allows them to easily penetrate microbial cell membranes and the biofilm EPS layer, resulting in irreversible cell damage and, eventually, cell death.¹⁵⁻¹⁷ Therefore, nanocoating materials are used to modify the surface properties of intrinsic medical devices/implants in order to prevent biofilm infections. To target the first step of biofilm formation, namely, the attachment of planktonic bacteria to implant surfaces, one approach is to make the surface resistant to biofilm formation and to obstruct the protein adsorption process. This can be accomplished by coating the surface with antibacterial nanomaterials that prevent bacterial adhesion and have antimicrobial or antifouling properties that kill microbes that come into contact with the device or implant surface.¹⁸

Herein, we present a comprehensive overview of the various types of nanomaterials used as antimicrobial nanocoatings on biomedical implants. A detailed survey is provided for metal and metal-oxide-nanoparticle-based coating; 2D nanomaterial-based antimicrobial coating; and hybrid nanocomposite structure for clinical coating. Furthermore, the biocompatibility and toxicity of nanomaterials are discussed in order to qualify a nanomaterial for use as a nanocoating agent for biomedical implants.

2. Metal and Metal-Oxide-Nanoparticle-Based Antimicrobial Coating

Metal-based nanoparticles are the most common inorganic nanoparticles and represent a promising solution to antibiotic resistance. Metal nanoparticles of various types have been shown to be effective antimicrobial agents. Silver (Ag), gold (Au), copper (Cu), titanium (Ti), nickel (Ni), magnesium (Mg), zinc (Zn), and their oxide-based nanoparticles are the most commonly used antimicrobial nanoagents.¹⁹ Due to their dimensions being smaller than bacteria and their large surface-to-volume ratio, they provide strong, targeted, and prolonged antimicrobial activity and antibiofilm interaction.^19,20 Not only do they employ mechanisms of action that differ significantly from those described for traditional antibiotics but also they target multiple biomolecules, limiting the development of resistant strains. The three major mechanisms of action include interactions with a phospholipid bilayer causing membrane damage; binding to cytosolic proteins and inhibiting the metabolic pathways; and the generation of free radicals and reactive oxygen species (ROS) for inducing damage to biomacromolecules (Figure 2).²⁰ Thus, metal and metal-oxide-based nanoparticle coatings of implant and medical device surfaces for antimicrobial and antibiofilm applications are discussed in detail below.

Figure 2.

Schematic representation of an (a) implant surface with biofilm depicting several HAIs and an (b) implant surface with surface-modified nanocoatings of metal and metal oxides and 2D material nanoparticles with their antibacterial/antibiofilm mechanisms of action. Created with BioRender.com.

2.1. Silver and Silver-Oxide-Based Nanomaterial Coating

Silver has been used since ancient times, and its nanoform has gained a lot of attention for decades for having antimicrobial properties. The antibacterial properties of silver have been known and explored since the ancient civilizations of the Greeks, Egyptians, and Romans.²¹ The medicinal use of silver originally started when Credé first published in the year 1881 that silver can be used to treat and cure eye infections in newborn children in Germany.²² Later in 1933, Gorden et al. showed the antimicrobial activity of silver on dental infection and root channels in the form of a metallic deposition in the infected area.²³ However, it has been reported that metallic-silver-based coating is less efficient, has high cytotoxicity, deteriorates quickly, and lacks sufficient antimicrobial activity. Furthermore, for metallic silver to act as an antimicrobial agent, moisture in the immediate environment is required for the formation and release of silver ions, which act as a potent antimicrobial agent.²⁴ As a result, silver nanoparticles (AgNPs) were preferred as nanocoating over metallic silver due to their ability for prolonged release of silver ions; large surface-to-volume ratio; low susceptibility to sequestration by other ions such as chloride, phosphate, or other cellular components; low risk of bacterial resistance development; and high antifouling and antibiofilm potential.^21,24,25 They have been identified as one of the best candidate materials for medical device coatings to prevent HAI caused by the extensive pathogenic biofilm network formed on such devices. To act as an antimicrobial agent, silver nanoparticles must be present in their oxidized form (Ag⁺), which is readily favorable due to the presence of oxygen, body fluids, and moisture at tissue sites.²⁵ It has also been used in several commercially available products for wound treatment—wound dressings, bandages used for burns and other chronic wounds; implant tubes and catheters; silver and silver nanoparticles as coatings; and other implantable medical devices.²⁵

Silver in ionic or nanoparticle forms has been seen to exhibit a broad spectrum of antimicrobial activity against biofilm-forming pathogenic microorganisms such as P. aeruginosa, S. epidermidis, MRSA, and methicillin-resistant S. epidermidis (MRSE) strains and several other microbial colonizations that are associated with biomedical infections.¹⁵ The mechanism of antimicrobial activity of AgNPs is well established, demonstrating that the mode of action of AgNPs is based on the (1) inactivation of DNA replication, (2) cellular protein denaturation, (3) damage to the cell membrane, and (4) generation of free radicals including ROS derived from the surface of nanoparticles which damage biomacromolecules.^26,27

The use of AgNPs as an antimicrobial coating material is a common practice to reduce HAIs and implant-induced infections. AgNP-based coating materials on medical implants and catheters have been highly explored on substrates such as glass-based, fabricated, plastic, and polyurethane (synthetic polymer for medical implants) catheters for their effectiveness toward antibacterial and antibiofilm applications. Agnihotri et al. explored the disinfectant properties of immobilized AgNPs on a glass substrate which displayed complete disinfection against both E. coli and B. subtilis, concluding its applicability as a coating material for surgical devices and medical implants.²⁸ A distinctive amount of research is carried out on the AgNP-fabricated catheters and their role in the prevention of CAUTI or catheter-related bloodstream infections (CBSIs). The formation of a biofilm or surface thrombus, on the catheter substrates, starts to adsorb proteins upon contact with the blood, which subsequently becomes the breeding place for microorganisms.^29,30 In 2007, Roe and colleagues developed AgNP-implanted plastic catheters that showed a significant reduction in the infection rate. In their report, they described antimicrobial properties of AgNPs that prevented biofilm formation against Escherichia coli, Enterococcus, Staphylococcus aureus, coagulate-negative Staphylococci, Pseudomonas aeruginosa, and Candida albicans. Similarly, another study done by them using AgNP–polyurethane catheters, over a 10 day experimental period, revealed good antimicrobial activity. However, interestingly, the tested animals lost an average of 8% of their body weight, which may be caused by silver toxicity from Ag-coated catheters.³¹

AgNP-based coating applications are also studied on catheters implanted for the vascular system and for urinary tract infections as well. AgNP-coated urinary catheters are more beneficial than normal catheters; moreover, the CAUTI was also much lower after introduction to the silver nanocoating, and the absolute risk of reduction values of catheter-associated bacteria ranged from 0.5% to 32%.^32,33 Seymour stated that the risk rate of CAUTI was higher (at 11.1%) than the standard (at 7.3%) during the control period, and the rate was decreased when the silver-alloy-coated Foley catheters were used which upon evaluation came out to be 3.2%, implicating that the silver-coated catheters are efficient in reducing UTIs by up to 20%.³⁴

Pollini and co-workers, in their study of hemodialysis catheters, showed that the use of polyurethane catheters for vascular access and filtration of blood causes infections which often lead to the death of patients, but when AgNPs were photochemically deposited at the site of infection, they caused inhibition of bacterial growth. They demonstrated that the developed surface coating can result in the manufacture of safe catheters with low infection rates that are also cost-effective.³⁵ Similarly, in the case of external ventricular drain (EVD) catheters used in neurosurgery for patients with acute hydrocephalus (a condition in which cerebrospinal fluid (CSF) accumulates in the brain), there is a high risk of microorganisms infecting the catheter surface and causing subsequent infections.³⁶ Fichtner et al. performed an extensive study on patients with and without silver-coated EVDs and concluded that silver-bearing catheters can act as a local infection barrier, which may be a safe and effective way to reduce the incidence of CSF infections.³⁶

Another important medical implant is the use of prosthetic joints. In modern times, there is an increase in the rate of orthopedic surgery, and complications related to prosthetic joints have increased over time, which eventually leads to morbidity and even death.³⁷ Thukkaram and colleagues investigated implant-associated infections in 2020 and developed a new solution: they coated titanium implants with an amorphous hydrocarbon (a-C:H) matrix containing AgNP nanocomposites.³⁸ The in vitro antimicrobial assay demonstrated good antibacterial activity against E. coli and S. aureus. Furthermore, it was biocompatible and improved osteoblast adhesion and proliferation. Because of its excellent properties, this coating has emerged as a viable option for orthopedic implants.³⁸ Thus, AgNPs as a coating on biomedical implants have a great potential and a promising future for controlling biofilm-associated HAIs.

2.2. Copper and Copper-Oxide-Based Antimicrobial Coating

Copper metal has been employed for its antibacterial properties since ancient times, just like silver. It was used to sheath ships during the Phoenician era, and vessels made of copper metal were used to sanitize drinking water as well as to treat chest wounds, infections, fresh wounds, scalds, burns, intestinal worms, ear infections, and itching.³⁹ As a result of its antibacterial characteristics, copper is frequently utilized on various surfaces, and it is claimed that this kills germs through a process known as “contact killing” (i.e., by contact-dependent inhibition of planktonic bacteria).⁴⁰ Because of its propensity to stick close to sulfhydryl groups, copper can interact with the thiols found in proteins. This renders copper and copper compounds their bactericidal property. However, it has been observed that bacteria have evolved a wide range of mechanisms to protect themselves from the toxic effects of metallic copper ions by sequestration of ions extracellularly, reducing the cell membrane permeability of copper ions, the active outflux of ions from the cell, and scavenging of ions by metallothionein-like scavenging proteins in the cell cytoplasm.⁴⁰ Copper and copper oxide nanoparticles have thus become increasingly popular because they are biocompatible, less expensive than other metals like silver and gold, more easily synthesized using environmentally friendly methods, dissolve more quickly than other metallic nanoparticles, have comparable antibacterial effects, and have a huge potential for surface functionalization to boost their primary antimicrobial activity.⁴¹ Further, the antibacterial activity of the oxides of copper, cuprous oxide (Cu₂O) and cupric oxide (CuO), is based on their semiconducting property, which has the capability of generating ROS.⁴² The following are the possible mechanisms by which Cu and CuO NPs exert their antibacterial effects: (1) interaction with −SH groups leading to protein denaturation; (2) binding to DNA molecules and creating cross-links between the strands; (3) adhesion to the cell membrane by electrostatic interaction causing damage and increasing cell permeability; (4) copper ions released interrupting several biochemical processes; and (5) causes oxidative stress by free radicals and ROS generation.^41,43 Due to their high capacity for absorption, adsorption, penetration, and availability in the aqueous biofilm environment, Cu and CuO NPs are also reported as crucial antibiofilm agents. In order to avoid nosocomial infections caused by bacterial contamination, they have therefore been utilized as antimicrobial coating agents on the surfaces of implants and medical devices.⁴⁴

Thus, use of Cu- and CuO-based nanocoatings on implant surfaces is explored by researchers to prevent HAIs and implant-associated infections. Li et al. developed a surface coating material based on mussel-inspired dendritic polyglycerol (MI-dPG) and integrated it with CuNPs (Figure 3). This cocktail matrix coating was studied on E. coli and S. aureus through an “attract–kill–release” strategy where the synergistic effect of contact-mediated killing and generation of ROS led to bacterial inactivation (Figure 3(I)).⁴⁵

Figure 3.

(I) Antibacterial potential of CuNP-incorporated MI-dPG surface coatings. (i) Biofilm observation by confocal laser microscopy via live/dead staining (green: viable bacteria, red: dead bacteria): MI-dPG coating [(a) live bacteria, (b) dead bacteria, (c) merged image of the live and dead bacteria, and (d) merged image of bacteria and coating], CuNP-incorporated MI-dPG coating [(e) live bacteria, (f) dead bacteria, (g) merged image of the live and dead bacteria, and (h) merged image of bacteria and coating]. Scale bar: 50 μm. (ii) (a) Antibacterial ability of different samples against S. aureus, kanamycin-resistant E. coli: CuNP-incorporated MI-dPG coating, MI-dPG coating, glass slides, and blank control (without materials). (b) In vitro cytotoxicity of the CuNP-incorporated MI-dPG coating leachate determined by the MTT assay against NIH/3T3 cells after 24 h incubation. (c) Typical photographs of the agar plate testing results: CuNP-incorporated MI-dPG coating, MI-dPG coating, glass slides, and blank control (without materials). Data are presented as mean ± SD, n = 5. Statistically significant differences at the same period are indicated by **p < 0.01 (S. aureus) or ^##p < 0.01 (kanamycin-resistant E. coli) compared with the blank control. Photograph courtesy of Mingjun Li et al. reproduced with permission from ref (45). Copyright 2017 American Chemical Society. (II) Antibiofilm properties of CuONP-coated teeth that restrict S. mutans biofilm formation. (i) Coated and uncoated teeth were incubated with S. mutans for 24 h. (A) The biofilm biomass that developed was stained by a crystal-violet staining as described in the experimental section. (B) Quantification of biofilm biomass. (ii) HR-SEM imaging of S. mutans biofilms on coated and uncoated teeth. Biofilms were grown for 24 h at 37 °C. Photograph courtesy of Michal Eshed et al. reproduced with permission from ref (46). Copyright 2012 American Chemical Society.

Interestingly, self-defending bone implants with intrinsic antibacterial properties have been demonstrated by Hengel and co-workers. For the study they used CuNPs as coating material along with Ag in varying ratios on the surface of a biofunctionalized TiO₂ layer with plasma electrolytic oxidation (PEO) of the implant surface and checked their antibacterial activity on the MRSA bacterial strain. They concluded that PEO-treated Ag (PT-Ag) and PEO-treated AgCu (PT-AgCu) implants with Ag and Cu ratios up to 75% and 25% completely eliminated the nonadherent planktonic and adherent bacteria both in vivo and ex vivo within 24 h with no cell cytotoxicity in the preosteoblastic cell line.⁴⁷ Furthermore, it was reported by Yoon et al. that CuNPs showed better antibacterial effects than AgNPs on pathogenic strains like E. coli, S. aureus, and Bacillus subtilis,⁴⁸ and the MIC, MBC, and disk diffusion test results demonstrated by Ruparelia et al. suggested that they have greater affinity to surface-active groups of bacteria like B. subtilis because they shows better bactericidal effect.⁴⁹

Polyurethane (PU) is used in various biomedical fields mostly in catheters, heart valves, and dental fillers, and from the study of Hoffman-Kim et al. it is established that the rough pattern surface generates topology for better cell adhesion.⁵⁰ Ahmad et al. made PU-CuO-coated biocompatible biomaterials, which are porous, are elastomeric, and also possess antimicrobial property, especially in the case of an epidemic methicillin-resistant strain of Staphylococcus aureus (EMRSA). CuO nanoparticles in the PU fibrous matrix have a significant impact on the level of MRSA inhibition, and the properties depend on the pore size and film thickness. These coatings are used in dental applications and catheters.⁵¹

For dental caries (the formation of biofilm on the teeth’s surface by most prominently Streptococcus mutans) to protect the teeth and make the antifouling coating, CuO NPs are widely used. These coatings used in dental protection either kill or prevent bacterial adhesion, thus preventing biofilm formation (Figure 3(II)).⁴⁶ Contact lenses are one of the important modern-day medical implants where bacterial biofilm formation increases the infection, and wearing lenses is the largest single risk factor for microbial keratitis.⁵² CuO is also used in combination with other metals for increased antimicrobial activity and have industrial application too. Nahum et al. used a conjugate nanoparticle of Zn-CuO and coated it in PureVision balafilcon—soft contact lens via sonochemical deposition using a high-intensity ultrasonic horn. It was tested against pathogenic strains S. epidermidis and P. aeruginosa that are involved in contact-lens-related infectious keratitis, and the antibacterial mechanism of action was based on ROS generation for bacterial membrane damage.⁵³

2.3. Zinc and Zinc-Oxide-Based Antimicrobial Coating

It is established that zinc is an essential trace element for cellular growth, development, DNA synthesis, enzymatic activity, and most importantly biomineralization in organisms.⁵⁴ Zinc is most commonly used as an antimicrobial agent in oral hygiene products such as in mouth rinses and toothpastes to prevent dental plaque, inhibit calculus formation, and reduce halitosis. Zinc nanoparticles (ZnNPs) are used for inhibition of S. mutans, the main cariogenic bacteria which is involved dental caries.⁵⁵ However, zinc oxide nanoparticles (ZnO NPs) are used more commonly than zinc nanoparticles because of their superior antibacterial properties.⁵⁵ ZnO nanoparticles are one of many metal oxides utilized for antimicrobial applications, but they have drawn much more attention due to their distinctive and intriguing features. These distinctive characteristics include a high surface-to-volume ratio, UV radiation absorption, transparency for visible light, electrical conductivity, piezoelectric properties, semiconductivity, nontoxicity to mammalian cells, widespread availability, cost effectiveness, and prolonged environmental stability.^56,57 Zinc gets absorbed excessively by the cells compared to copper and iron ions as Zn²⁺ acts as a strong Lewis acid, making it highly corrosive in nature and therefore preventing copper and iron from being absorbed by the cell.⁵⁸ ZnO NPs have been shown to be effective against both Gram-positive strains such as S. aureus, S. epidermidis, B. subtilis, and Enterococcus faecalis and Gram-negative strains such as E. coli, P. aeruginosa, and Campylobacter jejuni. However, they exhibit greater sensitivity and susceptibility to Gram-positive bacteria compared to Gram-negative ones.²⁰ Studies have also demonstrated that photoactivated ZnO NPs were effective against Listeria, E. coli, B. subtilis, and Lactobacillus helveticus.⁵⁹⁻⁶¹ Overall, ZnO NPs exhibit antimicrobial activity through three main mechanisms: cell membrane damage brought on by electrostatic interactions of nanoparticles with membranes, release of Zn ions that alter the bacterial microenvironment, and induction of oxidative stress brought on by the formation of free radicals and ROS.⁵⁷

ZnO NPs are currently used to functionalize surfaces to reduce biofilm formation on medical implants such as orthopedic, dental, and metallic and urinary catheters. Tejeda et al. suggested that ZnO particles inhibit the aggregation of cells on the surface, and the released Zn²⁺ ions generate ROS, thus preventing complex biofilm structure formation.⁶² Titanium implants are widely used for orthopedic surgeries, oral implantation, and other medical devices due to their noncorrosive nature and high mechanical strength; however, there are two major issues, implant-associated infection and low osteogenesis rates. ZnO NPs were demonstrated to be used as a surface modification material on such implants to function as antimicrobial-, anticorrosive-, and osteogenesis-promoting coatings.⁵⁴ Similarly, Schwartz et al. demonstrated the use of ZnO NPs to test their antimicrobial activity when doped on medical implants. The ZnO-doped implant revealed high antibacterial activity against S. aureus with about 37% higher antimicrobial effect than nondoped implant samples.⁶³ Memarzadeh et al. demonstrated the use of ZnO NPs as a coating- and osteogenesis-promoting material. Their results indicated that a 100% ZnO composite coated substrate showed significant antimicrobial activity against S. aureus and also assisted in promoting osteogenesis as ZnO immobilized on the substrate provided osteoblast cells with the ability to adhere, grow, and become metabolically active for proliferation. This evidence implicated that ZnO-based coating material on the implant surface could be used as a promising and potential agent for future medical device applications particularly for bone and dental implants.⁶⁴ Mcguffie et al. demonstrated a layer by layer anti-infective coating of ZnO NPs on the surface of implantable medical devices. They concluded that all three ZnO NP shapes—plate, sphere, and pyramid—caused a dose-dependent reduction in Gram-positive bacteria growth by causing cell membrane disruption and ROS generation, but Gram-negative strains required a higher dose of ZnO NPs to inhibit growth. Furthermore, the layer by layer ZnO NP coating increased the surface roughness (or bacterial surface hydrophobicity) of the implant substrate and had high stability to ZnO NP leaching over a 7-day period (Figure 5).⁶⁵ Jansson et al. investigated an alternative antibacterial coating approach on implants by using a ZnO nanorod surface coating that helps reduce bacterial cell adhesion and viability, thereby decreasing the incidences of implant-associated infections. They compared the effect of such ZnO nanorod surface coatings on common implant-associated pathogens P. aeruginosa and S. epidermidis using sputured ZnO substrates and glass substrates as controls. The study concluded that the ZnO nanorod substrate had a significant antibacterial effect on the P. aeruginosa strain and a greater antibacterial effect on S. epidermidis when compared to sputtered ZnO nanoparticles. As a result, it has a high potential for use as a surface coating on implantable medical devices to prevent bacterial adhesion and to demonstrate bactericidal activity.⁶⁶

Figure 5.

(I) Layer-by-layer coatings of ZnO nanoparticles inhibit Staphylococcal growth. (i) SEM micrographs of (A–C) bare polystyrene pegs, pegs coated in ZnO (D–F) spheres, (G–I) plates, and (J–L) pyramids cultured with (A, D, G, and J) E. coli, (B, E, H, and K) S. aureus, and (C, F, I, and L) S. epidermidis. Scale bars at the bottom of each image are equal to 10 μm. (ii) Growth curves for E. coli, K. pneumonia, S. aureus, and S. epidermidis in the presence of increasing concentration of ZnO NPs synthesized as pyramids, spheres, and plates. Photograph courtesy of Matthew J. McGuffie et al. Reproduced with permission from ref (65). Copyright 2016 Elsevier. (II) Antibacterial activity of nano-Mg(OH)₂. (i) (a) TEM and (b) SEM images of E. coli treated with 0.5 mg/mL Mg(OH)₂ slurries for 4 h, respectively. Inset images of b show the EDS analysis of bacteria. The size of all SEM images is 6.0 μm. (ii) Relationship between the antibacterial efficiency and the Mg(OH)₂ species. E. coli was exposed for 4 h to 0.1, 0.3, and 0.5 mg/mL of different types of Mg(OH)₂. Comparison with a positive control (bacteria in ddH₂O at pH ∼ 10 without nanoparticles) permitted to determine the percentage of antibacterial efficiency. Data presented as mean ± standard deviation (n = 3). Photograph courtesy of Xiaohong Pan et al. Reproduced with permission from ref (92). Copyright 2013 American Chemical Society.

2.4. Titanium- and Titanium-Oxide-Based Antimicrobial Coating

Titanium and titanium dioxides are now extensively used in various biomedical implants (mostly orthopedic and dental implants), and different studies have demonstrated their antibacterial and antifungal properties.⁶⁷ Titanium oxide is a nontoxic and inert metal oxide with self-disinfecting properties and is used for disinfection of drinking water and in packaging of food, in drugs, in cosmetics, and for orthodontic cosmetics.⁶⁸ It is a semiconductor transition metal oxide with several advantages over other antimicrobial metal oxides, including corrosion resistance, biocompatibility, high mechanical strength, low cost, large surface area, nontoxic nature, excellent surface morphological properties, and, most importantly, high photocatalytic properties with high oxidation ability and long-term stability.⁶⁹ This has led to an increased demand for titanium-coated implants. TiO₂ nanoparticles are used as an antimicrobial agent for a broad spectrum of pathogens. Some of the pathogens targeted by TiO₂ include MRSA, S. aureus, S. epidermidis, P. aeruginosa, Clostridium difficile, Klebsiella pneumoniae, E. coli, and Mycobacterium tuberculosis strains. The inhibitory properties of TiO₂ nanoparticles are based on the generation of ROS when activated by UV irradiation (wavelength less than 385 nm), solar irradiation, and absorption of the high-energy photons.⁷⁰ ROS are produced as a result of a redox reaction between electron acceptors and water and atmospheric oxygen molecules. This photocatalytic reaction produces free radicals such as hydroxyl radicals (OH^•) and superoxide anions (•O^2–), which cause cell damage and death (Figure 4).⁷¹ The ROS produced in the microbial cells leads to lipid peroxidation, oxidation of metabolic enzymes, DNA damage, and ultimately reduction of the activity of respiratory activity which leads to cell death.⁶⁹

Figure 4.

Schematic representing the antibacterial mechanism of action. (a) Titanium oxide nanoparticles by photoactivation leading to ROS production to kill bacterial cells. (b) Supermagnetic iron oxide nanoparticles (SPIONs) by inducing an external magnetic field leading to biofilm ECM disruption and ROS generation. Created with BioRender.com.

Interestingly, Liu et al. showed that the UV light excitation effect of TiO₂ can be maintained when submerged in the simulated body fluids.⁷² In dentistry, titanium is a widely used biomedical implant material as it is an integral part of dental therapy and has been an old practice since the 1960s.⁷³ Formation of the oxide layer on top of the Ti layer infers it with significantly higher biological activity than bare Ti because oxides contain surface hydroxyl groups on both the terminal OH and bridge OH in equal proportions that react not only with water molecules in an aqueous environment but also with moisture in the air.⁷⁴ A study conducted by Doran et al. on the neoplastic transformation of cells on orthopedic implants established that there are no effects of Ti ions or Ti particles on cells, and thus it is not lethal or carcinogenic to the cell.⁷⁵ Cranio-facial TiO₂ implants are now the commercially most used implant, lowering the cost and decreasing the chance of infection.⁷⁶ Liu et al. suggested that the antibacterial efficiency of titanium is increased when it is conjugated with AgNPs.⁷² Furthermore, Van et al. demonstrated that at a concentration of 20 ppm the antimicrobial activity of AgNPs loaded onto TiO₂ against S. aureus was 99.9% efficient in 60 min, whereas only TiO₂ can kill 95.9% S. aureus in 60 min.⁷⁷

2.5. Iron- and Iron-Oxide-Based Antimicrobial Coating

Iron (Fe) and iron oxide (Fe₃O₄, Fe₂O₃) magnetic nanoparticles are used in several therapeutic applications, and research has demonstrated that they have potent antibacterial properties as well.⁷⁸ Supermagnetic iron oxide nanoparticles (SPIONs) are known to be biocompatible, biodegradable, and nontoxic despite the fact that the pure forms of these metals are highly toxic, sensitive to the cell, and damaging.⁷⁹ SPIONs are effectively eliminated by the body through a variety of iron metabolism pathways.⁷⁹ These are shown to inhibit the growth of most common biofilm-associated pathogens like S. aureus, MRSA, E. coli, P. aeruginosa, S. epidermidis, K. pneumoniae, and B. subtilis. According to reports, the main mechanisms of action involve the destruction of cell membranes as a result of electrostatic interactions and the induction of oxidative stress in the cells through the production of free radicals and ROS (single oxygen, superoxide radicals, hydroxyl radicals, and hydrogen peroxide), which lead to the destruction of cellular proteins and DNA.⁸⁰ Additionally to these effects, the magnetic properties of SPIONs can be used to produce localized hyperthermia, which can physically shatter and disperse pathogen biofilm through static friction produced by an external magnetic field (Figure 4).⁸¹ Studies have discussed the use of these particles as a coating material for biomedical devices and suggest that SPIONs can be functionalized with various polymers and biomaterials, such as dextran, polyethylene glycol (PEG), and poly(vinyl alcohol) (PVA), or can even be attached with various functional groups, such as thiols, carboxyls, and amines, to cover a wide range of applications including antimicrobial action.⁸² According to Thukkaram et al., a combination of nonadhesive coatings, such as polymer brush and iron oxide nanoparticles, can significantly reduce the formation of biofilm by S. aureus, E. coli, and P. aeruginosa. It belongs to a particular family of metal oxides that can be created with a high surface to volume ratio with an unusual crystalline structure having increased number of edges and corners, which might cause a rise in the generation of ROS.^80,83 According to a study by Jie Li and colleagues, magnetic iron oxide nanoparticles, combined with AC and DC magnetic fields, can disrupt the biofilm. This combination works by inducing magnetic hyperthermia, and it also causes detachment of the matrix by applying pure brute force to magnetic nanoparticles by pulling them over the surface of the biofilm. They claimed that due to their capacity to effectively detach and produce local mechanical damage to the biofilm matrix, bigger nanoparticles and DC magnetic fields exhibited the highest efficacy of reduction in biofilm formation when compared to AC fields or direct contact nanoparticles.⁸⁴ According to reports published by Taylor et al., 56% of medical device related infections are caused by multidrug-resistant S. aureus (MRSA), and for it a superparamagnetic iron oxide nanoparticle (SPION) can be used as a highly comprehensive tool for antibacterial treatment, biofilm disruption, and site-specific targeting on such infection sites. SPIONs coated with dimercaptosuccinic acid (DMSA) have been shown to have antibacterial and antibiofilm activity by either adhering to bacterial surfaces to prevent the formation of biofilms or by penetrating bacterial cells to cause the intracellular release of metal ions, which inhibits bacterial growth.⁸⁵

2.6. Magnesium-Hydroxide-Based Antimicrobial Coating

Magnesium hydroxide (Mg(OH)₂) is becoming more significant in this era of green chemistry due to its environmental friendliness, broad antibacterial spectrum, low toxicity, biocompatibility, and low manufacturing cost.⁸⁶ It is reported to be the ideal biodegradable material which when used for orthopedic applications can easily get corroded without the generation of toxic end products.⁸⁷ It has been demonstrated that magnesium hydroxide NPs can impede the formation of biofilms by attaching to surfaces and diffusing into cells, where they raise lipid peroxidation rates, disrupt membrane potential, and bind to genomic DNA.⁸⁸ In addition to these, it blocks ATP synthase, prevents cell respiration, and prevents protein synthesis in the cells, thus inactivating pathogens.⁸⁷ Several reports have demonstrated that Mg(OH)₂ NPs are effective against E. coli, S. aureus, P. aeruginosa, K. pneumoniae, and B. phytofirmans in a size-dependent order as smaller particles can easily penetrate and relocate themselves between the cell compartments.^89,90 Dong and co-workers in their study checked the activity of magnesium hydroxide nanoparticles on Burkholderia phytofirmans and Escherichia coli and stated that the metal ions of this nanoparticle act on the bacterial cell through release of OH^– and Mg²⁺ ions and also established that pH plays an important role in killing the bacteria as adsorption of water moisture on the surface of the Mg(OH)₂ layer forms a thin highly alkaline water meniscus around the particles by a process of capillary condensation. When the nanoparticles are in contact with the bacteria this meniscus containing highly concentrated OH^– groups causes membrane damage, leading to cell death.⁹¹ In their study, Pan et al. synthesized nanoparticles using a variety of precursor molecules, including MgCl₂, MgSO₄, and MgO, to test the relative activity of those particles. Their findings suggested that (i) antimicrobial activity depends on particle size and (ii) lower toxicity of these particles is based on their electrostatic interaction, implying that the precursor and their hydrolysis are crucial at the time of synthesis. (iii) The mechanism of inhibitory action is based on the interaction of metal ions with the cell wall, which damages the integrity of the membrane and increases its permeability, leading to bacterial cell death (Figure 5).⁹² Another work by Halbus et al. used C. reinhardtii, S. cerevisiae, and E. coli as model organisms and examined the antibacterial activity of Mg(OH)₂ NPs synthesized using the direct precipitation approach. The antimicrobial coating had a favorable influence since Mg(OH)₂ NPs were effective against all of these organisms and produced very little toxicity and are safe for human use.⁹⁰ Meng et al. showed that magnesium hydroxide nanoparticles have potent antibacterial activity against Streptococcus mutans, a biofilm-forming bacteria for dental implants. In their report, they concluded that Mg(OH)₂ NPs cause bacterial cell death by attaching to cell wall proteins, increasing cell membrane permeability and causing leakage of intracellular components.⁹³

3. 2D-Nanomaterial-Based Nanocoating

In recent decades, the major problem associated with metal and metal oxide nanoparticles is that bacteria become resistant to it. Hence to be effective huge concentrations of nanoparticles are required which is again toxic to animal cells. To overcome the drawbacks related to metal and metal oxide nanoparticles, 2D nanomaterials are introduced. A variety of different 2D nanomaterials like graphene oxide, black phosphorus, MoS₂ (molybdenum disulfide), and boron nitride are widely used in different antimicrobial coating agents.

3.1. Graphene- and Graphene-Oxide-Based Coating

Graphene is a naturally occurring honeycomb lattice-like two-dimensional nanomaterial consisting of a monolayer of sp²-bonded carbon atoms which was first isolated in the year 2004.^94,95 Because of its magnificent thermal, electrical,⁹⁶ and mechanical properties,⁹⁷ graphene has gained immense importance among both academic and industrial researchers.^98,99 Graphene oxide is a cognate of graphene, composed of sp³-bonded carbon atoms connected via oxygen functional groups in the planes and edges of the sheets.¹⁰⁰ Graphene oxide because of its excellent biocompatibility, mechanical strength, high specific surface area, flexibility, electronic properties, excellent conductivity, good dispersion, astonishing thermal stability, and easy-to-fabricate nature has become one of the important materials in biomedical implants.^101,102 The hydroxyl groups present in the graphene sheet structure are responsible for its amphipathic nature. Furthermore, these groups also allow graphene to interact more with hydrogen and hydrophilic groups.^103,104 More importantly, graphene nanosheets due to their excellent properties act as a great base material for the attachment of different small molecules, insertion of functional groups, antimicrobial substances, polymers, or even other nanomaterials. The interaction between these molecules and graphene nanosheets is directed through strong intermolecular forces.¹⁰⁵ Another major property that has made graphene a material of choice for biomedical implant coatings is its innate antimicrobial properties¹⁰⁶ and lower cytotoxicity¹⁰⁷ toward eukaryotic cells. Studies showed that the bactericidal or bacteriostatic action of GO is based on the destruction of the cell wall (Figure 6). The sharp edges of GO damage the membrane, including the leakage of intracellular materials in the environment. This inhabitation mechanism causes membrane stress, thus leading to fatal death of the bacteria.¹⁰⁸ Another mechanism of inactivation of bacteria is related to light absorption and the oxidation of organic materials via generating ROS.¹⁰⁶ Thus, scientists around the world have developed plenty of implants using different forms of graphene on different materials. Shao et al. and his group have reported that surface free energy of the metal plates was modified when polyethylenimine (PEI) grafted graphene oxide (GO) nanosheets were incorporated into Ni–P coatings by an electroless plating technique, which has a significant effect on bacterial adhesion onto the surface.¹⁰⁹ Gelatin-functionalized graphene oxide (GOGel) was used to develop surface coatings on nitinol substrates where it showed antibacterial properties against E. coli, whereas it promoted mammalian cell growth.¹¹⁰ Another study has shown that GO and rGO exhibit antimicrobial activity against E. coli when surface coated on aluminum plates.¹¹¹ Ruibin Li et al. and co-workers have developed a hydrated graphene oxide coated catheter which showed exceptional antimicrobial activity depending on the density of carbon radicals.¹¹² Besides this, graphene oxide plays a huge role in dental implants. A recent study revealed that graphene oxide coatings on dental implants not only reduce bacterial adhesions but also help in osteoblast activation.¹¹³

Figure 6.

Schematic representation of different types of 2D nanomaterials used for fabricating antimicrobial biomedical implants and their mechanism of action. Created with BioRender.com.

3.2. Molybdenum-Disulfide-Based Coating

Molybdenum disulfide is one of the examples of transition metal dichalcogenides (TMDs) where molybdenum and sulfur atoms can be arranged in various orientations. According to these orientations, MoS₂ has four different crystalline structures.¹¹⁴ Being a TMD, MoS₂ also forms a layered structure where individual layers are stacked together via van der Waals interactions. Among various other properties like high mechanical strength, elasticity, biocompatibility, and thermal conductivity,¹¹⁵ one of the interesting properties is that thin nanosheets of MoS₂ have a direct band gap of approximately 1.8 eV, whereas the bulk form of it has an indirect band gap of approximately 1.6 eV.¹¹⁶ All these properties have made MoS₂ nanomaterials a suitable material for various biomedical applications like bioimaging, biosensing, drug delivery, etc.^117,118 Besides all the above-mentioned properties, MoS₂ nanosheets also have antimicrobial activities which have made MoS₂ one of the appropriate materials for antimicrobial fabrics¹¹⁹ and biomedical coatings because of its interesting interacting nature with different materials.¹²⁰ Xi Yang et al. have reported that the high surface area to volume ratio and high conductivity mainly provide MoS₂ nanosheets with their antimicrobial activity. According to their report, because of the high surface-to-volume ratio and conductivity, they were able to cause membrane damage to bacterial cells and oxidative stress which can be both ROS dependent (superoxide anion) and ROS independent.¹²¹ A detailed study on the antimicrobial mechanisms of chemically exfoliated MoS₂ nanosheets also showed that the mechanisms behind the antimicrobial activity of MoS₂ nanosheets are a collaborative effect of membrane depolarization and loss of membrane integrity, and as a consequence, protein leakage, inhibition of metabolic activities, and oxidative stress¹²² occur (Figure 6). Shin et al. have used MoS₂ nanoflakes as antimicrobial surface coatings on titanium dental implants. The antimicrobial activity was checked against E. coli, and they have observed that MoS₂ nanoflakes were able to damage the bacterial cell membrane and produced a significantly high amount of ROS which was another reason for their antimicrobial activity. Besides this, because of the aqueous stability of MoS₂, the coated surface was also able to provide a hydrophilic surface for easy attachment of cells and resulted in cell proliferation¹²³ (Figure 7). Zhang et al. have used photothermal therapy against MoS₂-coated implants to provide excellent antimicrobial activity. They have used a titanium implant and coated it with molybdenum disulfide (MoS₂)/polydopamine (PDA)-arginine-glycine-aspartic acid (RGD), and by taking advantage of photothermal therapy, upon NIR irradiation this implant showed great antibacterial property through damaging the bacterial cell membrane and producing oxidative stress in bacteria although it was ROS independent.¹²⁴ Feng et al. have used the photodynamic and photocatalytic activity of MoS₂ nanosheets for antimicrobial activity while using both visible light and NIR light against E. coli and S. aureus. They have observed almost 91% bacterial death upon 10 min of visible-light exposure, whereas upon NIR exposure the bacterial death rate was around 60%; however, when both the lights were used it showed a significantly high antibacterial effect.¹²⁵ Another group used the enhanced photocatalytic and photodynamic effect of MoS₂ nanosheets for antimicrobial activity. They have conjugated MoS₂ nanosheets with N-doping carbon quantum dots (CQDs) which act as a mediator of photogenerated electrons produced by MoS₂ nanosheets to intensify the photocatalytic and photodynamic activity and result in almost 99% bacterial death, but parallelly it was biocompatible as well¹²⁶ (Figure 7). Zhang et al. also used both 660 nm visible light and 880 nm NIR light irradiation for antimicrobial activity of a titanium implant coated with biofunctionalized titanium dioxide–molybdenum disulfide–polydopamine–arginine-glycine-aspartic acid nanorod arrays. Here also visible light resulted in antibacterial activity by producing reactive oxygen species and NIR light by increasing the local temperature.¹²⁷ Further, Weidong et al. also checked the antimicrobial activity of MoS₂ by 660 nm visible-light application. They have coated the titanium implant with AgBr nanoparticles and 2D MoS₂ nanosheets where AgBr nanoparticles act as an acceptor of photoelectrons released from the MoS₂ nanosheet upon light irradiation, hence providing great photocatalytic activity. The mechanism behind the antibacterial property was again ROS production and Ag⁺ leaching¹²⁸ (Figure 7). A recent study has showed that a self-activating implant modified with hydroxyapatite (HA)/MoS₂ coating was able to kill Staphylococcus aureus and Escherichia coli infections and as well as promote bone regeneration by stimulating osteoblastic differentiation of mesenchymal stem cells by changing cell membrane and mitochondrial membrane potentials¹²⁹ (Figure 7). Another research group prepared an antimicrobial implant by coating titanium implants by silver-nanoparticle-loaded chitosan-modified MoS₂ nanosheets where the photocatalytic activity of MoS₂ nanosheets was exploited for the antimicrobial property.¹³⁰ Very recently, Ma et al. developed antimicrobial leather by coating carbon quantum dots and MoS₂ quantum dot loaded dendritic fibrous nanosilica which has showed 95–99% antimicrobial activity not only against a broad range of bacteria but also against fungus.¹³¹

Figure 7.

(I) Antibacterial activity of MoS₂-coated specimens. (a) Photographs of E. coli growth on bacterial culture medium (overnight incubation at 37 °C) and (b) bacterial growth inhibition rate of each substrate at 37 °C for 6 h compared to the CP Ti substrates and compared with the MS-Ti substrate. (c) SEM images of E. coli exposed to the CP Ti and the MS-eECAP substrates at 37 °C for 6 h. The left and right side images are E. coli of the surface of the CP Ti and the MS-eECAP, respectively. Photographs courtesy of Myeong Hwan Shin et al. Reproduced with permission from ref (123). Copyright 2018. Scientific Reports. (II) Fluorescence microscopy images and FE-SEM images of bacteria on HA/MoS₂-coated implants. Photographs courtesy of Jieni Fu et al. Reproduced with permission from ref (129). Copyright 2021. Nature Communications. (III) Antimicrobial activity of MoS₂-nanosheet-conjugated carbon quantum dots on Ti surfaces on (A) S. aureus and (B) E. coli. Photographs courtesy of Donglin Han et al. Reproduced with permission from ref (126). Copyright 2019. American Chemical Society. (IV) Antimicrobial assay of Ti, MoS₂–Ti, and AgBr@MoS₂ coatings on the Ti surface under visible light on (a) S. aureus and (b) E. coli. (c) Antimicrobial activity of the conjugate against S. aureus and E. coli under both dark and light conditions. (d) Photocatalytic degradation of RhB on Ti, MoS₂–Ti, and AgBr@MoS₂–Ti under visible light for 20 min. Photograph courtesy of Weidong Zhu et al. Reproduced with permission from ref (128). Copyright 2019. American Chemical Society.

3.3. Black-Phosphorus-Based Coating

In comparison to white phosphorus and red phosphorus, black phosphorus (BP) is the most stable paragon of phosphorus elements.¹³² In recent times BP has attracted massive attention for a variety of applications^133,134 in the field of biomedicine, such as bioimaging, photothermal therapy, photodynamic therapy (PDT), biosensing, drug delivery, etc. The phosphorus atoms in BP connects to three phosphorus atoms surrounded by chemical bonds, and other layers of BP are connected by van der Waals forces.¹³⁵ Black phosphorus has a layered structure, and the band gap of BP is highly dependent on the total thickness of the layers. It has been reported that the band gap can be modulated from 0.3 eV (bulk) to 2.0 eV (single layer)¹³⁶ which results in strong absorption of BP in a whole range of spectra of visible light in comparison to the other 2D materials. Besides this, antimicrobial property, excellent biocompatible nature, as well as biodegradability of black phosphorus have made it a suitable material for various biomedical applicationa. According to Mao et al., black phosphorus nanoparticles are an ideal photocatalyst for generating ROS disinfection products originating from their thickness-dependent tunable band gap (0.3–2.0 eV).¹³⁷ They have prepared a hydrogel made of black phosphorus (BP) nanosheets and chitosan to treat bacteria-infected wounds. BP-based nanomaterials could be a potential alternative to treat the diseases caused by MDR bacteria. Xiong et al. exhibited the antibacterial activity of BP nanosheets against Escherichia coli and Bacillus subtilis, and the results indicated the time- and concentration-dependent antibacterial activity. Parallelly, BP-based nanocomposites have antibacterial activity independent of the bacterial membrane structure as it acts against both Gram-positive and Gram-negative strains, which proves that the bacteria are unable to express resistance against the nanocomposites. BP nanosheets have also an excellent membrane damageability of bacterial cells which suggests the application of its antibacterial activity¹³⁸ in biomedical implants. In the study of Sun et al. the BP nanoparticles expressed ROS-dependent oxidative stress and membrane damage. They synthesized BP nanosheets with N,N′- dimethylpropyleneurea (DMPU), and it showed excellent antimicrobial activity when compared to other 2D materials like graphene and MoS₂.¹³⁹ High surface-to-volume ratio plays an important role in the antimicrobial activity of BP-associated nanomaterials. Ouyang et al. also have demonstrated the antimicrobial activity of black phosphorus. They have conjugated the BP nanosheets with silver nanoparticles, and because of the tunable band gap of BP nanosheets, with NIR irradiation they were able to show antibacterial effects because of hyperthermia. It causes irreversible membrane damage of drug-resistant bacterial strains. Second, Ag⁺ ions were leached out and resulted in oxidative stress in bacterial strains. Hence, the synergistic effect of both hyperthermia and oxidative stress was able to reduce bacterial load in a very short period.¹⁴⁰ Photothermal effects of BP/Au nanocomposites against biofilm-forming bacteria were reported by Aksoy et al.¹⁴¹ A group of researchers have developed a NIR-triggered nano-antibiotic platform based on Au–ZnO-conjugated black phosphorus which was able to show its antimicrobial activities against multi-drug-resistant bacteria.¹⁴² Because of the above-discussed properties, researchers started using black phosphorus as a coating material for antimicrobial biomedical implants. Shaw et al. have reported a solvent-free layered black phosphorus coating when imparted on medically relevant surfaces that showed excellent antimicrobial properties as well as high biocompatibility.¹⁴³ Recently, Li et al. have shown that a polyetheretherketone composite modified by carbon fiber and black phosphorus exhibited higher wear resistance, low cytotoxicity, and magnificent antimicrobial properties against S. aureus, thus making it a suitable composite for implant material.¹⁴⁴ A similar study was also conducted by Sun et al. where the group demonstrated that a composite made of polyetheretherketone (PEEK)/polytetrafluoroethylene (PTFE) with black phosphorus (BP) nanosheets was able to increase the wear resistance and antimicrobial property while being much less cytotoxic.¹⁴⁵ Recently, Fang et al. have reported a phototherapeutic system by combining polydopamine (PDA)–black phosphorus nanosheets (BP NSs)/ZnO nanowires (NWs) on titanium (Ti) substrates which exhibited outstanding antibacterial properties against biofilm-causing bacteria.¹⁴⁶

3.4. Boron-Nitride-Based Coatings

Another graphite-like refractory 2D material made with sp² covalently bonded¹⁴⁷ boron and nitrogen is boron nitride (BN). BN can be present in many crystalline forms, but the hexagonal form is the analogue of graphite and also the most stable and softest form among other boron nitride polymorphs.¹⁴⁸ BN has no charge traps on their surface along with various interesting properties like high-temperature stability, high thermal conductivity,¹⁴⁷ remarkable antioxidation property,¹⁴⁹ and biocompatibility, and because of these properties BN has an extinctive biomedical application mainly in tissue engineering scaffolds and reinforcement of materials to increase the mechanical strength, etc.¹⁵⁰ Recently, antimicrobial properties of boron nitride have also been reported. Studies have shown that boron nitride is effective in the reduction of biofilm caused by bacteria like Escherichia coli, Pseudomonas aeruginosa, Staphylococcus epidermidis, and Staphylococcus aureus mainly via damaging the bacterial cell membrane by direct interaction with it.¹⁵¹ Ikram et al. have reported that the antimicrobial activity of chemically exfoliated BN nanosheets doped with zirconium can also be exhibited by ROS production. The study said that this composite shows great antimicrobial activity against both Gram-positive and Gram-negative bacteria.¹⁵² Considering all the above-mentioned properties, BN has been used as an antimicrobial coating material for biomedical implants. Yenal et al. have used wollastonite and BN-doped, hydroxyapatite-based HAp-Wo-BN composite coatings on the surface of Ti₆Al₄V where BN was used for its bone tissue healing and antimicrobial property.¹⁵³ A very recent study also has reported the antimicrobial action of BN in a coating material made of boron nitride nanosheets containing a chitosan-based coating on a Mg alloy where BN has showed its antimicrobial property against E. coli and S. aureus by causing irreversible membrane damage.¹⁵⁴

3.5. Other 2D Materials

MXenes are a group of 2D transition metal carbides and carbonitrides with various attractive properties among which antimicrobial activity is of interest. It has been reported that the main mechanism of action of antimicrobial activity of two-dimensional Ti₃C₂T_x (MXene) is membrane damage of bacterial cells, resulting in leakage of cytoplasmic material and finally death of the microorganism. It is active against both Gram-positive and Gram-negative bacteria,^155,156 but the cytotoxicity against mammalian cells is very low, making MXenes another suitable material for antimicrobial coatings.¹⁵⁷ Jie Yin et al. developed a surface coating made of MXene nanosheets, gelatin methacrylate (GelMA) hydrogels, and bioinert-sulfonated polyetheretherketone (SP) for after orthopedic implant surgery where the role of MXenes was mainly providing antimicrobial property against pathogenic bacteria.¹⁵⁸ Another recent study on a 2D niobium carbide (Nb₂C) MXene titanium plate (Nb₂C@TP)-based clinical implant has reported the excellent antimicrobial property of MXenes where they have stated that the antibiofilm formation property exhibited by the MXenes was produced by altering the ATP synthesis related metabolic pathways of the bacteria.¹⁵⁹

Layered silicates also known as nanoclay are another 2D nanomaterial with a plate-like structure of polyanions. Because of its unique shape, large surface to volume ratio, and biocompatibility, nanoclay has gained immense importance in the biomedical field.¹⁶⁰ Besides this, cetylpyridinium chloride montmorillonite, a type of 2D nanoclay, has been reported to have antimicrobial activities. Tsubasa et al. reported the antimicrobial activity of cetylpyridinium chloride montmorillonite against Candida albicans and Staphylococcus aureus.¹⁶¹ Another study by Kiichi et al. showed the application of a polyurethane gel sheet with cetylpyridinium chloride–montmorillonite in promoting facial and somato prosthesis where cetylpyridinium chloride–montmorillonite has provided the antimicrobial activity against bacterial colonization.¹⁶²

4. Other Nanoparticles and Nanocomposite-Based Antimicrobial Coating

Recent studies reveal that hybrid nanoparticles and nanocomposites are the most precise way to coat a medical implant. Such multifunctional nanocomposites are advantageous over other nanocoating materials as they function in synergy to provide bioactivity as well as antimicrobial activity. This prompted researchers to investigate the area in the quest of more efficient, enhanced, and cutting-edge methods to boost the antibacterial and antibiofilm activity for such surface-modified medical devices and implants.

An overwhelming number of studies have been performed on silver-based nanocomposite biomaterial in these past years due to their known ability to act as an antimicrobial¹⁶³ and antibiofilm agent for pathogens involved in nosocomial infections. In 2009, Stobie and co-workers investigated the role of silver-doped perfluoropolyether-urethane coatings as a surface modification for biofilm inhibition. Polyether polyurethanes are known for their biocompatible, elastomeric, low friction antifouling biomaterial nature for biomedical coatings. They performed antibacterial assays on opportunistic organisms including MRSA, E. coli, P. aeruginosa, Acinetobacter baumanni, and a biofilm-producing Gram-positive isolate of S. epidermidis. After 24 h of incubation with inoculated silver-doped perfluoropolyether-urethane coating, the results show a 99% reduction of bacterial colonies due to the improved free radical scavenging ability of silver ions and biostability provided by fluorocarbons against oxidation and hydrolysis. It was hence concluded that the proposed silver-releasing perfluoropolyether-urethane coating surfaces are a great choice for urinary catheters to reduce hospital-born infections (such as UTIs and other secondary infections).¹⁶⁴ Furthermore, in 2013, Afzal et al. used Ag-reinforced hydroxyapatites (HAs) and carbon nanotube (CNTs) to reduce the fatal infections caused after an orthopedic implant or prosthesis. HA is an orthopedic implant material extensively used for its exceptional bioactivity, and CNT is known for its outstanding mechanical and physiochemical properties for biomedical materials. In their study they synthesized nanocomposites of CNT–Ag and HA–Ag with 5% Ag via a spark plasma sintering (SPS) method. Using SEM analysis, it was observed that there was a decrease of 64.9% bacterial growth in HA–Ag and 85.7% in CNT–Ag composites for E. coli, while there was a 78.8% reduction of growth in HA–Ag and 67.4% reduction of growth in CNT–Ag observed in the case of the S. epidermidis strain of bacteria. Thus, it was concluded that the HA/CNT–Ag nanocomposite has the potential to act as an antimicrobial biocomposite for orthopedic medical implants.¹⁶⁵

Gold nanocomposites were also reported to be used for biofilm-associated infections to rescue patients from prosthetic joint infections (PJIs) as reported by Wickramasinghe and colleagues in 2020. According to the study, they developed a photothermal nanocomposite containing d-amino acids (d-tyrosine, d-tryptophan, and d-phenylalanine) and Au nanorods (AuNRs). These nanocomposites were fabricated on different metal alloys which were Ti-based, CoCr-, and Ta-based alloys that are most frequently used in prosthetic joint surgeries. Synthesized AuNRs were loaded with a glycol chitin hydrogel matrix, and a photoactivated d-amino acid loaded hydrogel nanocomposite was prepared. The antimicrobial effect of the nanocomposite was studied on S. aureus, and the nanotoxicity of the composite was checked on the HeLa cell line. Post evaluation of results, they concluded that the hybrid nanocomposites displayed less toxicity and were effective for biofilm disruption. The crystal violet assay showed that the two-step approach of using d-amino acids for biofilm disruption and AuNRs for photothermal treatment completely eradiated the biofilm on the medical implant surface. Their study further suggested that the biofilm-associated infections can be treated by the devised method of photothermal treatment on several metal alloy materials commercially exploited for orthopedic implants.¹⁶⁶

5. Conclusion

Biomedical implants have revolutionized the treatment methodologies in biomedical science starting from dental surgeries to orthopedics to catheters, but along with this comes another life-threatening problem which is bacterial infection. Biomedical implants are very prone to bacterial contamination, and to combat this the strategy was administration of antibiotics; however, soon after, infection-causing bacteria developed resistance against the antibiotics. Beside this, bacteria usually form biofilms on the surface of implants against which the same dosage of antibiotics is not effective, which eventually triggered further bacterial resistance. This review provides a detailed insight into the use of several commercially applied metal and metal oxide nanoparticles and 2D nanomaterials as a coating material on biomedical surfaces with their proposed mechanisms of action.

With the advancement in the field of nanobiotechnology, there has been a tremendous increase in research for the development of materials that are multifunctional, bioactive, and biocompatible and have improved physiochemical properties for implant surfaces and biomedical devices. To overcome the rise in resistance to traditional antibiotics, the use of metal and metal oxide nanoparticles and 2D nanomaterials has served as a novel alternative to antimicrobial therapies for biofilm-associated hospital-acquired infections. To use nanomaterials as antimicrobial coating materials, it is important to know the mechanism of antimicrobial action where it is observed that the most common mechanisms of action used are cell membrane damage, inducing oxidative stress, cellular protein damage, and DNA damage. Silver, copper, zinc, magnesium, and its oxide nanoparticles use cellular membrane damage as a primary mode of antibacterial mechanism of action, whereas titanium and iron metal oxide nanoparticles use oxidative damage to cellular proteins and DNA by ROS generation as the primary mode of action. The antimicrobial mechanism of action imparted by the 2D nanomaterials is also similar to metal and metal oxide nanoparticles which include membrane damage by direct interaction with the bacterial cell, production of ROS, or alternating metabolic pathways. In the case of metal and metal oxides, the leached ions play important roles, whereas in the case of 2D nanomaterials, the super sharp edges of the nanosheets provide antimicrobial actions. Besides this, even though the primary mechanism of action is the same in both metal oxides and 2D nanomaterials, the process used is sometimes quite different like in copper oxide based coatings. “Contact killing” is observed, whereas in MoS₂ nanosheet coated implants photothermal and photodynamic therapies are often exploited. Incorporating such nanomaterials in a coating for implants not only provides antimicrobial activity but also contributes additional properties like an increase in mechanical strength, wear resistance, and thermal stability.

Parallelly the major limitation of such studies is lack of data from in vivo studies which again becomes one big barrier to the translational rate of such research. Even though a lot of research is going on in various in vivo models with various fabricated implants, the long-term effects of such materials in animal models are not known, and also there is a possibility that different implants show different results in different animal models. Above all, this review provides a comprehensive idea about different types of metal and metal oxide nanoparticles and 2D nanomaterials with antimicrobial properties which have been used to prepare antimicrobial biomedical implants to encourage more in vivo studies that will increase the translational rate of new antimicrobial coatings which are potent, nontoxic, and cost-effective to maintain critical hygiene and control the rapidly increasing rates of hospital-acquired infections in the near future.

Biographies

Jyotirmayee Sahoo received her Bachelor of Technology degree in Biotechnology from Amity University, Mumbai, Maharashtra, India, in 2019, alongside receiving a “Gold Medal” for her academic accomplishments. She received a scholarship from the MHRD, India, after qualifying with the GATE Biotechnology exam in the 2021. Currently, she is pursuing a Master of Technology at the Indian Institute of Technology, Mandi, in the School of Biosciences and Bioengineering. She is currently working under the supervision of Dr. Amit Jaiswal on her Master’s thesis, and her research interest includes 2D nanomaterials and their antibiofilm and antimicrobial mechanisms of action.

Sanchita Sarkhel received her Bachelor’s degree in Microbiology at the University of Calcutta. After that, she completed her Master’s degree in Microbiology from St. Xavier’s College, Kolkata. She received a scholarship from the MHRD, India, after qualifying with the GATE Life science exam in 2020. Currently, she is pursuing her Ph.D. at the Indian Institute of Technology, Mandi, under the guidance of Dr. Amit Jaiswal. Her primary research interest lies in biomaterials with antimicrobial and wound healing properties.

Dr. Amit Jaiswal is presently an Associate Professor in the School of Biosciences and Bioengineering at Indian Institute of Technology Mandi, India. Dr. Jaiswal is also an associate of the Indian Academy of Sciences Bangalore, India. He completed his Ph.D. in Nanotechnology at Indian Institute of Technology Guwahati, India, and postdoctoral research at Washington University in St. Louis, USA, and Technion – Israel Institute of Technology, Haifa, Israel. His research interest is in the synthesis of nanomaterials for sensing, catalysis, drug delivery, and diagnostic applications.

Author Contributions

^† These authors contributed equally.

The authors declare no competing financial interest.