Introduction: biofilms and its association with respiratory diseases

Biofilms comprise a community of microorganisms which are embedded in a group of exopolysaccharides, diverse proteins, and nucleic acids. Such potentially pathogenic bacterial biofilms have been shown to develop on the surfaces of medical devices, such as endotracheal tubes [1] and tracheostomy tubes [2], primarily depending on the manufacturing material of these tubes [3]. These biofilms can lead to life-threatening infections [4–11] and pose a major concern for the patients with chronic respiratory diseases, including cystic fibrosis (CF) and chronic obstructive pulmonary disease (COPD) [12–16]. In cystic fibrosis patients, the biofilms have been demonstrated to be non-attached aggregates [17–19]. Notably, biofilms are also present on the lungs of patients with chronic pulmonary infections which are a major cause of morbidity and mortality [20].

The biofilm research in respiratory diseases is still not fully explored and needs immediate attention in order to maximize the drug effectiveness and treatment strategies. Most of the microflora which exists on the biofilms displays resistance to various antimicrobial agents which further increases threat to the patients with chronic pulmonary diseases. Moreover, the mechanisms contributing to this resistance are not illustrated, and it is largely phenotypic [9, 16, 21–27]. There are various factors and characteristics which affect the composition of microbial flora on the biofilm, some of which are attachment efficiency, cyclic stage, anti-effective hostile forces, physicochemical environment, mechanical factors such as shear forces, substratum, genotypic factors, nutrient sources, etc. [28].

The formation of biofilm on the treatment devices also exposes the patients to severe bacterial infections, thus worsening their disease condition [18]. Various approaches have been explored and investigated to encounter this problem such as surface modifications of the devices to alter bacterial adherence and incorporation of various antimicrobials onto the devices to prevent colonization of microflora [9]. Another approach includes use of electrical mode where the antimicrobials are released from the device surface so as to allow penetration of the antimicrobials through the biofilms [5, 29, 30].

Biofilm resistance: barrier to an effective drug delivery

There exists a difference of opinion regarding the efficacy of these approaches probably due to the biofilm resistance [31–34]. The phenomenon that has been attributed to this antimicrobial resistance includes the formation of stack of cells with various aqueous channels over and around the biofilm that acts as a penetration barrier resulting in impermeability [35]. The biofilm usually is coated with a polymer called the glycocalyx which is anionic in nature [36]. So it is also speculated that the chemical interaction between the biofilm polymer and the antimicrobial agent results in the formation of a penetration barrier which ultimately leads to a very low degree of antimicrobial absorption and effects [37]. Another postulated mechanism is heavy production of inactivating enzymes such as β-lactamases by the microflora which accumulates within the glycocalyx, thereby protecting the underlying cells [36, 38, 39]. Some studies have also questioned the microbial efflux pump system, oxygen deprivation [40–42], and colonization of anaerobic flora as contributors to the biofilm’s unresponsiveness to the antibiotics; however, certainly it requires further investigations [43]. Various other factors include oxygen deprivation [44–47] and enhanced growth of highly antibiotic resistant Pseudomonas aeruginosa strains within the airway mucus [48–50] of cystic fibrosis patients.

Approaches to prevent biofilm formation

Currently, treatment of respiratory diseases involves various drug delivery systems. Among all, inhalers (metered dose and dry powder) [51–55] are one of the most important and commonly employed drug delivery devices, which also significantly increase the chances of biofilm formation. This may lead to clinical complications, longer duration of treatment, and reduced patient compliance. Thus, it is crucial to regularly check biofilm formation, i.e., avoiding its deposition and subsequently dealing with any complications and potential hazards arising once the biofilm has established. Prevention of colonization and microflora deposition on diagnostic and drug delivery appliances remains the primary concern, followed by developing strategies for increasing accumulation of antimicrobials at the biofilm surface and also enhancing drug penetration into the biofilm [5]. Notably, the material properties of the medical device play a crucial role in preventing the formation of biofilms (Fig. 1).

Fig. 1
figure 1

Approaches in biofilm prevention

Full size image

Several recent reviews have shown that bacterial biofilm-associated chronic sinusitis in cystic fibrosis (CF) patients is primarily caused by P. aeruginosa infections and there is a lack of available treatments for such condition where the disease pathology is accompanied by opportunistic infections. It was also shown that the challenges with providing a suitable treatment include (i) identification of a suitable antimicrobial compound; (ii) selection of a suitable drug delivery device; and (iii) optimizing formulation variables to achieve effective targeted drug delivery (sinuses and nasal cavity) [5, 18, 56, 57]. One of such investigations involved the preparation of a novel inhaled combination powder containing amorphous colistin and crystalline rifapentine with enhanced antimicrobial activities against planktonic cells and biofilm of P. aeruginosa for respiratory infections [58]. Another research study has demonstrated the potential of amphibian AMP esculentin [1–21] as a new antibiotic formulation to treat infections caused by P. aeruginosa in sepsis and pulmonary infections, primarily by disrupting biofilms [59]. Remarkably, the results have also shown that esculentin [1–21] indeed prolonged survival of animals in both sepsis and pulmonary infection models [59].

Recent advances in controlling the biofilms are not only restricted to the material modifications of the drug delivery device but also include ultrasound enhancement of the antimicrobial transport that has been demonstrated to be effective against various microorganisms existing in the biofilms including Escherichia coli, P. aeruginosa, and Staphylococcus epidermidis [60–63]. In addition, various photodynamic approaches are highlighted to disrupt biofilms by generating reactive oxygen species [64]. Photodynamic approaches are most commonly employed for the pathogens associated with the oral cavity and skin [63, 65].

Formulation and drug delivery approaches to combat biofilm

Jones and colleagues have extensively investigated the capacity of liposome drug delivery in preventing growth and deposition of various pathogens [66–71]. They have explored various mechanistic insights into how the interaction between the liposomes and bacterial biofilms can avoid biofilm generation. Liposomal drug delivery applications have also been shown to be effective for the intracellular infections particularly with the reticulo-endothelial system where various pathogens like Francisella tularensis and Streptococcus pneumoniae [72, 73] were targeted. One of the advances with liposomal drug delivery includes stealth liposomes which are employed to deliver antibiotics such as gentamicin and have been demonstrated effective in a rat model of Klebsiella pneumoniae [74]. Other novel drug delivery approaches include polymer-based antimicrobial drug delivery including nanoparticles [75], microspheres and microparticles [76], hydrogels, micelles, fibrous scaffolds, and thermo-reversible gels [77–80].

In order to have effective delivery of antibiotics to the lungs for various pulmonary diseases, aerosolized systems are the best and effective approach for treatment and prophylaxis of pulmonary infections [81, 82]. Bacterial resistance in patients with cystic fibrosis is overcome by using high endobronchial concentration of tobramycin [83, 84]. Also, the antibiotic can be aerosolized in mechanically ventilated patients [85].

To design an effective drug delivery for pulmonary diseases, various critical factors need to be considered like physicochemical and mechanical characteristics of drug, intrinsic device characteristics and its performance, etc. The application of high-frequency ultrasound is another merit to be included on the aerosol systems which can help in eradicating the biofilms and delivering the drug with targeted approach and maximum therapeutic efficacy [86].

One of the recent advances include an infection-responsive system where the antibiotic is released from the drug delivery system once it receives the signals from the occurring infection such as the release of mediators, cytokines, etc. This approach is suitable for both prophylactic and prolonged use of antibiotics which in turn can avoid the associated side effects. One such investigation involves the delivery of gentamicin which was prepared as a drug delivery using polyvinyl alcohol as a drug-polymer conjugate for the treatment of wound infection where the signals appear in the form of levels of thrombin-like activity. The drug delivery system releases the gentamicin when it is incubated with thrombin and leucine aminopeptidase together, but not with any of the components alone. This system was found suitable and effective against Staphylococcus aureus in an animal model of infection [87]. Khun et al. investigated various conventional and new antifungal agents (triazoles, amphotericin B lipid (AMB) formulations, and echinocandins) for their antifungal activity against Candida albicans and C. parapsilosis biofilms which were grown on a bioprosthetic model where they demonstrated that Candida biofilms show unique susceptibilities to echinocandins and AMB lipid formulations [88]. Also, formulations for managing bacterial infections, comprising taurolidine in the form of gels, liquid, thixotropic gels, colloidal mixtures, dispensal suspensions, injectable polymers, or a microparticle, were prepared and were found effective for localized bacterial infections [89].

Various mathematical models and approaches have been established to understand the physiology of biofilms [90–94] and to investigate various novel drugs, antibiotics, antimicrobials, or biocides [95–100]. Khassehkhan and Eberl [101] guided the development of a robust mathematical modeling study that served as a platform for future models and numerical experiments to help in understanding the effect of probiotic cultures on the pathogenic films. Cipolla and colleagues [102] have shown the development of ciprofloxacin liposomal formulations (Lipoquin and Pulmaquin) which are in clinical trial for the treatment of lung infections. These liposomal formulations are believed to improve tolerability, increase patient compliance by reducing the dosing frequency, enhance penetration of biofilms, and promote treatment of intracellular infections [102]. Likewise, Halwani et al. [103] co-encapsulated gallium with gentamicin in liposomal formulation and demonstrated the combination to be more efficacious than the antibiotic alone, in eradicating antibiotic-resistant P. aeruginosa isolated from a growing planktonic or biofilm community [103].

Alhajlan et al. [104] have demonstrated the efficacy and safety of liposomal formulations containing clarithromycin with different surface charges against clinical isolates of P. aeruginosa from the lungs of cystic fibrosis patients [104, 105].

To develop a patient-compliant, effective drug delivery and treatment strategy to combat the bacterial colonization on the biofilms in the respiratory tract, various approaches like nebulization can be employed where the active moiety is aerosolized by inhalation to the respiratory tract [18]. Various compounds which are under investigation include antimicrobials [43, 106–112], silver efflux inhibitors, and certain enzymes [109, 113, 114]. These compounds mechanistically act on the extracellular polymeric substances (EPSs)/glycocalyx and various other structural components of the biofilms. This can be achieved by preparing the formulation in both aqueous solutions which can easily absorb and penetrate through the biofilm or in the form of various polymeric drug delivery systems [115] like nanoparticles and nanospheres [116–118], liposomes [119–122], microspheres [123, 124], hydrogels [125–127], micelles [128, 129], fibrous scaffolds [130–132], thermo-reversible gels [133, 134], etc., which can infiltrate through the cracks and consequently release the therapeutically active moiety over the passage of time [135, 136] (Fig. 2). Thomas et al. developed a new approach in the form of a dry aerosol where a blend of dispersion compound disintegrates bacterial colonies. One of such formulations includes ciprofloxacin HCl and glutamic acid as dispersion compound with l-leucine as an excipient. Live/dead assay along with confocal microscopy confirmed higher efficacy of the drug delivery system in eliminating biofilms in vitro in comparison to traditional antibiotic treatments [137].

Fig. 2
figure 2

Various approaches and drug delivery systems preventing biofilms

Full size image

Also, Sans-Serramitjana et al. prepared and evaluated nanocapsules containing colistin sulfate. The nanoencapsulated charged colistins have shown higher antimicrobial efficacy against biofilms comprising P. aeruginosa clinical isolates from CF patients as compared to the pure drug, colistin sulfate [104, 105]. Cheow et al. [138] demonstrated that inhaled formulations of levofloxacin-loaded polymeric nanoparticles have better antibacterial efficacy against the E. coli biofilm, thus likely to be a better treatment for respiratory infections with higher patient compliance and therapeutic activity [138]. Similarly, Loo et al. [139] presented that the combination of silver and curcumin nanoparticles possesses enhanced anti-biofilm activities against both P. aeruginosa and S. aureus [139].

One of the recent advancements to target the multidrug-resistant bacterial infections includes phage therapy, where the liposomal entrapment of phage has been shown to be highly effective in vitro as well as in vivo by overcoming the hurdles related to the clinical use of phage [140].

There are various lipid-based antibiotic delivery systems under experimental investigation which include drugs like ticarcillin [141], tobramycin [142], gentamicin [143], amikacin [122], ciprofloxacin [144], moxifloxacin [145], polymyxin B [146] colistin [147], vancomycin [148], clindamycin [149], and ceftaroline [149]. To overcome the challenges associated with oral antibiotic therapy in biofilm-related chronic pulmonary infections, especially in CF, several drugs are being assessed as potential candidates for inhalational antibiotic therapy which are currently in clinical trials, which include vancomycin and levofloxacin solution, vancomycin powder, liposomal amikacin [12], etc. These recent advances in optimizing the mode of antibiotic administration would certainly enhance the targeted and efficient delivery of antibiotics at high pulmonary concentrations necessary for disrupting complex biofilms.

Apart from the chemical moieties and drugs, the biofilms can also be targeted using natural compounds. Verkaik et al. have demonstrated comparable in vitro efficacy of various natural antimicrobials and chitosan-based formulations (toothpaste) with chlorhexidine against the bacterial flora comprising biofilms [150]. Such advances can further improve and strengthen the understanding of biofilm testing and designing of an ideal drug delivery system [151].

The choice and selection of the therapeutic moiety are very crucial as sometimes the active drug acts as both preventing the formation of biofilms and showing its therapeutic effectiveness in the treatment of disease pathology. However, in certain cases, a blend of active drug and a compound inhibiting the formation of biofilm can be employed which can be aerosolized simultaneously, exhibiting their application as an effective drug delivery system [86]. Recent developments indicate the application of high-frequency ultrasound as an effective addition to the aerosol systems, which would indeed improve targeted therapeutic effects.

Conclusions and prospects

Biofilm is a major concern in various respiratory diseases as well as associated medical devices and can cause life-threatening infections. The treatment strategies have various limitations that can be attributed to the antimicrobial resistance of the microflora comprising the biofilms, as well as the permeability of antimicrobials across the reasonably efficient barrier formed by biofilm. This leads to an unmet need of various advances, both in terms of treatment and prophylaxis, which can be utilized for the prevention of biofilm deposition and its associated effects. Preventive approaches to disrupt biofilms include development and validation of drugs that could induce/enhance pathways of biofilm self-destruction, such as energy limitation of microflora through inhibitors of oxidative phosphorylation. However, the major limitation is the lack of in-depth knowledge in the mechanisms involved in biofilm formation and persistence, which restricts the translation of the currently available technology into a clinically effective drug delivery system that can significantly overcome the complications due to biofilms in respiratory diseases. Various novel drug delivery systems such as nanoparticles, liposomes, niosomes, implantable matrices, fibrous scaffolds, micelles thermoreversible gels, etc., can be successfully and safely employed to target the biofilms. Also, recent discoveries like the infection-responsive system and application of high-frequency ultrasound with various aerosolized systems further open the horizons to develop in understanding advanced respiratory drug delivery systems with effective measures to overcome biofilm-associated problems. These novel approaches of drug delivery may open new vistas in the pulmonary clinics ensuring improved clinical outcome.