Anti-biofilm compounds for various clinically important bacteria.
Biofilm formation in clinical settings is an increasingly important issue particularly due to the emergence of multidrug-resistant strains, as it resulted in increased mortality, which poses a considerable financial burden on healthcare systems. The bacterial biofilms are quite resistant to the routine antimicrobial-based therapies; therefore, the novel strategies are desired in addition to the conventional antibiotics for the effective control of infections caused by biofilm-forming microbes. So far, the approaches being proposed to control the biofilm formation in clinical practice settings include the use of biofilm inhibitors and the use of modified biomaterials for the development of medical devices to thwart the formation of biofilms. In this chapter, we have focused on the latest developments in the anti-biofilm strategies through the interruption of the quorum-sensing system, which is crucial for biofilm formation and have summarized the various classes of antibacterial compounds for the control of biofilm formation. This agrees with the recent approaches suggested by the National Institute of Health (NIH) that advocates the use of combinational therapies based on the conventional methods and complementary treatment to explore the potential utility and safety concerns of the natural products. The studies regarding these emerging strategies could possibly lead to the establishment of better therapeutic alternates compared to conventional treatments.
- quorum sensing
Group of microbial cells that are surface-attached and embedded within the extracellular matrix (self-produced), and are strikingly resistant to antimicrobials are called biofilms . Biofilms can adhere to almost different types of surfaces like body tissue and plant, plastics, metals, implant objects as well as medical devices . Formation of biofilm on implants and medical equipment and implants, for example, vascular grafts, prosthetic joints, heart valves, catheters, intrauterine devices, pacemakers, and contact lenses can cause infection. Central line-associated bloodstream infection (CLABSI) can occur due to use of intravascular catheters, furthermore, CLABSI can cause an increased rate of mortality and morbidity, and every year in the USA almost 250,000 cases of bloodstream infections are reported .
When cells adhere and attach to surfaces biofilm formation begins. Several factors can promote the attachment of microorganisms to biomaterials including increased shear forces, bacterial motility, and electrostatic as well as hydrodynamic interactions between the surface and microbial cells . It has been observed that adherence of biomaterials to bacteria via biomaterial-surface interactions and cell-surface is facilitated by numerous factors, such as protein autolysin, surface, and adhesion proteins and capsular polysaccharides, etc. For example, ‘
Another worrying characteristic of infections associated with biofilm formation is increased biofilm cell tolerance to biocides. As biofilms provide an excellent niche for exchange of plasmid, so increased resistance to the drug can affect genes containing plasmids which results in multidrug resistance (MDR) phenotypes. Enhanced drug resistance mechanisms include incomplete or slow infiltration of antimicrobials within the extracellular matrix, the formation of dormant cells during the non-dividing phase, reduced cell’s growth rate within the biofilm, hence ultimately decreasing total targets for antimicrobial molecules . Furthermore, it is difficult to treat biofilm formation with the traditional antimicrobial approach and the therapy is further inhibited by increased resistance to the antibiotic because under antibiotic selective pressure microbial cells develop resistance. For instance, it has been observed that almost above 70% of hospital isolate of
2. The process of biofilm development
The production and maturation of biofilm are complex, subsequent and dynamic processes, depending upon several factors i.e. cellular metabolism, intrinsic properties of the cells, genetic control, the substratum, and the medium signaling molecules. Biofilm formation is introduced with a conditioning film of inorganic or organic material on the cell surface; furthermore, this layer modifies the surface feature of substratum which ultimately favors microbes for colonization on the cell surface. The formation of biofilm consist of several different steps: (i) initially the reversible attachment of microbial cells with biotic or abiotic surfaces through weak forces for example van der Waals forces, (ii) irreversible attachment to the cell surface with the help of different attachment structure i.e. lipopolysaccharides, flagella, adhesive proteins or fimbriae by hydrophobic or hydrophilic interactions, (iii) and then eventually biofilm architecture development due to the production and proliferation of extracellular polysaccharide (EPS) matrix which is self-produced and is made up of proteins, extracellular deoxyribonucleic acid (DNA) and polysaccharides  (iv) in the next step mature biofilm is formed which has water channels that are responsible for distribution of nutrients as well as signaling molecules within the biofilm , (v) and then due to extrinsic or intrinsic factors separation of biofilm cells occurs individually or in clumps and finally colonization of other niches and dispersal of the cells .
3. Inhibition of initial attachment
3.1 Altering physical properties of biomaterials
Biofilm development starts with a reversible weak adhesion of microbial cells to the exterior surface of medical equipment, however, if they are not removed from the exterior of devices, they adhered permanently through their adhesion structures i.e. fimbriae, pili and thereby forming biofilm matrix . Surface charge and hydrophobicity of implant constituents play a significant role in controlling the ability of microbes to anchor to cell surfaces. Therefore, alteration in the hydrophobicity and surface charge of polymeric constituents are proved as efficient for controlling biofilm formation by using numerous antimicrobial agents and backbone compounds . Poly N-vinylpyrrolidone and Hydrophilic polymers i.e. hyaluronic acid  on silicone shunt and polyurethane catheters have been widely used to decrease the adherence of ‘
3.2 Altering the chemical properties of biomaterials
There are several chemical approaches used to alter the exterior of biomedical equipment to inhibit the biofilm formation comprising ion coatings, biocides and also antibiotics . Catheters that are impregnated with antibiotics, for example, rifampin and minocycline have been revealed to reduce the occurrence of biofilm-based infections by
It has been observed that numerous antibacterial peptides also inhibit biofilm formation in several microbes. For instance, it is considered that peptide 1018 has inhibitory effects in different microbes such as in
Chelators hindering the role of metal ions in the production of biofilm are considered as biofilm inhibitors, for example, silver salts, metallic silver and also silver nanoparticles are commonly employed as antibacterial agents in clinical implants against
4. Quorum quenching
In the majority of Gram-negative and Gram-positive bacteria, an essential cellular communicating system is presently called as Quorum sensing, which regulates a variety of genes in accordance with the density of signaling molecules furthermore, signaling molecules are called autoinducers . On the bases of signaling molecules QS is classified into three i.e. autoinducing peptide (AIP-based) for Gram-positive bacteria, N-acyl homoserine lactones (AHLs-based) for Gram-negative bacteria and autoinducer-2 (AI-2-based) for both Gram-negative and Gram-positive bacteria . When the biofilm is formed, after the initial attachment, cells secrete QS molecules that alter the expression of the microbial gene, thus changing planktonic form into a sessile form. Furthermore, QS plays a significant role in biofilm development, so It has been observed that QS inhibition i.e. quorum quenching (QQ) would be a striking approach to control biofilm formation . QS system is thought to be a target for developing new antimicrobial agents, moreover, QS system plays a crucial role in regulating pathogenetic factors and also virulence factors production in several pathogens . The most important benefit of preventing biofilm formation by QQ is that this approach decreases the risk of multidrug resistance (MDR) and thus creating this approach noticeable to prevent biofilm-based infections in clinical settings. The different approaches for the inhibition or removal of biofilms are summarized in Table 1 and Figure 1.
|N-Acyl homoserine lactones||Transcriptional regulators (LuxR and LasR)||Decreased the production of QS signals and virulence factors|||
|Patriniae||Biofilm related genes||Reduced the production of exopolysaccharide|||
|Hordenine||Quorum sensing related genes||Blocked QS-controlled phenotypes like biofilm formation|||
|Quercetin||Transcriptional regulators of quorum sensing related genes||Inhibition of biofilm formation||[52, 53]|
|‘||Pyocyanin||Inhibited Pyocyanin production and reduced twitching ability|||
|Parthenolide||Extracellular polymeric substance and transcriptional regulators of quorum sensing related genes||Inhibition of the expression of QS related genes expression and downregulation of extracellular polymeric substance production|||
|Ginkgolic acids (GAs)||Curli gene expression, prophage genes||Biofilm formation was inhibited on the polystyrene, glass and nylon membrane|||
|Phloretin||Toxin genes, autoinducer-2 importer genes curli genes, prophage genes||Decreased biofilm formation and production of fimbria|||
|Cinnamaldehyde||LuxR-DNA-binding||Affected the biofilm formation and virulence|||
|‘||F-ATPase activity, virulence genes, surface protein antigen (SpaP)||Affected the cell-surface hydrophobicity index, Inhibited surface protein antigen (SpaP)|||
|Leaf extract of ‘||Exopolysaccharides (EPSs), glucosyltransferases (Gtfs)||Decreased adherence properties of bacterial cells|||
|Quercetin||pH||Disrupted the pH in biofilm|||
|‘||Not mentioned||Inhibition of biofilm formation and disruption of mature biofilm|||
|Phloretin||Efflux protein genes||Anti-biofilm formation at low|||
|‘||Adhesin proteins||Antimicrobial and anti-biofilm activity|||
|Quercetin||SrtA gene||The blockage of SrtA gene function, impairment of biofilm formation|||
|Quercetin||Protein translation and folding pathways||Blocked the protein translation and folding pathways|||
5. Removal of biofilms
Another anti-biofilm approach is the dissociation of the biofilm matrix which accounts for around 90% of biofilm dry mass. This dissociation will ultimately expose the sessile bacteria to the antibiotics as well as host immune defense. The enzymes majorly employed for biofilm matrix-degradation can be divided into three categories Proteases, nucleases and polysaccharide degrading enzymes .
Moreover, the surfactants also possess the antibiofilm activities as the cetyltrimethylammonium bromide (CTAB), sodium dodecyl sulfate (SDS) and Tween 20 have been found to promote the detachment or dispersal of biofilms. Surfactin; a biosurfactant produced by the
6. Bacteriophages as antibiofilm agents
Bacteriophages are considered as the largest creature in the biosphere, because of antibiotic resistance development, bacteriophages play an important role in the destruction of microbes. Use of bacteriophages is now considered as an alternative strategy to antibiotics, particularly for disruption or biofilm inhibition. Bacteriophages are beneficial than chemical agents and antibiotics. The isolation of bacteriophage is simple and fast, furthermore, its production is also cheap, and these are very distinct against a host or either host range, therefore, do not disrupt the normal flora. Bacteriophages are ecologically friendly, so with the persistence of host bacteria, they can replicate at the target site and have no adverse effects.
Bacteriophages also considered as potent antibiofilm mediators, e.g., phage T4 can cause infection and replicates within
Despite the several benefits of phage use, there are some disadvantages also, for example, the release of a considerable amount of bacterial membrane-bound endotoxins, decreased number of phages encoding toxins, insufficient pharmacokinetic data and conversion of lytic phages to prophages is also a big concern. Some of the above-mentioned problems have been well determined through different processes like designing a recombinant phage from
Different studies show some of the strongest inhibitions, for example, the existence of biofilm EPS matrix hindering the control of biofilm via antibacterial agents and higher antibiotic resistance can be controlled through phage use. Furthermore, there are many limitations of phage use such as microbial resistance to phages, virulence genes that are phage-encoded can incorporate inside the host bacterial genome and the narrow host range. Phage efficacy can also be reduced by the immune system, and phage preparations that are improperly obtained can also contain endotoxin. To control these obstacles engineered phages or phage mixtures can be an effective alternative. Moreover, after proper selection and several studies phages has become one of the most useful anti-biofilm agents.
7. Natural anti-biofilm strategies
7.1 Plant extracts
Many extracts of plants and their derivatives were widely studied to eliminate the ‘
Other two plant extracts Dandasa (
Another plant extract ‘casbane diterpene’ isolated from “
A natural product extracted by ‘honey’ bee from floral nectar is called as ‘honey’ however, ‘honey’ is generally common and is usually used for its remarkable activity in wound-healing, anti-inflammatory, and antibacterial activity and used as an antioxidant. It has antimicrobial activities against 60 species of fungi and bacteria. ‘Honey’ was reported as a useful agent to control the biofilm formation. Furthermore, it was described that ‘honey’ is effective in the prevention of
7.3 Essential oils
Naturally plant-derived volatile substances are called as essential oils (EOs). Because of their antibacterial and preservative properties, these are effective and favorable natural products for the food industry. These essential oils are commonly used against a wide diversity of microorganisms since ancient time. These oils exhibit antimicrobial impact on the cell wall of microbes, leading to the destruction of microbes. Furthermore, it is suggested that these oils are very effective in inactivating many microbes without producing antimicrobial resistance . Because of little mammalian toxicity, rapid degradation in the environment and availability of many essential oils make them beneficial antibiofilm agent .
The antimicrobial activity of “tea tree” essential oils scientifically named
Since biofilms are abundant in nature, the importance of biofilms in hospitals especially regarding their role in infections is often undervalued. Future studies should attempt to comprehend the biological forces controlling the colonization to develop innovative strategies for controlling biofilm biomass within a clinical context. Additionally, comprehensive research is required to recognize the potential of various synthetic and natural quorum sensing inhibitors (QSIs) for their applicability for humans. As these QSIs do not encourage the antibiotic resistance, therefore they can surely be the future therapeutic agents for the management of biofilm-based bacterial infections in clinical settings.
Conflict of interest
The authors declare no conflict of interest.