Natural Products as Antibiofilm Agents

Abstract

Biofilms, are vastly structured surface-associated communities of microorganisms, enclosed within a self-produced extracellular matrix. Microorganisms, especially bacteria are able to form complex structures known as biofilms. The presence of biofilms especially in health care settings increases resistance to antimicrobial agents which poses a major health problem. This is because biofilm-associated persistent infections are difficult to treat due to the presence of multidrug-resistant microorganisms. This chapter will give an idea about documented agents including isolated compounds, crude extracts, decoctions, fractions, etc. obtained from natural sources such as plants, bacteria, fungi, sponge and algae with antibiofilm activities. Furthermore, we have done phylogenetic analysis to identify plant families most prolific in producing plant species and compounds with good antibiofilm properties so as to aid in prioritizing plant species to investigate in future studies. The data in this chapter will help serve as valuable information and guidance for future antimicrobial development.

1. Introduction

The empirical approach to antimicrobial therapy among health care professionals and the concurrent patronage of over-the-counter antibiotics by patients have together caused an exponential rise in multidrug resistance among clinically relevant antimicrobials and with increasing trends for the past two decades [1]. Different mechanisms of antimicrobial resistance have been proposed, including the (i) alteration of the antibiotic target by genetic mutations or post-translational modification, (ii) deactivation of the antibiotic through hydrolysis or modification, such as phosphorylation by an enzyme, (iii) increased efflux of the antibiotic out of the cell by efflux pumps and porins, (iv) decreased influx/penetration of the antibiotic into the cell, through changes in cell wall structure; and overproduction of the antibiotic target through gene amplification [2]. However, one of bacteria’s preferred and commonly deployed strategies to overcome the effect of antimicrobials is the formation of biofilms. Over 90% of pathogenic bacterial species, including Staphylococcus aureus (S. aureus) and Pseudomonas aeruginosa (P. aeruginosa), possess an inherent ability to produce biofilms, making biofilms the leading cause of multidrug resistance among microorganisms [3, 4, 5].

Biofilm is a complex community of sessile microbial communities embedded in a self-producing polymeric matrix comprising exopolysaccharides, proteins, nucleic acids, and cell surface proteins [6, 7, 8]. As a community of microorganisms, biofilms constitute either a single microbial species or a combination of a different class of bacteria, fungi, protozoa, archaea, and yeast, with a unique ability to colonize almost any environmental niche, biotic or inert surfaces [9, 10, 11, 12, 13]. Biofilm enables microorganisms to withstand harsh environmental conditions such as nutrient deficiencies, high osmotic pressure, the low potential of hydrogen, oxidative stress, and antimicrobial insults [14]. The increased resistance of biofilms to antimicrobials arise from phenotypic cell variation and gene transcription. In particular, there is an exponential growth of microorganisms and genetic transfer of extrachromosomal elements via cell-to-cell communication system called quorum sensing [14, 15, 16, 17]. Quorum sensing is critical in the development and survival of biofilms; thus, it regulates the nutritional demands of microorganisms within the biofilm to meet the external supply of resources [18, 19]. In addition quorum sensing is essential for the biosynthesis and secretion of small molecule signals that activate a range of downstream processes including virulence and drug resistance mechanisms as seen in biofilms [20, 21].

The health risks of biofilms are enormous, which underscore their utilization in plant protection, bioremediation, wastewater treatment, and corrosion prevention in agricultural and industrial settings [22, 23, 24]. In particular, the biofilm grows on living human tissues such as the lungs and teeth and the surfaces of implanted biomedical devices, including contact lenses, central venous catheters [8, 25], prosthetic joints, pacemakers, and intrauterine devices [7]. Unlike single bacterial plankton cells, the treatment of biofilm-mediated infections is challenging owing to the decreased susceptibility to antimicrobial agents and other chemotherapeutics. The availability of qualitative (such as Congo red agar, microtitre plate, tube methods) and quantitative (including polymerase chain reaction (PCR)) techniques have enabled the detection and measurement of biofilms [26]. Conversely, the evaluation and screening of antimicrobials against biofilms are of great challenge. In particular, standard microdilution testing cannot evaluate the susceptibility of biofilms to antimicrobial drugs because these tests focus on planktonic (suspended) organisms rather than biofilm (surface-associated) organisms [7]. Instead, susceptibility must be determined directly against biofilm-associated organisms, preferably under conditions that mimic in vitro and/or in vivo conditions. In this light, several biofilm models systems have been developed to permit accurate screening and evaluation of novel agents for their antibiofilm activity [27, 28].

Although nature has provided a plethora of natural products with varying chemotherapeutic properties to fight human infectious diseases, discovering new and effective antimicrobials has been slow. The decline in the efficacy of existing chemotherapy and the surge in drug resistance has triggered an expedient exploration of natural products, especially from plants and microbial origin, for their antibiofilm activity against biofilm-mediated human infections. Plant extracts and plant-derived chemical products, such as essential oils, flavonoids, terpenoids, have been shown in vitro to have antimicrobial and antibiofilm activity [27, 28, 29, 30, 31]. Secondary metabolites and other peptidic compounds from microorganisms also exhibit antagonistic effects against biofilms [6, 32]. These chemical constituents exert their action by inhibiting critical elements within a biofilm and/or terminating biofilm formation processes [33]. Given the unique nature of plants and microbes, natural products derived from these sources could provide an avenue for developing newly efficacious and clinically desirable chemotherapies against biofilms-mediated infections and their associated health consequences.

This chapter aims to provide a comprehensive summary of natural products from plants and microbial sources as potential sources of antibiofilm agents. Again, it highlights the strategies and model organisms used to identify and evaluate the antibiofilm capacity of these naturally isolated chemical compounds.

2. Biofilm formation

Biofilm formation represents a survival mechanism deployed by microorganisms in response to unfavorable environmental conditions [34]. Structurally, biofilms are a collection of adherent microorganisms in a milieu of an extracellular matrix consisting of polysaccharides, proteins, nucleic acids, and lipids. This unique architecture enables biofilms to cling firmly to surfaces of implanted body organs and biomedical devices and, more importantly, increase their resistance to antimicrobial therapy. The presence of bacterial secreted glycocalyx and degrading matrix enzymes reduces the antimicrobial concentration of which individual plankton cells within the biofilm are exposed [35, 36].

The morphogenesis of biofilms constitutes five distinct stages; namely, reversible attachment, irreversible adhesion, production of extracellular polymeric substances, biofilm maturation, and dispersal/detachment. As the initial step in biofilm formation, reversible attachment is characterized by the interaction between plankton cells and the conditioned surface. Fewer plankton cells move to the surface of the substrate by convection, pedesis, or sedimentation [37]. Consequently, chemotaxis directs bacterial cells along a nutrient gradient [38]. Upon reaching the surface of the substratum, the interaction between the cell surfaces and the substratum is dependent on the net sum of repulsive or attractive forces generated by the two characters [39, 40]. The presence of fimbriae, flagella, pili, and glycocalyx enables the microorganisms to overcome the repulsive forces (such as electrostatic, hydrophobic, Van der Waals, and hydration interactions) from the substratum and subsequently cling [39, 41, 42]. The rate of biofilms formation is influenced by the substrate’s physicochemical properties, including the surface roughness, hydrophobicity, surface charge, and the presence of conditioning films [41, 43, 44].

Furthermore, bacterial cells transition into an irreversible adhesion phase. Irreversible attachment occurs through the combined effect of short-range forces of the substrate (such as dipole-dipole, hydrogen, ionic and covalent interactions) and adhesive structures of the bacterial cells. The flagella and pili, for instance, are critically important in the attachment process of various strains of microorganisms [45, 46, 47, 48]. For example, Vatanyoopaisarn et al. demonstrated the firm clinging ability of wild-type Listeria monocytogens (L. monocytogens) compared to the non-flagellated mutant type [45]. Similarly, Di Martino and colleagues showed the distinctive role of type one and type three fimbriae in initiating the attachment of Klebsiella pneumonia (K. pneumonia) to abiotic surfaces [46]. Alarcon and coworkers also observed the critical role of pilus in the twitching substrate movement of P. aeruginosa [48].

Moreover, the resident plankton cells produce extracellular polymeric substances (EPS), an essential biofilm component. Quorum sensing and cyclic-di-GMP mediated EPS formation [49, 50, 51, 52]. The formation of EPS promotes cohesion among bacteria and the adhesion of biofilms via hydrophobic and ionic interactions [49, 53, 54]. In addition, EPS is vital in constructing biofilms, maintaining biofilm architecture, quorum sensing, and genetic transfer among individual organisms within the biofilm [49, 55].

The resident bacterial cells proliferate into microcolonies mediated by autoinducers (AIs). AIs are chemical signaling molecules that permit intra-species and inter-species bacterial cell-to-cell communication [56, 57]. The surge in AIs activates critical enzymatic machinery in bacterial species for regulating the formation of microcolonies and the maturation of biofilms [52]. For example, the increase in AIs causes synchronous activation of the 15 gene-long epsA-O in Bacillus subtilis (B. subtilis) that causes an increased production of EPS. The proliferation of microcolonies and the increased accumulation of EPS trigger gene expression [52]. This alteration in gene expression reversibly stimulates additional EPS as adhesive molecules to bind individual plankton cells. In addition to EPS production, water channels are created to facilitate the inflow of nutrients to the individual cells within the biofilm [58]. During the maturation stage of biofilm formation, there is restricted motility of the bacterial cells together with characteristic variation in gene and protein expression between biofilm and plankton cells [59, 60].

The terminal phase of biofilm formation, delineated as detachment or dispersal, is regulated by a complex mechanism constituting signal transduction, effector, and environmental factors [61]. Detachment/dispersal represents a unique phase in the life cycle, where plankton cells segregate and escape from biofilms to establish microcolonies on fresh surfaces [62, 63]. Of note, the dispersal phase of a biofilm is characterized by the detachment of plankton cells from hitherto biofilm, seeding or passive movement of plankton to new uncolonized surfaces, and clinging or attachment to substrates [61, 64, 65].

3. Models for assessing antibiofilm activity

Several methods have been developed to study the antibiofilm activities of various compounds in vitro. However, only a few in vivo strategies for studying biofilms have been described. Given the importance of bacterial biofilm infections worldwide, we describie some models for assessing the efficacy of antibiofilm compounds in vivo.

3.1 The human organoid model

The human epidermis organoid model has a tough methicillin-resistant S. aureus (MRSA) USA300 and P. aeruginosa PAO1 biofilm system for studying host-microbe interplay and enable the screening of novel antibiofilm agents. This model allows the screening of synthetic host peptides to reveal their superior antibiofilm activity against MRSA compared to the antibiotic mupirocin. This model provides an exciting tool for elucidating disease pathology and testing novel drugs toxicities and efficacies. It also has the added advantage of reducing the use of animals in pre-clinical testing and replacing in vivo infection models with an ethical alternative that better reflects human disease [27].

This method involves establishing bacterial biofilm by seeding the center of the skin model with 5 µL of 2 $\times$ 10⁸ CFU/ml of MRSA or P. aeruginosa PA01 or fluorescently-tagged MRSA or PA01-mCherry or luminescent MRSA-lux or PA01-lux resuspended in PBS and cultured at 37$°\mathrm{C}$ and 7.3% CO₂. 30 µL of 1–4 mg/ml DJK-5 peptide was then added on top of the biofilm for 4 h, 1–3 days post inoculation. Luminisense signal are monitored daily after the establishment of infection until luminescence are observed in the culture medium underneath the skin. This is to study how long the skin could endure biofilm growth. ChemiDoc imaging system is used to visualize biofilms and bacterial counts quantified by sonicating, votexing and serially diluting excised skin samples on agar plates [27].

3.2 Wound models

Among the most widely used models to investigate antibiofilm compounds is the skin wound model. It involves either causing damage to the skin (abrasion, burns or surgical excisions) and subsequently infecting the injured region with biofilm-forming bacteria, or inducing the formation of absess or wounds by seeding high-density biofilm forming bacteria subcutaneously. The commonly used clinically relevant organisms are S. aureus, Stapylococcus epidermidis (Staphylococcus epidermidis) and P. aeuruginosa [66]. The inoculum can differ depending on the expected severity of the infection ranging from acute to chronic, with chronic infections mimicking biofilm infection in human more accurately. Recovery and/or healing of the infected wound therefore indicates antibiofilm activity. Effectiveness of antibiofilm compounds can also be assessed by (a) examining the infectious process and recovery via real-time imaging with an in vivo imaging system as well as wound size measurement using calipers and photographs, (b) tissue analysis to assess tissue regeneration process, (c) assessment of genetic fingerprints associated with the formation of biofilms such as pslD, mucC and quorum sensing related genes (d) analysis of inflammatory patterns (e) assessmet of underlying organs [67, 68].

3.3 Oral infections model

Various biofilms from disease and non-disease causing microorganisms results in the formation of dental caries. Dental caries results from the interation between diet and microbiota-matrix that occur on the oral surface [69]. This is mostly replicated in animal models using newly weaned rats. Prior treatmet with antbiotics is essential to elimintate existing microbiome. Subsequently, the animals are fed with cariogenic diet while also receiving the bacteria (e.g. Streptococcus mutans (S. mutans)) orally daily for period of 5–7 days. The infection is ascertained by sowing oral samples. The topical application of the compounds is carried out on the teeth, daily for 30–45 days and the mandibles and molars excised at the end of the study to evalauate the carious lesions [70].

Periodontitis can as well be replicated in animal models using its associated bacteria (e.g. Streptococcus gordoni (S. gordoni) and Porphyromonas gingivalis (P. gingivalis)) and confirmed by oral sowing or PCR analysis [71]. The treatment can be perfomed topically either to prevent or eradicate already formed biofilm infection. The animals are euthanized at the end of the experiment, and the skull excised for alveolar bone loss assay of the maxilla [71, 72].

3.4 Respiratory tract chronic infections model

The primary organism associated with biofilm lung infection in cystic fibrosis (CF) has been identified to be P. aeruginosa. In the cystic fibrosis murine model, bacteria are inoculated either intrathecally, intranasally or by instillation [73]. The inoculum and the frequency of inoculation underscores the severity of infection. Bacteria carriers such as alginate formed by the bacteria strain itself or by bacteria incorporation on agar beads can be used to establish chronic pulmonary infection. Intrathecal instillation is however, the most preferred route for inoculation of bacteria in this scenario [74].

Clinical isolates of P. aeuruginosa has also been used in some models. This model has an advantage of having a shorter time between establishment of infection and end of treatment than that described above. Since the bacteria is directly inoculated, it can result in severe acute respiratory distress (SARS) and eventually death even before treatment has been effective [67].

3.5 Foreign body infection model

The ability of biofilm forming bacteria to grow and multiply on the surfaces of certain medical devices [75] has led to the discovery of this model. The preformation of biofilm on these surgically implanted foreign bodies affect the activity of defense cells [25]. This model can be executed using two (2) approaches. These are Site Specific Device Model where biofilm forming bacteria are introduced at the injection site after devices are placed in particular organ or region in humans for evaluation of antibiofilm activity, and Subcutaneous Device Model where deliberately colonized foreign bodies are inserted in the subcutaneous layer, mostly at the back of the animals [76]. In Site Specific Device Model, antibiofim activity is measured at the part of the device that made contact with bacteria or measured by bacterial recovery at injection site [75]. In Subcutaneous Device Model, the mobility of antibiofilm peptides can be restricted with the aim of preventing bacterial contact and eventually biofilm development [75]. However other modes of assessment like histological analysis, imaging by IVIS, scanning microscopy, and inflammatory response detection can also be employed in evaluating antibiofilm activity in test organisms [75].

4. Methods used to determine anti-biofilm effects of natural products

Bacteria undergo an evolutionary mechanism to withstand harsh environmental conditions. The antibacterial agents derived from natural sources may serve as an effective alternative due to the presence of secondary metabolites, which possess selectional advantages against the biofilm-forming microorganisms [77, 78, 79]. Several methods have been reported as reliable protocols to investigate the anti-biofilm effects of natural products (Table 1) [88, 89]. Crystal violet assay is the widely accepted assay used to identify the anti-biofilm potentials of natural products despite the limitation, including the repeated washing that could lead to loss of cells and biofilm disruption [77, 88, 90, 91]. Other methods used to determine the antibiofilm effects of natural products are the Tissue Culture Plate (TCP) method [82], which exists as the most typical use standard method and is a comparatively reliable method to Congo Red Agar method (CRA) and Tube method [80]. Tube method and Congo red agar methods qualitatively detect biofilm formed, whiles the tissue culture plate method quantitatively determines the amount of biofilm formed [76]. Real time, conventional and multiplex PCR are other techniques used at molecular level to detect biofilm genes [92, 93, 94].

Method of biofilm detection	Principle	Aim
Tissue culture plate	It involves the staining of cells with crystal violet dye [77, 80, 81]	Biofilm detected quantitatively
Tube method	Crystal violet staining where visible lining forms at the bottom and wall of the tube [80]	Biofilm detected qualitatively
Congo red agar	Congo red staining formed black colonies crystals [81, 82, 83, 84]	Biofilm detected qualitatively
Crystal violet assay	Quantifies the dye bound to biofilm [77, 85]	Quantitative determination of biofilm
Real-time PCR, Multiplex PCR and conventional PCR	Amplification of DNA to the generation of fluorescence which can simply be detected [86, 87]	Detection of biofilm genes

Table 1.

Methods to determine anti-biofilm effects of natural products.

In measuring the anti-biofilm activities of natural products, viability and matrix biomass can be assessed, where resazurin and crystal violet staining are performed sequentially in the same plate. Wheat germ agglutinin-Alexa Fluor 488 fluorescent conjugate is mainly used to stain the matrix, which is essential to measure the biofilm matrix, biomass, and viability to investigate the potencies of anti-biofilm effects of natural products [95, 96].

5. Antibiofilm agents from nature

5.1 Plant-derived antibiofilm agents

Plants have since time immemorial served as a source of therapeutics for the treatment and prevention of a plethora of diseases. This practice continues today, with more than 80% of people globally reportedly using various herbal remedies as a source of primary healthcare [97]. In mainstream medicine, plants have proven to be a prolific source of novel chemical matter from which essential drugs used to treat various diseases have been developed [98]. Galvanized by the emergence and spread of the antimicrobial drug resistance phenomena, numerous plant species have been thoroughly investigated as novel sources of antibacterial agents. To complement these strategies, the search for agents that can reverse resistance (resistance breakers) or target alternative mechanisms of overcoming antibacterial resistance, including biofilms, is being pursued [99, 100]. Plants have been identified as a potential oasis of such agents, prompting many studies in the last decade inspired towards the search for antibiofilm agents from plants. This section summarizes current studies on the investigation of antibiofilm agents, including crude extracts, fractions thereof, and pure compounds from plants (Tables 2 and 3; Figure 1).

Plant species	Comment
Aralia spinosa (Araliaceae)	MBIC50 = 2 μg/ml against S. aureus [101]
Juglans regia (Juglandaceae)	MBIC50 = 7.21 μg/ml and MBEC50 = 57.71 μg/ml against S. epidermis [102]
Liriodendron tulipifera (Magnoliaceae)	MBIC50 = 32 μg/ml against S. aureus [101]
Citrus bergamia (Rutaceae)	Inhibited P. aeruginosa biofilm formation 79% at 1.56 μg/ml [103]
Gymnopodium floribundum (Polygonaceae)	IC50 = 53.6 μg/ml against S. aureus [104]
Zygophyllum coccineum (Zygophyllaceae)	MBEC = 15.63, 3.9, 15.63 and 15.63 μg/ml against Streptococcus pneumoniae, S. aureus, P. aeruginosa, and E. coli, respectively [105]
Ziziphus jujuba (Rhamnaceae)	50% inhibition at 1.41 μg/ml against S. aureus [106]
Matayba oppositifolia (Sapindaceae)	IC50 = 10.4 μg/ml against S. aureus [104]
Schoepfia schreberi (Schoepfiaceae)	IC50 = 17.7 μg/ml against S. aureus [104]

Table 2.

Potent antibiofilm plant species.

IC, inhibitory concentration; MBIC, minimum biofilm inhibitory concentration; MBEC, minimum biofilm eradication concentration.

Compound and plant source	Comment
Xanthohumol (Humulus lupulus)	100% inhibition of S. aureus biofilm formation at 9.8 μg/ml [107]
5-Hydroxymethylfurfural (Musa acuminata)	83% inhibition at 10 μg/ml against P. aeruginosa [108]
Lupulone (H. lupulus)	100% inhibition of S. aureus biofilm formation at 1.2 μg/ml [107]
Cyanidin 3-O-glucoside (Lonicera caerulea)	MICB50 = 3.3 μg/ml against Porphyromonas gingivalis [109]
Hodiendiol I (P. artemisioides)	78, 75 and 13% inhibition of Listeria monocytogenes, Pseudomonas aeruginosa, and Staphylococcus aureus biofilms at 4 μg/ml [110]
Negletein (S. oblonga)	72–88% reduction of biofilms of S. aureus, B. subtilis, P. aeruginosa and E. coli at 12 μg/ml [111]
Syringopicroside (Syringa oblata)	92% inhibition at 1,28 μg/ml against S. aureus [112]
Quercitin-3-glucoside (S. oblonga)	92–98% reduction of biofilms of S. aureus, B. subtilis, P. aeruginosa and E. coli at 12 μg/ml [111]
Panduratin A (Kaempferia pandurate)	Prevented S. mutans and S. sanguis biofilm growth by >50% at 8 μg/ml, and reduced the biofilms by >70% at 10 μg/ml [113]

Table 3.

Potent antibiofilm plant-derived compounds.

Figure 1.

Chemical structures of some active plant derived antibiofilm compounds.

5.1.1 Apiaceae

Despite being one of the least investigated, the Apiaceae plant family has produced some of the most prolific antibiofilm plant species. Among them is the annual herb Trachyspermum ammi popularly called bishop’s weed [114]. Investigations on its seed led the isolation of a potent novel naphthalene compound, (4aS, 5R, 8aS) 5, 8a-di-1-propyl-octahydronaphthalen-1-(2H)-one, which remarkably inhibited both adherence (IC₅₀ = 39.06 μg/ml) and formation of S. mutans biofilms (~60% inhibition at 78.13 μg/ml) in vitro (Figure 2). This activity was strikingly more pronounced than its parent compound’s bacteriostatic and bactericidal properties (MIC = 156.25 μg/ml; MBC = 312.5 μg/ml) against S. mutans [114]. Thymol, a monoterpenoid isolated from Carum copticum, showed good activity against three bacterial species, namely Klebsiella pneumoniae, Escherichia coli (E. coli), and Enterobacter cloacae (E. cloacea), at sub-MIC levels, reducing biofilm formation by 80, 78, and 83%, respectively at 50 μg/ml (Figure 2). The compound was approximately fourfold more potent than its parent species [115].

Figure 2.

Chemical structures of (4aS, 5R, 8aS) 5, 8a-di-1-propyl-octahydronaphthalen-1-(2H)-one and thymol.

5.1.2 Asteraceae

The Asteraceae is one of the most prominent species-rich plant families that produce highly active terpenoid compounds. A study on Helichrysum italicum led to the isolation of 21 compounds demonstrating varied activity of either inhibiting the formation or eradication of preformed P. aeruginosa biofilms. From the 21 compounds screened, chlorogenic acid emerged as the most active inhibiting biofilm formation (45% inhibition at 128 μg/ml). In contrast, biofilm eradication for all compounds was weak (<30%) [116]. Chondrillasterol, a terpenoid isolated from the plant Vernonia adoensis, has shown an intriguing activity profile being more potent in disrupting P. aeruginosa biofilms (complete disruption at 1.6 μg/ml) in comparison to inhibiting biofilm formation (wholly inhibited at 100 μg/ml) (Figure 3) [117].

Figure 3.

Chemical structure of chondrillasterol.

5.1.3 Burseraceae

An aqueous extract of Commiphora leptophloeos showed promising inhibition of Staphylococcus epidermidis biofilm formation on different surfaces. At a concentration of 4 mg/ml, an aqueous stem bark extract of C. leptophloeos showed equipotent activity on inhibiting S. epidermis biofilms on a polystyrene (84% inhibition) and glass surface (82% inhibition) [118].Boswellia papyrifera (B. papyrifera) is a deciduous tree 12 m high with a rounded crown, a white to pale brown bark that peels off in large flakes and exudes a fragrant resin [119]. Traditionally, as therapeutics, its leaves and roots are used to manage lymphadenopathy, while the resin serves as a febrifuge. The burnt leaves of B. papyrifera act as a mosquito repellent [120]. Essential oils obtained from B. papyrifera resin inhibited preformed S. epidermidis and S. aureus biofilms by 99–71%, and 95.3–59.1% at 217.3–6.8 μg/ml, respectively [121]. At a sub-MIC concentration of 0.27 μg/ml, the essential oil of B. papyrifera observed, under fluorescence microscopy, showed to inhibit the adhesion of stained S. epidermidis cells [122].

5.1.4 Combretaceae

The medicinal plant Terminalia bellerica (T. bellerica) is found predominantly in India, Sri Lanka, Bangladesh, and South-East Asia. Its fruits are traditionally used as a laxative, astringent, and antipyretic in treating menstrual disorder, piles, and leprosy. An investigation by Ahmed et al. [122] showed that the dried fruits of T. bellerica ethanol extracts could inhibit S. mutans biofilm formation in vitro on a glass surface by 92.2% at 250 μg/ml. Another Terminalia species, T. fagifolia, has been shown to have good antibiofilm properties. The ethanol stem bark extract of T. fagifolia inhibited the formation of preformed S. epidermis and S. aureus strains in vitro. It was particularly active against S. epidermis by inhibiting biofilm formation by ~70% at a sub-MIC concentration of 12.5 μg/ml compared to ~85% inhibition at 50 μg/ml against S. aureus [123]. Similarly, a water fraction of Combretum elaeagnoides showed potency against multiple species being able to reduce biofilm formation of S. aureus, Salmonella typhimurium (S. typhimurium), Salmonella enteritidis (S. enteriditis), Klebsiella pneumoniae, and Enterobacter cloacae by 80, 73, 63, 54, and 66%, respectively, at 1 mg/ml [124].

5.1.5 Fabaceae

Along with the Asteraceae, the Fabaceae family is one plant species that has received substantial interest as a source of antibiofilm agents. Copaifera pauper C. paupera) is a medicinal tree commonly found in South America that exhibits activity against monospecies and multispecies formed biofilms [125]. For the monospecies (Aggregatibacter actinomycetemcomitans and Porphyromonas gingivalis) produced biofilms, C. paupera oleoresins showed marked activity against the individual strains and with IC₅₀ (eradication of biofilm) values of 58.66 μg/ml and 104.9 μg/ml, respectively. Activity against the multispecies biofilms was marginally lower with a measured IC₅₀ (eradication of biofilm) of 594.5 μg/ml. Copaifera pubiflora oleoresins have shown a similar pattern of activity against individual A. actinomycetemcomitans [IC₅₀ (eradication of biofilm) = 189.4 μg/ml)] and P. gingivalis [IC₅₀ (eradication of biofilm) = 94.02 μg/ml)] strains and their combined multispecies biofilm [IC₅₀ (eradication of biofilm) = 556.8 μg/ml)]. Three compounds, namely polylactic acid, hardwikiic acid, and kaurenoic acid, have been isolated from a Copaifera spp. and also shown to have potency against both the monospecies and multispecies biofilms of A. actinomycetemcomitans and P. gingivalis [IC₅₀ (eradication of biofilm) ranging from 55.79 to 462 μg/ml)] [125]. Other species that have shown marked activity against multispecies biofilms include Pityrocarpa moniliformis, Anadenanthera colubrina, and Dioclea grandiflora [125].

Trigonella foenum-graceum (T. foenum-graceum), commonly called fenugreek, is an annual legume and a traditional spice crop native to the eastern Mediterranean. It has been known for its medicinal properties in the Mediterranean and Asian cultures for many years. Fenugreek seeds are traditionally used as laxative, expectorant, carminative, and demulcent [126]. The methanol extracts of T. foenum-graceum seeds inhibited P. aeruginosa biofilms in a dose-dependent pattern (24.1–68.7% at 125–1000 μg/ml) without affecting bacterial proliferation [127]. The extract caused a reduction to the exopolysaccharide (EPS) produced by P. aeruginosa biofilms. In addition to P. aeruginosa, T. foenum-graceum showed activity against the aquatic pathogen Aeromonas hydrophila reducing EPS production and biofilm formation by 46 and 76.9%, respectively, at 800 μg/ml [127].

5.1.6 Lamiaceae

The Lamiaceae is a family of flowering plants commonly known as the mint family with a cosmopolitan distribution containing about 236 genera and about 6900–7200 species. Many plants in this family are aromatic and include widely used culinary herbs like basil, mint, rosemary, and sage [128]. Several Lamiaceae species have been interrogated for their antibiofilm activity and have shown pronounced activity against different biofilm stages of various microorganisms. One such species is the plant Marrubium vulgare (M. vulgare), a perennial herb found right across the globe. The plant is well renowned for its medicinal properties and serves as a therapeutic agent for several ailments, including gastrointestinal disorders, asthma, pulmonary infections, and ulcers. The aqueous decoctions of M. vulgare inhibited adherence of methicillin-resistant S. aureus biofilms with IC₅₀ of 8 μg/ml and IC₉₀ of 128 μg/ml [129]. However, the plant was less effective in inhibiting S. aureus biofilm growth on a plastic surface (31% inhibition at 128 μg/ml). Surprisingly, at the highest test concentration of 128 μg/ml, M. vulgare showed no bacteriostatic activity suggesting the species is selectively more potent against biofilm mechanisms. Aqueous extract prepared from the aerial parts of Ballota nigra, mirrored this bioactivity profile. Specifically, inhibiting methicillin-resistant S. aureus biofilm formation and adherence by 45–90% at 8–128 μg/ml while demonstrating limited bacteriostatic activity at the highest test concentration [129].

The genus Salvia is well documented for its bacteriostatic and bactericidal properties. Various species within this genus possess dual antibiofilm properties. Hexane-soluble and dichloromethane soluble fractions and sub-fractions of Salvia officinalis (S. officinalis) have shown impeccable antibiofilm and bacteriostatic properties with an MBIC₅₀ and MIC values ranging from 3.668 to 200 μg/ml and 25 to 400 μg/ml, respectively, against P. gingivalis, F. nucleatum, P. melaninogenica, and A. actinomycetemcomitans. The labdane diterpenoid manool has been isolated and identified as the active principle from S. officinalis, showing pronounced activity with MBIC₅₀ and MIC values of 12.5 μg/ml and 3.12 μg/ml, respectively against A. actinomycetemcomitans (Figure 4) [130].

Figure 4.

Chemical structure of manool.

While Mentha piperita oil is considerably active against Chromobacterium violaceum (Inhibited biofilm formation by 72.5% at 0.049 μg/ml), it is inactive against P. aeruginosa at reasonably higher test concentrations of 6.25, 3.125 and 1.56 μg/ml. In the same study, Thymus vulgare essential oil showed marked potency against both species inhibiting their biofilm formation by 70% at 0.049 μg/ml (against C. violaceum) and 65% at 3.125 μg/ml (against P. aeruginosa) [103]. Equally impressive is the species Perovskia artemisioides, which has inhibited biofilm formation of L. monocytogenes, P. aeruginosa, S. aureus, Acinetobacter baumanii (A. baumanii), and Pectobacterium carotovorum by 92, 95, 71, 35, and 94% at 4 μg/ml. Subsequent work led to the identification of numerous antibiofilm compounds from P. artemisioides [110].

5.1.7 Malvaceae

Alcea longipedicellata (Aulonemia longipedicellata) is a member of the Alcea genus with over 80 flowering plants in the family Malvaceae, commonly known as the hollyhocks and native to Asia and Europe. The compound, malvin, isolated from the flowers of A. longipedicellata flower, exhibited about 55% inhibition of S. mutans biofilm adherence at 0.1% v/v (Figure 5) [131]. Hibiscus rosa-sinensis a tropical shrub used in folk medicine to treat respiratory disorders and diarrhea, among other ailments, has shown remarkable activity against drug-resistant strains of Helicobacter pylori (H. pylori). An ethyl acetate fraction of H. rosa-sinensis demonstrated strong biofilm formation inhibition against H. pylori at sub-MIC concentrations (79% inhibition at 125 μg/ml) [132].

Figure 5.

Chemical structure of malvin.

5.1.8 Myristicaceae

The Myristicaceae are flowering plants native to Africa, Asia, Pacific islands, and the Americas. The family consists of 20 genera and at least 500 species. Fruit of the Myristicaceae, particularly the lipid-rich aril surrounding the seed in some species, are essential as food for birds and mammals of tropical forests [133]. Plants in the family Myristicaceae with reported antibiofilm activities include Myristica fragrans (M. fragrans), Syzygium aromaticum, and Syzygium cumini. M. fragrans has been shown to inhibit Salmonella enterica biofilm formation by 88% at 50 μg/ml. Biosynthesised silver nanoparticles of M. fragrans showed marginally improved activity inhibiting the formation of S. enterica biofilm by 99.1% at 50 μg/ml [134]. Another study on M. fragrans led to the isolation of the compound macelignan, which reduced the formation of S. mutans and S. sanguis biofilm by >50% at 10 μg/ml (Figure 6) [113]. The methanol fruit extract of S. cumini disrupted Klebsiella pneumoniae biofilm biomass in a dose-dependent manner by 35.85, 64.03, and 79.94% at test concentrations of 0.1, 0.5, and 1 mg/ml, respectively [135]. Essential oils from the aerial parts of S. aromaticum reduced Staphylococcus epidermidis biofilm biomass by 50.3% at 20 μg/ml [136].

Figure 6.

Chemical structure of macelignan.

5.1.9 Amaryllidaceae

Extracts of Crinum asciaticum, a member of the family Amaryllidaceae, was investigated for its anti-tuberculosis, anti-efflux pump and antibiofilm activity. This study reealed the anti-infective activity of the extracts against Mycobacterium smegmatis (M. smegmatis) (NCTC 8159) and Mycobacterium aurum (M. aurum) (NCTC 10437) at MICs of 125 μg/ml and 250 μg/ml respectively. Also, efflux pump inhibition was observed for both M. smegmatis and M. aurum. Of great importance is the in vitro inhibition of M. smegmatis and M. aurum biofilms which was very significant at p < 0.005 [77].

5.2 Antibiofilm agents obtained from mushrooms

Research has shown that some species of macrofungi have various chemical components with antibacterial, antifungal, antiviral, antioxidant, anticancer and antiprotozoal properties [137]. The extracts of some species, including Laetiporus sulphureus, Ganoderma lucidum, and Lentinus edodes have demonstrated antibacterial activity [138]. Fistulina hepatica, Ramaria botrytis, and Russula delica extracts had promising antibacterial activity against multi-resistant microorganisms namely MRSA, E. coli and Proteus mirabilis.

In addition, some of these compounds were found to inhibit biofilm formation [137].

Studies on the aqueous extracts of Macrolepiota procera, Pleurotus ostreatus, Auricularia auricula-judae, Armillaria mellea, and Laetiporus sulphureus were shown to inhibit Staphylococcal spp biofilm formation. These extracts reduced biofilm formation by 47.72–70.87% without affecting bacterial growth [139].

A study by Borges et al demonstrated that ferulic and gallic acid inhibited biofilm formation in P. aeruginosa by interfering with cell motility and physico-chemical features on the cell surface. It also inhibited biofilm formation by E. coli due to phenolic compounds present therein [140]. Again, wild mushroom extracts had antibiofilm activity against E. coli, Leucopaxillus gigantes and Mycenus rosea. From this same study, extracts from Sarcodon imbricants, and Russula delica inhibited biofilm formation of P. mirabilis that is resistant to fluoroquinolones, ampicillin, and cephalosporins [138].

Extracts from Lentinus edodes, one of the mostly cultivated edible mushrooms, reacted negatively to biofilm proliferation by some bacteria in a study conducted by Lingström and colleagues [141]. Upon further fractionation and isolation, the compounds; oxalic acid, quinic acid, inosine and uridine (Figure 7) were discovered to be responsible for the various levels of antibiofilm activity against S. mutans, Actinomyces naeslundii, and Prevotella intermedia strains [141].

Figure 7.

Structures of compounds isolated from mushrooms with antibiofilm activities.

Melanin obtained from Auricularia aricula, an edible mushroom, has established antibiofilm properties [142]. This pigment exhibited significant antibiofilm inhibitory activity against E. coli K-12, P. aeruginosa PA01, and Pseudomonas fluorescens P-3 [142].

5.3 Sponges as antibiofilm agents

Marine sponges produce an array of secondary metabolites such as enzymes, enzyme inhibitors, and antibiotics and represent an untapped reservoir of bioactive compounds [143]. These compounds serve as defense against environmental threats like microbial infection, competition for space, or overgrowth by fouling organisms [144].

Phorbaketals isolated from the Korean marine sponge Phorbas spp. had antibiofilm activity against S. aureus [143]. Moreover, all six phorbaketals (phorbaketal A, phorbaketal B, phorbaketal C, phorbaketal A acetate, phorbaketal B acetate, phorbaketal C acetate, Figure 8) assessed for their antibiofilm activities revealed a minimum inhibitory concentration against S. aureus 6538 higher than 200 μg/ml. All six compounds significantly inhibited biofilm formation of methicillin-sensitive S. aureus in a dose-dependent manner, with Phorbaketal B and Phorbaketal C having the highest inhibitory effects, probably due to the presence of two hydroxyl groups in its structure. Phorbaketal B and C exerts their action via reduction of the expression of alpha-hemolysin (hla) and nuclease (nuc1). Phorbaketal C further reduced the expression of RNAIII (a regulatory molecule) which stimulates hla translation, thereby repressing the expression of hla [143].

Figure 8.

Chemical structures of phorbiketals isolated from Phorbas sp.

In addition, natural compounds such as collismycin, hydroxyl flavonoids, hydroxylbipyridine, and hydroxyl anthraquinones exhibited antibiofilm activity depending on the number and positions of hydroxyl groups in the backbone structures [145]. The planktonic cell growth of S. aureus was relatively unaffected by the six phorbaketals at <100 µg/ml [143].

In another study by Paul and Puglisi, cell-free supernatants (CFSs) isolated from the sponge-associated bacteria belonging to the genera Colwellia, Pseudoalteromonas, Shewanella and Winogradskyella were evaluated for antibiofilm activity at 4°C and 25°C against Antarctic strains of P. aeruginosa ATCC27853 and S. aureus ATCC29213. Inhibition of biofilm formation was observed differently among strains which was dependent on the incubation temperature. Significant antibiofilm activity was observed by CFSs at 4$°\mathrm{C}$ and 25$°\mathrm{C}$ respectively against S. aureus and P. auruginosa without exhibiting cidal activity on bacterial growth [146]. The different physico-chemical nature of exopolymers produced by the Colwellia sp. GW185, Shewanella sp. CAL 606 and Winogradsyella CAL396 is responsible for their antibiofilm activity (Table 4).

Species and strain	Major constituents	Antibiofilm activity against organisms
Colwellia spp. GW185	Glucose, mannose, galactose, galactosamine	P. aeruginosa, S. aureus
Shewanella spp. CAL606	Glucose, mannose, galactose, galactosamine
Winogradskyella spp. CAL396	Mannose, arabinose, galacturonic acid

Table 4.

Bacterial exopolysaccharide with antibiofilm activity against pathogenic bacteria [143].

In another study, marine sponge-derived Strepomyces sp. SBT343 extracts were investigated for their antibiofilm activity on Staphylococcal biofilm formation. Results from in vitro biofilm assay of an organic extract showed inhibition of biofilm formation on polysterene, glass and contact lens surfaces. This same extract inhibited biofilm formation of Staphylococcus epidermidis and S. aureus with no antibiofilm activity against Pseudomonas biofilms [147].

5.4 Algal sources of antibiofilm agents

Existing literature proves the existence of compounds obtained from algae that possess antibiofilm properties against human pathogenic microbes. The scientific research community however, continues to discover such natural antibiofilm agents. These compounds do not exist in their pure forms but are isolated from crude extracts through a series of processes [148].

Marine algae produce certain sulfated polysaccharides that exhibit antimicrobial and antibiofilm activities [149]. Fucoidan F85 (Figure 9), a sulfated polysaccharide extracted from Fucus vesiculosus upon observation was found to possess antimicrobial and antibiofilm properties against some dental plaque bacteria [149]. Fucoidans are made up of L-fucose and sulfate esters with other different molecules [150] and are normally extracted from brown algae using acid, solvent or water at a high temperature and a long reaction [151]. According to Yunhai and colleagues, Icelandic local seaweed species (Ascophyllum nodosum and Laminaria digitate), are sources of fucoidans with antibacterial activity [152].

Figure 9.

Structure of fucoidan.

A study conducted by Maggs et al proves that marine brown algae, Halidrys siliquosa produces compounds with antibiofilm activity against Staphylococcus sp, Streptococcus sp, Enterococcus sp, Pseudomonas sp, Stenotrophomonas sp, and Chromobacterium sp. Halidrys siliquosa can be found in rock pools and sometimes forests in the shallow subtidal zone [148].

Delisea pulchra red alga, produces halogenated furanones which show antibiofilm effects against B. subtilis, E. coli [153] and P. aueroginosa [154]. These furanones oppose the transmission of intracellular signals and speed up LuxR transcription turnover (Figure 10) [155].

Figure 10.

Structure of a halogenated furanones.

The algal fronds of Plocamium magga has been reported to produce an isolate, KS8 from the Pseudoalteromonas genus that shows antibiofilm activity against acyl homoserine lactone base reporter strains (Chromobacterium violaceum (CV) ATCC 12472 and CV026) [156].

Ethanolic extracts of Chlorella vulgaris and Dunaliella salina can inhibit biofilm formation by S. mutans and P. aueroginosa [157]. This antibiofilm characteristic may be associated with the activity of glucotransferases [157].

Methanol extract of Oscillatoria sp., green algae containing silver nanoparticles also showed strong antibiofilm activity against all test pathogens in an experiment conducted by Adebayo-Tayo and associates [158].

Silver nanoparticles associated with aqueous extract of Turbinaria conoides have been reported to possess antibiofilm activity via adherence inhibition against Salmonella typhi, E. coli and Serratialique faciens [159].

6. Miscellaneous agents with antibiofilm activities

Several agents from natural products such as essential oils, honey etc. have shown great potential as bacterial biofilm inhibitors. These have been described below;

6.1 Essential oil

Essential oils from medicinal plants have received attention in recent times for their potential exploitations. This is as a result of the increasing reports of their composition and biochemicals to possess medicinal properties. A number of in vitro evidences indicates that essential oils can act as antibacterial and antibiofilm agents against a large spectrum of pathogenic bacterial strains.

The effect of Lippia alba (L. alba) and Cymbopogon citratus (C. citratus) (lemon grass) essential oils on biofilms of S. mutans was tested by Tofiño-Rivera et al. in an attempt to find new compounds against dental caries using the MBEC-high-throughput (MBEC-HTP) assay. The L. alba essential oils demonstrated significant eradication activity against S. mutans biofilms of 95.8% in 0.01 mg/dL concentration, and C. citratus essential oils showed eradication activity of 95.4% at 0.1 and 0.01 mg/dL concentrations and of 93.1% in the 0.001 mg/dL concentration [160]. Further, geraniol and citral were later identified as the major components of the essential oils. A similar investigation by Ortega-Cuadros et al., showed 93.0% growth inhibition of S. mutans biofilms at a concentration of 1.00 μg/ml of C. citratus essential oil [161].

In an investigation to access the ability of Allium sativum fermented extract and cannabinol oil extract to inhibit and remove P. aeruginosa biofilms on soft contact lenses, the cannabinol oil extract inhibited biofilm formation by about 70% and eradicated preformed biofilms in both P. aeruginosa (ATCC 9027 strain) and P. aeruginosa clinical isolates from the ocular swabs tested [162]. Cannabigerol, a non-psychoactive cannabinoid which is also naturally present in trace amounts in the Cannabis plant was able to reduced the QS-regulated bioluminescence and biofilm formation of Vibrio harveyi (a marine quorum-sensing and biofilm-producing bacterial species) at concentrations not affecting the planktonic bacterial growth [163].

Essential oils from Cyclamen coam (C. coam) and Zataria multiflora (Zinnia multiflora) extracts inhibited biofilm formation on P. aeruginosa 214, a strong biofilm producing clinical strain [164]. C. coam and Z. multiflora essential oils inhibited biofilm formation completely at concentrations <0.062 mg/ml and 4 μl/ml, respectively. It is reported that carvacrol, a major constituent of Z. multiflora essential oil inhibits biofilm formation by preventing the initial adhesion of biofilm cells to the surface [165, 166].

6.2 Lectin

A study by Moura et al. reported the antibiofilm activity of a lectin extracted from Moringa oleifera (M. oleifera) seed. The lectin from this plant exhibited antibiofilm activity against Bacillus spp. and Serratia marcescens at concentrations of 20.8–41.6 μg/ml and 0.325–1.3 μg/ml respectively [167]. The antibiofilm activity of the M. oleifera seed lectin might be due to the ability of these lectins to damage the cell wall and cell membranes through its interactions with glycoconjugates and polysaccharides constituents within the bacterial cell wall [168].

Solanum tuberosum lectins had a varying biofilm inhibitory effect when evaluated against an isolate of P. aeruginosa PA01. At a concentration between 2.5 and 15 μg/ml, the lectins inhibited the biofilm formation by 5–20% [169].

Plant lectins are reported to also exhibit antibiofilm activities against pathogenic microorganisms. A typical example are, lectins extracted from Canavalia ensiformis, Calliandra surinamensis, Canavalia marítima and Alpinia purpurata [170].

6.3 Chitosan

Chitosan is a polysaccharide composed of units of glucosamine (2-amino-2-deoxy-d-glucose) and N-acetyl glucosamine (2-acetamido-2-deoxy-d-glucose) linked by β (1 → 4) bonds. Chitosan is produced as a result of partial deacetylation of chitin leads. Chitin is found on the shells of crustaceans, arthropods and fungal cell wall [171].

The antibiofilm activity of chitosan from crab and shrimp species indigenous to the Philippines was investigated against P. aeruginosa and S. aureus. Biofilm inhibitory activity for both crab and shrimp chitosan were not observed against S. aureus at the concentration used, but activity was observed for shrimp chitosan at a concentration of 2.5 g/L. A 2.5 g/L mixed (1:1) chitosan solution of the two extracts had the highest percentage antibiofilm formation inhibition in P. aeruginosa biofilms. S. aureus biofilm formation was sensitive to the 10 g/L mixed (1:1) solution. The same mixed solution produced an inhibition against P. aeruginosa [172].

Costa et al. also reported that chitosan demonstrated antibiofilm and biofilm eradication activity against the fungus Candida albicans [171].

6.4 Honey

The exploration of new antibiotics to combat biofilm formation in resistant microbes has led to an increase interest evaluating the antibiofilm properties of honey. Manuka honey have demonstrated good antibiofilm forming activity against a range of bacteria, including Streptococcus and Staphylococcus species, P. mirabilis, A. baumannii, E. coli, E. cloacae and P. aeruginosa [173, 174].

Lu and colleagues studied the antibiofilm properties of four New Zealand based honeys; monofloral manuka honey, Medihoney (a manuka-based medical-grade honey), manuka-kanuka blend, and a clover honey on two P. aeruginosa strains PAO1 and PA14 with different biofilm forming abilities. All the different types of honey used in the study were effective at inhibiting both the planktonic cell growth and biofilm formation of both strains. In the study of the biofilm eradication properties of the honey, they concluded that honey used at clinically obtainable concentrations completely eradicated established P. aeruginosa biofilms [175]. Similar results were obtained using different strains of S. aureus, including methicillin-resistant S. aureus (MRSA) strains. In this study, they demonstrated that honey is able to reduce biofilm mass and also to kill cells that remain embedded in the biofilm matrix; and planktonic cells released from biofilms following honey treatment do not have elevated resistance to honey [176].

The biofilm inhibitory effect of Costa Rican Meliponini stingless bee honeys has also been reported against S. aureus and P. aeruginosa biofilm formation. The meliponini stingless bee honeys in a concentration-dependent manner inhibited the planktonic growth and biofilm formation, and also caused the destruction of S. aureus biofilm [177].

Australian honey has also been reported to possess antibacterial and biofilm inhibitory activities. Sindi A and colleagues in their investigation reported that Western Australian honeys from Eucalyptus marginata (Jarrah) and Corymbia calophylla (Marri) trees exhibited antimicrobial activity against Gram-negative and Gram-positive pathogens. They reduced both the formation of biofilms and the production of bacterial pigments, which are both regulated by quorum sensing. The Western Australian honey when applied to preformed biofilms had biofilm eradication activity by reducing metabolic activity in the biofilms [178].

6.5 Peptides

Peptides are small molecules made of 10–100 amino acids that are part of the innate immune response, and found among all classes of life contributing to the first line of defense against infections. In the search for an effective agent that can treat chronic infections, antimicrobial peptides (AMPs) have been shown to demonstrate antimicrobial, antibiofilm and biofilm eradication properties. Although there has not been much studies on the biofilm inhibitory action of AMP compared to its antibacterial activity, some naturally occurring AMP’s have been reported to exhibit strong antibiofilm activities against multidrug resistant as well as clinically isolated bacterial biofilms [179].

Cathelicidin peptides are one of the most important classes of AMP. Investigation of cathelicidin AMP, indicates that SMAP-29, BMAP-28, and BMAP-27 have antimicrobial activity and are able to significantly reduce biofilm formation by multidrug-resistant (MDR) P. aeruginosa strains isolated from patients with cystic fibrosis. In addition, they were bactericidal in preformed biofilms [180]. Blower et al. also demonstrated that the SMAP-29 peptide is able to inhibit biofilm production in Burkholderia thailandensis by about 50% at peptide concentrations at or above 3 μg/ml [181].

Hepcidin 20 alters the biofilm architecture of Staphylococcus epidermidis by targeting the polysaccharide intercellular adhesin after it has reduced the extracellular matrix mass [182].

The peptides lactoferrin, conjugated lactoferricin, melimine and citropin 1.1 have all shown good anti-biofilm activity against S. aureus and P. aeruginosa infection in medical devices [183].

7. Conclusion

Microorganisms, though form biofilms as a defense mechanism for survival, this action poses a threat to the healthcare system by compromising the therapeutic efficacy of antimicrobial agents and causing ascendancies in antimicrobial resistance. Natural products from plants and microorganisms provide a plethora of chemical compounds with antibiofilm properties capable of disrupting pre-formed biofilms or inhibiting the formation of new biofilms. Identifying novel antibiofilm compounds from these sources is essential to mitigate biofilm-mediated infections. Similarly, the exploration of model systems is critical for evaluating the antibiofilm properties of newly identified medicinal compounds. Altogether, understanding the antibiofilm potential of these natural products could serve as an impetus in antimicrobial drug discovery.