Species of the genus Candida, despite part of the normal human microbiota, can cause important fungal infections ranging from superficial clinical manifestations to fatal invasive lesions. Most of these infections are related to the ability of Candida spp. To form biofilm, which results in increased resistance to antifungal agents, such as ketoconazole, amphotericin B and fluconazole, hindering proper treatment. The increase in these infections together with increased resistance to antifungal drugs has made it necessary to seek new therapeutic alternatives. Among the new alternatives is the search for essential oils of plants that have antibiofilm properties. A search was performed in the Science direct, Scopus, PubMed and Scielo databases. From this search, a total of 39 essential oils were found, related to antibiofilm activity in front of Candida species, among them are lemon grass, eucalyptus, cinnamon and tea tree oils. The data found in this review demonstrated the antibiofilm activity of most essential oils, suggesting that they can be used as new treatment alternatives and reinforcing the need for further studies to prove their efficacy.
Keywords: Candida; essential oils; antibiofilm; virulence; candidiasis
Candidiasis is one of the main causes of opportunistic fungal infections. The factors that imply the increased occurrence of candidiasis are immunosuppressive therapies, invasive surgical procedures and the use of broad-spectrum antimicrobials [1, 2]. Candida species infections range from mucosal colonization to invasive lesions that can be fatal [1].
Candida albicans is the most frequent fungal species and cause superficial, cutaneous, subcutaneous and systemic infections [3]. However, other species known as "nonalbicans", including Candida tropicalis, C. glabrata, C.
parapsilosis and others have been isolated regularly [4].
Candida species are found both in the environment and in the normal flora of humans, so their presence in the organism will not always indicate fungal infection. They are causatous agents that can reach high mortality rates, especially in immunocompromised individuals, intensive care patients, post-surgical units and neoplasms [3, 5]. Candida species can colonize and cause diseases in various sites of the body such as the skin, oral cavity, gastrointestinal tract, genital, vascular system and others [5, 6].
The pathogenesis of candidiasis depends on the health of the host, as well as the virulence factors that these species express in certain situations. Virulence factors include germ tube formation, adhesin production, adhesions, biofilm The expression of virulence factors may vary depending on the infecting species, geographic origin, type of infection, location and stage of infection, and host reaction. Knowledge of these factors is considered an important tool to understand the pathogenesis of fungal diseases caused by Candida spp. and to assist in the development and discovery of new therapeutic targets [7].
Most diseases caused by Candida species are related to biofilm formation [3]. Biofilm production varies according to Candida species and consists of a pathogenetic factor that plays an important role in tissue adhering and colonization by Candida spp. [8]. The biofilm corresponds to a group of microorganisms embedded in an extracellular matrix that form a three-dimensional structure on biotic and abiotic surfaces [6, 9]. Microbial biofilms are the main cause of hospital infections and the source of many diseases that become recurrent and persistent. In addition to biofilms being genetically resistant to some antifungals, including amphotericin B and fluconazole, studies show that biofilms of Candida species considerably reduce sensitivity to different antifungals, hindering their treatment [3, 4, 6]. The increased resistance of these pathogens to conventional antifungals as well as their side effects, such as nausea, vomiting, rash and others, encouraged the search for new therapeutic alternatives. There has been a growing interest in the study of secondary metabolites of plants as new therapeutic alternatives, especially essential oils, as they have antibacterial and antifungal properties [10, 11].
Essential oils (OEs) are natural compounds constituted of a complex mixture of volatile compounds extracted from medicinal and aromatic plants with promising biological properties [10, 12]. They have wide application in popular medicine, aromatization, food preservation and fragrance industries. In recent years, studies have shown that essential oils and their constituents have antimicrobial properties against some bacteria and fungi. However, there are still few studies on the action of essential oils against the biofilm formed by Candida spp. [13, 10].
In view of the need for new therapeutic alternatives for the treatment of infections caused by Candida spp. and, the use of essential oils from plants with antibiofilm activity appears as a promising therapeutic alternative. Thus, the objective of the present study was to conduct a literature review on the antibiofilm activity of some essential oils against Candida spp.
A literature review was carried out in the online databases Science direct, Scopus, PubMed and Scielo. Scientific articles related to essential oils from plants with antibiofilm activity against Candida spp were searched. The keywords used were Candida spp. AND oil AND antibiofilm for the last 10 years (2010 to 2020).
In this research 279 articles were found in Science direct, 48 in Scopus, 40 in PubMed and 3 articles in Scielo. These articles were analyzed in their entirety to verify the relevance for the purpose of this review. After reading and study, a total 31 articles of oils were found associated with the antibiofilm activity of Candida albicans, C. tropicalis, C. krusei, C. dublinienses, C. glabrata, C. parapsilosis, C. guillermondii, C. rugosa, C. lusitaniae, C. membranafaciens C. famata, C.boidinii, C. pelliculosae C. colliculosa (Table 1).
In the last 10 years, few studies have been done on the effectiveness of essential oils in inhibiting the biofilm formation of Candida species. The main results obtained from the antibiofilm activity of essential oils found in Candida species are listed below:
Elkhalifa et al. analyzed the antibiofilm activity of essential oil and methanolic extract of Anthemis stiparum on Candida albicans and other microorganisms. The essential oil of Anthemis stiparum showed moderate antibiofilm activity against C. albicans ATCC 10239 in the MIC (50 μL/mL). The methanolic extract demonstrated greater antibiofilm activity against C. albicans, inducing 80.02% biofilm inhibition when used tested by the MIC [14].
Mback et al. evaluated the antibiofilm activity of the essential oils of Aeollanthus cucullathus, Aeollanthus heliotropioides and Plectranthus glandulosus against the species of Candida albicans and C. glabrata. The results obtained demonstrated that there was inhibition of the mature biofilm at the tested concentrations (MIC/2; MIC/4; MIC/6, MIC/10), 0.06 mg/mL, 0.5 mg/mL, 3 mg/mL, respectively. The best result was obtained by the essential oil of A. helitropioides followed by the essential oil of P. glandulosus and A. cucullathus [15].
Francisconi et al. tested and demonstrated antibiofilm activity of tea tree oil and its main component terpinen-4-ol against two clinical strains of Candida albicans. These strains, identified by molecular typing as genotypes A and B, were isolated from diabetic patients with chronic periodontitis. Nystatin tests using the reference strains C. albicans ATCC 90028 and SC 5314 were also compared to the results. Mouthwash simulations containing the oil, terpinen-4-ol and nystatin were also performed. The researchers also observed that at concentrations of 17.92 mg/mL for tea tree oil and 8.86 mg/mL of terpinen-4-ol, both were effective against the biofilm formed in the samples when compared to treatment with nystatin. In addition, it was possible to observe that there was less antibiofilm effect of the oil and its main component against the strain of Candida albicans genotype A [16].
Candida rugosa; C9 - Candida colliculosa; C14 - Cinnamomum camphora C1 C2 C5 Cinnamomum tamala C1 C2 C5 Cinnamomum verum C1 C2 C5 Citrus hystrix C1 C3 Citrus x limon C2 Cymbopogon citratus C1 C2 C3 C4 Cymbopogon flexuosus C1 Cymbopogon winterianus C1 C3 Cuminum cyminum C1 C2 Eucalypthos globulus C1 C2 C5 Gaultheria procumbens L. C1 Hedychium coccineum C1 Longifolia mint C1 Lavandula augustifolia C1 Laurus nobilis C1 C2 C3 C5 Mint x piperita L. C1 C2 C3 C4 C5 Melaleuca alternifólia C1 C2 C5 Ocimum basil C1 C3 Ocimum tenuiflorum C1 C3 Pelargonium graveolens C1 C2 C3 C5 Plectranthus glandulosus C1 C5 Pogostemon heyneanus C1 C2 C5 Sage triloba L. C1 Satureja macrosiphon C1 C4 Satureja hortensis L. C1 Syzygium aromaticum C1 C2 C3 Thymbra capitata C1 C2 C5 Thymus capitatus C1 Thymus vulgaris C1 C2 C3 Turmeric longa L. C1 C3 Zingiber cassumunar C1 C3 Zingiber officinale C1 C3 Ziziphora Tenuior C1 C1 - Candida albicans; C2 - Candida tropicalis; C3 - Candia krusei; C4 - C6 - Candida parapsilosis; C7 - Candida guillermondii; C8 - C11 - Candida boidinii; C12 - Candida pelliculosa; C13 - Souza et al. tested the antibiofilm activity of tea tree essential oil and nanoparticles of tea tree oil to reduce problems due to the low solubility and high volatility that this oil presents. Both were tested for action on biofilms formed by C. albicans, C. glabrata, C. tropicalis, C.
parapsilosis and C. membranafaciens. The essential oil of tea tree showed a significant reduction in the biomass of the biofilm formed when compared to the positive control in all tested strains. The best result was obtained against C.
glabrata, both for oil and for nanoparticles. It was also observed that at the same concentration, nanoparticles show a greater reduction in biofilm formation compared to Candida species [17].
C6 C6 C6 C8 C9 C10 C11 C12 C13 C14 C6 C7 C6 C8 C9 C10 C11 C12 C13 C14 Candida dubliniensis; C5 - Candida glabrata; Candida lusitaniae; C10 - Candida famata; Candida membranafaciens.
Khan and Ahmad evaluated the antibiofilm activity of essential oil of Cymbopogon citrates (lemongrass) against Candida albicans at concentrations of 1/2 MIC and 1/4 MIC and obtained a reduction of 11.46 to 24.67% [18]. In the study of Sahal et al. the antibiofilm activity of lemon grass oil in three strains of Candida tropicalis. The results showed inhibition in the formation of biofilms in the three strains of Candida tropicalis tested at concentrations of 0, 2, 4 and 8% (w/w) [19]. Ngo-Mback et al. tested the antibiofilm activity of lemongrass essential oil in front of Candida glabrata. The oil was fractionated by silica gel chromatography in order to find the most active compounds. The results showed that the combination of the F7F10 fractions of lemongrass oil inhibited the formation of Candida glabrata biofilm in the dose-dependent form. A 30% decrease in biofilm formation was observed when compared to the control at a MIC/10 concentration [21].
Taweechaisupapong et al. tested the antibiofilm activity of lemon grass oil in biofilm formation and against preformed biofilm of Candida dubliniensis. At the same time, the same tests were also performed for nystatin. The results revealed that the inhibiting effect of lemongrass and nystatin oil is related to doses. Lemongrass oil was tested at concentrations between 0.11 and 27.6 mg/mL and exhibited 40 to 99% inhibition in biofilm formation. Nystatin at concentrations between 4 and 512 μg/mL and exhibited 20-93% inhibition in biofilm formation. Regarding antibiofilm activity in relation to the preformed biofilm of Candida dubliniensis, both lemongrass oil and nystatin were less efficient, in the same concentrations [13]. In another study by the same author was evaluated the antibiofilm activity of lemongrass essential oil against Candida albicans and C. krusei. The results revealed that the inhibiting effect of oils on biofilm formation and against preformed biofilm was dose-related. The essential oil of lemongrass showed inhibition in the formation of biofilm in Candida spp isolates, being higher for Candida krusei than in Candida albicans. In addition, it was observed that lemongrass oil and its main constituents were able to reduce the adhesion of Candida cells used [21]. Da Silva et al. evaluated the antibiofilm activity of lemongrass essential oil in front of Candida albicans biofilm simulating mouthwash applications to evaluate the best action of the oil as to inhibition of formation, biofilm removal or in both situations. This study aimed at a further development of mouthwash based on lemongrass oil. The results showed that there was inhibition in the formation of the biofilm of Candida albicans, however, there was no effect on the removal of the mature biofilm by the strains used [22].
Cymbopogon flexuosus essential oil (Indian lemongrass) was evaluated in the study developed by da Silva Gundel et al. This study verifies the antibiofilm activity of C. flexuosus (Indian lemongrass) free and nanoemulsion in front of the Candida albicans biofilm and other micro-organisms of interest. As a result, it was observed that the formation of the biofilm of C. albicans was inhibited at concentrations of 1.22 mg/mL and 2.56 mg/mL for free oil and at concentrations of 0.28 mg/mL and 0.58 mg/mL for nanoemulsion This result indicates that the nanoemulsion containing the oil has higher antibiofilm activity than the free oil against C. albicans. Further studies should be conducted to prove its effectiveness [23].
Cymbopogon winterianus (citronella) essential oil showed antibiofilm activity against Candida albicans isolates at concentrations of 5 and 10 MIC. The best results were in the inhibition of biofilm formation. For the biofilm of C. albicans, the lowest microbial counts were obtained in citronella oil concentration 10 MIC. The commercial mouthwashes tested showed lower antibiofilm activity in
Essid et al. analyzed the antibiofilm activity of the essential oil Thymus capitatus and their combined effect with amphotericin B and nystatin. The results showed that the essential oil of T. capitatus significantly inhibited the formation of the biofilm of C. albicans. The thyme essential oil at ½ MIC (62.5μg/mL) the inhibition in biofilm formation of C. albicans was 80.6%, In comparison, at 1/2 MIC, essential oils had a higher inhibiting effect than amphotericin B and nystatin. It was also observed that the combination of the essential oil of T. capitatus with amphotericin B or nystatin showed a significant increase in antibiofilm activity. Both oils showed great eradication activity of preformed biofilm at concentrations twice as high as inhibition in biofilm formation. In addition, the combination of essential oils with antifungal was used in an in vitro biofilm model of C. albicans associated with a venous catheter. The combinations showed almost complete rupture of the biofilm on the catheter surface, demonstrating synergistic effect between the oils and antifungal agents tested [25]. In another study with thyme oil, total inhibition of biofilm development was observed for C. tropicalis, C. pelliculosaand C. albicans strains in polypropylene and glass. Only a few strains of C. colliculosa and C. parapsilosis were resistant to thyme oil at the concentrations used [26].
Jafri; Ahmad evaluated the antibiofilm activity of thymus vulgaris essential oil and its main component, thymol, in Candida albicans and C. tropicalis isolates. These were evaluated for biofilm inhibition and eradication of preformed biofilm at different concentrations, including subinhibition concentrations. The variable level of attenuation of the biofilms of C. albicans and C. tropicalis obtained in the presence of T. vulgaris and timol indicated that there was inhibition of the biofilm, preventing adhesion and its development [27].
Alves et al. evaluated the ability of the essential oils of Thymus carnosus and T. camphoratus to decrease the mass and viability of the Candida albicans biofilm. As a result, both oils were effective at concentrations close to their MIC, and the essential oil of T. carnosus was slightly more active in reducing the viability of the biofilm [28].
Rajkowska et al. reported the antibiofilm activity of clove essential oils in different strains of C.albicans, C. krusei, C. lusitaniae, C. rugosa, C. tropicalis, C. parapsilosis, C. famata, C. boidinii, C. pelliculosa and C. colliculosa. The results showed that the essential clove oil at MIC resulted in the total inhibition of the biofilm formation of the strains of C. lusitaniae, C. krusei, C. tropicalis, C. rugosa and C. albicans on glass surface and polypropylene (PP). On polyethylene terophalate (PET) surface, clove oil completely inhibited the biofilm formation of the strains of C. lusitaniae, C. boidinii, C. tropicalis, C. rugosa, and C. albicans. Only pelliculosa, showed resistance to clove oil in the tested materials. Biofilms not completely inhibited by clove oil were significantly reduced in the materials analyzed [26]. In the study by Khan and Ahmad, clove oil at the concentration of 1/2 MIC inhibited 50% of Candida albicans biofilm formation [18].
Manoharan et al. evaluated the antibiofilm activity of the essential oil of cedar leaves and their main components against Candida albicans. The result shows that the essential oil of the cedar leaf inhibited, in a dose-dependent manner, the formation of the biofilm of C. albicans in 87% at a concentration of 0.01%. The components examined at concentrations of 0.01% significantly inhibited the formation of the biofilm. In addition, the metabolic activity of biofilm cells and their adherence were reduced [29].
Quatrin et al. analyzed the antibiofilm activity of eucalyptus essential oil by comparing the activity of free oil and the nanoemulsion containing the oil against isolates of Candida albicans, C. tropicalis and C. glabrata. The results obtained proved the antibiofilm activity of eucalyptus oil. In the comparison of the forms presented, it was observed that there were no significant difference between the activity of eucalyptus essential oil and their nanoemulsion against C. albicans, C. tropicalis and C. glabrata, since the variations found in the tests were the same between the species [30].
Essential oil of Gaultheria procumbens L.
Nikolic et al. tested the antibiofilm activity of the essential oil of Gaultheria procumbens L. against the biofilm of Candida albicans and Streptococcus mutans. For this, they used the essential oil of Gautheria procumbens L., Hexoral® and Tebodont®, (mouthwashes). It was observed that Hexoral® and Gaultheria procumbens oil showed good antibiofilm activity, and Hexoral ® most effective. It is suggested that the antibiofilm activity of mouthwashes is due to the presence of its components, especially methyl salicylate, which is the main component of essential oil of Gaultheria procumbens L. Further studies are needed to prove such antibiofilm activity [31].
Peixoto et al.analyzed the inhibition of adhesion, biofilm formation and reduction of mature biofilm of Candida albicans by bay essential oil. The results showed that essential oil of L. nobilis inhibited the initial adhesion of C. albicans at 1000μg/mL (2 MIC). At the tested concentrations (MIC, 2 MIC and 4 MIC), the essential oil inhibited the formation of the biofilm. In addition, when the oil was applied for 1 min every 8h, for 24h and 48h, a decrease in mature biofilm of Candida albicans was observed. There was no significant difference when compared to nystatin. This indicates that essential oil may be a treatment alternative for Candida spp infections however, more studies should be carried out [10].
Pazarci and Tutar analyzed the antibiofilm activity of Mentha longifólia (wild mint) against Candida albicans and others biofilm-producing microorganisms in vitro implant surfaces. The surfaces tested were titanium and stainless steel, which showed no variation in biofilm growth. The results obtained showed good inhibition in the formation of biofilm, mainly of C. albicans which was susceptible to M. longifolia oil. In the highest concentrations of oil, a better eradication of the biofilm was observed, which may demonstrate that the antibiofilm activity is related to the dose of the applied oil. In addition, differences were observed in relation to the eradication of biofilm on titanium and stainless-steel surfaces at the different concentrations of essential oil used [32].
Saharkhiz et al. evaluated the antibiofilm activity of essential oil of Mentha piperita L. (peppermint) against Candida albicans and C. dubliniensis. Peppermint essential oil completely inhibited the biofilm formation of C. albicans and C. dubliniensis in a dose-dependent form at concentrations of 1 μL/mL and 2 μL/mL, respectively [33].
Giongo et al. evaluated the antibiofilm activity of the essential oil of Pelargonium graveolens and its nanoemulsion against Candida species. The geranium essential oil and its nanoemulsion inhibited the formation of the biofilm of C. albicans, C. tropicalis, C. glabrata and C. krusei. The nanoemulsion containing the oil presented better antibiofilm activity for C. albicans and C. krusei when compared to the action of the essential oil. Against C. glabrata and C. tropicalis was not difference, both, the nanoemulsion and the essential oil have a similar antibiofilm activity. When tested on catheters containing biofilm formed from the Candida species, the nanoemulsion containing the oil significantly increased activity in all species, when compared to the control of biofilm formation. In addition, in the presence of oil and nanoemulsion, a reduction in protein was observed, which is used as a measure of biofilm biomass, in plates and catheters, confirming their antibiofilm activity in the Candida species used [34].
Banu et al. tested the inhibition of biofilm formation and the rupture of preformed biofilm of the Essential Oils of Cinnamomum tamala (Indian cassia) and Cinnamomum camphora (Camphor) against Candida albicans, C. tropicalis and C. glabrata. All essential oils tested were able to inhibit the biofilm formation of the Candida species used and significantly reduced preformed biofilms of C. albicans, C. it was observed that essential oils reduced cell viability in preformed biofilms [12]. In other study Essid et al. analyzed the antibiofilm activity of the essential oils of Cinnamomum verum and their combined effect with amphotericin B and nystatin. The results showed that the essential oils of C. verum significantly inhibited the formation of the biofilm of C. albicans. At ½ MIC (31.25 μg/mL) inhibition in biofilm formation of C. albicans was 85.57%. The combination of C. verum with amphotericin B and nystatin showed no significant increase in inhibition [25].
Banu et al. verified the antibiolme activity of the essential oil of Pogostemon heyneanus (patchouli). All essential oils tested were able to inhibit the biofilm of the Candida species used. Regarding the preformed biofilm, all three reduced 60% in the biofilm formation of C. albicans, C. tropicalis and C. glabrata. In addition, it was observed that essential essentials reduced cell viability in preformed biofilms [12].
Al-Bakri; Othman and Afifi tested the antifungal activity of seven salvia species (Salvia ceratophylla L., S. dominica L., S. hierosolymitana Boiss., S. indica L., S. syriaca L., S. fruticosa Mill. [syn. S. triloba L.] and S. verbenaca L. in Candida albicans and other microorganisms. For biofilm inhibition, the essential oil of S. triloba L. The essential oil showed good prevention and control of biofilm, presenting anti-adhesion and antibiofilm activities [35].
Motamedi et al. tested the antibiofilm activity of the essential oil of Satureja macrosiphon against Candida albicans and C. dubliniensis. The essential oil showed effective antibiofilm activity against C. albicans and C. dubliniensis at concentrations of 4 and 8 μg/mL, respectively. It was observed that inhibition was dosedependent [36].
Sharifzadeh, khosravi and Ahmadian tested the antibiofilm activity of the essential oil of Satureja hortensis L. in preformed biofilms of Candida albicans isolates from oral lesions of HIV+ patients. The inhibiting effect of essential oil is related to the dose used. At concentrations of 4800, 3200, 2400 and 1600 μg/mL, biofilm formation was reduced on average 87.1%, 73.6%, 69.4% and 67%, respectively. At concentrations below the reduction rate was less than 50%. The decrease in metabolic activity of mature biofilms of C. albicans ranged from 7.1 to 80%. The morphology of the biofilm of C. albicans, presented morpho structural changes, shrinkage in the cell membranes of sessis cells and the destruction of the three-dimensional structure of the biofilm [37].
Palmeira de Oliveira et al evaluated the antibiofilm activity of Thymbra capitata essential oil in 15 strains of Candida spp. (Candida albicans, C. glabrata, C. tropicalis, C. guilliermondii and C. parapsilosis). The essential oil of Thymbra capitata showed an inhibitative effect against Candida spp. biofilms. For C. albicans and other tested species, the inhibiting effect of biofilm biomass was significant at 2x MIC (0.64 μL/mL).C. glabrata was the most resistant to the biomass reduction of the biofilm being less than 50%. However, in relation to the metabolic activity of C. glabrata biofilm, a reduction effect of more than 80% was observed at the same concentration, demonstrating that the remaining cells were metabolically inactive or dead. C. glabrata, the essential oil when used in C. albicans had a higher inhibiting effect for biomass reduction than for metabolism activity. In short, biofilm biomass and metabolic activity were reduced by up to 50% in all species in the MIC value and the most significant effects were observed at 2x MIC [38].
Rinanda et al. analyzed the antibiofilm activity of the essential oil of red ginger, whose main components are monoterpenes, on Candida albicans isolates for inhibition and degradation of biofilm. The results showed that at the lowest concentration (0.125%) the essential oil has already demonstrated activity in the degradation of the biofilm and this increased with the increase of its concentration. For inhibition in the biofilm formation of C. albicans the concentration of red ginger oil found was 0.5%, thus demonstrating that this essential oil showed significant degradation activity and inhibition of C. albicans biofilm [39].
Dolatabadi, Salari and Mahboubi evaluated the antibiofilm activity of the essential oils of Ziziphora tenuior, Lavandula angustifólia and Cuminum cyminum against Candida albicans isolates of vaginal secretion. The antibiofilm activity of the oils was related to the dose of the applied oil, that is, the highest concentrations used showed greater elimination of the biofilm. The essential oils of C. cyminume L. angustifolia showed the greatest elimination effects of C. albicans biofilm, followed by the essential oil of Z. Tenuior [40].
The search for therapeutic alternatives for the treatment of infections caused by Candida species is extremely important, especially when biofilm is formed, resulting in increased resistance to conventional antifungals. The data found in this literature review demonstrated antibiofilm activity of essential oils, including tea tree oil that showed good antibiofilm activity for Candida species and lemongrass oil that considerably inhibited the biofilm formation of Candida albicans, suggesting that they can be used as treatment alternatives. Further studies are needed before its use in the therapeutic clinic.
All the authors have contributed equally in designing, drafting the manuscript as per the journal submission format. All authors read and approved the final manuscript.