Onychomycosis is a difficult-to-treat chronic fungal infection of the nail. The chronic nature of onychomycosis, with relevance to current treatment practices, could be attributed to host anergy, development of increased virulence in causal agents (multidrug resistance efflux pump), and biofilms. Biofilms must be disrupted prior to antifungal treatment suggesting the necessity of combination treatment. Once the biofilm has been disrupted, further techniques in addition to antifungal usage are suggested to ensure a positive prognosis including use of antimicrobial photodynamic therapy or low-frequency surface acoustic waves. Overall, with continued success in developing antibiofilm treatment for bacterial and yeast pathogens, therapy can be more quickly expanded to dermatophytes. With a rise in predisposing factors, it is important to preemptively address treatment for this disease with continued investigation into antibiofilm therapy including optimal treatment combinations and dosages targeted specifically at dermatophytes.

Despite their ubiquitous presence, fungal pathogens rarely lead to disease in immunocompetent individuals. Moreover, in the event of successful infection, many are superficial with effective therapy available. In contrast, dermatophyte infection of the nail, onychomycosis, is difficult to treat and often leads to recurrences or relapse [1]. As onychomycosis is believed to affect 7-14% of North Americans [2,3,4,5] and contributes to 50% of nail disease [6], possible reasons for the chronic nature of the disease with relevance to current treatment practices were investigated.

Dermatophyte pathogens are normally susceptible to antifungal treatment; thus, it is unclear why infection of the nail or onychomycosis often results in chronic disease. Drug malabsorption and degradation by microsomal enzymes has been proposed as a contributing factor [7]. Additional explanations for the discrepancy that have been explored include host anergy and development of increased virulence in causal agents.

Evidence of anergy, or lack of immune response to specific fungal antigens, has been observed in patients with chronic (greater than 5 years) dermatophytosis [8]. The constitutive presence of dermatophytes and residual fungal elements is believed to induce selective immune tolerance, suggesting the importance of early treatment (within 5 years of infection) to prognosis [8]. Alternatively, complete elimination of fungal presence such as that achieved with oral terbinafine treatment can restore cell-mediated immunity [8]. Immune restoration is not observed in all patients while some require additional treatments, allowing for recurrence or chronic disease to continue. It is hypothesized that refraction to immunological conversion may be genetically determined [8].

In combination with anergy, pathogen virulence is also believed to influence disease progression and treatment recalcitrance. For instance, a proposed multidrug resistance efflux pump has been identified in dermatophytes that leads to resistance against both griseofulvin and tioconazole [9]. It is expressed in response to diverse toxic compounds including ethidium bromide, ketoconazole, cycloheximide, fluconazole, griseofulvin, imazalil, and itraconazole [9], suggesting a mechanism for strains carrying the corresponding gene to escape diverse antifungal treatments. Further investigation, however, has revealed that acquisition of genetic resistance to antifungals is not likely the reason for treatment recalcitrance. Genetically identical isolates of Trichophyton rubrum leading to chronic nail disease remain susceptible to antifungals such as griseofulvin, ketoconazole, itraconazole, and fluconazole when grown in vitro [10]. Therefore unknown contributing factors of the nail environment remain to be elucidated.

Another explanation for the chronic nature of onychomycosis is the presence of biofilms. Biofilm formation is known to lead to chronic infection with recalcitrance to traditional treatment [11,12], similar to what has been described for onychomycosis. Biofilms are also known to lead to similar symptoms including persistent inflammation and tissue damage [11]. Moreover a significant amount of microbial infections are known to be a result of biofilm formation [13].

The presence of biofilms can also explain the difference in treatment response as phenotypic characteristics of the same organism change (including growth rate and gene activation) when converting from planktonic to biofilm lifestyles [14]. For example, microbes can form communities insulated from the environment within biofilms. In addition, increased communication, increased virulence, improved metabolic cooperation, and cooperative regulation of gene expression have been observed [15] leading to increased resistance to antimicrobial agents and increased protection from host defenses (Fig. 1) [13].

Fig. 1

Planktonic growth versus biofilm formation. As microbial numbers reach a minimum density, biofilm formation begins with surface attachment and development of the protective extracellular matrix (ECM) resulting in differential phenotypic characteristics that lead to resistance to both antimicrobial agents (including antifungals) and the host immune system.

Fig. 1

Planktonic growth versus biofilm formation. As microbial numbers reach a minimum density, biofilm formation begins with surface attachment and development of the protective extracellular matrix (ECM) resulting in differential phenotypic characteristics that lead to resistance to both antimicrobial agents (including antifungals) and the host immune system.

Close modal

To further build on this theory, dermatophytes such as T. rubrum are known to be able to form biofilms on nails [14]. This can be established in part through quorum sensing. Quorum sensing, a method of communication between microbial cells used to establish population density, can act to coordinate a group response including biofilm formation and virulence. When a minimum density of bacteria/fungi accumulate, genes involved in biofilm formation are induced, thus increasing the odds of successfully establishing a mature biofilm prior to immune detection [16,17,18].

Through the process of biofilm formation, microbes have the ability to bind irreversibly to a surface. Therefore, once formed, surgical removal of a mature biofilm may be necessary and is used in the case of implanted devices or live tissue [19]. Similarly, microbes encased in a mature biofilm are not as accessible to the immune system, which can lead to tissue damage without clearing of the infection [20,21]. This is believed to be accomplished through the development of a protective layer, the extracellular matrix (ECM). Presence of the ECM also limits the efficacy of antimicrobial agents such as antifungal treatment, either through physically inhibiting contact with microbes or slowing the penetration rate [22]. As a result, increased resistance up to 1,000 fold is observed [12,23].

Therefore, biofilms must be disrupted prior to antifungal treatment suggesting the necessity of combination treatment. Components of the ECM which could act as targets for treatment include polysaccharides, proteins, and extracellular microbial DNA [24]. Use of enzymes including deoxyribonuclease I (DNase I), α-amylase, and lyase as well as lactic acid, chitosan, terpinen-4-ol-loaded lipid nanoparticles, and povidone-iodine (PVP-I) have all been investigated for an ability to disrupt biofilms.

In response to bacteria, DNase I degraded 50% of common biofilms, which increased efficacy of the antibiotics azithromycin, rifampin, levofloxacin, ampicillin, and cefotaxime [25], while up to 95% reduction in biofilm was observed in a second study [26]. Use of α-amylase was effective in detaching biofilms for some bacterial species (Staphylococcus aureus) but not others (S. epidermidis) [27]. Lyase in combination with gentamycin reduced viable counts of Pseudomonas aeruginosa by 2-3 log10 units [28], while lactonase increased penetration of ciprofloxacin and gentamycin [29]. A combination of lactic acid and cetrimide eliminated Enterococcus faecalis and E. duran biofilms [30]. Chitosan reduced biofilms of Streptococcus mutans, Listeria monocytogens, Bacillus cereus, S. aureus, Salmonella enterica, and P. fluorescens [31,32]. Finally, PVP-I demonstrated antibiofilm activity against Porphyromonas gingivalis and Fusobacterium nucleatum with a 4-6 log10 reduction in viable counts [33].

In response to Candida albicans biofilm, use of DNase I improved amphotericin B treatment by a 1-5 log10 reduction in viability depending on dosage but did not improve caspofungin and fluconazole efficacy [34]. Likewise terpinen-4-ol-loaded lipid nanoparticles were observed to eradicate C. albicans biofilms [35].

Once the biofilm has been disrupted, further techniques in addition to antifungal usage are suggested to ensure a positive prognosis including use of antimicrobial photodynamic therapy (PDT) [24]. Use of PDT for bacterial biofilms led to an almost complete disruption and clearance of P. aeruginosa [36], a reduction of greater than 6-7 log10 units depending on treatment protocol for P. aerugenosa and methicillin-resistant S. aureus [37,38], and a reduction of 1.9 log10 in the viable counts of E. faecalis [39]. Use of PDT for C. albicans biofilms led to a reduction of 0.33-0.85 log10[40]. Similarly, in mouse models, PDT has also been successful in reducing bacterial density [41,42,43] and C. albicans biofilms [44]. For best results, an increase in light dose is recommended when treating a biofilm which can help maintain efficacy against this resilient target without raising photosensitizer concentration, which could increase toxicity to patients [24].

A second option includes the use of low-frequency surface acoustic waves (SAW). SAW were previously shown to reduce bacterial biofilms (Escherichia coli, S. epidermidis, and P. aeruginosa) formed on catheters by greater than 85% when used in combination with antibiotics [45].

Overall, with continued success in developing antibiofilm treatment for bacterial and yeast pathogens, therapy can be more quickly expanded to dermatophytes.

Observational evidence suggests that the abnormally difficult-to-treat nature of onychomycosis may be caused by the formation of biofilms. The phenotypic changes, differential gene regulation, and increased virulence of sessile microbes in comparison to planktonic growth correlates with genetically identical dermatophyte strains observed to be recalcitrant to treatment in patients with nail disease but susceptible to antifungals when grown in vitro. To address the possible role of biofilms in onychomycosis, antibiofilm treatment is suggested in combination with traditional antifungals. This includes disruption of the biofilm, possible through use of enzymes and exogenous treatments along with additional therapy such as PDT or SAW. With a rise in predisposing factors (diabetes, immunosuppression, obesity, smoking, and advancing age [46,47,48,49,50]) it is important to preemptively address treatment for this disease with continued investigation into antibiofilm therapy including optimal treatment combinations and dosages targeted specifically at dermatophytes.

The authors have no conflicts of interest. There were no funding sources for this work.

Westerberg DP, Voyack MJ: Onychomycosis: current trends in diagnosis and treatment. Am Fam Physician 2013;88:762-770.
de Berker D: Clinical practice. Fungal nail disease. N Engl J Med 2009;360:2108-2116.
Elewski BE, Charif MA: Prevalence of onychomycosis in patients attending a dermatology clinic in northeastern Ohio for other conditions. Arch Dermatol 1997;133:1172-1173.
Gupta AK, Jain HC, Lynde CW, Macdonald P, Cooper EA, Summerbell RC: Prevalence and epidemiology of onychomycosis in patients visiting physicians' offices: a multicenter Canadian survey of 15,000 patients. J Am Acad Dermatol 2000;43:244-248.
Ghannoum MA, Hajjeh RA, Scher R, Konnikov N, Gupta AK, Summerbell R, Sullivan S, Daniel R, Krusinski P, Fleckman P, Rich P, Odom R, Aly R, Pariser D, Zaiac M, Rebell G, Lesher J, Gerlach B, Ponce-De-Leon GF, Ghannoum A, Warner J, Isham N, Elewski B: A large-scale North American study of fungal isolates from nails: the frequency of onychomycosis, fungal distribution, and antifungal susceptibility patterns. J Am Acad Dermatol 2000;43:641-648.
Scher RK, Nakamura N, Tavakkol A: Luliconazole: a review of a new antifungal agent for the topical treatment of onychomycosis. Mycoses 2014;57:389-393.
Jones HE: Problems of resistant dermatophytes. J Am Acad Dermatol 1990;23:779-781.
Elewski BE, El Charif M, Cooper KD, Ghannoum M, Birnbaum JE: Reactivity to trichophytin antigen in patients with onychomycosis: effect of terbinafine. J Am Acad Dermatol 2002;46:371-375.
Cervelatti EP, Fachin AL, Ferreira-Nozawa MS, Martinez-Rossi NM: Molecular cloning and characterization of a novel ABC transporter gene in the human pathogen Trichophyton rubrum. Med Mycol 2006;44:141-147.
Cordeiro RA, Brilhante RSN, Rocha MFG, Rabenhorsch SHB, Moreira JLB, Grangeiro TB, Sidrim JJ: Antifungal susceptibility and genetic similarity of sequential isolates of Trichophyton rubrum from an immunocompetent patient with chronic dermatophytosis. Clin Exp Dermatol 2006;31:122-124.
Costerton W, Veeh R, Shirtliff M, Pasmore M, Post C, Ehrlich G: The application of biofilm science to the study and control of chronic bacterial infections. J Clin Invest 2003;112:1466-1477.
Costerton JW, Stewart PS, Greenberg EP: Bacterial biofilms: a common cause of persistent infections. Science 1999;284:1318-1322.
Ramage G, Mowat E, Jones B, Williams C, Lopez-Ribot J: Our current understanding of fungal biofilms. Crit Rev Microbiol 2009;35:340-355.
Costa-Orlandi CB, Sardi JCO, Santos CT, Fusco-Almeida AM, Mendes-Giannini MJS: In vitro characterization of Trichophyton rubrum and T. mentagrophytes biofilms. Biofouling 2014;30:719-727.
Percival SL, Emanuel C, Cutting KF, Williams DW: Microbiology of the skin and the role of biofilms in infection. Int Wound J 2012;9:14-32.
Donlan RM, Costerton JW: Biofilms: survival mechanisms of clinically relevant microorganisms. Clin Microbiol Rev 2002;15:167-193.
Donlan RM: Biofilms: microbial life on surfaces. Emerg Infect Dis 2002;8:881-890.
Hooshangi S, Bentley WE: From unicellular properties to multicellular behavior: bacteria quorum sensing circuitry and applications. Curr Opin Biotechnol 2008;19:550-555.
Costerton JW, Lewandowski Z, Caldwell DE, Korber DR, Lappin-Scott HM: Microbial biofilms. Annu Rev Microbiol 1995;49:711-745.
Cochrane DM, Brown MR, Anwar H, Weller PH, Lam K, Costerton JW: Antibody response to Pseudomonas aeruginosa surface protein antigens in a rat model of chronic lung infection. J Med Microbiol 1988;27:255-261.
Khoury AE, Lam K, Ellis B, Costerton JW: Prevention and control of bacterial infections associated with medical devices. ASAIO J Am Soc Artif Intern Organs 1992;38:M174-M178.
Jefferson KK, Goldmann DA, Pier GB: Use of confocal microscopy to analyze the rate of vancomycin penetration through Staphylococcus aureus biofilms. Antimicrob Agents Chemother 2005;49:2467-2473.
Costerton JW: Introduction to biofilm. Int J Antimicrob Agents 1999;11:217-221; discussion 237-239.
Taraszkiewicz A, Fila G, Grinholc M, Nakonieczna J: Innovative strategies to overcome biofilm resistance. BioMed Res Int 2013;2013:150653.
Tetz GV, Artemenko NK, Tetz VV: Effect of DNase and antibiotics on biofilm characteristics. Antimicrob Agents Chemother 2009;53:1204-1209.
Hall-Stoodley L, Nistico L, Sambanthamoorthy K, Dice B, Nguyen D, Mershon WJ, Johnson C, Hu FZ, Stoodley P, Ehrlich GD, Post JC: Characterization of biofilm matrix, degradation by DNase treatment and evidence of capsule downregulation in Streptococcus pneumoniae clinical isolates. BMC Microbiol 2008;8:173.
Craigen B, Dashiff A, Kadouri DE: The use of commercially available alpha-amylase compounds to inhibit and remove Staphylococcus aureus biofilms. Open Microbiol J 2011;5:21-31.
Alkawash MA, Soothill JS, Schiller NL: Alginate lyase enhances antibiotic killing of mucoid Pseudomonas aeruginosa in biofilms. APMIS Acta Pathol Microbiol Immunol Scand 2006;114:131-138.
Kiran S, Sharma P, Harjai K, Capalash N: Enzymatic quorum quenching increases antibiotic susceptibility of multidrug resistant Pseudomonas aeruginosa. Iran J Microbiol 2011;3:1-12.
Arias-Moliz M-T, Baca P, Ordóñez-Becerra S, González-Rodríguez M-P, Ferrer-Luque C-M: Eradication of enterococci biofilms by lactic acid alone and combined with chlorhexidine and cetrimide. Med Oral Patol Oral Cirugia Bucal 2012;17:e902-e906.
Chávez de Paz LE, Resin A, Howard KA, Sutherland DS, Wejse PL: Antimicrobial effect of chitosan nanoparticles on Streptococcus mutans biofilms. Appl Environ Microbiol 2011;77:3892-3895.
Orgaz B, Lobete MM, Puga CH, San Jose C: Effectiveness of chitosan against mature biofilms formed by food related bacteria. Int J Mol Sci 2011;12:817-828.
Hosaka Y, Saito A, Maeda R, Fukaya C, Morikawa S, Makino A, Ishihara K, Nakagawa T: Antibacterial activity of povidone-iodine against an artificial biofilm of Porphyromonas gingivalis and Fusobacterium nucleatum. Arch Oral Biol 2012;57:364-368.
Martins M, Henriques M, Lopez-Ribot JL, Oliveira R: Addition of DNase improves the in vitro activity of antifungal drugs against Candida albicans biofilms. Mycoses 2012;55:80-85.
Sun L, Zhang C, Li P: Characterization, antibiofilm, and mechanism of action of novel PEG-stabilized lipid nanoparticles loaded with terpinen-4-ol. J Agric Food Chem 2012;60:6150-6156.
Collins TL, Markus EA, Hassett DJ, Robinson JB: The effect of a cationic porphyrin on Pseudomonas aeruginosa biofilms. Curr Microbiol 2010;61:411-416.
Biel MA, Sievert C, Usacheva M, Teichert M, Wedell E, Loebel N, Rose A, Zimmermann R: Reduction of endotracheal tube biofilms using antimicrobial photodynamic therapy. Lasers Surg Med 2011;43:586-590.
Biel MA, Sievert C, Usacheva M, Teichert M, Balcom J: Antimicrobial photodynamic therapy treatment of chronic recurrent sinusitis biofilms. Int Forum Allergy Rhinol 2011;1:329-334.
Meire MA, Coenye T, Nelis HJ, De Moor RJG: Evaluation of Nd:YAG and Er:YAG irradiation, antibacterial photodynamic therapy and sodium hypochlorite treatment on Enterococcus faecalis biofilms. Int Endod J 2012;45:482-491.
Junqueira JC, Jorge AOC, Barbosa JO, Rossoni RD, Vilela SFG, Costa ACBP, Primo FL, Gonçalves JM, Tedesco AC, Suleiman JM: Photodynamic inactivation of biofilms formed by Candida spp., Trichosporon mucoides, and Kodamaea ohmeri by cationic nanoemulsion of zinc 2,9,16,23-tetrakis(phenylthio)-29H, 31H-phthalocyanine (ZnPc). Lasers Med Sci 2012;27:1205-1212.
Park J-H, Moon Y-H, Bang I-S, Kim Y-C, Kim S-A, Ahn S-G, Yoon JH: Antimicrobial effect of photodynamic therapy using a highly pure chlorin e6. Lasers Med Sci 2010;25:705-710.
Hashimoto MCE, Prates RA, Kato IT, Núñez SC, Courrol LC, Ribeiro MS: Antimicrobial photodynamic therapy on drug-resistant Pseudomonas aeruginosa-induced infection. An in vivo study. Photochem Photobiol 2012;88:590-595.
Lu Z, Dai T, Huang L, Kurup DB, Tegos GP, Jahnke A, Wharton T, Hamblin MR: Photodynamic therapy with a cationic functionalized fullerene rescues mice from fatal wound infections. Nanomed 2010;5:1525-1533.
Dai T, Bil de Arce VJ, Tegos GP, Hamblin MR: Blue dye and red light, a dynamic combination for prophylaxis and treatment of cutaneous Candida albicans infections in mice. Antimicrob. Agents Chemother 2011;55:5710-5717.
Kopel M, Degtyar E, Banin E: Surface acoustic waves increase the susceptibility of Pseudomonas aeruginosa biofilms to antibiotic treatment. Biofouling 2011;27:701-710.
Gupta AK, Gupta MA, Summerbell RC, Cooper EA, Konnikov N, Albreski D, MacDonald P, Harris KA: The epidemiology of onychomycosis: possible role of smoking and peripheral arterial disease. J Eur Acad Dermatol Venereol JEADV 2000;14:466-469.
Gupta AK, Taborda P, Taborda V, Gilmour J, Rachlis A, Salit I, Gupta MA, MacDonald P, Cooper EA, Summerbell RC: Epidemiology and prevalence of onychomycosis in HIV-positive individuals. Int J Dermatol 2000;39:746-753.
Güleç AT, Demirbilek M, Seçkin D, Can F, Saray Y, Sarifakioglu E, Haberal M: Superficial fungal infections in 102 renal transplant recipients: a case-control study. J Am Acad Dermatol 2003;49:187-192.
Baran R: The nail in the elderly. Clin Dermatol 2011;29:54-60.
Döner N, Yaşar Ş, Ekmekçi TR: Evaluation of obesity-associated dermatoses in obese and overweight individuals. Turkderm 2011;45:146-151.
Copyright / Drug Dosage / Disclaimer
Copyright: All rights reserved. No part of this publication may be translated into other languages, reproduced or utilized in any form or by any means, electronic or mechanical, including photocopying, recording, microcopying, or by any information storage and retrieval system, without permission in writing from the publisher.
Drug Dosage: The authors and the publisher have exerted every effort to ensure that drug selection and dosage set forth in this text are in accord with current recommendations and practice at the time of publication. However, in view of ongoing research, changes in government regulations, and the constant flow of information relating to drug therapy and drug reactions, the reader is urged to check the package insert for each drug for any changes in indications and dosage and for added warnings and precautions. This is particularly important when the recommended agent is a new and/or infrequently employed drug.
Disclaimer: The statements, opinions and data contained in this publication are solely those of the individual authors and contributors and not of the publishers and the editor(s). The appearance of advertisements or/and product references in the publication is not a warranty, endorsement, or approval of the products or services advertised or of their effectiveness, quality or safety. The publisher and the editor(s) disclaim responsibility for any injury to persons or property resulting from any ideas, methods, instructions or products referred to in the content or advertisements.