A Review of Onychomycosis Due to Aspergillus Species

Aspergillus spp. are emerging causative agents of non-dermatophyte mould onychomycosis (NDMO). New Aspergillus spp. have recently been described to cause nail infections. The following criteria are required to diagnose onychomycosis due to Aspergillus spp.: (1) positive direct microscopy and (2) repeated culture or molecular detection of Aspergillus spp., provided no dermatophyte was isolated. A review of 42 epidemiological studies showed that onychomycosis due to Aspergillus spp. varies between < 1 and 35% of all cases of onychomycosis in the general population and higher among diabetic populations accounting for up to 71% and the elderly; it is very uncommon among children and adolescence. Aspergillus spp. constitutes 7.7–100% of the proportion of NDMO. The toenails are involved 25 times more frequently than fingernails. A. flavus, A. terreus and A. niger are the most common aetiologic species; other rare and emerging species described include A. tubingensis, A. sydowii, A. alliaceus, A. candidus, A. versicolor, A. unguis, A. persii, A. sclerotiorum, A. uvarum, A. melleus, A. tamarii and A. nomius. The clinical presentation of onychomycosis due to Aspergillus spp. is non-specific but commonly distal–lateral pattern of onychomycosis. A negative culture with a positive KOH may point to a NDM including Aspergillus spp., as the causative agent of onychomycosis. Treatment consists of systemic therapy with terbinafine or itraconazole.


Introduction
Onychomycosis are caused by dermatophytes, nondermatophyte (saprophytic) moulds (NDMs) or yeasts. Aspergillus spp., Scopulariopsis spp., Alternaria spp., Acremonium spp. and Fusarium spp. are the most common NDM agents reported to be responsible for approximately 2-25% of all the causes of onychomycoses [1][2][3]. NDM onychomycosis presents with clinical features mimicking dermatophytic onychomycosis, making clinical diagnosis difficult and unreliable [4]. Very little is known regarding the ability of NDM to invade an intact nail plate. [5] Aspergillus spp. are increasingly being reported as primary causative agents of onychomycosis worldwide with prevalence as high as 34.4% in Guatemala [6], 69.3% in Iran [4] and up to 71% among diabetic patients in Sri Lanka [7]. It has been previously thought that same species of Aspergillus were responsible for both superficial (e.g. onychomycosis) and systemic infections; however, many new species of Aspergillus are increasingly being reported to cause onychomycosis and these were not previously reported to cause systemic infections [8,9].
The isolation of Aspergillus spp. from nail specimens may mean several things: causative agent, coloniser or contaminant. Aspergillus spp. isolated from nail specimens are not susceptible to most of the topical and systemic antifungals used to treat dermatophytes [2]. Resistance to triazole antifungals occurs among the Aspergilli [10] and inadequate treatment may lead to resistance and recurrence of infection. Proper clinical diagnosis, laboratory workup and adequate antifungal therapy are thus the standard of care for Aspergillus infections.
The prevalence, clinical manifestations and mycological characteristics of onychomycosis caused by Aspergillus spp. are poorly understood. This review evaluates the clinicomycological characteristics and epidemiology of onychomycosis due to Aspergillus. We use the species name to describe both that species and closely related, often cryptic species, for the commonest species complex.

Epidemiology of Onychomycosis due to Aspergillus Species
Onychomycosis is a common condition accounting for up to 18-50% of all nail diseases and 30% of cutaneous fungal infections [11]. The global burden of fungal nail, skin and hair infections is about 1 billion cases [12], translating to nearly 300 million cases of onychomycosis globally. Aspergillus species accounts for 0.5-3% of all cases of onychomycosis [13]; therefore, about 10 million cases of onychomycosis are attributable to Aspergillus spp.
We reviewed data from 42 epidemiological studies from 19 countries across the globe between 1974 and 2017. The prevalence of onychomycosis due to Aspergillus spp., both as percentage of all causes of onychomycosis and as percentage of NDM, shows a marked geographical variation among countries, different regions of same country, over time in the same region and underlying co-morbid conditions or occupational predisposition ( Table 1). As an overall cause of onychomycosis, the prevalence of onychomycosis due to Aspergillus spp. varies between\ 1 and 35% in the general population and higher among diabetic populations at 71% (Table 1). Aspergillus spp. constitutes 7.7-100% of the proportion of NDM onychomycosis. Over 50% (23/42) of the reviewed epidemiological studies reportedly isolated Aspergillus spp. in 50-100% of the NDMs. Onychomycosis due to Aspergillus spp. is thus more prevalent and emerging cause of onychomycosis than previously thought.
Generally, A. niger (complex) and A. flavus (complex) are the commonest species group of Aspergillus isolated from abnormal nail specimens. A. fumigatus, A. terreus and A. nidulans are also common, as reported from the epidemiological studies (Table 1). Rare and newly described species from case reports include A. tubingensis, A. sydowii, A. alliaceus, A. candidus, A. versicolor, A. unguis, A. persii, A. sclerotiorum, A. uvarum, A. melleus, A. tamarii, A. nomius and others, some of which are with the main pathogenic species complexes [9,14].

Predisposing Factors
Factors such as increasing age, nail trauma, immunodeficiency, hyperhidrosis, socio-economic status, poor hygiene, climatic conditions, occupational exposures such as gardening and house chores, barefoot walking and paronychia predispose to onychomycosis [15]. Furthermore, damage can also be induced by hormonal disturbances (diabetes mellitus, Cushing's syndrome and hypothyroidism) or by HIV/AIDS immunosuppression or ongoing biological (immunosuppressive) therapies [16]. Among diabetics with onychomycosis in Sri Lanka, onychomycosis due to Aspergillus spp. occurred in 71%, among which A. niger (76%) and A. flavus (12%) were the most predominant species isolated [7]. The same study showed that the risk of having Aspergillus onychomycosis among diabetics increased with age and duration of diabetes. None of the above predisposing factors is specific for Aspergillus spp. However, Aspergillus onychomycosis is seen more among individuals with occupational exposures such as vegetable vendors [17] and among babassu coconut breakers [18], diabetics and the elderly [19]. Some individuals diagnosed with onychomycosis due to Aspergillus spp. do not have identifiable predisposing conditions/occupational risk factors. In fact, Soltani and colleagues in their study reported that up to 70% of patients with Aspergillus onychomycosis had no predisposing conditions [20]. Onychomycosis due to Aspergillus spp. is very uncommon in children. [21,22] Pathophysiology Aspergillus spp. are ubiquitous environmental moulds found in soil, decaying vegetation and water and are not transmitted from person to person [23]. Infection starts under the nail near the hyponychium where spores may have lodged or at the lateral nail folds, or on a diseased nail plate colonised by Aspergillus spp. [24]. Once the fungus starts to grow, the infection spreads back towards the cuticle. It looks much the same as any fungal nail infection, discolouring the nail, causing it to become thick, distorted and flaky [25]. An early experimental study with A. versicolor using healthy nail samples showed that A. versicolor could only grow on the surface of the nail without penetrating the nail plate [5]. An evidence of the nonkeratinolytic potentials of these moulds. Aspergillus species growing in nature often produces colourful pigments; therefore, an Aspergillus nail infection may well appear greenish, black, brown or various other shades [17]. The fungus will not, however, spread to the surrounding skin like some other fungal causes of nail infection [17].

Clinical Manifestation
Onychomycosis due to Aspergillus spp. is usually a distal-lateral subungual onychomycosis (DLSO). The toenails are involved 25 times more frequently than fingernails due to increased exposure to soil, water and decaying vegetation where Aspergillus moulds thrive [17].
Few studies have been carried out correlating the clinical classification with the causative fungal species. One of such studies from Saudi Arabia showed that 17 of the 23 (74%) cases of onychomycosis due to Aspergillus spp. were DLSO, 5/23 (22%)  [28]. In A. terreus onychomycosis (Fig. 1), the observed clinical patterns in fingernail were DLSO (33.3%), SWO (33.3%) and onycholysis (33.3%), and in toenail, SWO (52.9%) was the most frequent clinical pattern followed by DLSO (42.0%) and DLSO plus SWO (5.9%) [29]. Another study showed that among those with onychomycosis due to Aspergillus spp., 93% manifested with hard nails, 89% with brittle nails and 85% had discoloured nails. Involvement of surrounding skin is not common [17]. Subungual hyperkeratosis is almost always present in all patients with mould onychomycosis regardless of the genus of the fungi isolated [30]. Onychomycosis due to Aspergillus spp. are sometimes associated with subungual dermatophytoma (''fungal ball'') formation. [6] Diagnosis NDM onychomycosis and therefore onychomycosis due to Aspergillus spp. may be considered in patients with fungal infection in a diseased or traumatised nail, without associated skin involvement, unresponsive to commonly used antifungal agents [31]. It may be suspected in cases which were KOH positive and culture negative for dermatophytes [31]. The differential diagnoses for onychomycosis due to Aspergillus spp. are very broad and include yeast nail infections, tinea unguium, non-Aspergillus spp. NDMO and other non-fungal nail infections and disorders. Therefore, the diagnosis of onychomycosis due to Aspergillus spp. is both clinical and mycological. Since there are no specific signs associated with onychomycosis due to Aspergillus spp., it is not possible to diagnose it based solely on physical appearance.
Determining the mycological cause of onychomycosis is helpful in guiding antifungal treatment and preventing complications [32]. Identification of the fungal agent directs the treatment plan, as well as prognosis. Since culture of NDMs from nail specimens does not always translate to causation, Gupta et al. [2] proposed that at least 3 of the following criteria should be satisfied: (1) KOH positive, (2) culture of NDM, (3) repeated culture (2-3) of NDMs, provided no dermatophyte was isolated, (4) histopathology using periodic acid-Schiff staining positive for fungal elements, (5) culture of NDM from 5 of 20 nail inoculated nail fragments and (6) NDM identification through molecular techniques. However, a positive direct microscopy, and repeated culture or molecular detection of Aspergillus spp., provided no dermatophyte was isolated is sufficient to diagnose Aspergillus onychomycosis. Aspergillary heads may be observed with direct microscopic examination of nail specimens, especially in very chronic cases or with onycholysis. [1]  Mycological culture on Sabouraud's dextrose agar with or without cycloheximide yields fungal isolates in less than 50% of the cases. However, combining KOH preparation and culture, sensitivity is increased to 85.8%. [33] Isolation rate is higher (83%) for nail samples obtained by drilling compared to scraping (67%) [34].

Treatment
Treatment options for NDM onychomycosis are still limited; however, onychomycosis caused by Aspergillus spp. responds well to systemic antifungal agents, with itraconazole performing better than terbinafine in vitro [31]. Affected fingernails typically require 3-month therapy and toenails at least 6 months. Terbinafine given as pulse (500 mg per day for 1 week every month for 3 months) produced complete cure in 30 of 34 cases on the 12-month follow-up [2]. Tosti et al. recommend either daily terbinafine (250 mg per day) or pulse itraconazole (400 mg per day for 1 week per month) for 2-4 months, completely curing 5 of 5 patients with Aspergillus onychomycosis who accepted treatment [35]. Interestingly, the nail discolorations in Aspergillus onychomycosis often persist despite evidence of mycological cure [17]. Systemic antifungals are best combined with chemical nail avulsion using 40% urea ointment for hyperkeratotic nails and topical ciclopirox olamine nail lacquers for SWO [13]. Terbinafine resistance has been reported with A. candidus onychomycosis, and mycological cure was achieved following 10 weeks of itraconazole therapy [36]. Total nail avulsion followed by topical antifungal post-operatively has also been shown to be an effective management option (clinical cure rate 88% and mycological cure rate 100%) for patients with single or oligo-onychomycosis [37]. However, it should be noted that comparative clinical trials on the treatment of Aspergillus onychomycosis have not been done to date and that recommendations have been based on case studies.

Conclusion
Onychomycosis due to Aspergillus spp. is more prevalent and emerging cause of onychomycosis than previously thought. The prevalence ranges from 7.7 to 100% of all NDMO and between \ 1 and 35% in the general population. Since the clinical presentation of onychomycosis due to Aspergillus spp. is non-specific, it is necessary to perform laboratory procedures such as KOH, culture, histopathology and molecular techniques to diagnose it. A positive direct microscopy, and repeated culture or molecular detection of Aspergillus spp., provided no dermatophyte was isolated is the required criteria for the diagnosis of onychomycosis due to Aspergillus spp. Treatment consists of systemic therapy with terbinafine or itraconazole.
Funding Global Action Fund for Fungal Infections (GAFFI) supports Felix Bongomin.

Compliance with Ethical Standards
Conflict of interest Dr. Denning and family hold Founder shares in F2G Ltd, a University of Manchester spin-out antifungal discovery company. He acts or has recently acted as a consultant to Astellas, Sigma Tau, Basilea, Scynexis, Cidara, Biosergen, Quintiles, Pulmatrix, Pulmocide and Zambon. In the last 3 years, he has been paid for talks on behalf of Astellas, Dynamiker, Gilead, Merck and Pfizer. He is a long-standing member of the Infectious Disease Society of America Aspergillosis Guidelines group, the European Society for Clinical Microbiology and Infectious Diseases Aspergillosis Guidelines group and the British Society for Medical Mycology Standards of Care committee. FB, CRB and MDR reports no conflicts of interest.
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