Forgotten Fungi: The Importance of the Skin Mycobiome

Abstract

The mosaic ecosystems of microbes that live on our skin encompasses not only bacteria but also fungi, microeukaryotes, and viruses. As the second most prevalent group, unique fungal communities are found across the dry, moist, and oily microenvironments of human skin, and alterations of these communities is largely driven by changes in skin physiology throughout an individual’s lifespan. Fungi have also been associated with infection and dermatological disorders, resulting from the disrupted balance between fungal-bacterial networks on the skin. Mechanisms of colonization resistance towards fungi in the skin microbiome of animals have advanced our understanding in conservation strategies yet in the human skin, the fungal microbiome (mycobiome) remains vastly unexplored. Here, we review recent studies on the role of fungi in the skin microbiome, emphasizing how fungal-bacterial interactions at the skin surface play an important ecological function in vertebrate hosts.

Introduction

Diverse microbial communities composed of bacteria, fungi, viruses and microeukaryotes live on human skin, which is collectively called the skin microbiome. Skin pores and appendages such as hair follicles and sebaceous glands provide safe harbor for these communities, representing one of the largest direct epithelial interfaces facilitating host-microbe interactions. The skin microbiome serves to fortify the primary barrier function of the skin both by directly warding off invading pathogens and tuning the immune system starting early in life [1,2]. A unique feature of the skin is the diversity of microenvironments across the body. Sites such as the forearm are drier and more desiccate, while the umbilicus (navel) and axilla (arm pit) retain a high moisture content, and sites like the forehead are oily. In turn, these features help shape the structure of associated microbial communities [3] and offer an unprecedented opportunity to study microbial ecology on the human body. Indeed, the skin is an ideal system because it is amenable to repeat sampling and longitudinal sampling over the lifespan of an individual and across the biogeography of the body.

While the skin microbiome is relatively understudied compared to the gut microbiome, significant advances have been made in recent years highlighting the importance of skin-associated microbial communities to dermatological health [4–6]. Bacteria comprise the largest proportion of the microbiome, however fungi are ubiquitous on the skin but at a lower diversity than their bacterial counterparts. Thus, interkingdom interactions within the skin microbiome are omnipresent and represent a rich source of unexplored biology. Dysbiosis induced by fungal overgrowth is a phenomenon increasingly understood to contribute to chronic diseases in both the skin and gut. Moreover, antifungal therapeutics are severely lacking in the clinic making fungal infection among one of the most difficult diseases to manage and being associated to one of the skin diseases with the highest economic burden.

Although we have only begun to shed light on how fungi function within the skin microbiome, fungal colonization has already been shown to be important during early life, throughout key developmental stages, and into adulthood as we age, impacting dermatological health and disease outcomes in both humans and non-human vertebrates. Here, we review recent advances towards the study of fungi within the skin microbiome and their association health and disease. We further highlight the gaps in fundamental knowledge regarding host-microbe and microbe-microbe interactions that still exist.

Early Life and Development of the Skin Mycobiome

Microbial colonization during infancy is critical in the development of skin homeostasis. From birth, delivery mode determines whether acquired microbiota resemble those of maternal skin or vaginal microbiota [7,8]. Characterization of fungal communities overtime has revealed that delivery mode influences colonization by Candida species, where the relative abundances of Candida albicans is higher on the skin of infants delivered vaginally while Candida orthopsilosis is higher in the oral cavity of C-section-born infants over the first 30 days of life [9]. However, a study of term and preterm neonates examined clinical and host determinants that may influence the skin mycobiome [10] and found Malassezia, Candida, Cladosporium, Fusarium, and Cryptococcus were the most abundant genera and did not differ between the groups but fungal alpha and beta diversity varied with both clinical and host factors [10]. This suggests initial colonization by birth mode is more important than gestational age at birth. Conversely, constant changes in the skin physiology and a rapidly evolving infant microbiome over the first few months of life contribute to skin disorders, such as infant dermatitis (i.e., diaper rash). In a clinical study examining healthy infants and those with diaper dermatitis, C. albicans was more abundant in areas with skin-to-skin contact, whereas Cladosporium and Alternaria were more abundant in less occluded areas. Candida abundance directly correlated with disease severity [11]. Together, these findings reveal that shifts in clinical and host factors and skin microenvironments directly shape the skin mycobiome and dynamics during early life development.

To understand how the skin microbiome transitions towards adulthood, longitudinal studies have examined puberty-associated shifts [12]. Significant shifts in microbial community composition are associated with Tanner sexual maturation stages (Figure 1). Further, microbial changes appear to be sex specific with Cutibacterium and Malassezia as the dominant taxa in female children, whereas Corynebacterium, Staphylococcus, Streptococcus, and Haemophilus are the dominant taxa in male children. Fungal communities between sexes exhibited no significant differences, however sebaceous skin showed tighter clustering that is driven by Malassezia in female children compared to male children. Lipophilic species, including Cutibacterium and Malassezia, also exhibit strong association with several serum sex hormones [12], suggesting that the microbial transitions during puberty are mediated by sex hormone concentrations that subsequently increase sebaceous gland activity. Skin aging also influences the microbiome. It was recently reported that fungal communities on the forehead and cheek are significantly different and impacted by age (Figure 1) when including healthy young (19–28 years old) and older (60–63 years old) Korean women [13]. Further, the younger group exhibited higher network densities on the cheek and associations were found between low-abundance bacteria and fungi, including M. sympodialis, Gemella, and Methylobacterium, indicating the potential role of these species in aging. Overall, throughout the life span, the skin fungal and bacterial communities are dynamic and are influenced by numerous factors, working harmoniously to form a stable and homeostatic skin barrier. Due to the ease by which skin can be repeatedly sampled across diverse microenvironments, continued study of fungal-bacterial behaviors on the skin during normal human development presents an immense opportunity towards understanding species interactions in a community context and their role in host development. Furthermore, general principles gleaned from the cutaneous ecosystem can be generally applied to other host-associated microbial communities.

Figure 1.

Overview of fungal colonization throughout the lifespan. Fungal colonization is dependent on mode of delivery. Fungal composition is diverse during early life, and as an individual ages, fungal diversity decreases. During puberty, relative abundance of lipophilic taxa (i.e., Malassezia and Cutibacterium) gradually increased as a result of increased sex hormone concentrations, including estradiol, estrone, and testosterone, and sebaceous gland activity. Transitioning towards adulthood, the skin microbiome becomes dominated by Malassezia, Cutibacterium, Staphylococcus, and Corynebacterium. Finally, aging causes skin physiological changes, and thus fungal diversity varies depending on the skin region.

Skin Associated Fungi and Dermatological Diseases

Although overall fungal phylogenetic diversity on the skin is low, shifts in fungal abundance and expansion have been associated with numerous dermatological diseases. Malassezia species are the most prevalent fungal taxa found on human skin, acquiring lipids exogenously from skin secretions and generally living in a commensal state. However, this is context-dependent with several skin disorders associated with Malassezia, the most common being pityriasis versicolor and seborrheic dermatitis. The causal role of Malassezia in skin barrier diseases such as atopic dermatitis (AD) and psoriasis is less clear. Certain species including M. sloofiae, obtusa, and yamatoensis were found to be exclusively associated with AD samples along with an increase in the overall fungal diversity compared to healthy controls [4]. This is in contrast to a loss of bacterial diversity in AD patients, usually driven by the expansion of Staphylococcal bacteria species, i.e. S. aureus [5]. Thus, in AD, increased prevalence of Malassezia may simply be a consequence of disrupted bacterial communities. Concurrently, Malassezia will aggravate cutaneous inflammation through IL-23 and IL-17 dependent mechanisms only under conditions where skin integrity is impaired, such as during an AD flare [6]. The observation that fungal diversity increases as overall bacterial diversity decreases suggests a dynamic, yet stable ecosystem composed of fungal-bacterial networks on the skin. Fungal expansion may drive inflammatory responses during a skin barrier breach. The mechanisms governing these interactions remain largely unknown and represent an important area of future research.

Although Malassezia dominates human skin, Candida spp. are opportunistic fungal pathogens also commonly found on human skin. The presence of fungi in chronic wounds such as diabetic foot ulcers can have major consequences. We have shown that fungi are significantly associated with tissue necrosis and amputation. They can also serve as a scaffold for colonizing bacteria to bind and develop biofilms [14,15]. Skin-associated bacterial species have been shown to directly interact with C. albicans through cell surface macromolecules, such as fungal agglutin-like sequence (Als) proteins targeting Staphylococcal spp. [16]. We recently demonstrated that bacterial mannose binding lectins (fim operon) expressed by a wound isolate of Citrobacter freundii mediate attachment to the fungal cell wall [15]. We investigated the growth dynamics of C. albicans with C. freundii and observed an increase in fungal filamentation during biofilm formation [15]. C. albicans biofilm extracellular matrix can inhibit the release of neutrophil extracellular traps, or NETs, which are web-like structures consisting of DNA and antimicrobial proteins [17]. When human neutrophils are exposed to C. albicans biofilms alone or in combination with C. freundii, a significant increase in NET release under the co-culture exposure is observed [15]. Together, this suggests the fungal-bacterial interaction mediates the host inflammatory response. Thus, interkingdom interaction can result in emergent traits that influence pathogenesis. Identifying the signals driving these interactions will provide a framework to develop novel antimicrobial strategies targeted at polymicrobial infections.

The Emerging Pathogen Candida auris

In recent years, the fungal pathogen Candida auris has made headlines for its rapid emergence and alarming drug resistance profile. Multi-drug resistant and highly transmissive strains are commonly isolated from patients within care settings. Asymptomatic skin colonization is considered the primary mode of carriage, dissemination of C. auris, and a risk factor for subsequent bloodstream infection. C. auris can persistently colonize skin and environmental surfaces for long periods of time. Nett and colleagues used an ex vivo skin model to show that sweat and sebum secretions further enhance colonization and biofilm formation of C. auris on the skin surface compared to other Candida species [18]. Thus, decolonization, using antiseptics, represents a first line of defense to protect against invasive infection. Studies have demonstrated that C. auris colonizes mammalian skin better than C. albicans and has the ability to reside on the surface and within deeper tissue layers [18,19]. Strains from distinct clades can be further differentiated by their ability to colonize skin from diverse species including mice, pigs, and humans [18,19]. Studying this emerging pathogen to identify clade-specific signatures will permits reconstruction of traits conferring enhanced virulence and drug resistance on the skin though an evolutionary lens.

C. auris has been hypothesized to have emerged from marine environments. Strains related to clinical isolates have been found on the sandy beach and tidal swamp in the Andaman Islands, however they do not exhibit the same drug resistance profile of human-associated strains. A recent study screened seasonal tropical fruit as a potential reservoir of environmental C. auris and found the surfaces of stored apples, as opposed to freshly harvested apples, were colonized with high levels of antifungal resistant C. auris [20]. Apples are often treated with wax and agricultural fungicides acting as a potential source of selection pressure for antifungal resistance that could then disseminate into human populations. It is possible C. auris was transferred between fruit and skin of individuals during processing, raising important future research questions regarding the role of agricultural fungicides and food chain dynamics in the spread of emerging pathogens. Further, it remains unclear whether the healthy skin microbiome plays a role in host susceptibility to C. auris colonization, particularly in vulnerable older populations. One of the most common types of fungal infections worldwide is dermatophytic infection, affecting the skin and its appendages. Topical azoles, one of the largest antifungal groups, are often used for dermatophytoses. Studies have elucidated some possible mechanisms of azoles resistance in dermatophytes, including drug efflux, drug target modification, and upregulation of stress response [21]. The reliance of azoles to treat superficial mycoses has the potential to drive the emergence of drug resistance in C. auris in the skin environment. Further investigation into the use of topical antifungals and whether associations exist with the carriage C. auris is a critical gap that should be addressed.

Non-human Skin Mycobiome

Although the human skin microbiome is not as well studied as the gut microbiome, significant advances have been made in the study of non-human skin microbiomes. The amphibian skin microbiome has been best characterized within the last decade largely driven by conservation strategies to prevent species extinction due to devastating fungal infection. In addition to colonization resistance, it is also important to consider host-microbe interactions and their impact on the host’s health. Characterizing fungal-host associations, a recent study examined transition of the amphibian fungal community during host metamorphosis on the Colorado boreal toad [22]. From ITS fungal sequences, communities significantly varied across life stages of the toads and shared taxa between the host and habitat were observed. Interestingly, host-specific taxa were only observed during the egg life cycle, a sensitive period in the life cycle of this animal. As in other animals, fungal communities stabilize through maturity [22], highlighting similarities in the fungal communal shifts during maturation processes across different host species. In fish, fungi colonize at low abundance compared to bacteria, and the fungal community structure is clade-specific, where each clade harbors unique fungal communities. Further, cartilaginous fish show patterns of phylosymbiosis, while bony fish do not, which could stem from clade-specific microbial selection and maintenance [23]. Establishing a stable and unique skin microbiome could be impacted by changing ocean conditions and biological factors, such as sea surface temperature, chlorophyll a, and fish age, that has been shown in wild fish like the Pacific chub mackerel [24]. Unlike the human skin microbiome which exhibits temporal stability regardless of perturbations [25,26], the skin microbiome of vertebrates is affected by habitat, geographic location, season, and disease state. The effort of sampling vertebrate skin microbiota along with lifestyle and environmental changes will be crucial for future conservation strategies and broaden our view on how these key elements drive ecological functions across different hosts and environments (Table 1).

Table 1.

Summary of key literature focused on fungi in the skin microbiome.

Host	Authors, date	Summary	Reference
Human	Findley et al., 2013	Surveyed fungal community based on topography of the human skin.	[41]
	Oh et al., 2013	Shift in the bacterial and fungal communities in patients with primary immunodeficiencies.	[42]
	Oh et al., 2014	The first multi-site metagenomic study of the human skin microbiome.	[3]
	Findley and Grice, 2014	Review of bacterial and fungal pathogens and their roles in skin disease.	[43]
	Takemoto et al., 2015	Molecular survey of skin fungal microbiota in patients with psoriasis.	[44]
	Oh et al., 2016	Explored temporal stability of bacteria, fungi and viruses on human skin.	[25]
	Leung et al., 2016	Characterization of the skin mycobiome of healthy, urban Chinese individuals.	[45]
	Johnson et al., 2016	C. albicans biofilms interactions with host neutrophil response.	[17]
	Ward et al., 2018	Temporal and topographical development of human mycobiome during the first month of life.	[9]
	Tipton et al., 2018	Fungi stabilize community network in the skin microbial ecosystems.	[46]
	Han et al., 2018	Analysis of the skin mycobiome in patients with atopic dermatitis.	[4]
	Sparber et al., 2019	The role of Malassezia spp. in regulating antifungal immunity and promoting skin inflammation in inflammatory skin disorders.	[6]
	Paul et al., 2020	Skin fungal communities of preterm and term neonates.	[10]
	Flowers and Grice, 2020	A review of colonization resistance in the human skin microbiome, highlighting the balance between risk and reward of commensal skin bacteria.	[2]
	Horton et al., 2020	Sweat and sebum secretions enhanced colonization and biofilm formation of C. auris on the skin surface compared to other Candida species using an ex vivo porcine skin model.	[18]
	Teufel et al., 2021	Comparison of skin mycobiome between healthy and damage skin in infants with diaper dermatitis.	[11]
	Barber and Zhang, 2021	A review of small molecules produced by member of the human nasal/oral microbiome.	[34]
	Cheong et al., 2021	Priority effects and competitive exclusion on biofilm community structure in a community comprised of bacteria and fungi isolated from chronic wounds.	[15]
	Huang et al., 2021	Murine skin model to investigate C. auris colonization, host risk factors, and potential therapies.	[19]
	Park et al., 2022	Transitions of skin fungal and bacterial communities in healthy children during puberty.	[12]
	Yadav et al., 2022	Stored fruits as an environment contributing to development and spread of antifungal resistance C. auris.	[20]
	Kim et al., 2022	Age related changes in the skin microbiome and mycobiome in Korean women.	[13]
Animal	Kearns et al., 2017	Explored antifungal properties of amphibian skin mycobiome.	[47]
	2018–2021	Snake skin bacterial and fungal communities.	[48–50]
	Chermprapai et al., 2019	Comparison between healthy and atopic dermatitis dogs and their bacterial and fungal microbiota.	[51]
	Niederle et al., 2019	Skin-associated lactic acid bacteria from bullfrogs as potential biocontrol agents for fungal pathogens.	[37]
	Greenspan et al., 2019	Environmental effects on skin microbial assemblages and subsequent effects on host resistance to fungal pathogens.	[38]
	Woodhams et al., 2020	Modulations of amphibian skin defensive antifungal peptide and antifungal bacteria growth by probiotics.	[40]
	2020	Skin fungal microbiome of fish.	[23,24]
	Vanderwolf et al., 2021	Identifying fungal assemblages and key species based on susceptibility to white-nose syndrome in bats.	[36]
	Alexiev et al., 2021	Survey of skin fungal communities throughout metamorphic toad life stages.	[22]
	2012–2019	Antifungal metabolites derived from amphibian skin microbiome.	[52–56]

Colonization Resistance in the Skin Microbiome

One function of the skin is to block pathogenic invasion. The diversity-invasibility hypothesis states that more diverse communities are more resistant to invasion than poorly diverse communities [27–29]. Resistance mechanisms against invading pathogens include production of antimicrobials [30–32], priming of the immune system [1], and utilization of metabolic niches [33]. Novel bioactive metabolites have been discovered from distinct locations on the human body. For example, in the oral microbiome, mutanobactin, mutanamide, and farnesol regulate C. albicans growth and development [34]. On the skin, bacterial-bacterial competition via metabolite production is well described between skin residents and pathogenic S. aureus [2,30–32], while the fungus M. globosa has been shown to secrete a protease, cleaving a major virulence factor of S. aureus and subsequently disrupting biofilm formation [2]. However, examples of bacterial antagonism towards fungi on human skin are lacking. This is somewhat surprising given the propensity of the skin microbiome to demonstrate antifungal colonization resistance in other animals; a recent example is in bats susceptible to white-nose syndrome (WNS) caused by the fungus Pseudogymnoascus destructans, a dermatophyte that manifests a ‘vampire-like’ lifestyle due to the lack of UV repair mechanisms [35,36]. Examining the skin microbiome in these animals showed that fungal community structure did not differ based on WNS-susceptibility, but several fungal species were differentially abundant on WNS-resistant hosts. Further, the yeast species Cutaneotrichosporon moniliiforme was found to inhibit P. destructans in vitro [36], highlighting the role of key members in the skin fungal assemblages in modulating disease resistance and colonization of pathogens.

Skin bacterial assemblages also contribute to colonization resistance in amphibians. For example, lactic acid bacteria that live on the skin of bullfrogs (i.e. Enterococcus gallinarum), have been shown to produce a wide range of anti-Batrachochytrium dendrobatidis (Bd) factors, the causative agent of the devastating fungal disease chytridiomycosis [37]. Further, environmental bacterial diversity and bioclimatic factors, such as temperature, precipitation, and elevation shape microbial community and structure of the skin microbiome, which in turn, are correlated to Bd infection intensity [38,39]. Metabolite-producing bacteria can also tune the host microbiome to dial up secondary defense mechanisms. For example, a novel skin defense peptide brevinine-1Ma was downregulated in probiotic-treated frogs, promoting growth of bacteria that had inhibitory activity against Bd [40]. Thus, to maintain and optimize colonization resistance, the host must acquire key beneficial bacterial and fungal taxa that engage in mutualistic and antagonistic interactions, while the environmental conditions must be preserved. Clear evidence is emerging of defensive symbiosis via production of antifungal factors in mammalian and amphibian skin microbiomes. Thus, the complexity and intricacy of the human skin microbiome makes it an excellent niche to discover novel bacterial-fungal interactions and small molecules that mediate these networks. Building off work in amphibians to explore bacterial antagonism towards fungi on human skin is an exciting direction of research to expedite the development of novel therapeutic drugs with minimal toxicity for superficial fungal infections.

Conclusions and Future Outlook

While the skin microbiome is composed of a large percentage of bacteria, fungi are key members participating in core functions related to the development, maintenance, and repair of the skin in vertebrate hosts. For the human skin microbiome, fungal communities are less diverse but universally present and change along with skin physiology and age. The question remains whether the bacterial communities contribute to the structure of the mycobiome including limiting pathogen expansion and promoting a less diverse community as one transitions into adulthood. It is likely that these changes in the skin fungal community serve an important ecological role. Understanding these shifts will provide insights into how the human host resists colonization of potential fungal invaders, a topic that has been well-studied in the context of conservation strategies for animals. Discovering these interactions could provide deeper insight into crucial players in the skin microbial community and their ecological functions. Potentially, this effort will lead to novel therapeutics and improve fungi-related dermatological outcomes.