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The impact of the Fungus-Host-Microbiota interplay upon Candida albicans infections: current knowledge and new perspectives


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Candida albicans is a major fungal pathogen of humans. It exists as a commensal in the oral cavity, gut or genital tract of most individuals, constrained by the local microbiota, epithelial barriers and immune defences. Their perturbation can lead to fungal outgrowth and the development of mucosal infections such as oropharyngeal or vulvovaginal candidiasis, and patients with compromised immunity are susceptible to life-threatening systemic infections. The importance of the interplay between fungus, host and microbiota in driving the transition from C. albicans commensalism to pathogenicity is widely appreciated. However, the complexity of these interactions, and the significant impact of fungal, host and microbiota variability upon disease severity and outcome, are less well understood. Therefore, we summarise the features of the fungus that promote infection, and how genetic variation between clinical isolates influences pathogenicity. We discuss antifungal immunity, how this differs between mucosae, and how individual variation influences a person's susceptibility to infection. Also, we describe factors that influence the composition of gut, oral and vaginal microbiotas, and how these affect fungal colonisation and antifungal immunity. We argue that a detailed understanding of these variables, which underlie fungal-host-microbiota interactions, will present opportunities for directed antifungal therapies that benefit vulnerable patients.
A combination of virulence factors and fitness attributes promote C. albicans virulence. Polymorphism: The ability of C. albicans to undergo morphological transitions allows it to adapt to different growth conditions, adhere to biotic and abiotic surfaces, invade cells and tissue, and escape from immune cells. Invasion and damage: A combination of induced endocytosis and active penetration promote fungal invasion of host tissues, and the accumulation of the toxin, candidalysin, in the invasion pocket leads to pore formation and host cell damage. Adhesion/biofilm formation: The battery of adhesins promotes fungal adhesions to biological and abiotic surfaces, which can lead to the development of biofilms, for example on medical devices such as catheters. Genetic and metabolic plasticity: Candida albicans displays a high degree of metabolic flexibility, which allows it to adapt rapidly to diverse host niches. This fungus also displays great genetic plasticity, which permits rapid evolutionary adaptation to selective pressures and stresses such as exposure to antifungal drugs. Stress responses: Candida albicans activates robust stress responses following exposure to host imposed stresses, including ROS and RNS, which enhances fungal survival following immune attack, for example. Cell wall: As well as maintaining cell morphology, the robust cell wall provides protection against host-imposed stresses including changes in osmolarity. Immune evasion: Candida albicans has evolved a variety of immune evasion strategies that include the modulation of PAMP exposure at the cell surface to evade immune recognition, and phagocytic escape mechanisms to evade killing by innate immune cells. See text.
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FEMS Microbiology Reviews, fuaa060, 44, 2020, 1
doi: 10.1093/femsre/fuaa060
Advance Access Publication Date: 0 2020
Review article
The impact of the Fungus-Host-Microbiota interplay
upon Candida albicans infections: current knowledge
and new perspectives
Christophe d’Enfert1,*,, Ann-Kristin Kaune2, Leovigildo-Rey Alaban3,4,
Sayoni Chakraborty5,6, Nathaniel Cole7, Margot Delavy1,4, Daria Kosmala1,4,
ıt Marsaux8,9,RicardoFr
ois-Martins10,11, Moran Morelli12,
Diletta Rosati13,, Marisa Valentine5, Zixuan Xie14, Yoan Emritloll1, Peter
eric Bequet3, Marie-Elisabeth Bougnoux1, Stephanie Bornes16,
Mark S. Gresnigt5, Bernhard Hube5,IlseD.Jacobsen
elanie Legrand1,
e Leibundgut-Landmann10,11, Chaysavanh Manichanh14,Carol
A. Munro2, Mihai G. Netea13, Karla Queiroz12, Karine Roget17,
Vincent Thomas3, Claudia Thoral17, Pieter Van den Abbeele8,Alan
W. Walker7and Alistair J.P. Brown18,*
e Biologie et Pathog´
e Fongiques, D´
epartement de Mycologie, Institut Pasteur, USC 2019 INRA, 75015
Paris, France, 2Aberdeen Fungal Group, Institute of Medical Sciences, University of Aberdeen, Foresterhill,
Aberdeen AB25 2ZD, UK, 3BIOASTER Microbiology Technology Institute, 40 avenue Tony Garnier, 69007 Lyon,
France, 4Universit´
e de Paris, Sorbonne Paris Cit´
e, Paris, France, 5Microbial Immunology Research Group, Emmy
Noether Junior Research Group Adaptive Pathogenicity Strategies, and the Department of Microbial
Pathogenicity Mechanisms, Leibniz Institute for Natural Product Research and Infection Biology – Hans Kn ¨
Institute, Beutenbergstraße 11a, 07745 Jena, Germany, 6Institute of Microbiology, Friedrich Schiller University,
Neugasse 25, 07743 Jena, Germany, 7Gut Microbiology Group, Rowett Institute, University of Aberdeen,
Foresterhill, Aberdeen AB25 2ZD, UK, 8ProDigest, Technologiepark 94, B-9052 Gent, Belgium, 9Center for
Microbial Ecology and Technology (CMET), Department of Biotechnology, Faculty of Bioscience Engineering,
Ghent University, 9000 Ghent, Belgium, 10Immunology Section, Vetsuisse Faculty, University of Zurich,
Winterthurerstrasse 266a, Zurich 8057, Switzerland, 11Institute of Experimental Immunology, University of
Zurich, Winterthurerstrasse 190, Z¨
urich 8057, Switzerland, 12Mimetas, J.H. Oortweg 19, Leiden, The
Netherlands, 13Department of Internal Medicine and Radboud Center for Infectious Diseases, Radboud
University Medical Center, Geert Grooteplein 28, 6525 GA Nijmegen, The Netherlands, 14Gut Microbiome
Group, Vall d’Hebron Institut de Recerca (VHIR), Vall d’Hebron Hospital Universitari, Vall d’Hebron Barcelona
Received: 3 October 2020; Accepted: 18 November 2020
2020. Published by Oxford University Press on behalf of FEMS. This is an Open Access article distributed under the terms of the Creative Commons
Attribution License (, which permits unrestricted reuse, distribution, and reproduction in any medium,
provided the original work is properly cited.
2FEMS Microbiology Reviews 2020, Vol. 00, No. 00
Hospital Campus, Passeig Vall d’Hebron 119–129, 08035 Barcelona, Spain, 15Magic Bullet Consulting, London,
WC2H 9JQ, UK, 16Universit ´
e Clermont Auvergne, INRAE, VetAgro Sup, UMRF, 15000 Aurillac, France, 17Biose, 22
ee Alan Turing, 63000 Clermont-Ferrand, France and 18MRC Centre for Medical Mycology, University of
Exeter, Geoffrey Pope Building, Stocker Road, Exeter EX4 4QD, UK
Corresponding authors: Christophe d’Enfert: Unit ´
e Biologie et Pathog´
e Fongiques, D´
epartement de Mycologie, Institut Pasteur, 25–28 rue du
Docteur Roux, 75015 Paris, France. E-mail:; Alistair J P Brown: MRC Centre for Medical Mycology, University of Exeter,
Geoffrey Pope Building, Stocker Road, Exeter EX4 4QD, UK. E-mail:
One sentence summary: The complexity and variability of FunHoMic interactions between the fungal pathogen, its human host and the Microbiota
strongly inuence the development and outcomes of the supercial and systemic Candida albicans infections that plague human health worldwide.
Editor: Bart Thomma
Christophe d’Enfert, 6235-3886
Diletta Rosati, 2992-2503
§Alistair J.P. Brown, 1406-4251
Candida albicans is a major fungal pathogen of humans. It exists as a commensal in the oral cavity, gut or genital tract of
most individuals, constrained by the local microbiota, epithelial barriers and immune defences. Their perturbation can lead
to fungal outgrowth and the development of mucosal infections such as oropharyngeal or vulvovaginal candidiasis, and
patients with compromised immunity are susceptible to life-threatening systemic infections. The importance of the
interplay between fungus, host and microbiota in driving the transition from C. albicans commensalism to pathogenicity is
widely appreciated. However, the complexity of these interactions, and the signicant impact of fungal, host and
microbiota variability upon disease severity and outcome, are less well understood. Therefore, we summarise the features
of the fungus that promote infection, and how genetic variation between clinical isolates inuences pathogenicity. We
discuss antifungal immunity, how this differs between mucosae, and how individual variation inuences a person’s
susceptibility to infection. Also, we describe factors that inuence the composition of gut, oral and vaginal microbiotas, and
how these affect fungal colonisation and antifungal immunity. We argue that a detailed understanding of these variables,
which underlie fungal-host-microbiota interactions, will present opportunities for directed antifungal therapies that benet
vulnerable patients.
Keywords: Candida;Candida infections; antifungal immunity; microbiota; mycobiota; fungus-host-microbiota interactions;
patient variability; fungal variability; microbiota variability
Fungal pathogens have a major global impact upon human
health. Estimates suggest that, at any given time, over a quar-
ter of the world’s population have a fungal infection of the skin,
that 75% of women suffer at least one episode of vulvovaginal
candidiasis during their lifetime, and that over a million peo-
ple die each year from an invasive fungal infection (Brown et al.
2012). Mortality rates for those suffering systemic fungal infec-
tions are unacceptably high, reaching 50% in many cases. This
is because fungal infections are often difcult to diagnose, and
are particularly challenging to treat (Perlroth, Choi and Spell-
berg 2007;Brownet al. 2012;K¨
ohler, Casadevall and Perfect 2014).
There is a clear and urgent medical need for more accurate
diagnostics, for safer and more effective antifungal drugs, and
for host-directed therapies. The search for antifungal drug tar-
gets is somewhat constrained by the fact that, as eukaryotes,
fungi share fundamental mechanisms of cell growth and divi-
sion with humans. The search for diagnostic markers that can
distinguish infection from fungal commensalism is especially
challenging. Therefore, the development of potent new clinical
tools is dependent upon a comprehensive understanding of fun-
gal pathogenicity and antifungal immunity.
Candida species are amongst the top fungal killers (Brown
et al. 2012). Of these, Candida albicans remains the most com-
mon cause of life-threatening systemic candidiasis, although
the frequent prophylactic use of azole antifungal drugs has led
to the emergence of other Candida species with intrinsic resis-
tance to these drugs (Nguyen et al. 1996; Silva et al. 2012; Chowd-
hary, Sharma and Meis 2017). Nevertheless, in this review we
focus on C. albicans, because a combination of three main factors
arguably makes this species unique amongst fungal pathogens:
(a) its lifestyle as both a commensal and potent pathogen; (b)
the range and frequency of infections that it causes; and (c) its
pathobiology has been studied in greater depth than most other
fungal pathogens.
Candida albicans is an opportunistic pathogen that exists as
a commensal in most individuals, and is a frequent cause of
mucosal and systemic infections (See The Fungus). Unlike most
fungal pathogens, C. albicans is generally considered to be obli-
gately associated with warm-blooded animals (Odds 1988). Envi-
ronmental isolates of C. albicans continue to be reported (Bensas-
son et al. 2019;Macielet al. 2019; Opulente et al. 2019). However,
although the existence of an environmental reservoir cannot be
excluded, it is apparently not necessary for human colonisation.
Candida albicans is transmitted vertically from mother to
child, and infections arise predominantly from the endogenous
microbiota rather than other sources (d Enfert 2009; Miranda
et al. 2009;Zhaiet al. 2020) (see The Microbiota). This contrasts
with other major pathogens such as Aspergillus, Cryptococcus
and Histoplasma species,which are fundamentally environmen-
tal fungi that have evolved traits that promote pathogenicity in
humans, possibly through their transient passage in niches that
have similarities with those encountered in the human host, for
d’Enfert et al. 3
Figure 1. Three-way interactions between the fungus, the host and the local
microbiota strongly inuence the likelihood and severityof C. albicans infections.
See text.
example, their association with rodents or contact and evasion
of amoebic predation in the environment (Steenbergen, Shu-
man and Casadevall 2001; Malliaris, Steenbergen and Casade-
vall 2004; Van Waeyenberghe et al. 2013; Hillmann et al. 2015).
Pneumocystis jirovecii is obligately associated with humans, but
this major pathogen differs from C. albicans in that it is unable
to thrive outside its host (Liu, Fahle and Kovacs 2018). Con-
sequently, key aspects of Pneumocystis jirovecii biology remain
unexplored. The lifestyle of C. albicans even differs considerably
from its distant cousin, C. (Brunke and Hube 2013;Kasper,Sei-
der and Hube 2015). Genetic evidence suggests that, although it
is often presumed to be a human commensal such as C. albicans,
C. glabrata seems to be only secondarily associated with humans
and is likely to have environmental reservoirs (Gabald ´
on and
Fairhead 2019).
The biology, epidemiology, pathogenicity and immunology
of C. albicans have been studied in greater depth than for any
other fungal pathogen. This depth of knowledge provides a
strong platform for studies of the relationships between the
fungal pathogen, host immunity and local microbiota that lie
at the heart of fungal infection (Casadevall and Pirofski 1999,
2003,2015;Jabra-Rizket al. 2016)(Fig.1). Other major fungal
pathogens infect humans by different routes to C. albicans,but
many principles that are emerging for C. albicans may be applica-
ble to these pathogens. Therefore, we present underlying princi-
ples of C. albicans colonisation and infection, antifungal immune
defences, and the protective properties of the local microbiota
in the gastrointestinal (GI) tract, oral cavity and vagina. We also
address the variability that inuences the Fungus-Host-Microbiota
interplay and how this impacts infection. A detailed under-
standing of this tripartite interplay is essential to optimise ther-
apeutic strategies for individual patients (d Enfert 2009; Pirofski
and Casadevall 2020).
C. albicans commensalism and pathogenicity
C. albicans frequently inhabits the oral, vaginal and GI mucosa of
healthy individuals as a harmless commensal (Ghannoum et al.
2010;Drellet al. 2013; Nash et al. 2017)(Fig.2). Indeed, C. albicans
is present on the mucosa of most people in most human pop-
ulations (Neville, d Enfert and Bougnoux 2015; Prieto et al. 2016;
Mishra and Koh 2018). However, this fungus can cause infections
if the local microbiota becomes perturbed, normal tissue barri-
ers are weakened or immune defences become compromised.
Mucosal infections, characterised by fungal colonisation (i.e.
overgrowth) associated with an inammatory host response, are
extremely common and can have a major impact upon the qual-
ity of life for many individuals (Fig. 2). For instance, most women
of reproductive age (75%) will experience at least one episode of
VVC (‘thrush’) in their lifetime, and up to 9% suffer from recur-
rent VVC, as dened by multiple episodes of vaginitis per annum
(Foxman et al. 2013;Yanoet al. 2019; Rosati, Bruno, Jaeger, Ten
Oever et al. 2020). Risk factors for VVC include high estrogen
levels, the use of oral contraceptives and uncontrolled diabetes.
However, episodes can be idiopathic (i.e. of unknown cause) and
VVC, unlike oral candidiasis, can occur in apparently healthy
individuals (see Innate antifungal responses).
Oropharyngeal candidiasis (OPC) can broadly be classied
into three main conditions, namely acute, chronic and chronic
mucocutaneous candidiasis syndromes (Vila et al. 2020)(Fig.2).
Predisposing factors include nutritional deciencies, local dys-
biosis, salivary hypo-function, smoking, wearing dentures and
dysfunctional T-cell immunity due to genetic alterations or
other infections. Indeed, OPC is the most frequently diag-
nosed oral opportunistic infection in HIV-positive individuals
and many acute cases are caused by broad-spectrum antibiotic
treatments (Samaranayake 1992; Vila et al. 2020).
Life-threatening systemic C. albicans infections can arise
when the fungus enters the bloodstream (Fig. 2). Candidaemia
is the fourth most common nosocomial bloodstream infection
in North America (Pfaller and Diekema 2010), but the incidence
of invasive candidiasis in European countries is generally lower
(Meyer et al. 2013; Yapar 2014). The presence of a central venous
catheter, dialysis, antibiotic treatment, lengthy stays in inten-
sive care units (ICUs), recent major surgery, and receiving total
parenteral nutrition are among the predisposing factors for sys-
temic candidiasis (Pappas et al. 2018). Most disseminated infec-
tions arise from Candida escaping the patient’s own GI tract
(Miranda et al. 2009; Gouba and Drancourt 2015;Zhaiet al. 2020).
Systemic infections arise when host defences are compromised
by, for example, damage to the intestinal barrier (e.g. surgery
or trauma), medically induced immunosuppression (corticos-
teroids or chemotherapy-induced neutropenia), or the use of
broad-spectrum antibiotics (Pappas et al. 2018). A combination
of these factors is typically needed to allow C. albicans to translo-
cate from the gut (Koh et al. 2008; Papon, Bougnoux and d Enfert
2020). Once in the blood, C. albicans can disseminate to almost
all organs including kidney, liver, and spleen (Pappas et al. 2018).
The mortality rate for these infections, which varies across geo-
graphical regions, is reported to lie between 10% and 47% despite
the availability of antifungal therapies (Brown et al. 2012). This
is unacceptably high.
Clearly, knowledge about the factors and conditions that pro-
mote C. albicans commensalism or opportunism is important for
an understanding of the mechanisms that underlie the tran-
sition from commensalism to pathogenicity. Much work has
4FEMS Microbiology Reviews 2020, Vol. 00, No. 00
Figure 2. Sites of C. albicans commensalism and disease on the human body.Sites of C. albicans commensalism (left side) include the oral cavity, gastrointestinal tract
(gut) and the genital tract. C. albicans can infect these sites (right side) to cause oropharyngeal or vulvovaginal candidiasis. C. albicans can also cause systemic infections
of the blood and internal organs, which often arise via translocation of C. albicans from the gut into the bloodstream. Candida albicans also causes mucocutaneous
infections of the skin and nails. Factors that predispose individuals to such infections are listed. See text.
focussed on the virulence factors and tness attributes that
promote C. albicans infection (see Virulence Factors and Fitness
attributes). However, the pathogenesis of C. albicans also depends
on the host site of colonisation (Fidel et al. 2020). Candida albi-
cans asymptomatically inhabits the oral mucosa and only causes
infection when host defences are weakened. In contrast, C. albi-
cans is an immunoreactive coloniser during vulvovaginal infec-
tion, eliciting host damage via a hyperactive immune response.
Meanwhile, systemic infections are mostly nosocomial and are
generally associated with predisposing conditions. The fungus
is able to cause these different types of infection by tuning
the expression of its arsenal of virulence factors and tness
attributes to the local niche.
Virulence factors
Cellular polymorphism
The polymorphic nature of C. albicans is integral to both com-
mensalism and pathogenesis. This fungus is able to switch
reversibly between different growth forms and morphologies
(Noble, Gianetti and Witchley 2017)(Fig.3). Depending upon
the environmental conditions, C. albicans can grow as unicellu-
lar yeast cells, pseudohyphae, or true hyphae that lack invagi-
nations at septal junctions (Sudbery, Gow and Berman 2004).
Also, depending on the presence of certain environmental cues,
C. albicans can undergo phenotypic switching to interchange
reversibly between white, grey and opaque phenotypes, each
of which displays distinct yeast cell and colony morphologies,
and gene expression proles. Furthermore, a gastrointestinally
induced transition (GUT) phenotype has been described for C.
albicans cells that ectopically overexpress the Wor1 regulator
which, together with Efg1, controls white-grey-opaque switch-
ing (Pande, Chen and Noble 2013). Phenotypic switching is a
strictly regulated process that seems to be associated with com-
mensalism, host niche adaptation, mating, immune evasion and
virulence (Miller and Johnson 2002; Morschh¨
auser 2010; Pande,
Chen and Noble 2013; Xie et al. 2013;Taoet al. 2014). Finally,
C. albicans can differentiate to form chlamydospores, enlarged
thick-walled cells, under nutrient limitation, low temperature
and microaerophilia (Staib and Morschh¨
auser 2007;B
et al. 2016)(Fig.3).
Both yeast and hyphal morphologies are necessary for the
full virulence of C. albicans (Lo et al. 1997; Murad, Leng, et al.
2001; Saville et al. 2003; Jacobsen et al. 2012)(Fig.4). However,
it is generally thought that yeast cells are well suited to dis-
semination, and hyphal cells to tissue invasion (Gow, Brown and
Odds 2002). The yeast-to-hypha transition is accompanied by an
extensive change in gene expression prole, in cell wall struc-
ture, and by the expression of many virulence factors (Jacobsen
et al. 2012; Mayer, Wilson and Hube 2013; Chen et al. 2020). The
change in morphology can be triggered by many environmen-
tal factors present in host niches, such as physiological temper-
atures (>36C), starvation, an ambient pH of >7, the presence
of serum, N-acetylglucosamine, or elevated CO2levels (Mayer,
Wilson and Hube 2013). Furthermore, hyphal development is
triggered by the bacterial cell wall component, peptidoglycan
(Xu et al. 2008), which is of particular relevance to fungus-host-
microbiota interactions. Not surprisingly given the complexity
of environmental inputs and cellular outputs, yeast-hypha mor-
phogenesis is regulated by a complex signalling network that
includes the cAMP-protein kinase A, Efg1, Cph1, Czf1, Hog1 and
Nrg1 pathways (Basso et al. 2019; Kadosh 2019; Kornitzer 2019).
d’Enfert et al. 5
Figure 3. Candida albicans is polymorphic, displaying a range of cellular growth forms.C. albicans yeast cells can undergo phenotypic switching between white, grey
and opaque growth forms that present with different shapes and cell surface characteristics (Gow, Brown and Odds 2002; Sudbery, Gow and Berman 2004;Xuet al.
2008; Huang et al. 2009; Mayer, Wilson and Hube 2013;Taoet al. 2014;Sunet al. 2015). These forms are induced in response to different environmental inputs, and
hence are associated with different types of infection (Gow, Brown and Odds 2002). Signicantly, the opaque form is associated with efcient mating in C. albicans
(Miller and Johnson 2002),with grey cells displaying an intermediate mating competence between opaque and white cells (Tao et al. 2014). The gastrointestinally
induced transition (GUT) phenotype is observed in C. albicans cells that ectopically express WOR1 (Pande, Chen and Noble 2013), a key regulator of commensalism. The
transition from (white) yeast cells to pseudohyphae or hyphae is stimulated by a wide variety of environmental inputs, which include elevated temperatures, pH and
peptidoglycan. Pseudohyphae can be distinguished from hyphae on the basis of the position of the septal junction between a mother yeast cell and its lamentous
daughter, and by the presence of invaginations at these septal junctions in pseudohyphae, but not hyphae (Merson-Davies and Odds 1989; Sudbery 2001; Sudbery, Gow
and Berman 2004). Candida albicans can be induced to form chlamydospores under specic environmental conditions (Jansons and Nickerson 1970), but the biological
signicance of this growth form remains obscure (Staib and Morschh¨
auser 2007). See text.
During experimental colonisation of the murine GI tract, C.
albicans was found to thrive in the yeast form (Vautier et al. 2015).
The basis for the predominance of the yeast morphology during
gut colonisation remains unclear, but unknown selective pres-
sures favour growth in the yeast form during experimental GI
colonisation in mice during GI dysbiosis (Tso et al. 2018). Further-
more, mucus covering the epithelium, tight junctions between
epithelial cells, and the lamina propria serve as physical barri-
ers that limit C. albicans translocation and dissemination from
the gut (Yan, Yang and Tang 2013; Arevalo and Nobile 2020).
Mucin, the main component of mucus, prevents hyphal forma-
tion (Kavanaugh et al. 2014) and reduces the adherence of C. albi-
cans to epithelial cells (de Repentigny et al. 2000). Similarly, saliva
can exert anti-Candida effects in the oral cavity (Hibino et al.
2009) (see Oral cavity). More recent work suggests that lamen-
tous forms can exist in certain parts of the GI tract where the
microenvironment favours hyphal development (Witchley et al.
2019). Only under certain circumstances, for example when a
perturbed microbiota and a compromised immune system lose
control over C. albicans growth (see The Host and The Microbiota),
6FEMS Microbiology Reviews 2020, Vol. 00, No. 00
Figure 4. A combination of virulence factors and tness attributes promote C. albicans virulence.Polymorphism: The ability of C. albicans to undergo morphological
transitions allows it to adapt to different growth conditions, adhere to biotic and abiotic surfaces, invade cells and tissue, and escape from immune cells. Invasion and
damage: A combination of induced endocytosis and active penetration promote fungal invasion of host tissues, and the accumulation of the toxin,candidalysin, in the
invasion pocket leads to pore formation and host cell damage. Adhesion/biolm formation: The battery of adhesins promotes fungal adhesions to biological and abiotic
surfaces, which can lead to the development of biolms, for example on medical devices such as catheters. Genetic and metabolic plasticity:Candida albicans displays
a high degree of metabolic exibility, which allows it to adapt rapidly to diverse host niches. This fungus also displays great genetic plasticity, which permits rapid
evolutionary adaptation to selective pressures and stresses such as exposure to antifungal drugs. Stress responses:Candida albicans activates robust stress responses
following exposure to host imposed stresses, including ROS and RNS, which enhances fungal survival following immune attack, for example. Cell wall:Aswellas
maintaining cell morphology, the robust cell wall provides protection against host-imposed stresses including changes in osmolarity. Immune evasion:Candida albicans
has evolved a variety of immune evasion strategies that include the modulation of PAMP exposure at the cell surface to evade immune recognition, and phagocytic
escape mechanisms to evade killing by innate immune cells. See text.
the fungus can switch from commensalism to pathogenicity
(Gow et al. 2011).
Signicantly, the host can exploit the yeast-to-hypha tran-
sition to discriminate between colonisation and infection. This
involves a biphasic innate immune response at the epithe-
lial barrier (Moyes et al. 2010; Roselletti et al. 2019). The rst
signalling event is triggered by fungal cell wall components,
notably β-glucans and mannans, irrespective of cell morphology
(Moyes et al. 2010). The second, danger response, is only induced
once a high fungal burden is achieved, hypha formation occurs,
and the hypha-associated toxin candidalysin is expressed (see
Host damage) (Moyes et al. 2010,2016). This leads to the secretion
of pro-inammatory cytokines and phagocyte inltration, which
promote fungal clearance. In addition, phagocytes can distin-
guish hyphae from yeast cells based on the shorter cell wall
mannan brils of hyphal cells (Cheng et al. 2011). Macrophages
also respond to hyphal load, in part through the degree of
metabolic competition between host and pathogen, displaying
reduced activation of the NLRP3-inammasome pathway at low
hyphal burdens (Tucey et al. 2020; Westman et al. 2020). Thus,
while hypha formation is critical for invasion (see Invasion mech-
anisms), the host has developed mechanisms to recognise the
invasive form of C. albicans. Therefore, hypha formation seems
to be detrimental for C. albicans commensalism.
Adhesion to abiotic and biotic surfaces
Candida albicans cells can adhere to each other as well as to host
cells and abiotic surfaces, such as catheters or dental implants,
which promotes colonisation and the formation of biolms (de
Groot et al. 2013;Lohseet al. 2018)(Fig.4). Candida albicans forms
hyphae upon sensing contact to a surface (Kumamoto 2008)and
hyphae express specic adhesins that promote adhesion to such
surfaces (de Groot et al. 2013).
The Agglutinin-Like Sequence (ALS) genes represent one
family of adhesins in C. albicans, some of which are morpho-
genetically regulated (Hoyer and Cota 2016). Analogous adhesin
families are present in other pathogenic and non-pathogenic
fungi (Butler et al. 2009). Als adhesins have a three-domain struc-
ture: the N-terminal ligand-binding domain (Lin et al. 2014);
internal tandem repeats; and the C-terminal domain, which
binds the cell wall via a modied glycosylphosphatidylinisotol
(GPI)-anchor. In C. albicans,theALS gene family has nine mem-
bers, each of which displays a high degree of variability between
alleles and strains, particularly in the length of the central repet-
itive domain (Hoyer and Cota 2016). Als3, the best-studied Als
d’Enfert et al. 7
family member, has multiple functions. It binds heterogenous
ligands including cadherins, ferritin and a Streptococcus gordonii
surface protein (Phan et al. 2007; Almeida et al. 2008; Bamford
et al. 2015). Als3 also acts as an invasin that promotes fungal
invasion of host cells (Phan et al. 2007) and iron assimilation
(Almeida et al. 2008). This makes Als3 an asset for the fungus
during infection, but also a potential target for anti-Candida ther-
apies (Edwards et al. 2018;Marcet al. 2018; Kioshima et al. 2019).
The hyphal wall protein 1 (Hwp1), is specically expressed
during hyphal growth (Staab, Ferrer and Sundstrom 1996), and
is the founding member of a second family of ve adhesins
in C. albicans (de Groot et al. 2013).Members of the Hwp fam-
ily are required for both virulence and mating. The N-terminus
of Hwp1 is enriched in glutamine residues that become cross-
linked to the host extracellular matrix by host transglutami-
nases (Staab et al. 1999). In contrast, Yeast wall protein 1 (Ywp1)
appears to counteract adhesion leading to the release of yeast
cells from surfaces, which might promote fungal dissemination
during systemic candidiasis (Granger 2012).
A third family of putative adhesins is encoded by the twelve-
member HYR gene family (de Groot et al. 2013).The founding
member of this family, HYR1, like ALS3 and HWP1, is expressed
during hyphal development (Bailey et al. 1996). This HYR family
has been less well characterised than the ALS and HWP families.
Nevertheless, it adds to the adhesins that C. albicans expresses
to promote robust adhesion to each other, abiotic surfaces or the
The cell wall
Both cellular polymorphism and adhesion are intimately asso-
ciated with the C. albicans cell wall, the organelle that maintains
the morphology of the C. albicans cell and that supplies the scaf-
fold for most adhesin proteins (Klis, de Groot and Hellingwerf
2001; de Groot et al. 2004; Gow, Latge and Munro 2017)(Fig.4).
The cell wall also provides osmotic stability and protects against
environmental stresses. It is robust in exerting control of cell
shape, and yet elastic during responses to acute osmotic stress
(Ene et al. 2015). Furthermore, the cell wall is a highly exi-
ble organelle, in that it displays a high capacity to adapt and
remodel itself in response to environmental challenges or anti-
fungal drugs (Sosinska et al. 2008;Eneet al. 2012; Childers et al.
The C. albicans cell wall is a two-layered structure. The inner
layer consists of chitin, β-1,3- and β-1,6-glucans and manno-
proteins. The outer layer is enriched in mannan brils that are
anchored to mannoproteins cross-linked to the inner layer of
the wall (Kapteyn et al. 2000;Gowet al. 2011; Gow, Latge and
Munro 2017). Chitin comprises about 2%–3% of the mass of the
yeast cell wall, but represents an important structural compo-
nent that is essential for the integrity of the cell wall. The main
structural polysaccharide of the C. albicans cell wall is β-glucan,
which accounts for 50%–60% of the mass of the yeast cell wall
(Shepherd 1987; Klis, de Groot and Hellingwerf 2001). The β-1,3-
glucan network provides the platform for covalent attachment
of chitin, β-1,6-glucan and mannoproteins.
Two main classes of cell wall mannoproteins have been
dened in C. albicans. GPI-anchored proteins are the more abun-
dant class. As their name suggests, these are linked via modi-
ed GPI anchors to β-1,6-glucan which, in turn, are covalently
attached to β-1,3-glucan (Kapteyn et al. 2000). Pir proteins (pro-
teins with internal repeats) are covalently attached to β-1,3-
glucan directly (Kapteyn et al. 2000). C. albicans cell wall manno-
proteins contribute 30–40% of the mass of the yeast cell wall
(Kapteyn et al. 2000) and are adorned with N-and/orO-linked
oligosaccharides. The O-linked oligosaccharides are often linked
to serine-threonine-rich repeats (e.g. in ALS adhesins: see Adhe-
sion to abiotic and biotic surfaces) and are thought to confer rod-
like structures to these domains (Gatti et al. 1994). N-linked man-
nans are highly branched structures that form the brils in the
outer layer of the wall (Gow, Latge and Munro 2017; Childers et al.
2019). The functions of about 70% of cell wall mannoproteins
remain obscure, but some are known or suspected to be involved
in the infection process (De Groot, Ram and Klis 2005;Richard
and Plaine 2007).
The cell wall is an attractive target for antifungal therapy
because it is essential for fungal viability and not present on
human cells. Consequently, β-1,3-glucan synthesis is the tar-
get for a major class of antifungal drugs in clinical use—the
echinocandins (Odds, Brown and Gow 2003). Signicantly, in the
context of this review, the cell wall is also the rst point of direct
contact with the host, and therefore a prime target for immune
recognition (see Fungal recognition) (Netea et al. 2008; Erwig and
Gow 2016).
Biolm formation
Candida albicans can form orid biolms on biological surfaces
and also abiotic surfaces such as catheters, dentures and pros-
thetic joints (Fig. 4). Biolms are a common source of nosocomial
infection (Ramage et al. 2005; Nobile and Johnson 2015), and they
increase therapeutic challenges by enhancing the resistance to
antifungal drugs (Taff et al. 2013).
Biolm formation is initiated by adhesion of C. albicans cells
to the surface (see Adhesion to abiotic and biotic surfaces). Surface
contact stimulates hyphal growth (see Cellular polymorphism), the
development of the biolm and the production of extracellular
matrix, and the biolm matures into an organised and robust
structure (Nobile and Johnson 2015). Biolm formation is a com-
plex process that is controlled by a network of transcription
factors and that integrates the expression of adhesins, cellu-
lar morphogenesis and the production of extracellular matrix.
Accordingly, biolm formation is controlled by a complex tran-
scriptional network of over 1000 genes (Finkel and Mitchell 2011;
Nobile et al. 2012;Lohseet al. 2018). These target genes include
members of the ALS family,which are essential for biolm for-
mation and enhance aggregation between fungal cells via amy-
loid formation (Dehullu et al. 2019;VidaHoet al. 2019).
Biolm maturation is followed by the dispersal of yeast cells
from the biolm, which promotes fungal dissemination. Candida
albicans cells dispersed from biolms are distinct from plank-
tonically grown yeast. These dispersed cells are transcription-
ally reprogrammed to utilise alternative carbon sources and they
acquire nutrients, such as zinc and amino acids, with higher ef-
ciency (Uppuluri et al. 2018).
Candida albicans clinical isolates display a high degree of het-
erogeneity with respect to their capacity to form biolms and
the underlying regulatory network (Sherry et al. 2017;Huanget al.
2019), and biolm-forming ability has been associated with high
mortality rates in patients (Rajendran et al. 2016). In the clini-
cal setting, the situation is further complicated by the forma-
tion of multispecies biolms. For example, C. albicans is com-
monly associated with Streptococcus and Actinomyces species in
dental samples, with Lactobacillus species in vaginal specimens,
and with Pseudomonas in the lungs of cystic brosis patients
(Hogan, Vik and Kolter 2004; Falagas, Betsi and Athanasiou 2006;
Bamford et al. 2009;Bandaraet al. 2009; Cruz et al. 2013; Bam-
ford et al. 2015) (see Synergistic and antagonistic interactions between
kingdoms). These inter-kingdom associations affect C. albicans
8FEMS Microbiology Reviews 2020, Vol. 00, No. 00
growth, morphogenesis and drug resistance (Hogan, Vik and
Kolter 2004).
Invasion mechanisms
The invasion of host cells and tissues provides an effective
strategy to access more nutrients, avoid competition with other
members of the microbiota, and potentially escape antimicro-
bial treatment (Fig. 4). Two distinct routes for the invasion of
epithelia and endothelia are known for C. albicans: induced
endocytosis and active penetration (Dalle et al. 2010;W
et al. 2012). Induced endocytosis is mediated by the fungal pro-
teins Ssa1 and Als3 (the adhesin-invasin, mentioned above),
both of which are present on the cell wall. These proteins bind to
E- and N- cadherins on epithelial and endothelial cells, as well as
to the epithelial growth factor receptor of oral epithelial cells, to
induce the uptake of fungal cells through remodelling of the host
cytoskeleton (Phan et al. 2007; Moreno-Ruiz et al. 2009;Sunet al.
2010; Solis et al. 2017). Active penetration is achieved through the
growth of hyphae into host tissue. This is the dominant route of
fungal invasion into oral epithelial cells and the only observed
route in enterocytes (Dalle et al. 2010;W
achtler et al. 2012).
As stated, the GI tract is a major reservoir for resident C.
albicans (Nucci and Anaissie 2001; Gouba and Drancourt 2015),
and hence fungal translocation across intestinal barriers is a
common source of systemic candidiasis. This translocation can
be promoted by injury, GI pathologies or medical interventions.
Nevertheless, the translocation of C. albicans cells through ente-
rocytes in a transcellular manner, and subsequent necrotic host
cell death, is a major mechanism by which the fungus crosses
the epithelial barrier (Allert et al. 2018). C. albicans directs physi-
cal force against cell membranes to stretch and rupture host cell
membranes via a combination of hyphal growth and secreted
virulence factors (W¨
achtler et al. 2012). Meanwhile, host cells
employ several mechanisms to expand and repair membranes
to limit this damage (Westman, Hube and Fairn 2019). This leads
to the formation of the so-called ‘invasion pocket’ where the
invading hypha is surrounded by host membrane (Moyes et al.
2016). The conned space around the hypha, within the inva-
sion pocket, permits the accumulation of C. albicans secreted
virulence factors to high local concentrations that cause further
damage and stress to the host (Dalle et al. 2010;Moyeset al. 2016;
Allert et al. 2018).
Host damage
The ability to damage host cells provides C. albicans with access
to cytoplasmic nutrients, and the fungus possesses an exten-
sive weaponry to impose damage (Fig. 4). Damaging factors that
accumulate in the invasion pocket include secreted hydrolases
such as phospholipase B1, lipases and secreted aspartic pro-
teases (Saps) that degrade host membranes, proteins and extra-
cellular matrix releasing nutrients (Mukherjee et al. 2001; Naglik,
Challacombe and Hube 2003; Schoeld et al. 2005). Candida albi-
cans also expresses candidalysin—a pore forming α-helical pep-
tide toxin that is encoded by the ECE1 gene (Moyes et al. 2016).
Pores formed in the host cell membrane by candidalysin proba-
bly leak cytoplasmic contents into the invasion pocket, thereby
providing additional nutrients for the fungus. This may include
access to essential micronutrients such as iron and zinc. Spe-
cic proteins bind these micronutrients, which are then endo-
cytosed or transported across the fungal cell membrane via
specic transporters. For example, members of the Rbt5-family
transport heme across the cell wall (Kuznets et al. 2014; Nasser
et al. 2016). Also, zinc is acquired via the zincophore Pra1 (pH-
regulated antigen 1), which is released into the extracellular
space and then, when loaded with zinc, is transported back into
the fungus by the zinc transporter Ztr1 (Citiulo et al. 2012).
Fitness attributes
Fitness attributes are factors that promote fungal virulence by
enhancing the physiological robustness of the fungus in host
niches, rather than by interacting directly with the host. In C.
albicans, tness attributes include metabolic exibility combined
with potent nutrient acquisition systems, and robust stress
response mechanisms (Mayer, Wilson and Hube 2013; Brown,
Budge et al. 2014; Brown, Brown, et al. 2014). These promote the
success of C. albicans both as a commensal and as a pathogen of
Flexible metabolic adaptation
Metabolic adaptability is critical during C. albicans transitions
between commensalism and pathogenicity (Fig. 4). This was
highlighted by an elegant screen for regulatory circuitry that
drives the commensal and pathogenic states in C. albicans (P´
Kumamoto and Johnson 2013). Much of this circuitry is involved
in the regulation of metabolism. Metabolic regulation in C. albi-
cans is integrated with the control of virulence factors and stress
resistance through major regulatory hubs such as Efg1, Tup1,
Nrg1, Hog1 and Gcn4 (Murad, d Enfert et al. 2001; Tripathi et al.
2002; Doedt et al. 2004; Alonso-Monge et al. 2009). Therefore,
metabolic adaptation is essential for commensalism and viru-
lence, and is intimately linked with other pathogenicity traits
(Mayer, Wilson and Hube 2013; Brown, Brown, et al. 2014).
Glucose is a preferred carbon source for C. albicans, but under
glucose-limiting conditions, such as in the colon or after entrap-
ment in the phagosome, C. albicans tunes its metabolism to feed
on alternative carbon sources (Lorenz, Bender and Fink 2004;
Barelle et al. 2006). Even when glucose becomes available, C.
albicans can simultaneously utilise alternative carbon sources
through multiple pathways (Sandai et al. 2012; Childers et al.
2016). This metabolic exibility allows the fungus to adapt to
contrasting host niches. Signicantly, it also inuences the tol-
erance of C. albicans to antifungal drugs and environmental
stresses (Ene et al. 2012). For example, growth on lactate protects
against osmotic and cell wall stresses while utilisation of amino
acids and N-acetylglucosamine (GlcNAc) increases fungal resis-
tance to reactive oxygen and nitrogen species (ROS and RNS,
respectively) (Williams and Lorenz 2020). These alternative car-
bon sources appear to serve as niche-specic signals that prime
the fungus for impending challenges, pointing to the dexterity
of C. albicans not only to adapt, but also to anticipate, local stress
conditions (Brown, Budge et al. 2014; Alistair J P Brown et al. 2019;
Williams and Lorenz 2020). The metabolic exibility of C. albi-
cans extends well beyond carbon metabolism to include nitro-
gen, phosphate and micronutrient assimilation (Lorenz, Bender
and Fink 2004;Yinet al. 2004;Vylkovaet al. 2011;Eneet al. 2014;
Ikeh et al. 2016).
Micronutrients, such as iron and zinc, are essential for
structural integrity and physiological processes in C. albicans.
However, in response to infection, through a process called
nutritional immunity, the host limits the availability of these
micronutrients and exposes the fungus to toxic levels of other
species such as copper ions (Noble 2013; Potrykus et al. 2013;
Mackie et al. 2016; Sprenger et al. 2018). In response, the fun-
gus activates efcient micronutrient acquisition strategies. High
afnity iron uptake involving a cyclic iron reduction pathway
(iron reductase, multicopper ferroxidase and iron permease)
is activated to take over from low afnity ferritin-iron uptake
d’Enfert et al. 9
via the protein Als3, which is operational in hyphae during
iron-replete conditions (Wilson, Naglik and Hube 2016; Bairwa,
Hee Jung and Kronstad 2017). Candida albicans can also assim-
ilate iron from heme and hemoglobin using Common in Fun-
gal Extracellular Membrane (CFEM) proteins, and can scavenge
siderophores synthesised by other microorganisms using the
Arn1/Sit1 ferrichrome transporter (Bairwa, Hee Jung and Kro-
nstad 2017). Transcriptional circuitry involving Sef1, Sfu1 and
Hap43 control iron homeostasis by activating iron assimila-
tion mechanisms when iron is limiting, and by repressing iron
uptake when it is in excess (Chen et al. 2011;Noble2013). Can-
dida albicans utilises two uptake mechanisms to scavenge zinc.
The rst, which operates mainly at acidic pHs, involves uptake
via the Zrt2 transporter into the cytoplasm (Crawford et al.
2018). The second, which is functional at neutral pHs, entails
zincophore-mediated zinc scavenging through a secreted pro-
tein, Pra1 and uptake via the transporter Zrt1 (Citiulo et al. 2012;
Wilson 2015; Crawford et al. 2018). C. albicans responds to zinc
limitation by forming goliath cells (enlarged and spherical yeasts
that exhibit enhanced adhesion) and avoids zinc toxicity by
rapidly compartmentalizing zinc in storage vacuoles called zin-
cosomes (Malavia et al. 2017; Crawford et al. 2018).
Robust stress responses
Fungal pathogens generally display robust responses to cer-
tain stresses, particularly oxidative stress (Brown et al. 2017)
(Fig. 4). Candida albicans is resistant to signicantly higher levels
of ROS than its distant cousin, Saccharomyces cerevisiae(Jamieson,
Stephen and Terri`
ere 1996; Nikolaou et al. 2009) and this helps
the fungus to counter toxic ROS produced by innate immune
cells, before and during phagocytic attack (Miram´
on et al. 2012).
C. albicans and other fungal pathogens counteract acute exoge-
nous oxidative stresses by inducing genes involved in ROS
detoxication (e.g. catalase and superoxide dismutases), the
synthesis of antioxidants (e.g. glutathione and thioredoxin),
and the repair of ROS-mediated damage to DNA, proteins and
lipids (Enjalbert, Nantel and Whiteway 2003,Enjalbertet al.
2006;Znaidiet al. 2009). The inactivation of key regulators of
the response in C. albicans (Cap1, Skn7 and Hog1) compromises
oxidative stress resistance (Alarco and Raymond 1999;Singh
et al. 2004;Smithet al. 2004). Virulence is attenuated by the
inactivation of the Hog1 stress activated protein kinase (Alonso-
Monge et al. 1999; Cheetham et al. 2011), but only to a minor
extent by the loss of Cap1 or Skn7 (Singh et al. 2004;Jainet al.
2013). The overexpression of catalase, which detoxies hydrogen
peroxide, enhances oxidative stress resistance in vitro, and yet,
counterintuitively, reduces the virulence of C. albicans (Rom ´
et al. 2016;Pradhanet al. 2017). This is because overexpres-
sion of this abundant ferroprotein places an undue demand for
the essential micronutrient, iron, under iron limiting conditions
in vivo (Pradhan et al. 2017). Clearly, numerous and potentially
opposing, selective pressures must be balanced to optimise fun-
gal tness in a particular host niche.
While much attention has focussed on oxidative stress,
C. albicans faces other forms of environmental stress in the
host, including nitrosative, osmotic and thermal stresses. Innate
immune cells expose C. albicans to RNS) in an attempt to kill
and clear the fungus. C. albicans responds by activating genes
involved in RNS detoxication (such as the avohemoglobin,
Yhb1), glutathione synthesis and recycling, and the repair of
RNS-mediated damage (Hromatka, Noble and Johnson 2005; Till-
mann et al. 2015). The response to nitrosative stress is driven by
the transcription factor Cta4 and Hog1 (Chiranand et al. 2008;
Herrero-de-Dios et al. 2018). The inactivation of YHB1,CTA4
or HOG1 attenuates nitrosative stress resistance and virulence
(Alonso-Monge et al. 1999; Hromatka, Noble and Johnson 2005;
Chiranand et al. 2008; Cheetham et al. 2011; Miram ´
on et al. 2012).
Candida albicans cells thrive in niches with different osmo-
larities (e.g. on skin, in the oral cavity or GI tract), and yet must
maintain osmo-homeostasis to grow. Hypo- and hyper-osmotic
challenges are countered by modulating the levels of intracel-
lular osmolytes. For example, C. albicans upregulates the syn-
thesis and accumulation of glycerol and arabitol in response
to hyperosmotic challenges (San Jos´
eet al. 1996; Kayingo and
Won g 2005). This response is regulated at both transcriptional
and post-transcriptional levels by the evolutionarily conserved
Hog1 MAP kinase signalling pathway (Smith et al. 2004;Enjalbert
et al. 2006).
Candida albicans must also restore and maintain proteostasis
in the face of thermal challenges, even within the mammalian
host (Nicholls et al. 2011). Even mild increases in temperature
lead to activation of the so-called heat shock response (Leach,
Ty c et al. 2012), which is regulated by an autoregulatory circuit
involving the heat shock transcription factor (Hsf1) and heat
shock protein 90 (Hsp90) (Leach, Budge et al. 2012). The response
involves the induction of functions involved in protein refolding
and protein degradation to repair or recycle damaged proteins
(Nicholls et al. 2009; Leach et al. 2016). The heat shock response
is integrated with key virulence attributes in C. albicans such as
yeast-hypha morphogenesis, adhesion and the ability to dam-
age epithelial cells (Shapiro et al. 2009; Leach et al. 2016). Con-
sequently, the inactivation of the response attenuates virulence
(Nicholls et al. 2011).
Candida albicans can thrive over an extremely wide range of
ambient pHs, from pH 2 to 10 (Vylkova et al. 2011). pH responses
are particularly relevant given the ability of C. albicans to colonise
host niches with contrasting pHs such as the vagina (acidic), GI
tract (acidic to mildly alkaline) and blood (neutral). These pH
responses, which are regulated in part by the evolutionarily con-
served Rim101 pathway (Davis, Wilson and Mitchell 2000), are
tightly integrated with metabolic adaptation, nutrient acquisi-
tion and morphogenesis (Davis et al. 2000). Yeast-hypha mor-
phogenesis in C. albicans is regulated in response to ambient pH
(Buffo, Herman and Soll 1984; Porta et al. 1999; Chen et al. 2020;
Villa et al. 2020). Ambient pH also affects trace metal solubil-
ity, and consequently, micronutrient assimilation strategies in
C. albicans are regulated in response to pH (Noble 2013; Wilson
2015; Crawford et al. 2018). Signicantly, C. albicans is not sim-
ply reactive to pH: it can proactively alkalinise its microenviron-
ment through the catabolism of polyamines and amino acids,
leading to the release of ammonia and/or CO2(Mayer et al. 2012;
Vylkova and Lorenz 2014; Danhof et al. 2016;Vylkova2017). Inter-
estingly, lactate production by a co-commensal in the oral cavity,
Streptococcus mutans, provides carboxylic acid substrates that are
sufcient to promote C. albicans-mediated alkalinisation of the
microenvironment (Danhof et al. 2016; Willems et al. 2016).
Immune evasion
Immune evasion can be viewed as an additional type of tness
attribute because it promotes the physiological robustness of
the fungus in the host (Fig. 4). Candida albicans has evolved a
variety of mechanisms through which it can reduce recognition
by immune cells, decrease the efcacy of antimicrobial killing
mechanisms, escape immune cells following engulfment, and
manipulate the immune system (see Innate antifungal responses
and Fungal countermeasures for more detail). During co-evolution
with its host, C. albicans has even developed mechanisms by
10 FEMS Microbiology Reviews 2020, Vol. 00, No. 00
which it can anticipate, and protect itself against, imminent
immune attack.
Clearly, C. albicans possesses an array of powerful tness
attributes through which this fungus tunes its physiology to
counter environmental challenges presented by the host. Sig-
nicantly, the fungus not only adapts to host-dened conditions,
but can also anticipate impending challenges, and actively mod-
ulate its microenvironment.
Candida albicans epidemiology and variability
The exibility of C. albicans, which underlies its success as a
commensal and a pathogen, is also reected at the genetic level
(Fig. 4). Clinical isolates of C. albicans are generally diploid, with
a haploid genome size of 16 Mb, organised into eight chro-
mosomes. However, isolates display high levels of sequence
heterozygosity between homologous chromosomes (Selmecki,
Forche and Berman 2006; Ford et al. 2014;Hirakawaet al. 2015)
and a high degree of genome plasticity driven by ploidychanges,
karyotypic variations due to partial and whole chromosome
aneuploidies, point mutations, short and long-range loss of het-
erozygosity (LOH) events and copy number variations (Chibana,
Beckerman and Magee 2000; Selmecki, Forche and Berman 2006;
Ford et al. 2014;Hirakawaet al. 2015; Ropars et al. 2018; Sitterl´
et al. 2019). Furthermore, haploid and tetraploid strains have
been observed both in vitro and in vivo (Hull, Raisner and Johnson
2000; Magee and Magee 2000; Hickman et al. 2013).
Multilocus sequence typing (MLST) and genome sequencing
studies have revealed that C. albicans isolates are distributed
amongst at least 23 genetic clusters (1–18, A-E) (Bougnoux et al.
2006; Odds et al. 2007; Odds 2010; Ropars et al. 2018). In general,
there are no clear phenotypic associations with these clusters
(Bougnoux et al. 2006; MacCallum et al. 2009). However, some
clusters do exhibit geographical enrichment (Odds et al. 2007;
MacCallum et al. 2009;Shinet al. 2011), suggesting indepen-
dent recent evolutionary histories for these clusters. Cluster 13
is somewhat exceptional in that it represents a highly clonal
lineage of isolates that exhibit low heterozygosity (Ropars et al.
2018). Isolates in cluster 13 are distributed worldwide (Fakhim
et al. 2020), despite being called Candida africana strains (Tietz
et al. 2001). They are isolated predominantly from the genital
niche and display unusual morphological and phenotypic fea-
tures that include slow growth, an inability to produce chlamy-
dospores and assimilate aminosugars, and decreased virulence
(Tietz et al. 2001; Romeo, De Leo and Criseo 2011; Borman et al.
2013). In contrast to other C. albicans clusters, cluster 13 isolates
harbour a unique pattern of single nucleotide polymorphisms
(SNPs) and a signicantly lower level of heterozygosity (Ropars
et al. 2018). In addition, in cluster 13 isolates, genes important
for morphogenesis and virulence have undergone pseudogeni-
sation, which probably explains the decreased virulence and
apparent genital niche restriction of these isolates (Ropars et al.
Once thought to be an asexual obligate diploid organism, C.
albicans has been shown to undergo a parasexual cycle (Magee
and Magee 2000; Bennett and Johnson 2003; Ene and Bennett
2014). The majority of C. albicans diploid strains are incapable
of mating, being heterozygous at the mating type-like (MTL)
locus. However, mating can occur mainly between strains that
have become homozygous at the MTL locus on chromosome 5,
and have complementary MTL genotypes (i.e. are MTLa/a and
MTLα/α). Additionally, mating in C. albicans is also dependent
on a phenotypic switch from the mainly sterile ‘white’ pheno-
type to the mating competent ‘opaque’ phenotype (Miller and
Johnson 2002). Mating between competent isolates of opposite
mating-type results in tetraploid cells. These can subsequently
undergo concerted chromosome loss, which can restore the
diploid state in a meiosis-independent manner (Bennett and
Johnson 2003; Hickman et al. 2015). However, this process yields
diverse intermediate aneuploid states (Hickman et al. 2015).
Hence, this mode of parasexual reproduction provides a means
of generating genetic and phenotypic diversity in C. albicans
(Forche et al. 2008; Hickman et al. 2015). Indeed, recombination
has been shown to occur three orders of magnitude more fre-
quently during concerted chromosome loss than during mitosis
(Anderson et al. 2019). Interestingly, recombination during con-
certed chromosome loss is highly dependent on two meiosis-
specic genes, SPO11 and REC8 (Forche et al. 2008;Andersonet al.
2019). The involvement of meiosis-specic genes in concerted
chromosome loss has led to the suggestion that this process
blurs the boundaries’ between meiosis and mitosis, and that this
parameiosis’ might provide insight into the evolution of meiosis
(Anderson et al. 2019).
The view that the parasexual cycle rarely occurs in the host
is supported by population genetics, which shows that C. albi-
cans populations are predominantly clonal (Pujol et al. 1993;
McManus and Coleman 2014). Nevertheless, the conservation
of mating genes suggests that this process is associated with
an evolutionary advantage. Furthermore, because the parasex-
ual cycle is stimulated by environmental stress, it may be a
diversity-enhancing process that enhances adaptation and sur-
vival under hostile conditions (Selmecki, Forche and Berman
2010; Zhang et al. 2015;Hirakawaet al. 2017; Popp et al. 2019). This
idea is corroborated by evidence of recombination and gene ow
in natural isolates, despite the largely clonal structures of C. albi-
cans populations (Odds et al. 2007; Bougnoux et al. 2008; Zhang
et al. 2015; Ropars et al. 2018). This could explain why C. albicans
isolates maintain a high degree of genetic diversity despite their
predominantly clonal reproduction.
The diversity of C. albicans populations has arisen partly
through changes in ploidy and aneuploidy. These mechanisms
have provided C. albicans with a means of evolving rapidly in
response to environmental challenges (Selmecki, Forche and
Berman 2006;Diogoet al. 2009; Bennett, Forche and Berman
2014). The association of genome rearrangements with anti-
fungal resistance acquisition has been well documented, with
genomes of antifungal-resistant strains often exhibiting copy
number variations and chromosome aneuploidies (Selmecki,
Forche and Berman 2010). Indeed, a striking example of seg-
mental aneuploidy was reported in uconazole resistant strains,
consisting of an isochromosome composed of the two left arms
of chromosome 5 (Selmecki, Forche and Berman 2006, Selmecki
et al. 2008). Trisomy of chromosome 2 or R has also been reported
to enhance antifungal drug resistance in C. albicans (Xingxing Li
et al. 2015;Yanget al. 2019). Large-scale chromosome rearrange-
ments occur in C. albicans as an adaptation mechanism in both
oral and GI niches (Ene et al. 2018; Forche et al. 2018). Similar
observations have been made in isolates collected from a sin-
gle human individual (Sitterl´
eet al. 2019). Genome sequencing
of clinical isolates from patients that received antifungal ther-
apy revealed that eight of the 21 isolates underwent karyotypic
changes, with the majority being trisomic for chromosome 4 or
7(Hirakawaet al. 2015). However, a more recent study of 182 clin-
ical isolates might suggest that both segmental and whole chro-
mosome aneuploidies are relatively infrequent events (Ropars
et al. 2018). Changes in ploidy are known to provide a selec-
tive advantage under stress conditions, but can confer long-term
tness defects when grown under nonselective conditions, as
d’Enfert et al. 11
illustrated by decreased growth and virulence (Hickman et al.
2015,2013;Hirakawaet al. 2015). Therefore, the extent to which
these events are observed in the genomes of C. albicans isolates
must reect the frequency of these types of genetic event and
the nature of the selective pressures that these isolates recently
Diversity has also arisen through high rates of mutation at
the nucleotide level (SNPs, insertions and deletions). Candida
albicans isolates display high levels of natural heterozygosity,
with one heterozygous SNP occurring per 200–300 bp on average
(Jones et al. 2004; Butler et al. 2009;Hirakawaet al. 2015; Ropars
et al. 2018). The levels of heterozygosity are inuenced by large
LOH events, which can affect all chromosomes and are com-
mon in C. albicans isolates. LOH events are signicantly elevated
under stress conditions, such as exposure to antifungal agents,
heat or oxidative stress (Forche et al. 2011; Ropars et al. 2018).
Rapid phenotypic and genetic changes have been observed in
various infection and colonisation models as well as in clinical
isolates (Forche, May and Magee 2005; Bougnoux et al. 2006, 2009;
Cheng et al. 2007; Bougnoux et al. 2008;Diogoet al. 2009;L¨
et al. 2013;Eneet al. 2018; Forche et al. 2018; Sitterl´
eet al. 2020).
This microevolution is driven primarily by de novo base substitu-
tion and short-range LOH events (Ene et al. 2018), and can clearly
impact the relationship between fungus and host (Wartenberg
et al. 2014; Tso et al. 2018; Liang and Bennett 2019) as well as resis-
tance to antifungal therapy (Coste et al. 2006; Ford et al. 2014).
Mammals are constantly exposed to microbes on the skin and
mucosal surfaces of the GI, respiratory and reproductive tracts.
Therefore, epithelial surfaces in the mucosal tissues repre-
sent primary sites of interaction between C. albicans and the
host (Lim et al. 2012). To prevent microbial overgrowth on the
epithelial barriers and microbial invasion of tissues, the host
actively surveys and protects its barrier surfaces via two dis-
tinct, complementary and cooperating branches of the immune
system: innate and adaptive immunity (Fig. 5). As well as form-
ing a physical barrier, epithelial cells contribute to the host
response through active recognition of microbes and evalua-
tion of their pathogenic potential. This is complemented by
myeloid cells of the innate immune system, which exploit
evolutionarily conserved pattern recognition receptors (PRRs)
to recognise microbial pathogen-associated molecular patterns
(PAMPs). Recognition of PAMPs by PRRs triggers phagocytosis
of the microbial target and/or antimicrobial effector responses
with the purpose to eradicate the pathogen. In addition, the
innate immune system, and dendritic cells (DCs) in particular,
activate the adaptive immune system. T helper (Th) cells are
activated in an antigen-specic manner to coordinate epithe-
lial defenses, improve innate immune function, activate anti-
body responses, and ultimately control the fungal load and
resolve inammation. Through the development of immuno-
logical memory, adaptive immunity provides long-lasting pro-
tection against microbes. We address the cellular and molecu-
lar mechanisms of innate and adaptive immunity that provide
critical protection against C. albicans infection at epithelial bar-
riers where interactions between the fungus, host and micro-
biota play out. These interactions are dependent on tissue type
and are inuenced by variations between individuals that affect
susceptibility to fungal infection.
Innate immunity
Fungal recognition
The innate immune system is the rst line of defense against
C. albicans infection (Fig. 5). Epithelial cells (Richardson, Ho and
Naglik 2018;Nikouet al. 2019; Swidergall 2019) combine with
innate immune cells (Naglik et al. 2017; Verma, Gaffen and
Swidergall 2017; Richardson et al. 2019) to provide this defense
system, initiating anti-Candida immunity in response to fungal
Tissue-resident phagocytes, such as macrophages and DCs,
are crucial in maintaining mucosal homeostasis (Ramirez-Ortiz
and Means 2012; Xu and Shinohara 2017; Watanabe et al. 2019).
However, innate immune cell populations differ between tis-
sues, resulting in tissue-specic variation in the induction of
innate and adaptive immune responses (see Variability in the
immune response). Following hypha formation and C. albicans
invasion, neutrophils and monocytes are rapidly recruited to the
site of infection to mediate pathogen clearance through various
antifungal responses (see Antifungal response) (Richardson et al.
Myeloid cells recognise specic microbial PAMPs using spe-
cic PRRs that fall into four main families: Toll-like receptors
(TLRs), C-type lectin receptors (CLRs), nucleotide oligomerisa-
tion domain (NOD)-like receptors (NLRs) and RigI-helicase recep-
tors (RLRs). CLRs are critical for fungal recognition (Hardison
and Brown 2012). Several types of CLR recognise C. albicans,
including Dectin-1, Dectin-2, Mincle, DC-Sign, and the man-
nose receptor (MR) (Hardison and Brown 2012; Dambuza et al.
2017;Goyalet al. 2018; Swidergall 2019). Dectin-1 recognises
fungal β-glucans, which triggers the Card9-Syk pathway, lead-
ing to Nuclear Factor-kappa B (NFκB) activation and consequent
cytokine and chemokine release (Drummond et al. 2011). In addi-
tion, dectin-1 induces phagocytosis and inammasome activa-
tion (Kankkunen et al. 2010; Goodridge, Underhill and Touret
2012; Swidergall 2019). Dectin-2 recognises α-mannans (McGreal
et al. 2006; Saijo et al. 2010) and induces the formation of Neu-
trophil Extracellular Traps (NETs) after recognising unopsonised
C. albicans cells (Wu et al. 2019). In addition, Dectin-2 forms het-
erodimers with Dectin-3 and binds α-mannans on the surfaces
of C. albicans hyphae (Zhu et al. 2013). Mannans are also recog-
nised by Mincle, DC-Sign and the MR (Hardison and Brown 2012;
Erwig and Gow 2016; Dambuza et al. 2017).
TLR-mediated PAMP recognition activates MyD88-dependent
and TRIF signalling pathways in innate immune cells to regulate
the inammatory response (Kawasaki and Kawai 2014; Swider-
gall 2019). TLR2 and TLR4 recognise mannoproteins, while TLR9
recognises fungal DNA (Naglik et al. 2017). In addition, together
with TLR9, the cytosolic NLR receptor NOD2 senses chitin parti-
cles (Wagener et al. 2014). NOD2-mediated recognition of chitin
was found to down-regulate inammatory responses (Wagener
et al. 2014), which explains why NOD2 was initially described as
being redundant for the induction of inammatory responses
against C. albicans (van der Graaf et al. 2006; van de Veerdonk
et al. 2009). Recently, the epithelial Ephrin type-A receptor 2
(EphA2) was described as a non-classical PRR that recognises β-
glucan (Swidergall et al. 2018). This receptor is expressed on neu-
trophils and stimulates antifungal activity during oropharyngeal
candidiasis (OPC) (Swidergall, Solis et al. 2019). Meanwhile, the
melanoma differentiation-associated factor 5 (MDA5), a member
of the RIG-I-like receptor (RLR) family that senses viral RNA, has
been reported to also trigger an antifungal immune response,
although its ligand remains obscure (Jaeger, van der Lee et al.
12 FEMS Microbiology Reviews 2020, Vol. 00, No. 00
Figure 5. Immune recognition of, and immune responses against, C. albicans.Candida albicans yeast and hyphal cells are recognised by neutrophils, macrophages
and dendritic cells via pattern recognition receptors (see key). This recognition activates the expression and release of proinammatory cytokines and chemokines
that promote the recruitment of macrophages and neutrophils to the site of infection. Epithelial cells respond to hypha formation and the subsequent secretion of
candidalysin by the fungus, by activating the expression and release of AMPs, DAMPs, chemokines and cytokines via p38/cFos and ERK/MKP1 signalling. The AMPs
attenuate fungal growth and invasion, while DAMPs and cytokines promote inammation. Myeloid cells promote fungal killing and clearance through a combination
of phagocytosis and NETosis in the case of neutrophils. Fungal recognition leads to the maturation of dendritic cells, and their surface presentation of fungal antigens
to na¨
ıve T-cells, which stimulates adaptive immunity. The interactions between antigen-presenting dendritic cells and na¨
ıve T-cells induces T-cell activation and
differentiation into various effector T cell subsets that regulate mucosal immunity largely via IL-17 and IL-22 secretion,and stimulate macrophages via IFN-γ. See text.
Tab l e 1 . Pattern recognition receptors in epithelial and innate immune cells that recognise C. albicans pathogen-associated molecular patterns.
PRR family PRR Fungal PAMP Expressed in Reference
TLRs TLR2 Phospholipomannans Neutrophils, macrophages, DCs,
Epithelial cells (oral, vaginal,
(Kurt-Jones et al. 2002; Jouault et al. 2003; Fazeli,
Bruce and Anumba 2005;D
ecanis, Savignac and
Rouabhia 2009; McClure and Massari 2014)
TLR4 O-linked mannans Neutrophils, monocyte,
(oral, vaginal, intestinal)
(Netea et al. 2006; Hyung Sook Kim et al. 2016;
Fazeli, Bruce and Anumba 2005;Weindlet al.
2007; McClure and Massari 2014)
TLR9 Fungal DNA Chitin DCs, Neutrophils, macrophages,
epithelial cells (oral, vaginal,
(Miyazato et al. 2009; Kasperkovitz et al. 2011;
McClure and Massari 2014; Wagener et al. 2014)
CLRs Dectin-1 β-glucans Macrophages, monocytes,
neutrophils, DCs, epithelial cells
(oral, intestinal)
(Brown and Gordon 2001;Brownet al. 2002;Taylor
et al. 2002; Ariizumi, Shen, Shikano, Xu et al. 2000;
Cohen-Kedar et al. 2014)
Dectin-2 Mannoproteins
Macrophages, DCs (Taylor et al. 2005; Ariizumi, Shen, Shikano, Ritter,
et al. 2000)
Dectin-3 Mannoproteins
Macrophages, (Zhu et al. 2013)
DC SIGN Mannoproteins Macrophages, DCs (Cambi et al. 2003; Rappocciolo et al. 2006)
Mincle Mannoproteins Neutrophils, macrophages, DCs (Wells et al. 2008; Vijayan et al. 2012;
ınez-L ´
opez et al. 2019)
MR Mannoproteins Chitin DCs, macrophages (van de Veerdonk et al. 2009; Martinez-Pomares
2012; Wagener et al. 2014)
NA EphA2 β-glucans Oral epithelial cells, neutrophils (Swidergall, Solis, et al. 2019)
Galectin-3 β-mannosides Monocytes, macrophages, DCs,
neutrophils, epithelial cells
(Jouault et al. 2006)
RLRs MDA5 Unknown Monocytes, DCs, macrophages,
epithelial cells
(Plato, Hardison and Brown 2015)
NLRs NOD2 Chitin Monocytes, DCs, macrophages (Wagener et al. 2014)
PRRs involved in the recognition of C. albicans by myeloid
cells have been well characterised (above), but less is known
about epithelial cell PRRs that recognise C. albicans. Epithelial
cells use several types of PRR to sense C. albicans, including TLR2,
TLR4, dectin-1 and EphA2 (Weindl et al. 2007;D
ecanis, Savignac
and Rouabhia 2009; Cohen-Kedar et al. 2014; Swidergall et al.
2018). Despite its primordial role in the recognition of C. albi-
cans by myeloid cells, dectin-1 is thought to play a limited role
in epithelial cells (Moyes et al. 2010; Verma et al. 2017;Richard-
son, Ho and Naglik 2018). Rather, sensing of fungal β-glucans
d’Enfert et al. 13
by epithelial cells is achieved mainly by EphA2, which activates
MAPK and STAT3 signalling to induce the secretion of inamma-
tory cytokines and antimicrobial peptides by oral epithelial cells
(Swidergall et al. 2018). PRR expression patterns vary amongst
epithelial cell types and this, together with differential myeloid
cell types, contributes to niche-specic variations in mucosal
responses against C. albicans (Nikou et al. 2019; Swidergall 2019)
(see Tissue-specic immune responses).
Epithelial cells can be activated by the C. albicans peptide
toxin, candidalysin, as well as through PRR-PAMP interactions.
This cytolytic peptide damages epithelial cells and activates the
epithelial growth factor receptor (EGFR) (Jemima Ho et al. 2019).
This, in turn, activates p38/cFos and ERK/MKP1 signalling, lead-
ing to the initiation of various effector responses (see Innate anti-
fungal responses). The epithelial response to candidalysin is par-
ticularly relevant to the transition of C. albicans from commen-
salism to pathogenicity, because candidalysin is synthesised
during hyphal growth and accumulates in the invasion pocket
as the fungus invades the epithelial surface (Moyes et al. 2016)
(see Invasion mechanisms). This response to candidalysin endows
epithelial cells with the ability to respond to the invasive hyphal
form of C. albicans, rather than its relatively benign commensal
state (Moyes et al. 2010; Naglik et al. 2017).
Innate antifungal responses
Following recognition of C. albicans by phagocytic receptors,
phagocytes such as neutrophils and macrophages can engulf
the target C. albicans cell by phagocytosis, the purpose being to
entrap and kill the pathogen (Brown 2011)(Fig.5). Phagocyto-
sis involves rapid reorganisation of the plasma membrane and
cytoskeleton, and the imposition of mechanical force to engulf
the fungal cell and entrap it within a phagosome (Ostrowski,
Grinstein and Freeman 2016; Huse 2017). The phagosome then
undergoes a series of plasma-membrane phosphoinositide-
and Rab-dependent membrane fusion and ssion events with
endolysosomal compartments that promote the assimilation of
microbicidal and lytic enzymes, and the progressive acidica-
tion of the organelle, to form the mature phagolysosome (Brown
2011; Fairn and Grinstein 2012; Miram ´
on, Kasper and Hube 2013;
Erwig and Gow 2016; Walpole, Grinstein and Westman 2018). In
an attempt to kill the fungus, the phagocyte exposes its fun-
gal cargo to a low pH, a nutrient limiting microenvironment
and a potent mix of proteases, reactive chemical species ROS
and RNS, cation uxes and AMPs (Lorenz, Bender and Fink 2004;
Brown 2011; Miram ´
on, Kasper and Hube 2013