Biofilms and vulvovaginal candidiasis
Graphical abstract
Introduction
Despite therapeutic advances, vulvovaginal candidiasis (VVC) continues to be a global problem, affecting all social strata. Vaginal defence mechanisms are not yet fully understood; therefore, although numerous risk factors have been identified, the pathogenic mechanisms of VVC remain to be elucidated. The absence of a rapid, simple, and inexpensive diagnostic test results in both over- and under-diagnosis. According to Brandolt et al., VVC is generally misdiagnosed, while studies carried out on VVC patients are not representative of the general population and are peppered with diagnostic errors [1]. VVC affects between 70 and 75% of women of childbearing age, and it is estimated that 40–50 % will experience a recurrence [1,2].
According to Gonçalves et al., the causes of VVC are multifactorial, and genetic factors (polymorphisms in blood group), hormones, the use of antibacterial agents, age, sexual activity, pathologies such as diabetes mellitus, and idiopathic causes, are all thought to be predisposing factors that contribute to the colonization and development of VVC [3]. Sources of infection may include Candida acquired from the intestine, through sexual contact, or from a relapse resulting from a previous treatment that failed to eradicate the yeast completely.
Candida spp. are commensal fungi and are a normal part of the human mycoflora found in the lower genital tract in 10–20% of healthy women of childbearing age. Infections by Candida spp. are usually asymptomatic; however, the evolution from colonization to symptomatic infection involves different host factors, such as susceptibility and inflammatory responses and/or an imbalance in the vaginal microbiota [1,4].
C. albicans has been the species most frequently implicated in the different forms of invasive candidiasis. However, as reported by Brandolt et al. [1] and Nakamura-Vasconcelos et al. [5], infections by different species such as C. glabrata, C. parapsilosis, C. krusei, and C. tropicalis have emerged over the last two decades, resulting in an increase in the number of species that cause VVC. Two of the factors that have made the most significant contributions to this increase in the number of causative species have been the use of azoles and the increased use of intravascular devices, resulting in an increase in candidaemia caused by commensal species from the skin, such as C. parapsilosis and C. glabrata. There are differences not only in the pathogenicity of the different Candida spp., but also in their resistance profiles. For example, C. albicans is more virulent than C. krusei, and yet it is notably more susceptible to antifungal drugs [1].
Until recently, C. glabrata was considered to be an avirulent species that is part of the normal microbiota found on skin and mucous membranes. However, with the use of immunosuppressants and broad-spectrum antibiotics, infections due to this species have increased exponentially [6].
A more complicated form of VVC is referred to as recurrent VVC (RVVC), which is defined as at least four confirmed episodes in a year, with the following clinical symptoms: vulvar erythema, vaginal discharge, oedema, and burning/soreness. Culturing of Candida spp. and/or wet mount microscopy is mandatory, and these tests should be positive (as is the case with the symptoms listed above). It is estimated that 5–8 % of women develop these symptoms, making RVVC an important public health problem [7,8].
Muzny et al. showed [9] that Candida spp. are able to form biofilms, and numerous studies have been performed to characterise biofilm formation, virulence, and the antifungal susceptibility profiles in clinical isolates from VVC and RVVC patients [[10], [11], [12], [13]]. Currently, it is known that virulence and the type of resistance to conventional treatments for VVC depend on the formation of Candida biofilms. Biofilm formation occurs when planktonic microorganisms encounter a surface and adhere to it. Assuming that the location offers nutrients or other advantages, the adherent cells up-regulate the expression of genes involved in matrix production, and biofilm formation begins. Biofilms are characterized as highly organized sessile microbial communities that decrease susceptibility to antimicrobial agents and enhance the spread of antimicrobial resistance, thus providing a safe haven for other opportunistic pathogens.
The mechanisms underlying biofilm resistance include: [1] an incomplete penetration by antibiotics and host immune cells through the matrix [2], physiological changes in the biofilm microenvironment due to low growth and starvation responses [3], phenotypic changes in biofilm cells [4], quorum sensing (QS) between biofilm microorganisms [5], expression of efflux pumps which can transport antimicrobial agents out of cells, and [6] the presence of “persister cells,” a small fraction of microorganisms that are able to survive antibiotics. Although some antimicrobial agents can readily permeate biofilms, there exist several mechanisms by which they can be deactivated or antagonized (e.g. the presence of catalytic enzymes, hypoxic conditions, and pH changes). Additionally, the expression of efflux pumps that are implicated in regulation by quorum sensing (QS) allows for the uptake or efflux of molecules by biofilm cells [9].
This article presents an exhaustive review of the relationship between the treatment and monitoring of VVC, and the formation of biofilms.
Section snippets
General characteristics of biofilms
Biofilms (Fig. 1) are formed when a microbial community binds irreversibly to a substrate that then becomes embedded in a self-produced extracellular matrix. The formation of biofilms, as pointed out by Paiva et al., requires a series of steps that depend on the type of microorganism involved, but always begins with the adherence of the microorganism to a surface [11]. There are several factors that influence the formation of biofilms:
Fluids: The flow of fluids allows for the exchange of
Specific genes/pathways that regulate biofilm formation on mucosal surfaces and on abiotic surfaces: genes involved in resistance
The expression pattern of a diverse range of genes has been reported by Palkova et al. in cells that are part of the biofilm (either sessile or adherent), as well as in planktonic (free) cells, from different samples including mucous membranes [38]. Variations in the level of gene expression, such as genes encoding ribosomal proteins involved in protein synthesis, genes related to protein turnover, and translation, were clearly observed between the two different types of cells.
Experiments
Susceptibility of VVC isolates to antifungal treatment
The resistance and tolerance of biofilm cells to conventional antifungal agents (azoles, pyrimidine analogues, and polyenes) have led to the development of other strategies to combat them. Among these new strategies are the use of caspofungin, anidulafungin, and chelating agents. Combination therapies of several antifungals, such as amphotericin B, ethanol, and echinocandins have also been tried, with good results. Other combinations include lipid formulations of amphotericin B and
Probiotics in VVC/RVVC
According to Kang et al., lactobacilli form the most important defensive barrier against candidiasis infection, as evidenced by in vitro studies [149]. These act through different mechanisms such as competition between fungi for available nutrients, prevention of binding to epithelial receptors by blocking and the formation of co-aggregates, generation of substances such as H2O2, lactacins, and acidolines that are capable of inhibiting the germination of mycelia, enhancement of the immune
Future lines of research in VVC/RVVC-related studies
The formation of biofilms on the vaginal mucosa has been observed in in vivo models of VVC, in ex vivo models of reconstituted vaginal tissue, and in in vitro cultures of epithelial and fungal cell lines [13].
Within the virulence factors in Candida spp. the production of hydrolytic enzymes, their regulation, and the mechanisms by which they exert their effects have been the subject of study for a long time. Clinical and experimental evidence demonstrate that the secretory aspartyl proteinases
Conclusions
Biofilms represent the most common form of microbial growth in nature; their presence is essential for the development of VVC and RVVC, where C. albicans remains the most dominant species studied, though recently, non-C. albicans have been shown to be involved in an increasing number of these infections. Thus, a key role for the morphogenic changes experienced by the fungus, and the production of Sap as a virulence factor that activates the local inflammatory response, associated with the
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