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Vulvovaginal candidiasis (VVC) is a common clinical condition with symptoms and signs of vaginal inflammation in the presence of Candida species. At least one episode of VVC is experienced in up to 75% of women in the reproductive age group during their lifetime, and 5% to 8% of such women suffer from the chronic form. Most cases of VVC are still caused by C. albicans. However, the incidence of VVC cases by non-albicans Candida (NAC) species, such as C. parapsilosis, is continuously increasing. Despite the prevalence of VVC from NAC, little is known about these species and almost nothing about the mechanisms that trigger the VVC. Lactobacillus spp. are the most widely before represented microorganisms in the vaginal microbiota of healthy women. Here, cell-free supernatants (CFS) obtained from L. acidophilus, L. plantarum, L. rhamnosus, and L. reuteri were assessed for their effect on C. parapsilosis virulence traits. Moreover, we assessed if such an effect persisted even after the removal of the CFS (CFS preincubation effect). Moreover, a transwell coculture system was employed by which the relevant antifungal effect was shown to be attributable to the compounds released by lactobacilli. Our results suggest that lactobacilli can work (i) by reducing C. parapsilosis virulence traits, as indicated by the reduced fungal proliferation, viability, and metabolic activity, and (ii) by improving epithelial resistance to the fungus. Overall, these data suggest that, in the context of the vaginal microbiota, the lactobacilli may play a role in preventing the onset of mucosal C. parapsilosis infection. IMPORTANCE The incidence of VVC by non-albicans Candida (NAC) species, such as C. parapsilosis, is increasing. Treatment failure is common in NAC-VVC because some species are resistant or poorly susceptible to the antifungal agents normally employed. Research on C. parapsilosis's pathogenic mechanisms and alternative treatments are still lacking. C. albicans triggers the VVC by producing hyphae, which favor the loss of epithelial tolerance. Differently, C. parapsilosis only produces pseudohyphae. Hence, different virulence factors may trigger the VVC. Likewise, the therapeutic options could also involve different fungal targets. Substantial in vitro and in vivo studies on the pathogenicity mechanisms of C. parapsilosis are lacking. The data presented here ascribe a novel beneficial role to different Lactobacillus spp., whose CFS provides a postbiotic-like activity against C. parapsilosis. Further studies are needed to unravel the mechanisms involved in the bioactivities of such compounds, to better understand the role of single postbiotics in the CFS.

Vulvovaginal candidiasis (VVC), a major gynecological disease with high recurrence rate, increases the risk of abortion, intrauterine infection, premature rupture of membranes, and premature birth in pregnancy. However, the exact pathogenesis of this disease has yet to be elucidated. To facilitate understanding of the pathogenesis of VVC in pregnancy, this study sought to establish an animal model of vaginal infection with Candida albicans in pregnant mice. Female mice were mated with male mice, and female mice were infected with C. albicans at E4.5 (embryonic day 4.5). The weight and abortion rate of pregnant mice at E0.5, E4.5, E8.5, E11.5, and E18.5 were recorded, respectively, as well as the weights of fetus and placenta on E18.5. Fetal weight at E18.5 and the weight growth rate in the experimental mice was lower than those in the control mice, but the placenta weight at E18.5 and the abortion rate in the experimental mice were increased with those of the control mice. Hematoxylin-eosin (H&E) staining, Gomori-Grocott staining and vaginal lavage culturing were conducted to verify that the experimental mice were infected with C. albicans. Differentially expressed gene IL-15 was screened out by polymerase chain reaction (PCR) array between the two groups. Enzyme-linked immunosorbent assay (ELISA) showed that IL-15 expression in plasma of the mice was decreased in the experimental group compared with the control group. RT-qPCR confirmed that IL-15 mRNA expression was increased in placental tissues, while mRNA expression of IL-15R/JAK1-JAK3/PI3K/PDK1/AKT/P70S6K-mTOR was decreased in placental tissues. In conclusion, this study demonstrated that VVC in BALB/c pregnant mice led to a series of adverse pregnancy outcomes that were related to changes in IL-15 and its downstream signaling pathways, which may indicate a potential therapy for VVC during pregnancy in humans., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Candida albicans is a commensal of the urogenital tract and the predominant cause of vulvovaginal candidiasis (VVC). Factors that increase circulatory estrogen levels such as pregnancy, the use of oral contraceptives, and hormone replacement therapy predispose women to VVC, but the reasons for this are largely unknown. Here, we investigate how adaptation of C. albicans to estrogen impacts the fungal host-pathogen interaction. Estrogen promotes fungal virulence by enabling C. albicans to avoid the actions of the innate immune system. Estrogen-induced innate immune evasion is mediated via inhibition of opsonophagocytosis through enhanced acquisition of the human complement regulatory protein, Factor H, on the fungal cell surface. Estrogen-induced accumulation of Factor H is dependent on the fungal cell surface protein Gpd2. The discovery of this hormone-sensing pathway might pave the way in explaining gender biases associated with fungal infections and may provide an alternative approach to improving women's health., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Vulvovaginal candidiasis (VVC) is a common mucosal disease, caused primarily by Candida albicans that affects up to 75% of women of childbearing age. The pathogenesis of VVC and recurrent VVC (RVVC) is largely understood after decades of research. In this regard, an immunopathological response involving the migration of neutrophils that become dysfunctional (anergic) in the vaginal environment leads to the symptomatic conditions. However, immunotherapeutic strategies to correct the immunopathogenesis are still elusive. Much of the mechanistic discoveries have been uncovered using the established mouse model of chronic VVC. This chapter details the methods widely used for the mouse model of experimental VVC and associated outcome measures of the immunopathologic response and resulting symptomatic condition and focuses further on assays used to demonstrate "neutrophil anergy" in the model. These methods may serve as a source or resource for further experimentation with the ultimate goal to reduce or eliminate VVC/RVVC., (© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Vulvovaginal candidiasis (VVC), caused by Candida albicans, is a common infection in women affecting their quality of life. Standard antifungal drugs (e.g., fluconazole, itraconazole) are typically fungistatic or rendered ineffective due to drug resistance indicating an urgent need to build an arsenal of novel antifungal agents. To surmount this issue, we tested the hypothesis that the organoselenium compound ebselen (EB) possesses antifungal efficacy in a mouse model of VVC. EB is a poorly water-soluble drug and DMSO as a vehicle has the potential to exhibit cytotoxic effects when administered in vivo. EB loaded self-nanoemulsifying preconcentrate (EB-SNEP) was developed, characterized in vitro, and tested in a mouse model of VVC. In vivo studies carried out with EB-SNEP (12.5 mg/kg) showed a remarkable decrease in infection by ~562-fold compared to control (infected, untreated animals). Taken together, EB nanoemulsion proved to be an effective and promising antifungal agent., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Marine natural products are recognised as one among the major contributors of several important biological functions. The arguments has made to utilization of natural products against different kinds of infectious diseases. In the present study, Callophycin A was successfully prepared and its anti-candidal activity was evaluated through in-vitro and in-vivo methods. The in-vitro results revealed that, Callophycin A significantly inhibits the azole resistant and sensitive C. albicans. Further, in-vivo animal experiments have shown the effective reduction in CFU of C. albicans from its beginning day of the treatment as compared to the disease control group. At the end of Callophycin A administration, there was a decrease in inflammatory response and immune molecules such as IL-6, IL-12, IL-17, IL-22, TNF-α, macrophages, CD4 and CD8 cells were observed. Whereas the animals in the disease control group expressed all the parameters with the elevated level as compared to the control group. There are no hematological abnormalities such as neutropenia, lymphocytosis and eosinophilia was observed in any animal groups except the disease control group. Finally, the evidence based prediction of anti-candidal efficacious of Callophycin A was demonstrated., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The present study aimed to understand the killing effects of seaweed derived metabolite Callophycin A (Cal A). In vitro studies confirmed that the beneficial effects of Cal A on the viability of C. albicans. To enhance the biological activity, we used to demonstrated that chitosan and spicules as a drug carrier. The Callophycin A loading was confirmed by spectral variation of FT-IR and morphological variation by SEM. Moreover, around 65% and 38% of Cal A was successfully loaded in chitosan and spicules respectively. Further, VVC induced animal model experiments confirmed that the candidicidal activity of 1% clotrimazole, Cal A, Cal@Chi and Cal@Spi. After 6 days of treatment Cal@Chi produces a significant reduction in the fungal burden of vaginal lavage. The histo-morphological alterations also evidenced that the protective role of Cal@Chi in VVC model. The present investigations are known to be the first and foremost study to discriminate the potentiality of Cal A composites. Cal A loaded chitosan nanoparticles could be used as an alternative strategy for the development of the novel marine natural product based topical applications., Competing Interests: Declaration of competing interest We wish to confirm that there are no known conflicts of interest associated with this publication and there has been no significant financial support for this work that could have influenced its outcome., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Objective:  To identify clinical, microscopic, and biochemical characteristics that differentiate cytolytic vaginosis (CV) from vulvovaginal candidiasis (VVC)., Methods:  The present cross-sectional study analyzed the vaginal contents of 24 non-pregnant women aged 18 to 42 years who were attended at the Genital Infections Clinic at Centro de Atenção Integral à Saúde da Mulher da Universidade Estadual de Campinas (CAISM-UNICAMP). They were diagnosed either with (CV = 8, VVC = 8) or without vulvovaginitis or vaginal dysbiosis (controls). The socio-demographic, clinical, and gynecological data were obtained from a detailed patient interview. Samples of the vaginal contents were collected for analysis of vaginal pH, gram stain, and specific fungal culture. The Kruskal-Wallis and Fisher exact tests were used to compare the differences between the groups. Odds ratios were used to compare the categorical variables. The significance level was considered at p  < 0.05., Results:  Both women with CV and VVC had a lumpy vaginal discharge ( p  = 0,002) and vaginal hyperemia ( p  = 0.001), compared with controls. The inflammatory process was more intense in the VVC group ( p  = 0.001). In the CV group, there was statistical significance for the lactobacillus amount ( p  = 0.006), vaginal epithelium lysis ( p  = 0.001), and vaginal pH ( p  = 0.0002)., Conclusion:  Cytolytic vaginosis and VVC diagnoses rarely differ on clinical characteristics but have different laboratorial findings. The present study highlights the importance of conducting an accurate investigation through laboratory tests rather than clinical criteria to avoid misdiagnosis., Competing Interests: The authors have no conflict of interests to declare., (The Author(s). This is an open access article published by Thieme under the terms of the Creative Commons Attribution License, permitting unrestricted use, distribution, and reproduction so long as the original work is properly cited. (https://creativecommons.org/licenses/by/4.0/).)

Purposes: To investigate the role of 17β-estrogen in Candida albicans (C. albicans) adhesion on human vaginal epithelial cells in vulvovaginal candidiasis (VVC)., Methods: The vaginal epithelial cell line, VK2/E6E7, was used to study the estrogen-induced molecular events between C. albicans and cells. An adhesion study was performed to evaluate the involvement of the estrogen-dependent focal adhesion kinase (FAK) activation in cell adhesion. The phosphorylation status of FAK and estrogen receptor α (ERα) upon estrogen challenge was assessed by western blotting. Specific inhibitors for ERα were used to validate the involvement of ERα-FAK signaling cascade., Results: A transient activation of ERα and FAK was observed following the stimulation with 1000 nM estrogen for 48 h, as well as the increased average number of C. albicans adhering to each vaginal epithelial cell. Estrogen-induced activation of ERa and FAK was inhibited by the specific inhibitor of ERα, especially when the inhibitor reached a 10 μM concentration and allowed to act for 12 h. Simultaneously, a decrease in the number of adherent C. albicans was observed. However, this inhibitory effect diminished as the concentration of estrogen increased., Conclusion: FAK and ERα signaling cascades were involved in the early interaction between the vaginal epithelial cells and C. albicans, which appeared to be linked with the enhanced cell adhesion leading to VVC and promoted by a certain concentration of estrogen.

Candida albicans, a ubiquitous commensal fungus that colonizes human mucosal tissues and skin, can become pathogenic, clinically manifesting most commonly as oropharyngeal candidiasis and vulvovaginal candidiasis (VVC). Studies in mice and humans convincingly show that T-helper 17 (Th17)/interleukin 17 (IL-17)-driven immunity is essential to control oral and dermal candidiasis. However, the role of the IL-17 pathway during VVC remains controversial, with conflicting reports from human data and mouse models. Like others, we observed induction of a strong IL-17-related gene signature in the vagina during estrogen-dependent murine VVC. As estrogen increases susceptibility to vaginal colonization and resulting immunopathology, we asked whether estrogen use in the standard VVC model masks a role for the Th17/IL-17 axis. We demonstrate that mice lacking IL-17RA, Act1, or interleukin 22 showed no evidence for altered VVC susceptibility or immunopathology, regardless of estrogen administration. Hence, these data support the emerging consensus that Th17/IL-17 axis signaling is dispensable for the immunopathogenesis of VVC., (© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Recurrent vulvovaginal infections (RVVI) has not only become an epidemiological and clinical problem but also include large social and psychological consequences. Understanding the mechanisms of both commensalism and pathogenesis are necessary for the development of efficient diagnosis and treatment strategies for these enigmatic vaginal infections. Through this review, an attempt has been made to analyze vaginal microbiota (VMB) from scratch and to provide an update on its current understanding in relation to health and common RVVI i.e. bacterial vaginosis, vulvovaginal candidiaisis and Trichomoniasis, making the present review first of its kind. For this, potentially relevant studies were retrieved from data sources and critical analysis of the literature was made. Though, culture-independent methods have greatly unfolded the mystery regarding vaginal bacterial microbiome, there are only a few studies regarding the composition and diversity of vaginal mycobiome and different Trichomonas vaginalis strains. This scenario suggests a need of further studies based on comparative genomics of RVVI pathogens to improve our perceptive of RVVI pathogenesis that is still not clear (Fig. 5). Besides this, the review details the rationale for Lactobacilli dominance and changes that occur in healthy VMB throughout a women's life. Moreover, the list of possible agents continues to expand and new species recognised in both health and VVI are updated in this review. The review concludes with the controversies challenging the widely accepted dogma i.e. "VMB dominated with Lactobacilli is healthier than a diverse VMB". These controversies, over the past decade, have complicated the definition of vaginal health and vaginal infections with no definite conclusion. Thus, further studies on newly recognised microbial agents may reveal answers to these controversies. Conversely, VMB of women could be an answer but it is not enough to just look at the microbiology. We have to look at the woman itself, as VMB which is fine for one woman may be troublesome for others. These differences in women's response to the same VMB may be determined by a permutation of behavioural, cultural, genetic and various other anonymous factors, exploration of which may lead to proper definition of vaginal health and disease.

Objectives: This study evaluated use of long-term fluconazole beyond an initial 6-month course of weekly fluconazole in premenopausal patients with idiopathic recurrent vulvovaginal candidiasis (RVVC) due to Candida albicans., Materials and Methods: A retrospective chart review was performed of women seen in Wayne State University Vaginitis Clinic with culture-confirmed idiopathic RVVC due to Candida albicans during a 10-year period (January 2006 to December 2015). Only patients without risk factors for secondary VVC and who initiated a 6-month course of weekly fluconazole therapy were selected. Data included long-term use of fluconazole therapy, treatment efficacy, and development of fluconazole resistance. Questionnaires were mailed to evaluate patient's experience after fluconazole therapy., Results: Of 883 patients with RVVC based on clinical records, 191 with culture positive idiopathic RVVC due to C. albicans were started on the maintenance fluconazole regimen, and 147 (77.0%) completed 6 months of therapy. Of these, 107 (72.8%) continued or received maintenance past 6 months. The most common reason for additional fluconazole therapy was culture-confirmed VVC recurrence (55.1%), unconfirmed but possible VVC recurrence (16.8%), and patient preference (10.3%). The mean duration of fluconazole maintenance was 35.7 (range = 7-288) months. Fluconazole resistance emerged in 7.5% completing 6-month therapy. Upon questionnaire follow-up, 93.6% of 51 respondents reported benefit during maintenance regimen; however, 80.9% described relapse after discontinuing weekly therapy., Conclusions: Fluconazole suppression therapy was highly effective in preventing VVC symptoms but was rarely curative and VVC relapse occurred frequently after discontinuation of maintenance therapy. The development of drug resistance in C. albicans isolates after long-term fluconazole maintenance therapy although uncommon is a previously unrecognized complication.

Vulvovaginal candidiasis (VVC) is a common mucosal infection caused by Candida spp., most frequently by Candida albicans , which may become recurrent and severely impacting the quality of life of susceptible women. Although it is increasingly being recognized that mucosal damage is mediated by an exaggerated inflammatory response, current therapeutic approaches are only based on antifungals that may relieve the symptomatology, but fail to definitely prevent recurrences. The unrestrained activation of the NLRP3 inflammasome with continuous production of IL-1β and recruitment of neutrophils is recognized as a pathogenic factor in VVC. We have previously shown that IL-22 is required to dampen pathogenic inflammasome activation in VVC via the NLRC4/IL-1Ra axis. However, IL-22 also regulates IL-18, a product of the inflammasome activity that regulates IL-22 expression. Here we describe a cross-regulatory circuit between IL-18 and IL-22 in murine VVC that is therapeutically druggable. We found that IL-18 production was dependent on IL-22 and NLRC4, and that IL-18, in turn, contributes to IL-22 activity. Like in IL-22 deficiency, IL-18 deficiency was associated with an increased susceptibility to VVC and unbalanced Th17/Treg response, suggesting that IL-18 can regulate both the innate and the adaptive responses to the fungus. Administration of the microbial metabolite indole-3-aldehyde, known to stimulate the production of IL-22 via the aryl hydrocarbon receptor (AhR), promoted IL-18 expression and protection against Candida infection. Should low levels of IL-18 be demonstrated in the vaginal fluids of women with recurrent VVC, targeting the AhR/IL-22/IL-18 pathway could be exploited for future therapeutic approaches in VVC. This study suggests that a deeper understanding of the mechanisms regulating inflammasome activity may lead to the identification of novel targets for intervention in VVC., (Copyright © 2019 Borghi, Pariano, Solito, Puccetti, Bellet, Stincardini, Renga, Vacca, Sellitto, Mosci, Brancorsini, Romani and Costantini.)

Background: Vaginal candidiasis is common disease affecting women; however, how Candida albicans shift from commensalism towards a pathogenic status remains poorly understood. The present study investigated the vaginal epithelial cell (EC) response dynamics under various conditions., Methods: Healthy women, asymptomatic C. albicans carriers, and symptomatic patients with vaginal candidiasis were enrolled in this study. ECs in vaginal swabs were analyzed with cytofluorimetric analysis for pattern recognition receptors and intracellular signals, with lactate dehydrogenase assay performed for cell damage, and an enzyme-linked immunosorbent assay for cytokine expression., Results: The level of toll-like receptor 4 (TLR4), TLR2, and erythropoietin-producing hepatoma A2 (EphA2) expression was significantly higher in ECs from asymptomatic and symptomatic subjects compared to healthy subjects. Activation of transcription factors, nuclear factor-κB (NF-κB) and c-Fos-p-38, was observed in ECs from symptomatic and asymptomatic pseudohyphae/hyphae carriers but not from the asymptomatic yeast carriers. EC damage was only observed in symptomatic patients., Conclusions: The presence of pseudohyphae/hyphae is required to determine vaginal candidiasis; however, it may be not sufficient to induce the pathologic process associated with neutrophil recruitment and EC damage. This study sheds light on the ambiguous role of the hyphal form during vaginal human commensalism., (© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Vulvovaginal candidiasis (VVC) affects approximately 75% of all women of during their reproductive years. Previously, we reported that recombinant human IFN α-2b (rhIFNα-2b) protects vaginal epithelial cells from candidal injury in vitro. In the current study, we examined the effects of rhIFNα-2b (1.25 mg/mL, 10% inhibition concentration) on fungal clearance, immunocompetent cytokine responses, non-B IgG production, and tissue repair in a rat model of VVC. Following rhIFNα-2b treatment, the negative pathogen conversion rate reached 50.0% (3/6). Although rhIFNα-2b exhibited a limited ability to decrease inflammation and injury progression (P > 0.05), the Flameng mitochondrial injury scores were significantly reduced (P < 0.001) compared with those of the Model rats. After rhIFNα-2b treatment, the levels of IFN-γ and epithelial-derived IgG (tested by RP215) in vaginal tissues were significantly increased with those in the Control and Model groups (both P < 0.001), while there were no significant differences in the levels of IL-4 and IL-17 (P > 0.05). This is the first study to address the efficacy of rhIFNα-2b in treating VVC in a rat model, providing a theoretical basis for development of this promising treatment for clinical use., (Copyright © 2019. Published by Elsevier Ltd.)

Aim: The primary objective of this study was to evaluate the effects of polypeptide-enriched Gastrodia elata extracts (GE) on vulvovaginal candidiasis (VVC). Materials & methods: A VVC model induced by Candida albicans ( C. albicans ) infection was successfully developed in BALB/c mice. After treatment, the colony-forming unit (CFU) of vaginal lavage was measured by plating. The extent of the inflammatory response was assessed by hematoxylin-eosin (H&E) staining and enzyme-linked immunosorbent assay (ELISA). Results: GE had an inhibitory effect on the proliferation of C. albicans and inflammatory reaction. Meanwhile, it had a potentially beneficial effect on the growth of Lactobacillus . Conclusion: These results showed the potential application of GE as an antifungal agent in VVC treatment.

Background: Vulvovaginal candidiasis (VVC) is a recurrent vaginal condition in childbearing women., Objectives: The aim of this study was to assess the efficacy of an oral formulation containing Lactobacillus acidophilus GLA-14, Lactobacillus rhamnosus HN001 and bovine lactoferrin on symptoms and recurrence of VVC as adjuvant therapy to topical clotrimazole., Patients/methods: Forty-eight women positive for C. albicans, symptoms of VVC and documented history of recurrences were randomised into 2 groups receiving verum or placebo (2 capsules/day for 5 days followed by 1 capsule/day for additional 10 days) as adjuvant treatment to clotrimazole (induction phase) followed by a maintenance cycle of 6 months (1 capsule/day verum or placebo for 10 consecutive days each month). Symptoms, overall cure rate and recurrence rate were assessed., Results: After clotrimazole therapy, a significant improvement of symptoms was shown in both groups. However, only women treated with probiotics and lactoferrin showed a significant improvement of itching and discharge at 3 and 6 months. During the six-month follow-up, recurrences were significantly less in the intervention group vs placebo (33.3% vs 91.7% after 3 months and 29.2% vs 100% after 6 months)., Conclusions: The results show that the investigated lactobacilli mixture in combination with lactoferrin represents a safe and effective adjuvant approach for reducing symptoms and recurrences of RVVC., (© 2018 Blackwell Verlag GmbH.)

Candida species, including C. albicans, are part of the mucosal flora of most healthy women, and inhabit the gastrointestinal and genitourinary tracts. Under favourable conditions, they can colonize the vulvovaginal mucosa, giving rise to symptomatic vulvovaginal candidiasis (VVC). The mechanism by which Candida spp. produces inflammation is unknown. Both, the blastoconidia and the pseudohyphae are capable of destroying the vaginal epithelium by direct invasion. Although the symptoms are not always related to the fungal burden, in general, VVC is associated with a greater number of yeasts and pseudohyphae. Some years ago, C. albicans was the species most frequently involved in the different forms of VVC. However, infections by different species have emerged during the last two decades producing an increase in causative species of VVC such as C. glabrata, C. parapsilosis, C. krusei and C. tropicalis. Candida species are pathogenic organisms that have two forms of development: planktonic and biofilm. A biofilm is defined as a community of microorganisms attached to a surface and encompassed by an extracellular matrix. This form of presentation gives microorganisms greater resistance to antifungal agents. This review, about Candia spp. with a special emphasis on Candida albicans discusses specific areas such as biofilm structure and development, cell morphology and biofilm formation, biofilm-associated gene expression, the cell surface and adherence, the extracellular matrix, biofilm metabolism, and biofilm drug resistance in vulvovaginitis biofilms as an important virulence factor in fungi., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Vulvovaginal candidiasis (VVC) is caused mainly by the opportunistic fungus Candida albicans, and its yeast to hyphae transition is considered a major virulence factor. Farnesol is a molecule that inhibits yeast to hyphae transition. The increased incidence of VVC has influenced a need for developing new therapeutic strategies. The objective was to develop a mucoadhesive nanostructured system composed of miconazole and farnesol co-encapsulated within chitosan nanoparticles. The miconazole presented a minimal inhibitory concentration (MIC) of 1 μg/ml against C. albicans. The farnesol was capable of inhibiting yeast to hyphae transition at levels greater or equal to 300 μM. The combination of miconazole and farnesol showed no change in miconazole MIC. Chitosan nanoparticles containing miconazole and farnesol were prepared by ionic gelation and showed favorable characteristics for use on mucous membranes. They showed size variation and polydispersion index (PDI) after 30 days, but the efficiency of drug encapsulation was maintained. Regarding toxicity in cultured fibroblasts (BALB/c 3T3) the nanoparticles were considered nontoxic. The nanoparticles showed antifungal activity against the C. albicans strain used with MICs of 2.5 μg/ml and 2 μg/ml for nanoparticles containing miconazole or miconazole/farnesol, respectively. Nanoparticles containing farnesol inhibited yeast to hyphae transition at concentrations greater than or equal to 240 μM. The in vivo antifungal activity was assessed in the murine model for VVC. The results suggested that chitosan nanoparticles containing miconazole and farnesol were effective at inhibiting fungal proliferation. Additionally, chitosan nanoparticles containing farnesol were capable of decreasing the pathogenicity of infection, demonstrated through the absence of inflammation.

Background: The recent demonstration of a vaginal biofilm in bacterial vaginosis and its postulated importance in the pathogenesis of recurrent bacterial vaginosis, including relative resistance to therapy, has led to the hypothesis that biofilms are crucial for the development of vulvovaginal candidiasis. The histopathology and microbial architecture of vulvovaginal candidiasis have not been previously defined; neither has Candida, containing biofilm been reported in situ. The present study aimed at clarifying the histopathology of vulvovaginal candidiasis including the presence or absence of vaginal biofilm., Study Design: In a cross-sectional study, vaginal tissue biopsies were obtained from 35 women with clinically, microscopically, and culture-proven vulvovaginal candidiasis and compared with specimens obtained from 25 healthy women and 30 women with active bacterial vaginosis. Vaginal Candida infection was visualized using fluorescent in situ hybridization with ribosomal gene-based probes., Results: Candida microorganisms were confirmed in 26 of 35 biopsies obtained from women with vulvovaginal candidiasis; however, Candida containing biofilm were not detected in any of the cases. Histopathological lesions were exclusively invasive and accompanied by co-invasion with Gardnerella or Lactobacillus species organisms., Conclusion: Histopathological lesions of vulvovaginal candidiasis are primarily invasive in nature and polymicrobial and do not resemble biofilms. The clinical significance of Candida tissue invasion is unknown., (Copyright © 2018 Elsevier Inc. All rights reserved.)

The human fungal pathogen Candida albicans is the major etiological agent of vulvovaginal candidiasis (VVC). Despite this fact, other non- albicans Candida (NAC) species have frequently been reported, as well. Despite their presence in the vaginal environment, little is known about their capacities to elicit immune responses classically associated with C. albicans -mediated immunopathology, including neutrophil recruitment and proinflammatory cytokine signaling. Therefore, using a combination of in vitro and in vivo approaches, we undertook a comparative analysis to determine whether a representative panel of NAC species could colonize, induce immunopathological markers, or cause damage at the vaginal mucosa. Using a murine model of VVC, C. albicans was found to induce robust immunopathology (neutrophils and interleukin 1β [IL-1β]) and elicit mucosal damage. However, all the NAC species tested (including C. dubliniensis , C. tropicalis , C. parapsilosis , C. krusei , C. glabrata , and C. auris ) induced significantly less damage and neutrophil recruitment than C. albicans , despite achieving similar early colonization levels. These results largely correlated with a notable lack of ability by the NAC species (including C. dubliniensis and C. tropicalis ) to form hyphae both in vitro and in vivo Furthermore, both C. dubliniensis and C. tropicalis induced significantly less expression of the ECE1 gene encoding candidalysin, a key fungal virulence determinant driving VVC immunopathology. In order to determine the relative capacities of these species to elicit inflammasome-dependent IL-1β release, both wild-type and NLRP3 -/- THP-1 cells were challenged in vitro While most species tested elicited only modest amounts of IL-1β, challenge with C. albicans led to significantly elevated levels that were largely NLRP3 dependent. Collectively, our findings demonstrate that although NAC species are increasingly reported as causative agents of VVC, C. albicans appears to be exceedingly vaginopathogenic, exhibiting robust immunopathology, hypha formation, and candidalysin expression. Thus, this study provides mechanistic insight into why C. albicans is overwhelmingly the major pathogen reported during VVC., (Copyright © 2018 American Society for Microbiology.)

Vulvovaginal candidiasis (VVC) is a common fungal infection caused by Candida albicans . The antifungal therapy represents the standard of care but due to the high costs of treatment and to the inability to prevent recurrences, the development of alternative therapeutic approaches is much-awaited. Recently, we have shown that the pathogenesis of C. albicans in the gut is modulated by IL-9, a pleiotropic cytokine able to promote both inflammation and tolerance during C. albicans infection. Herein, by using a mouse model of VVC, we similarly demonstrated that IL-9 might exert a dual role in VVC by contributing to inflammation during the initial immune activation and promoting resolution thereafter. Specifically, IL-9 has a pro-inflammatory activity at the onset of VVC by promoting NLRP3 inflammasome activity and mucosal mast cells expansion but a tolerogenic role in the resolution phase by promoting IL-1Ra production and connective tissue mast cells activation. We further show that a timely IL-9 neutralization at the onset of the inflammatory response ameliorated symptoms and vaginal pathology. Given that vaginal fluids from patients with recurrent VVC had higher levels of IL-9, these findings, by providing novel insights into the pathogenesis of VVC, may pave the way for alternative therapeutic strategies based on IL-9 neutralization.

Recurrent vulvovaginal candidiasis is a debilitating, long-term condition that can severely affect the quality of life of affected women. No estimates of the global prevalence or lifetime incidence of this disease have been reported. For this systematic review, we searched PubMed, Embase, and Web of Science databases for population-based studies published between 1985 and 2016 that reported on the prevalence of recurrent vulvovaginal candidiasis, defined as four or more episodes of the infection every year. We identified 489 unique articles, of which eight were included, consisting of 17 365 patients from 11 countries. We generated estimates of annual global prevalence, estimated lifetime incidence and economic loss due to recurrent vulvovaginal candidiasis, and predicted the number of women at risk to 2030. Worldwide, recurrent vulvovaginal candidiasis affects about 138 million women annually (range 103-172 million), with a global annual prevalence of 3871 per 100 000 women; 372 million women are affected by recurrent vulvovaginal candidiasis over their lifetime. The 25-34 year age group has the highest prevalence (9%). By 2030, the population of women with recurrent vulvovaginal candidiasis each year is estimated to increase to almost 158 million, resulting in 20 240 664 extra cases with current trends using base case estimates in parallel with an estimated growth in females from 3·34 billion to 4·181 billion. In high-income countries, the economic burden from lost productivity could be up to US$14·39 billion annually. The high prevalence, substantial morbidity, and economic losses of recurrent vulvovaginal candidiasis require better solutions and improved quality of care for affected women., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Owing to the growing resistance among isolates of Candida species to usual antifungal agents and the well-known therapeutic potential of curcumin, the purpose of this study was to develop and validate a vaginal formulation containing this substance and to evaluating its effectiveness in the treatment of experimental vulvovaginal candidiasis. Curcumin was incorporated in a vaginal cream in three concentrations (0.01%, 0.1% and 1.0%). The different concentrations of the cream and its controls were intravaginally administered in an immunosuppressed rat model to evaluate the efficacy in the treatment of experimental vulvovaginal candidiasis. Samples of the cream were also subjected to centrifugation and physical stability tests and an analytical method for quantification of curcumin was validated based on HPLC. The formulation was stable and the HPLC method could be considered suitable for the quantitative determination of curcumin in the cream. After 6 days of preclinical study, the number of infected animals was 1/6 in all groups treated with curcumin vaginal cream and the fungal burden showed a progressive reduction. Reduction in the inflammatory infiltrate was observed in the group treated with 1.0% cream. Vaginal cream containing curcumin could be considered a promising effective antifungal medicine in the treatment of vulvovaginal candidiasis., (© 2018 Blackwell Verlag GmbH.)

Ethnopharmacological Relevance: Pulsatilla decoction (PD) is a classical prescription in Traditional Chinese Medicine (TCM) and has been reported to have inhibitory effects on Candida albicans proliferation., Study Aim: To investigate the therapeutic effects of PD in the treatment of Vulvovaginal candidiasis (VVC) and elucidate the potential mechanism., Materials and Methods: Female BALB/c mice (N = 90) were randomized to six treatment groups, including the Control group, Model group, three PD groups and Fluconazole group which served as a positive control (20 mg/kg weekly). The three PD groups (low dose group, medium dose group and high dose group) were given a daily intragastric gavage of PD at doses of 5, 10 and 20 g/kg, respectively. Five animals from each group were euthanized on Day 4, Day 7 and Day 14 after treatment. Colony forming unit (CFU) was measured by the serial dilution method. The degree of infection was assessed by Gram staining, Periodic acid schiff (PAS) staining, Hematoxylin and eosin (H&E) staining and Scanning electron microscopy (SEM). The serum inflammation levels were determined by a Luminex assay. Gene and protein expression levels of components of the Dectin-1 signaling pathway were determined by Real-time PCR, Western-blot and immunohistochemistry, respectively., Results: The administration of PD significantly decreased the fungal load from Day 7 post-infection onwards and decreased the number of visible microorganisms based on findings from Gram staining, PAS staining and SEM. H&E staining indicated that the impaired histological profiles were improved in all three PD groups. PD led to a significantly lower level of IL-23 in the serum; the levels of IL-10 and TNF-α were also decreased, although the differences were not significant. Furthermore, a substantial downregulation of Dectin-1, CARD9 and NF-κB mRNA levels and Dectin-1, Syk, CARD9 and NF-κB protein levels was observed after the administration of PD., Conclusion: This study suggests that PD exerts inhibitory effects on C. albicans proliferation, adhesion and inflammation and simultaneously downregulates the expression levels of important genes and proteins associated with the Dectin-1 pathway, highlighting the potential application of PD to improve the clinical management of VVC., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Objective: To characterize the lipid profile in vaginal discharge of women with vulvovaginal candidiasis, cytolytic vaginosis, or no vaginal infection or dysbiosis., Design: Cross-sectional study., Setting: Genital Infections Ambulatory, Department of Tocogynecology, University of Campinas, Campinas, São Paulo-Brazil., Sample: Twenty-four women were included in this study: eight with vulvovaginal candidiasis, eight with cytolytic vaginosis and eight with no vaginal infections or dysbiosis (control group)., Methods: The lipid profile in vaginal discharge of the different study groups was determined by liquid chromatography-mass spectrometry and further analyzed with MetaboAnalyst 3.0 platform., Main Outcome Measures: Vaginal lipids concentration and its correlation with vulvovaginal candidiasis and cytolytic vaginosis., Results: PCA, PLS-DA and hierarchical clustering analyses indicated 38 potential lipid biomarkers for the different groups, correlating with oxidative stress, inflammation, apoptosis and integrity of the vaginal epithelial tissue. Among these, greater concentrations were found for Glycochenodeoxycholic acid-7-sulfate, O-adipoylcarnitine, 1-eicosyl-2-heptadecanoyl-glycero-3-phosphoserine, undecanoic acid, formyl dodecanoate and lipoic acid in the vulvovaginal candidiasis group; N-(tetradecanoyl)-sphinganine, DL-PPMP, 1-oleoyl-cyclic phosphatidic, palmitic acid and 5-aminopentanoic acid in the cytolytic vaginosis group; and 1-nonadecanoyl-glycero-3-phosphate, eicosadienoic acid, 1-stearoyl-cyclic-phosphatidic acid, 1-(9Z,12Z-heptadecadienoyl)-glycero-3-phosphate, formyl 9Z-tetradecenoate and 7Z,10Z-hexadecadienoic acid in the control group., Conclusions: Lipids related to oxidative stress and apoptosis were found in higher concentrations in women with vulvovaginal candidiasis and cytolytic vaginosis, while lipids related to epithelial tissue integrity were more pronounced in the control group. Furthermore, in women with cytolytic vaginosis, we observed higher concentrations of lipids related to bacterial overgrowth., Competing Interests: The authors have declared that no competing interests exist.

Vaginal candidiasis is a common disorder in women of childbearing age, caused primarily by the dimorphic fungus Candida albicans. Since C. albicans is a normal commensal of the vaginal mucosa, a long-standing question is how the fungus switches from being a harmless commensal to a virulent pathogen. Work with human subjects and in mouse disease models suggests that host inflammatory processes drive the onset of symptomatic infection. Fungal cell wall molecules can induce inflammation through activation of epithelial and immune receptors that trigger pro-inflammatory cytokines and chemokines, but pathogenic fungi can evade recognition by masking these molecules. Knowledge about which cell wall epitopes are available for immune recognition during human infection could implicate specific ligands and receptors in the symptoms of vaginal candidiasis. To address this important gap, we directly probed the surface of fungi present in fresh vaginal samples obtained both from women with symptomatic Candida vaginitis and from women that are colonized but asymptomatic. We find that the pro-inflammatory cell wall polysaccharide β-glucan is largely masked from immune recognition, especially on yeast. It is only exposed on a small percentage of hyphal cells, where it tends to co-localize with enhanced levels of chitin. Enhanced β-glucan availability is only found in symptomatic patients with strong neutrophil infiltration, implicating neutrophils as a possible driver of these cell wall changes. This is especially interesting because neutrophils were recently shown to be necessary and sufficient to provoke enhanced β-glucan exposure in C. albicans, accompanied by elevated immune responses. Taken together, our data suggest that the architecture of C. albicans cell wall can be altered by environmental stress during vaginal candidiasis., Competing Interests: The authors have declared that no competing interests exist.

Introduction: Clinicians are increasingly challenged by patients with refractory vulvovaginal candidiasis (VVC) caused by azole-resistant Candida species. Fluconazole resistant C.albicans is a growing and perplexing problem following years of indiscriminate drug prescription and unnecessary drug exposure and for which there are few therapeutic alternatives. Regrettably, although the azole class of drugs has expanded, new classes of antifungal drugs have not been forthcoming, limiting effective treatment options in patients with azole resistant Candida vaginitis., Areas Covered: This review covers published data on epidemiology, pathophysiology and treatment options for women with azole-resistant refractory VVC., Expert Opinion: Fluconazole resistant C.albicans adds to the challenge of azole resistant non-albicans Candida spp. Both issues follow years of indiscriminate drug prescription and unnecessary fluconazole exposure. Although an understanding of azole resistance in yeast has been established, this knowledge has not translated into useful therapeutic advantage. Treatment options for such women with refractory symptoms are extremely limited. New therapeutic options and strategies are urgently needed to meet this challenge of azole drug resistance.

Vulvovaginal candidiasis is a common mucosal infection affecting a large proportion of women with some of them affected by recurrent often intractable forms of the disease. Thus, there is an increasing interest in understanding the pathogenesis of this disease. The aim of our work was to characterize, in animal models of vaginal candidiasis, the components of the host-fungus interaction at the mucosal level.The evidence of an immune response in the vaginal compartment was very encouraging to identify the proper targets for new strategies for vaccination or immunotherapy of vaginal candidiasis. Aspartyl-proteinase (Sap2), which is an important immunodominant antigens and virulence factors of C.albicans acting in mucosal infections, was assembled with virosomes and a vaccine PEV7 was obtained. The results obtained in the mouse model and in the clinical trial conducted by Pevion on women have evidenced that the vaccine PEV7, intravaginally administered, has an encouraging therapeutic potential for the treatment of recurrent vulvovaginal candidiasis. This opens the way to a modality for anti-Candida protection at mucosal level.

Previously we demonstrated that the treatment with live Saccharomyces cerevisiae exerts beneficial therapeutic effects against vaginal candidiasis. Here, we address potential mechanisms particularly examining the probiotic capacity to modulate both fungus and host-related factors. We show that the S. cerevisiae-based probiotic markedly affects the expression of virulence traits of Candida albicans such as aspartyl proteinases (SAPs) as well as hyphae-associated proteins Hwp1 and Ece1 in the vaginal cavity. On the host side, the probiotic suppression of the influx of neutrophils caused by the fungus into the vaginas of the mice is likely related to: (1) lower production of interleukin-8; and (2) inhibition of SAPs expression. However, these neutrophils displayed reactive oxygen species hyperproduction and increased killing activity as compared to the neutrophils of placebo-treated mice. There was no evidence of any cytotoxic effect by the probiotic, either when used in vivo on vaginal epithelial cell and organ architecture, or in in vitro in human vaginal epithelium. Inactivated yeast cells did not affect any of the factors above. In summary, the data suggest that the beneficial effect exerted by this S. cerevisiae-based probiotic is the result of its interference with the expression of fungus virulence factors coupled with the modulation of the inflammatory response of the host.

For over 3 decades, investigators have studied the pathogenesis of vulvovaginal candidiasis (VVC) and recurrent VVC (RVVC) through clinical studies and animal models. While there was considerable consensus that susceptibility was not associated with any apparent deficiencies in adaptive immunity, protective immune mechanisms and the role of innate immunity remained elusive. It was not until an innovative live-challenge design was conducted in women that a fuller understanding of the natural history of infection/disease was achieved. These studies revealed that symptomatic infection is associated with recruitment of polymorphonuclear neutrophils (PMNs) into the vaginal lumen. Subsequent studies in the established mouse model demonstrated that infiltrating PMNs were incapable of reducing the fungal burden, which supported the hypothesis that VVC/RVVC was an immunopathology, whereby Candida and the host response drive symptomatic disease. Further studies in mice revealed the requirement for C. albicans hyphae and identified pattern recognition receptors (PRRs) and proinflammatory mediators responsible for the PMN response, all of which are critical pieces of the immunopathogenesis. However, a mechanism explaining PMN dysfunction at the vaginal mucosa remained an enigma. Ultimately, by employing mouse strains resistant or susceptible to chronic VVC, it was determined that heparan sulfate (HS) in the vaginal environment of susceptible mice serves as a competitive ligand for Mac-1 on PMNs, which effectively renders the PMNs incapable of binding to Candida to initiate killing. Hence, the outcome of symptomatic VVC/RVVC is postulated to be dependent on a PMN-mediated immunopathogenic response involving HS that effectively places the neutrophils in a state of functional anergy., (Copyright © 2018 American Society for Microbiology.)

The female reproductive tract (FRT) presents a unique challenge to the mucosal immune system as it needs to monitor constantly for the presence of opportunistic pathogens amidst its commensal flora. During infection, autophagy plays a critical role in pathogen clearance, presentation of antigens and production of pro-inflammatory cytokines. However, no information is available that describes the role of autophagy in mouse vaginal infection of Candida albicans. The objective of our study is to evaluate the effect of autophagy gene, ATG5 knockout in vaginal cells in response to vaginal C. albicans infection. Mice having knockout of ATG5 in the vaginal cells (PR-ATG5-KO mice) were infected intra-vaginally with the yeast form of Candida albicans. Vaginal lavages were collected once in a week until the infection was cleared. We detected the expression of autophagy marker genes (LC3, ATG5 and LAMP1) in the vaginal cells. We determined the levels of various cytokines (IL-1α, IL-1β, IL-6, IL-10, IL-17A, IL-22, IL-23p19, TNF-α and G-CSF) involved in anti-candida response. The levels of cytokines in the vaginal lavages were quantified using Aimplex Premixed analyte kit. The vaginal lavages were checked for polymorphonuclear leucocytes (PMNLs) infiltration. The candida clearance rate from the vaginal lumen was determined by Colony Forming Units (CFUs) assay. The results revealed that PR-ATG5-KO mice failed to induce the expression of LC3, ATG5 and LAMP1 indicating an impaired autophagy pathway. The levels of all the cytokines (except IL-10) in C. albicans infected PR-ATG5-KO mice were significantly reduced as compared to the wild type infected C57BL/6 mice. The number of PMNLs infiltrated into the vaginal lavages of infected PR-ATG5-KO mice was reduced. The clearance of C. albicans from the vaginal lumen was also considerably delayed in PR-ATG5-KO mice. In conclusion, the results revealed that impaired autophagy in vaginal cells influences host response during vaginal infection of C. albicans by affecting anti-Candida cytokine levels in the vaginal lavage resulting in reduction of pathogen clearance rate., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Background/aims: Vulvovaginal candidiasis (VVC) is a disease commonly occurring in sexually active women. The involvement of microRNAs in several kinds of infectious diseases has been highlighted in a number of researches. Therefore, we conducted the present study in order to investigate whether microRNA-1192 (miR-1192) would significantly target CXCR4 in Th17 cells as well as inflammatory factors in mouse models suffering from VVC., Methods: Seventy-five mice were selected as test subjects for this study, of which twenty-five were used as the normal control, while the rest were treated with estradiol or oil-treated in order to establish VVC mouse models (each n = 25). Protein expressions of CXCR4, IL-6, IL-17, and IL-23 were all measured using both an immunohistochemistry and ELISA. The Th17 cell percentage in peripheral blood and the expression of RORγt in Th17 cells were detected using a flow cytometry. Mouse vaginal epithelial cells were isolated from normal mice, after which the mice were treated with estradiol to regulate their estrogen, followed by treatments involving the miR-1192 mimic, miR-1192 inhibitor, siRNA-CXCR4, and miR-1192 inhibitor + si-CXCR4. The cell cycle, apoptosis, and proliferation were all examined by using an additional flow cytometry as well as the employment of the MTT assay. The miR-1192, CXCR4, IL-6, IL-17, and IL-23 expressions in tissues and cells were both measured using both RT-qPCR and western blot assay techniques., Results: The mice treated with either estradiol or oil had presented to us lowered levels in miR-1192 expression as well as higher levels in both Th17 cell percentage and expression of RORγt in Th17 cells, along with mRNA and protein expressions of CXCR4, IL-6, IL-17, and IL-23. In cell experiments, the mouse vaginal epithelial cells that had been treated with miR-1192 inhibitor had shown us a decreased cell proliferation rate and contrarily increased expressions of CXCR4, IL-6, IL-17, and IL-23 mRNA, protein, and cell apoptosis rate; these results were opposite to the ones found in the mice treated with miR-1192 mimic., Conclusion: Our results provided significant evidence that miR-1192 could directly development and progression of VVC by restraining the CXCR4 gene in the VVC mice., (© 2018 The Author(s). Published by S. Karger AG, Basel.)

Objective: The aim of this study was to assess the correlation between atypical squamous cells (ASC) and inflammatory infiltrate and vaginal microbiota using cervical liquid-based cytological (SurePath®) and high-risk human papillomavirus (HR-HPV) tests., Study Design: A cross-sectional study was conducted using a 6-year database from a laboratory in Fortaleza (Brazil). Files from 1,346 ASC cases were divided into subgroups and results concerning inflammation and vaginal microorganisms diagnosed by cytology were compared with HR-HPV test results., Results: An absence of specific microorganisms (ASM) was the most frequent finding (ASC of undetermined significance, ASC-US = 74%; ASC - cannot exclude high-grade squamous intraepithelial lesion, ASC-H = 68%), followed by bacterial vaginosis (ASC-US = 20%; ASC- H = 25%) and Candida spp. (ASC-US = 6%; ASC-H = 5%). Leukocyte infiltrate was present in 71% of ASC-US and 85% of ASC-H (p = 0.0040), and in these specific cases HR-HPV tests were positive for 65 and 64%, respectively. A positive HR-HPV test was relatively more frequent when a specific microorganism was present, and Candida spp. was associated with HR-HPV-positive results (p = 0.0156), while an ASM was associated with negative HR-HPV results (p = 0.0370)., Conclusion: ASC-US is associated with an absence of inflammation or vaginosis, while ASC-H smears are associated with Trichomonas vaginalis and inflammatory infiltrate. A positive HR-HPV is associated with Candida spp. in ASC cytology., (© 2017 S. Karger AG, Basel.)

Candida spp. are part of the normal vaginal microflora in 20-30% of healthy women. However, if the balance between these yeasts and the host is disturbed, Candida spp. can cause vulvovaginal candidiasis (VVC), with Candida albicans being the major causative agent. Different studies have been performed in order to better understand Candida's dual lifestyle in the vagina. The potential of C. albicans to switch from the yeast cell morphology to its hyphal form is considered a key element in VVC pathogenesis. Candida spp. also express other virulence factors, such as hydrolytic extracellular enzymes and heat shock proteins and can form biofilms., (Copyright © 2017 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.)

Histatin5 (Hst-5) is a member of the histatin family of antimicrobial peptides secreted by human parotid and submandibular glands. With the natural antibacterial activity, it plays an important role in the first-line barrier of oral cave against pathogens, especially for the fungal intrusion. In this study, we explored the utility of Hst-5 in the treatment of vulvovaginal candidiasis, a common condition of women of the childbearing age. We used a synthesized Hst-5 over five consecutive days as the topical treatment in a murine model of vulvovaginal candidiasis. According to the fungal colony counts, fungal burden in the vagina lavage dropped remarkably after treatment with Hst-5. Furthermore, cytological analysis of the lavage fluid indicated that the number of cast-off cells including cornified epithelial cells and inflammatory cells also decreased; histological evaluation of the vagina tissue revealed less fungi adhering to the vaginal wall in treated animals than in controls. Combined, these results suggested for the first time the potential utility of Hst-5 as a topical treatment for vulvovaginal candidiasis, uncovering the possibility of exploiting the natural antibiotic peptides in other aspects., (© FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Problem: Autophagy plays an important role in clearance of intracellular pathogens. However, no information is available on its involvement in vaginal infections such as vulvo-vaginal candidiasis (VVC). VVC is intimately associated with the immune status of the human vaginal epithelial cells (VECs). The objective of our study is to decipher if autophagy process is involved during Candida albicans infection of VECs., Methods of Study: In this study, C. albicans infection system was established using human VEC line (VK2/E6E7). Infection-induced change in the expression of autophagy markers like LC3 and LAMP-1 were analyzed by RT-PCR, q-PCR, Western blot, immunofluorescence and transmission electron microscopy (TEM) studies were carried out to ascertain the localization of autophagosomes. Multiplex ELISA was carried out to determine the cytokine profiles., Results: Analysis of LC3 and LAMP-1 expression at mRNA and protein levels at different time points revealed up-regulation of these markers 6 hours post C. albicans infection. LC3 and LAMP-1 puncti were observed in infected VECs after 12 hours. TEM studies showed C. albicans entrapped in autophagosomes. Cytokines-TNF-α and IL-1β were up-regulated in culture supernatants of VECs at 12 hours post-infection., Conclusion: The results suggest that C. albicans invasion led to the activation of autophagy as a host defense mechanism of VECs., (© 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Despite acute inflammation by polymorphonuclear neutrophils (PMNs) during vulvovaginal candidiasis (VVC), clearance of Candida fails to occur. The purpose of this study was to uncover the mechanism of vaginal PMN dysfunction. Designs included assessing PMN migration, proinflammatory mediators, and tissue damage (by analysis of the activity of lactate dehydrogenase [LDH]) in mice susceptible (C3H/HeN-C57BL/6) or resistant (CD-1) to chronic VVC (CVVC-S or CVVC-R) and testing morphology-specific Candida albicans strains under conditions of preinduced PMN migration (CVVC-S mice) or PMN depletion (CVVC-R mice). In vitro designs included evaluation of C. albicans killing by elicited vaginal or peritoneal PMNs in standard or vaginal conditioned medium (VCM). Results showed that despite significant migration of PMNs and high levels of vaginal beta interleukin-1 (IL-1β) and alarmin S100A8, CVVC-S mice failed to reduce vaginal fungal burden irrespective of morphology or whether PMNs were present pre- or postinoculation, and had high LDH levels. In contrast, CVVC-R mice had reduced fungal burden and low LDH levels following PMN recruitment and IL-1β/S100A8 production, but maintained colonization in the absence of PMNs. Elicited vaginal and peritoneal PMNs showed substantial killing activity in standard media or VCM from CVVC-R mice but not in VCM from CVVC-S mice. The inhibitory effect of VCM from CVVC-S mice was unaffected by endogenous or exogenous estrogen and was ablated following depletion/neutralization of Mac-1 ligands using Mac-1 +/+ PMNs or recombinant Mac-1. Heparan sulfate (HS) was identified as the putative inhibitor as evidenced by the rescue of PMN killing following heparanase treatment of VCM, as well as by inhibition of killing by purified HS. These results suggest that vaginal HS is linked to PMN dysfunction in CVVC-S mice as a competitive ligand for Mac-1. IMPORTANCE Vaginal candidiasis, caused by Candida albicans , affects a significant number of women worldwide. Despite an acute inflammatory response by neutrophils during infection, the response fails to reduce the organism. Instead, the response is considered a key process underlying the symptoms of vaginitis. Therefore, it is important to determine the mechanism(s) associated with the lack of vaginal neutrophil antifungal activity. The established mouse model of Candida vaginitis was used to uncover the mechanism of neutrophil dysfunction. Results revealed that heparan sulfate present in the vagina of mice susceptible to chronic vaginitis served as a competitive ligand for the receptor (Mac-1) necessary for fungal recognition and neutrophil-mediated killing. This inhibitory function of heparan sulfate, confirmed through several approaches, provides the first evidence to explain the lack of antifungal immune reactivity during vaginal candidiasis. This finding paves the way for design of therapeutic strategies to reduce/eliminate symptomatic vaginal candidiasis and restore quality of life to those affected., (Copyright © 2017 Yano et al.)

Objectives: This study aimed to determine if vulvar cutaneous candidosis and dermatophytosis can be distinguished by routine histopathology., Materials and Methods: Twenty-four cases of periodic acid-Schiff-stained vulvar biopsies with a diagnosis of cutaneous mycosis were reviewed and histopathological characteristics on both periodic acid-Schiff and hematoxylin and eosin were recorded. Data were collected on age, clinical impression, microbiological results, and treatment, and all specimens underwent multiplex polymerase chain reaction analysis., Results: The mean age was 60 years, and all but 3 women had at least 1 risk factor for mycosis including 15 (62.5%) with lichen sclerosus and/or planus managed with topical corticosteroids. A clinical suspicion of tinea or candidosis was documented in 12 (50%) of the cases. Vulvovaginal swabs showed Candida species in 9 women; one skin scraping was positive for Trichophyton rubrum. Microbiology was not obtained in 8 patients, 5 had a negative swab, and 1 had negative skin scrapings. No histopathological or morphological features distinguished Candida species from dermatophytes. Organisms appeared as basophilic structures in the stratum corneum in 15 (62.5%) hematoxylin and eosin-stained slides. Polymerase chain reaction results were positive for Candida species in 5 (21%) and for dermatophytes in 3 (13%), negative in 13, and unassessable in 3 cases., Conclusions: Vulvar cutaneous candidosis and dermatophytosis cannot be reliably distinguished by routine histopathology or specific polymerase chain reaction. A high index of suspicion combined with adequate microbiological testing remains the best approach to differentiating between the 2, which impacts on counseling, treatment, and prognosis.

Vulvovaginal candidiasis (VVC) is a high-incidence disease seriously affecting the quality of life of women worldwide, particularly in its chronic, recurrent forms (RVVC), and with no definitive cure or preventive measure. Experimental studies in currently used rat and mouse models of vaginal candidiasis have generated a large mass of data on pathogenicity determinants and inflammation and immune responses of potential importance for the control of human pathology. However, reflection is necessary about the relevance of these rodent models to RVVC. Here we examine the chemical, biochemical, and biological factors that determine or contrast the forms of the disease in rodent models and in women and highlight the differences between them. We also appeal for approaches to improve or replace the current models in order to enhance their relevance to human infection., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)

Mouse model is an appropriate tool for pathogenic determination and study of host defenses during the fungal infection. Here, we established a mouse model of candidiasis with concurrent oral and vaginal mucosal infection. Two C. albicans strains sourced from clinical candidemia (SC5314) and mucosal infection (ATCC62342) were tested in ICR mice. The different combinational panels covering estrogen and immunosuppressive agents, cortisone, prednisolone and cyclophosphamide were used for concurrent oral and vaginal candidiasis establishment. Prednisolone in combination with estrogen proved an optimal mode for concurrent mucosal infection establishment. The model maintained for 1 week with fungal burden reached at least 10(5) cfu/g of tissue. This mouse model was evaluated by in vivo pharmacodynamics of fluconazole and host mucosal immunity of IL-17 and IL-23. Mice infected by SC5314 were cured by fluconazole. An increase in IL-23 in both oral and vaginal homogenates was observed after infection, while IL-17 only had a prominent elevation in oral tissue. This model could properly mimic complicated clinical conditions and provides a valuable means for antifungal assay in vivo and may also provide a useful method for the evaluation of host-fungal interactions.

Candida glabrata is the second most common organism isolated from women with vulvovaginal candidiasis (VVC), particularly in women with uncontrolled diabetes mellitus. However, mechanisms involved in the pathogenesis of C. glabrata-associated VVC are unknown and have not been studied at any depth in animal models. The objective of this study was to evaluate host responses to infection following efforts to optimize a murine model of C. glabrata VVC. For this, various designs were evaluated for consistent experimental vaginal colonization (i.e., type 1 and type 2 diabetic mice, exogenous estrogen, varying inocula, and co-infection with C. albicans). Upon model optimization, vaginal fungal burden and polymorphonuclear neutrophil (PMN) recruitment were assessed longitudinally over 21 days post-inoculation, together with vaginal concentrations of IL-1β, S100A8 alarmin, lactate dehydrogenase (LDH), and in vivo biofilm formation. Consistent and sustained vaginal colonization with C. glabrata was achieved in estrogenized streptozotocin-induced type 1 diabetic mice. Vaginal PMN infiltration was consistently low, with IL-1β, S100A8, and LDH concentrations similar to uninoculated mice. Biofilm formation was not detected in vivo, and co-infection with C. albicans did not induce synergistic immunopathogenic effects. This data suggests that experimental vaginal colonization of C. glabrata is not associated with an inflammatory immunopathogenic response or biofilm formation.

Several important classes of antifungal agents, including the azoles, act by blocking ergosterol biosynthesis. It was recently reported that the azoles cause massive disruption of the fungal vacuole in the prevalent human pathogen Candida albicans. This is significant because normal vacuolar function is required to support C. albicans pathogenicity. This study examined the impact of the morpholine antifungals, which inhibit later steps of ergosterol biosynthesis, on C. albicans vacuolar integrity. It was found that overexpression of either the ERG2 or ERG24 gene, encoding C-8 sterol isomerase or C-14 sterol reductase, respectively, suppressed C. albicans sensitivity to the morpholines. In addition, both erg2Δ/Δ and erg24Δ/Δ mutants were hypersensitive to the morpholines. These data are consistent with the antifungal activity of the morpholines depending upon the simultaneous inhibition of both Erg2p and Erg24p. The vacuoles within both erg2Δ/Δ and erg24Δ/Δ C. albicans strains exhibited an aberrant morphology and accumulated large quantities of the weak base quinacrine, indicating enhanced vacuolar acidification compared with that of control strains. Both erg mutants exhibited significant defects in polarized hyphal growth and were avirulent in a mouse model of disseminated candidiasis. Surprisingly, in a mouse model of vaginal candidiasis, both mutants colonized mice at high levels and induced a pathogenic response similar to that with the controls. Thus, while targeting Erg2p or Erg24p alone could provide a potentially efficacious therapy for disseminated candidiasis, it may not be an effective strategy to treat vaginal infections. The potential value of drugs targeting these enzymes as adjunctive therapies is discussed., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Unlabelled: Vaginal inflammation (vaginitis) is the most common disease caused by the human-pathogenic fungus Candida albicans. Secretory aspartyl proteinases (Sap) are major virulence traits of C. albicans that have been suggested to play a role in vaginitis. To dissect the mechanisms by which Sap play this role, Sap2, a dominantly expressed member of the Sap family and a putative constituent of an anti-Candida vaccine, was used. Injection of full-length Sap2 into the mouse vagina caused local neutrophil influx and accumulation of the inflammasome-dependent interleukin-1β (IL-1β) but not of inflammasome-independent tumor necrosis factor alpha. Sap2 could be replaced by other Sap, while no inflammation was induced by the vaccine antigen, the N-terminal-truncated, enzymatically inactive tSap2. Anti-Sap2 antibodies, in particular Fab from a human combinatorial antibody library, inhibited or abolished the inflammatory response, provided the antibodies were able, like the Sap inhibitor Pepstatin A, to inhibit Sap enzyme activity. The same antibodies and Pepstatin A also inhibited neutrophil influx and cytokine production stimulated by C. albicans intravaginal injection, and a mutant strain lacking SAP1, SAP2, and SAP3 was unable to cause vaginal inflammation. Sap2 induced expression of activated caspase-1 in murine and human vaginal epithelial cells. Caspase-1 inhibition downregulated IL-1β and IL-18 production by vaginal epithelial cells, and blockade of the IL-1β receptor strongly reduced neutrophil influx. Overall, the data suggest that some Sap, particularly Sap2, are proinflammatory proteins in vivo and can mediate the inflammasome-dependent, acute inflammatory response of vaginal epithelial cells to C. albicans. These findings support the notion that vaccine-induced or passively administered anti-Sap antibodies could contribute to control vaginitis., Importance: Candidal vaginitis is an acute inflammatory disease that affects many women of fertile age, with no definitive cure and, in its recurrent forms, causing true devastation of quality of life. Unraveling the fungal factors causing inflammation is important to be able to devise novel tools to fight the disease. In an experimental murine model, we have discovered that aspartyl proteinases, particularly Sap2, may cause the same inflammatory signs of vaginitis caused by the fungus and that anti-Sap antibodies and the protease inhibitor Pepstatin A almost equally inhibit Sap- and C. albicans-induced inflammation. Sap-induced vaginitis is an early event during vaginal infection, is uncoupled from fungal growth, and requires Sap and caspase-1 enzymatic activities to occur, suggesting that Sap or products of Sap activity activate an inflammasome sensor of epithelial cells. Our data support the notion that anti-Sap antibodies could help control the essence of candidal vaginitis, i.e., the inflammatory response., (Copyright © 2015 Pericolini et al.)

Unlabelled: Treatment of vulvovaginal candidiasis (VVC), caused most frequently by Candida albicans, represents a significant unmet clinical need. C. albicans, as both a commensal and a pathogenic organism, has a complex and poorly understood interaction with the vaginal environment. Understanding the complex nature of this relationship is necessary for the development of desperately needed therapies to treat symptomatic infection. Using transcriptome sequencing (RNA-seq), we characterized the early murine vaginal and fungal transcriptomes of the organism during VVC. Network analysis of host genes that were differentially expressed between infected and naive mice predicted the activation or repression of several signaling pathways that have not been previously associated with VVC, including NLRP3 inflammasome activation. Intravaginal challenge of Nlrp3(-/-) mice with C. albicans demonstrated severely reduced levels of polymorphonuclear leukocytes (PMNs), alarmins, and inflammatory cytokines, including interleukin-1β (IL-1β) (the hallmarks of VVC immunopathogenesis) in vaginal lavage fluid. Intravaginal administration of wild-type (WT) mice with glyburide, a potent inhibitor of the NLRP3 inflammasome, reduced PMN infiltration and IL-1β to levels comparable to those observed in Nlrp3(-/-) mice. Furthermore, RNA-seq analysis of C. albicans genes indicated robust expression of hypha-associated secreted aspartyl proteinases 4, 5, and 6 (SAP4-6), which are known inflammasome activators. Despite colonization similar to that of the WT strain, ΔSAP4-6 triple and ΔSAP5 single mutants induced significantly less PMN influx and IL-1β during intravaginal challenge. Our findings demonstrate a novel role for the inflammasome in the immunopathogenesis of VVC and implicate the hypha-associated SAPs as major C. albicans virulence determinants during vulvovaginal candidiasis., Importance: Vaginitis, most commonly caused by the fungus Candida albicans, results in significant quality-of-life issues for all women of reproductive age. Recent efforts have suggested that vaginitis results from an immunopathological response governed by host innate immunity, although an explanatory mechanism has remained undefined. Using comprehensive genomic, immunological, and pharmacological approaches, we have elucidated the NLRP3 inflammasome as a crucial molecular mechanism contributing to host immunopathology. We have also demonstrated that C. albicans hypha-associated secreted aspartyl proteinases (SAP4-6 and SAP5, more specifically) contribute to disease immunopathology. Ultimately, this study enhances our understanding of the complex interplay between host and fungus at the vaginal mucosa and provides proof-of-principle evidence for therapeutic targeting of inflammasomes for symptomatic vulvovaginal candidiasis., (Copyright © 2015 Bruno et al.)

Objective: This study aimed to observe the morphological characteristic of vaginal discharge in patients with cytolytic vaginosis (CV) under the microscope and to identify it in patients with CV and in patients with vulvovaginal candidiasis (VVC)., Methods: A total of 108 subjects including 21 healthy women, 33 patients with CV, and 54 patients with VVC were enrolled in the present morphological study. Vaginal discharge was collected and made into smear. The morphological characteristics of these vaginal smears with Gram staining were observed under the microscope. The smears were assessed for the quantity of lactobacilli, epithelial cell morphology, and the absence or presence of Candida species, Trichomonas vaginalis, and clue sells., Results: First, the age, the level of education, and especially the status of pregnancy of patients with CV were significantly different from those of the patients with VVC. Second, the morphological characteristics of patients with CV consisted of overgrowth of lactobacilli, the presence of naked nuclei and fragments of the epithelial cells, a paucity of leukocytes, and the absence of Candida species and other pathogens. However, the morphological characteristic of patients with VVC consisted of the presence or absence of lactobacilli and the presence of normal epithelial cells, candidal spores, blastospores, hyphae, or other pathogens such as T. vaginalis and Gardnerella vaginalis., Conclusions: Both CV and VVC can be identified based on the quantity of lactobacilli, the morphology of the epithelial cells, and the absence or presence of Candida species and other pathogens, and the misdiagnosis of CV as VVC can be avoided.

Objective: Is it feasible to learn the basics of wet mount microscopy of vaginal fluid in 10 hours?, Materials and Methods: This is a pilot project wherein 6 students with different grades of education were invited for being tested on their ability to read wet mount microscopic slides before and after 10 hours of hands-on training. Microscopy was performed according to a standard protocol (Femicare, Tienen, Belgium). Before and after training, all students had to evaluate a different set of 50 digital slides. Different diagnoses and microscopic patterns had to be scored. κ indices were calculated compared with the expert reading., Results: All readers improved their mean scores significantly, especially for the most important types of altered flora (p < .0001). The mean increase in reading concordance (κ from 0.64 to 0.75) of 1 student with a solid previous experience with microscopy did not reach statistical significance, but the remaining 5 students all improved their scores from poor performance (all κ < 0.20) to moderate (κ = 0.53, n = 1) to good (κ > 0.61, n = 4) concordance. Reading quality improved and reached fair to good concordance on all microscopic items studied, except for the detection of parabasal cells and cytolytic flora., Conclusions: Although further improvement is still possible, a short training course of 10 hours enables vast improvement on wet mount microscopy accuracy and results in fair to good concordance of the most important variables of the vaginal flora compared to a reference reader.

The concept of infertility as a result of asymptomatic microbial colonization of the female reproductive tract has been neglected to date. However, increasing incidence of infertility and advanced research has drawn attention towards this idea. Many of these micro-organisms have been reported to bring about adverse changes in sperm parameters in vitro, but their in vivo potential to cause infertility is still a controversy. The present study was carried out to observe what effect the intravaginal inoculation of sperm-agglutinating Serratia marcescens and sperm-immobilizing Candida albicans had in the reproductive tract and consequently in fertility outcome. When these strains were intravaginally inoculated into female BALB/c mice at 10(4), 10(6) and 10(8) c.f.u. in 20 µl PBS for 10 consecutive days, with mating of mice on day 12, the results showed 100 % decrease in fertility in all groups as compared with control mice receiving PBS alone. Furthermore, no clinical or histopathological changes were observed in the reproductive organs (i.e. ovary, uterus and vagina), suggesting that colonization of the genital tract with sperm-impairing micro-organisms could be a feasible reason for female infertility., (© 2015 The Authors.)

Candida africana, an emerging yeast pathogen, is closely related to Candida albicans and most commonly involved in vulvovaginal candidiasis (VVC). However, its prevalence in candidal balanoposthitis is still unclear. In this study, the prevalence of C. africana in both candidal balanoposthitis and VVC in a sexually transmitted diseases (STD) clinic in Shanghai, China, was analyzed, and the molecular characterization and susceptible profiles of C. africana isolates were investigated. As results, C. africana was only isolated in 5 out of 79 (6.3%) cases of candidal balanoposthitis rather than cases with vulvovaginal candidiasis. Among them, 4 out of 5 isolates share the same genotype of DST 782 with an isolate from vaginal swab in Japan published previously. All C. africana isolates were susceptible to amphotericin B, flucytosine, fluconazole, itraconazole, voriconazole, posaconazole, caspofungin, and micafungin.