Search

Purpose: Loop-mediated isothermal amplification (LAMP) is a simple and rapid nucleic acid method for DNA amplification at a constant temperature. The "gold standard" culture method for yeast detection, has low sensitivity with severe consequences, increasing morbidity and mortality rates. Here, we report the development of a LAMP method for the specific detection of C. glabrata. Methodology: The specific LAMP primers for C. glabrata detection were designed and evaluated. Results: The LAMP assay accurately detected C. glabrata with no cross-reactivity with other Candida species. Conclusion: The developed molecular method would be a promising tool in the management of invasive candidiasis. [ABSTRACT FROM AUTHOR]

Background: Candida africana is an emergent variant that has been listed as a new species or variety within the Candida albicans complex since 2001. It has a worldwide intra-albicans complex pooled prevalence of 1.67% and varies between 0 and 8% depending on geographical region. We present the results of a pilot study on its prevalence in Uganda. Methodology: We conducted a cross-sectional study between March and June 2023. We recruited 4 pregnant women from Mulago Specialized Women and Neonatal Hospital, 102 from Kawempe National Referral Hospital, and 48 from Sebbi Hospital. Vaginal swabs were tested using microscopy, culture and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF). Results: The prevalence of C. africana was zero. Out of the 103 isolates, the majority (81.553%) were identified as Candida albicans, followed by Nakeseomyces glabrata (13.592%) and Pichia kudriavzevii (1.942%). Cyberlindnera jadinii, Candida tropicalis, and Candida parapsilosis each accounted for 0.971% of the isolates. Conclusion: The prevalence of C. africana in Uganda is zero. However, large-scale cross-sectional studies, including studies involving the collection of vaginal samples from both urban and rural settings in Uganda and the use of both MALDI-TOF- and PCR-based laboratory methods, are needed to fully describe the public health burden of C. africana infections. [ABSTRACT FROM AUTHOR]

Abstract Background Candida africana is an emergent variant that has been listed as a new species or variety within the Candida albicans complex since 2001. It has a worldwide intra-albicans complex pooled prevalence of 1.67% and varies between 0 and 8% depending on geographical region. We present the results of a pilot study on its prevalence in Uganda. Methodology We conducted a cross-sectional study between March and June 2023. We recruited 4 pregnant women from Mulago Specialized Women and Neonatal Hospital, 102 from Kawempe National Referral Hospital, and 48 from Sebbi Hospital. Vaginal swabs were tested using microscopy, culture and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF). Results The prevalence of C. africana was zero. Out of the 103 isolates, the majority (81.553%) were identified as Candida albicans, followed by Nakeseomyces glabrata (13.592%) and Pichia kudriavzevii (1.942%). Cyberlindnera jadinii, Candida tropicalis, and Candida parapsilosis each accounted for 0.971% of the isolates. Conclusion The prevalence of C. africana in Uganda is zero. However, large-scale cross-sectional studies, including studies involving the collection of vaginal samples from both urban and rural settings in Uganda and the use of both MALDI-TOF- and PCR-based laboratory methods, are needed to fully describe the public health burden of C. africana infections.

Abstract Background Biofilm formation is viewed as a vital mechanism in C. glabrata pathogenesis. Although, it plays a significant role in virulence but transcriptomic architecture and metabolic pathways governing the biofilm growth mode of C. glabrata remain elusive. The present study intended to investigate the genes implicated in biofilm growth phase of C. glabrata through global transcriptomic approach. Results Functional analysis of Differentially expressed genes (DEGs) using gene ontology and pathways analysis revealed that upregulated genes are involved in the glyoxylate cycle, carbon-carbon lyase activity, pre-autophagosomal structure membrane and vacuolar parts whereas, down- regulated genes appear to be associated with glycolysis, ribonucleoside biosynthetic process, ribosomal and translation process in the biofilm growth condition. The RNA-Seq expression of eight selected DEGs (CgICL1, CgMLS1, CgPEP1, and CgNTH1, CgERG9, CgERG11, CgTEF3, and CgCOF1) was performed with quantitative real-time PCR (RT-qPCR). The gene expression profile of selected DEGs with RT-qPCR displayed a similar pattern of expression as observed in RNA-Seq. Phenotype screening of mutant strains generated for genes CgPCK1 and CgPEP1, showed that Cgpck1∆ failed to grow on alternative carbon substrate (Glycerol, Ethanol, Oleic acid) and similarly, Cgpep1∆ unable to grow on YPD medium supplemented with hydrogen peroxide. Our results suggest that in the absence of glucose, C. glabrata assimilate glycerol, oleic acid and generate acetyl coenzyme-A (acetyl-CoA) which is a central and connecting metabolite between catabolic and anabolic pathways (glyoxylate and gluconeogenesis) to produce glucose and fulfil energy requirements. Conclusions The study was executed using various approaches (transcriptomics, functional genomics and gene deletion) and it revealed that metabolic plasticity of C. glabrata (NCCPF-100,037) in biofilm stage modulates its virulence and survival ability to counter the stress and may promote its transition from commensal to opportunistic pathogen. The observations deduced from the present study along with future work on characterization of the proteins involved in this intricate process may prove to be beneficial for designing novel antifungal strategies.

In the face of increased frequency of non-albicans Candida vulvovaginitis (VVC) reported worldwide, there is a paucity of effective oral and topical antifungal drugs available. Drug selection is further handicapped by an absence of data of clinical efficacy of available antifungal drugs for these infections. In this review, attention is directed at the cause of drug shortage as well as increased frequency of non-albicans Candida (NAC) vulvovaginitis. There is widespread recognition of reduced in vitro azole drug susceptibility in NAC species. Moreover, antifungal susceptibility tests have not been standardized or validated for NAC isolates, hence clinicians rely on an element of empiricism especially given the absence of randomized controlled comparative studies targeting NAC species. Clinical spectrum of NAC species isolates is highly variable with ongoing difficulty in determining a causal role in symptomatic patients. We have entered the era of demand for Candida species-specific therapy and although consensus treatment guidelines are emerging, new antifungal agents that target these multiple-azole resistant or relatively resistant vaginal NAC species are urgently needed. [ABSTRACT FROM AUTHOR]

Background: Biofilm formation is viewed as a vital mechanism in C. glabrata pathogenesis. Although, it plays a significant role in virulence but transcriptomic architecture and metabolic pathways governing the biofilm growth mode of C. glabrata remain elusive. The present study intended to investigate the genes implicated in biofilm growth phase of C. glabrata through global transcriptomic approach. Results: Functional analysis of Differentially expressed genes (DEGs) using gene ontology and pathways analysis revealed that upregulated genes are involved in the glyoxylate cycle, carbon-carbon lyase activity, pre-autophagosomal structure membrane and vacuolar parts whereas, down- regulated genes appear to be associated with glycolysis, ribonucleoside biosynthetic process, ribosomal and translation process in the biofilm growth condition. The RNA-Seq expression of eight selected DEGs (CgICL1, CgMLS1, CgPEP1, and CgNTH1, CgERG9, CgERG11, CgTEF3, and CgCOF1) was performed with quantitative real-time PCR (RT-qPCR). The gene expression profile of selected DEGs with RT-qPCR displayed a similar pattern of expression as observed in RNA-Seq. Phenotype screening of mutant strains generated for genes CgPCK1 and CgPEP1, showed that Cgpck1∆ failed to grow on alternative carbon substrate (Glycerol, Ethanol, Oleic acid) and similarly, Cgpep1∆ unable to grow on YPD medium supplemented with hydrogen peroxide. Our results suggest that in the absence of glucose, C. glabrata assimilate glycerol, oleic acid and generate acetyl coenzyme-A (acetyl-CoA) which is a central and connecting metabolite between catabolic and anabolic pathways (glyoxylate and gluconeogenesis) to produce glucose and fulfil energy requirements. Conclusions: The study was executed using various approaches (transcriptomics, functional genomics and gene deletion) and it revealed that metabolic plasticity of C. glabrata (NCCPF-100,037) in biofilm stage modulates its virulence and survival ability to counter the stress and may promote its transition from commensal to opportunistic pathogen. The observations deduced from the present study along with future work on characterization of the proteins involved in this intricate process may prove to be beneficial for designing novel antifungal strategies. [ABSTRACT FROM AUTHOR]

Fungal infections caused by Candida are still a public health concern. Particularly, the resistance to traditional chemotherapeutic agents is a major issue that requires efforts to develop new therapies. One of the most interesting approaches to finding new active compounds is drug repurposing aided by computational methods. In this work, two databases containing anticandidal agents and drugs were studied employing cheminformatics and compared by similarity methods. The results showed 36 drugs with high similarities to some candicidals. From these drugs, trimetozin, osalmid and metochalcone were evaluated against C. albicans (18804), C. glabrata (90030), and miconazole‐resistant strain C. glabrata (32554). Osalmid and metochalcone were the best, with activity in the micromolar range. These findings represent an opportunity to continue with the research on the potential antifungal application of osalmid and metochalcone as well as the design of structurally related derivatives. [ABSTRACT FROM AUTHOR]

Candida glabrata and Candida albicans, the most frequently isolated candidiasis species in the world, have developed mechanisms of resistance to treatment with azoles. Among the clinically used antifungal drugs are statins and other compounds that inhibit 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR), resulting in decreased growth and ergosterol levels in yeasts. Ergosterol is a key element for the formation of the yeast cell membrane. However, statins often cause DNA damage to yeast cells, facilitating mutation and drug resistance. The aim of the current contribution was to synthesize seven series of compounds as inhibitors of the HMGR enzyme of Candida ssp., and to evaluate their effect on cellular growth, ergosterol synthesis and generation of petite mutants of C. glabrata and C. albicans. Compared to the reference drugs (fluconazole and simvastatin), some HMGR inhibitors caused lower growth and ergosterol synthesis in the yeast species and generated fewer petite mutants. Moreover, heterologous expression was achieved in Pichia pastoris, and compounds 1a, 1b, 6g and 7a inhibited the activity of recombinant CgHMGR and showed better binding energy values than for α-asarone and simvastatin. Thus, we believe these are good candidates for future antifungal drug development. [ABSTRACT FROM AUTHOR]

Researchers have explored natural products to combat the antibiotic resistance of various microorganisms. Cinnamaldehyde (CIN), a major component of cinnamon essential oil (CC-EO), has been found to effectively inhibit the growth of bacteria, fungi, and mildew, as well as their production of toxins. Therefore, this study aimed to create a delivery system for CIN using PLGA microparticles (CIN-MPs), and to compare the antifungal activity of the carried and free CIN, particularly against antibiotic-resistant strains of Candida spp. The first part of the study focused on synthesizing and characterizing the PLGA MPs, which had no toxic effects in vivo and produced results in line with the existing literature. The subsequent experiments analyzed the antifungal effects of MPs-CIN on Candida albicans and Candida glabrata, both resistant (R) and sensitive (S) strains and compared its efficacy with the conventional addition of free CIN to the culture medium. The results indicated that conveyed CIN increased the antifungal effects of the product, particularly towards C. albicans R. The slow and prolonged release of CIN from the PLGA MPs ensured a constant and uniform concentration of the active principle within the cells. [ABSTRACT FROM AUTHOR]

Reboxetine (REB) and sertraline (SER) are antidepressants. The antifungal potential of these drugs against planktonic Candida has been recently reported with limited data about their effects on Candidal biofilms. Biofilms are self-derived extracellular matrixes produced by the microbial population that is attached to biotic surfaces, such as vaginal and oral mucosa, or abiotic surfaces, such as biomedical devices, resulting in persistent fungal infections. The commonly prescribed antifungals, azoles, are usually less effective when biofilms are formed, and most of the prescribed antifungals are only fungistatic. Therefore, the current study investigates the antifungal potentials of REB and SER, alone and in combination with fluconazole (FLC) and itraconazole (ITR) against Candidal biofilms. Using proper controls, Candida species (Candida albicans, C. albicans; Candida krusei, C. krusei; and Candida glabrata, C. glabrata) were used to form biofilms in 96-well microplates. Serial dilutions corresponding to concentrations ranging from 2 to 4096 µg/mL of the target drugs (REB, SER, FLC, ITR) were prepared and added to the plates. Impairment of the biofilm biomass and biofilm metabolic viability was detected using the crystal violet (CV) assay and 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay, respectively. In the checkerboard assay, the sessile fractional inhibitory concentration index (SFICI) was calculated to evaluate the effects of drug combinations. SER was more effective in reducing the biomass than REB for C. albicans and C. glabrata, but both were equal for C. krusei. For the reduction in metabolic activity in C. albicans and C. glabrata, SER had a slight advantage over REB. In C. krusei, REB was slightly more potent. Overall, FLC and ITR were almost equal and produced more significant reductions in metabolic activity when compared to SER and REB, except for C. glabrata, where SER was almost equal to FLC. Synergism was detected between REB + FLC and REB + ITR against biofilm cells of C. albicans. Synergism was detected between REB + ITR against biofilm cells of C. krusei. Synergism was detected between REB + FLC and REB + ITR against biofilm cells of C. albicans, C. krusei, and C. glabrata. The results of the present study support the potential of SER and REB as anti-Candidal biofilm agents that are beneficial as a new antifungal to combat Candidal resistance. [ABSTRACT FROM AUTHOR]

Background and Purpose: Candidiasis is considered as one of the most important fungal infections in hematological malignancy. Candidiasis is considered as the most important cause of disease and mortality in hematological malignancy and antifungal prophylaxis is important (remove highlighted text).Objectives: The aim of study is to identify and evaluated antifungal susceptibility pattern in patients with hematological malignancyMaterial and Methods: In this study, samples were collected from the oral cavity of 100 patients and Candida colonization was confirmed by fungal culture. Isolated Candida strains were identified by ITS-PCR. In vitro antifungal susceptibility tests against fluconazole (replace “fluconazole” with “azoles”), amphotericinB and caspofungin were performed according to CLSI M27-A3/S4. Results: Demographics, comorbidities, distribution of Candida species, antifungal susceptibility were analyzed. Study participants included 100 patients. The mean age of patients was 15.48% ±48.74 years were in the range of 17 to 84 years and in terms of sex distribution, 64% were male and 36% were female. In terms of the distribution of underlying hematologic malignancy, 27% had lymphoma. The most commonly isolated species among patients were 49% C. albicans, 31% C. glabrata, 10% Co-colonization of C. albicans and C. with C. glabrata .The overall resistance of C.albicance was %5 to fluconazole%2 to amphotericinB.C. glabrata showed %11 resistance to fluconazole and was susceptible to amphotericinB.All candida spp.isolated from patients were susceptible to caspofungin.Conclusions: The high rate of colonization of Candida species, especially the significant increase in the frequency of Candida glabrata in patients with blood malignancies and the gradual increase in resistance to fluconazole, necessitate a change in the use of antifungal drugs for the prevention and experimental treatment of hematological malignancy.

Antimicrobial resistance related to the misuse of antibiotics is a well-known current topic. Their excessive use in several fields has led to enormous selective pressure on pathogenic and commensal bacteria, driving the evolution of antimicrobial resistance genes with severe impacts on human health. Among all the possible strategies, a viable one could be the development of medical features that employ essential oils (EOs), complex natural mixtures extracted from different plant organs, rich in organic compounds showing, among others, antiseptic properties. In this work, green extracted essential oil of Thymus vulgaris was included in cyclic oligosaccharides cyclodextrins (CD) and prepared in the form of tablets. This essential oil has been shown to have a strong transversal efficacy both as an antifungal and as an antibacterial agent. Its inclusion allows its effective use because an extension of the exposure time to the active compounds is obtained and, therefore, a more marked efficacy, especially against biofilm-producing microorganisms such as P. aeruginosa and S. aureus, was registered. The efficacy of the tablet against candidiasis opens their possible use as a chewable tablet against oral candidiasis and as a vaginal tablet against vaginal candidiasis. Moreover, the registered wide efficacy is even more positive since the proposed approach can be defined as effective, safe, and green. In fact, the natural mixture of the essential oil is produced by the steam current method; therefore, the manufacturer employs substances that are not harmful, with very low production and management costs. [ABSTRACT FROM AUTHOR]

As an alternative to fossil volatile hydrocarbon solvents used nowadays in perfumery, investigation on essential oil of Commiphora wildii Merxm. oleo gum resin as a source of heptane is reported here. Heptane, representing up to 30 wt-% of this oleo gum resin, was successfully isolated from the C. wildii essential oil, using an innovative double distillation process. Isolated heptane was then used as a solvent in order to extract some noble plants of perfumery. It was found that extracts obtained with this solvent were more promising in terms of sensory analysis than those obtained from fossil-based heptane. In addition, in order to valorize the essential oil depleted from heptane, chemical composition of this oil was found to obtain, and potential biological activity properties were studied. A total of 172 different compounds were identified by GC-MS in the remaining oil. In vitro tests—including hyaluronidase, tyrosinase, antioxidant, elastase and lipoxygenase, as well as inhibitory tests against two yeasts and 21 bacterial strains commonly found on the skin—were carried out. Overall, bioassays results suggest this heptane-depleted essential oil is a promising active ingredient for cosmetic applications. [ABSTRACT FROM AUTHOR]

ABSTRACT: Objectives: We evaluated the azole resistance mechanisms and epidemiology of fluconazole-resistant Candida glabrata from a global survey. Methods: A total of 2992 Candida spp. isolates collected during 2018–2019 were susceptibility tested by the broth microdilution reference method following CLSI guidelines. Fluconazole-resistant C. glabrata isolates were submitted to whole genome sequencing and gene expression assays using qRT-PCR. Results: Among 561 CGLA isolates tested, 34 (6.1%) were fluconazole resistant. These isolates were collected from 11 countries and mainly recovered from bloodstream infections (79.4%). All fluconazole-resistant C. glabrata isolates were non-wild type for voriconazole, 24/34 were non-wild type for posaconazole, but only 2/34 were non-wild type for itraconazole. Isavuconazole MIC values ranged from 0.25 to >4 mg/L. Fluconazole-resistant C. glabrata isolates belonged to 14 different sequence types (ST). None of the isolates exhibited alterations in ERG3 or ERG11, the target of azoles. All but two fluconazole-resistant isolates displayed overexpression of CgCDR1 (22/34; 64.7%) and/or CgCDR2 (26/34; 76.5%), while 16 isolates had both genes overexpressed. Overexpression of CgSNQ2 or ERG11 was not observed. Gain of function (GoF) alterations in the transcription factor CgPDR1 were noted in 14 isolates. Four (11.8%) isolates that were nonsusceptible to one or more echinocandins had FKS2 HS1 alterations (2 S663P and 2 F659Y/deletion). Conclusion: Fluconazole-resistant C. glabrata was driven by overexpression of CgCDR1 and/or CgCDR2. GoF alterations in PDR1 that have been associated with increased virulence were observed. Susceptibility results and surveillance data are needed to guide treatment for these isolates.

Rezafungin, a new echinocandin with an extended half‐life, exhibits potent activity against Candida spp. Aside from the MIC, specific interactions between antifungal and isolate, including the duration of anti‐infective activity, may impact dose interval choices and infection outcome. We evaluated rezafungin and micafungin post‐antifungal effect (PAFE) against C. albicans, C. parapsilosis and C. glabrata. Six Candida spp. isolates were tested, including two of each species, C. albicans, C. parapsilosis and C. glabrata. Antifungal susceptibility testing was performed using the CLSI reference broth microdilution method. Antifungal concentrations of 1x, 4x and 16x the baseline MIC were used for PAFE determinations. Colony counts were performed at T0 (pre‐exposure), after the 1‐h drug exposure, after the cell wash (T1), and at T2, T4, T8, T12, T24 and T48 h. Rezafungin PAFE results were equivalent to micafungin PAFE values for one C. albicans (>14.9 h) and both C. glabrata (>40 h) isolates for all concentrations tested. The rezafungin and micafungin PAFEs could not be determined against one C. albicans isolate. Prolonged PAFE results were also noted for rezafungin (range, 18.4 to >40 h) against both C. parapsilosis isolates at all concentrations, while no micafungin PAFE or a short PAFE (range, 1.8 to 7.4 h) was observed against these organisms, except at 16x bMIC. Rezafungin showed sustained growth inhibition following drug removal and displayed equivalent or longer PAFE values than micafungin against all tested Candida spp. [ABSTRACT FROM AUTHOR]

Infections by non-albicans Candida species have increased drastically in the past few decades. Candida glabrata is one of the most common opportunistic fungal pathogens in immunocompromised individuals, owing to its capability to attach to various human cell types and medical devices and being intrinsically weakly susceptible to azoles. Immunotherapy, including the development of antifungal vaccines, has been recognized as an alternative approach for preventing and treating fungal infections. Secretory proteins play a crucial role in establishing host–pathogen interactions and are also responsible for eliciting an immune response in the host during candidiasis. Therefore, fungal secretomes can provide promising protein candidates for antifungal vaccine development. This study attempts to uncover the presence of immunodominant antigenic proteins in the C. glabrata secretome and delineate their role in various biological processes and their potency in the development of antifungal vaccines. LC–MS/MS results uncovered that C. glabrata secretome consisted of 583 proteins, among which 33 were identified as antigenic proteins. The protection ability of secretory proteins against hematogenously disseminated infection caused by C. glabrata was evaluated in BALB/c mice. After immunization and booster doses, all the animals were challenged with a lethal dose of C. glabrata. All the mice showing signs of distress were sacrificed post-infection, and target organs were collected, followed by histopathology and C. glabrata (CFU/mg) estimation. Our results showed a lower fungal burden in target organs and increased survival in immunized mice compared to the infection control group, thus revealing the immunogenic property of secreted proteins. Thus, identified secretome proteins of C. glabrata have the potential to act as antigenic proteins, which can serve as potential candidates for the development of antifungal vaccines. This study also emphasizes the importance of a mass-spectrometry approach to identifying the antigenic proteins in C. glabrata secretome. [ABSTRACT FROM AUTHOR]

Simple Summary: The large antibiotic consumption in the clinical, veterinary, and agricultural fields has resulted in a tremendous flow of antibiotics into the environment. This has led to enormous selective pressures driving the evolution of antimicrobial resistance in bacteria and yeasts. For this reason, the World Health Organization is promoting research to discover new natural products competitive with synthetic drugs in clinical performances. Compared with conventional drugs, the production of natural pharmaceuticals often has a lower environmental impact and lower economic costs of processes, especially when they originate from agricultural wastes. In the context of a circular economy, we aimed to successfully present preliminary results for the valorization of agricultural waste produced in cabbage cultivation by isolating a highly performant antibacterial and antifungal lipophilic natural mixture from cabbage leaves. As dramatically experienced in the recent world pandemic, viral, bacterial, fungal pathogens constitute very serious concerns in the global context of human health. Regarding this issue, the World Health Organization has promoted research studies that aim to develop new strategies using natural products. Although they are often competitive with synthetic pharmaceuticales in clinical performance, they lack their critical drawbacks, i.e., the environmental impact and the high economic costs of processing. In this paper, the isolation of a highly performant antibacterial and antifungal lipophilic natural mixture from leaves of savoy and white cabbages is proposed as successful preliminary results for the valorization of agricultural waste produced in cabbage cultivation. The fraction was chemically extracted from vegetables with diethyl ether and tested against two Candida species, as well as Pseudomonas aeruginosa, Klebsiella pneumoniae and Staphylococcus aureus reference strains. All the different fractions (active and not active) were chemically characterized by vibrational FT-IR spectroscopy and GC-MS analyses. The extracts showed high growth-inhibition performance on pathogens, thus demonstrating strong application potential. We think that this work, despite being at a preliminary stage, is very promising, both from pharmaceutical and industrial points of view, and can be proposed as a proof of concept for the recovery of agricultural production wastes. [ABSTRACT FROM AUTHOR]

Researchers have explored natural products to combat the antibiotic resistance of various microorganisms. Cinnamaldehyde (CIN), a major component of cinnamon essential oil (CC-EO), has been found to effectively inhibit the growth of bacteria, fungi, and mildew, as well as their production of toxins. Therefore, this study aimed to create a delivery system for CIN using PLGA microparticles (CIN-MPs), and to compare the antifungal activity of the carried and free CIN, particularly against antibiotic-resistant strains of Candida spp. The first part of the study focused on synthesizing and characterizing the PLGA MPs, which had no toxic effects in vivo and produced results in line with the existing literature. The subsequent experiments analyzed the antifungal effects of MPs-CIN on Candida albicans and Candida glabrata, both resistant (R) and sensitive (S) strains and compared its efficacy with the conventional addition of free CIN to the culture medium. The results indicated that conveyed CIN increased the antifungal effects of the product, particularly towards C. albicans R. The slow and prolonged release of CIN from the PLGA MPs ensured a constant and uniform concentration of the active principle within the cells.

Reboxetine (REB) and sertraline (SER) are antidepressants. The antifungal potential of these drugs against planktonic Candida has been recently reported with limited data about their effects on Candidal biofilms. Biofilms are self-derived extracellular matrixes produced by the microbial population that is attached to biotic surfaces, such as vaginal and oral mucosa, or abiotic surfaces, such as biomedical devices, resulting in persistent fungal infections. The commonly prescribed antifungals, azoles, are usually less effective when biofilms are formed, and most of the prescribed antifungals are only fungistatic. Therefore, the current study investigates the antifungal potentials of REB and SER, alone and in combination with fluconazole (FLC) and itraconazole (ITR) against Candidal biofilms. Using proper controls, Candida species (Candida albicans, C. albicans; Candida krusei, C. krusei; and Candida glabrata, C. glabrata) were used to form biofilms in 96-well microplates. Serial dilutions corresponding to concentrations ranging from 2 to 4096 µg/mL of the target drugs (REB, SER, FLC, ITR) were prepared and added to the plates. Impairment of the biofilm biomass and biofilm metabolic viability was detected using the crystal violet (CV) assay and 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay, respectively. In the checkerboard assay, the sessile fractional inhibitory concentration index (SFICI) was calculated to evaluate the effects of drug combinations. SER was more effective in reducing the biomass than REB for C. albicans and C. glabrata, but both were equal for C. krusei. For the reduction in metabolic activity in C. albicans and C. glabrata, SER had a slight advantage over REB. In C. krusei, REB was slightly more potent. Overall, FLC and ITR were almost equal and produced more significant reductions in metabolic activity when compared to SER and REB, except for C. glabrata, where SER was almost equal to FLC. Synergism was detected between REB + FLC and REB + ITR against biofilm cells of C. albicans. Synergism was detected between REB + ITR against biofilm cells of C. krusei. Synergism was detected between REB + FLC and REB + ITR against biofilm cells of C. albicans, C. krusei, and C. glabrata. The results of the present study support the potential of SER and REB as anti-Candidal biofilm agents that are beneficial as a new antifungal to combat Candidal resistance.

Incidence of vulvovaginal candidiasis are strikingly high and treatment options are limited with nearly 50% Candida glabrata cases left untreated or experience treatment failures. The vaginal microenvironment is rich in lactic acid (LA), and the adaptation of C. glabrata to LA is the main reason for clinical treatment failure. In the present study, C. glabrata and its vaginal clinical isolates were comprehensively investigated for their growth response, metabolic adaptation and altered cellular pathway to LA using different biochemical techniques, metabolic profiling and transcriptional studies. Candida glabrata shown considerable variations in its topological and biochemical features without compromizing growth in LA media. Chemical profiling data highlighted involvement of cell wall/membrane, ergosterol and oxidative stress related pathways in mediating adaptative response of C. glabrata towards LA. Furthermore, one dimensional proton (1H) NMR spectroscopy based metabolic profiling revealed significant modulation in 19 metabolites of C. glabrata cells upon growth in LA. Interestingly myo-inositol, xylose, putrescine, and betaine which are key metabolites for cell growth and viability were found to be differentially expressed by clinical isolates. These observations were supported by the transcriptional expression study of selected genes evidencing cell wall/membrane re-organization, altered oxidative stress, and reprogramming of carbon metabolic pathways. Collectively, the study advances our understanding on adaptative response of C. glabrata in vaginal microenvironment to LA for survival and virulence. Lay Summary In vaginal tract, LA present as a natural carbon source is a potentiating factor for vulvovaginal candidiasis caused by C. glabrata is highest. The present article delineates the lactic acid adaptation in vaginal clinical isolates of C. glabrata using a comprehensive approach of biochemical, metabolic, and transcriptional studies. [ABSTRACT FROM AUTHOR]

Background and Purpose: Despite advances in burn care and management, infections are still a major contributor to morbidity and mortality rates in patients with burn injuries. Regarding this, the present study was conducted to investigate the prevalence and importance of candidemia in pediatric burn patients. Materials and Methods: Blood samples were collected from the patients and cultured in an automated blood culture system. Candida species were identified using specific culture media. The relationship between candidemia and possible risk factors was evaluated and compared to a control group. Results: A total of 71 patients with the mean age of 4.52±3.63 years were included in the study. Blood cultures showed candidemia in 19 (27%) patients. Based on the results, C. albicans was the most common fungus among patients with and without candidemia. The results of statistical analysis also showed that candidemia was significantly correlated with total body surface area (TBSA), mechanical ventilation, duration of total parenteral nutrition, length of intensive care unit (ICU) stay, presence of neutropenia, and R-Baux score (all P≤0.001). In this regard, TBSA, length of ICU stay, R-Baux score, and Candida score were identified as the determinant factors for mortality due to candidemia. Conclusion: Candidemia increases the mortality and morbidity rates associated with burn injuries. Prompt diagnostic and prevention measures can reduce the unfortunate outcomes via controlling the possible risk factors.

Antimicrobial resistance related to the misuse of antibiotics is a well-known current topic. Their excessive use in several fields has led to enormous selective pressure on pathogenic and commensal bacteria, driving the evolution of antimicrobial resistance genes with severe impacts on human health. Among all the possible strategies, a viable one could be the development of medical features that employ essential oils (EOs), complex natural mixtures extracted from different plant organs, rich in organic compounds showing, among others, antiseptic properties. In this work, green extracted essential oil of Thymus vulgaris was included in cyclic oligosaccharides cyclodextrins (CD) and prepared in the form of tablets. This essential oil has been shown to have a strong transversal efficacy both as an antifungal and as an antibacterial agent. Its inclusion allows its effective use because an extension of the exposure time to the active compounds is obtained and, therefore, a more marked efficacy, especially against biofilm-producing microorganisms such as P. aeruginosa and S. aureus, was registered. The efficacy of the tablet against candidiasis opens their possible use as a chewable tablet against oral candidiasis and as a vaginal tablet against vaginal candidiasis. Moreover, the registered wide efficacy is even more positive since the proposed approach can be defined as effective, safe, and green. In fact, the natural mixture of the essential oil is produced by the steam current method; therefore, the manufacturer employs substances that are not harmful, with very low production and management costs.

As an alternative to fossil volatile hydrocarbon solvents used nowadays in perfumery, investigation on essential oil of Commiphora wildii Merxm. oleo gum resin as a source of heptane is reported here. Heptane, representing up to 30 wt-% of this oleo gum resin, was successfully isolated from the C. wildii essential oil, using an innovative double distillation process. Isolated heptane was then used as a solvent in order to extract some noble plants of perfumery. It was found that extracts obtained with this solvent were more promising in terms of sensory analysis than those obtained from fossil-based heptane. In addition, in order to valorize the essential oil depleted from heptane, chemical composition of this oil was found to obtain, and potential biological activity properties were studied. A total of 172 different compounds were identified by GC-MS in the remaining oil. In vitro tests—including hyaluronidase, tyrosinase, antioxidant, elastase and lipoxygenase, as well as inhibitory tests against two yeasts and 21 bacterial strains commonly found on the skin—were carried out. Overall, bioassays results suggest this heptane-depleted essential oil is a promising active ingredient for cosmetic applications.

Background:Candida species have long been recognised as aetiological agents of opportunistic infections of the oral mucosa, and more recently, as players of polymicrobial interactions driving caries, periodontitis and oral carcinogenesis. Methods: We studied the clonal structure of Candida spp. at oral niche resolution in patients (n = 20) with a range of oral health profiles over 22 months. Colonies from oral micro-environments were examined with multilocus sequencing typing. Results:Candida spp. identified were C. albicans, C. glabrata and C. dubliniensis. Increased propensity for micro-variations giving rise to multiple diploid strain types (DST), as a result of loss of heterozygosity, was observed among C. albicans clade 1 isolates compared to other clades. Micro-variations among isolates were also observed in C. dubliniensis contra to expectations of stable population structures for this species. Multiple sequence types were retrieved from patients without clinical evidence of oral candidosis, while single sequence types were isolated from oral candidosis patients. Conclusion: This is the first study to describe the clonal population structure, persistence and stability of Candida spp. at oral niche level. Future research investigating links between Candida spp. clonality and oral disease should recognise the propensity to micro-variations amongst oral niches in C. albicans and C. dubliniensis identified here. [ABSTRACT FROM AUTHOR]

Invasive candidiasis is a common, severe and often difficult to treat complication in various categories of patients. Despite the fact that in most cases Candida albicans retains a leading position as an etiological cause, a pathogen such as Candida glabrata is becoming an increasingly relevant species, especially in oncohematology patients who have previously received therapy with azole antimycotics. For the treatment of systemic infections caused by C. glabrata, according to current versions of practical guidelines, echinocandins are the drugs of choice. Possessing high fungicidal activity, unlike azoles, they allow in most cases to achieve the clinical efficacy of therapy when this pathogen is isolated. However, there is an evidence of the selection of C. glabrata strains with reduced susceptibility or even resistance to echinocandins. This article briefly presents data on the problem of resistance of C. glabrata to drugs of this class, as well as the results of Russian in vitro studies.

Infections by non-albicans Candida species have increased drastically in the past few decades. Candida glabrata is one of the most common opportunistic fungal pathogens in immunocompromised individuals, owing to its capability to attach to various human cell types and medical devices and being intrinsically weakly susceptible to azoles. Immunotherapy, including the development of antifungal vaccines, has been recognized as an alternative approach for preventing and treating fungal infections. Secretory proteins play a crucial role in establishing host–pathogen interactions and are also responsible for eliciting an immune response in the host during candidiasis. Therefore, fungal secretomes can provide promising protein candidates for antifungal vaccine development. This study attempts to uncover the presence of immunodominant antigenic proteins in the C. glabrata secretome and delineate their role in various biological processes and their potency in the development of antifungal vaccines. LC–MS/MS results uncovered that C. glabrata secretome consisted of 583 proteins, among which 33 were identified as antigenic proteins. The protection ability of secretory proteins against hematogenously disseminated infection caused by C. glabrata was evaluated in BALB/c mice. After immunization and booster doses, all the animals were challenged with a lethal dose of C. glabrata. All the mice showing signs of distress were sacrificed post-infection, and target organs were collected, followed by histopathology and C. glabrata (CFU/mg) estimation. Our results showed a lower fungal burden in target organs and increased survival in immunized mice compared to the infection control group, thus revealing the immunogenic property of secreted proteins. Thus, identified secretome proteins of C. glabrata have the potential to act as antigenic proteins, which can serve as potential candidates for the development of antifungal vaccines. This study also emphasizes the importance of a mass-spectrometry approach to identifying the antigenic proteins in C. glabrata secretome.

Infections with Candida glabrata have recently gained worldwide attention owing to its association with long hospitalizations and high mortality rates. This problem is highlighted when the infection is associated with echinocandin resistance, which is used for first-line therapy. Echinocandin resistance is exclusively attributed to functional mutations in FKS genes, and especially in hot spot (HS) regions. Unfortunately, few studies have focused on the rapid identification of FKS mutations associated with echinocandin resistance in C. glabrata. This study was intended to evaluate and validate the use of Surveyor nuclease assay (SN) for detection of FKS gene mutations. SN was evaluated against three segments of FKS1 and FKS2 genes including whole gene, regions including all HSs, and the region including only HS1. Our results showed that SN results are basically dependent on the type of gene as well as the segment type. Interestingly, SN can detect mutations in the region containing HS1 in both FKS1 and FKS2 genes. Furthermore, SN can detect mutations in the segment containing all HS regions for FKS1 but not FKS2. SN was unable to detect mutations in the whole FKS1 and FKS2 genes. As far as we know, this is the first study to validate SN for rapid identification of FKS gene mutations. This assay could be used as a sample for rapid identification of mutations associated with HS1 region in FKS genes, which have a predominant role for echinocandin resistance induction in C. glabrata. [ABSTRACT FROM AUTHOR]

Background: Candida species have long been recognised as aetiological agents of opportunistic infections of the oral mucosa, and more recently, as players of polymicrobial interactions driving caries, periodontitis and oral carcinogenesis. Methods: We studied the clonal structure of Candida spp. at oral niche resolution in patients (n = 20) with a range of oral health profiles over 22 months. Colonies from oral micro-environments were examined with multilocus sequencing typing. Results: Candida spp. identified were C. albicans, C. glabrata and C. dubliniensis. Increased propensity for micro-variations giving rise to multiple diploid strain types (DST), as a result of loss of heterozygosity, was observed among C. albicans clade 1 isolates compared to other clades. Micro-variations among isolates were also observed in C. dubliniensis contra to expectations of stable population structures for this species. Multiple sequence types were retrieved from patients without clinical evidence of oral candidosis, while single sequence types were isolated from oral candidosis patients. Conclusion: This is the first study to describe the clonal population structure, persistence and stability of Candida spp. at oral niche level. Future research investigating links between Candida spp. clonality and oral disease should recognise the propensity to micro-variations amongst oral niches in C. albicans and C. dubliniensis identified here.

This study was designed to analyze the interaction of 21 antifungal combinations consisting of seven major antifungal agents against 11 echinocandin- susceptible and six-resistant C. glabrataisolates. The combinations were divided into five major groups and were evaluated by checkerboard,disc diffusion, and time-killing assays. Synergy based on the fractional inhibitory concentration indexof≤0.50 was observed in 17.65–29.41% of the cases for caspofungin combinations with azoles oramphotericin B. Amphotericin B combination with azoles induced synergistic interaction in a rangeof 11.76–29.41%. Azole combinations and 5-flucytosine combinations with azoles or amphotericin Bdid not show synergistic interactions. None of the 21 combinations showed antagonistic interactions.Interestingly, 90% of the detected synergism was among the echinocandin-resistant isolates. Disk diffusion assays showed that the inhibition zones produced by antifungal combinations were equal toor greater than those produced by single drugs. The time-killing assay showed the synergistic actionof caspofungin combination with fluconazole, voriconazole, and posaconazole, and the amphotericinB-5-flucytosine combination. Furthermore, for the first time, this assay confirmed the fungicidal activity of caspofungin-voriconazole and amphotericin B-5-flucytosine combinations. The combination interactions ranged from synergism to indifference and, most importantly, no antagonism was reported and most of the synergistic action was among echinocandin-resistant isolates. [ABSTRACT FROM AUTHOR]

Purpose: Chronic infections of Candida albicans are characterised by the embedding of budding and entwined filamentous fungal cells into biofilms. The biofilms are refractory to many drugs and Candida biofilms are associated with ocular fungal infections. The objective was to test the activity of nanoparticulate amphotericin B (AmB) against Candida biofilms. Methods: AmB was encapsulated in the Molecular Envelope Technology (MET, N-palmitoyl-N-monomethyl-N,N-dimethyl-N,N,N-trimethyl-6-O-glycolchitosan) nanoparticles and tested against Candida biofilms in vitro. Confocal laser scanning microscopy (CLSM) imaging of MET nanoparticles’ penetration into experimental biofilms was carried out and a MET-AmB eye drop formulation was tested for its stability. Results: MET-AmB formulations demonstrated superior activity towards C. albicans biofilms in vitro with the EC50 being ~30 times lower than AmB alone (EC50 MET-AmB = 1.176 μg mL−1, EC50 AmB alone = 29.09 μg mL−1). A similar superior activity was found for Candida glabrata biofilms, where the EC50 was ~10× lower than AmB alone (EC50 MET-AmB = 0.0253 μg mL−1, EC50 AmB alone = 0.289 μg mL−1). CLSM imaging revealed that MET nanoparticles penetrated through the C. albicans biofilm matrix and bound to fungal cells. The activity of MET-AmB was no different from the activity of AmB alone against C. albicans cells in suspension (MET-AmB MIC90 = 0.125 μg mL−1, AmB alone MIC90 = 0.250 μg mL−1). MET-AmB eye drops were stable at room temperature for at least 28 days. Conclusions: These biofilm activity findings raise the possibility that MET-loaded nanoparticles may be used to tackle Candida biofilm infections, such as refractory ocular fungal infections.

In the budding yeast Saccharomyces cerevisiae, Svl3 and Pam1 proteins work as functional homologues. Loss of their function causes increased levels of chitin deposition in the cell wall and temperature sensitivity, suggesting their involvement in cell wall formation. We found that the N‐ and C‐termini of these proteins have distinctive and critical functions. They contain an N‐terminal part that has a probable 2‐dehydropantoate 2‐reductase domain. In Svl3, this part can be replaced with the yeast 2‐dehydropantoate 2‐reductase, Pan5, suggesting that Svl3 and its homologues may be able to mediate 2‐dehydropantoate 2‐reductase function. On the other hand, Svl3 is recruited to the bud tip and bud neck via multiple localization signals in the C‐terminal part. One of such signals is the lysine‐rich region located in the C‐terminal end. The function and localization of Svl3 are significantly disrupted by the loss of this lysine‐rich region; however, its localization is not completely abolished by the mutation because another localization signal enables appropriate transport. Svl3 and Pam1 orthologues are found in cells across fungal species. The Svl3 orthologues of Candida glabrata can complement the loss of Svl3 and Pam1 in S. cerevisiae. C. glabrata cells lacking the SVL3 and PAM1 orthologue genes exhibit phenotypes similar to those observed in svl3∆pam1∆ S. cerevisiae cells. Thus, Svl3 homologues may be generally required for the assembly of the cell wall in fungal cells. [ABSTRACT FROM AUTHOR]

β-citronellol, an acyclic monoterpenoid, inhibits dose-dependent growth of C. glabrata and C. tropicalis with MIC/MFC values of 400 µg/ml/600 µg/ml and 100 µg/ml/200 µg/ml, respectively. Below MIC, the compound was more toxic to C. glabrata but the inhibition of extracellular proteinases was higher in C. tropicalis. Significant phospholipase activity was not observed in both cases. Membrane ergosterol content decreased at concentrations above MIC/4; inhibition being greater in C. tropicalis. Interestingly at MIC, β-citronellol showed significantly better ergosterol inhibition in comparison to FLC. At MIC/2, β-citronellol destabilized fungal cell membrane and cell wall in the presence of cell wall perturbing agents (caspofungin and calcoflour white). Of the two species, C. tropicalis was affected more by DTT and salt-induced stress in the presence of β-citronellol, severely affecting fungal endoplasmic reticulum and cell surface integrity. β-citronellol thus alters metabolic activities of both Candida species decreasing growth and viability projecting it as a potential antifungal. [ABSTRACT FROM AUTHOR]

Candida glabrata causes difficult to treat invasive candidiasis due to its antifungal resistance, mainly to azoles. The aim of the present work was to study the role of the genes ERG11 , CDR1 , CDR2, and SNQ2 on the resistance to voriconazole (VRC) in a set of C. glabrata strains with known in vitro and in vivo susceptibility to this drug. Eighteen clinical isolates of C. glabrata were exposed in vitro to VRC, and the expression of the cited genes was quantified by real time quantitative polymerase chain reaction (q-PCR). In addition, the ERG11 gene was amplified and sequenced to detect possible mutations. Ten synonymous mutations were found in 15 strains, two of them being reported for the first time; however, no amino acid changes were detected. ERG11 and CDR1 were the most expressed genes in all the strains tested, while the expression of CDR2 and SNQ2 was modest. Our results show that gene expression does not directly correlate with the VRC MIC. In addition, the expression profiles of ERG11 and efflux pump genes did not change consistently after exposure to VRC. Although individual analysis did not result in a clear correlation between MIC and gene expression, we did observe an increase in ERG11 and CDR1 expression in resistant strains. It is of interest that considering both in vitro and in vivo results, the slight increase in such gene expression correlates with the observed resistance to VRC. [ABSTRACT FROM AUTHOR]

Vulvovaginal candidiasis (VVC) alongside with antifungal resistance are becoming a major clinical problem in recent years. A prospective study aimed to evaluate the diversity of yeast strains associated with VVC in Tebessa city (northeastern Algeria) and investigate their susceptibility patterns. Over two months, yeasts were isolated on chromogenic medium from twenty-nine non-pregnant women with symptomatic VVC. The isolates were characterized with MALDI-TOF MS and antifungal susceptibility testing was performed for nine antifungal drugs using SensititreTM YeastOneTM YO10. Twenty-nine non-duplicate yeasts were recovered and the mass spectrometry profiles showed reliable scores of which four genera and five different species were identified. Candida albicans accounted for 65.5 % (n = 19) of the total number of isolates, followed by C. glabrata with 20.7% (n = 6). For the remaining non-albicans Candida (NCA) species, Kluyveromyces marxianus with 6.9% (n = 2), Pichia kudriavzevii and Saccharomyces cerevisiae with one isolate each. The antifungal susceptibilities showed wild type MICs of C. albicans to amphotericin B, azoles and echinocandins. In addition, four C. albicans isolates were resistant to flucytosine. For C. glabrata isolates, 100% non-WT phenotype was found for both posaconazole and itraconazole. For the very first time, the obtained outcomes bring out new data concerning the epidemiology of yeasts causing VVC in Algeria and their antimicrobial susceptibility profiles.

Candida glabrata is a genetically diverse human pathogenic yeast, whose subpopulations have been documented to vary geographically. Here, we report MLST genotypes and antifungal drug susceptibility of C. glabrata isolates from Africa. Among 47 mostly urogenital isolates, we found 13 sequence types, amounting to a 27% genetic population difference. More than half of the isolates were of novel sequence types. ST18 was most predominant and had reduced susceptibility to fluconazole. There was clear segregation of STs between urine and vaginal specimen. In Tanzania, the C. glabrata population is genetically diverse, and divergent from those seen in other countries. [ABSTRACT FROM AUTHOR]

Highlights • The recombinant enzyme CgHMGR was used as a model to test new compounds with possible antifungal activity. • Compounds 3 and 4 inhibited CgHMGR enzyme activity, with an IC 50 lower than that for α-asarone. • Molecular docking studies revealed that compounds 3 and 4 bind to CgHMGR active site. • Effect caused by simvastatin and α-asarone-based compounds were reversed with the addition of sterols. Abstract Due to increasing resistance of Candida species to antifungal drugs, especially azoles, new drugs are needed. The proposed compounds 3 and 4 are analogous to α-asarone (2) , a naturally occurring potent inhibitor of HMGR with hypolipidemic and antifungal activity. We used the recombinant enzyme 3-hydroxy-3-methylglutaryl coenzyme A reductase of Candida glabrata (CgHMGR) as a model to test the effectiveness of the test compounds. Compounds 3 and 4 demonstrated inhibitory kinetics, having lower IC 50 values (42.65 μM and 28.77 μM, respectively) than compound 2 (>100 μM). The docking studies showed better binding energies for compounds 3 and 4 (−5.35 and −6.1 kcal/mol, respectively) than for compound 2 (−4.53 kcal/mol). These findings suggest that the tested compounds are better than their natural analogue. Plaque assays were performed on the C. glabrata strain CBS138 by applying ergosterol or cholesterol to evaluate the possible reversal of the inhibition induced by compounds 2 , 3 and 4. Inhibition was easily suppressed in all three cases, recovering the viability of C. glabrata. These results reveal that the CgHMGR model is excellent for testing antifungals. Compound 4 produced the best effect and is herein proposed as a new potent antifungal agent. [ABSTRACT FROM AUTHOR]

Candida species are opportunistic fungal pathogens that cause superficial or invasive infections. Recently, the incidence of infection by non-Candida albicans species, especially Candida glabrata, has increased. In this study, we analyzed the adhesion and cytotoxicity of various Candida spp. that are part of the normal human microbiota. C. albicans adheres well to cell culture plates and to cultured cells. C. glabrata selectively adheres to epithelial cells rather than to cell culture plates. Candida parapsilosis insufficiently adheres to confluent monolayers of human lung epithelial A549 and keratinocyte HaCaT cells. We then analyzed the cytotoxicity of C. albicans and C. glabrata, which adhered well to epithelial cells. C. glabrata has been found to cause more damage to A549 cells than to HaCaT cells, suggesting that resident Candida spp. have distinct cytotoxic effects in different tissues. It is important to clarify the properties of Candida spp. as there is evidence that normal microbiota can cause infections. Our data suggest that it is necessary to use appropriate cell lines for characterizing the adherence and cytotoxicity of Candida spp. [ABSTRACT FROM AUTHOR]

This study was designed to analyze the interaction of 21 antifungal combinations consisting of seven major antifungal agents against 11 echinocandin- susceptible and six-resistant C. glabrata isolates. The combinations were divided into five major groups and were evaluated by checkerboard, disc diffusion, and time-killing assays. Synergy based on the fractional inhibitory concentration index of ≤0.50 was observed in 17.65–29.41% of the cases for caspofungin combinations with azoles or amphotericin B. Amphotericin B combination with azoles induced synergistic interaction in a range of 11.76–29.41%. Azole combinations and 5-flucytosine combinations with azoles or amphotericin B did not show synergistic interactions. None of the 21 combinations showed antagonistic interactions. Interestingly, 90% of the detected synergism was among the echinocandin-resistant isolates. Disk diffusion assays showed that the inhibition zones produced by antifungal combinations were equal to or greater than those produced by single drugs. The time-killing assay showed the synergistic action of caspofungin combination with fluconazole, voriconazole, and posaconazole, and the amphotericin B-5-flucytosine combination. Furthermore, for the first time, this assay confirmed the fungicidal activity of caspofungin-voriconazole and amphotericin B-5-flucytosine combinations. The combination interactions ranged from synergism to indifference and, most importantly, no antagonism was reported and most of the synergistic action was among echinocandin-resistant isolates.

Background: Increasing numbers of immunocompromised patients have resulted in greater incidence of invasive fungal infections with high mortality. Candida albicans infections dominate, but during the last decade, Candida glabrata has become the second highest cause of candidemia in the United States and Northern Europe. Reliable and early diagnosis, together with appropriate choice of antifungal treatment, is needed to combat these challenging infections. Objectives: To confirm the identity of 183 Candida glabrata isolates from different human body sites using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) and VITEK®2, and to analyze isolate protein profiles and antifungal susceptibility. The minimum inhibitory concentration (MIC) of seven antifungal drugs was determined for the isolates to elucidate susceptibility. Design: A total of 183 C. glabrata isolates obtained between 2002 and 2012 from Norwegian health-care units were analyzed. For species verification and differentiation, biochemical characterization (VITEK®2) and mass spectrometry (MALDI–TOF) were used. MIC determination for seven antifungal drugs was undertaken using E-tests®. Results: Using VITEK®2, 92.9% of isolates were identified as C. glabrata, while all isolates (100%) were identified as C. glabrata using MALDI-TOF. Variation in protein spectra occurred for all identified C. glabrata isolates. The majority of isolates had low MICs to amphotericin B (≤1 mg/L for 99.5%) and anidulafungin (≤0.06 mg/L for 98.9%). For fluconazole, 18% of isolates had MICs >32 mg/L and 82% had MICs in the range ≥0.016 mg/L to ≤32 mg/L. Conclusions: Protein profiles and antifungal susceptibility characteristics of the C. glabrata isolates were diverse. Clustering of protein profiles indicated that many azole resistant isolates were closely related. In most cases, isolates had highest susceptibility to amphotericin B and anidulafungin. The results confirmed previous observations of high MICs to fluconazole and flucytosine. MALDI-TOF was more definitive than VITEK®2 for C. glabrata identification.

Background and objectives: Oral moisturizers have been used to treat dry mouth. This study aimed to investigate the effects of storage temperature and pH on the antifungal effects of oral moisturizers against Candida albicans and Candida glabrata. Materials and Methods: Thirty-one oral moisturizers and amphotericin B (AMPH-B) were stored at 25 and 37 °C for 1 week. Subsequently, they were added to cylindrical holes in 50% trypticase soy agar plates inoculated with C. albicans and C. glabrata (107 cells/ml). The antifungal effects were evaluated based on the sizes of the growth-inhibitory zones formed. Two-way analysis of variance was used to determine the effects of storage temperature and pH on the growth-inhibitory zones. Results: Significant differences in the effects of storage temperature and pH of the moisturizers were observed against C. albicans and C. glabrata. The growth-inhibitory zones of samples stored at 37 °C and with neutral pH were significantly larger than those stored at 25 °C and with acidic pH, respectively. The sizes of the zones formed by most of the oral moisturizers were larger than those formed by AMPH-B (concentration, 0.63 µg/ml). Conclusion: The antifungal effects of oral moisturizers against C. albicans and C. glabrata were affected by their storage temperature and pH.

Abstract In recent years, C. albicans and C. glabrata have been identified as the main cause of candidemia and invasive candidiasis in hospitalized and immunocompromised patients. In order to colonize the human host, these fungi express several virulence factors such as the response to oxidative stress and the formation of biofilms. In the expression of these virulence factors, the cell wall of C. albicans and C. glabrata is of fundamental importance. As the outermost structure of the yeast, the cell wall is the first to come in contact with the reactive oxygen species (ROS) generated during the respiratory outbreak, and in the formation of biofilms, it is the first to adhere to organs or medical devices implanted in the human host. In both processes, several cell wall proteins (CWP) are required, since they promote attachment to human cells or abiotic surfaces, as well as to detoxify ROS. In our working group we have identified moonlighting CWP in response to oxidative stress as well as in the formation of biofilms. Having identified moonlighting CWP in Candida species in response to two virulence factors indicates that these proteins may possibly be immunodominant. The aim of the present work was to evaluate whether proteins of this type such as fructose-bisphosphate aldolase (Fba1), phosphoglycerate kinase (Pgk) and pyruvate kinase (Pk), could confer protection in a mouse model against C. albicans and C. glabrata. For this, recombinant proteins His 6 -Fba1, His 6 -Pgk and His 6 -Pk were constructed and used to immunize several groups of mice. The immunized mice were infected with C. albicans or C. glabrata , and subsequently the liver, spleen and kidney were extracted and the number of CFU was determined. Our results showed that Pk confers immunity to mice against C. albicans , while Fba1 to C. glabrata. This data allows us to conclude that the moonlighting CWP, Fba1 and Pk confer in vivo protection in a specific way against each species of Candida. This makes them promising candidates for developing specific vaccines against these pathogens. Highlights • C. albicans and C. glabrata in response to oxidative stress (OSR) and the formation of biofilms express several cell wall proteins (CWP). • C. albicans and C. glabrata change the expression of CWP in OSR and biofilm formation. • C. albicans and C. glabrata differentially regulates the expression of moonlighting CWP, such as Fba1, Pgk and Pk. • Fba1, Pgk and Pk could confer protection in a mouse model against C. albicans and C. glabrata. • Our results showed that Pk confers immunity to mice against C. albicans , while Fba1 to C. glabrata. [ABSTRACT FROM AUTHOR]

Although combinatorial regulation is a common feature in gene regulatory networks, how it evolves and affects network structure and function is not well understood. In S. cerevisiae, the phosphate starvation (PHO) responsive transcription factors Pho4 and Pho2 are required for gene induction and survival during phosphate starvation. In the related human commensal C. glabrata, Pho4 is required but Pho2 is dispensable for survival in phosphate starvation and is only partially required for inducing PHO genes. Phylogenetic survey suggests that reduced dependence on Pho2 evolved in C. glabrata and closely related species. In S. cerevisiae, less Pho2-dependent Pho4 orthologs induce more genes. In C. glabrata, its Pho4 binds to more locations and induces three times as many genes as Pho4 in S. cerevisiae does. Our work shows how evolution of combinatorial regulation allows for rapid expansion of a gene regulatory network’s targets, possibly extending its physiological functions.

The aim of the study was to assess the antifungal activity of preparation „fin Candimis” (oregano essential oil) against yeast-like strains belonging to the genus Candida. During the investigation, there were used up nine Candida albicans strains and ten C. glabrata strains isolated from different clinical material, along with one C. albicans demonstration strain ATCC 90028. The oregano essential oil, utilized in the study, was obtained from fresh leaves of Origanum vulgare L. and bore a trade name „fin Candimis”. According to data yielded by its manufacturer, concentration of pure oregano essential oil in preparation „fin Candimis” totals up to 210 mg/ml. The susceptibility of the Candida strains to preparation „fin Candimis” was assessed by means of the disc-diffusion method, upon the Sabouraud solid medium (after a 24-hour incubation of the cultures at temperature of 37 degrees centigrade); the oregano essential oil had been diluted in 1 ml of DMSO, according to the geometrical progression. A measure of the antifungal activity of preparation „fin Candimis” was the minimal inhibitory concentration (MIC), in terms of the fungus growth. Preparation „fin Candimis” is capable of being applied in the prevention and treatment of candidiasis – alone, or as a natural adjunctive agent. The C. albicans strains are more susceptible to preparation „fin Candimis” in comparison to the C. glabrata ones.