Yeast Form Research Articles

Histoplasmosis associated hemophagocytosis- a rare cause of pancytopenia in an immunocompetent patient

Pancytopenia as a result of hemophagocytosis in a middle-aged man presenting with high-grade fever of unknown etiology could be a rare consequence of Histoplasmosis infection. In the past, many cases of patients with human immunodeficiency virus infection contracting histoplasmosis-induced hemophagocytosis have been well documented, however, little data has been entered in the setting of HIV non-reactive status. Interestingly, we report a case of histoplasmosis in an immunocompetent male with the presence of yeast forms of H. capsulatum in the bone marrow smears. The patient was subsequently placed on Amphotericin B and corticosteroids but lost to the infectious disease. Cases like this, highlight the importance of taking into account the early decision for bone marrow sampling with careful attention to microscopic details as incidental pathologies may be elicited which could have significant effects on patient’s prognosis.

Auranofin is active against Histoplasma capsulatum and reduces the expression of virulence-related genes.

Auranofin is an approved anti-rheumatic drug that has a broad-range inhibitory action against several microorganisms, including human pathogenic fungi. The auranofin activity against Histoplasma capsulatum, the dimorphic fungus that causes histoplasmosis, has not been properly addressed. Since there are few therapeutic options for this life-threatening systemic mycosis, this study evaluated the effects of auranofin on H. capsulatum growth and expression of virulence factors. Minimal inhibitory and fungicidal concentrations (MIC and MFC, respectively) of auranofin against 15 H. capsulatum strains with distinct genetic backgrounds were determined using the yeast form of the fungus and a microdilution protocol. Auranofin activity was also assessed on a macrophage model of infection and on a Tenebrio molitor invertebrate animal model. Expression of virulence-related genes was compared between auranofin treated and untreated H. capsulatum yeast cells using a quantitative PCR assay. Auranofin affected the growth of different strains of H. capsulatum, with MIC and MFC values ranging from 1.25 to 5.0 μM and from 2.5 to >10 μM, respectively. Auranofin was able to kill intracellular H. capsulatum yeast cells and conferred protection against the fungus in the experimental animal model of infection. Moreover, the expression of catalase A, HSP70, superoxide dismutase, thioredoxin reductase, serine proteinase, cytochrome C peroxidase, histone 2B, formamidase, metallopeptidase, Y20 and YPS3 proteins were reduced after six hours of auranofin treatment. CONCLUSIONS/SIGNIFICANCE: Auranofin is fungicidal against H. capsulatum and reduces the expression of several virulence-related genes, which makes this anti-rheumatic drug a good candidate for new medicines against histoplasmosis.

Antifungal Drug Resistance in Candida albicans: Identifying Novel Targets for the Development of Effective Antifungal Agents.

Candidiasis, a fungal infection initiated primarily through Candida albicans, affects various portions of the human body and is particularly prevalent in immunocompromised individuals. The pathogenicity of C. albicans is facilitated by numerous virulence causes, including adhesins, morphogenesis, and phenotypic switching. The organism’s capability in the direction of switching between yeast and hyphal forms contributes to the severity of infections. The appearance of resistant strains has rendered current treatments less effective, necessitating the exploration of new drug targets and the progress of novel antifungal agents. Antifungal drug resistance is a multifaceted phenomenon involving genetic mutations, overexpression of efflux pumps, epigenetic changes, and biofilm formation, all regulated by complex genetic and transcriptional networks. These resistance mechanisms can cause treatment failures, highlighting the need for new antifungal agents and improved diagnostic tools. The identification of potential drug targets in C. albicans is critical due to increasing resistance to existing antifungal agents. Recent studies have identified promising targets, for example the riboflavin metabolic pathway and unique protein kinases involved in regulating virulence and pathogenicity. Developing new antifungals is difficult due to C. albicans’ eukaryotic nature and resistance. Ongoing research is essential to find novel targets and strategies, especially with the limited antifungal drug classes available.

Ume6-dependent pathways of morphogenesis and biofilm formation in Candida auris

ABSTRACT Candida auris is a yeast pathogen causing nosocomial outbreaks of candidemia. Its ability to adhere to inert surfaces and to be transmitted from one patient to another via medical devices is of particular concern. Like other Candida spp., C. auris has the ability to transition from the yeast form to pseudohyphae and to build biofilms. Moreover, some isolates have a unique capacity to form aggregates. These morphogenetic changes may impact virulence. In this study, we demonstrated the role of the transcription factor Ume6 in C. auris morphogenesis. Genetic hyperactivation of Ume6 induced filamentation and aggregation. The Ume6-hyperactivated strain ( UME6 HA ) also exhibited increased adhesion to inert surface and formed biofilms of higher biomass compared to the parental strain. Transcriptomic analyses of UME6 HA revealed enrichment of genes encoding for adhesins, proteins involved in cell wall organization, sterol biosynthesis, and aspartic protease activities. The three most upregulated genes compared to wild-type were those encoding for the agglutin-like sequence adhesin Als4498, the C. auris -specific adhesin Scf1, and the hypha-specific G1 cyclin-related protein Hgc1. The deletion of these genes in the UME6 HA background showed that Ume6 controls filamentation via Hgc1 and aggregation via Als4498 and Scf1. Adhesion to inert surface was essentially triggered by Scf1. However, Als4498 and Hgc1 were also crucial for biofilm formation. Our data show that Ume6 is a universal regulator of C. auris morphogenesis via distinct modulators. IMPORTANCE C. auris represents a public health threat because of its ability to cause difficult-to-treat infections and hospital outbreaks. The morphogenetic plasticity of C. auris , including its ability to filament, to form aggregates or biofilms on inert surfaces, is important to the fungus for interhuman transmission, skin or catheter colonization, tissue invasion, antifungal resistance, and escape of the host immune system. This work deciphered the importance of Ume6 in the control of distinct pathways involved in filamentation, aggregation, adhesion, and biofilm formation of C. auris . A better understanding of the mechanisms of C. auris morphogenesis may help identify novel antifungal targets.

Biochemical Characterization of Recombinant Enterococcus faecalis EntV Peptide to Elucidate Its Antihyphal and Antifungal Mechanisms against Candida albicans.

Candida albicans is a common opportunistic fungus in humans, whose morphological switch between yeast and hyphae forms represents a key virulence trait. Developing strategies to inhibit C. albicans hyphal growth may provide insights into designs of novel antivirulent therapeutics. Importantly, the gut commensal bacterium, Enterococcus faecalis, secretes a bacteriocin EntV which has potent antivirulent and antifungal effects against C. albicans in infection models; however, hampered by the challenges to access large quantities of bioactive EntV, the detailed understanding of its mechanisms on C. albicans has remained elusive. In this work, we biochemically reconstituted the proteolytic cleavage reaction to obtain recombinant EntV88-His6 on a large preparative scale, providing facile access to the C-terminal EntV construct. Under in vitro C. albicans hyphal assay with specific inducers, we demonstrated that EntV88-His6 exhibits potent bioactivity against GlcNAc-triggered hyphal growth. Moreover, with fluorescent FITC-EntV88-His6, we revealed that EntV88-His6 enters C. albicans via endocytosis and perturbs the proper localization of the polarisome scaffolding Spa2 protein. Our findings provide important clues on EntV's mechanism of action. Surprisingly, we showed that EntV88-His6 does not affect C. albicans yeast cell growth but potently exerts cytotoxicity against C. albicans under hyphal-inducing conditions in vitro. The combination of EntV88-His6 and GlcNAc displays rapid killing of C. albicans, rendering it a promising antivirulent and antifungal agent.

The yeast Dothiora sorbi IOJ-3 naturally produced various filamentous sectors with distinct abilities by undergoing DNA demethylation

Some fungi have demonstrated the ability to adapt rapidly to changing environments by exhibiting morphological plasticity, a trait influenced by species and environmental factors. Here, an anamorphic yeast strain IOJ-3 exhibited unique sectorization characteristics, naturally producing diverse filamentous sectors when cultivated on potato dextrose agar (PDA) medium or natural culture medium for durations exceeding 13 days. The strain IOJ-3 and its filamentous sectors were identified as Dothiora sorbi. The morphology of the sectors was consistent and heritable. The life cycle of strain IOJ-3 was investigated through microscopic observation, emphasizing the development of conidiogenous cells as a crucial stage, from which filamentous sectors originate. Some physiological characteristics of IOJ-3 and filamentous sectors are compared, and strain IOJ-3 has a higher antibiotic tolerance than two filamentous sectors, IOJ-3a expands faster on the culture medium, and IOJ-3b can penetrate cellophane. A transcriptomic analysis was conducted to investigate the differentially expressed genes between the yeast form IOJ-3 and its two filamentous sectors, revealing a total of 594 genes that exhibited consistent differential expression relative to IOJ-3, including 44 silencing genes in IOJ-3 that were activated. Gene Ontology analysis indicated that these differentially expressed genes were primarily associated with the cellular component category. Furthermore, adding 5-Azacytidine accelerated filamentous sectorization and increased the proportion of filamentous cells of strain IOJ-3 in PD liquid media, suggesting that the filamentous sectorization observed in strain IOJ-3 is linked to processes of DNA demethylation. In conclusion, this study sheds light on the biological characteristics of D. sorbi regarding morphological transitions and provides substantial direction for exploring genes related to fungal filamentous development.

Engineering Adhesion of the Probiotic Strain Escherichia coli Nissle to the Fungal Pathogen Candida albicans.

Engineering live biotherapeutic products against fungal pathogens such as Candida albicans has been suggested as a means to tackle the increasing threat of fungal infections and the development of resistance to classical antifungal treatments. One important challenge in the design of live therapeutics is to control their localization inside the human body. The specific binding capability to target organisms or tissues would greatly increase their effectiveness by increasing the local concentration of effector molecules at the site of infection. In this study, we utilized surface display of carbohydrate binding domains to enable the probiotic E. coli Nissle 1917 to adhere specifically to the pathogenic yeast Candida albicans. Binding was quantified using a newly developed method based on the automated analysis of microscopic images. In addition to a rationally selected chitin binding domain, a synthetic peptide of identical length but distinct sequence also conferred binding. Efficient binding was specific to fungal hyphae, the invasive form of C. albicans, while the yeast form, as well as abiotic cellulose and PET particles, was only weakly recognized.

Sinefungin, a natural nucleoside analog of S-adenosyl methionine, impairs the pathogenicity of Candida albicans

Candida albicans, an opportunistic fungal pathogen, causes life-threatening infections in immunocompromised patients. Current antifungals are limited by toxicity, drug-drug interactions, and emerging resistance, underscoring the importance of identifying novel treatment approaches. Here, we elucidate the impact of sinefungin, an analog of S-adenosyl methionine, on the virulence of C. albicans strain SC5314 and clinical isolates. Our data indicate that sinefungin impairs pathogenic traits of C. albicans including hyphal morphogenesis, biofilm formation, adhesion to epithelial cells, and virulence towards Galleria mellonella, highlighting sinefungin as an avenue for therapeutic intervention. We determine that sinefungin particularly disturbs N6-methyladenosine (m6A) formation. Transcriptome analysis of C. albicans hyphae upon sinefungin treatment reveals an increase in transcripts related to the yeast form and decrease in those associated with hyphae formation and virulence. Collectively, our data propose sinefungin as a potent molecule against C. albicans and emphasize further exploration of post-transcriptional control mechanisms of pathogenicity for antifungal design.

An overview of various stages and morphology of Histoplasma capsulatum var. farciminosum in the horse.

This article describes the development of the pathogenic dimorphic fungus Histoplasma capsulatum var. farciminosum (HCF), which is the causative agent of Equine epizootic lymphangitis (EEL), from the mycelial form in the soil to the yeast form in the horse. In this study, the stages and morphology of HCF were identified through histopathological analysis and culture with various samples collected in Ethiopia from 15 horses showing clinical signs of EEL. In equids, especially cart horses in Ethiopia, poor-quality harnesses cause cutaneous wounds, which often attract flies facilitating the transmission of the fungus. Also, HCF infection occurs through open wounds or ocular mucous membranes when horses roll on contaminated damp soil. Respiratory histoplasmosis can occur through inhaling fungal spores, which is rare. HCF microconidia enter the lungs and skin wounds and are phagocytized by tissue-resident macrophages. The spores undergo intracellular replication within the macrophages transitioning into yeasts. The infected macrophages undergo lysis releasing pathogenic yeast cells into the surrounding tissue. Consequently, yeast-rich purulent exudate is produced, contaminating the soil in stables where yeast cells germinate into the mycelial form, and the entire process starts from the beginning.

Yeast and filamentous Candida auris stimulate distinct immune responses in the skin.

Candida auris, an emerging multidrug-resistant fungal pathogen, predominately colonizes the human skin long term leading to subsequent life-threatening invasive infections. Fungal morphology is believed to play a critical role in modulating mucocutaneous antifungal immunity. In this study, we used an intradermal mouse model of C. auris infection to examine fungal colonization and the associated innate and adaptive immune response to yeast and filamentous C. auris strains. Our results indicate that mice infected with a filamentous C. auris had significantly decreased fungal load compared to mice infected with the yeast form. Mice infected with yeast and filamentous forms of C. auris stimulated distinct innate immune responses. Phagocytic cells (CD11b+Ly6G+ neutrophils, CD11b+Ly6Chi inflammatory monocytes, and CD11b+MHCII+CD64+ macrophages) were differentially recruited to mouse skin tissue infected with yeast and filamentous C. auris. The percentage and absolute number of interleukin 17 (IL-17) producing innate lymphoid cells, TCRγδ+, and CD4+ T cells in the skin tissue of mice infected with filamentous C. auris were significantly increased compared to the wild-type of yeast strain. Furthermore, complementation of filamentous mutant strain of C. auris (Δelm1 + ELM1) strain exhibited wild-type yeast morphology in vivo and induced comparable level of skin immune responses similar to mice infected with yeast strain. Collectively, our findings indicate that yeast and filamentous C. auris induce distinct local immune responses in the skin. The decreased fungal load observed in mouse skin infected with filamentous C. auris is associated with a potent IL-17 immune response induced by this morphotype.IMPORTANCECandida auris is a globally emerging fungal pathogen that transmits among individuals in hospitals and nursing home residents. Unlike other Candida species, C. auris predominantly colonizes and persists in skin tissue resulting in outbreaks of systemic infections. Understanding the factors that regulate C. auris skin colonization and host immune response is critical to develop novel preventive and therapeutic approaches against this emerging pathogen. We identified that yeast and filamentous forms of C. auris induce distinct skin immune responses in the skin. These findings may help explain the differential colonization and persistence of C. auris morphotypes in skin tissue. Understanding the skin immune responses induced by yeast and filamentous C. auris is important to develop novel vaccine strategies to combat this emerging fungal pathogen.

A Proteogenomic Pipeline for the Analysis of Protein Biosynthesis Errors in the Human Pathogen Candida albicans

Candida albicans is a diploid pathogen known for its ability to live as a commensal fungus in healthy individuals but causing both superficial infections and disseminated candidiasis in immunocompromised patients where it is associated with high morbidity and mortality. Its success in colonizing the human host is attributed to a wide range of virulence traits that modulate interactions between the host and the pathogen, such as optimal growth rate at 37°C, the ability to switch between yeast and hyphal forms, and a remarkable genomic and phenotypic plasticity. A fascinating aspect of its biology is a prominent heterogeneous proteome that arises from frequent genomic rearrangements, high allelic variation, and high levels of amino acid misincorporations in proteins. This leads to increased morphological and physiological phenotypic diversity of high adaptive potential, but the scope of such protein mistranslation is poorly understood due to technical difficulties in detecting and quantifying amino acid misincorporation events in complex protein samples. We have developed and optimized mass spectrometry and bioinformatics pipelines capable of identifying rare amino acid misincorporation events at the proteome level. We have also analyzed the proteomic profile of an engineered C.albicans strain that exhibits high level of leucine misincorporation at protein CUG sites and employed an invivo quantitative gain-of-function fluorescence reporter system to validate our LC-MS/MS data. C.albicans misincorporates amino acids above the background level at protein sites of diverse codons, particularly at CUG, confirming our previous data on the quantification of leucine incorporation at single CUG sites of recombinant reporter proteins, but increasing misincorporation of Leucine at these sites does not alter the translational fidelity of the other codons. These findings indicate that the C.albicans statistical proteome exceeds prior estimates, suggesting that its highly plastic phenome may also be modulated by environmental factors due to translational ambiguity.

Cold atmospheric plasma therapy for Malassezia folliculitis: Laboratory investigations and a randomized clinical trial.

Current treatment options for Malassezia folliculitis (MF) are limited. Recent research has demonstrated the inhibitory effect of cold atmospheric plasma (CAP) on the growth of Malassezia pachydermatis in vitro, suggesting CAP as a potential therapeutic approach for managing MF. The objective of our study is to assess the in vitro antifungal susceptibility of Malassezia yeasts to CAP. Additionally, we aim to evaluate the efficacy and tolerability of CAP in treating patients with MF. We initially studied the antifungal effect of CAP on planktonic and biofilm forms of Malassezia yeasts, using well-established techniques such as zone of inhibition, transmission electron microscopy, colony count assay and 2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide salt assay. Subsequently, a randomized (1:1 ratio), active comparator-controlled, observer-blind study was conducted comparing daily CAP therapy versus itraconazole 200mg/day for 2weeks in 50 patients with MF. Efficacy outcomes were measured by success rate, negative microscopy rate and changes in Dermatology Life Quality Index (DLQI) and Global Aesthetic Improvement Scale (GAIS) scores. Safety was assessed by monitoring adverse events (AEs) and local tolerability. In laboratory investigations, CAP time-dependently inhibited the growth of Malassezia yeasts in both planktonic and biofilm forms. Forty-nine patients completed the clinical study. At week 2, success was achieved by 40.0% of subjects in the CAP group versus 58.3% in the itraconazole group (p=0.199). The negative direct microscopy rates of follicular samples were 56.0% in the CAP group versus 66.7% in the itraconazole group (p=0.444). No significant differences were found in the proportion of subjects achieving DLQI scores of 0/1 (p=0.456) or in the GAIS responder rates (p=0.588) between the two groups. Three patients in the CAP group and one patient in the itraconazole group reported mild AEs. CAP demonstrated significant antifungal activity against Malassezia yeasts in vitro and exhibited comparable efficacy to itraconazole in treating MF patients. Without the associated adverse effects of oral antifungal drugs, CAP can be considered a promising and safe treatment modality for MF.

Genotypic diversity, virulence, and molecular genetic tools in Histoplasma.

SUMMARYHistoplasmosis is arguably the most common fungal respiratory infection worldwide, with hundreds of thousands of new infections occurring annually in the United States alone. The infection can progress in the lung or disseminate to visceral organs and can be difficult to treat with antifungal drugs. Histoplasma, the causative agent of the disease, is a pathogenic fungus that causes life-threatening lung infections and is globally distributed. The fungus has the ability to germinate from conidia into either hyphal (mold) or yeast form, depending on the environmental temperature. This transition also regulates virulence. Histoplasma and histoplasmosis have been classified as being of emergent importance, and in 2022, the World Health Organization included Histoplasma as 1 of the 19 most concerning human fungal pathogens. In this review, we synthesize the current understanding of the ecological niche, evolutionary history, and virulence strategies of Histoplasma. We also describe general patterns of the symptomatology and epidemiology of histoplasmosis. We underscore areas where research is sorely needed and highlight research avenues that have been productive.

Anti-adhesive, anti-biofilm and fungicidal action of newly synthesized gemini quaternary ammonium salts

Newly synthesized gemini quaternary ammonium salts (QAS) with different counterions (bromide, hydrogen chloride, methylcarbonate, acetate, lactate), chain lengths (C12, C14, C16) and methylene linker (3xCH2) were tested. Dihydrochlorides and dibromides with 12 carbon atoms in hydrophobic chains were characterized by the highest biological activity against planktonic forms of yeast and yeast-like fungi. The tested gemini surfactants also inhibited the production of filaments by C. albicans. Moreover, they reduced the adhesion of C. albicans cells to the surfaces of stainless steel, silicone and glass, and slightly to polystyrene. In particular, the gemini compounds with 16-carbon alkyl chains were most effective against biofilms. It was also found that the tested surfactants were not cytotoxic to yeast cells. Moreover, dimethylcarbonate (2xC12MeCO3G3) did not cause hemolysis of sheep erythrocytes. Dihydrochlorides, dilactate and diacetate showed no mutagenic potential.

Farnesol‐Imprinted Nanospheres (FINs) as Quorum Sensing Modulators of Candida albicans Hyphae Formation

AbstractCandida albicans (C. albicans) produces farnesol as a quorum sensing molecule that suppresses its filamentation. In this study, we synthesized farnesol‐imprinted nanospheres (FINs) to investigate their impact on the morphogenic transition of C. albicans, a fungus known for its pathogenicity in superficial and systemic candidiasis. FINs, synthesized for this purpose, exhibited a significantly higher binding capacity for farnesol compared to control polymers, with a 6.5‐fold increase observed. These polymers displayed a spherical morphology with diameters ranging from 500–700 nm. Treatment with FINs resulting in an increase in hyphae formation in C. albicans, indicating effective capture farnesol by the FINs. This increase in hyphae formation corresponded to a suppressed yeast form of the fungus. Consequently, FINs hold promise as a potential compound to mitigate the severity of disseminated candidiasis by depleting yeast forms. Interestingly, pre‐incubated of FINs with 0.1 mM of farnesol led to a reduction in C. albicans filamentation. This finding suggests that FINs have the capability to adsorb and release farnesol. Therefore, FINs could serve as a delivery agent for farnesol, offering protection against C. albicans filamentation. Overall, the synthesis of FINs presents a promising approach to combating C. albicans infection, whether by reducing disseminated candidiasis or preventing fungal superficial localization.

Assessment of transaminase enzymes and effect of potassium iodide on its production in the yeast form of Sporothrix schenckii

Assessment of transaminase enzymes and effect of potassium iodide on its production in the yeast form of Sporothrix schenckii

Differential alteration in Lactiplantibacillus plantarum subsp. plantarum quorum-sensing systems and reduced Candida albicans yeast survival and virulence gene expression in dual-species interaction.

Candida albicans (C. albicans) and Lactiplantibacillus plantarum subsp. plantarum (L. plantarum) are frequently identified in various niches, but their dual-species interaction, especially with C. albicans in yeast form, remains unclear. This study aimed to investigate the dual-species interaction of L. plantarum and C. albicans, including proliferation, morphology, and transcriptomes examined by selective agar plate counting, microscopy, and polymicrobial RNA-seq, respectively. Maintaining a stable and unchanged growth rate, L. plantarum inhibited C. albicans yeast cell proliferation but not hyphal growth. Combining optical microscopy and atomic force microscopy, cell-to-cell direct contact and co-aggregation with L. plantarum cells surrounding C. albicans yeast cells were observed during dual-species interaction. Reduced C. albicans yeast cell proliferation in mixed culture was partially due to L. plantarum cell-free culture supernatant but not the acidic environment. Upon polymicrobial transcriptomics analysis, interesting changes were identified in both L. plantarum and C. albicans gene expression. First, two L. plantarum quorum-sensing systems showed contrary changes, with the activation of lamBDCA and repression of luxS. Second, the upregulation of stress response-related genes and downregulation of cell cycle, cell survival, and cell integrity-related pathways were identified in C. albicans, possibly connected to the stress posed by L. plantarum and the reduced yeast cell proliferation. Third, a large scale of pathogenesis and virulence factors were downregulated in C. albicans, indicating the potential interruption of pathogenic activities by L. plantarum. Fourth, partial metabolism and transport pathways were changed in L. plantarum and C. albicans. The information in this study might aid in understanding the behavior of L. plantarum and C. albicans in dual-species interaction.IMPORTANCEThe anti-Candida albicans activity of Lactiplantibacillus plantarum has been explored in the past decades. However, the importance of C. albicans yeast form and the effect of C. albicans on L. plantarum had also been omitted. In this study, the dual-species interaction of L. plantarum and C. albicans was investigated with a focus on the transcriptomes. Cell-to-cell direct contact and co-aggregation with L. plantarum cells surrounding C. albicans yeast cells were observed. Upon polymicrobial transcriptomics analysis, interesting changes were identified, including contrary changes in two L. plantarum quorum-sensing systems and reduced cell survival-related pathways and pathogenesis determinants in C. albicans.

Exophiala amphawaensis sp. nov., a novel black yeast isolated from the gut of amphipods in Thailand.

The genus Exophiala is polymorphic, able to transition between yeast, hyphal and pseudohyphal forms. Species of the genus Exophiala are ubiquitous fungi that are distributed in various environments around the world. During a survey of fungal diversity in the gut of amphipods (Floresorchestia amphawaensis and undescribed Dogielinotid amphipods) from the Amphawa estuary, Samut Songkhram province, Thailand, five black yeast strains (DMKU-MG01, DMKU-MG07, DMKU-MG08, DMKU-HG10 and DMKU-FG04) were identified as representing a novel taxon on the basis of a combination of morphological and molecular phylogenetic features. The five strains did not produce filamentous hyphae or pseudohyphae. Only budding yeast cells were observed. On the basis of the phenotypic characteristics and the results of molecular analyses of the D1/D2 region of the large subunit (LSU) rRNA gene and the internal transcribed spacer (ITS) region, the five strains were identified as representing a novel species via applied nucleotide pairwise analysis. They differed from the most closely related species Exophiala alcalophiala by 3.54 % nucleotide substitutions (20 nucleotide substitutions in 572 bp) in the D1/D2 domains of the LSU rRNA gene. Moreover, the sequences of the ITS region of the five strains differed from those of the most closely related species E. alcalophiala, by 7.44-9.62 % nucleotide substitutions, and Exophiala halophiala, by 7.2-7.53 % nucleotide substitutions. The results of phylogenetic analyses based on the concatenated sequences of the ITS regions and the D1/D2 domains of the LSU rRNA gene confirmed that the five black yeast strains represented a single novel species of the genus Exophiala. In this study, Exophiala amphawaensis sp. nov. is proposed to accommodate these strains. The holotype is TBRC 15626T and the isotype is PYCC9020. The MycoBank accession number of the novel species is MB 851477.

Segmentation of hyphae and yeast in fungi-infected tissue slice images and its application in analyzing antifungal blue light therapy.

Candida albicans is a pathogenic fungus that undergoes morphological transitions between hyphal and yeast forms, adapting to diverse environmental stimuli and exhibiting distinct virulence. Existing research works on antifungal blue light (ABL) therapy have either focused solely on hyphae or neglected to differentiate between morphologies, obscuring potential differential effects. To address this gap, we established a novel dataset of 150 C. albicans-infected mouse skin tissue slice images with meticulously annotated hyphae and yeast. Eleven representative convolutional neural networks were trained and evaluated on this dataset using seven metrics to identify the optimal model for segmenting hyphae and yeast in original high pixel size images. Leveraging the segmentation results, we analyzed the differential impact of blue light on the invasion depth and density of both morphologies within the skin tissue. U-Net-BN outperformed other models in segmentation accuracy, achieving the best overall performance. While both hyphae and yeast exhibited significant reductions in invasion depth and density at the highest ABL dose (180J/cm2), only yeast was significantly inhibited at the lower dose (135J/cm2). This novel finding emphasizes the importance of developing more effective treatment strategies for both morphologies.

Multifactorial inhibition of Candida albicans by combinations of lactobacilli and probiotic Saccharomyces cerevisiae CNCM I-3856

Strategies against the opportunistic fungal pathogen Candida albicans based on probiotic microorganisms represent a promising alternative to traditional antifungals. Here, we investigated the effects of Lactobacillaceae isolates from fermented foods or the human vagina, alone or in combination with the probiotic yeast Saccharomyces cerevisiae CNCM I-3856, against C. albicans in vitro. Nine out of nineteen tested strains of Lactobacillaceae inhibited growth of C. albicans with inhibition zones of 1–3 mm in spot assays. Five out of nineteen lactobacilli tested as such or in combination with S. cerevisiae CNCM I-3856 also significantly inhibited C. albicans hyphae formation, including Limosilactobacillus fermentum LS4 and L. fermentum LS5 resulting in respectively 62% and 78% hyphae inhibition compared to the control. Thirteen of the tested nineteen lactobacilli aggregated with the yeast form of C. albicans, with Lactiplantibacillus carotarum AMBF275 showing the strongest aggregation. The aggregation was enhanced when lactobacilli were combined with S. cerevisiae CNCM I-3856. No significant antagonistic effects were observed between the tested lactobacilli and S. cerevisiae CNCM I-3856. The multifactorial activity of Lactobacillaceae strains alone or combined with the probiotic S. cerevisiae CNCM I-3856 against C. albicans without antagonistic effects between the beneficial strains, paves the way for developing consortium probiotics for in vivo applications.