Human gut mycobiota tune immunity via CARD9-dependent induction of anti-fungal IgG antibodies.

Antibodies mediate natural and vaccine-induced immunity against viral and bacterial pathogens, whereas fungi represent a widespread kingdom of pathogenic species for which neither vaccine nor neutralizing antibody therapies are clinically available. Here, using a multi-kingdom antibody profiling (multiKAP) approach, we explore the human antibody repertoires against gut commensal fungi (mycobiota). We identify species preferentially targeted by systemic antibodies in humans, with Candida albicans being the major inducer of antifungal immunoglobulin G (IgG). Fungal colonization of the gut induces germinal center (GC)-dependent B cell expansion in extraintestinal lymphoid tissues and generates systemic antibodies that confer protection against disseminated C. albicans or C. auris infection. Antifungal IgG production depends on the innate immunity regulator CARD9 and CARD9+CX3CR1+ macrophages. In individuals with invasive candidiasis, loss-of-function mutations in CARD9 are associated with impaired antifungal IgG responses. These results reveal an important role of gut commensal fungi in shaping the human antibody repertoire through CARD9-dependent induction of host-protective antifungal IgG.

Diverse mechanisms control amino acid-dependent environmental alkalization by Candida albicans.

Candida albicans has the capacity to neutralize acidic growth environments by releasing ammonia derived from the catabolism of amino acids. The molecular components underlying alkalization and its physiological significance remain poorly understood. Here, we present an integrative model with the cytosolic NAD+-dependent glutamate dehydrogenase (Gdh2) as the principal ammonia-generating component. We show that alkalization is dependent on the SPS-sensor-regulated transcription factor STP2 and the proline-responsive activator Put3. These factors function in parallel to derepress GDH2 and the two proline catabolic enzymes PUT1 and PUT2. Consistently, a double mutant lacking STP2 and PUT3 exhibits a severe alkalization defect that nearly phenocopies that of a gdh2-/- strain. Alkalization is dependent on mitochondrial activity and in wild-type cells occurs as long as the conditions permit respiratory growth. Strikingly, Gdh2 levels decrease and cells transiently extrude glutamate as the environment becomes more alkaline. Together, these processes constitute a rudimentary regulatory system that counters and limits the negative effects associated with ammonia generation. These findings align with Gdh2 being dispensable for virulence, and based on a whole human blood virulence assay, the same is true for C. glabrata and C. auris. Using a transwell co-culture system, we observed that the growth and proliferation of Lactobacillus crispatus, a common component of the acidic vaginal microenvironment and a potent antagonist of C. albicans, is unaffected by fungal-induced alkalization. Consequently, although Candida spp. can alkalinize their growth environments, other fungal-associated processes are more critical in promoting dysbiosis and virulent fungal growth.

Defining Optimal Doses of Liposomal Amphotericin B Against Candida auris: Data From an In Vitro Pharmacokinetic/Pharmacodynamic Model.

BACKGROUND: Candida auris isolates exhibit elevated amphotericin B (AMB) minimum inhibitory concentrations (MICs). As liposomal AMB (L-AMB) can be safely administered at high doses, we explored L-AMB pharmacodynamics against C. auris isolates in an in vitro pharmacokinetic/pharmacodynamic (PK/PD) dilution model. METHODS: Four C. auris isolates with Clinical and Laboratory Standards Institute (CLSI) AMB MICs = 0.5-2 mg/L were tested in an in vitro PK/PD model simulating L-AMB pharmacokinetics. The in vitro model was validated using a Candida albicans isolate tested in animals. The peak concentration (Cmax)/MIC versus log10 colony-forming units (CFU)/mL reduction from the initial inoculum was analyzed with the sigmoidal model with variable slope (Emax model). Monte Carlo analysis was performed for the standard (3 mg/kg) and higher (5 mg/kg) L-AMB doses. RESULTS: The in vitro PK/PD relationship Cmax/MIC versus log10 CFU/mL reduction followed a sigmoidal pattern (R2 = 0.91 for C. albicans, R2 = 0.86 for C. auris). The Cmax/MIC associated with stasis was 2.1 for C. albicans and 9 for C. auris. The probability of target attainment was >95% with 3 mg/kg for wild-type C. albicans isolates with MIC ≤2 mg/L and C. auris isolates with MIC ≤1 mg/L whereas 5 mg/kg L-AMB is needed for C. auris isolates with MIC 2 mg/L. CONCLUSIONS: L-AMB was 4-fold less active against C. auris than C. albicans. Candida auris isolates with CLSI MIC 2 mg/L would require a higher L-AMB dose.

Impact of micafungin on Candida auris ß-glucan masking and neutrophil interactions.

Invasive fungal pathogen Candida auris has become a public health threat causing outbreaks of high mortality infections. Drug resistance often limits treatment options. For Candida albicans, subinhibitory concentrations of echinocandins unmask immunostimulatory ß-glucan, augmenting immunity. Here we analyze the impact of echinocandin treatment of C. auris on ß-glucan exposure and human neutrophil interactions. We show subinhibitory concentrations lead to minimal glucan unmasking and only subtle influences on neutrophil functions for the isolates belonging to circulating clades. The data suggest that echinocandin treatment will not largely alter phagocytic responses. Glucan masking pathways appear to differ between C. auris and C. albicans.

Identification of the Most Immunoreactive Antigens of Candida auris to IgGs from Systemic Infections in Mice.

The delay in making a correct diagnosis of Candida auris causes concern in the healthcare system setting, and immunoproteomics studies are important to identify immunoreactive proteins for new diagnostic strategies. In this study, immunocompetent murine systemic infections caused by non-aggregative and aggregative phenotypes of C. auris and by Candida albicans and Candida haemulonii were carried out, and the obtained sera were used to study their immunoreactivity against C. auris proteins. The results showed higher virulence, in terms of infection signs, weight loss, and histopathological damage, of the non-aggregative isolate. Moreover, C. auris was less virulent than C. albicans but more than C. haemulonii. Regarding the immunoproteomics study, 13 spots recognized by sera from mice infected with both C. auris phenotypes and analyzed by mass spectrometry corresponded to enolase, phosphoglycerate kinase, glyceraldehyde-3-phosphate dehydrogenase, and phosphoglycerate mutase. These four proteins were also recognized by sera obtained from human patients with disseminated C. auris infection but not by sera obtained from mice infected with C. albicans or Aspergillus fumigatus. Spot identification data are available via ProteomeXchange with the identifier PXD049077. In conclusion, this study showed that the identified proteins could be potential candidates to be studied as new diagnostic or even therapeutic targets for C. auris.

A correlative study of the genomic underpinning of virulence traits and drug tolerance of Candida auris.

Candida auris is an opportunistic fungal pathogen with high mortality rates which presents a clear threat to public health. The risk of C. auris infection is high because it can colonize the body, resist antifungal treatment, and evade the immune system. The genetic mechanisms for these traits are not well known. Identifying them could lead to new targets for new treatments. To this end, we present an analysis of the genetics and gene expression patterns of C. auris carbon metabolism, drug resistance, and macrophage interaction. We chose to study two C. auris isolates simultaneously, one drug sensitive (B11220 from Clade II) and one drug resistant (B11221 from Clade III). Comparing the genomes, we confirm the previously reported finding that B11220 was missing a 12.8 kb region on chromosome VI. This region contains a gene cluster encoding proteins related to alternative sugar utilization. We show that B11221, which has the gene cluster, readily assimilates and utilizes D-galactose and L-rhamnose as compared to B11220, which harbors the deletion. B11221 exhibits increased adherence and drug resistance compared to B11220 when grown in these sugars. Transcriptomic analysis of both isolates grown on glucose or galactose showed that the gene cluster was upregulated when grown on D-galactose. These findings reinforce growing evidence of a link between metabolism and drug tolerance. B11221 resists phagocytosis by macrophages and exhibits decreased ß-1,3-glucan exposure, a key determinant that allows Candida to evade the host immune system, as compared to B11220. In a transcriptomic analysis of both isolates co-cultured with macrophages, we find upregulation of genes associated with transport and transcription factors in B11221. Our studies show a positive correlation between membrane composition and immune evasion, alternate sugar utilization, and drug tolerance in C. auris.

Factors Associated with Poor Clinical and Microbiologic Outcomes in C. auris Bloodstream Infection: A Multicenter Retrospective Cohort Study.

BACKGROUND: C. auris has become a growing concern worldwide due to increases in incidence of colonization and reports of invasive infections. There are limited data on clinical factors associated with poor outcomes in patients with C. auris bloodstream infection (BSI). METHODS: We assembled a multicenter retrospective cohort of patients with C. auris BSI from two geographics areas in US healthcare settings. We collected data on demographic, clinical, and microbiologic characteristics to describe the cohort and constructed multivariate logistic regression models to understand risk factors for two clinical outcomes, all-cause mortality during facility admission, and blood culture clearance. RESULTS: Our cohort consisted of 187 patients with C. auris BSI (56.1% male, 55.6% age >65 years); 54.6% died by facility discharge and 66.9% (of 142 with available data) experienced blood culture clearance. Pitt bacteremia score at infection onset was associated with mortality (odds-ratio [95% confidence interval]: 1.19 [1.01,1.40] per 1-point increase). Hemodialysis was associated with a reduced odds of microbiologic clearance (0.15 [0.05,0.43]) and with mortality (3.08 [1.27,7.50]). CONCLUSIONS: The Pitt bacteremia score at the onset of C. auris BSI may be a useful tool in identifying patients at risk for mortality. Targeted infection prevention practices in patients receiving hemodialysis may be useful to limit poor outcomes.

Candida auris in US Correctional Facilities.

Candida auris is an emerging fungal pathogen that typically affects patients in healthcare settings. Data on C. auris cases in correctional facilities are limited but are needed to guide public health recommendations. We describe cases and challenges of providing care for 13 patients who were transferred to correctional facilities during January 2020-December 2022 after having a positive C. auris specimen. All patients had positive specimens identified while receiving inpatient care at healthcare facilities in geographic areas with high C. auris prevalence. Correctional facilities reported challenges managing patients and implementing prevention measures; those challenges varied by whether patients were housed in prison medical units or general population units. Although rarely reported, C. auris cases in persons who are incarcerated may occur, particularly in persons with known risk factors. Measures to manage cases and prevent C. auris spread in correctional facilities should address setting-specific challenges in healthcare and nonhealthcare correctional environments.

Candida auris Updates: Outbreak Evaluation through Molecular Assays and Antifungal Stewardship-A Narrative Review.

Candida auris was reported by the WHO as second to Cryptococcus neoformans, in the list of nineteen fungal priority pathogens, along with two species with a new nomenclature, Nakaseomyces glabrata (Candida glabrata) and Pichia kudriavzevii (Candida krusei). This novel classification was based on antifungal resistance, the number of deaths, evidence-based treatment, access to diagnostics, annual incidence, and complications and sequelae. We assessed which molecular assays have been used to diagnose Candida auris outbreaks in the last five years. Using "Candida auris; outbreak; molecular detection" as keywords, our search in PubMed revealed 32 results, from which we selected 23 original papers published in 2019-2024. The analyzed studies revealed that the detection methods were very different: from the VITEK® 2 System to MALDI TOF (Matrix-Assisted Laser Desorption Ionization-Time of Flight), NGS (Next-Generation Sequencing), WGS (Whole Genome Sequencing), and commercially available real-time PCR (Polymerase Chain Reaction) assays. Moreover, we identified studies that detected antifungal resistance genes (e.g., FKS for echinocandins and ERG11 for azoles). The analyzed outbreaks were from all continents, which confirms the capability of this yeast to spread between humans and to contaminate the environment. It is important that real-time PCR assays were developed for accurate and affordable detection by all laboratories, including the detection of antifungal resistance genes. This will allow the fast and efficient implementation of stewardship programs in hospitals.

Liposomal formulation of a new antifungal hybrid compound provides protection against Candida auris in the ex vivo skin colonization model.

The newly emerged pathogen, Candida auris, presents a serious threat to public health worldwide. This multidrug-resistant yeast often colonizes and persists on the skin of patients, can easily spread from person to person, and can cause life-threatening systemic infections. New antifungal therapies are therefore urgently needed to limit and control both superficial and systemic C. auris infections. In this study, we designed a novel antifungal agent, PQA-Az-13, that contains a combination of indazole, pyrrolidine, and arylpiperazine scaffolds substituted with a trifluoromethyl moiety. PQA-Az-13 demonstrated antifungal activity against biofilms of a set of 10 different C. auris clinical isolates, representing all four geographical clades distinguished within this species. This compound showed strong activity, with MIC values between 0.67 and 1.25 µg/mL. Cellular proteomics indicated that PQA-Az-13 partially or completely inhibited numerous enzymatic proteins in C. auris biofilms, particularly those involved in both amino acid biosynthesis and metabolism processes, as well as in general energy-producing processes. Due to its hydrophobic nature and limited aqueous solubility, PQA-Az-13 was encapsulated in cationic liposomes composed of soybean phosphatidylcholine (SPC), 1,2-dioleoyloxy-3-trimethylammonium-propane chloride (DOTAP), and N-(carbonyl-methoxypolyethylene glycol-2000)-1,2-distearoyl-sn-glycero-3-phosphoethanolamine, sodium salt (DSPE-PEG 2000), and characterized by biophysical and spectral techniques. These PQA-Az-13-loaded liposomes displayed a mean size of 76.4 nm, a positive charge of +45.0 mV, a high encapsulation efficiency of 97.2%, excellent stability, and no toxicity to normal human dermal fibroblasts. PQA-Az-13 liposomes demonstrated enhanced antifungal activity levels against both C. auris in in vitro biofilms and ex vivo skin colonization models. These initial results suggest that molecules like PQA-Az-13 warrant further study and development.

In vitro activity of taurolidine against clinical Candida auris isolates: relevance to catheter-related bloodstream infections.

Candida auris is an evolving and concerning global threat. Of particular concern are bloodstream infections related to central venous catheters. We evaluated the activity of taurolidine, a broad-spectrum antimicrobial in catheter lock solutions, against 106 C. auris isolates. Taurolidine was highly active with a MIC50/MIC90 of 512/512 mg/L, over 20-fold lower than lock solution concentrations of ≥13,500 mg/L. Our data demonstrate a theoretical basis for taurolidine-based lock solutions for prevention of C. auris catheter-associated infections.

Validation of a qualitative real-time PCR assay for the detection of Candida auris in hospital inpatient screening.

Candida auris is a multidrug-resistant opportunistic fungal pathogen capable of causing serious infections and healthcare-associated outbreaks. Screening for colonization with C. auris has become routine and is recommended in many hospitals and healthcare facilities as an infection control and prevention strategy. Subsequently, and since there are currently no FDA-approved tests for this purpose, clinical microbiology laboratories have become responsible for developing protocols to detect C. auris using axial and inguinal screening swabs. In a College of American Pathologists-accredited large academic healthcare center setting, we implemented a laboratory-developed nucleic-acid amplification test for the detection of C. auris DNA. Our test validation evaluated the performance of the DiaSorin C. auris primer set used in a real-time qualitative PCR assay on the LIAISON MDX thermocycler with the Simplexa Universal Disc. The assay was highly sensitive and specific, with a limit of detection of 1-2 CFU/reaction, with no observed cross-reactivity with other Candida spp., bacterial skin commensal organisms or commonly encountered viruses. When run in parallel with a culture-based detection method, the PCR assay was 100% sensitive and specific. The assay was precise, with low variability between replicates within and between runs. Lastly, pre-analytical factors, including swab storage time, temperature, and transport media, were assessed and found to have no significant effect on the detection of C. auris at variable concentrations. Taken together, this study expands the available options for nucleic acid detection of C. auris and characterizes pre-analytical factors for implementation in both high- and low-volume laboratory settings. IMPORTANCE: This study overviews the validation and implementation of a molecular screening tool for the detection of Candida auris in a College of American Pathologist-accredited clinical laboratory. This molecular laboratory-developed test is both highly sensitive and specific and has significant health-system cost-savings associated with significantly reduced turn-around-time compared to traditional standard-of-care culture-based work up. This method and workflow is of interest to support clinical microbiology diagnostics and to help aid in hospital inpatient, and infection prevention control screening.

Evaluation of the Vitek 2 system for antifungal susceptibility testing of Candida auris using a representative international panel of clinical isolates: overestimation of amphotericin B resistance and underestimation of fluconazole resistance.

Although the Vitek 2 system is broadly used for antifungal susceptibility testing of Candida spp., its performance against Candida auris has been assessed using limited number of isolates recovered from restricted geographic areas. We therefore compared Vitek 2 system with the reference Clinical and Laboratory Standards Institute (CLSI) broth microdilution method using an international collection of 100 C. auris isolates belonging to different clades. The agreement ±1 twofold dilution between the two methods and the categorical agreement (CA) based on the Centers for Disease Control and Prevention's (CDC's) tentative resistance breakpoints and Vitek 2-specific wild-type upper limit values (WT-ULVs) were determined. The CLSI-Vitek 2 agreement was poor for 5-flucytosine (0%), fluconazole (16%), and amphotericin B (29%), and moderate for voriconazole (61%), micafungin (67%), and caspofungin (81%). Significant interpretation errors were recorded using the CDC breakpoints for amphotericin B (31% CA, 69% major errors; MaEs) and fluconazole (69% CA, 31% very major errors; VmEs), but not for echinocandins (99% CA, 1% MaEs for both micafungin and caspofungin) for which the Vitek 2 allowed correct categorization of echinocandin-resistant FKS1 mutant isolates. Discrepancies were reduced when the Vitek 2 WT-ULV of 16 mg/L for amphotericin B (98% CA, 2% MaEs) and of 4 mg/L for fluconazole (96% CA, 1% MaEs, 3% VmEs) were used. In conclusion, the Vitek 2 system performed well for echinocandin susceptibility testing of C .auris. Resistance to fluconazole was underestimated whereas resistance to amphotericin B was overestimated using the CDC breakpoints of ≥32 and ≥2 mg/L, respectively. Vitek 2 minimun inhibitory concentrations (MICs) >4 mg/L indicated resistance to fluconazole and Vitek 2 MICs ≤16 mg/L indicated non-resistance to amphotericin B.

A simple and sensitive test for Candida auris colonization, surveillance, and infection control suitable for near patient use.

Candida auris is a multidrug-resistant fungal pathogen with a propensity to colonize humans and persist on environmental surfaces. C. auris invasive fungal disease is being increasingly identified in acute and long-term care settings. We have developed a prototype cartridge-based C. auris surveillance assay (CaurisSurV cartridge; "research use only") that includes integrated sample processing and nucleic acid amplification to detect C. auris from surveillance skin swabs in the GeneXpert instrument and is designed for point-of-care use. The assay limit of detection (LoD) in the skin swab matrix was 10.5 and 14.8 CFU/mL for non-aggregative (AR0388) and aggregative (AR0382) strains of C. auris, respectively. All five known clades of C. auris were detected at 2-3-5× (31.5-52.5 CFU/mL) the LoD. The assay was validated using a total of 85 clinical swab samples banked at two different institutions (University of California Los Angeles, CA and Wadsworth Center, NY). Compared to culture, sensitivity was 96.8% (30/31) and 100% (10/10) in the UCLA and Wadsworth cohorts, respectively, providing a combined sensitivity of 97.5% (40/41), and compared to PCR, the combined sensitivity was 92% (46/50). Specificity was 100% with both clinical (C. auris negative matrix, N = 31) and analytical (non-C. auris strains, N = 32) samples. An additional blinded study with N = 60 samples from Wadsworth Center, NY yielded 97% (29/30) sensitivity and 100% (28/28) specificity. We have developed a completely integrated, sensitive, specific, and 58-min prototype test, which can be used for routine surveillance of C. auris and might help prevent colonization and outbreaks in acute and chronic healthcare settings. IMPORTANCE: This study has the potential to offer a better solution to healthcare providers at hospitals and long-term care facilities in their ongoing efforts for effective and timely control of Candida auris infection and hence quicker response for any potential future outbreaks.

Chitinase-functionalized UiO-66 framework nanoparticles active against multidrug-resistant Candida Auris.

Candida auris (C. auris) is a yeast that has caused several outbreaks in the last decade. Cell wall chitin plays a primary role in the antifungal resistance of C. auris. Herein, we investigated the potential of chitinase immobilized with UiO-66 to act as a potent antifungal agent against C. auris. Chitinase was produced from Talaromyces varians SSW3 in a yield of 8.97 U/g dry substrate (ds). The yield was statistically enhanced to 120.41 U/g ds by using Plackett-Burman and Box-Behnken design. We synthesized a UiO-66 framework that was characterized by SEM, TEM, XRD, FTIR, a particle size analyzer, and a zeta sizer. The produced framework had a size of 70.42 ± 8.43 nm with a uniform cubic shape and smooth surface. The produced chitinase was immobilized on UiO-66 with an immobilization yield of 65% achieved after a 6 h loading period. The immobilization of UiO-66 increased the enzyme activity and stability, as indicated by the obtained Kd and T1/2 values. Furthermore, the hydrolytic activity of chitinase was enhanced after immobilization on UiO-66, with an increase in the Vmax and a decrease in the Km of 2- and 38-fold, respectively. Interestingly, the antifungal activity of the produced chitinase was boosted against C. auris by loading the enzyme on UiO-66, with an MIC50 of 0.89 ± 0.056 U/mL, compared to 5.582 ± 0.57 U/mL for the free enzyme. This study offers a novel promising alternative approach to combat the new emerging pathogen C. auris.

Screening of the global health priority BoxⓇ reveals potential new disinfectants against the emerging multidrug-resistant pathogen Candida auris.

BACKGROUND: Candida auris has been identified by the World Health Organization as a critical pathogen due to its invasive nature, resistance to multiple drugs, and high mortality rates in hospital outbreaks. This fungus can persist on surfaces and human skin for extended periods, complicating infection control efforts. The need for effective disinfection strategies is urgent, as current disinfectants are often ineffective against C. auris biofilms. OBJECTIVE: The study aimed to identify potential disinfectants from a collection of 240 compounds in the Global Health Priority Box® that are effective against C. auris, particularly strains resistant to existing options. METHODS: The research employed a screening protocol using a fluconazole-resistant strain of C. auris (149/23). Antifungal activity was assessed using the microdilution method to determine Minimum Inhibitory Concentrations (MICs) and Minimum Fungicidal Concentrations (MFCs). Additional assays were conducted to evaluate biofilm inhibition, biofilm eradication, cell membrane integrity, nucleotide leakage, sorbitol protection assay, efflux pump inhibition, and hemolysis assay. RESULTS: Two compounds, Hydramethylnon (MMV1577471) and Flufenerim (MMV1794206), demonstrated significant inhibitory effects against C. auris. Hydramethylnon exhibited potent antifungal activity, inhibiting up to 93 % of fungal growth with an MFC of 16 µg/mL. Flufenerim inhibited up to 58 % of fungal growth, showing fungistatic action with an MFC greater than 4 µg/mL. Biofilm inhibition tests showed that both compounds significantly inhibited biofilm formation, with increased efficacy at higher concentrations. Both compounds showed eradication rates in both stages. Furthermore, Hydramethylnon and Flufenerim did not affect cell membrane integrity or nucleotide leakage, suggesting a mode of action not reliant on disrupting these cellular components. The sorbitol protection assay revealed that neither compound caused cell wall damage. In the efflux pump inhibition assay, Hydramethylnon did not activate efflux pumps, while Flufenerim activated efflux pumps, reducing its effectiveness. Hemocompatibility assay showed safety. CONCLUSION: The study highlights Hydramethylnon and Flufenerim as promising candidates for further development as disinfectants, offering potential solutions to the urgent need for effective disinfection agents against C. auris. The findings underscore the value of screening compound collections to identify novel antifungal agents and understand their mechanisms of action, thereby contributing to the advancement of new disinfection strategies in healthcare settings.

Antifungal activity of synthetic xanthenone against fluconazole-resistant Candida auris and its mechanism of action.

Candida auris, an emerging multidrug-resistant fungal pathogen discovered in Japan in 2009, poses a significant global health threat, with infections reported in about 25 countries. The escalation of drug-resistant strains underscores the urgent need for new treatment options. This study aimed to investigate the antifungal potential of 2,3,4,4a-tetrahydro-1H-xanthen-1-one (XA1) against C. auris, as well as its mechanism of action and toxic profile. The antifungal activity of XA1 was first evaluated by determining the minimum inhibitory concentration (MIC), time-kill kinetics and biofilm inhibition. In addition, structural changes, membrane permeability, reactive oxygen species (ROS) production, and in vitro and in vivo toxicity of C. auris after exposure to XA1 were investigated. The results indicated that XA1 exhibited an MIC of 50 µg/mL against C. auris, with time-kill kinetics highlighting its efficacy. Field emission scanning electron microscopy (FE-SEM) showed structural damage in XA1-treated cells, supported by increased membrane permeability leading to cell death. Furthermore, XA1 induced ROS production and significantly inhibited biofilm formation. Importantly, XA1 exhibited low cytotoxicity in human epidermal keratinocytes (HaCaT), with a cell viability of over 90 % at 6.25 µg/mL. In addition, an LD50 of 17.68 µg/mL was determined in zebrafish embryos 24 h post fertilization (hpf), with developmental delay observed at prolonged exposure at 6.25 µg/mL (48-96 hpf). These findings position XA1 as a promising candidate for further research and development of an effective antifungal agent.

Sphingolipid diversity in Candida auris: unraveling interclade and drug resistance fingerprints.

In this study, we explored the sphingolipid (SL) landscape in Candida auris, which plays pivotal roles in fungal biology and drug susceptibility. The composition of SLs exhibited substantial variations at both the SL class and molecular species levels among clade isolates. Utilizing principal component analysis, we successfully differentiated the five clades based on their SL class composition. While phytoceramide (PCer) was uniformly the most abundant SL class in all the isolates, other classes showed significant variations. These variations were not limited to SL class level only as the proportion of different molecular species containing variable number of carbons in fatty acid chains also differed between the isolates. Also a comparative analysis revealed abundance of ceramides and glucosylceramides in fluconazole susceptible isolates. Furthermore, by comparing drug-resistant and susceptible isolates within clade IV, we uncovered significant intraclade differences in key SL classes such as high PCer and low long chain base (LCB) content in resistant strains, underscoring the impact of SL heterogeneity on drug resistance development in C. auris. These findings shed light on the multifaceted interplay between genomic diversity, SLs, and drug resistance in this emerging fungal pathogen.

Suppressing the virulence factors of Candida auris with baicalein through multifaceted mechanisms.

Candida auris, a rapidly spreading multi-drug-resistant fungus, is causing lethal infections under certain conditions globally. Baicalin (BE), an active ingredient extracted from the dried root of Scutellaria baicalensis Georgi, exhibits antifungal activity. However, studies have shown the distinctive advantages of Traditional Chinese medicine in combating fungal infections, while the effect of BE, an active ingredient extracted from the dried roots of Scutellaria baicalensis Georgi, on C. auris, remains unknown. Therefore, this study aims to evaluate the potential of BE as an antifungal agent against the emerging multidrug-resistant C. auris. Various assays and models, including microbroth dilution, time growth curve analysis, spot assays, adhesion tests, flocculation test, cell surface hydrophobicity assay, hydrolase activity assays, XTT assay, violet crystal assay, scanning electron microscope (SEM), confocal laser scanning microscope (CLSM), flow cytometry, Live/dead fluorescent staining, reactive oxygen species (ROS), cell wall assay, aggregation assay, porcine skin model, Galleria mellonella larvae (G. mellonella larvae) infection model, and reverse transcription-quantitative polymerase chain reaction (RT-PCR) were utilized to investigate how baicalein suppresses C. auris through possible multifaceted mechanisms. The findings indicate that BE strongly inhibited C. auris growth, adhesion, and biofilm formation. It also effectively reduced drug resistance and aggregation by disrupting the cell membrane and cell wall while reducing colonization and invasion of the host. Transcriptome analysis showed significant modulation in gene expression related to different virulence factors post-BE treatment. In conclusion, BE exhibits significant effectiveness against C. auris, suggesting its potential as a viable treatment option due to its multifaceted suppression mechanisms.

In silico genome wide identification of long non-coding RNAs differentially expressed during Candida auris host pathogenesis.

Candida auris is an invasive fungal pathogen of high concern due to acquired drug tolerance against antifungals used in clinics. The prolonged persistence on biotic and abiotic surfaces can result in onset of hospital outbreaks causing serious health threat. An in depth understanding of pathology of C. auris is highly desirable for development of efficient therapeutics. Non-coding RNAs play crucial role in fungal pathology. However, the information about ncRNAs is scanty to be utilized. Herein our aim is to identify long noncoding RNAs with potent role in pathobiology of C. auris. Thereby, we analyzed the transcriptomics data of C. auris infection in blood for identification of potential lncRNAs with regulatory role in determining invasion, survival or drug tolerance under infection conditions. Interestingly, we found 275 lncRNAs, out of which 253 matched with lncRNAs reported in Candidamine, corroborating for our accurate data analysis pipeline. Nevertheless, we obtained 23 novel lncRNAs not reported earlier. Three lncRNAs were found to be under expressed throughout the course of infection, in the transcriptomics data. 16 of potent lncRNAs were found to be coexpressed with coding genes, emphasizing for their functional role. Noteworthy, these ncRNAs are expressed from intergenic regions of the genes associated with transporters, metabolism, cell wall biogenesis. This study recommends for possible association between lncRNA expression and C. auris pathogenesis.