Similar evolutionary trajectories in an environmental Cryptococcus neoformans isolate after human and murine infection.

A pet cockatoo was the suspected source of Cryptococcus neoformans recovered from an immunocompromised patient with cryptococcosis based on molecular analyses available in 2000. Here, we report whole genome sequence analysis of the clinical and cockatoo strains. Both are closely related MATα strains belonging to the VNII lineage, confirming that the human infection likely originated from pet bird exposure. The two strains differ by 61 single nucleotide polymorphisms, including eight nonsynonymous changes involving seven genes. To ascertain whether changes in these genes are selected for during mammalian infection, we passaged the cockatoo strain in mice. Remarkably, isolates obtained from mouse tissue possess a frameshift mutation in one of the seven genes altered in the human sample (LQVO5_000317), a gene predicted to encode an SWI-SNF chromatin-remodeling complex protein. In addition, both cockatoo and patient strains as well as mouse-passaged isolates obtained from brain tissue had a premature stop codon in a homologue of ZFC3 (LQVO5_004463), a predicted single-zinc finger containing protein, which is associated with larger capsules when deleted and reverted to a full-length protein in the mouse-passaged isolates obtained from lung tissue. The patient strain and mouse-passaged isolates show variability in virulence factors, with differences in capsule size, melanization, rates of nonlytic expulsion from macrophages, and amoeba predation resistance. Our results establish that environmental strains undergo genomic and phenotypic changes during mammalian passage, suggesting that animal virulence can be a mechanism for genetic change and that the genomes of clinical isolates may provide a readout of mutations acquired during infection.

The yeast genus Tardiomyces gen. nov. with one new species and two new combinations.

PURPOSE: Rare yeasts species are increasingly reported as causative agents of invasive human infection. Proper identification and antifungal therapy are essential to manage these infections. Candida blankii is one of these emerging pathogens and is known for its reduced susceptibility to multiple antifungals. METHODS: To obtain more insight into the characteristics of this species, 26 isolates reported as C. blankii were investigated using genetic and phenotypical approaches. RESULTS: Among the 26 isolates, seven recovered either from blood, sputum, urine, or the oral cavity, displayed substantial genetic and some phenotypical differences compared to the other isolates, which were confirmed as C. blankii. We consider these seven strains to represent a novel species, Tardiomyces depauwii. Phylogenomics assigned C. blankii, C. digboiensis, and the novel species in a distinct branch within the order Dipodascales, for which the novel genus Tardiomyces is erected. The new combinations Tardiomyces blankii and Tardiomyces digboiensis are introduced. Differences with related, strictly environmental genera Sugiyamaella, Crinitomyces, and Diddensiella are enumerated. All three Tardiomyces species share the rare ability to grow up to 42 °C, display slower growth in nutrient-poor media, and show a reduced susceptibility to azoles and echinocandins. Characteristics of T. depauwii include high MIC values with voriconazole and a unique protein pattern. CONCLUSION: We propose the novel yeast species Tardiomyces depauwii and the transfer of C. blankii and C. digboiensis to the novel Tardiomyces genus.

Differential lipase virulence in Malassezia furfur dimorphism isolated from pityriasis versicolor patients and healthy individuals.

BACKGROUND: Malassezia furfur is a member of the human skin microbiomes that can cause various skin diseases. Dimorphism plays a role as the yeast phase predominates during skin colonisation whereas mycelial forms are observed in the scales of patients with pityriasis versicolor (PV). However, due to their condition-dependence for growth, it is difficult to culture M. furfur and this is an additional challenge for studying the pathogenicity of this fungus. OBJECTIVE: To describe different media suitable for culturing Malassezia from the yeast phase into mycelial forms, with a particular focus on nutritional supplements and pH conditions. METHODS: Clinical M. furfur isolates from patients with PV and healthy individuals were used to investigate Malassezia dimorphism as well as the activity and expression of lipase enzymes. RESULTS: Our experimental media were significantly more likely to promote mycelial growth in strains from healthy individuals compared to those from patients with PV. Lipase activity was increased in the mycelial phase cells compared to yeast forms for all strains tested. Assessment of the relative transcriptional expression of lipase within M. furfur revealed that LIP-coding genes were upregulated in mycelium relative to yeast forms for the strains tested. However, the increases in LIP3, LIP5 and LIP6 gene expressions were significantly greater in strains from healthy individuals compared to those from patients with PV. CONCLUSION: Overall, this study validated effective growth conditions to study M. furfur virulence factors and demonstrated that lipase is associated with M. furfur dimorphism.

Methamphetamine Enhances Cryptococcus neoformans Melanization, Antifungal Resistance, and Pathogenesis in a Murine Model of Drug Administration and Systemic Infection.

Methamphetamine (METH) is a major public health and safety problem in the United States. Chronic METH abuse is associated with a 2-fold-higher risk of HIV infection and, possibly, additional infections, particularly those that enter through the respiratory tract or skin. Cryptococcus neoformans is an encapsulated opportunistic yeast-like fungus that is a relatively frequent cause of meningoencephalitis in immunocompromised patients, especially in individuals with AIDS. C. neoformans melanizes during mammalian infection in a process that presumably uses host-supplied compounds such as catecholamines. l-3,4-Dihydroxyphenylalanine (l-Dopa) is a natural catecholamine that is frequently used to induce melanization in C. neoformans. l-Dopa-melanized cryptococci manifest resistance to radiation, phagocytosis, detergents, and heavy metals. Using a systemic mouse model of infection and in vitro assays to critically assess the impact of METH on C. neoformans melanization and pathogenesis, we demonstrated that METH-treated mice infected with melanized yeast cells showed increased fungal burdens in the blood and brain, exacerbating mortality. Interestingly, analyses of cultures of METH-exposed cryptococci supplemented with l-Dopa revealed that METH accelerates fungal melanization, an event of adaptation to external stimuli that can be advantageous to the fungus during pathogenesis. Our findings provide novel evidence of the impact of METH abuse on host homeostasis and increased permissiveness to opportunistic microorganisms.

Development of an Interferon-Gamma Release Assay (IGRA) to Aid Diagnosis of Histoplasmosis.

Establishing diagnosis of latent and active histoplasmosis is challenging. Interferon gamma-release assays (IGRAs) may provide evidence of latent and active infection. An enzyme-linked immunospot (ELISpot) assay was developed using yeast cell lysate (YCL) antigen prepared from a representative North American Histoplasma capsulatum strain. Assay parameters were optimized by measuring responses in healthy volunteers with and without Histoplasma infection. Assay performance as an aid for diagnosing histoplasmosis was assessed in a prospective cohort of 88 people with suspected or confirmed infection, and 44 healthy controls enrolled in two centers in North America (2013 to 2018). Antigen specificity of IFN-γ release was demonstrated using ELISpot and enzyme-linked immunosorbent assay (ELISA). Antigen-evoked, single-cell mRNA expression by memory T cells was shown using flow cytometry. The area under the receiver operating characteristic curve (AUC) was estimated at 0.89 (95% confidence interval [CI]: 78.5% to 99.9%). At optimal cutoff, sensitivity was 77.2% (95% CI: 54.6% to 92.2%) and specificity was 100% (95% CI: 89.7% to 100%). Sixteen of 44 healthy volunteers (36.4%) from a region of hyperendemicity had positive responses, suggesting detection of previously unrecognized (latent) infection. The ELISpot assay is sensitive and specific as an aid to diagnose H. capsulatum infection and disease, supporting proof of concept and further development.

Host cell membrane microdomains and fungal infection.

Lipid microdomains or lipid rafts are dynamic and tightly ordered regions of the plasma membrane. In mammalian cells, they are enriched in cholesterol, glycosphingolipids, Glycosylphosphatidylinositol-anchored and signalling-related proteins. Several studies have suggested that mammalian pattern recognition receptors are concentrated or recruited to lipid domains during host-pathogen association to enhance the effectiveness of host effector processes. However, pathogens have also evolved strategies to exploit these domains to invade cells and survive. In fungal organisms, a complex cell wall network usually mediates the first contact with the host cells. This cell wall may contain virulence factors that interfere with the host membrane microdomains dynamics, potentially impacting the infection outcome. Indeed, the microdomain disruption can dampen fungus-host cell adhesion, phagocytosis and cellular immune responses. Here, we provide an overview of regulatory strategies employed by pathogenic fungi to engage with and potentially subvert the lipid microdomains of host cells. TAKE AWAY: Lipid microdomains are ordered regions of the plasma membrane enriched in cholesterol, glycosphingolipids (GSL), GPI-anchored and signalling-related proteins. Pathogen recognition by host immune cells can involve lipid microdomain participation. During this process, these domains can coalesce in larger complexes recruiting receptors and signalling proteins, significantly increasing their signalling abilities. The antifungal innate immune response is mediated by the engagement of pathogen-associated molecular patterns to pattern recognition receptors (PRRs) at the plasma membrane of innate immune cells. Lipid microdomains can concentrate or recruit PRRs during host cell-fungi association through a multi-interactive mechanism. This association can enhance the effectiveness of host effector processes. However, virulence factors at the fungal cell surface and extracellular vesicles can re-assembly these domains, compromising the downstream signalling and favouring the disease development. Lipid microdomains are therefore very attractive targets for novel drugs to combat fungal infections.

Identification of four compounds from the Pharmakon library with antifungal activity against Candida auris and species of Cryptococcus.

There is an urgent need to develop novel antifungals. In this study, we screened 1600 compounds for antifungal activity against Cryptococcus neoformans and Candida auris. We evaluated 4 promising compounds against 24 additional isolates of Cr. neoformans, Ca. auris, Cr. deuterogattii, and Cr. gattii. The four compounds, dequalinium chloride (DQC), bleomycin sulfate (BMS), pentamidine isethionate salt (PIS), and clioquinol (CLQ), varied in their efficacy against these pathogens but were generally more effective against cryptococci. The compounds exerted their antifungal effect via multiple mechanisms, including interference with the capsule of cryptococci and induction of hyphal-like morphology in Ca. auris. Our results indicate that DQC, BMS, PIS, and CLQ represent potential prototypes for the future development of antifungals. LAY SUMMARY: Fungal infections can be lethal and the options to fight them are scarce. We tested 1600 molecules for their ability to control the growth of two important fungal pathogens, namely Candida auris and species of Cryptococcus. Four of these compounds showed promising antifungal activities.

Media matters! Alterations in the loading and release of Histoplasma capsulatum extracellular vesicles in response to different nutritional milieus.

Histoplasma capsulatum is a dimorphic fungus that most frequently causes pneumonia, but can also disseminate and proliferate in diverse tissues. Histoplasma capsulatum has a complex secretion system that mediates the release of macromolecule-degrading enzymes and virulence factors. The formation and release of extracellular vesicles (EVs) are an important mechanism for non-conventional secretion in both ascomycetes and basidiomycetes. Histoplasma capsulatum EVs contain diverse proteins associated with virulence and are immunologically active. Despite the growing knowledge of EVs from H. capsulatum and other pathogenic fungi, the extent that changes in the environment impact the sorting of organic molecules in EVs has not been investigated. In this study, we cultivated H. capsulatum with distinct culture media to investigate the potential plasticity in EV loading in response to differences in nutrition. Our findings reveal that nutrition plays an important role in EV loading and formation, which may translate into differences in biological activities of these fungi in various fluids and tissues.

Candida parapsilosis: from Genes to the Bedside.

Patients with suppressed immunity are at the highest risk for hospital-acquired infections. Among these, invasive candidiasis is the most prevalent systemic fungal nosocomial infection. Over recent decades, the combined prevalence of non-albicans Candida species outranked Candida albicans infections in several geographical regions worldwide, highlighting the need to understand their pathobiology in order to develop effective treatment and to prevent future outbreaks. Candida parapsilosis is the second or third most frequently isolated Candida species from patients. Besides being highly prevalent, its biology differs markedly from that of C. albicans, which may be associated with C. parapsilosis' increased incidence. Differences in virulence, regulatory and antifungal drug resistance mechanisms, and the patient groups at risk indicate that conclusions drawn from C. albicans pathobiology cannot be simply extrapolated to C. parapsilosis Such species-specific characteristics may also influence their recognition and elimination by the host and the efficacy of antifungal drugs. Due to the availability of high-throughput, state-of-the-art experimental tools and molecular genetic methods adapted to C. parapsilosis, genome and transcriptome studies are now available that greatly contribute to our understanding of what makes this species a threat. In this review, we summarize 10 years of findings on C. parapsilosis pathogenesis, including the species' genetic properties, transcriptome studies, host responses, and molecular mechanisms of virulence. Antifungal susceptibility studies and clinician perspectives are discussed. We also present regional incidence reports in order to provide an updated worldwide epidemiology summary.

Histoplasma Capsulatum: Mechanisms for Pathogenesis.

Histoplasmosis, caused by the dimorphic environmental fungus Histoplasma capsulatum, is a major mycosis on the global stage. Acquisition of the fungus by mammalian hosts can be clinically silent or it can lead to life-threatening systemic disease, which can occur in immunologically intact or deficient hosts, albeit severe disease is more likely in the setting of compromised cellular immunity. H. capsulatum yeast cells are highly adapted to the mammalian host as they can effectively survive within intracellular niches in select phagocytic cells. Understanding the biological response by both the host and H. capsulatum will facilitate improved approaches to prevent and/or modify disease. This review presents our current understanding of the major pathogenic mechanisms involved in histoplasmosis.

Host membrane glycosphingolipids and lipid microdomains facilitate Histoplasma capsulatum internalisation by macrophages.

Recognition and internalisation of intracellular pathogens by host cells is a multifactorial process, involving both stable and transient interactions. The plasticity of the host cell plasma membrane is fundamental in this infectious process. Here, the participation of macrophage lipid microdomains during adhesion and internalisation of the fungal pathogen Histoplasma capsulatum (Hc) was investigated. An increase in membrane lateral organisation, which is a characteristic of lipid microdomains, was observed during the first steps of Hc-macrophage interaction. Cholesterol enrichment in macrophage membranes around Hc contact regions and reduced levels of Hc-macrophage association after cholesterol removal also suggested the participation of lipid microdomains during Hc-macrophage interaction. Using optical tweezers to study cell-to-cell interactions, we showed that cholesterol depletion increased the time required for Hc adhesion. Additionally, fungal internalisation was significantly reduced under these conditions. Moreover, macrophages treated with the ceramide-glucosyltransferase inhibitor (P4r) and macrophages with altered ganglioside synthesis (from B4galnt1-/- mice) showed a deficient ability to interact with Hc. Coincubation of oligo-GM1 and treatment with Cholera toxin Subunit B, which recognises the ganglioside GM1, also reduced Hc association. Although purified GM1 did not alter Hc binding, treatment with P4 significantly increased the time required for Hc binding to macrophages. The content of CD18 was displaced from lipid microdomains in B4galnt1-/- macrophages. In addition, macrophages with reduced CD18 expression (CD18low ) were associated with Hc at levels similar to wild-type cells. Finally, CD11b and CD18 colocalised with GM1 during Hc-macrophage interaction. Our results indicate that lipid rafts and particularly complex gangliosides that reside in lipid rafts stabilise Hc-macrophage adhesion and mediate efficient internalisation during histoplasmosis.

Cryptococcus neoformans Secretes Small Molecules That Inhibit IL-1ß Inflammasome-Dependent Secretion.

Cryptococcus neoformans is an encapsulated yeast that causes disease mainly in immunosuppressed hosts. It is considered a facultative intracellular pathogen because of its capacity to survive and replicate inside phagocytes, especially macrophages. This ability is heavily dependent on various virulence factors, particularly the glucuronoxylomannan (GXM) component of the polysaccharide capsule. Inflammasome activation in phagocytes is usually protective against fungal infections, including cryptococcosis. Nevertheless, recognition of C. neoformans by inflammasome receptors requires specific changes in morphology or the opsonization of the yeast, impairing proper inflammasome function. In this context, we analyzed the impact of molecules secreted by C. neoformans B3501 strain and its acapsular mutant Δcap67 in inflammasome activation in an in vitro model. Our results showed that conditioned media derived from B3501 was capable of inhibiting inflammasome-dependent events (i.e., IL-1ß secretion and LDH release via pyroptosis) more strongly than conditioned media from Δcap67, regardless of GXM presence. We also demonstrated that macrophages treated with conditioned media were less responsive against infection with the virulent strain H99, exhibiting lower rates of phagocytosis, increased fungal burdens, and enhanced vomocytosis. Moreover, we showed that the aromatic metabolite DL-Indole-3-lactic acid (ILA) and DL-p-Hydroxyphenyllactic acid (HPLA) were present in B3501's conditioned media and that ILA alone or with HPLA is involved in the regulation of inflammasome activation by C. neoformans. These results were confirmed by in vivo experiments, where exposure to conditioned media led to higher fungal burdens in Acanthamoeba castellanii culture as well as in higher fungal loads in the lungs of infected mice. Overall, the results presented show that conditioned media from a wild-type strain can inhibit a vital recognition pathway and subsequent fungicidal functions of macrophages, contributing to fungal survival in vitro and in vivo and suggesting that secretion of aromatic metabolites, such as ILA, during cryptococcal infections fundamentally impacts pathogenesis.

The putative flippase Apt1 is required for intracellular membrane architecture and biosynthesis of polysaccharide and lipids in Cryptococcus neoformans.

Flippases are responsible for the asymmetric distribution of phospholipids in biological membranes. In the encapsulated fungal pathogen Cryptococcus neoformans, the putative flippase Apt1 is an important regulator of polysaccharide secretion and pathogenesis in mice by unknown mechanisms. In this study, we analyzed the role of C. neoformans Apt1 in intracellular membrane architecture and synthesis of polysaccharide and lipids. Analysis of wild type (WT), apt1Δ (mutant) and apt1Δ::APT1 (complemented) strains by transmission electron microscopy revealed that deletion of APT1 resulted in the formation of irregular vacuoles. Disorganization of vacuolar membranes in apt1Δ cells was accompanied by a significant increase in the amounts of intra-vacuolar and pigment-containing vesicles. Quantitative immunogold labeling of C. neoformans cells with a monoclonal antibody raised to a major capsular component suggested impaired polysaccharide synthesis. APT1 deletion also affected synthesis of phosphatidylserine, phosphatidylethanolamine, inositolphosphoryl ceramide, glucosylceramide and ergosterylglycoside. These results reveal novel functions of Apt1 and are in agreement with the notion that this putative flippase plays an important role in the physiology of C. neoformans.

Anti-biofilm activity of garlic extract loaded nanoparticles.

The emergence and widespread distribution of multi-drug resistant bacteria are considered as a major public health concern. The inabilities to curb severe infections due to antibiotic resistance have increased healthcare costs as well as patient morbidity and mortality. Bacterial biofilms formed by drug-resistant bacteria add additional challenges to treatment. This study describes a solgel based nanoparticle system loaded with garlic extract (GE-np) that exhibits: i) slow and sustained release of garlic components; ii) stabilization of the active components; and iii) significant enhancement of antimicrobial and antibiofilm activity relative to the free garlic extract. Also, GE-np were efficient in penetrating and disrupting the well-established methicillin-resistant Staphylococcus aureus (MRSA) biofilms. Overall, the study suggests that GE-np might be a promising candidate for the treatment of chronic infections due to biofilm forming drug-resistant bacteria.

A case of sporotrichosis caused by different Sporothrix brasiliensis strains: mycological, molecular, and virulence analyses.

BACKGROUND: Sporotrichosis is a subcutaneous mycosis caused by dimorphic pathogenic fungi belonging to the Sporothrix genus. Pathogenic Sporothrix species typically produce melanin, which is known to be a virulence factor. OBJECTIVES: The aim of this study was to perform phenotypic, genotypic, and virulence analyses of two distinct Sporothrix brasiliensis strains isolated from the same lesion on a patient from Rio de Janeiro. METHODS AND FINDINGS: Genotypic analyses by partial sequencing of the calmodulin, ß-tubulin, and chitin synthase genes, as well as polymerase chain reaction (PCR)-fingerprinting by T3B, M13, and GACA, showed that the isolates were very similar but not identical. Both isolates had similar phenotypic characteristics and effectively produced melanin in their yeast forms, accounting for their ability of causing disease in a murine sporotrichosis model. Remarkably, isolate B was albino in its environmental form but caused more severe disease than the pigmented A isolate. CONCLUSIONS: These findings indicate that the patient was infected by two genetically and biologically distinct S. brasiliensis that vary in their production of melanin in their environmental forms. The results underscore the importance of characterizing phenotypically different isolates found in the same clinical specimen or patient.

Production of melanin pigments in saprophytic fungi in vitro and during infection.

Melanins are one of the great natural pigments produced by a wide variety of fungal species that promote fitness and cell survival in diverse hostile environments, including during mammalian infection. In this study, we sought to demonstrate the production of melanin in the conidia and hyphae of saprophytic fungi, including dematiaceous and hyaline fungi. We showed that a melanin-specific monoclonal antibody (MAb) avidly labeled the cell walls of hyphae and conidia, consistent with the presence of melanin in these structures, in 14 diverse fungal species. The conidia of saprophytic fungi were treated with proteolytic enzymes, denaturant, and concentrated hot acid to yield dark particles, which were shown to be stable free radicals, consistent with their identification as melanins. Samples obtained from patients with fungal keratitis due to Fusarium falciforme, Aspergillus fumigatus, Aspergillus flavus, Curvularia lunata, Exserohilum rostratum, or Fonsecaea pedrosoi were found to be intensely labeled by the melanin-specific MAb at the fungal hyphal cell walls. These results support the hypothesis that melanin is a common component that promotes survival under harsh conditions and facilitates fungal virulence. Increased understanding of the processes of melanization and the development of methods to interfere with pigment formation may lead to novel approaches to combat these complex pathogens that are associated with high rates of morbidity and mortality.

Nitric Oxide Releasing Nanoparticles as a Strategy to Improve Current Onychomycosis Treatments.

Topical antimicrobials are the ideal mode of onychomycosis treatment for efficient drug delivery and avoidance of sytemic effects associated with oral medications. However, high treatment costs, tissue penetration limitations, and low cure rates have continued to pose major challenges. To capitalize on the progress made by topical efinaconazole solution, efinaconazole was combined with inexpensive, previously-characterized nitric oxide releasing nanoparticles (NO-np), which have been shown to offer sustained nitric oxide release over time and enhanced barrier penetration, while exerting broad spectrum antimicrobial and immunomodulating properties. NO-np were combined with efinaconazole in varying concentrations and applied against reference strains of Trichophyton rubrum using a checkerboard method. Results demonstrated synergism of NO-np+efinaconazole against T. rubrum, which is noteworthy given the barriers present in the topical treatment of onychomycosis, and the multiple potential benefits offered by NO-np. Overall, this study illustrates the untapped potential of nanotechnology in the treatment of disorders of the skin, hair, and nails where drug delivery remains a challenge. J Drugs Dermatol. 2018;17(7):717-720.

Sustained Nitric Oxide-Releasing Nanoparticles Interfere with Methicillin-Resistant Staphylococcus aureus Adhesion and Biofilm Formation in a Rat Central Venous Catheter Model.

Staphylococcus aureus is frequently isolated in the setting of infections of indwelling medical devices, which are mediated by the microbe's ability to form biofilms on a variety of surfaces. Biofilm-embedded bacteria are more resistant to antimicrobial agents than their planktonic counterparts and often cause chronic infections and sepsis, particularly in patients with prolonged hospitalizations. In this study, we demonstrate that sustained nitric oxide-releasing nanoparticles (NO-np) interfere with S. aureus adhesion and prevent biofilm formation on a rat central venous catheter (CVC) model of infection. Confocal and scanning electron microscopy showed that NO-np-treated staphylococcal biofilms displayed considerably reduced thicknesses and bacterial numbers compared to those of control biofilms in vitro and in vivo, respectively. Although both phenotypes, planktonic and biofilm-associated staphylococci, of multiple clinical strains were susceptible to NO-np, bacteria within biofilms were more resistant to killing than their planktonic counterparts. Furthermore, chitosan, a biopolymer found in the exoskeleton of crustaceans and structurally integrated into the nanoparticles, seems to add considerable antimicrobial activity to the technology. Our findings suggest promising development and translational potential of NO-np for use as a prophylactic or therapeutic against bacterial biofilms on CVCs and other medical devices.

Topical nitric oxide releasing nanoparticles are effective in a murine model of dermal Trichophyton rubrum dermatophytosis.

Systemic therapies are preferred for treating dermal dermatophytosis due to inadequate penetration of topical agents. However, systemic antifungals are associated with off-target effects and limited tissue penetration, and antimicrobial resistance is a growing concern. To address this, we investigated topical nitric oxide-releasing nanoparticles (NO-np), which have been used against superficial fungal infections and bacterial abscesses. In addition to enhanced penetration and permeation conferred by nanoparticles, nitric oxide, a broad-spectrum multi-mechanistic antimicrobial agent, offers decreased likelihood of resistance development. In the current study, NO-np inhibited Trichophyton rubrum in vitro, as well as in a murine model of dermal dermatophytosis. In mice, NO-np reduced fungal burden after three days, with complete clearance after seven. Furthermore, NO-np decreased tissue IL-2, 6, 10 and TNFα, indicating earlier attenuation of the host inflammatory response and decreased tissue morbidity. Thus, topical NO-np represent an attractive alternative to systemic therapy against dermal T. rubrum infection.