Chromoblastomycosis Caused by Fonsecaea monophora Mimicking Lichen Planus.

Chromoblastomycosis is a rare fungal infection acquired by traumatic inoculation of pigmented fungi from an environmental source. The polymorphic presentation of chromoblastomycosis may mimic other dermatologic conditions, leading to delays in diagnosis. Thus, histopathology is critical in identifying the presence of fungi and confirming the diagnosis. We present a case of chromoblastomycosis caused by the organism Fonsecaea monophora mimicking a lesion of lichen planus to highlight the importance of histopathology in the diagnosis of this condition.

Comparison of the antifungal activity of the pyrimidine analogs flucytosine and carmofur against human-pathogenic dematiaceous fungi.

Chromoblastomycosis (CBM) and pheohyphomycosis (PHM) are the most common implantation mycoses caused by dematiaceous fungi. In the past, flucytosine (5-FC) has been used to treat CBM, but development of resistance is common. Carmofur belongs to the same class as 5-FC and has in vitro inhibitory activity against the main agents of CBM and PHM. The aim of this study was to compare the action of these two pyrimidine analog drugs against CBM and PHM agents. The minimum inhibitory concentration (MIC) and the selectivity index based on cytotoxicity tests of these two drugs against some agents of these mycoses were determined, with carmofur presenting a higher selectivity index than 5-FC. Carmofur demonstrated here synergistic interactions with itraconazole and amphotericin B against Exophiala heteromorpha, Fonsecaea pedrosoi, Fonsecaea monophora, and Fonsecaea nubica strains. Additionally, carmofur plus itraconazole demonstrated here synergism against a Phialophora verrucosa strain. To evaluate the development of carmofur resistance, passages in culture medium containing subinhibitory concentrations of this pyrimidine analog were carried out, followed by in vitro susceptibility tests. Exophiala dermatitidis quickly developed resistance, whereas F. pedrosoi took seven passages in carmofur-supplemented medium to develop resistance. Moreover, resistance was permanent in E. dermatitidis but transient in F. pedrosoi. Hence, carmofur has exhibited certain advantages, albeit accompanied by limitations such as the development of resistance, which was expected as with 5-FC. This underscores its therapeutic potential in combination with other drugs, emphasizing the need for a meticulous evaluation of its application in the fight against dematiaceous fungi.

Chromoblastomycosis in French Guiana: Epidemiology and Practices, 1955-2023.

Chromoblastomycosis (CBM) is a chronic neglected fungal disease, usually met in tropical areas. French Guiana is a South American territory with limited epidemiological data. This retrospective study concerned all patients with CBM proven by at least one paraclinical examination and diagnosed in French Guiana between 1950 and 2023. In total, 23 patients were included, mostly males (87%) of Creole origin, living in the coastal region (87%) and involved in outdoor occupations (74%). Lesions were mostly observed on the lower limbs (78.3%), with a median time to diagnosis of four years. Laboratory tests included positive direct microscopic examinations (78.3%) and mycological cultures (69.6%), identifying 14 cases of Fonsecaea pedrosoi and one case of Exophiala janselmei. Various treatments were employed, including antifungals, surgery and combinations of both. In conclusion, CBM in French Guiana involves a different population than other subcutaneous mycoses such as Lobomycosis or Paracoccidioidomycosis, mostly found in the forest hinterland. Surgery should be recommended for recent and limited lesions. Itraconazole and terbinafine should systematically be proposed, either in monotherapy or in combination with surgery or cryotherapy.

Galleria mellonella in vitro model for chromoblastomycosis shows large differences in virulence between isolates.

BACKGROUND: Chromoblastomycosis is the World Health Organization (WHO)-recognized fungal implantation disease that eventually leads to severe mutilation. Cladophialophora carrionii (C. carrionii) is one of the agents. However, the pathogenesis of C. carrionii is not fully investigated yet. METHODS: We investigated the pathogenic potential of the fungus in a Galleria mellonella (G. mellonella) larvae infection model. Six strains of C. carrionii, and three of its environmental relative C. yegresii were tested. The G. mellonella model was also applied to determine antifungal efficacy of amphotericin B, itraconazole, voriconazole, posaconazole, and terbinafine. RESULTS: All strains were able to infect the larvae, but virulence potentials were strain-specific and showed no correlation with clinical background of the respective isolate. Survival of larvae also varied with infection dose, and with growth speed and melanization of the fungus. Posaconazole and voriconazole exhibited best activity against Cladophialophora, followed by itraconazole and terbinafine, while limited efficacy was seen for amphotericin B. CONCLUSION: Infection behavior deviates significantly between strains. In vitro antifungal susceptibility of tested strains only partly explained the limited treatment efficacy in vivo.

Resazurin to determine the minimum inhibitory concentration on antifungal susceptibility assays for Fonsecaea sp. using a modified EUCAST protocol.

Chromoblastomycosis is a fungal chronic disease, which affects humans, especially in cutaneous and subcutaneous tissues. There is no standard treatment for Chromoblastomycosis, and it is a therapeutic challenge, due natural resistance of their causative agents, inadequate response of patients and common cases of relapse. Protocols for determination of antifungal drugs susceptibility are not standardized for chromoblastomycosis agents and endpoint definition is usually based on visual inspection, which depends on the analyst, making it sometimes inaccurate. We presented a colorimetric and quantitative methodology based on resazurin reduction to resofurin to determine the metabolic status of viable cells of Fonsecaea sp. Performing antifungal susceptibility assay by a modified EUCAST protocol allied to resazurin, we validated the method to identify the minimum inhibitory concentrations of itraconazole, fluconazole, amphotericin B, and terbinafine for eight Fonsecaea clinical isolates. According to our data, resazurin is a good indicator of metabolic status of viable cells, including those exposed to antifungal drugs. This work aimed to test resazurin as an indicator of the metabolic activity of Fonsecaea species in susceptibility assays to antifungal drugs. Species of this genus are the main causative agents of Chromoblastomycosis, which affects humans.

Neutrophil extracellular traps (NETs) and Th-2 dominant immune responses in chronic granulomatous chromobalstomycosis.

Chromoblastomycosis (CBM), a chronic, granulomatous, suppurative mycosis of the skin and subcutaneous tissue, is caused by several dematiaceous fungi. The formation of granulomas, tissue proliferation, and fibrosis in response to these pathogenic fungi is believed to be intricately linked to host immunity. To understand this complex interaction, we conducted a comprehensive analysis of immune cell infiltrates, neutrophil extracellular traps (NETs) formation, and the fibrosis mechanism in 20 CBM lesion biopsies using immunohistochemical and immunofluorescence staining methods. The results revealed a significant infiltration of mixed inflammatory cells in CBM granulomas, prominently featuring a substantial presence of Th2 cells and M2 macrophages. These cells appeared to contribute to the production of collagen I and III in the late fibrosis mechanism, as well as NETs formation. The abundance of Th2 cytokines may act as a factor promoting the bias of macrophage differentiation toward M2, which hinders efficient fungal clearance while accelerates the proliferation of fibrous tissue. Furthermore, the expression of IL-17 was noted to recruit neutrophils, facilitating subsequent NETs formation within CBM granulomas to impede the spread of sclerotic cells. Understanding of these immune mechanisms holds promise for identifying therapeutic targets for managing chronic granulomatous CBM.

Zinc Starvation Induces Cell Wall Remodeling and Activates the Antioxidant Defense System in Fonsecaea pedrosoi.

The survival of pathogenic fungi in the host after invasion depends on their ability to obtain nutrients, which include the transition metal zinc. This essential micronutrient is required to maintain the structure and function of various proteins and, therefore, plays a critical role in various biological processes. The host's nutritional immunity limits the availability of zinc to pathogenic fungi mainly by the action of calprotectin, a component of neutrophil extracellular traps. Here we investigated the adaptive responses of Fonsecaea pedrosoi to zinc-limiting conditions. This black fungus is the main etiological agent of chromoblastomycosis, a chronic neglected tropical disease that affects subcutaneous tissues. Following exposure to a zinc-limited environment, F. pedrosoi induces a high-affinity zinc uptake machinery, composed of zinc transporters and the zincophore Pra1. A proteomic approach was used to define proteins regulated by zinc deprivation. Cell wall remodeling, changes in neutral lipids homeostasis, and activation of the antioxidant system were the main strategies for survival in the hostile environment. Furthermore, the downregulation of enzymes required for sulfate assimilation was evident. Together, the adaptive responses allow fungal growth and development and reveals molecules that may be related to fungal persistence in the host.

Fonsecaea pedrosoi produces ferricrocin and can utilize different host iron sources.

The survival of living organisms depends on iron, one of the most abundant metals in the Earth's crust. Nevertheless, this micronutrient is poorly available in our aerobic atmosphere as well as inside the mammalian host. This problem is circumvented by the expression of high affinity iron uptake machineries, including the production of siderophores, in pathogenic fungi. Here we demonstrated that F. pedrosoi, the causative agent of the neglected tropical disease chromoblastomycosis, presents gene clusters for siderophore production. In addition, ten putative siderophore transporters were identified. Those genes are upregulated under iron starvation, a condition that induces the secretion of hydroxamates, as revealed by chrome azurol S assays. RP-HPLC and mass spectrometry analysis allowed the identification of ferricrocin as an intra- and extracellular siderophore. F. pedrosoi can grow in different iron sources, including the bacterial ferrioxamine B and the host proteins ferritin, hemoglobin and holotransferrin. Of note, addition of hemoglobin, lactoferrin and holotransferrin to the growth medium of macrophages infected with F. pedrosoi enhanced significantly fungal survival. The ability to produce siderophores in iron limited conditions added to the versatility to utilize different sources of iron are strategies that certainly may contribute to fungal survival inside the host.

New Immunological Markers in Chromoblastomycosis-The Importance of PD-1 and PD-L1 Molecules in Human Infection.

The pathogenesis of chromoblastomycosis (CBM) is associated with Th2 and/or T regulatory immune responses, while resistance is associated with a Th1 response. However, even in the presence of IFN-γ, fungi persist in the lesions, and the reason for this persistence is unknown. To clarify the factors associated with pathogenesis, this study aimed to determine the polarization of the cellular immune response and the densities of cells that express markers of exhaustion in the skin of CBM patients. In the skin of patients with CBM, a moderate inflammatory infiltrate was observed, characterized primarily by the occurrence of histiocytes. Analysis of fungal density allowed us to divide patients into groups that exhibited low and high fungal densities; however, the intensity of the inflammatory response was not related to mycotic loads. Furthermore, patients with CBM exhibited a significant increase in the number of CD4+ and CD8+ cells associated with a high density of IL-10-, IL-17-, and IFN-γ-producing cells, indicating the presence of a chronic and mixed cellular immune response, which was also independent of fungal load. A significant increase in the number of PD-1+ and PD-L1+ cells was observed, which may be associated with the maintenance of the fungus in the skin and the progression of the disease.

From Child to Old Man: A Slowly Evolving Case of Chromoblastomycosis Caused by Cladosporium cladosporioides.

Chromoblastomycosis is a chronic granulomatous mycosis of the skin and subcutaneous tissue caused by traumatic inoculation with dematiaceous fungi. This disease primarily affects agricultural workers, who are mostly men. We present a case of chromoblastomycosis in a 63-year-old male farmer patient with dermatosis over 50 years of evolution, with warty, erythematous, and scaly plaques that predominate on the left hemithorax. Direct examination with potassium hydroxide (KOH) revealed numerous fumagoid cells. Amplification and sequencing of the internal transcribed spacer (ITS) and translation elongation factor 1-alpha (TEF-1a) gene revealed that chromoblastomycosis was caused by Cladosporium cladosporioides. The chromoblastomycosis was treated with itraconazole and fluconazole without any improvement, and amphotericin B was administered with partial improvement.

Chromoblastomycosis: New Perspective on Adjuvant Treatment with Acitretin.

Chromoblastomycosis (CBM) is a neglected human disease, caused by different species of pigmented dematiaceous fungi that cause granulomatous and suppurative dermatosis. This infection is difficult to treat and there are limited therapeutic options, including terbinafine, itraconazole, and tioconazole. Classic treatment is administered for a long period of time, but some patients do not respond properly, and therefore, such therapeutic approaches possess low cure rates. Therefore, it is vital to develop new strategies for the treatment of CBM. In this regard, it has been observed that the association of immunomodulatory molecules such as glucan with therapy carried out with antifungal drugs improves cutaneous lesions in comparison to treatment with antifungal drugs alone, suggesting that drug association may be an interesting and significant approach to incorporate into CBM therapy. Thus, the aim of this work was to associate classical antifungal therapy with the adjuvants imiquimod and acitretin. In the present case, we reported a patient with extensive CBM caused by Fonsaecae pedrosoi, that affected an extensive area of the right leg, that was left without treatment for 11 years. He was treated with a classical combination of itraconazole and terbinafine via the oral route plus topical imiquimod and oral acitretin, as an adjuvant therapy. After five months of treatment, a significant regression of verrucous plaques was observed, suggesting that the use of these adjuvants combined with the classical antifungal drugs, intraconazole plus terbinafine, can reduce treatment time and rapidly improve the patient's quality of life. This result confirms that the use of coadjuvant drugs may be effective in the treatment of this infectious disease.

Chromoblastomycosis Presenting with Sporotrichoid Distribution and Bony Destruction: A Rare Presentation.

Chromoblastomycosis (CBM) presenting with linear distribution and with underlying bony destruction is rare. Herein, we report such a presentation in a farmer who presented with ulcerated nodules over the right leg and swelling of the right foot. Potassium hydroxide (KOH) preparation and histopathological examination of biopsy from nodule revealed characteristic sclerotic bodies on Gomori methenamine silver and periodic acid Schiff stain (PAS), which confirmed the diagnosis of chromoblastomycosis. X-ray of right foot revealed osteolytic destruction of right third metatarsophalangeal joint. Work-up for systemic involvement did not reveal any involvement. He was placed on combination therapy of itraconazole and terbinafine and is under follow-up.

Chromoblastomycosis: A Case Series and Literature Review.

Chromoblastomycosis is a subcutaneous mycosis caused by a variety of dematiaceous fungi. Fonsecaea (F.) pedrosoi is the most common causative agent. Majority of cases have been reported from tropical and subtropical regions with rural and agricultural background. It is a chronic disease with low incidence of complications but is very refractory to therapies. This is a case series of 22 cases of chromoblastomycosis from two health-care facilities in India. Information regarding the history, clinical presentations, diagnostic methods, therapy, and outcome of treatment were retrieved. Preponderance was seen among the males and in the age group of 41-60 years. Manual and agricultural laborers were commonly affected. Lower extremities were the most common sites affected. Morphological patterns like verrucous plaque, psoriasiform plaque, and verrucous nodules were seen. Direct microscopy with potassium hydroxide (KOH) mount was positive in all the cases. Histopathology in all cases displayed suppurative granulomatous inflammation with pigmented fungal cells. Fungal culture was positive in 10 cases with F. pedrosoi being the commonest agent. Antifungal treatment alone was instituted in 10 cases, cryotherapy along with antifungal therapy was given in 9 cases, and surgical excision was done in 3 cases. Complete clinical cure was achieved in seven cases. Chromoblastomycosis is characterized by chronicity, diverse clinical presentations, and therapeutic recalcitrance. Direct KOH mount of the black dots forms an important bedside tool in the diagnosis. Long-term antifungal therapy along with adjuvant cryotherapy may be the best option for the management.

Repurposing Benzimidazoles against Causative Agents of Chromoblastomycosis: Albendazole Has Superior In Vitro Activity Than Mebendazole and Thiabendazole.

Chromoblastomycosis (CBM) is a neglected human implantation mycosis caused by several dematiaceous fungal species. Currently available therapy is usually associated with physical methods, especially surgery, and with high refractoriness. Therefore, drug discovery for CBM is essential. Drug repositioning is a strategy used to facilitate the discovery of new treatments for several diseases. The aim of this study was to discover substances with antifungal activity against CBM agents from a collection of drugs previously approved for use in human diseases. A screening was performed with the NIH Clinical Collection against Fonsecaea pedrosoi. Ten substances, with clinical applicability in CBM, inhibited fungal growth by at least 60%. The minimum inhibitory concentration (MIC) of these substances was determined against other CBM agents, and the benzimidazoles albendazole, mebendazole and thiabendazole presented the lowest MIC values. The selectivity index, based on MIC and cytotoxicity of these substances, revealed albendazole to be more selective. To investigate a possible synergism of this benzimidazole with itraconazole and terbinafine, the chequerboard method was used. All interactions were classified as indifferent. Our current results suggest that benzimidazoles have repositioning potential against CBM agents. Albendazole seems to be the most promising, since it presented the highest selectivity against all dematiaceous fungi tested.

MALDI-TOF MS identification of Exophiala species isolated in Japan: Library enrichment and faster sample preparation.

Exophiala species cause chromoblastomycosis, mycetoma, and phaeohyphomycosis, which are occasionally fatally in immunocompromised patients. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) provides rapid and accurate examination of isolated bacteria and some fungal isolates, but the preparation method for filamentous fungi is complicated. In this study, 31 clinical isolates of Exophiala spp. in Japan were identified by MALDI-TOF MS with a library enriched by adding data. To simplify the sample preparation method, two modified methods were compared with the standard method for filamentous fungi. The agar cultivation sample preparation method reduced the time required for liquid culture and was considered suitable for clinical use. In 30 of 31 clinical isolates of Exophiala spp., the species identified by MALDI-TOF MS with the highest score matched the species identified by sequencing the internal transcribed spacer region. Exophiala dermatitidis, E. lecanii-corni, and E. oligosperma were identified above the genus level, while E. jeanselmei and E. xenobiotica were often not identified at the species level. The identification scores tended to be lower for less-registered strains in the in-house library. It is suggested that library enrichment and the modified preparation method may facilitate early diagnosis of rare fungal infections by Exophiala spp. in clinical laboratories using MALDI-TOF MS.

Cromoblastomicosis verrugosa-cicatricial multifocal: una entidad desatendida en Colombia

La cromoblastomicosis es una micosis subcutánea crónica de presentación clínica heterogénea que afecta principalmente a poblaciones de escasos recursos, lo que sumado al acceso limitado a los servicios de salud condiciona el retraso en el diagnóstico y tratamiento, ocasionando secuelas físicas graves. Se describe el caso de un hombre de 50 años con lesiones cutáneas verrugosas y cicatriciales de 30 años de evolución en la extremidad inferior y mano izquierda, además en cara en los últimos cinco años. Se realizó el diagnóstico de cromoblastomicosis de presentación multifocal y clínica mixta, con examen directo e histopatología compatibles, y cultivo para hongos positivo para Fonsecaea pedrosoi. Se indicó tratamiento oral con itraconazol y seguimiento clínico. Reconocer esta entidad es crucial para un diagnóstico y tratamiento oportunos, con el fin de evitar secuelas físicas y estigmatización secundaria. Se debe fortalecer en la atención primaria el reconocimiento de patologías desatendidas y de incidencia subestimada en Colombia, con una presentación multifocal mixta atípica y de un tiempo de evolución prolongado. El examen directo KOH es un herramienta accesible y económica en los primeros niveles de atención que puede contribuir al enfoque diagnóstico.

Chromoblastomycosis is a chronic subcutaneous mycosis with heterogeneous clinical presentation. It mainly affects low-income populations, which added to limited access to health services delays diagnosis and treatment, causing serious physical sequelae. We describe the case of a 50-year-old man with warty and cicatricial skin lesions of 30 years of evolution, at the level of left lower limb and left hand, with face involvement in the last five years. A diagnosis of chromoblastomycosis with multifocal and mixed clinical presentation was made, based on compatible direct examination and histopathology, and positive fungal culture for Fonsecaea pedrosoi. Oral treatment with itraconazole and clinical followup were stablished. Recognizing this entity is crucial for timely diagnosis and treatment, to avoid physical sequelae and secondary stigmatization. Primary health care should be strengthened for the recognition of neglected diseases whose incidence is underestimated in Colombia, with an atypical mixed multifocal presentation and a long evolution time. The KOH direct examination is an accessible and economical tool in the first levels of care that can contribute to the diagnostic approach.

An Experimental Model of Chromoblastomycosis Caused by Fonsecaea sp. Species.

The experimental rodent models for the fungal disease are a handy tool for understanding host-fungus interactions. To Fonsecaea sp., one of the causative agents of chromoblastomycosis, there is an extra challenge because the animals preferably used show a spontaneous cure; so until now, there is no model to reproduce the long-term disease similar to human chronic disease. In this chapter, we described an experimental model using rats and mice with a subcutaneous route, with the checkpoints of acute-like and chronic-like lesion analysis comparable with human lesions, the fungal burden, and the lymphocytes investigation.

New methylene blue-mediated photodynamic inactivation of multidrug-resistant Fonsecaea nubica infected chromoblastomycosis in vitro.

Chromoblastomycosis is a fungal disease presented with local warty papule, plaque, and verrucous nodules. In addition, the incidence and drug resistance of chromoblastomycosis are increasing each year worldwide. Photodynamic therapy is a promising method to treat mycoses. The purpose of this study was to evaluate the effect of new methylene blue (NMB)-induced PDT on multidrug-resistant chromoblastomycosis in vitro. We isolated one wild-type strain pathogen from one clinical patient diagnosed with chromoblastomycosis for over 27 years. The pathogen was identified by histopathology, the morphology of fungal culture, and genetic testing. Drug susceptibility testing was performed on the isolate. It was cultured with logarithmic growth phase spore in vitro and incubated with different concentrations of NMB for 30 min, and received illumination by red light-emitted diode with different light doses. After photodynamic treatment, the scanning electron microscopy (SEM) and transmission electron microscopy (TEM) were conducted. The pathogen was Fonsecaea nubica, and it was resistant to itraconazole, terbinafine, amphotericin B, voriconazole andcaspofungin. At the same NMB concentration, the sterilization efficiency of NMB-photodynamic therapy (PDT) on F. nubica increased with increasing light intensity; F. nubica was completely killed at 25 µmol/L NMB with a light dose of 40 J/cm2 or 50 µmol/L NMB and light doses of ≥ 30 J/cm2. SEM and TEM observed ultrastructural changes after PDT. NMB-PDT inactivates the survival of multidrug-resistant F. nubica in vitro; it therefore has the potential to become an alternative or adjuvant treatment for refractory chromoblastomycosis.