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.

Preservation Methods in Isolates of Sporothrix Characterized by Polyphasic Approach.

Sporotrichosis is a subcutaneous mycosis with worldwide distribution and caused by eight pathogenic species of the Sporothrix genus. Different ex situ preservation methods are used around the world to maintain the survival, morphophysiological and genetic traits of fungal strains isolated from patients with sporotrichosis for long terms. The main aim of the present study was to evaluate the survival, phenotypic and genotypic stability of Sporothrix strains after preservation on PDA slant stored at 4 °C, sterile water and cryopreservation at -80 °C, for a period of time of 6, 12, 18 and 24 months of storage. Eight clinical Sporothrix isolates were identified based on a polyphasic approach consisting of classical macro- and micro-morphological traits, biochemical assays, proteomic profiles by MALDI-TOF MS and molecular biology. According to the final identification, one strain was identified as S. schenckii (CMRVS 40428) and seven strains were re-identified as S. brasiliensis (CMRVS 40421, CMRVS 40423, CMRVS 40424, CMRVS 40425, CMRVS 40426, CMRVS 40427 and CMRVS 40433). In addition, it was observed that the isolates survived after the different time points of storage in distilled water, PDA slant and cryopreservation at -80 °C. For fungi preserved in water, low polymorphisms were detected by the partial sequencing of ß-tubulin. Cryopreservation at -80 °C induced morphological changes in one single isolate. The proteomic profiles obtained by MALDI-TOF MS after preservation showed differences among the methods. In conclusion, preservation on agar slant stored at 4 °C was the most effective method to preserve the eight clinical Sporothrix strains. This method produced less change in the phenotypic traits and kept the genetic integrity of all strains. Agar slant stored at 4 °C is a simple and inexpensive method and can be especially used in culture collections of limited funding and resources.

New possibilities for chromoblastomycosis and phaeohyphomycosis treatment: identification of two compounds from the MMV Pathogen Box® that present synergism with itraconazole

BACKGROUND Black fungi of the Herpotrichiellaceae family are agents of chromoblastomycosis and phaeohyphomycosis. There are few therapeutic options for these infections and it is common to associate antifungal drugs in their treatment. OBJECTIVES To investigate the Medicines for Malaria Venture (MMV) Pathogen Box® for possible compounds presenting synergism with antifungal drugs used to treat black fungal infections. METHODS An initial screening of the Pathogen Box® compounds was performed in combination with itraconazole or terbinafine at sub-inhibitory concentrations against Fonsecaea pedrosoi. Hits were further tested against eight Herpotrichiellaceae using the checkerboard method. FINDINGS No synergism was observed with terbinafine. MMV687273 (SQ109) and MMV688415 showed synergism with itraconazole against F. pedrosoi. Synergism of these compounds was confirmed with some black fungi by the checkerboard method. SQ109 and itraconazole presented synergism for Exophiala dermatitidis, F. pedrosoi, F. monophora and F. nubica, with fungicidal activity for F. pedrosoi and F. monophora. MMV688415 presented synergism with itraconazole only for F. pedrosoi, with fungicidal activity. The synergic compounds had high selectivity index values when combined with itraconazole. MAIN CONCLUSIONS These compounds in combination, particularly SQ109, are promising candidates to treat Fonsecaea spp. and E. dermatitidis infections, which account for most cases of chromoblastomycosis and phaeohyphomycosis.

A case-series of bloodstream infections caused by the Meyerozyma guilliermondii species complex at a reference center of oncology in Brazil.

Bloodstream infections (BSI) caused by Candida species are the fourth cause of healthcare associated infections worldwide. Non-albicans Candida species emerged in the last decades as agents of serious diseases. In this study, clinical and microbiological aspects of six patients with BSI due to the Meyerozyma (Candida) guilliermondii species complex from an oncology reference center in Brazil, were evaluated. To describe demographic and clinical characteristics, medical records of the patients were reviewed. Molecular identification of the isolates was performed by ITS1-5.8S-ITS2 region sequencing. Antifungal susceptibility was evaluated by the EUCAST method and the minimal inhibitory concentrations (MIC) assessed according to the epidemiological cutoff values. Virulence associated phenotypes of the isolates were also studied. Ten isolates from the six patients were evaluated. Five of them were identified as Meyerozyma guilliermondii and the others as Meyerozyma caribbica. One patient was infected with two M. caribbica isolates with different genetic backgrounds. High MICs were observed for fluconazole and echinocandins. Non-wild type isolates to voriconazole appeared in one patient previously treated with this azole. Additionally, two patients survived, despite infected with non-wild type strains for fluconazole and treated with this drug. All isolates produced hemolysin, which was not associated with a poor prognosis, and none produced phospholipases. Aspartic proteases, phytase, and esterase were detected in a few isolates. This study shows the reduced antifungal susceptibility and a variable production of virulence-related enzymes by Meyerozyma spp. In addition, it highlights the poor prognosis of neutropenic patients with BSI caused by this emerging species complex. LAY ABSTRACT: Our manuscript describes demographic, clinical and microbiological characteristics of patients with bloodstream infection by the Meyerozyma guilliermondii species complex at a reference center in oncology in Brazil.

Molecular identification and antifungal susceptibility testing of Pucciniomycotina red yeast clinical isolates from Rio de Janeiro, Brazil.

Infections caused by Rhodotorula spp. are increasing worldwide. This study identified, through the light of the new taxonomic advances on the subphylum Pucciniomycotina, 16 isolates from blood cultures and compared their antifungal susceptibility on microdilution and gradient diffusion methods. Internal transcriber spacer sequencing identified Rhodotorula mucilaginosa (n = 12), Rhodotorula toruloides (n = 2), Rhodotorula dairenensis (n = 1), and Cystobasidium minutum (n = 1). Amphotericin B was the most effective drug. A good essential agreement was observed on MIC values of amphotericin B and voriconazole determined by the two methods. Therefore, the gradient method is useful for susceptibility tests of R. mucilaginosa against these drugs.

Fatal septic shock caused by Paracoccidioides brasiliensis phylogenetic species S1 in a young immunocompetent patient: a case report.

The authors report the first case of fatal septic shock, a rare clinical presentation of paracoccidioidomycosis (PCM) caused by Paracoccidioides brasiliensis S1. We also provide an immunological evaluation of the patient. Severe clinical signs such as organ dysfunction and digital gangrene occurred in this case. The patient presented a remarkable cell activation profile and diminished percentage of peripheral blood T regulatory cells. A decrease in anti-inflammatory IL-1RA plasma level showed the potential for endothelium damage, probably contributing to a vasculitis process. Together with P. lutzii, P. brasiliensis appears to be involved in severe cases of PCM.

Paracoccidioidomycosis due to Paracoccidioides brasiliensis S1 plus HIV co-infection

BACKGROUND Paracoccidioidomycosis (PCM) is one of the most important systemic mycoses in Latin America and the leading fungal cause of mortality in non-immunosuppressed individuals in Brazil. However, HIV/PCM co-infection can increase the clinical severity in these co-infected patients. This co-infection is rarely reported in the literature mainly because of the different epidemiological profiles of these infections. Furthermore, PCM is a neglected and non-notifiable disease, which may underestimate the real importance of this disease. The advent of molecular studies on the species of the genus Paracoccidioides has expanded the knowledge regarding the severity and the clinical spectrum in PCM. In this context, the development of studies to describe the association of the Paracoccidioides phylogenetic cryptic species in vulnerable populations, such as HIV-infected patients, appears relevant. OBJECTIVE To describe the clinical, epidemiological, therapeutic and prognostic aspects in HIV/PCM co-infected patients, along with the molecular identification of the Paracoccidioides species involved in these cases. METHODS The investigators performed a molecular and clinical retrospective study involving HIV/PCM co-infected patients, from a reference centre for PCM care in the endemic area of Rio de Janeiro, Brazil, from 1998 to 2015. Molecular identification of the fungal strains was done by amplification of partial sequences of arf and gp43 genes. FINDINGS Of 89 patients diagnosed with PCM by fungal isolation in the culture, a viable isolate was recovered for molecular analysis from 44 patients. Of these 44 patients, 28 (63.6%) had their serum samples submitted for enzyme immunoassay tests for screening of HIV antibodies, and 5 (17.9%) had a positive result. All cases were considered severe, with a variable clinical presentation, including mixed, acute/subacute clinical forms and a high rate of complications, requiring combination therapy. Paracoccidioides brasiliensis S1 was the species identified in all cases. CONCLUSIONS HIV/PCM co-infection can change the natural history of this fungal disease. The authors reinforce the need to include HIV screening diagnostic tests routinely for patients with PCM.

Paracoccidioidomycosis due to Paracoccidioides brasiliensis S1 plus HIV co-infection.

BACKGROUND: Paracoccidioidomycosis (PCM) is one of the most important systemic mycoses in Latin America and the leading fungal cause of mortality in non-immunosuppressed individuals in Brazil. However, HIV/PCM co-infection can increase the clinical severity in these co-infected patients. This co-infection is rarely reported in the literature mainly because of the different epidemiological profiles of these infections. Furthermore, PCM is a neglected and non-notifiable disease, which may underestimate the real importance of this disease. The advent of molecular studies on the species of the genus Paracoccidioides has expanded the knowledge regarding the severity and the clinical spectrum in PCM. In this context, the development of studies to describe the association of the Paracoccidioides phylogenetic cryptic species in vulnerable populations, such as HIV-infected patients, appears relevant. OBJECTIVE: To describe the clinical, epidemiological, therapeutic and prognostic aspects in HIV/PCM co-infected patients, along with the molecular identification of the Paracoccidioides species involved in these cases. METHODS: The investigators performed a molecular and clinical retrospective study involving HIV/PCM co-infected patients, from a reference centre for PCM care in the endemic area of Rio de Janeiro, Brazil, from 1998 to 2015. Molecular identification of the fungal strains was done by amplification of partial sequences of arf and gp43 genes. FINDINGS: Of 89 patients diagnosed with PCM by fungal isolation in the culture, a viable isolate was recovered for molecular analysis from 44 patients. Of these 44 patients, 28 (63.6%) had their serum samples submitted for enzyme immunoassay tests for screening of HIV antibodies, and 5 (17.9%) had a positive result. All cases were considered severe, with a variable clinical presentation, including mixed, acute/subacute clinical forms and a high rate of complications, requiring combination therapy. Paracoccidioides brasiliensis S1 was the species identified in all cases. CONCLUSIONS: HIV/PCM co-infection can change the natural history of this fungal disease. The authors reinforce the need to include HIV screening diagnostic tests routinely for patients with PCM.

Fatal septic shock caused by Paracoccidioides brasiliensis phylogenetic species S1 in a young immunocompetent patient: a case report

Abstract The authors report the first case of fatal septic shock, a rare clinical presentation of paracoccidioidomycosis (PCM) caused by Paracoccidioides brasiliensis S1. We also provide an immunological evaluation of the patient. Severe clinical signs such as organ dysfunction and digital gangrene occurred in this case. The patient presented a remarkable cell activation profile and diminished percentage of peripheral blood T regulatory cells. A decrease in anti-inflammatory IL-1RA plasma level showed the potential for endothelium damage, probably contributing to a vasculitis process. Together with P. lutzii, P. brasiliensis appears to be involved in severe cases of PCM.