The fungal genus Sporothrix includes at least four human pathogenic species. One of these species, S. brasiliensis, is the causal agent of a major ongoing zoonotic outbreak of sporotrichosis in Brazil. Elsewhere, sapronoses are caused by S. schenckii and S. globosa. The major aims on this comparative genomic study are: 1) to explore the presence of virulence factors in S. schenckii and S. brasiliensis; 2) to compare S. brasiliensis, which is cat-transmitted and infects both humans and cats with S. schenckii, mainly a human pathogen; 3) to compare these two species to other human pathogens (Onygenales) with similar thermo-dimorphic behavior and to other plant-associated Sordariomycetes. The genomes of S. schenckii and S. brasiliensis were pyrosequenced to 17x and 20x coverage comprising a total of 32.3 Mb and 33.2 Mb, respectively. Pair-wise genome alignments revealed that the two species are highly syntenic showing 97.5% average sequence identity. Phylogenomic analysis reveals that both species diverged about 3.8-4.9 MYA suggesting a recent event of speciation. Transposable elements comprise respectively 0.34% and 0.62% of the S. schenckii and S. brasiliensis genomes and expansions of Gypsy-like elements was observed reflecting the accumulation of repetitive elements in the S. brasiliensis genome. Mitochondrial genomic comparisons showed the presence of group-I intron encoding homing endonucleases (HE’s) exclusively in S. brasiliensis. Analysis of protein family expansions and contractions in the Sporothrix lineage revealed expansion of LysM domain-containing proteins, small GTPases, PKS type1 and leucin-rich proteins. In contrast, a lack of polysaccharide lyase genes that are associated with decay of plants was observed when compared to other Sordariomycetes and dimorphic fungal pathogens, suggesting evolutionary adaptations from a plant pathogenic or saprobic to an animal pathogenic life style. Comparative genomic data suggest a unique ecological shift in the Sporothrix lineage from plant-association to mammalian parasitism, which contributes to the understanding of how environmental interactions may shape fungal virulence. . Moreover, the striking differences found in comparison with other dimorphic fungi revealed that dimorphism in these close relatives of plant-associated Sordariomycetes is a case of convergent evolution, stressing the importance of this morphogenetic change in fungal pathogenesis.

Nosocomial infections caused by fungi have increased greatly in recent years, mainly due to the rising number of immunocompromised patients. However, the available antifungal therapeutic arsenal is limited, and the development of new drugs has been slow. Therefore, the search for alternative drugs with low resistance rates and fewer side effects remains a major challenge. Plants produce a variety of medicinal components that can inhibit pathogen growth. Studies of plant species have been conducted to evaluate the characteristics of natural drug products, including their sustainability, affordability, and antimicrobial activity. A considerable number of studies of medicinal plants and alternative compounds, such as secondary metabolites, phenolic compounds, essential oils and extracts, have been performed. Thus, this review discusses the history of the antifungal arsenal, surveys natural products with potential antifungal activity, discusses strategies to develop derivatives of natural products, and presents perspectives on the development of novel antifungal drug candidates.

Background: The prevalence of invasive fungal infections (IFIs) has increased steadily worldwide in the last few decades. Particularly, there has been a global rise in the number of infections among immunosuppressed people. These patients present severe clinical forms of the infections, which are commonly fatal, and they are more susceptible to opportunistic fungal infections than non-immunocompromised people. IFIs have historically been associated with high morbidity and mortality, partly because of the limitations of available antifungal therapies, including side effects, toxicities, drug interactions and antifungal resistance. Thus, the search for alternative therapies and/or the development of more specific drugs is a challenge that needs to be met. Genomics has created new ways of examining genes, which open new strategies for drug development and control of human diseases.Results: In silicoanalyses and manual mining selected initially 57 potential drug targets, based on 55 genes experimentally confirmed as essential forCandida albicansorAspergillus fumigatusand other 2 genes (kre2anderg6) relevant for fungal survival within the host. Orthologs for those 57 potential targets were also identified in eight human fungal pathogens (C. albicans,A. fumigatus,Blastomyces dermatitidis,Paracoccidioides brasiliensis,Paracoccidioides lutzii, Coccidioides immitis,Cryptococcus neoformansandHistoplasma capsulatum). Of those, 10 genes were present in all pathogenic fungi analyzed and absent in the human genome. We focused on four candidates:trr1that encodes for thioredoxin reductase,rim8that encodes for a protein involved in the proteolytic activation of a transcriptional factor in response to alkaline pH,kre2that encodes for α-1,2-mannosyltransferase anderg6that encodes for Δ(24)-sterol C-methyltransferase.Conclusions: Our data show that the comparative genomics analysis of eight fungal pathogens enabled the identification of four new potential drug targets. The preferred profile for fungal targets includes proteins conserved among fungi, but absent in the human genome. These characteristics potentially minimize toxic side effects exerted by pharmacological inhibition of the cellular targets. From this first step of post-genomic analysis, we obtained information relevant to future new drug development.

Candida albicans is the most common species isolated from nosocomial bloodstream infections. Due to limited therapeutic arsenal and increase of drug resistance, there is an urgent need for new antifungals. Therefore, the antifungal activity against C. albicans and in vivo toxicity of a 1,3,4-oxadiazole compound (LMM6) was evaluated. This compound was selected by in silico approach based on chemical similarity. LMM6 was highly effective against several clinical C. albicans isolates, with minimum inhibitory concentration values ranging from 8 to 32 µg/mL. This compound also showed synergic effect with amphotericin B and caspofungin. In addition, quantitative assay showed that LMM6 exhibited a fungicidal profile and a promising anti-biofilm activity, pointing to its therapeutic potential. The evaluation of acute toxicity indicated that LMM6 is safe for preclinical trials. No mortality and no alterations in the investigated parameters were observed. In addition, no substantial alteration was found in Hippocratic screening, biochemical or hematological analyzes. LMM6 (5 mg/kg twice a day) was able to reduce both spleen and kidneys fungal burden and further, promoted the suppresses of inflammatory cytokines, resulting in infection control. These preclinical findings support future application of LMM6 as potential antifungal in the treatment of invasive candidiasis.

Paracoccidioidomycosis (PCM) is a neglected disease present in Latin America with difficulty in treatment and occurrence of serious sequelae. Thus, the development of alternative therapies is imperative. In the current work, two oxadiazole compounds (LMM5 and LMM11) presented fungicidal activity against Paracoccidioides spp. The minimum inhibitory and fungicidal concentration values ranged from 1 to 32 μg/mL, and a synergic effect was observed for both compounds when combined with Amphotericin B. LMM5 and LMM11 were able to reduce CFU counts (≥2 log₁₀) on the 5^th and 7^th days of time-kill curve, respectively. The fungicide effect was confirmed by fluorescence microscopy (FUN-1/FUN-2). The hippocratic screening and biochemical analysis were performed in Balb/c male mice that received a high dose of each compound, and the compounds showed no in vivo toxicity. The treatment of experimental PCM with the new oxadiazoles led to significant reduction in CFU (≥1 log₁₀). Histopathological analysis of the groups treated exhibited control of inflammation, as well as preserved lung areas. These findings suggest that LMM5 and LMM11 are promising hits structures, opening the door for implementing new PCM therapies. Paracoccidioidomycosis (PCM) is a granulomatous fungal infection with clinically severe forms and serious pulmonary sequelae. The current limited arsenal and prolonged treatment regimen demonstrate the need for new antifungals. This study reveals two fungicidal oxadiazole compounds for PCM treatment. The in vitro assay showed the antifungal activity and the synergic effect with Amphotericin B. The high doses administration in mice showed absence of toxicity, which allowed to demonstrate the in vivo antifungal activity.