Ergosterol Biosynthesis Inhibitors Research Articles

The development of fluconazole resistance in Candida albicans - an example of microevolution of a fungal pathogen.

The yeast Candida albicans is a member of the microbiota in the gastrointestinal and urogenital tracts of most healthy persons, but it can also cause symptomatic infections, especially in immunocompromised patients. During the life-long association with its human host, C. albicans generates genetically altered variants that are better adapted to changes in their environment. A prime example of this microevolution is the development of resistance to the commonly used drug fluconazole, which inhibits ergosterol biosynthesis, during antimycotic therapy. Fluconazole resistance can be caused by mutations in the drug target, by changes in the sterol biosynthesis pathway, and by gain-of-function mutations in transcription factors that result in the constitutive upregulation of ergosterol biosynthesis genes and multidrug efflux pumps. Fluconazole also induces genomic rearrangements that result in gene amplification and loss of heterozygosity for resistance mutations, which further increases drug resistance. These genome alterations may affect extended chromosomal regions and have additional phenotypic consequences. A striking case is the loss of heterozygosity for the mating type locus MTL in many fluconazole-resistant clinical isolates, which allows the cells to switch to the mating-competent opaque phenotype. This, in turn, raises the possibility that sexual recombination between different variants of an originally clonal, drug-susceptible population may contribute to the generation of highly fluconazole-resistant strains with multiple resistance mechanisms. The gain-of-function mutations in transcription factors, which result in deregulated gene expression, also cause reduced fitness. In spite of this, many clinical isolates that contain such mutations do not exhibit fitness defects, indicating that they have overcome the costs of drug resistance with further evolution by still unknown mechanisms.

Isavuconazole: an orphan drug for treating invasive candidiasis

Introduction: Infections by Candida species are the most frequent invasive fungal infections in critically ill patients, and available licensed antifungals are restricted and show several limitations such as a narrow spectrum of activity, toxicity, and drug interactions. Hence, there is an urgent need for new antifungal drugs with favorable pharmacokinetic (PK), improved efficacy and safety profile. Isavuconazole is new broad-spectrum antifungal available as an oral and intravenous formulation; the prodrug isavuconazonium sulfate (BAL8557) is a water-soluble triazole and its active moiety, BAL4815, is a potent inhibitor of ergosterol biosynthesis. Isavuconazole offers several advantages over other azole antifungals, including high prodrug water solubility, high oral bioavailability, predictable and linear PK with no relevant food effect. As with other azoles, a loading dose is required.Areas covered: Studies and reviews were collected through a MEDLINE search and from http://www.clinicaltrials.gov, Food and Drug Administration briefing documents, program abstracts from international symposia, and the manufacturer’s website with focus on isavuconazole and Candida spp.Expert opinion: Clinical data regarding isavuconazole and outcome of Candida infections are limited, hence a clear position of the drug in any treatment-regimens are still open; the results of the ACTIVE-study showed that isavuconazole did not meet the primary objective of demonstrating non-inferior efficacy of isavuconazole versus caspofungin at the end of iv therapy.

Antifungal properties and inhibitory effects upon aflatoxin production by Zingiber officinale essential oil in Aspergillus flavus

SummaryIn this study, the efficacy of ginger (Zingiber officinale Roscoe) essential oil (GEO) in reducing A. flavus growth and aflatoxin production was investigated. Gas chromatography coupled to mass spectrometry and nuclear magnetic resonance spectroscopy showed that the major components of GEO were α‐zingiberene (23.85%) and geranial (14.16%). Mycelial growth of Aspergillus flavus was reduced significantly at a GEO concentration of 150 μg mL−1, and complete inhibition of conidial germination was observed at a concentration of 10 μg mL−1. Statistically significant inhibition of ergosterol biosynthesis was detected at a GEO concentration of 10 μg mL−1. GEO was capable of fully inhibiting aflatoxin production by A. flavus at a concentration of 15 μg mL−1. The results suggest that low concentrations of GEO are capable of inhibiting aflatoxin production; such ability could be valuable in the upcoming future for agricultural companies to better control aflatoxigenic fungi in agricultural products.

The activity of RTA2, a downstream effector of the calcineurin pathway, is required during tunicamycin-induced ER stress response in Candida albicans.

In this study, we demonstrate a novel function of a downstream effector molecule of the calcineurin pathway, RTA2 (Resistance To Aminocholesterol), in ER stress response. The deletion of RTA2 increases susceptibility to the ER stressor tunicamycin and morpholine-like drug, 7-aminocholesterol. Additionally, the expression of RTA2 is also transcriptionally induced by ergosterol biosynthesis inhibitors and cell-wall-damaging agents. As tunicamycin induces the unfolded protein response pathway (UPR) via the transcription factor, HAC1, we monitored the expression of a subset of HAC1-dependent UPR target genes in rta2Δ/Δ cells. Upon tunicamycin exposure, rta2Δ/Δ cells displayed a significantly reduced expression of UPR genes, in spite of only a moderate decrease in the HAC1 spliced mRNA levels and no change in Hac1 protein levels. Furthermore, hac1Δ/Δrta2Δ/Δ cells display an exacerbated sensitivity to tunicamycin compared to the single mutants. We propose that functional RTA2 is requisite for the regulation of Hac1p-dependent UPR target genes to maximal levels, thereby assisting survival during ER stress. Collectively, this study proposes, for the first time, existence of an interplay between the Hac1p- and calcineurin- controlled networks via a downstream effector molecule of the latter, RTA2, to facilitate survival during ER stress in Candida albicans.

ChemInform Abstract: One‐Pot Three‐Component Domino Protocol for the Synthesis of Novel Pyrano[2,3‐d]pyrimidines as Antimicrobial and Anti‐biofilm Agents.

A simple and facile synthesis of a series of novel pyrano[2,3-d]pyrimidines has been achieved successfully via the one-pot three-component reaction of 2-amino-7-methyl-5-oxo-4-phenyl-4,5-dihydropyrano[4,3-b]pyran-3-carbonitriles, DMF-DMA and arylamines in the presence of 1-butyl-3-methylhydrogensulphate [Bmim]HSO4 ionic liquid. This method has several advantages such as high yields, clean reaction, simple methodology and short reaction times. The synthesized compounds were evaluated for their antimicrobial activity against Gram-positive, Gram-negative and different Candida strains. Among the derivatives screened, compounds 4c, 4d, 4h and 4l were found to be active against both bacterial and Candida strains with MIC values ranging from 3.9 to 31.2 μg mL−1. In addition, compound 4l showed a good minimum bactericidal concentration, minimum fungicidal concentration and anti-biofilm activities. Furthermore, the mode of the antifungal action for the promising compound 4l was evaluated in C. albicans MTCC 1637 through an ergosterol biosynthesis inhibition process.

Fluorinated Sterols Are Suicide Inhibitors of Ergosterol Biosynthesis and Growth in Trypanosoma brucei

Trypanosoma brucei, the causal agent for sleeping sickness, depends on ergosterol for growth. Here, we describe the effects of a mechanism-based inhibitor, 26-fluorolanosterol (26FL), which converts invivo to a fluorinated substrate of the sterol C24-methyltransferase essential for sterol methylation and function of ergosterol, and missing from the human host. 26FL showed potent inhibition of ergosterol biosynthesis and growth of procyclic and bloodstream forms while having no effect on cholesterol biosynthesis or growth of human epithelial kidney cells. During exposure of cloned TbSMT to 26-fluorocholesta-5,7,24-trienol, the enzyme is gradually killed as a consequence of the covalent binding of the intermediate C25 cation to the active site (kcat/kinact= 0.26min(-1)/0.24min(-1); partition ratio of 1.08), whereas 26FL is non-productively bound. These results demonstrate that poisoning of ergosterol biosynthesis by a 26-fluorinated Δ(24)-sterol is a promising strategy for developing a new treatment for trypanosomiasis.

Elevated ergosterol protects Leishmania parasites against antimony-generated stress.

Parasite lipids can serve as signaling molecules, important membrane components, energy suppliers, and pathogenesis factors critical for survival. Functional roles of lipid changes in response to drug-generated stress in parasite survival remains unclear. To investigate this, Leishmania donovani parasites, the causative agents of kala-azar, were exposed to the antileishmanial agent potassium antimony tartrate (PAT) (half-maximal inhibitory concentration ∼ 284 µg/ml). Analysis of cell extracts using gas chromatography-mass spectrometry showed significant increases in very long-chain fatty acids (VLCFAs) prior to an increase in ergosterol in PAT-treated parasites as compared with vehicle-treated controls. Ergosterol biosynthesis inhibition during PAT treatment decreased cell viability. VLCFA inhibition with specific inhibitors completely abrogated ergosterol upsurge followed by a reduction in cell viability. Following PAT-induced VLCFA increase, an upsurge in reactive oxygen species (ROS) occurred and inhibition of this ROS with antioxidants abrogated ergosterol increase. Genetically engineered parasites expressing low constitutive ergosterol levels showed more susceptibility to PAT as compared with wild-type control cells but ergosterol supplementation during PAT treatment increased cell viability. In conclusion, we propose that during antimony treatment, the susceptibility of parasites is determined by the levels of cellular ergosterol that are regulated by oxidative stress generated by VLCFAs.

Heterologous expression of the Monilinia fructicola CYP51 (MfCYP51) gene in Pichia pastoris confirms the mode of action of the novel fungicide, SYP-Z048.

The novel agricultural fungicide 3-[5-(4-chlorophenyl)-2,3-dimethyl-3-isoxazolidinyl] pyridine (SYP-Z048) developed by China Shenyang Research Institute of Chemical Industry has been confirmed to be an ergosterol biosynthesis inhibitor (EBI). Previous studies have shown that EBIs target the proteins from a range of genes, including CYP51, ERG2 and/or ERG24, and ERG27, which are involved in the ergosterol biosynthesis pathway. In the current study the ERG2, ERG24, and ERG27 genes were cloned from wild type and resistant mutants of Monilinia fructicola in an attempt to clarify the target site of SYP-Z048. Comparative analysis of the deduced aa sequence of these genes, as well as CYP51, revealed several point mutations that resulted in amino acid variation among the sensitive and resistant isolates. However, sensitivity assays indicated that only one, the substitution of phenylalanine (F) for the tyrosine (Y) at 136 in CYP51, was correlated with reduced sensitivity to SYP-Z048. Heterologous expression of MfCYP51-136Y (MfCYP136Y) and MfCYP51-136F (MfCYP136F) in Pichia pastoris revealed that MfCYP136F significantly reduced sensitivity to SYP-Z048, increasing the average EC50 of the transformants 11-fold relative to those carrying MfCYP136Y. However, neither the additional copy of MfCYP136Y nor multiple copies of MfCYP136F were found to reduce sensitivity relative to the empty vector control or single copy transformants, respectively. Molecular docking experiments using SYP-Z048 with HsCYP145Y and the mutated version HsCYP145F as substitutes for MfCYP136Y and MfCYP136F, respectively, indicated that the reduced affinity of HsCYP145F for SYP-Z048 resulted from the loss of a hydrogen bond between the fungicide and the active site. Taken together these results indicate that MfCYP51 is the major target site of SYP-Z048 in M. fructicola, which has important implications for the resistance management of this fungicide in the field.

QTL mapping of fungicide sensitivity reveals novel genes and pleiotropy with melanization in the pathogen Zymoseptoria tritici

A major problem associated with the intensification of agriculture is the emergence of fungicide resistance. Azoles are ergosterol biosynthesis inhibitors that have been widely used in agriculture and medicine since the 1970s, leading to emergence of increasingly resistant fungal populations. The known genetic mechanisms underlying lower azole sensitivity include mutations affecting the CYP51 gene that encodes the target protein, but in many cases azole resistance is a more complex trait with an unknown genetic basis. We used quantitative trait locus (QTL) mapping to identify genes affecting azole sensitivity in two crosses of Zymoseptoria tritici, the most damaging wheat pathogen in Europe. Restriction site associated DNA sequencing (RADseq) was used to genotype 263 (cross 1) and 261 (cross 2) progeny at ∼8500 single nucleotide polymorphisms (SNP) and construct two dense linkage maps. Azole sensitivity was assessed using high-throughput digital image analysis of colonies growing on Petri dishes with or without the fungicide propiconazole. We identified three QTLs for azole sensitivity, including two that contained novel fungicide sensitivity genes. One of these two QTLs contained only 16 candidate genes, among which four most likely candidates were identified. The third QTL contained ERG6, encoding another protein involved in ergosterol biosynthesis. Known genes in QTLs affecting colony growth included CYP51 and PKS1, a gene affecting melanization in Z. tritici. PKS1 showed compelling evidence for pleiotropy, with a rare segregating allele that increased melanization while decreasing growth rate and propiconazole sensitivity. This study resolved the genetic architecture of an important agricultural trait and led to identification of novel genes that are likely to affect azole sensitivity in Z. tritici. It also provided insight into fitness costs associated with lowered azole sensitivity and suggests a novel fungicide mixture strategy.

Antifungal Activity of Ellagic Acid In Vitro and In Vivo.

Ellagic acid (EA) has been shown to have antioxidant, antibacterial, and anti-inflammatory activities. In Uighur traditional medicine, Euphorbia humifusa Willd is used to treat fungal diseases, and recent studies suggest that it is the EA content which is responsible for its therapeutic effect. However, the effects of EA on antifungal activity have not yet been reported. This study aimed to investigate the inhibitory effect of EA on fungal strains both in vitro and in vivo. The minimal inhibitory concentration (MIC) was determined by the National Committee for Clinical Laboratory Standards (M38-A and M27-A2) standard method in vitro. EA had a broad spectrum of antifungal activity, with MICs for all the tested dermatophyte strains between 18.75 and 58.33 µg/ml. EA was also active against two Candida strains, with MICs between 25.0 and 75.0 µg/ml. It was inactive against Candida glabrata. The susceptibility of six species of dermatophytes to EA was comparable with that of the commercial antifungal, fluconazole. The most sensitive filamentous species was Trichophyton rubrum (MIC = 18.75 µg/ml). Studies on the mechanism of action using an HPLC-based assay and an enzyme linked immunosorbent assay showed that EA inhibited ergosterol biosynthesis and reduced the activity of sterol 14α-demethylase P450 (CYP51) in the Trichophyton rubrum membrane, respectively. An in vivo test demonstrated that topical administration of EA (4.0 and 8.0 mg/cm(2) ) significantly enhanced the cure rate in a guinea-pig infection model of Trichophyton rubrum. The results suggest that EA has the potential to be developed as a natural antifungal agent.

Trafficking through the late endosome significantly impacts Candida albicans tolerance of the azole antifungals.

The azole antifungals block ergosterol biosynthesis by inhibiting lanosterol demethylase (Erg11p). The resulting depletion of cellular ergosterol and the accumulation of "toxic" sterol intermediates are both thought to compromise plasma membrane function. However, the effects of ergosterol depletion upon the function of intracellular membranes and organelles are not well described. The purpose of this study was to characterize the effects of azole treatment upon the integrity of the Candida albicans vacuole and to determine whether, in turn, vacuolar trafficking influences azole susceptibility. Profound fragmentation of the C. albicans vacuole can be observed as an early consequence of azole treatment, and it precedes significant growth inhibition. In addition, a C. albicans vps21Δ/Δ mutant, blocked in membrane trafficking through the late endosomal prevacuolar compartment (PVC), is able to grow significantly more than the wild type in the presence of several azole antifungals under standard susceptibility testing conditions. Furthermore, the vps21Δ/Δ mutant is able to grow despite the depletion of cellular ergosterol. This phenotype resembles an exaggerated form of "trailing growth" that has been described for some clinical isolates. In contrast, the vps21Δ/Δ mutant is hypersensitive to drugs that block alternate steps in ergosterol biosynthesis. On the basis of these results, we propose that endosomal trafficking defects may lead to the cellular "redistribution" of the sterol intermediates that accumulate following inhibition of ergosterol biosynthesis. Furthermore, the destination of these intermediates, or the precise cellular compartments in which they accumulate, may be an important determinant of their toxicity and thus ultimately antifungal efficacy.

Synergistic activity of magnolol with azoles and its possible antifungal mechanism againstCandida albicans

The goal of this study was to investigate the synergic effects between magnolol and azoles, and the potential antifungal mechanisms. Microdilution checkerboard, time-kill and agar diffusion assay were employed to evaluate the synergic effects between magnolol and fluconazole (FLC). Magnolol significantly decreased the efflux of rhodamine 123 (Rh123), leading to greater intracellular accumulation of Rh123 in Candida albicans cells. Compared to the Candida drug resistance (cdr) 2 or multidrug resistance (mdr) 1 deletion mutant, the growth of cdr1 strain was most sensitive to magnolol exposure. In the presence of magnolol, MDR1 overexpressing cells were sensitive to FLC, whereas CDR1 and CDR2 overexpressing cells displayed tolerance to FLC. Magnolol treatment correlated with up-regulation of transporter and ergosterol biosynthesis pathway genes, analyzed by real-time reverse transcription-polymerase chain reaction. The ergosterol content of C.albicansSC5314 was significantly decreased after magnolol exposure. Magnolol synergizes with azoles for targeting of C.albicans by inducing a higher intracellular content of antifungals, by tapping into the competitive effect of ABC transporter Cdr1p substrates, and enhancing the effect by targeting of the ergosterol biosynthesis pathway. Our results provide the first evidence that magnolol may function as a Cdr1p substrate and as an inhibitor of ergosterol biosynthesis. This function can thus be exploited in combination with azoles to reverse multidrug resistance of C.albicans strains.

The evolution of drug resistance in clinical isolates of Candida albicans.

Candida albicans is both a member of the healthy human microbiome and a major pathogen in immunocompromised individuals. Infections are typically treated with azole inhibitors of ergosterol biosynthesis often leading to drug resistance. Studies in clinical isolates have implicated multiple mechanisms in resistance, but have focused on large-scale aberrations or candidate genes, and do not comprehensively chart the genetic basis of adaptation. Here, we leveraged next-generation sequencing to analyze 43 isolates from 11 oral candidiasis patients. We detected newly selected mutations, including single-nucleotide polymorphisms (SNPs), copy-number variations and loss-of-heterozygosity (LOH) events. LOH events were commonly associated with acquired resistance, and SNPs in 240 genes may be related to host adaptation. Conversely, most aneuploidies were transient and did not correlate with drug resistance. Our analysis also shows that isolates also varied in adherence, filamentation, and virulence. Our work reveals new molecular mechanisms underlying the evolution of drug resistance and host adaptation.

Lead Identification to Clinical Candidate Selection: Drugs for Chagas Disease

Chagas disease affects 8 million people worldwide and remains a main cause of death due to heart failure in Latin America. The number of cases in the United States is now estimated to be 300,000, but there are currently no Food and Drug Administration (FDA)–approved drugs available for patients with Chagas disease. To fill this gap, we have established a public-private partnership between the University of California, San Francisco and the Genomics Institute of the Novartis Research Foundation (GNF) with the goal of delivering clinical candidates to treat Chagas disease. The discovery phase, based on the screening of more than 160,000 compounds from the GNF Academic Collaboration Library, led to the identification of new anti-Chagas scaffolds. Part of the screening campaign used and compared two screening methods, including a colorimetric-based assay using Trypanosoma cruzi expressing β-galactosidase and an image-based, high-content screening (HCS) assay using the CA-I/72 strain of T. cruzi. Comparing molecules tested in both assays, we found that ergosterol biosynthesis inhibitors had greater potency in the colorimetric assay than in the HCS assay. Both assays were used to inform structure-activity relationships for antiparasitic efficacy and pharmacokinetics. A new anti–T. cruzi scaffold derived from xanthine was identified, and we describe its development as lead series.

One-pot three-component domino protocol for the synthesis of novel pyrano[2,3-d]pyrimidines as antimicrobial and anti-biofilm agents.

A simple and facile synthesis of a series of novel pyrano[2,3-d]pyrimidines has been achieved successfully via the one-pot three-component reaction of 2-amino-7-methyl-5-oxo-4-phenyl-4,5-dihydropyrano[4,3-b]pyran-3-carbonitriles, DMF-DMA and arylamines in the presence of 1-butyl-3-methylhydrogensulphate [Bmim]HSO4 ionic liquid. This method has several advantages such as high yields, clean reaction, simple methodology and short reaction times. The synthesized compounds were evaluated for their antimicrobial activity against Gram-positive, Gram-negative and different Candida strains. Among the derivatives screened, compounds 4c, 4d, 4h and 4l were found to be active against both bacterial and Candida strains with MIC values ranging from 3.9 to 31.2 μg mL(-1). In addition, compound 4l showed a good minimum bactericidal concentration, minimum fungicidal concentration and anti-biofilm activities. Furthermore, the mode of the antifungal action for the promising compound 4l was evaluated in C. albicans MTCC 1637 through an ergosterol biosynthesis inhibition process.

Novel fatty acid esters of apocynin oxime exhibit antimicrobial and antioxidant activities

Apocynin (4‐hydroxy‐3‐methoxy acetophenone), a well known phytochemical, was converted to its oxime and further esterified with fatty acids of different chain lengths (C4‐C18) to obtain the oxime esters in about 80% yields. The structures of the oxime esters were confirmed using NMR, IR and mass spectroscopy. The prepared oxime esters of fatty acids and apocynin were evaluated for their in vitro antimicrobial and antioxidant activities. It was observed that the oxime ester of undecenoic acid was the most active apocynin oxime ester that exhibited both antibacterial and antifungal activities. From a mechanistic perspective, it was found that the oxime ester of undecenoic acid acted as an ergosterol biosynthesis inhibitor, confirming its antifungal property. All the compounds exhibited moderate antioxidant activity based on different assays such as DPPH radical scavenging, superoxide free radical scavenging and inhibition of lipid peroxidation.Practical application: The fatty oxime ester could be promising as novel antifungal agent, in view of the increasing resistance of pathogenic fungi to antifungal agents.Oxime esters of apocynin.

Biodirected synthesis of Miconazole-conjugated bacterial silver nanoparticles and their application as antifungal agents and drug delivery vehicles

The recent strategy to improve the efficacy of drugs is to combine them with metal nanoparticles for the control of microbial infections. Considering this fact, we developed a low cost and eco-friendly method for silver nanoparticles synthesis using the cell free supernatant of Delftia sp. strain KCM-006 and their application as antifungal agents and as a drug carrier. Transmission electron microscopy (TEM) and dynamic light scattering (DLS) analysis revealed the formation of spherical and monodispersed silver nanoparticles with an average size of 9.8nm. The synthesized nanoparticles were found to be photoluminescent, highly stable and crystalline in nature having a zeta potential of −31mV. The silver nanoparticles exhibited very good antifungal activity against various pathogenic Candida strains. Furthermore, the efficacy of nanoparticles was increased by conjugating the antifungal drug Miconazole to silver nanoparticles which exhibited significant fungicidal activity, inhibition of ergosterol biosynthesis and biofilm inhibition by increasing ROS levels. In addition, the cell viability and immunocytochemistry analysis against different normal cell lines including Chinese hamster ovary cells (CHO), human lung cell line (MRC5) and human vascular endothelial cells (HUVEC) demonstrated that these nanoparticles were non-toxic up to a concentration of 20μM. In conclusion, these results suggest that the synthesized nanoparticles find application as both antifungal agents and drug delivery vehicles. This is a first report on the preparation of silver nanoparticles using culture supernatant from Delftia sp. and also on the conjugation of Miconazole, an antifungal drug, to the bacterial silver nanoparticles.

Antifungal activity of geraniol and citronellol, two monoterpenes alcohols, against Trichophyton rubrum involves inhibition of ergosterol biosynthesis

Context: Trichophyton rubrum is the most common fungus causing chronic dermatophytosis in humans. Antifungal activity of promising agents is of great interest. Geraniol and citronellol are monoterpenes with antimicrobial properties.Objective: This study aimed to investigate the inhibitory effects and possible mechanism of antifungal activity of geraniol and citronellol against strains of T. rubrum.Materials and methods: The minimum inhibitory concentration (MIC) of each drug against 14 strains was determined by broth microdilution. The effects of the drugs on dry mycelial weight, conidial germination, infectivity on human nail fragments, and morphogenesis of T. rubrum were analyzed. The effects on the cell wall (test with sorbitol) and cell membrane (release of intracellular material and ergosterol biosynthesis) were investigated.Results: MIC values of geraniol ranged between 16 and 256 µg/mL while citronellol showed MIC values from 8 to 1024 µg/mL. The drugs (MIC and 2 × MIC) inhibited the mycelial growth, conidia germination, and fungal growth on nail fragments. The drugs (half of MIC) induced the formation of wide, short, and crooked hyphae in T. rubrum morphology. With sorbitol, geraniol MIC was increased by 64-fold and citronellol by 32-fold. The drugs caused leakage of intracellular material and inhibited ergosterol biosynthesis.Discussion: The results suggest that the drugs damage cell wall and cell membrane of T. rubrum through a mechanism that seems to involve the inhibition of the ergosterol biosynthesis.Conclusion: This study confirms that geraniol and citronellol can be regarded as potential drugs for controlling T. rubrum growth, with great potential against agents of dermatophytosis.

Recent clinical trials for the etiological treatment of chronic chagas disease: advances, challenges and perspectives.

Chagas disease, a chronic systemic parasitosis caused by the Kinetoplastid protozoon Trypanosoma cruzi, is the first cause of cardiac morbidity and mortality in poor rural and suburban areas of Latin America and the largest parasitic disease burden in the continent, now spreading worldwide due to international migrations. A recent change in the scientific paradigm on the pathogenesis of chronic Chagas disease has led to a consensus that all T. cruzi-seropositive patients should receive etiological treatment. This important scientific advance has spurred the rigorous evaluation of the safety and efficacy of currently available drugs (benznidazole and nifurtimox) as well as novel anti-T. cruzi drug candidates in chronic patients, who were previously excluded from such treatment. The first results indicate that benznidazole is effective in inducing a marked and sustained reduction in the circulating parasites' level in the majority of these patients, but adverse effects can lead to treatment discontinuation in 10-20% of cases. Ergosterol biosynthesis inhibitors, such as posaconazole and ravuconazole, are better tolerated but their efficacy at the doses and treatment duration used in the initial studies was significantly lower; such results are probably related to suboptimal exposure and/or treatment duration. Combination therapies are a promising perspective but the lack of validated biomarkers of response to etiological treatment and eventual parasitological cures in chronic patients remains a serious challenge.

Antifungal properties and inhibitory effects upon aflatoxin production of Thymus vulgaris L. by Aspergillus flavus Link

The antifungal and antiaflatoxigenic properties of Thymus vulgaris essential oil (TEO) were evaluated upon Aspergillus flavus “in vitro”. Suspension containing 106 of A. flavus were cultivated with TEO in concentrations ranging from 50 to 500μg/mL. TEO reached minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) at 250μg/mL. Inhibition of ergosterol biosynthesis was detected at a concentration of 100μg/mL of TEO. Morphological evaluation performed by both light microscopy and scanning electron microscopy showed that antifungal activity of TEO could be detected starting at a concentration of 50μg/mL and the fungicide effect at a concentration of 250μg/mL. TEO completely inhibited production of both B1 and B2 aflatoxins (AFB1 and AFB2) at a concentration of 150μg/mL. This way, fungal biomass development and aflatoxin production were dependent on TEO concentration. Therefore, TEO was capable of controlling the growth of A. flavus and its production of aflatoxins.