Fungal Virulence Factors Research Articles

Insight into the transcriptome of Arthrobotrys conoides using high throughput sequencing.

Arthrobotrys conoides is a nematode-trapping fungus belonging to Orbiliales, Ascomycota group, and traps prey nematodes by means of adhesive network. Fungus has a potential to be used as a biocontrol agent against plant parasitic nematodes. In the present study, we characterized the transcriptome of A. conoides using high-throughput sequencing technology and characterized its virulence unigenes. Total 7,255 cDNA contigs with an average length of 425 bp were generated and 6184 (61.81%) transcripts were functionally annotated and characterized. Majority of unigenes were found analogous to the genes of plant pathogenic fungi. A total of 1749 transcripts were found to be orthologous with eukaryotic proteins of KOG database. Several carbohydrate active enzymes and peptidases were identified. We also analyzed classically and nonclassically secreted proteins and confirmed by BLASTP against fungal secretome database. A total of 916 contigs were analogous to 556 unique proteins of Pathogen Host Interaction (PHI) database. Further, we identified 91 unigenes homologous to the database of fungal virulence factor (DFVF). A total of 104 putative protein kinases coding transcripts were identified by BLASTP against KinBase database, which are major players in signaling pathways. This study provides a comprehensive look at the transcriptome of A. conoides and the identified unigenes might have a role in catching and killing prey nematodes by A. conoides.

Gene expression patterns and dynamics of the colonization of common bean (Phaseolus vulgaris L.) by highly virulent and weakly virulent strains of Fusarium oxysporum

The dynamics of root and hypocotyl colonization, and the gene expression patterns of several fungal virulence factors and plant defense factors have been analyzed and compared in the interaction of two Fusarium oxysporum f. sp. phaseoli strains displaying clear differences in virulence, with a susceptible common bean cultivar. The growth of the two strains on the root surface and the colonization of the root was quantitatively similar although the highly virulent (HV) strain was more efficient reaching the central root cylinder. The main differences between both strains were found in the temporal and spatial dynamics of crown root and hypocotyl colonization. The increase of fungal biomass in the crown root was considerably larger for the HV strain, which, after an initial stage of global colonization of both the vascular cylinder and the parenchymal cells, restricted its growth to the newly differentiated xylem vessels. The weakly virulent (WV) strain was a much slower and less efficient colonizer of the xylem vessels, showing also growth in the intercellular spaces of the parenchyma. Most of the virulence genes analyzed showed similar expression patterns in both strains, except SIX1, SIX6 and the gene encoding the transcription factor FTF1, which were highly upregulated in root crown and hypocotyl. The response induced in the infected plant showed interesting differences for both strains. The WV strain induced an early and strong transcription of the PR1 gene, involved in SAR response, while the HV strain preferentially induced the early expression of the ethylene responsive factor ERF2.

Potential roles of laccases on virulence of Heterobasidion annosum s.s.

Laccases, multi-copper-containing proteins, can catalyze the oxidation of phenolic substrates and have diverse functions such as a virulence factor in fungi. However, limited information can be found on the role of laccases in the interaction of Heterobasidion annosum s.s. to its host plant. Due to genome availability of the close-related species Heterobasidion irregulare, which contains 18 predicted laccase-encoding genes, phylogenetic analysis and gene expression profiling were performed. Eighteen laccase genes could be classified into 4 groups based on protein domains and phylogenetic analysis. However, there is no clear indication between phylogeny and domain compositions in laccases, and lifestyles of fungal species. The results of qRT-PCR showed that the expression of 8 laccase genes was highly up-regulated in Scots pine seedlings at 1 wpi. These data suggested that they might be involved in early stage of host infection. In addition, up-regulation of gene expression under glucose condition as a sole carbon source suggests that those laccases are not under carbon catabolite repression. Higher activities of laccase were observed in culture media containing cellulose, sucrose, or glucose compared to that of cellobiose as a sole carbon source. The highest mortality of Scots pine seedlings was observed when infected by H. annosum s.s. on extra carbon source as glucose. This was supported by the facts that glucose plays significant roles on up-regulation of laccase genes in planta and higher activity of laccase in H. annosum s.s.. Taking all together, laccases in H. annosum s.s. have diverse functions and a group of laccases may play a role during interactions with Scots pine seedlings.

Novel mode of action of plant defense peptides - hevein-like antimicrobial peptides from wheat inhibit fungal metalloproteases.

The multilayered plant immune system relies on rapid recognition of pathogen-associated molecular patterns followed by activation of defense-related genes, resulting in the reinforcement of plant cell walls and the production of antimicrobial compounds. To suppress plant defense, fungi secrete effectors, including a recently discovered Zn-metalloproteinase from Fusarium verticillioides, named fungalysin Fv-cmp. This proteinase cleaves class IV chitinases, which are plant defense proteins that bind and degrade chitin of fungal cell walls. In this study, we investigated plant responses to such pathogen invasion, and discovered novel inhibitors of fungalysin. We produced several recombinant hevein-like antimicrobial peptides named wheat antimicrobial peptides (WAMPs) containing different amino acids (Ala, Lys, Glu, and Asn) at the nonconserved position 34. An additional Ser at the site of fungalysin proteolysis makes the peptides resistant to the protease. Moreover, an equal molar concentration of WAMP-1b or WAMP-2 to chitinase was sufficient to block the fungalysin activity, keeping the chitinase intact. Thus, WAMPs represent novel protease inhibitors that are active against fungal metalloproteases. According to in vitro antifungal assays WAMPs directly inhibited hyphal elongation, suggesting that fungalysin plays an important role in fungal development. A novel molecular mechanism of dynamic interplay between host defense molecules and fungal virulence factors is suggested.

Effects of temperature and incubation time on the in vitro expression of proteases, phospholipases, lipases and DNases by different species of Trichosporon.

Fungi produce a broad spectrum of enzymes capable of degrading different substrates in nature. When the substrate is the tissue of a vertebrate host, these enzymes acts as a fungal virulence factor that increases the pathogenicity of the fungus. Trichosporon yeasts are emerging pathogens that infect immunocompromised patients. Little is known about the virulence characteristics of these fungi. The aim of this research was to characterize the behavior of protease, phospholipase, lipase and DNase production in different species of Trichosporon, with a focus on the influence of incubation temperature on the expression of these enzymes. Classical methodologies were used in all of the experiments, and the results were statistically analyzed. The proportions of the samples that produced each type of enzyme were as follows: lipases (95.5%), phospholipases (56.8%), proteases (50,0%) and DNases (38.6%). The incubation temperature was an important factor in the expression of enzymatic activity, and it influenced the incubation period of each species. Although these data concerning the enzymatic activity expressed by isolates of Trichosporon are valuable, further research is warranted to completely characterize this new pathogen, as well as in vivo studies to determine the roles of these enzymes in the pathogenesis of trichosporonosis.

Klinik Aspergillus spp. İzolatlarında Virülans Faktörü Olarak Asit Proteinaz ve Fosfolipaz Aktivitelerinin Araştırılması

Aspergillus spp. are widespread in nature and cause severe infections especially in immunocompromised patients. Aspergillus fumigatus complex is the most common species that causes infections in humans; however Aspergillus niger complex and Aspergillus flavus complex are the emerging agents that are isolated frequently from clinical specimens more recently. Besides the host factors such as immunosuppression, hematologic malignancy and corticosteroid use, fungal virulence factors such as elastase, acid protease and phospholipase enzymes are considered among the factors that affect the development of invasive aspergillosis. The aim of this study was to detect the acid proteinase and phospholipase enzyme activities in 30 A.fumigatus complex, nine A.flavus complex and four A.niger complex strains isolated from clinical specimens. Acid proteinase and phospholipase activities of the isolates were investigated by using bovine serum albumin agar (BSA), and egg yolk agar plates, respectively. Acid proteinase and phospholipase activity was detected in 76.7% (23/30) and 93.3% (28/30) of A.fumigatus complex isolates, respectively. None of the nine A.flavus complex isolates exhibited acid proteinase or phospholipase activity. Acid proteinase activity was not detected in any of the A.niger complex isolates (n= 4), however phospholipase activity was detected in one (25%) isolate. All of the acid proteinase positive A.fumigatus complex strains (n= 23) were also positive for phospholipase activity. In conclusion, further larger scale multicenter studies supported by clinical data, are needed to enlighten the roles of acid proteinase and phospholipase in the pathogenesis of infections due to Aspergillus spp.

Draft Genome Sequence of Penicillium expansum Strain R19, Which Causes Postharvest Decay of Apple Fruit.

Among the species that cause blue mold, isolates of Penicillium expansum are the most prevalent and virulent species, causing more than 50 percent of postharvest decay. We report the draft genome sequence of P. expansum R19 in order to identify fungal virulence factors and to understand the mechanism of infection.

Invasion of the central nervous system by Cryptococcus neoformans requires a secreted fungal metalloprotease.

ABSTRACTCryptococcus spp. cause life-threatening fungal infection of the central nervous system (CNS), predominantly in patients with a compromised immune system. Why Cryptococcus neoformans has this remarkable tropism for the CNS is not clear. Recent research on cerebral pathogenesis of C. neoformans revealed a predominantly transcellular migration of cryptococci across the brain endothelium; however, the identities of key fungal virulence factors that function specifically to invade the CNS remain unresolved. Here we found that a novel, secreted metalloprotease (Mpr1) that we identified in the extracellular proteome of C. neoformans (CnMpr1) is required for establishing fungal disease in the CNS. Mpr1 belongs to a poorly characterized M36 class of fungalysins that are expressed in only some fungal species. A strain of C. neoformans lacking the gene encoding Mpr1 (mpr1Δ) failed to breach the endothelium in an in vitro model of the human blood-brain barrier (BBB). A mammalian host infected with the mpr1Δ null strain demonstrated significant improvement in survival due to a reduced brain fungal burden and lacked the brain pathology commonly associated with cryptococcal disease. The in vivo studies further indicate that Mpr1 is not required for fungal dissemination and Mpr1 likely targets the brain endothelium specifically. Remarkably, the sole expression of CnMPR1 in Saccharomyces cerevisiae resulted in a robust migration of yeast cells across the brain endothelium, demonstrating Mpr1’s specific activity in breaching the BBB and suggesting that Mpr1 may function independently of the hyaluronic acid-CD44 pathway. This distinct role for Mpr1 may develop into innovative treatment options and facilitate a brain-specific drug delivery platform.

Patterns of variation at Ustilago maydis virulence clusters 2A and 19A largely reflect the demographic history of its populations.

The maintenance of an intimate interaction between plant-biotrophic fungi and their hosts over evolutionary times involves strong selection and adaptative evolution of virulence-related genes. The highly specialised maize pathogen Ustilago maydis is assigned with a high evolutionary capability to overcome host resistances due to its high rates of sexual recombination, large population sizes and long distance dispersal. Unlike most studied fungus-plant interactions, the U. maydis – Zea mays pathosystem lacks a typical gene-for-gene interaction. It exerts a large set of secreted fungal virulence factors that are mostly organised in gene clusters. Their contribution to virulence has been experimentally demonstrated but their genetic diversity within U. maydis remains poorly understood. Here, we report on the intraspecific diversity of 34 potential virulence factor genes of U. maydis. We analysed their sequence polymorphisms in 17 isolates of U. maydis from Europe, North and Latin America. We focused on gene cluster 2A, associated with virulence attenuation, cluster 19A that is crucial for virulence, and the cluster-independent effector gene pep1. Although higher compared to four house-keeping genes, the overall levels of intraspecific genetic variation of virulence clusters 2A and 19A, and pep1 are remarkably low and commensurate to the levels of 14 studied non-virulence genes. In addition, each gene is present in all studied isolates and synteny in cluster 2A is conserved. Furthermore, 7 out of 34 virulence genes contain either no polymorphisms or only synonymous substitutions among all isolates. However, genetic variation of clusters 2A and 19A each resolve the large scale population structure of U. maydis indicating subpopulations with decreased gene flow. Hence, the genetic diversity of these virulence-related genes largely reflect the demographic history of U. maydis populations.

Sulfone derivatives reduce growth, adhesion and aspartic protease SAP2 gene expression.

Fungal virulence factors represent a strategy for the design of new compounds with effective activities against Candida spp. Dichloromethyl-4-chloro-3-nitrophenylsulfone (named Compound 1) and chlorodibromomethyl-4-hydrazino-3-nitrophenylsulfone (Compound 2) versus Candida albicans virulence factors (SAP2 expression and adhesion to Caco-2 cell line) were investigated. Candida albicans SC5314 and its mutants: Δsap9, Δsap10, Δsap9/10 were used. MICs of the Compounds (concentrated at 0.0313-16 µg/ml) were determined using M27-A3. Percentage of cell inhibition was assessed spectrophotometrically (OD405) after 48 h at 35 °C. The SAP2 expression was analyzed with the use of RT-PCR; relative quantification was normalized against ACT1 in cells grown in YEPD and on Caco-2. Adherence assay of C. albicans to Caco-2 was performed in a 24-well-plate. Compound 1 showed higher activity (% = 100 at 4 µg/ml) than Compound 2 (MIC90 = 16 µg/ml). Dichloromethyl at the para position of the phenyl ring exerted anti-Candidal potential. Under Compound 1, SAP2 was down-regulated in all the strains (P ≤ 0.05). Conversely, SAP2 was over-expressed in Δsap9-10 (untreated cells) compared with the wild-type. The Compounds significantly affected adherence to epithelium (P ≤ 0.05). The tested sulfones interfered with the adhesion of C. albicans cells to the epithelial tissues without affecting their viability after 90-min of incubation. The Compounds' mode of action was attributed to the reduced adhesiveness and the lower SAP2 expression. Saps9-10 play a role in C. albicans adhesion and they can be involved in the sulfone resistance mechanisms.

Evaluation of bakanae disease progression caused by Fusarium fujikuroi in Oryza sativa L.

Bakanae disease caused by Fusarium fujikuroi is an important fungal disease in rice. Among the seven strains isolated from symptomatic rice grains in this study, one strain, FfB14, triggered severe root growth inhibition and decay in the crown and root of rice seedlings. The remaining six strains caused typical Bakanae symptoms such as etiolation and abnormal succulent rice growth. To reveal the relationship between mycelial growth in the infected tissues and Bakanae disease progression, we have established a reliable quantification method using real time PCR that employs a primer pair and dual-labeled probe specific to a unigene encoding F. fujikuroi PNG1 (FfPNG1), which is located upstream of the fumonisin biosynthesis gene cluster. Plotting the crossing point (CP) values from the infected tissue DNAs on a standard curve revealed the active fungal growth of FfB14 in the root and crown of rice seedlings, while the growth rate of FfB20 in rice was more than 4 times lower than FfB14. Massive infective mycelial growth of FfB14 was evident in rice stems and crown; however, FfB20 did not exhibit vigorous growth. Our quantitative evaluation system is applicable for the identification of fungal virulence factors other than gibberellin.

The bZIP Protein MeaB Mediates Virulence Attributes in Aspergillus flavus

LaeA is a fungal specific virulence factor of both plant and human pathogenic fungi. Transcriptional profiles of laeA mutants have been successfully exploited to identify regulatory mechanisms of secondary metabolism in fungi; here we use laeA mutants as tools to elucidate virulence attributes in Aspergillus flavus. Microarray expression profiles of ΔlaeA and over-expression laeA (OE::laeA) were compared to wild type A. flavus. Strikingly, several nitrogen metabolism genes are oppositely mis-regulated in the ΔlaeA and OE::laeA mutants. One of the nitrogen regulatory genes, the bZIP encoding meaB, is up-regulated in ΔlaeA. Significantly, over-expression of meaB (OE::meaB) phenocopies the decreased virulence attributes of a ΔlaeA phenotype including decreased colonization of host seed, reduced lipase activity and loss of aflatoxin B1 production in seed. However, a double knock-down of laeA and meaB (KD::laeA,meaB) demonstrated that KD::laeA,meaB closely resembled ΔlaeA rather than wild type or ΔmeaB in growth, aflatoxin biosynthesis and sclerotia production thus suggesting that meaB does not contribute to the ΔlaeA phenotype. MeaB and LaeA appear to be part of regulatory networks that allow them to have both shared and distinct roles in fungal biology.

Biochemical Characterization of Paracoccidioides brasiliensis α-1,3-Glucanase Agn1p, and Its Functionality by Heterologous Expression in Schizosaccharomyces pombe

α-1,3-Glucan is present as the outermost layer of the cell wall in the pathogenic yeastlike (Y) form of Paracoccidioides brasiliensis. Based on experimental evidence, this polysaccharide has been proposed as a fungal virulence factor. To degrade α-1,3-glucan and allow remodeling of the cell wall, α-1,3-glucanase is required. Therefore, the study of this enzyme, its encoding gene, and regulatory mechanisms, might be of interest to understand the morphogenesis and virulence process in this fungus. A single gene, orthologous to other fungal α-1,3-glucanase genes, was identified in the Paracoccidioides genome, and labeled AGN1. Transcriptional levels of AGN1 and AGS1 (α-1,3-glucan synthase-encoding gene) increased sharply when the pathogenic Y phase was cultured in the presence of 5% horse serum, a reported booster for cell wall α-1,3-glucan synthesis in this fungus. To study the biochemical properties of P. brasiliensis Agn1p, the enzyme was heterologously overexpressed, purified, and its activity profile determined by means of the degradation of carboxymethyl α-1,3-glucan (SCMG, chemically modified from P. brasiliensis α-1,3-glucan), used as a soluble substrate for the enzymatic reaction. Inhibition assays, thin layer chromatography and enzymatic reactions with alternative substrates (dextran, starch, chitin, laminarin and cellulose), showed that Agn1p displays an endolytic cut pattern and high specificity for SCMG. Complementation of a Schizosaccharomyces pombe agn1Δ strain with the P. brasiliensis AGN1 gene restored the wild type phenotype, indicating functionality of the gene, suggesting a possible role of Agn1p in the remodeling of P. brasiliensis Y phase cell wall. Based on amino acid sequence, P. brasiliensis Agn1p, groups within the family 71 of fungal glycoside hydrolases (GH-71), showing similar biochemical characteristics to other members of this family. Also based on amino acid sequence alignments, we propose a subdivision of fungal GH-71 into at least five groups, for which specific conserved sequences can be identified.

Identifying pathogenicity genes in the rubber tree anthracnose fungus Colletotrichum gloeosporioides through random insertional mutagenesis

To gain more insight into the molecular mechanisms of Colletotrichum gloeosporioides pathogenesis, Agrobacterium tumefaciens-mediated transformation (ATMT) was used to identify mutants of C. gloeosporioides impaired in pathogenicity. An ATMT library of 4128 C. gloeosporioides transformants was generated. Transformants were screened for defects in pathogenicity with a detached copper brown leaf assay. 32 mutants showing reproducible pathogenicity defects were obtained. Southern blot analysis showed 60.4% of the transformants had single-site T-DNA integrations. 16 Genomic sequences flanking T-DNA were recovered from mutants by thermal asymmetric interlaced PCR, and were used to isolate the tagged genes from the genome sequence of wild-type C. gloeosporioides by Basic Local Alignment Search Tool searches against the local genome database of the wild-type C. gloeosporioides. One potential pathogenicity genes encoded calcium-translocating P-type ATPase. Six potential pathogenicity genes had no known homologs in filamentous fungi and were likely to be novel fungal virulence factors. Two putative genes encoded Glycosyltransferase family 28 domain-containing protein and Mov34/MPN/PAD-1 family protein, respectively. Five potential pathogenicity genes had putative function matched with putative protein of other Colletotrichum species. Two known C. gloeosporioides pathogenicity genes were also identified, the encoding Glomerella cingulata hard-surface induced protein and C. gloeosporioides regulatory subunit of protein kinase A gene involved in cAMP-dependent PKA signal transduction pathway.

Brandpilz mit Pep: Ustilago maydis und das pflanzliche Immunsystem

The basidiomycete Ustilago maydis causes smut disease on maize, which is characterized by plant tumors that develop on all aerial parts of the host plant. As a biotrophic pathogen, U. maydis exclusively colonizes living host tissue and therefore depends on the efficient suppression of plant defense responses. Our research aims to understand this modulation of the plant immune system both by endogenous plant factors as well as by secreted fungal virulence factors, termed effector proteins.

Susceptibility of Intact Germinating Arabidopsis thaliana to Human Fungal Pathogens Cryptococcus neoformans and C. gattii

The fungus Cryptococcus contributes a large global burden of infectious death in both HIV-infected and healthy individuals. As Cryptococcus is an opportunistic pathogen, much of the evolutionary pressure shaping virulence occurs in environments in contact with plants and soil. The present studies investigated inoculation of intact seeds of the common weed Arabidopsis thaliana with fungal cells over a 21-day period. C. gattii was the more virulent plant pathogen, resulting in disrupted germination as well as increased stem lodging, fungal burden, and plant tissue colocalization. C. neoformans was a less virulent plant pathogen but exhibited prolonged tissue residence within the cuticle and vascular spaces. Arabidopsis mutants of the PRN1 gene, which is involved in abiotic and biotic signaling affecting phenylalanine-derived flavonoids, showed altered susceptibility to cryptoccocal infections, suggesting roles for this pathway in cryptococcal defense. The fungal virulence factor laccase was also implicated in plant pathogenesis, as a cryptococcal lac1Δ strain was less virulent than wild-type fungi and was unable to colonize seedlings. In conclusion, these studies expand knowledge concerning the ecological niche of Cryptococcus by demonstrating the pathogenic capacity of the anamorphic form of cryptococcal cells against healthy seedlings under physiologically relevant conditions. In addition, an important role of laccase in plant as well as human virulence may suggest mechanisms for laccase retention and optimization during evolution of this fungal pathogen.

A Peptide Derived from the Highly Conserved Protein GAPDH Is Involved in Tissue Protection by Different Antifungal Strategies and Epithelial Immunomodulation

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) plays an important role in glycolysis but also in non-metabolic processes, including transcription activation and apoptosis. We report the isolation of an hGAPDH (2-32) fragment peptide from human placental tissue exhibiting antimicrobial activity. The peptide was internalized by cells of the pathogenic yeast Candida albicans and initiated a rapid apoptotic mechanism, leading to killing of the fungus. Killing was dose-dependent, with 10 µg/ml (3.1 µM) and 100 µg/ml hGAPDH (2-32) depolarizing 45% and 90% of the fungal cells in a population, respectively. Experimental C. albicans infection induced epithelial hGAPDH (2-32) expression. Addition of the peptide significantly reduced the tissue damage as compared to untreated experimental infection. Secreted aspartic proteinases (Saps) activity of C. albicans was inhibited by the fragment at higher concentrations with an ED50 of 160 mg/l (50 μM) for Sap1p and 200 mg/l (63 μM) for Sap2p while Sap3 was not inhibited at all. Interestingly, hGAPDH (2-32) induced significant epithelial IL-8 and GM-CSF secretion and stimulated TLR4 expression at low concentrations independently of the presence of C. albicans without any toxic mucosal effects.In the future, the combination of different antifungal strategies, e.g. a conventional fungicidal with immunomodulatory effects and the inhibition of fungal virulence factors might be a promising treatment option.

Approaching the Functional Annotation of Fungal Virulence Factors Using Cross-Species Genetic Interaction Profiling

In many human fungal pathogens, genes required for disease remain largely unannotated, limiting the impact of virulence gene discovery efforts. We tested the utility of a cross-species genetic interaction profiling approach to obtain clues to the molecular function of unannotated pathogenicity factors in the human pathogen Cryptococcus neoformans. This approach involves expression of C. neoformans genes of interest in each member of the Saccharomyces cerevisiae gene deletion library, quantification of their impact on growth, and calculation of the cross-species genetic interaction profiles. To develop functional predictions, we computed and analyzed the correlations of these profiles with existing genetic interaction profiles of S. cerevisiae deletion mutants. For C. neoformans LIV7, which has no S. cerevisiae ortholog, this profiling approach predicted an unanticipated role in the Golgi apparatus. Validation studies in C. neoformans demonstrated that Liv7 is a functional Golgi factor where it promotes the suppression of the exposure of a specific immunostimulatory molecule, mannose, on the cell surface, thereby inhibiting phagocytosis. The genetic interaction profile of another pathogenicity gene that lacks an S. cerevisiae ortholog, LIV6, strongly predicted a role in endosome function. This prediction was also supported by studies of the corresponding C. neoformans null mutant. Our results demonstrate the utility of quantitative cross-species genetic interaction profiling for the functional annotation of fungal pathogenicity proteins of unknown function including, surprisingly, those that are not conserved in sequence across fungi.

DFVF: database of fungal virulence factors

Fungal pathogens cause various diseases for plant and animal hosts. Despite the extensive impact of fungi on human health and life, the threats posed by emerging fungal pathogens are poorly understood. Specifically, there exist few fungal virulence gene databases, which prevent effective bioinformatics studies on fungal pathogens. Therefore, we constructed a comprehensive online database of known fungal virulence factors, which collected 2058 pathogenic genes produced by 228 fungal strains from 85 genera. This database creates a pivotal platform capable of stimulating and facilitating further bench studies on fungal pathogens.Database URL: http://sysbio.unl.edu/DFVF/

Alternative approaches to antifungal therapies

The expansive use of immunosuppressive medications in fields such as transplantational medicine and oncology, the higher frequency of invasive procedures in an ageing population and the HIV/AIDS pandemic have increased the frequency of systemic fungal infections. At the same time, increased resistance of pathogenic fungi to classical antifungal agents has led to sustained research efforts targeting alternative antifungal strategies. In this review, we focus on two promising approaches: cationic peptides and the targeting of fungal virulence factors. Cationic peptides are small, predominantly positively charged protein fragments that exert direct and indirect antifungal activities, one mechanism of action being the permeabilization of the fungal membrane. They include lysozyme, defensins and cathelicidins as well as novel synthetic peptides. Among fungal virulence factors, the targeting of candidal secreted aspartic proteinases seems to be a particularly promising approach.