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  • Nitric oxide homeostasis is required for light-dependent regulation of conidiation in Aspergillus
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2020-01-25
    Ana T. Marcos; María S. Ramos; Thorsten Schinko; Joseph Strauss; David Cánovas

    Nitric oxide (NO) can be biologically synthesized from nitrite or from arginine. Although NO is involved as a signal in many biological processes in bacteria, plants, and mammals, still little is known about the role of NO in fungi. Here we show that NO levels are regulated by light as an environmental signal in Aspergillus nidulans. The flavohaemoglobin-encoding fhbB gene involved in NO oxidation to nitrate, and the arginine-regulated arginase encoded by agaA, which controls the intracellular concentration of arginine, are both up-regulated by light. The phytochrome fphA is required for the light-dependent induction of fhbB and agaA, while the white-collar gene lreA acts as a repressor when arginine is present in the media. The intracellular arginine pools increase upon induction of both developmental programs (conidiation and sexual development), and the increase is higher under conditions promoting sexual development. The presence of low concentrations of arginine does not affect the light-dependent regulation of conidiation, but high concentrations of arginine overrun the light signal. Deletion of fhbB results in the partial loss of the light regulation of conidiation on arginine and on nitrate media, while deletion of fhbA only affects the light regulation of conidiation on nitrate media. Our working model considers a cross-talk between environmental cues and intracellular signals to regulate fungal reproduction.

    更新日期:2020-01-26
  • The novel Huntiella omanensis mating gene, MAT1-2-7, is essential for ascomatal maturation
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2020-01-17
    Andi M. Wilson; P. Markus Wilken; Magriet A. van der Nest; Michael J. Wingfield; Brenda D. Wingfield

    Sexual reproduction is a highly conserved feature of the eukaryotes, yet sexual compatibility is determined by a wide variety of mechanisms. In ascomycete fungi, sexual development is controlled by genes at the mating type (MAT) locus that confer either MAT1-1 or MAT1-2 mating identity. Although the locus harbours, at minimum, a single gene, the individual MAT loci of certain species, including Huntiella omanensis, encode for two or more genes. The MAT1-2 idiomorph of H. omanensis is made up of MAT1-2-1, a primary MAT gene that is highly conserved in the Pezizomycotina and possesses a well-characterized DNA binding motif, the HMG-box domain. The idiomorph also harbours a novel secondary MAT gene, named MAT1-2-7, with no recognizable functional domains. In this study, we developed a transformation and CRISPR-Cas9-based genome editing protocol to characterize the MAT1-2-7 gene with respect to its function in mating. We have shown that MAT1-2-7 is essential for sexual reproduction and that isolates carrying the truncated MAT1-2-7 gene are incapable of ascomatal maturation and further sexual development. MAT1-2-7 was also shown to influence the vegetative radial growth rate of H. omanensis, illustrating the pleiotropic effects often associated with MAT genes.

    更新日期:2020-01-17
  • Increased activity of the sterol branch of the mevalonate pathway elevates glycosylation of secretory proteins and improves antifungal properties of Trichoderma atroviride
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2020-01-17
    Sebastian Graczyk; Urszula Perlińska-Lenart; Wioletta Górka-Nieć; Renata Lichota; Sebastian Piłsyk; Patrycja Zembek; Jacek Lenart; Przemysław Bernat; Elżbieta Gryz; Justyna Augustyniak; Grażyna Palamarczyk; Joanna S. Kruszewska

    Some Trichoderma spp. have an ability to inhibit proliferation of fungal plant pathogens in the soil. Numerous compounds with a proven antifungal activity are synthesized via the terpene pathway. Here, we stimulated the activity of the mevalonate pathway in T. atroviride P1 by expressing the Saccharomyces cerevisiae ERG20 gene coding for farnesyl pyrophosphate (FPP) synthase, a key enzyme of this pathway. ERG20-expressing Trichoderma strains showed higher activities of FPP synthase and squalene synthase, the principal recipient of FPP in the mevalonate pathway. We also observed activation of dolichyl phosphate mannose (DPM) synthase, an enzyme in protein glycosylation, and significantly increased O- and N-glycosylation of secreted proteins. The hyper-glycosylation of secretory hydrolases could explain their increased activity observed in the ERG20 transformants. Analysis of the antifungal properties of the new strains revealed that the hydrolases secreted by the transformants inhibited growth of a plant pathogen, Pythium ultimum more efficiently compared to the control strain. Consequently, the biocontrol activity of the transgenic strains, determined as their ability to protect bean seeds and seedlings against harmful action of P. ultimum, was also improved substantially.

    更新日期:2020-01-17
  • Comparative genomic and transcriptomic analyses reveal different pathogenicity-related genes among three eucalyptus fungal pathogens
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2020-01-08
    Samuel A. Santos; Pedro M.P. Vidigal; Amali Thrimawithana; Blanca M.L. Betancourth; Lúcio M.S. Guimarães; Matthew D. Templeton; Acelino C. Alfenas

    Ceratocystis fimbriata is an important plant pathogen known to cause Ceratocystis Wilt (CW), a prevalent fungal disease known to affect Eucalyptus spp. plantations in Brazil. To better understand the molecular mechanisms related to pathogenicity in eucalyptus, we generated a high-quality assembly and annotation of the Ce. fimbriata LPF1912 isolate (LPF1912) genome, as well as the first transcriptome of LPF1912 from 16 eucalyptus clones at three infection incubation periods (12, 18, and 24h). The LPF1912 genome assembly contains 805 scaffolds, totaling 31.8 Mb, with 43% of the genome estimated to be coding sequence comprised of 7,390 protein-coding genes of which 626 (8.5%) were classified as secreted proteins, 120 ribosomal RNAs, and 532 transfer RNAs. Comparative genomic analysis among three eucalyptus fungal pathogens (Ce. fimbriata, Ce. eucalypticola, and Calonectria pseudoreteaudii), showed high similarity in the proteome (21.81%) and secretome (52.01%) of LPF1912 and Ce. eucalypticola. GO annotation of pathogenicity-related genes of LPF1912 and Ce. eucalypticola, revealed enrichment in cell wall degrading enzymes (CWDEs), and lipid/cutin metabolism for Ca. pseudoreteaudii. Additionally, a transcriptome analysis between resistant and susceptible eucalyptus clones to CW infection indicated that a majority (11) of LPF1912 differentially expressed genes had GO terms associated with enzymatic functions, such as the polygalacturonase gene family, confirming the crucial role of CWDEs for Ce. fimbriata pathogenicity. Finally, our genomic and transcriptomic analysis approach provides a better understanding of the mechanisms involved in Ce. fimbriata pathogenesis, as well as a framework for further studies.

    更新日期:2020-01-08
  • Yeasts as probiotics: mechanisms, outcomes, and future potential
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2020-01-07
    Swastik Sen; Thomas J. Mansell

    The presence of commensal fungal species in the human gut indicates that organisms from this kingdom have the potential to benefit the host as well. Saccharomyces boulardii, a yeast strain isolated about a hundred years ago, is the most well-characterized probiotic yeast. Though for the most part it genetically resembles Saccharomyces cerevisiae, specific phenotypic differences make it better suited for the gut microenvironment such as better acid and heat tolerance. Several studies using animal hosts suggest that S. boulardii can be used as a biotherapeutic for use in humans. Clinical trials in humans indicate that it can alleviate symptoms from gastrointestinal (GI) tract infections to some extent, but further trials are needed to understand the full therapeutic potential of S. boulardii. Improvement on probiotic function using engineered yeast is an attractive future direction, though genome modification tools for use in S. boulardii have been limited until recently. However, some tools available for S. cerevisiae should be applicable for S. boulardii as well. In this review, we summarize the observed probiotic effect of this yeast and the state of the art for genome engineering tools that could help enhance its probiotic properties.

    更新日期:2020-01-07
  • Rab GDP-dissociation inhibitor gdiA is an essential gene required for cell wall chitin deposition in Aspergillus niger
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-12-27
    Tim M. van Leeuwe; Anne Gerritsen; Mark Arentshorst; Peter J. Punt; Arthur F.J. Ram

    The cell wall is a distinctive feature of filamentous fungi, providing them with structural integrity and protection from both biotic and abiotic factors. Unlike plant cell walls, fungi rely on structurally strong hydrophobic chitin core for mechanical strength together with alpha- and beta-glucans, galactomannans and glycoproteins. Cell wall stress conditions are known to alter the cell wall through the signaling cascade of the cell wall integrity (CWI) pathway and can result in increased cell wall chitin deposition. A previously isolated set of Aspergillus niger cell wall mutants was screened for increased cell wall chitin deposition. UV-mutant RD15.8#16 was found to contain approximately 60% more cell wall chitin than the wild type. In addition to the chitin phenotype, RD15.8#16 exhibits a compact colony morphology and increased sensitivity towards SDS. RD15.8#16 was subjected to classical genetic approach for identification of the underlying causative mutation, using co-segregation analysis and SNP genotyping. Genome sequencing of RD15.8#16 revealed eight SNPs in open reading frames (ORF) which were individually checked for co-segregation with the associated phenotypes, and showed the potential relevance of two genes located on chromosome IV. In situ re-creation of these ORF-located SNPs in a wild type background, using CRISPR/Cas9 genome editing, showed the importance Rab GTPase dissociation inhibitor A (gdiA) for the phenotypes of RD15.8#16. An alteration in the 5’ donor splice site of gdiA reduced pre-mRNA splicing efficiency, causing aberrant cell wall assembly and increased chitin levels, whereas gene disruption attempts showed that a full gene deletion of gdiA is lethal.

    更新日期:2019-12-27
  • LmCBP1, a secreted chitin-binding protein, is required for the pathogenicity of Leptosphaeria maculans on Brassica napus
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-12-19
    Fei Liu; Carrie Selin; Zhongwei Zou; W.G. Dilantha Fernando

    Leptosphaeria maculans is the causal agent of blackleg disease on Brassica napus. Determining the underlying functions of genes required for pathogenesis is essential for understanding the infection process. A chitin-binding protein (LmCBP1) was discovered as a pathogenicity factor for the infection of B. napus by L. maculans through gene knockout using the CRISPR-Cas9 system. Chitin-binding activity was demonstrated through a chitin-protein binding assay. A secreted signal peptide was detected using a yeast secreted-signal peptide trap assay. An increased expression level during the infection stage was also observed, suggesting that LmCBP1 is a secreted protein. The knockout mutants showed decreased infection on B. napus, with reduced pathogenicity on ten cultivars with/without diverse R genes. The mutants were more sensitive to H2O2 compared to wild type blackleg isolate JN3. This study provides evidence of the virulence of a novel chitin-binding protein LmCBP1 on B. napus through mutants created via the CRISPR-Cas9 system.

    更新日期:2019-12-19
  • Role of the XylA gene, encoding a cell wall degrading enzyme, during common wheat, durum wheat and barley colonization by Fusarium graminearum
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-12-11
    F. Tini, G. Beccari, A.H. Benfield, D.M. Gardiner, L. Covarelli

    Fusarium graminearum is the main causal agent of fusarium head blight (FHB) of wheat and barley. This filamentous fungus is able to produce hydrolytic enzymes, such as xylanases, that cause cell wall degradation, permitting host colonization. This study investigated the role of the F. graminearum XylA (FGSG_10999) gene during infection, using a knockout mutant in strain CS3005. Assays were carried out on common wheat, durum wheat and barley to compare virulence of a XylA knockout to that of wild type strain. These assays were conducted on wheat and barley seedling roots, seedling stem bases and heads. Furthermore, additional in vitro experiments were conducted to investigate the role of XylA gene in the utilisation of D-xylose, the main component of cereals cell wall. In planta assays showed the importance of XylA gene for F. graminearum virulence towards its main hosts. A positive correlation between symptom incidence and fungal biomass development was also observed for both the wild type and the knockout strains. Finally, gene expression studies performed in a liquid medium enriched with D-xylose, a known xylanase inducer in other fungi, showed that the absence of the gene in the FGSG_10999 locus was not compensated by two other F. graminearum xylanase encoding genes analysed (loci FGSG_06445 and FGSG_11478).

    更新日期:2019-12-11
  • Gain and loss of a transcription factor that regulates late trichothecene biosynthetic pathway genes in Fusarium
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-12-11
    Daren W. Brown, Alessandra Villani, Antonia Susca, Antonio Moretti, Guixia Hao, Hye-Seon Kim, Robert H. Proctor, Susan P. McCormick

    Trichothecenes are among the mycotoxins of most concern to food and feed safety and are produced by species in two lineages of Fusarium: the F. incarnatum-equiseti (FIESC) and F. sambucinum (FSAMSC) species complexes. Previous functional analyses of the trichothecene biosynthetic gene (TRI) cluster in members of FSAMSC indicate that the transcription factor gene TRI6 activates expression of other TRI cluster genes. In addition, previous sequence analyses indicate that the FIESC TRI cluster includes TRI6 and another uncharacterized transcription factor gene (hereafter TRI21) that was not reported in FSAMSC. Here, gene deletion analysis indicated that in FIESC TRI6 functions in a manner similar to FSAMSC, whereas TRI21 activated expression of some genes that function late in the trichothecene biosynthetic pathway but not early-pathway genes. Consistent with this finding, TRI21 was required for formation of diacetoxyscripenol, a late-trichothecene-pathway product, but not for isotrichodermin, an early-pathway product. Although intact homologs of TRI21 were not detected in FSAMSC or other trichothecene-producing fungal genera, TRI21 fragments were detected in some FSAMSC species. This suggests that the gene was acquired by Fusarium after divergence from other trichothecene-producing fungi, was subsequently lost in FSAMSC, but was retained in FIESC. Together, our results indicate fundamental differences in regulation of trichothecene biosynthesis in FIESC versus FSAMSC.

    更新日期:2019-12-11
  • Developing a tetO/TetR system in Neurospora crassa
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-12-09
    Tinh-Suong Nguyen, Eugene Gladyshev

    The development of a tetO/TetR system in the fungus Neurospora crassa is described. The system includes (i) a synthetic gene encoding a TetR variant fused to GFP, and (ii) a standard tetO array integrated homologously, as a proof of principle, near the his-3 gene. The localization of TetR-GFP at the tetO array (observed by fluorescence microscopy) can be disrupted by the application of tetracycline. The full-length array is stable during vegetative growth, but it triggers strong repeat-induced point mutation (RIP) by the RID-dependent as well as the DIM-2-dependent pathways during the sexual phase. Thus, both RIP pathways must be inactivated to allow the faithful inheritance of the unmodified construct. In summary, this study introduces a new molecular tool into Neurospora research, and suggests that the standard tetO array can self-engage in recombination-independent homologous pairing.

    更新日期:2019-12-09
  • CLR1 and CLR2 are light dependent regulators of xylanase and pectinase genes in Trichoderma reesei
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-12-06
    Sabrina Beier, Wolfgang Hinterdobler, Hoda Bazafkan, Lukas Schillinger, Schmoll Monika
    更新日期:2019-12-07
  • Fusaristatin A production negatively affects the growth and aggressiveness of the wheat pathogen Fusarium pseudograminearum
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-12-04
    Mohammed Khudhair, Kemal Kazan, Louise F. Thatcher, Friday Obanor, Anca Rusu, Jens L. Sørensen, Rasmus D. Wollenberg, Alan McKay, Danièle Giblot-Ducray, Steven Simpfendorfer, Elizabeth Aitken, Donald M. Gardiner

    Fusarium pseudograminearum (Fp), the causative fungal pathogen of the diseases Fusarium crown rot, is an important constraint to cereals production in many countries including Australia. Fp produces a number of secondary metabolites throughout its life cycle. One of these metabolites, the cyclic lipopeptide fusaristatin A, is encoded by a specific gene cluster containing a polyketide synthase and a three-module non-ribosomal peptide synthetase. However, a recent survey of Fp populations across Australia suggests that this cluster may only be present in a subset of isolates from Western Australia (WA). In this study, we screened 319 Fp isolates from WA and 110 Fp isolates from the Australian eastern states of New South Wales, Victoria, Queensland and South Australia to examine the distribution of this gene cluster among Australian Fp populations. The fusaristatin A gene cluster was found to be present in ∼50% of Fp isolates from WA but completely absent in Fp isolates from eastern states. To determine its potential function, mutants of the fusaristatin A gene cluster were generated by disrupting the non-ribosomal peptide synthetase and polyketide synthase genes simultaneously in two different parental backgrounds. The mutants showed increased growth rates and were significantly more aggressive than their respective parental strains on wheat in crown rot pathogenicity assays. This suggested that fusaristatin A has a negative effect on fungal development and aggressiveness. The possible reasons for the geographically restricted presence of the fusaristatin A gene cluster and its role in fungal biology are discussed.

    更新日期:2019-12-04
  • The construction of CRISPR-Cas9 system for endophytic Phomopsis liquidambaris and its PmkkA-deficient mutant revealing the effect on rice
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-11-22
    Peng-Wei Huang, Qian Yang, Ya-Li Zhu, Jun Zhou, Kai Sun, Yan-Zhen Mei, Chuan-Chao Dai

    The endophytic fungus Phomopsis liquidambaris efficiently promotes the nitrogen metabolism and growth of host plants such as rice and peanut. However, a lack of genetic tools limits further research regarding the mechanisms of interaction between P. liquidambaris and its host plants. Herein, a CRISPR/Cas9 system for targeted gene disruption in this strain was first constructed and optimized. The knock-out efficiency increased to over 60% when the ku70 or ku80 gene (involved in nonhomologous end-joining, NHEJ) was disrupted. Furthermore, the CRISPR/Cas9 system was applied to disrupt the PmkkA gene, encoding a mitogen-activated protein kinase kinase (MAPKK) in the cell-wall integrity (CWI) MAPK pathway of the strain. The ΔPmkkA mutant strain induced higher ROS production, chitinase activity and glucanase activity in rice seedlings than wild-type P. liquidambaris (WT), resulting in growth inhibition and strong resistance on rice. These results suggested that the PmkkA gene is crucial during the interaction with rice and may play a role in inhibiting the immune system of host plants. The CRISPR-Cas9 system will be of great use for the study of the interaction between P. liquidambaris and its host plants.

    更新日期:2019-11-22
  • The MAPK Hog1 mediates the response to amphotericin B in Candida albicans
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-11-19
    José Pedro Guirao-Abad, Ruth Sánchez-Fresneda, Elvira Román, Jesús Pla, Juan Carlos Argüelles, Rebeca Alonso-Monge

    The HOG MAP kinase pathway plays a crucial role in the response to different stresses in the opportunistic pathogen Candida albicans. The polyene amphotericin B (AMB) has been reported to trigger oxidative stress in several pathogenic fungi, including C. albicans. In the present work, we have analyzed the role of the MAPK Hog1 in sensing and survival to AMB treatment. Mutants lacking Hog1 are more susceptible to AMB than their parental strains and Hog1 became phosphorylated in the presence of this polyene. A set of mutated versions of Hog1 revealed that both the kinase activity and phosphorylation of Hog1 are required to cope with AMB treatment. Flow cytometry analysis showed that AMB induced intracellular ROS accumulation in both parental and hog1 null mutant strains. In addition, AMB triggered a Hog1-independent synthesis of trehalose. The addition of rotenone to AMB-treated cells improved cell viability, decreased intracellular ROS and prevented intracellular trehalose accumulation, suggesting that AMB-induced ROS is associated to a functional electron transport chain but the presence of rotenone did not impair Hog1 phosphorylation in AMB-treated cells. Our results indicate that Hog1 is necessary during AMB treatment to increase its survival.

    更新日期:2019-11-19
  • Identification of milRNAs and their target genes in Ganoderma lucidum by high-throughput sequencing and degradome analysis
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-11-18
    Junjie Shao, Liqiang Wang, Yang Liu, Qianru Qi, Bin Wang, Shanfa Lu, Chang Liu

    MicroRNAs (miRNAs in animals and plants or milRNAs in fungi) are endogenous noncoding RNAs that can regulate gene expression. However, little information is known about milRNAs and their target genes in Ganoderma lucidum. Here, we systematically predicted and characterised the milRNAs and their target genes across the three developmental stages of G. lucidum. A total of 168 unique milRNAs were predicted using a small RNA sequencing method. For them, 1,612 target sequences corresponding to 1,311 unique genes were predicted by degradome sequencing. We selected 42 predicted milRNAs and performed RT-PCR amplification and Sanger sequencing of the products. Five products were found to have sequences similar to those predicted, confirming the presence of milRNAs in G. lucidum, but demonstrating the difficulty in their validation. Among the 168 milRNAs, 111 were found to be significantly differentially expressed across the three developmental stages (q ≤ 0.05). The expression levels of 12 milRNAs were measured by stem-loop quantitative real-time polymerase chain reaction. Eight of them were in line with the sequencing results (r ≥ 0.9, p ≤ 0.05). These 12 milRNAs and their target genes form 16 milRNA-target gene pairs. The expression profiles of 8 of these 16 miRNA-target pairs were negatively correlated, according to real-time quantitative analysis, whereas the other eight pairs were positively correlated. Furthermore, the results of functional enrichment analysis showed that the target genes of milRNAs mapped to the Gene Ontology terms ‘GTP binding’ and ‘FAD binding’ were enriched in specific developmental stages. These target genes were related to the biosynthesis of triterpenes and polysaccharides and lignin degradation pathway in G. lucidum. In summary, this study has indicated that milRNAs may play crucial regulatory roles in various biological processes of G. lucidum for the first time and open up new avenues for research on milRNAs’ biosyntheses and functions in basidiomycetes.

    更新日期:2019-11-18
  • DsEcp2-1 is a polymorphic effector that restricts growth of Dothistroma septosporum in pine
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-11-12
    Yanan Guo, Lukas Hunziker, Carl.H. Mesarich, Pranav Chettri, Pierre-Yves Dupont, Rebecca J. Ganley, Rebecca L. McDougal, Irene Barnes, Rosie E. Bradshaw

    The detrimental effect of fungal pathogens on forest trees is an increasingly important problem that has implications for the health of our planet. Despite this, the study of molecular plant-microbe interactions in forest trees is in its infancy, and very little is known about the roles of effector molecules from forest pathogens. Dothistroma septosporum causes a devastating needle blight disease of pines, and intriguingly, is closely related to Cladosporium fulvum, a tomato pathogen in which pioneering effector biology studies have been carried out. Here, we studied D. septosporum effectors that are shared with C. fulvum, by comparing gene sequences from global isolates of D. septosporum and assessing effector function in both host and non-host plants. Many of the effectors were predicted to be non-functional in D. septosporum due to their pseudogenization or low expression in planta, suggesting adaptation to lifestyle and host. Effector sequences were polymorphic among a global collection of D. septosporum isolates, but there was no evidence for positive selection. The DsEcp2-1 effector elicited cell death in the non-host plant Nicotiana tabacum, whilst D. septosporum DsEcp2-1 mutants showed increased colonization of pine needles. Together these results suggest that DsEcp2-1 might be recognized by an immune receptor in both angiosperm and gymnosperm plants. This work may lead to the identification of plant targets for DsEcp2-1 that will provide much needed information on the molecular basis of gymnosperm-pathogen interactions in forests, and may also lead to novel methods of disease control.

    更新日期:2019-11-13
  • TBRG-1 a ras-like protein in Trichoderma virens involved in conidiation, development, secondary metabolism, mycoparasitism, and biocontrol unveils a new family of Ras-GTPases
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-11-12
    Mitzuko Dautt-Castro, Magnolia Estrada-Rivera, Ignacio Olguin-Martínez, Ma. del Carmen Rocha-Medina, María Auxiliadora Islas-Osuna, Sergio Casas-Flores

    Ras-GTPases are nucleotide hydrolases involved in key cellular processes. In fungi, Ras-GTPases regulate conidiation, development, virulence, and interactions with other fungi or plants. Trichoderma spp. are filamentous saprophytic fungi, widely distributed along all latitudes, characterized by their rapid growth and metabolic diversity. Many species of this genus interact with other fungi, animals or plants. Furthermore, these fungi are used as biocontrol agents due to their ability to antagonize phytopathogenic fungi and oomycetes, through competence, antibiosis, and mycoparasitism. However, the genetic and molecular regulation of these processes is scarcely described in these fungi. In this work, we investigated the role of the gene tbrg-1 product (GenBank accession number XP_013956100; JGI ID: Tv_70852) of T. virens during its interaction with other fungi and plants. Sequence analyses predicted that TBRG-1 bears the characteristic domains of Ras-GTPases; however, its size (1011 aa) is 3- to 4-times bigger compared with classical GTPases. Interestingly, phylogenetic analyses grouped the TBRG-1 protein with hypothetical proteins of similar sizes, sharing conserved regions; whereas other known Ras GTPases were perfectly grouped with their respective families. These facts led us to classify TBRG-1 into a new family of Ras-GTPases, the Big Ras GTPases. Therefore, the gene was named tbrg-1 (Trichoderma Big Ras GTPase). Quantification of conidia and scanning electron microscopy showed that the mutants-lacking tbrg-1 produced less conidia, as well as a delayed conidiophore development compared to the wild-type (wt). Moreover, a deregulation of conidiation-related genes (con-10, con-13, and stuA) was observed in tbrg-1-lacking strains, which indicates that TBRG-1 is necessary for proper conidiophore and conidia development. Furthermore, the lack of tbrg-1 affected positively the antagonistic capability of T. virens against the phytopathogens Rhizoctonia solani, Sclerotium rolfsii, and Fusarium oxysporum, which was consistent with the expression patterns of mycoparasitism-related genes, sp1 and cht1, that code for a protease and for a chitinase, respectively. Furthermore, the antibiosis effect of mycelium-free culture filtrates of Δtbrg-1 against R. solani was considerably enhanced. The expression of secondary metabolism-related genes, particularly gliP, showed an up-regulation in Δtbrg-1, which paralleled an increase in gliotoxin production as compared to the wt. These results indicate that TBRG-1 plays a negative role in secondary metabolism and antagonism. Unexpectedly, the biocontrol activity of Δtbrg-1 was ineffective to protect the tomato seeds and seedlings against R. solani. On the contrary, Δtbrg-1 behaved like a plant pathogen, indicating that TBRG-1 is probably implicated in the recognition process for establishing a beneficial relationship with plants.

    更新日期:2019-11-13
  • The glycoside hydrolase 18 family chitinases are associated with development and virulence in the mosquito pathogen Pythium guiyangense
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-11-07
    Danyu Shen, Jing Wang, Yumei Dong, Meiqian Zhang, Zhaoyang Tang, Qingyue Xia, Karani T. Nyawira, Maofeng Jing, Daolong Dou, Ai Xia

    Chitinases, the enzymes responsible for the biological degradation of chitin, participate in numerous physiological processes such as nutrition, parasitism, morphogenesis and immunity in various organisms. However, the genome-wide distribution, evolution and biological functions of chitinases are rarely reported in oomycetes. This study systematically investigated the glycoside hydrolase 18 (GH18) family of chitinases from the mosquito pathogenic oomycete, Pythium guiyangense using bioinformatics and experimental assays. A total of 3 pairs of GH18 chitinase genes distributed in three distinct phylogenic clusters were identified from P. guiyangense genome, which is consistent with the ones in plant pathogenic oomycetes. Further transcriptional analysis revealed that Pgchi1/2 was highly expressed at the development stages, while Pgchi3/4 and Pgchi5/6 were up-regulated at the infection stages. The biological function analysis of chitinase genes using genetic transformation silencing method showed that silencing of Pgchi1/2 resulted in reduced zoospore production, without affecting the virulence. However, attenuation of Pgchi3/4 and Pgchi5/6 genes regulate not only oxidative stress responses, but also led to decreased infection rates to mosquito larvae. Taken together, this study provides a comprehensive overview of P. guiyangense chitinase family and reveals their diverse roles in the development, stress response, and virulence, which would elucidate insightful information on the molecular mechanism of chitinase in entomopathogenic pathogens.

    更新日期:2019-11-08
  • FpDep1, a component of Rpd3L histone deacetylase complex, is important for vegetative development, ROS accumulation, and pathogenesis in Fusarium pseudograminearum
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-11-06
    Yinshan Zhang, Limin Wang, Shen Liang, Panpan Zhang, Ruijiao Kang, Mengjuan Zhang, Min Wang, Linlin Chen, Hongxia Yuan, Shengli Ding, Honglian Li

    Histone deacetylases (HDACs) play essential roles in modulating chromatin structure to provide accessibility to gene regulators. Increasing evidence has linked HADCs to pathogenesis control in the filamentous plant fungi. However, its function remains unclear in Fusarium pseudograminearum, which has led to the emergence of the disease Fusarium crown rot in China. Here we identified the FpDEP1 gene, an orthologue of Saccharomyces cerevisiae DEP1 encoding a component of the Rpd3 histone deacetylase complex in F. pseudograminearum. The gene deletion 0mutant, ΔFpdep1, showed significantly retarded growth on PDA plates with reduced aerial hyphae formation. Pathogenicity tests displayed no typical leaf lesions and limited expansion capability of coleoptiles. Histopathological analysis indicated the ΔFpdep1 deletion mutant differentiated infectious hyphae and triggered massive reactive oxygen species (ROS) accumulation during the early infection stage, resulting in limited expansion to neighbor cells which was concurring with sensitivity to H2O2 and SDS tests in vitro. FM4-64 staining revealed that the ΔFpdep1 deletion mutant was delayed in endocytosis. The FpDEP1-GFP transgene complemented the mutant phenotypes and the fusion protein co-localized with DAPI staining, indicating that the FpDEP1 gene product is localized to the nucleus in spores and mycelia. Immunoprecipitation coupled with LC-MS/MS and yeast two-hybrid screening identified the Rpd3L-like HDAC complex containing at least FpDep1, FpSds3, FpSin3, FpRpd3, FpRxt3, FpCti6, FpRho23, and FpUme6. These results suggest that FpDep1 is involved in a HDAC complex functioning on fungal development and pathogenesis in F. pseudograminearum.

    更新日期:2019-11-06
  • The roles of FgPEX2 and FgPEX12 in virulence and lipid metabolism in Fusarium graminearum
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-11-05
    Lina Wang, Li Zhang, Chunjie Liu, Shaohua Sun, Aixin Liu, Yuancun Liang, Jinfeng Yu

    Fusarium head blight (FHB) is a wheat disease with a worldwide prevalence, caused by Fusarium graminearum. Peroxisomes are ubiquitous in eukaryotic cells and are involved in various biochemical phenomena. FgPEX2 and FgPEX12 encode RING-finger peroxins PEX2 and PEX12 in F. graminearum. This study aimed to functionally characterize FgPEX2 and FgPEX12 in F. graminearum. We constructed deletion mutants of FgPEX2 and FgPEX12 via homologous recombination. The ΔPEX2 and ΔPEX12 mutants displayed defects in sexual and asexual development, virulence, cell wall integrity (CWI), and lipid metabolism. Deletion of FgPEX2 and FgPEX12 significantly decreased deoxynivalenol production. Furthermore, fluorescence microscopic analysis of the subcellular localization of GFP-PMP70 and GFP-HEX1 revealed that FgPEX2 and FgPEX12 maintain Woronin bodies. These results show that FgPEX2 and FgPEX12 are required for growth, conidiation, virulence, cell wall integrity, and lipid metabolism in F. graminearum and do not influence their peroxisomes.

    更新日期:2019-11-06
  • CgHog1 controls the adaptation to both sorbitol and fludioxonil in Colletotrichum gloeosporioides
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-11-05
    Yangfan Li, Puhuizhong He, Chengming Tian, Yonglin Wang

    The HOG ((high-osmolarity glycerol)) pathway is critical for the appropriate adaptation to adverse conditions. Here, we demonstrated that the deletion of CgHog1 resulted in enhanced sensitivity to osmotic stress and increased resistance to fludioxonil in the poplar anthracnose fungus Colletotrichum gloeosporioides. The accumulation of chitin around hyphal tips was obviously decreased in the ΔCgHog1 strain under sorbitol, whereas it strongly increased in the response to fludioxonil compared with the wild type. To investigate the underlying mechanism of CgHog1-mediated adaption to osmotic stress and fludioxonil, transcriptomic profiles were performed in both the ΔCgHog1 strain and the wild type under the treatment of sorbitol and fludioxonil, respectively. Under sorbitol, genes associated with glycolysis, lipid metabolism, and accumulation of soluble sugars and amino acids were differentially expressed; under fludioxonil, vesicle trafficking-related genes were highly downregulated in the ΔCgHog1 strain, which was consistent with abnormal vacuoles distribution and morphology of hyphae, indicating that the growth defect caused by fludioxonil may be associated with disruption of endocytosis. Taken together, we elucidated the adaptation mechanisms of how CgHog1 regulates appropriate response to sorbitol and fludioxonil via different metabolism pathways. These findings extend our insights into the HOG pathway in fungi.

    更新日期:2019-11-06
  • Septins are required for reproductive propagule development and virulence of the maize pathogen Cochliobolus heterostrophus
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-11-04
    Xianghui Zhang, Jonathan B. González, B. Gillian Turgeon

    Septins are highly conserved GTP-binding proteins that function in cell cytokinesis, polarity and morphogenesis. To evaluate the roles of these proteins in inoculum health and disease, mutants deleted for each of five septin proteins (Cdc3, Cdc10, Cdc11, Cdc12, and Cdc100) were characterized in the ascomycete Cochliobolus heterostrophus for ability to develop asexual and sexual spores and for virulence to the host maize. Strains deleted for CDC3, CDC10, CDC11, and CDC12 genes showed significant changes in hyphal growth, and in development of conidia and ascospores compared to the wild-type strain. Conidia had dramatically reduced numbers of septa and rates of germination, while ascospore development was blocked in the meiotic process. Although asci were produced, wild type ascospores were not. When equal numbers of conidia from wild-type and mutants were used to inoculate maize, cdc10 mutants showed reduced virulence compared to the wild-type strain and other mutants. This reduced virulence was demonstrated to be correlated with lower germination rate of cdc10 mutant conidia. When adjusted for germination rate, virulence was equivalent to the wild-type strain. Double mutants (cdc3cdc10, cdc3cdc11) showed augmented reduced growth phenotypes. CDC100 mutants were wild type in all assays. Taken together, these findings indicate that all four conserved septin proteins play a major role in reproductive propagule formation and that mutants with deletions of CDC10 are reduced in virulence to the host maize.

    更新日期:2019-11-04
  • Class V chitin synthase and β(1,3)-glucan synthase co-travel in the same vesicle in Zymoseptoria tritici
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-10-28
    Martin Schuster, Celia Guiu-Aragones, Gero Steinberg

    The fungal cell wall consists of proteins and polysaccharides, formed by the co-ordinated activity of enzymes, such as chitin or glucan synthases. These enzymes are delivered via secretory vesicles to the hyphal tip. In the ascomycete Neurospora crassa, chitin synthases and β(1,3)-glucan synthase are transported in different vesicles, whereas they co-travel along microtubules in the basidiomycete Ustilago maydis. This suggests fundamental differences in wall synthesis between taxa. Here, we visualize the class V chitin synthase ZtChs5 and the β(1,3)-glucan synthase ZtGcs1 in the ascomycete Zymoseptoria tritici. Live cell imaging demonstrate that both enzymes co-locate to the apical plasma membrane, but are not concentrated in the Spitzenkörper. Delivery involves co-transport along microtubules of the chitin and glucan synthase. Live cell imaging and electron microscopy suggest that both cell wall synthases locate in the same vesicle. Thus, microtubule-dependent co-delivery of cell wall synthases in the same vesicle is found in asco- and basidiomycetes.

    更新日期:2019-10-29
  • “Candida glabrata peroxiredoxins, Tsa1 and Tsa2, and sulfiredoxin, Srx1, protect against oxidative damage and are necessary for virulence”
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-10-22
    Guadalupe Gutiérrez-Escobedo, Oscar Hernández-Carreón, Brenda Morales-Rojano, Brenda Revuelta-Rodríguez, Norma Vázquez-Franco, Irene Castaño, Alejandro De Las Peñas
    更新日期:2019-10-23
  • Genetical and O-glycoproteomic analyses reveal the roles of three protein O-mannosyltransferases in phytopathogen Fusarium oxysporum f.sp. cucumerinum
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-10-21
    Yueqiang Xu, Hui Zhou, Guangya Zhao, Jinghua Yang, Yuanming Luo, Shutao Sun, Zhongfu Wang, Shaojie Li, Cheng Jin

    Protein O-mannosyltransferases (PMTs) have been identified in fungi but not in plants and nematodes, which makes PMTs become attractive targets for developing a new strategy against phytopathogens. Three PMTs have been identified in Fusarium oxysporum, a fungal pathogen that causes vascular wilt in a broad range of economical crops. By deletion or suppression of the pmt genes, we showed that all mutants displayed retarded growth, reduced conidiation, cell wall defects, ER stress and attenuated virulence in F. oxysporum f.sp. cucumerinum. In addition, the Δpmt1 exhibited reduced thermotolerance, while the Δpmt4 and the pmt2 conditional mutant exhibited abnormal polarized growth. Comparative glycoproteome analysis of these pmt mutants revealed that PMTs preferentially modified random coils with flanking regions rich in Ser, Thr, Ala, Glu, Asp and Lys at the stem region of membrane proteins, the N-terminal region close to signal peptide of secreted proteins, or surface of soluble proteins. PMT1 specifically acted on nuclear proteins and proteins that are responsible for protein folding, which might contribute to thermotolerance. PMT4 specifically acted on the membrane and soluble proteins in secretory pathways, especially the GPI anchoring pathway, which might contribute to synthesis and transportation of GPI anchored proteins and thus polarized growth. PMT2 was responsible for modification of proteins that are required for protein folding and cell wall synthesis, which might make PMT2 essential. Our results gave an insight to understanding of the roles of each O-mannosyltransferase in F. oxysporum f.sp. cucumerinum and provide a new perspective to prevent Fusarium wilt.

    更新日期:2019-10-22
  • A comparative study of liquid holding restitution of viability after oxidative stress in Ustilago maydis and Saccharomyces cerevisiae cell populations
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-10-18
    Mira Milisavljevic, Milorad Kojic

    The ability of Saccharomyces cerevisiae to reconstitute viability after strong peroxide-induced oxidative stress during liquid holding (LH) in non-nutrient medium has been compared with that of Ustilago maydis. It was found that like U. maydis, S. cerevisiae was capable of reconstituting viability through multiplication of the survivors. However, differences were observed in the pattern of their response: (i) the reconstitution of viability was slower in S. cerevisiae; (ii) before the viability was progressively increasing the treated samples of this fungus reproducibly passed through a phase of additional decrease of the surviving fraction and (iii) the final yields of viable cells attained in S. cerevisiae were below those achieved by U. maydis. The reason for the relative superiority of U. maydis is twofold: (1) early initiated and faster degradation and leakage of the intracellular biomolecules and (2) greater ability of U. maydis cells to recycle damaged and released intracellular compounds. Conceptually similar studies extended to another oxidative-stress-inducing condition, namely desiccation, indicated that the marked differences between these fungi in their patterns of the post-stress regrowth, cellular leakage and macromolecule decomposition are reproduced during LH of desiccated cells. The concordance of the findings obtained upon these two approaches was also corroborated by an analysis of the post-desiccation LH response of U. maydis mutants (adr1, did4, kel1 and tbp1) that were previously identified as defective in post-peroxide LH restitution of viability. We discuss the findings in terms of their possible relevance to the mechanisms of the ecological and evolutionary adaptation of free-living microorganisms to fluctuating and severely inhospitable environments.

    更新日期:2019-10-19
  • The TRP Ca2+ channel Yvc1 regulates hyphal reactive oxygen species gradient for maintenance of polarized growth in Candida albicans
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-10-16
    Liping Peng, Qilin Yu, Henan Wei, Nali Zhu, Tongtong Ren, Chao Liang, Jiachun Xu, Lei Tian, Mingchun Li
    更新日期:2019-10-17
  • Exploring links between antisense RNAs and pathogenesis in Ustilago maydis through transcript and gene characterization
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-10-16
    Kristi M. Goulet, Emilee R.M. Storfie, Barry J. Saville

    Biotrophic basidiomycete plant pathogens cause billions of dollars in losses to cereal crops annually. The model for this group of fungi is the corn smut pathogen Ustilago maydis. Annotation of its genome identified antisense RNAs (asRNAs) complementary to over half of the coded mRNAs, some of which are present at high levels in teliospores but detected at very low levels or not at all in other cell types, suggesting they have a function in the teliospore or during teliospore formation. Expression of three such asRNAs (as-UMAG_02150, ncRNA1, and as-UMAG_02151) is controlled by two adjacent genomic regions. Deletion of these regions increased transcript levels of all three asRNAs and attenuated pathogenesis. This study investigated the reason for this marked reduction in pathogenesis by: 1) using deletion analyses to assess the involvement of genes, complementary to the asRNAs, in pathogenesis; 2) determining that one of the linked genes encodes a putative xylitol dehydrogenase; and 3) identifying and functionally characterizing asRNAs that could influence expression of protein-coding genes. The results presented suggest that the influence of the asRNAs on pathogenesis occurs through their action at unlinked loci.

    更新日期:2019-10-17
  • Trichoderma harzianum transcriptome in response to cadmium exposure
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-10-15
    Letícia Harumi Oshiquiri, Karina Roterdanny Araújo dos Santos, Sidnei Alves Ferreira Junior, Andrei Stecca Steindorff, Jomal Rodrigues Barbosa Filho, Thuana Marcolino Mota, Cirano José Ulhoa, Raphaela de Castro Georg

    Cadmium (Cd) is a heavy metal present in the environment mainly as a result of industrial contamination that can cause toxic effects to life. Some microorganisms, as Trichoderma harzianum, a fungus used in biocontrol, are able to survive in polluted environments and act as bioremediators. Aspects about the tolerance to the metal have been widely studied in other fungi although there are a few reports about the response of T. harzianum. In this study, we determined the effects of cadmium over growth of T. harzianum and used RNA-Seq to identify significant genes and processes regulated in the metal presence. Cadmium inhibited the fungus growth proportionally to its concentration although the fungus exhibited tolerance as it continued to grow, even in the highest concentrations used. A total of 3,767 (1,993 up and 1,774 down) and 2,986 (1,606 up and 1,380 down) differentially expressed genes were detected in the mycelium of T. harzianum cultivated in the presence of 1.0 mg.mL-1 or 2.0 mg.mL-1 of CdCl2, respectively, compared to the absence of the metal. Of these, 2,562 were common to both treatments. Biological processes related to cellular homeostasis, transcription initiation, sulfur compound biosynthetic and metabolic processes, RNA processing, protein modification and vesicle-mediated transport were up-regulated. Carbohydrate metabolic processes were down-regulated. Pathway enrichment analysis indicated induction of glutathione and its precursor’s metabolism. Interestingly, it also indicated an intense transcriptional induction, especially by up-regulation of spliceosome components. Carbohydrate metabolism was repressed, especially the mycoparasitism-related genes, suggesting that the mycoparasitic ability of T. harzianum could be affected during cadmium exposure. These results contribute to the advance of the current knowledge about the response of T. harzianum to cadmium exposure and provide significant targets for biotechnological improvement of this fungus as a bioremediator and a biocontrol agent.

    更新日期:2019-10-16
  • Improvement of the CRISPR-Cas9 mediated gene disruption and large DNA fragment deletion based on a chimeric promoter in Acremonium chrysogenum
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-10-14
    Chang Chen, Jiajia Liu, Chengbao Duan, Yuanyuan Pan, Gang Liu

    Acremonium chrysogenum has been employed in the industrial production of cephalosporin C (CPC). However, there are still some impediments to understanding the regulation of CPC biosynthesis and improving strains due to the difficulty of genetic manipulation in A. chrysogenum, especially in the CPC high-producing strain C10. Here, an improved CRISPR-Cas9 system was constructed based on an U6/tRNA chimeric promoter. Using this system, high efficiency for single gene disruption was achieved in C10. In addition, double loci were simultaneously targeted when supplying with the homology-directed repair templates (donor DNAs). Based on this system, large DNA fragments up to 31.5 kb for the yellow compound sorbicillinoid biosynthesis were successfully deleted with high efficiency. Furthermore, CPC production was significantly enhanced when the sorbicillinoid biosynthetic genes were knocked out. This study provides a powerful tool for gene editing and strain improvement in A. chrysogenum.

    更新日期:2019-10-15
  • 更新日期:2019-10-14
  • MrMid2, encoding a cell wall stress sensor protein, is required for conidium production, stress tolerance, microsclerotium formation and virulence in the entomopathogenic fungus Metarhizium rileyi
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-10-11
    Caiyan Xin, Xiaorui Xing, Fen Wang, Jiexing Liu, Zhuonan Ran, Wenbi Chen, Guangxi Wang, Zhangyong Song

    Transmembrane proteins as sensors encoded by fungal genes activate specific intracellular signal pathways in response to stress cues to help the fungus survive in a changing environment. To better understand the role of the cell wall integrity (CWI) pathway in the entomopathogenic fungus Metarhizium rileyi, an ortholog encoding the transmembrane protein Mid2, MrMid2, was identified and characterized functionally. Transcriptional analysis indicated that MrMid2 was involved in dimorphic transition, conidiation, and microsclerotium formation. After a targeted deletion of MrMid2, all three traits were impaired. Compared with the wild-type strain, the △MrMid2 mutants were hypersensitive to thermal stress, and cell wall and oxidative stress. Insect bioassays revealed that △MrMid2 mutants had decreased virulence levels following topical (22.5 %) and injection bioassays (38.7 %). Furthermore, transcription analysis showed that other genes of the CWI pathway, with the exception of another major sensor protein encoding gene, MrWsc1, were down-regulated in △MrMid2 mutants. These results suggest that MrMid2 plays important roles in dimorphic transition, conidiation, the stress response, virulence, and microsclerotium development in M. rileyi.

    更新日期:2019-10-12
  • Two 14-3-3 proteins contribute to nitrogen sensing through the TOR and glutamine synthetase-dependent pathways in Fusarium graminearum
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-10-09
    Elizabeth K. Brauer, Nimrat Manes, Christopher Bonner, Rajagopal Subramaniam

    Fusarium graminearum responds to environmental cues to modulate its growth and metabolism during wheat pathogenesis. Nitrogen limitation activates virulence-associated behaviours in F. graminearum including mycotoxin production and penetrative growth. In other filamentous fungi, nitrogen sensing is mediated by both the Target of Rapamycin (TOR) and the glutamine synthetase (GS)-dependent signaling pathways. While TOR-dependent nitrogen responses have been demonstrated in F. graminearum, the involvement of GS remains unclear. Our study indicates that both the TOR and GS signalling pathways are involved in nitrogen sensing in F. graminearum and contribute to glutamine-induced mycelial growth. However, neither pathway is required for glutamine-induced repression of the mycotoxin deoxynivalenol (DON) indicating that an additional nitrogen sensing pathway must exist. Further, two genes FgBMH1 and FgBMH2 encoding 14-3-3 proteins regulate nitrogen responses with effects on gene expression, DON production and mycelial growth. Unlike yeast, where 14-3-3s function redundantly in regulating nitrogen sensing, the 14-3-3 proteins have differing functions in F. graminearum. While both FgBMH1 and FgBMH2 regulate early glutamine-induced DON repression, only FgBMH2 is involved in regulating reproduction, virulence and glutamine-induced AreA repression. Together, our findings help to clarify the nitrogen sensing pathways in F. graminearum and highlight the involvement of 14-3-3s in the nitrogen response of filamentous fungi.

    更新日期:2019-10-10
  • Specialized infection strategies of falcate and oval conidia of Colletotrichum graminicola
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-09-21
    Daniela E. Nordzieke, Alina Sanken, Luis Antelo, Anja Raschke, Holger B. Deising, Stefanie Pöggeler
    更新日期:2019-09-22
  • Increased expression of Phytophthora sojae genes encoding membrane-degrading enzymes appears to suggest an early onset of necrotrophy during Glycine max infection
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-09-10
    Nicholas Grams, Manuel Ospina-Giraldo

    Phytophthora sojae is an oomycete pathogen that causes root, stem, and leaf rot in soybean plants, frequently leading to massive economic losses. Despite its importance, the mechanism by which P. sojae penetrates the host is not yet fully understood. Evidence indicates that P. sojae is not capable of penetrating the plant cell wall via mechanical force, suggesting that alternative factors facilitate breakdown of the host cell wall and membrane. Members of the carbohydrate esterase (CE) family 10 (carboxylesterases, arylesterases, sterol esterases and acetylcholine esterases, collectively known as CE10), are thought to be important for this penetration process. To gain insight into the potential role of CE10-coding genes in P. sojae pathogenesis, the newly revised version of the P. sojae genome was searched for putative CE10-coding genes, and various bioinformatic analyses were conducted using their amino acid and nucleotide sequences. In addition, in planta infection assays were conducted with P. sojae Race 4 and soybean cultivars Williams and Williams 82, and the transcriptional activity of P. sojae CE10-coding genes was evaluated at different time points during infection. Results suggest that these genes are important for both the biotrophic and necrotrophic stages of the P. sojae infection process and provide molecular evidence for stage distinction during infection progression. Furthermore, bioinformatic analyses have identified several conserved gene and protein sequence features that appear to have a significant impact on observed levels of expression during infection. Results agree with previous reports implicating other carbohydrate-active enzymes in P. sojae infection.

    更新日期:2019-09-11
  • Members of the Euwallacea fornicatus species complex exhibit promiscuous mutualism with ambrosia fungi in Taiwan
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-09-10
    Joseph D. Carrillo, Paul F. Rugman-Jones, Deena Husein, Jason E. Stajich, Matt T. Kasson, Daniel Carrillo, Richard Stouthamer, Akif Eskalen

    Carrillo, J.D., Rugman-Jones, PF., Husein, D., Stajich, J.E., Kasson, MT., Carrillo, D., Stouthamer, R., and Eskalen, A. 2019. Members of the Euwallacea fornicatus species complex exhibit promiscuous mutualism with ambrosia fungi in Taiwan A number of ambrosia beetles have come to prominence in recent years because of the damage they inflict on a variety of trees within invaded habitats across the globe. Ambrosia beetles rely on symbiotic microorganisms, mainly fungi, as a dedicated food source and carry those microorganisms around with them within specialized organs termed mycangia. Investigation of members of the Euwallacea fornicatus species complex and their fungal symbionts in Taiwan revealed promiscuous symbioses with ambrosial Fusaria clade (AFC) members, Graphium spp., and Paracremonium spp. based on co-phylogenetic analyses. For AFC members, a novel diagnostic PCR assay targeting mating type genes MAT1-1-1 and MAT1-2-1 was developed and validated by amplicon size and sequencing. Mating types screening of AFC members revealed the isolates screened are all heterothallic (self-sterile), with both MAT types represented and recovered from fungi vectored by E. fornicatus, E. kuroshio, and E. whitfordiodendrus in Taiwan. Members of the Euwallacea fornicatus species complex and the variety of ambrosia fungi they utilize further confirms that their relationship with fungi are more likely promiscuous in native areas, as opposed to strictly obligate to a specific combination of fungi as observed in invaded areas.

    更新日期:2019-09-11
  • Exploring the resistance mechanisms in Trichosporon asahii: triazoles as the last defense for invasive trichosporonosis
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-09-09
    Ana Carolina Barbosa Padovan, Walicyranison Plinio da Silva Rocha, Ana Caroline de Moraes Toti, Daniel Felipe Freitas de Jesus, Guilherme Maranhão Chaves, Arnaldo Lopes Colombo

    Trichosporon asahii has recently been recognized as an emergent fungal pathogen able to cause invasive infections in neutropenic cancer patients as well as in critically ill patients submitted to invasive medical procedures and broad-spectrum antibiotic therapy. T. asahii is the main pathogen associated with invasive trichosporonosis worldwide. Treatment of patients with invasive trichosporonosis remains a controversial issue, but triazoles are mentioned by most authors as the best first-line antifungal therapy. There is mounting evidence supporting the claim that fluconazole (FLC) resistance in T. asahii is emerging worldwide. Since 2000, 15 publications involving large collections of T. asahii isolates described non-wild type isolates for FLC and/or voriconazole. However, very few papers have addressed the epidemiology and molecular mechanism of antifungal resistance in Trichosporon spp. Data available suggest that continuous exposure to azoles can induce mutations in the ERG11 gene, resulting in resistance to this class of antifungal drugs. A recent report characterizing T. asahii azole-resistant strains found several genes differentially expressed and highly mutated, including genes related to the Target of Rapamycin (TOR) pathway, indicating that evolutionary modifications on this pathway induced by FLC stress may be involved in developing azole resistance. Finally, we provided new data suggesting that hyperactive efflux pumps may play a role as drug transporters in FLC resistant T. asahii strains.

    更新日期:2019-09-10
  • A unique multidrug-resistant clonal Trichophyton population distinct from Trichophyton mentagrophytes / Trichophyton interdigitale complex causing an ongoing alarming dermatophytosis outbreak in India: genomic insights and resistance profile
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-09-03
    Ashutosh Singh, Aradhana Masih, Juan Monroy-Nieto, Pradeep Kumar Singh, Jolene Bowers, Jason Travis, Ananta Khurana, David M. Engelthaler, Jacques F. Meis, Anuradha Chowdhary

    There has been a considerable upsurge of extensive, treatment recalcitrant, dermatophytosis presenting as tinea corporis and tinea cruris in India since the past few years. Genome analysis of Trichophyton species causing severe superficial dermatophytosis in North India confirmed a unique clade most recently related to the T. mentagrophytes/interdigitale complex, seeming to belong to an early diverging clade of the complex. The Indian Trichophyton species genomes were highly related showing only up to 42 SNPs between any two isolates confirming their clonal origin. Other genetic approaches such as ITS sequencing and multigene phylogeny used in this study were contradictory or inconclusive to show the differentiation of these isolates from T. mentagrophytes/T. interdigitale. Remarkably, high rates of resistance to all three commonly used oral antifungals, i.e., 36% for terbinafine (MICs 4 to ≥32 mg/L), 39.5% for fluconazole (MIC range 32 to ≥64 mg/L) and griseofulvin (Geometric mean MIC ≥4 mg/L) were observed. Two important amino acid substitutions (Leu393Phe or Phe397Leu) leading to a terbinafine resistant phenotype were found in the squalene epoxidase protein of all tested terbinafine resistant isolates. All 20 examined genomes presented a high mobility group (HMG) domain transcription factor gene corresponding to mating type (+). Of these, three isolates also showed positivity for both alpha-box and HMG in the genome which might indicate hybridization or an incomplete sexual cycle. Therefore, we highlight the potential of this organism to rapidly spread alleles that might be driving antifungal resistance among its population. This new population of Trichophyton with high rates of in vitro antifungal resistance seems to be driving an ongoing outbreak of dermatophytosis in India. Our study highlights difficulties in identifying isolates from the Trichophyton mentagrophytes/interdigitale clade of the genus using currently available molecular tools. High resistance rates of terbinafine warrant further clinical studies to assess its utility in the treatment of dermatophytosis caused by this strain.

    更新日期:2019-09-03
  • Genomic perspective of triazole resistance in clinical and environmental Aspergillus fumigatus isolates without cyp51A mutations
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-08-26
    Cheshta Sharma, Shijulal Nelson-Sathi, Ashutosh Singh, M. Radhakrishna Pillai, Anuradha Chowdhary

    Aspergillus fumigatus is the most common etiologic agent of primarily all clinical manifestations of aspergillosis. A steady increase in the number of azole resistant A. fumigatus (ARAF) isolates from environment and clinical samples leading to therapeutic failures in clinical settings have alarmed the mycologists and clinicians worldwide. Although mutations in azole target cyp51A gene have been implicated in conferring azole resistance in A. fumigatus, recent studies have demonstrated occurrence of azole resistant strains without cyp51A mutations. In this study, next generation sequencing techniques and the expression profiling of transporter genes with single nucleotide polymorphisms (SNPs) in clinical and environmental ARAF isolates with (G54E) and without known cyp51A mutations was undertaken to understand the genetic background and role of transporters in azole resistance. The raw reads of four ARAF strains when mapped to Af293 reference genome (>100X depth) covered at least 93.1% of the reference genome. Among all four strains, a total of 212,711 SNPs was identified with 37,829 were common in at least two isolates. The expression analysis suggested the overexpression of MFS transporter, namely, MfsC in all ARAF isolates. None of the resistant strain showed significant upregulation of cyp51A and cyp51B gene. On the other hand, AbcD was upregulated (5-fold) in the isolates with cyp 51A mutation (G54E). The whole genome sequence analysis showed the presence of two previously described amino acid substitutions S269F and F390Y in HMG1 gene in a clinical panazole resistant strain without cyp51A mutations. These mutations have been previously associated with azole resistance in A. fumigatus strains without cyp51A mutations. Further, several punctual mutations and a large-segment deletion among different strains were observed suggesting the involvement of resistance mechanisms other than cyp51A.

    更新日期:2019-08-27
  • Role and dynamics of an agmatinase-like protein (AGM-1) in Neurospora crassa
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-08-26
    Luis L. Pérez-Mozqueda, Rafael Vazquez-Duhalt, Ernestina Castro-Longoria
    更新日期:2019-08-26
  • Altered nitrogen metabolism in biocontrol strains of Penicillium rubens
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-08-13
    E.A. Espeso, M. Villarino, M. Carreras, L. Alonso-Guirado, J.M. Alonso, P. Melgarejo, I. Larena

    The importance of the metabolic route of nitrogen in the fungus Penicillium rubens (strain PO212) is studied in relation to its biocontrol activity (BA). PO212 can resist a high concentration of chlorate anion and displays a classical nitrate-deficiency (nit-) phenotype resulting in poor colonial growth when nitrate is used as the main source of nitrogen. Analyses of genes implicated in nitrate assimilation evidenced the strong sequence conservation of PO212 and CH8 genome with penicillin producers such as reference strain P. rubens Wisconsin 54-1255, P2niaD18 and Pc3, however also revealed the presence of mutations. PO212 carries a mutation in the gene coding for zinc-binuclear cluster transcription factor NirA that specifically mediates the regulation of genes involved in nitrate assimilation. The nirA1 mutation causes an early stop of NirA factor, losing 66% of its sequence. The NirA1 mutant form is unable to mediate a nitrate-dependent regulation of nitrate and nitrite reductase coding genes. In this study, we study another isolate, CH8, with potential BA and nit- phenotype. A mutation in the nitrate permease coding gene crnA was found in CH8. An insertion of a guanine in the coding sequence cause a frameshift in CrnA with the loss of the last two transmembrane domains. Analysis of PO212 and CH8 isolates and complementation strains show the importance of NirA regulator in maintaining correct transcriptional levels of nitrate and nitrite reductases and suggest CrnA as the main nitrate transporter. the presence of alternative transporter for chlorate and the existence of a mechanism for preventing nitrite derived toxicity in Penicillum. BA of PO212 is partially altered when nirA1 mutation was complemented. This result and the finding of CH8, a novel biocontrol P. rubens strain with a nit- phenotype, suggest that nitrogen metabolism is a component of biocontrol capacity.

    更新日期:2019-08-14
  • Species borderlines in Fusarium exemplified by F. circinatum / F. subglutinans
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-08-12
    Liang Zhao, Sybren de Hoog, Ferry Hagen, YingQian Kang, Abdullah M.S. Al-Hatmi

    Fusarium species are known as cross-kingdom pathogens, causing infections in both plants and animals. This ecological variation challenges the species concept of closely similar lineages in the genus. The present paper describes various types of genetic interaction between strains of two neighboring model species with different predilection, F. circinatum and F. subglutinans. Parameters include sequencing of the translation elongation factor 1α (TEF1) and the second largest subunit of RNA polymerase (RPB2), sexual crossing, and vegetative compatibility groups (VCGs). Successful interspecific crosses resulted in either recombination or in homothallic fruiting, the latter being limited to F. subglutinans MAT1 parents. Crossings were skewed, as Fusarium circinatum recombined more often than F. subglutinans. We hypothesize that genetic exchange in Fusarium species is finely regulated with an arsenal of options, which are applied when partners are phylogenetically closely related, leading to fluent species borderlines.

    更新日期:2019-08-12
  • Are Macrophages the Heroes or Villains During Cryptococcosis?
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-08-12
    Jacob Rudman, Robert J. Evans, Simon Andrew Johnston

    Cryptococcus infections represents a major healthcare burden, with over 200,000 cases globally a year and even with treatment, mortality remains as high as 80%. There is a clear need for new classes of treatment, especially with the global threat of antifungal resistance. Several groups are investigating the potential of immunotherapy - circumventing many of the issues with current treatments. Macrophages are a cell type known to be heavily associated with cryptococcal infection, from the innate immune response through to the later stage chronic adaptive response – making these an ideal target for manipulation. However, it is currently debated whether macrophage activity is positive or negative for host outcomes. Here, we discuss the current literature surrounding the role of macrophages during Cryptococcus infection, and makes cases for and against macrophage enhancement. Finally, we discuss which pressing questions in the field still remain that require answers in order to safely design an immunotherapeutic with high efficacy.

    更新日期:2019-08-12
  • “When worlds collide and smuts converge”: Tales from the 1st International Ustilago/Smut Convergence”
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-08-05
    Barry J. Saville, Michael H. Perlin

    From the evening of March 12, till dinner on March 13, 2017, the 1st International Ustilago/Smut Convergence took place as a workshop prior to the start of the 29th Fungal Genetics Conference, in Asilomar, California. The overall goals of the meeting were to expand the smut model systems being used and to expand participation by the next generations of scientists with these fungi. These goals were implemented through a combination of emphasis on student and post-doc presentations, mentoring of such individuals, and active recruitment of participation by groups under-represented at such meetings in recent years in the US, especially those from Latin America and other Spanish-speaking countries. Work was presented at the first workshop on U. maydis, Sporosorium reilianum, Microbotryum violaceum, U. esculenta, and Thecaphora thlaspeos. Students and post-doctoral researchers were encouraged to present their “just-in-time,” as-yet-unpublished data, in a safe environment, with the understanding of those attending the meeting that this early access was a privilege not to be taken advantage of. The result was lively and constructive discussion, including a variety of presentations by these young scientists on putative and characterized smut effector proteins, clearly at the forefront of such research, even considering the advances presented later that week at the Fungal Genetics Conference. This review also briefly compares the first meeting with the events of the recent 2nd International Ustilago/Smut Convergence (March 11-12, 2019), which ended with a tribute to Prof. Dr. Regine Kahmann, in honor of her career, and especially for her contributions to the field of smut genetics.

    更新日期:2019-08-05
  • The peroxisomal SspA protein is redundant for purine utilization but essential for peroxisome localization in septal pores in Aspergillus nidulans
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-08-05
    Sofia Dimou, Anezia Kourkoulou, Sotiris Amillis, Riccardo Percudani, George Diallinas

    In an in silico search for correlated gene loss with fungal peroxisomal uric acid oxidase (UOX), we identified PMP22-like proteins, some of which function as promiscuous channels in organellar membranes. To investigate whether PMP22 channels have a role in peroxisomal uric acid transport and catabolism, we functionally analyzed the closest homologue in Aspergillus nidulans, named SspA. We confirmed that SspA is a peroxisomal membrane protein that co-localizes significantly with PTS1-tagged mRFP, UOX or HexA, the latter considered a protein of Woronin bodies (WB), organelles originating from peroxisomes that dynamically plug septal pores in ascomycetes. Our results suggest that in A. nidulans, unlike some other ascomycetes, there is no strict protein segregation of peroxisomal and WB-specific proteins. Importantly, genetic deletion of sspA, but not of hexA, led to lack of peroxisomal localization at septal pores, suggesting that SspA is a key factor for septal pore functioning. Additionally, ΔsspA resulted in increased sensitivity to oxidative stress, apparently as a consequence of not only the inability to plug septal pores, but also a recorded reduction in peroxisome biogenesis. However, deleting sspA had no effect on uric acid or purine utilization, as we hypothesized, a result also in line with the observation that expression of SspA was not affected by regulatory mutants and conditions known to control purine catabolic enzymes. Our results are discussed within the framework of previous studies of SspA homologues in other fungi, as well as, the observed gene losses of PMP22 and peroxisomal uric acid oxidase.

    更新日期:2019-08-05
  • Reduced phagocytosis and killing of Cryptococcus neoformans biofilm-derived cells by J774.16 macrophages is associated with fungal capsular production and surface modification
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-07-26
    Hiu Ham Lee, Jaclyn Del Pozzo, Sergio A. Salamanca, Hazael Hernandez, Luis R. Martinez

    Cryptococcus neoformans is an opportunistic encapsulated pathogen that causes life-threatening meningoencephalitis in individuals with immunosuppression. We compared the interactions of C. neoformans planktonic and biofilm-derived cells with J774.16 macrophage-like cells. Planktonic cells are more phagocytized and killed by J774.16 cells than biofilm-derived fungal cells. Biofilm-derived cryptococci possess larger capsule size and release significantly more capsular polysaccharide than planktonic cells in culture. Biofilm-derived fungi exhibited upregulation of genes involved in capsular production. Capsular-specific monoclonal antibody 18B7 demonstrated differential binding to the surface of planktonic and biofilm-derived cryptococci providing a plausible strategy for fungal evasion of macrophages and persistence. Future studies are necessary to elucidate how C. neoformans biofilm-derived cells regulate their virulence factors when interacting with cells of the immune system.

    更新日期:2019-07-27
  • The Mitochondrial Translocase of the Inner Membrane PaTim54 is involved in Defense Response and Longevity in Podospora anserina
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-07-24
    Alex Mercier, Colin Clairet, Robert Debuchy, David Morais, Philippe Silar, Sylvain Brun
    更新日期:2019-07-25
  • A metabolomics-guided approach to discover Fusarium graminearum metabolites after removal of a repressive histone modification
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-07-22
    Donovon A. Adpressa, Lanelle R. Connolly, Zachary M. Konkel, George F. Neuhaus, Xiao L. Chang, Brett R. Pierce, Kristina M. Smith, Michael Freitag, Sandra Loesgen

    Many secondary metabolites are produced by biosynthetic gene clusters (BGCs) that are repressed during standard growth conditions, which complicates the discovery of novel bioactive compounds. In the genus Fusarium, many BGCs reside in chromatin enriched for trimethylated histone 3 lysine 27 (H3K27me3), a modification correlated with transcriptional gene silencing. Here we report on our progress in assigning metabolites to genes by using a strain lacking the H3K27 methyltransferase, Kmt6. To guide isolation efforts, we coupled genetics to multivariate analysis of liquid chromatography–mass spectrometry (LCMS) data from both wild type and kmt6, which allowed identification of compounds previously unknown from F. graminearum. We found low molecular weight, amino acid-derived metabolites (N-ethyl anthranilic acid, N- phenethylacetamide, N-acetyltryptamine). We identified one new compound, protofusarin, as derived from fusarin biosynthesis. Similarly, we isolated large amounts of fusaristatin A, gibepyrone A, and fusarpyrones A and B, simply by using the kmt6 mutant, instead of having to optimize growth media. To increase the abundance of metabolites underrepresented in wild type, we generated kmt6 fus1 double mutants and discovered tricinolone and tricinolonoic acid, two new sesquiterpenes belonging to the tricindiol class. Our approach allows rapid visualization and analyses of the genetically induced changes in metabolite production, and discovery of new molecules by a combination of chemical and genetic dereplication. Of 22 fungal metabolites identified here, 10 compounds had not been reported from F. graminearum before. We show that activating silent metabolic pathways by mutation of a repressive chromatin modification enzyme can result in the discovery of new chemistry even in a well-studied organism, and helps to connect new or known small molecules to the BGCs responsible for their production.

    更新日期:2019-07-23
  • Antifungal resistance in dermatophytes: recent trends and therapeutic implications
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-07-19
    Ananta Khurana, Kabir Sardana, Anuradha Chowdhary

    Dermatophytoses or tinea refers to superficial fungal infection of keratinized tissues. Although generally considered easy to treat, recalcitrant infections, presenting as extensive and difficult to treat tinea corporis and cruris, are on the rise in some parts of the world. The situation demands an understanding of the pharmacokinetic and pharmacodynamic properties of the available antifungals against dermatophytes and the possible contribution of drug resistance and other factors to the present scenario. In this review, we provide the readers a comprehensive account of the available literature on in-vitro resistance to clinically used antifungals among dermatophytes and apprise them of the relevant skin pharmacokinetics of each. The established and postulated mechanisms of drug resistance are discussed and aspects on lack of in vivo correlation of in vitro resistance is presented. Finally, the lacunae in our existing knowledge on the topic and the arenas for future research are highlighted.

    更新日期:2019-07-20
  • Facilitators of adaptation and antifungal resistance mechanisms in clinically relevant fungi
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-07-19
    Margriet W.J. Hokken, B.J. Zwaan, W.J.G. Melchers, P.E. Verweij

    Opportunistic fungal pathogens can cause a diverse range of diseases in humans. The increasing rate of fungal infections caused by strains that are resistant to commonly used antifungals results in difficulty to treat diseases, with accompanying high mortality rates. Existing and newly emerging molecular resistance mechanisms rapidly spread in fungal populations and need to be monitored. Fungi exhibit a diversity of mechanisms to maintain physiological resilience and create genetic variation; processes which eventually lead to the selection and spread of resistant fungal pathogens. To prevent and anticipate this dispersion, the role of evolutionary factors that drive fungal adaptation should be investigated. In this review, we provide an overview of resistance mechanisms against commonly used antifungal compounds in the clinic and for which fungal resistance has been reported. Furthermore, we aim to summarize and elucidate potent generators of genetic variability across the fungal kingdom that aid adaptation to stressful environments. This knowledge can lead to recognizing potential niches that facilitate fast resistance development and can provide leads for new management strategies to battle the emerging resistant populations in the clinic and the environment.

    更新日期:2019-07-19
  • Epigenetic mechanisms of drug resistance in fungi
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-07-17
    Zanetta Chang, Vikas Yadav, Soo Chan Lee, Joseph Heitman

    The emergence of drug-resistant fungi poses a continuously increasing threat to human health. Despite advances in preventive care and diagnostics, resistant fungi continue to cause significant mortality, especially in immunocompromised patients. Therapeutic resources are further limited by current usage of only four major classes of antifungal drugs. Resistance against these drugs has already been observed in pathogenic fungi requiring the development of much needed newer antifungal drugs. Epigenetic changes such as DNA or chromatin modifications alter gene expression levels in response to certain stimuli, including interaction with the host in the case of fungal pathogens. These changes can confer resistance to drugs by altering the expression of target genes or genes encoding drug efflux pumps. Multiple pathogens share many of these epigenetic pathways; thus, targeting epigenetic pathways might also identify drug target candidates for the development of broad-spectrum antifungal drugs. In this review, we discuss the importance of epigenetic pathways in mediating drug resistance in fungi as well as in the development of anti-fungal drugs.

    更新日期:2019-07-18
  • Stage-specific functional relationships between Tub1 and Tub2 beta-tubulins in the wheat scab fungus Fusarium graminearum
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-07-15
    Huan Wang, Daipeng Chen, Chengliang Li, Neng Tian, Ju Zhang, Jin-Rong Xu, Chenfang Wang

    The filamentous ascomycete Fusarium graminearum contains two β-tubulin genes TUB1 and TUB2 that differ in functions during vegetative growth and sexual reproduction. To further characterize their functional relationship, in this study we determined the co-localization of Tub1 and Tub2 and assayed their expression levels in different mutants and roles in DON production. Tub1 co-localized with Tub2 to the same regions of microtubules in conidia, hyphae, and ascospores. Whereas deletion of TUB1 had no obvious effect on the transcription of TUB2 and two α-tubulin genes (TUB4 and TUB5), the tub2 mutant was up-regulated in TUB1 transcription. To assay their protein expression levels, polyclonal antibodies that could specifically detect four α- and β-tubulin proteins were generated. Western blot analyses showed that the abundance of Tub1 proteins was increased in tub2 but reduced in tub4 and tub5 mutants. Interestingly, protein expression of Tub4 and Tub5 was decreased in the tub1 mutant in comparison with the wild type, despite a lack of obvious changes in their transcription. In contrast, deletion of TUB2 had no effect on translation of TUB4 and TUB5. Ectopic expression of Tub2-mCherry partially recovered the growth defect of the tub1 mutant but did not rescue its defect in sexual reproduction. Expression of Tub1-GFP in the tub2 mutant also partially rescued its defects in vegetative growth, suggesting that disturbance in the balance of α- and β-tubulins contributes to mutant defects. The tub2 but not tub1 mutant was almost blocked in DON biosynthesis. Expression of TRI genes, toxisome formation, and DON-related cellular differentiation were significantly reduced in the tub2 mutant. Overall, our results showed that Tub1 and Tub2 share similar subcellular localization and have overlapping functions during vegetative growth but they differ in functions in DON production and ascosporogenesis in F. graminearum.

    更新日期:2019-07-16
  • Multidrug transporters of Candida species in clinical azole resistance
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-07-11
    Rajendra Prasad, Remya Nair, Atanu Banerjee

    Over-expression of the human P-glycoprotein (P-gp) in tumor cells is a classic example of an ABC protein serving as a hindrance to effective chemotherapy. The existence of proteins homologous to P-gp in organisms encompassing the entire living kingdom highlights extrusion of drugs as a general mechanism of multidrug resistance. Infections caused by opportunistic human fungal pathogens such as Candida species are very common and has intensified in recent years. The typical hosts, who possess suppressed immune systems due to conditions such as HIV and transplantation surgery etc., are prone to fungal infections. Prolonged chemotherapy induces fungal cells to eventually develop tolerance to most of the antifungals currently in clinical use. Amongst other prominent mechanisms of antifungal resistance such as manipulation of the drug target, rapid efflux achieved through overexpression of multidrug transporters has emerged as a major resistance mechanism for azoles. Herein, the azole-resistant clinical isolates of Candida species utilize a few select efflux pump proteins belonging to the ABC and MFS superfamilies, to deter the toxic accumulation of therapeutic azoles and thus, facilitating cell survival. In this article, we summarize and discuss the clinically relevant mechanisms of azole resistance in Candida albicans and non-albicans Candida (NAC) species, specifically highlighting the role of multidrug efflux proteins in the phenomenon.

    更新日期:2019-07-12
  • Functional analysis of the mitochondrial gene mitofilin in the filamentous entomopathogenic fungus Beauveria bassiana
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-07-04
    Jia-Jia Wang, Yue-Jin Peng, Ming-Guang Feng, Sheng-Hua Ying
    更新日期:2019-07-05
  • Fungal species boundaries in the genomics era
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-07-04
    Daniel R. Matute, Victoria E. Sepúlveda

    Genomic data has opened new possibilities to understand how organisms change over time, and could enable the discovery of previously undescribed species. Although taxonomy used to be based on phenotypes, molecular data has frequently revealed that morphological traits are insufficient to describe biodiversity. Genomics holds the promise of revealing even more genetic discontinuities, but the parameters on how to describe species from genomic data remain unclear. Fungi have been a successful case in which the use of molecular markers has uncovered the existence of genetic boundaries where no crosses are possible. In this minireview, we highlight recent advances, propose a set of standards to use genomic sequences to uncover species boundaries, point out potential pitfalls, and present possible future research directions.

    更新日期:2019-07-05
  • Heterologous Expression of Intact Biosynthetic Gene Clusters in Fusarium graminearum
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-07-04
    Mikkel Rank Nielsen, Rasmus Dam Wollenberg, Klaus Ringsborg Westphal, Teis Esben Sondergaard, Reinhard Wimmer, Donald Max Gardiner, Jens Laurids Sørensen
    更新日期:2019-07-05
  • Global distribution of mating types shows limited opportunities for mating across populations of fungi causing boxwood blight disease
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-06-27
    Martha Malapi-Wight, Daniel Veltri, Bjorn Gehesquière, Kurt Heungens, Yazmín Rivera, Catalina Salgado-Salazar, Jo Anne Crouch

    Boxwood blight is a disease threat to natural and managed landscapes worldwide. To determine mating potential of the fungi responsible for the disease, Calonectria pseudonaviculata and C. henricotiae, we characterized their mating-type (MAT) loci. Genomes of C. henricotiae, C. pseudonaviculata and two other Calonectria species (C. leucothoes, C. naviculata) were sequenced and used to design PCR tests for mating-type from 268 isolates collected from four continents. All four Calonectria species have a MAT locus that is structurally consistent with the organization found in heterothallic ascomycetes, with just one idiomorph per individual isolate. Mating type was subdivided by species: all C. henricotiae isolates possessed the MAT1-1 idiomorph, whereas all C. pseudonaviculata isolates possessed the MAT1-2 idiomorph. To determine the potential for divergence at the MAT1 locus to present a barrier to interspecific hybridization, evolutionary analysis was conducted. Phylogenomic estimates showed that C. henricotiae and C. pseudonaviculata diverged approximately 2.1 Mya. However, syntenic comparisons, phylogenetic analyses, and estimates of nucleotide divergence across the MAT1 locus and proximal genes identified minimal divergence in this region of the genome. These results show that in North America and parts of Europe, where only C. pseudonaviculata resides, mating is constrained by the absence of MAT1-1. In regions of Europe where C. henricotiae and C. pseudonaviculata currently share the same host and geographic range, it remains to be determined whether or not these two recently diverged species are able to overcome species barriers to mate.

    更新日期:2019-06-27
  • Antifungal resistance in Aspergillus terreus: A current scenario
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-06-25
    Roya Vahedi-Shahandashti, Cornelia Lass-Flörl

    Invasive aspergillosis caused by intrinsically resistant non-fumigatus Aspergillus species displays a poor outcome in immunocompromised patients. The polyene antifungal amphotericin B (AmB) remains to be “gold standard” in the treatment of invasive fungal infections. Aspergillus terreus is innately resistant to AmB, in vivo and in vitro. Till now, the exact mode of action in polyene resistance is not well understood. This review highlights the underlying molecular basis of AmB resistance in A. terreus, displaying data obtained from AmB susceptible A. terreus and AmB resistant A. terreus strains. The effect of AmB on main cellular and molecular functions such as fungal respiration and stress response pathways will be discussed in detail and resistance mechanisms will be highlighted. The fungal stress response machinery seems to be a major player in the onset of AmB resistance in A. terreus.

    更新日期:2019-06-25
  • Candida auris and multidrug resistance: defining the new normal
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-06-20
    Shawn R. Lockhart

    Candida auris is an emerging species of yeast characterized by colonization of skin, persistence in the healthcare environment, and antifungal resistance. C. auris was first described in 2009 from a single isolate but has since been reported in more than 25 countries worldwide. Resistance to fluconazole and amphotericin B is common, and resistance to the echinocandins is emerging in some countries. Antifungal resistance has been shown to be acquired rather than intrinsic and the primary mechanisms of resistance to the echinocandins and azoles have been determined. There are a number of new antifungal agents in phase 2 and phase 3 clinical trials and many have activity against C. auris. This review will discuss what is currently known about antifungal resistance in C. auris, limitations to antifungal susceptibility testing, the mechanisms of resistance, and the new antifungals that are on the horizon.

    更新日期:2019-06-20
  • Effects on hyphal morphology and development by the putative copper radical oxidase glx1 in Trichoderma virens suggest a novel role as a cell wall associated enzyme
    Fungal Genet. Biol. (IF 3.314) Pub Date : 2019-06-20
    Frankie K. Crutcher, Maria E. Moran-Diez, Inna V. Krieger, Charles M. Kenerley

    Trichoderma spp. have been characterized for their capacity to act as biological control agents against several pathogens through the activity of secondary metabolites and cell wall degrading enzymes. However, only T. reesei has been widely studied for the ability to assimilate lignocellulose substrates. Protein analysis by SDS-PAGE of culture filtrate of T. virens revealed the presence of an unknown ∼ 77 kDa band protein (GLX1) that showed sequence homology to glyoxal-like oxidase genes involved in lignin degradation. The analysis and biochemical characterization of the 1,119 amino acid coded protein showed the presence of five carbohydrate-binding modules (CBMs) with affinity for colloidal chitin, and a functional glyoxal oxidase catalytic domain that is involved in the production of hydrogen peroxide when methylglyoxal was used as a substrate. The silencing of the glx1 gene resulted in mutants with more than 90% expression reduction and the absence of glyoxal oxidase catalytic activity. These mutants showed delayed hyphal growth, reduced colony and conidial hydrophobicity, but showed no changes in their biocontrol ability. Most significantly, mutants exhibited a loss of growth directionality resulting in a curled phenotype that was eliminated in the presence of exogenous H2O2. Here we present evidence that in T. virens, glx1 is not involved in the breakdown of lignin but instead is responsible for normal hyphal growth and morphology and likely does this through free radical production within the fungal cell wall. This is the first time that a glyoxal oxidase protein has been isolated and characterized in ascomycete fungi.

    更新日期:2019-06-20
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