Frontiers in fungal biology最新文献

Mycoparasitic interactions are common in nature, form part of the microbiota of plants, and are considered significant contributors to fungus-fungus antagonism. Mycoparasites kill plant pathogens, protect the plant from abiotic and biotic stressors, and reduce disease incidence and severity at the plant population level. Their exploitation as biocontrol agents in agriculture is well documented. However, mycoparasites may potentially affect classical fungal biocontrol agents of invasive weed species. Classical biological control, or biocontrol, of invasive weeds involves the intentional introduction of exotic, usually co-evolved plant pathogens and insects, for permanent establishment and long-term control of the target plant. Agent establishment, effectiveness, and safety are the critical elements for a successful weed biocontrol programme. Establishment and effectiveness of agents on the invasive plant often vary throughout the invaded range with about two-thirds of weed biocontrol agents failing to suppress their target weed. There are many documented reasons why weed biocontrol agents do not establish or are ineffective when they do, and the presence and accumulation of natural enemies in the invaded range is one of them. Endophyte-enriched, invasive weeds and those forming mutualistic associations with indigenous, native endophytes could explain the lack of consistency of some classical biological control introductions. However, another variable could be factored into the mix: mycoparasitism, where one fungus parasitises another, the natural enemies of the plant's natural enemies. In this review article, we introduce the concept of invasive weed biocontrol and the history of using plant pathogens as biocontrol agents. We discuss the success and failure of fungal agent programmes and delve into the patterns of success or failure, with a focus on the potential antagonistic role of endophytes and mycoparasites.

Two models of increasing complexity were constructed to simulate the interactions between the root-knot nematode (RKN) Meloidogyne incognita and the biocontrol fungus Pochonia chlamydosporia var. catenulata in a rhizosphere microcosm. The models described discrete population dynamics at hourly rates over a 6-month period and were validated using real parasitism and nematode or fungus data. A first, general Pochonia-nematode-root model (GPNR) used five functions and 16 biological constants. The variables and constants describing the RKN life cycle included the rates of egg production, hatching, juvenile (J2), and mature female development, including root or nematode self-density-dependent factors. Other constants accounted for egg parasitism, nematode-induced root losses, growth, and mortalities. The relationship between nematodes and fungal propagules showed density dependence and cyclic variations in time, including an attractor on the propagules and J2 phases space. The simulations confirmed a P. chlamydosporia optimal initial density of 5 · 10³ propagules · cc soil^-1, as usually applied in assays. The constants used in GPNR showed adherence to the nematode biology, with 10³ eggs per egg mass, a 10-day average lifespan of J2, with 2 days required to enter roots, and adult lifespan lasting 24 days. The fungus propagule lifespan was 25 days, with an average feeder root lifespan lasting around 52 days. A second, more complex Pochonia-nematode-root detailed model (GPNRd) was then constructed using eight functions and 23 constants. It was built as GPNR did not allow the evaluation of host prevalence. GPNRd allowed simulations of all RKN life stages and included non-parasitic and parasitic fungus population fractions. Both GPNR and GPNRd matched real J2 and fungus density data observed in a RKN biocontrol assay. Depending on the starting conditions, simulations showed stability in time, interpreted as effective host regulation. GPNRd showed a fungus cyclic relationship with the J2 numbers, with prevalence data close to those observed (38.3 vs. 39.4%, respectively). This model also showed a further density-independent nematode regulation mechanism based on the P. chlamydosporia switch from a non-parasitic to a parasitic trophic behavior. This mechanism supported the biocontrol of M. incognita, also sustained by a concomitant increase of the root density.

Aspergillus fumigatus is the main cause of life-threatening invasive aspergillosis. Despite the availability of various antifungals, therapy remains challenging and requires further studies. Accordingly, the clinical A. fumigatus isolate NIH4215 deriving from a fatal case of human pulmonary aspergillosis has frequently been used in drug efficacy studies. Unexpectedly, our initial attempts to generate a bioluminescent reporter of strain NIH4215 for in vivo drug efficacy studies failed, as NIH4215 was unable to grow on defined minimal medium. Subsequent analyses discovered a previously undescribed thiamine auxotrophy of strain NIH4215 and transformation with thiamine biosynthesis genes from A. fumigatus strain Af293 identified the nmt1 gene as cause of the thiamine auxotrophy. Sequencing of the defective nmt1 gene revealed the loss of a cysteine codon within an essential iron-binding motif. Subsequently, the wild-type nmt1 gene was successfully used to generate a bioluminescent reporter strain in NIH4215 by simultaneously deleting the akuB locus. The resulting bioluminescent ΔakuB strains showed a high frequency of homologous integration as confirmed by generation of pyrG and niaD deletion mutants. When tested in a Galleria mellonella infection model, neither thiamine auxotrophy nor the deletion of the akuB locus had a significant effect on virulence. However, besides thiamine auxotrophy, sectors with altered morphology and albino mutants frequently arose on colony edges of strain NIH4215 and its derivatives, and stable albino mutants were successfully isolated. A proposed increased mutation rate of NIH4215 was confirmed by screening for spontaneous occurrence of fluoorotic acid resistant mutants. Independent mutations in the pyrG and pyrE gene were identified in the fluoroorotic acid resistant NIH4215 isolates and the frequency of mutation was by at least one order of magnitude higher than that observed for the clinical A. fumigatus isolate CBS144.89. In summary, despite its virulence in animal models, strain NIH4215 is a thiamine auxotroph and prone to accumulate mutations. Our results suggest that thiamine biosynthesis is dispensable for host infection and mutation-prone strains such as NIH4215 could potentially facilitate the evolution of azole resistant strains as increasingly observed in the environment.

Numerous plant-pathogenic fungi secrete necrotrophic effectors (syn. host-selective toxins) that are important determinants of pathogenicity and virulence in species that have a necrotrophic lifestyle. Corynespora cassiicola is a necrotrophic fungus causing emerging target spot epidemics in the southeastern United States (US). Previous studies revealed that populations of C. cassiicola from cotton, soybean, and tomato are clonal, host specialized and genetically distinct. Additionally, cassiicolin - the necrotrophic effector identified in some C. cassiicola isolates - is an important toxin for virulence on rubber. It is encoded by seven Cas gene variants. Our goal was to conduct comparative genomic analyses to identify variation among putative necrotrophic effector genes and to determine if lack of one of the mating-types explained clonal populations in C. cassiicola causing outbreaks in the southeastern US and the apparent absence of sexual reproduction worldwide. A total of 12 C. cassiicola genomes, with four each from isolates from tomato, soybean, and cotton, were sequenced using an Illumina Next Seq platform. Each genome was assembled de novo, compared with the reference genome from rubber, and searched for known Cas, and other gene clusters with homologs of secondary metabolites. Cas2 and/or Cas6 were present in isolates from soybean in the southeastern US, whereas Cas1 and Cas2 were present in isolates from cotton in the southeastern US. In addition, several toxin genes, including the T-toxin biosynthetic genes were present in all C. cassiicola from cotton, soybean, and tomato. The mating-type locus was identified in all of the sequenced genomes, with the MAT1-1 idiomorph present in all cotton isolates and the rubber isolate, whereas the MAT1-2 idiomorph was present in all soybean isolates. We developed a PCR-based marker for mating-type in C. cassiicola. Both mating types were present in isolates from tomato. Thus, C. cassiicola has both mating-types necessary for sexual reproduction, but the absence of both mating-types within soybean and cotton populations could explain clonality in these populations. Variation in necrotrophic effectors may underlie host specialization and disease emergence of target spot on cotton, soybean, and tomato in the southeastern US.

Grapevine fungal trunk diseases (GTDs) have become a serious problem for grapevines worldwide. Nursery vines infected during the propagation process are considered one of the main ways of dissemination of GTD pathogens. In this study, we examined the status of GTDs in grapevine planting material, from rootstocks and scion mother cuttings to grafted rooted vines ready to plant, according to the local nursery propagation process. During 2018-2019, internal symptoms of GTDs were examined in 2400 propagation materials and fungal isolations were carried out from a subsample of 1026 selected materials. Our results revealed that nursery grapevine plants produced in Uruguay have a high incidence of GTDs, regardless of the scion/rootstock combination. Typical brown to black streaks and sectorial wood necrosis were observed in materials on all propagation stages, with a markedly increasing incidence throughout the nursery process, reaching almost 100% in grafted rooted vines ready to plant. Botryosphaeria dieback, Petri disease and black-foot disease were the main GTDs found. The results showed that Botryosphaeria dieback and Petri disease pathogens infect materials from the early stages of the process, with a marked increase towards the end of the plant production process, whereas black-foot disease pathogens were found exclusively in vines ready to plant. Diaporthe dieback pathogens were also detected in materials in all stages but in a low proportion (less than 10% of infected material). Based on single locus analysis, the 180 isolates selected were placed into eight genera and 89% identified within 22 fungal species associated with GTDs, with Phaeoacremonium oleae and Diaporthe terebinthifolii as new records on grapevine worldwide. Our results have concluded that locally produced vines are one of the main ways of dissemination of GTD pathogens and showed that a nursery sanitation programme is required to reduce the incidence of these diseases.

The possibility to sequence the entire genome of an organism revolutionized the fields of biology and biotechnology. The first genome sequence of the important filamentous fungus Aspergillus niger was obtained in 2007, 11 years after the release of the first eukaryotic genome sequence. From that moment, genomics of A. niger has seen major progresses, facilitated by the advances in the sequencing technologies and in the methodologies for gene function prediction. However, there are still challenges to face when trying to obtain complete genomes, equipped with all the repetitive sequences that they contain and without omitting the mitochondrial sequences. The aim of this perspective article is to discuss the current status of A. niger genomics and draw attention to the open challenges that the fungal community should address to move research of this important fungus forward.

Fungal mating-type loci (MAT) encode transcription factors (TFs) MAT1-1-1 and MAT1-2-1, which govern sexual reproduction as well as other developmental processes. In Penicillium chrysogenum, the major producer of the beta-lactam antibiotic penicillin, a recent chromatin immunoprecipitation followed by sequencing (ChIP-seq) analysis identified 254 genes as direct targets of MAT1-1-1, many of which encode thus far uncharacterized proteins. Here, we characterized one of the major targets of MAT1-1-1, the tom1 gene, which encodes a protein highly conserved within the group of Eurotiomycetes fungi. Using fluorescence microscopy, we demonstrated binding of MAT1-1-1 to the tom1 promoter by reporter gene analysis. Extensive electrophoretic mobility shift assays (EMSAs) further showed that the promoter sequence of tom1 is bound in vitro by both MAT1-1-1 and MAT1-2-1. This indicated an interaction between the two TFs, which was verified by yeast two-hybrid analysis. The sequence of tom1 carries a nuclear localization sequence, and indeed its nuclear localization was verified by fluorescence microscopy. The in vivo function of tom1 was investigated using tom1 deletion strains, as well as a complementing strain where the wild-type tom1 gene was reintroduced. We found a clear sporulation defect in the deletion strain, which became more evident when the fungi were grown at an elevated temperature of 31°C.

Plant communities and fungi inhabiting their phyllospheres change along precipitation gradients and often respond to changes in land use. Many studies have focused on the changes in foliar fungal communities on specific plant species, however, few have addressed the association between whole plant communities and their phyllosphere fungi. We sampled plant communities and associated phyllosphere fungal communities in native prairie remnants and post-agricultural sites across the steep precipitation gradient in the central plains in Kansas, USA. Plant community cover data and MiSeq ITS2 metabarcode data of the phyllosphere fungal communities indicated that both plant and fungal community composition respond strongly to mean annual precipitation (MAP), but less so to land use (native prairie remnants vs. post-agricultural sites). However, plant and fungal diversity were greater in the native remnant prairies than in post-agricultural sites. Overall, both plant and fungal diversity increased with MAP and the communities in the arid and mesic parts of the gradient were distinct. Analyses of the linkages between plant and fungal communities (Mantel and Procrustes tests) identified strong correlations between the composition of the two. However, despite the strong correlations, regression models with plant richness, diversity, or composition (ordination axis scores) and land use as explanatory variables for fungal diversity and evenness did not improve the models compared to those with precipitation and land use (ΔAIC < 2), even though the explanatory power of some plant variables was greater than that of MAP as measured by R². Indicator taxon analyses suggest that grass species are the primary taxa that differ in the plant communities. Similar analyses of the phyllosphere fungi indicated that many plant pathogens are disproportionately abundant either in the arid or mesic environments. Although decoupling the drivers of fungal communities and their composition - whether abiotic or host-dependent - remains a challenge, our study highlights the distinct community responses to precipitation and the tight tracking of the plant communities by their associated fungal symbionts.

Parastagonospora nodorum causes glume and leaf blotch of wheat, a harmful disease resulting in serious losses in grain yield. In many countries including Russia, fungicidal formulations based on triazoles and on triazoles combined with strobilurins are used to control this fungus. However, their prolonged application may promote the selection of fungicide-resistant strains of P. nodorum leading to significant attenuation or even loss of fungicidal effect. Chemosensitization of plant pathogenic fungi with natural compounds represents a promising strategy for mitigating fungicide resistance and other negative impacts of fungicides. In this work, we applied a chemosensitization approach towards P. nodorum strains non-resistant or resistant to tebuconazole or azoxystrobin using 6-demethylmevinolin (6-DMM), a metabolite of Penicillium citrinum. The resistant strains were obtained by the mutagenesis and subsequent culturing on agar media incorporated with increasing doses of Folicur^® EC 250 (i.e., tebuconazole) or Quadris^® SC 250 (i.e., azoxystrobin). Test strains m8-4 and kd-18, most resistant to tebuconazole and azoxystrobin, respectively, were selected for sensitization experiments. These experiments demonstrated that combining 6-DMM with Folicur^® enhanced fungicidal effectiveness in vitro and in vivo in addition to attenuating the resistance of P. nodorum to tebuconazole in vitro. 6-DMM was also found to augment Quadris^® efficacy towards kd-18 when applied on detached wheat leaves inoculated with this strain. Experiments on P. nodorum sensitization under greenhouse conditions included preventive (applying test compounds simultaneously with inoculation) or post-inoculation spraying of wheat seedlings with 6-DMM together with Folicur^® at dose rates (DR) amounting to 10% and 20% of DR recommended for field application (RDR). Combined treatments were run in parallel with using the same DR of the fungicide and sensitizer, alone. A synergistic effect was observed in both preventive and post-inoculation treatments, when the sensitizer was co-applied with the fungicide at 10% of the RDR. In this case, disease reduction significantly exceeded the protective effect of Folicur^® at 10% or 20% of the RDR, alone, and also a calculated additive effect. Collectively, our findings suggest that 6-DMM is promising as a putative component for formulations with triazole and strobilurin fungicides. Such new formulations would improve fungicide efficacy and, potentially, lower rates of fungicides needed for plant pathogen control.

The filamentous ascomycete Bipolaris maydis is a plant pathogen that causes corn leaf blight and has been used in cytological studies of sexual reproduction. In this fungus, when null mutants of each septin are crossed with the wild-type strain, all ascospores derived from the same asci show abnormal morphology. The phenomenon was remarkably similar to the event known as "ascus dominance" in Neurospora crassa, which is known to be caused by MSUD (meiotic silencing by unpaired DNA). However, it is not clear whether B. maydis possesses functional MSUD. The object of this study is to elucidate whether this fungus carries a functional MSUD system that causes ascus dominance in the crosses of septin mutants and the wild-type strain. The results of homozygous and heterozygous crossing tests with mutants, having the insertional CDC10-septin gene sequence into the genome, suggested that the ascus dominance in B. maydis is triggered by the unpaired DNA as in N. crassa. To investigate whether MSUD is caused by the same mechanism as in N. crassa, an RNA-dependent RNA polymerase, one of the essential factors in MSUD, was identified and disrupted (Δrdr1) in B. maydis. When the Δrdr1 strain was crossed with each mutant of the septins, ascus dominance did not occur in all crosses. These results suggest that this ascus dominance is caused by RNA silencing triggered by an unpaired gene, as in N. crassa, and septin genes were affected by this silencing. To date, although MSUD has been found only in Fusarium graminearum and N. crassa, which are classified as Sordariomycetes, this study showed that MSUD is also functional in B. maydis, which is classified as a Dothideomycete. These results showed the possibility that this posttranscriptional regulation is extensively conserved among filamentous ascomycetes.