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1.
J Agric Food Chem ; 67(31): 8573-8580, 2019 Aug 07.
Artigo em Inglês | MEDLINE | ID: mdl-31293156

RESUMO

Glycosylation endows both natural and synthetic small molecules with modulated physicochemical and biological properties. Plant and bacterial glycosyltransferases capable of decorating various privileged scaffolds have been extensively studied, but those from kingdom Fungi still remain underexploited. Here, we use a combination of genome mining and heterologous expression techniques to identify four novel glycosyltransferase-methyltransferase (GT-MT) functional modules from Hypocreales fungi. These GT-MT modules display decent substrate promiscuity and regiospecificity, methylglucosylating a panel of natural products such as flavonoids, stilbenoids, anthraquinones, and benzenediol lactones. Native GT-MT modules can be split up and regrouped into hybrid modules with similar or even improved efficacy as compared with native pairs. Methylglucosylation of kaempferol considerably improves its insecticidal activity against the larvae of oriental armyworm Mythimna separata (Walker). Our work provides a set of efficient biocatalysts for the combinatorial biosynthesis of small molecule glycosides that may have significant importance to the pharmaceutical, agricultural, and food industries.


Assuntos
Proteínas Fúngicas/química , Glicosiltransferases/química , Hypocreales/enzimologia , Metiltransferases/química , Fenóis/química , Animais , Biocatálise , Cristalografia por Raios X , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Glicosilação , Glicosiltransferases/genética , Glicosiltransferases/metabolismo , Hypocreales/genética , Inseticidas/química , Inseticidas/farmacologia , Metilação , Metiltransferases/genética , Metiltransferases/metabolismo , Mariposas/efeitos dos fármacos , Fenóis/farmacologia , Especificidade por Substrato
2.
J Agric Food Chem ; 67(26): 7266-7273, 2019 Jul 03.
Artigo em Inglês | MEDLINE | ID: mdl-31244199

RESUMO

Chemical investigation of fungus Pochonia chlamydosporia strain 170, derived from rice fermentation sediment samples, afforded seven radicicol analogues, including two new compounds, monocillin VI (1) and monocillin VII (2), and five known compounds, monocillin II (3), monorden D (4), monocillin IV (5), monocillin V (6), and pochonin M (7). The structures of compounds 1-7 were established primarily by analysis of nuclear magnetic resonance data, and the absolute configurations of the secondary alcohol in compounds 1 and 2 were assigned by the modified Mosher method. All seven compounds have modest antibacterial activities, with a minimal inhibitory concentration (MIC) of 25.6 µg/mL for compounds 1 and 3-7 and 51.2 µg/mL for compound 2, on inhibition of the growth of the plant pathogen Xanthomonas campestris (the positive control ampicillin showed a MIC value of 12.8 µg/mL), indicating that the fungus has the potential to control bacterial disease. The biosynthetic gene cluster and putative biosynthetic pathways of these radicicol analogues in the P. chlamydosporia genome were proposed. These findings increase our knowledge of the chemical potential of P. chlamydosporia and may allow us to better utilize the fungus as a biological control agent.


Assuntos
Antibacterianos/química , Hypocreales/metabolismo , Macrolídeos/química , Antibacterianos/biossíntese , Antibacterianos/farmacologia , Vias Biossintéticas , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Hypocreales/química , Hypocreales/genética , Macrolídeos/metabolismo , Macrolídeos/farmacologia , Testes de Sensibilidade Microbiana , Família Multigênica , Xanthomonas campestris/efeitos dos fármacos , Xanthomonas campestris/crescimento & desenvolvimento
3.
Plant Dis ; 103(8): 1865-1875, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31161921

RESUMO

Black root rot of avocado is a severe disease of nursery trees and young orchard transplants, causing tree death within a year after planting. In Australia, key pathogens include species complexes Calonectria ilicicola and Dactylonectria macrodidyma; however, several other Dactylonectria species also cause the disease. Rapid detection of these pathogens in planta is important to speed up implementation of disease management and reduce loss. The purpose of this study was to develop three loop-mediated isothermal amplification (LAMP) diagnostic assays to rapidly identify species within the C. ilicicola and D. macrodidyma complexes and species in the Dactylonectria genus in avocado roots. Primers were designed from ß-tubulin sequence data of C. ilicicola and from histone H3 of D. macrodidyma and the Dactylonectria genus. The LAMP primers were tested for specificity and sensitivity with 82 fungal isolates, which included the target species complexes C. ilicicola and D. macrodidyma; species within the target Dactylonectria genus viz. D. macrodidyma, D. anthuriicola, D. novozelandica, D. pauciseptata, and D. vitis; and isolates of nontarget species, including Calonectria sp., Cylindrocladiella sp., Gliocladiopsis forsbergii, G. peggii, G. whileyi, Ilyonectria sp., Mariannaea sp., Fusarium sp., and Phytophthora cinnamomi. The species-specific LAMP assays were sensitive and specific at DNA concentrations of 1 pg/µl for C. ilicicola and 0.01 ng/µl for D. macrodidyma, whereas the Dactylonectria genus-wide assay was sensitive to 0.1 ng/µl. Detection of C. ilicicola occurred within 10 to 15 or 15 to 30 min when the template was pure DNA or crude extracts obtained from suspending fungal cultures in sterile water, respectively. Detection of D. macrodidyma was between 12 to 29 min with pure DNA and 16 to 30 min with crude extracts. Dactylonectria spp. were detected within 6 to 25 min with pure DNA and 7 to 23 min with crude extracts. The specificity of the assays was found to be dependent on time and isothermal amplification temperature, with optimal specificity occurring in reactions of <30 min and at temperatures of 67°C for C. ilicicola and D. macrodidyma assays and 69°C for Dactylonectria genus-wide assays. The assays were modified to accommodate a DNA extraction step and use of avocado roots as DNA templates. Detection in avocado roots ranged between 12 to 25 min for C. ilicicola, 12 to 26 min for D. macrodidyma, and 14 to 30 min for species in the Dactylonectria genus. The LAMP assays are applicable across multiple agricultural industries, because C. ilicicola, D. macrodidyma, and Dactylonectria spp. are also important pathogens of various crops and ornamental plants.


Assuntos
Agricultura/métodos , Hypocreales , Técnicas de Amplificação de Ácido Nucleico , Persea , Austrália , DNA Fúngico/genética , Hypocreales/genética , Persea/microbiologia , Doenças das Plantas/microbiologia
4.
Pestic Biochem Physiol ; 157: 99-107, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31153482

RESUMO

The entmopathogenic fungus Lecaniicillium lecanii is a naturally available biological control and it is considered to be one of the best mycoinsecticide agents against the destructive insect pest Diaphorina citri Kuwayama. The present study aimed to extract and characterize the toxic insecticidal protein from L. lecanii and to assess the toxicity level against the Asian citrus psyllid the vector of Huanglongbing disease (HLB), also called citrus greening. Extracts of a toxic substance from submerged batch culture examined by sodium dodecyl sulfate-poly-acrylamide (SDS-PAGE), had a molecular weight of 45 kDa. The most abundant toxic metabolite was subjected to HPLC to purify and identified it by mass spectrometry. Subsequently, metabolite toxicity was tested against D. citri at three different concentrations (1%, 2%, and 3%). The results showed that the highest concentration had a significant maximum mortality at 120 h post application. Furthermore, we investigated the expression of the GAS1 gene which was previously identified to have a role in pathogenicity in in vivo studies in adult insect psyllids. Results of this study indicated that expression of the virulence factor gene was present at three concentrations of the fungal suspension post inoculation. This is the first study to provide this novel approach for the characterization of fungal mediated synthesis of a cuticle degrading soluble protein against the insect D. citri. The present results provide strong information on the in vivo expression of the GAS1 gene involved in fungal virulence pertaining to penetration of the insect cuticle, but not to inhibiting the growth of the host.


Assuntos
Hemípteros/microbiologia , Hypocreales/metabolismo , Hypocreales/patogenicidade , Animais , Eletroforese em Gel de Poliacrilamida , Hypocreales/genética , Virulência
5.
Fungal Biol ; 123(4): 274-282, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30928036

RESUMO

The fungus Purpureocillium lavendulum (formally Paecilomyces lilacinus) is a natural enemy of insects and plant-parasitic nematodes, and has been used as an important bio-control agent against agricultural pests all over the world. In order to understand the genetic mechanisms governing its biocontrol efficiency and other biological processes, an effective gene disruption system is needed. Here we report the development of an efficient system which integrates selective markers that differ from Purpureocillium lilacinum, a one-step construction method for gene knockout plasmids, and a ku80 knockout strain for efficient homologous recombination. With this system, we effectively disrupted the transcription factors in the central regulation pathway of sporulation and a serine protease which were contributed to nematode infection, demonstrating this system as an efficient gene disrupting system for further characterization of genes involved in the development and pathogenesis of this fungus.


Assuntos
Técnicas de Inativação de Genes/métodos , Genética Microbiana/métodos , Hypocreales/genética , Biologia Molecular/métodos , Vetores Genéticos , Recombinação Homóloga , Plasmídeos , Seleção Genética
6.
Plant Dis ; 103(6): 1337-1346, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30958105

RESUMO

Woody plants of the Buxaceae, including species of Buxus, Pachysandra, and Sarcococca, are widely grown evergreen shrubs and groundcovers. Severe leaf spot symptoms were observed on S. hookeriana at the U.S. National Arboretum in Washington, DC, in 2016. Affected plants were growing adjacent to P. terminalis exhibiting Volutella blight symptoms. Fungi isolated from both hosts were identical based on morphology and multilocus phylogenetic analysis and were identified as Coccinonectria pachysandricola (Nectriaceae, Hypocreales), causal agent of Volutella blight of Pachysandra species. Pathogenicity tests established that Co. pachysandricola isolated from both hosts caused disease symptoms on P. terminalis and S. hookeriana, but not on B. sempervirens. Artificial inoculations with Pseudonectria foliicola, causal agent of Volutella blight of B. sempervirens, did not result in disease on P. terminalis or S. hookeriana. Wounding enhanced infection by Co. pachysandricola and Ps. foliicola on all hosts tested but was not required for disease development. Genome assemblies were generated for the Buxaceae pathogens that cause Volutella diseases: Co. pachysandricola, Ps. buxi, and Ps. foliicola; these ranged in size from 25.7 to 28.5 Mb. To our knowledge, this foliar blight of S. hookeriana represents a new disease for this host and is capable of causing considerable damage to infected plants.


Assuntos
Buxaceae , Hypocreales , Buxaceae/microbiologia , Genoma Fúngico/genética , Especificidade de Hospedeiro , Hypocreales/classificação , Hypocreales/citologia , Hypocreales/genética , Filogenia , Washington
7.
Mycologia ; 111(2): 299-318, 2019 Mar-Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30924725

RESUMO

Aquanectria and Gliocladiopsis are two closely related genera of Hypocreales. They are also morphologically similar, forming hyaline, penicillate conidiophores and hyaline, straight to sinuous, 0-1-septate phialoconidia. During a revision of gliocladiopsis-like isolates originating from rain forest areas of South America (Ecuador, French Guiana) and Southeast Asia (Singapore), multilocus phylogenetic inferences, based on DNA sequences encoding partial ß-tubulin (TUB2), translation elongation factor 1-α (TEF1- α), histone H3 (HIS3) genes and the nuc rDNA internal transcribed spacer region (ITS1-5.8S-ITS2 = ITS), revealed the occurrence of seven new phylogenetic species. These phylogenetic species also revealed unique combinations of phenotypes, allowing morphological distinction from their closest phylogenetic relatives. Four new species of Aquanectria and three new species of Gliocladiopsis are described and illustrated. Three of the four Aquanectria species deviate from the other species in the genus by having shorter conidia, which are in the size range observed in Gliocladiopsis species. They are placed in Aquanectria based on the phylogenetic analysis, but this also makes the morphological distinction between these two genera obsolete.


Assuntos
Hypocreales/classificação , Hypocreales/isolamento & purificação , Filogenia , Análise por Conglomerados , DNA Fúngico/química , DNA Fúngico/genética , DNA Ribossômico/química , DNA Ribossômico/genética , DNA Espaçador Ribossômico/química , DNA Espaçador Ribossômico/genética , Equador , Microbiologia Ambiental , Guiana Francesa , Histonas/genética , Hypocreales/genética , Hypocreales/crescimento & desenvolvimento , Microscopia , Fator 1 de Elongação de Peptídeos/genética , RNA Ribossômico 5,8S/genética , Análise de Sequência de DNA , Singapura , Tubulina (Proteína)/genética
8.
BMC Genomics ; 20(1): 120, 2019 Feb 07.
Artigo em Inglês | MEDLINE | ID: mdl-30732559

RESUMO

BACKGROUND: Genes involved in production of secondary metabolites (SMs) in fungi are exceptionally diverse. Even strains of the same species may exhibit differences in metabolite production, a finding that has important implications for drug discovery. Unlike in other eukaryotes, genes producing SMs are often clustered and co-expressed in fungal genomes, but the genetic mechanisms involved in the creation and maintenance of these secondary metabolite biosynthetic gene clusters (SMBGCs) remains poorly understood. RESULTS: In order to address the role of genome architecture and chromosome scale structural variation in generating diversity of SMBGCs, we generated chromosome scale assemblies of six geographically diverse isolates of the insect pathogenic fungus Tolypocladium inflatum, producer of the multi-billion dollar lifesaving immunosuppressant drug cyclosporin, and utilized a Hi-C chromosome conformation capture approach to address the role of genome architecture and structural variation in generating intraspecific diversity in SMBGCs. Our results demonstrate that the exchange of DNA between heterologous chromosomes plays an important role in generating novelty in SMBGCs in fungi. In particular, we demonstrate movement of a polyketide synthase (PKS) and several adjacent genes by translocation to a new chromosome and genomic context, potentially generating a novel PKS cluster. We also provide evidence for inter-chromosomal recombination between nonribosomal peptide synthetases located within subtelomeres and uncover a polymorphic cluster present in only two strains that is closely related to the cluster responsible for biosynthesis of the mycotoxin aflatoxin (AF), a highly carcinogenic compound that is a major public health concern worldwide. In contrast, the cyclosporin cluster, located internally on chromosomes, was conserved across strains, suggesting selective maintenance of this important virulence factor for infection of insects. CONCLUSIONS: This research places the evolution of SMBGCs within the context of whole genome evolution and suggests a role for recombination between chromosomes in generating novel SMBGCs in the medicinal fungus Tolypocladium inflatum.


Assuntos
Cromossomos Fúngicos/genética , Ciclosporina/metabolismo , Rearranjo Gênico , Variação Genética , Hypocreales/genética , Hypocreales/metabolismo , Metabolismo Secundário/genética , Duplicação Cromossômica , Evolução Molecular , Genoma Fúngico/genética , Família Multigênica/genética , Recombinação Genética , Especificidade da Espécie
9.
Mycologia ; 111(1): 78-102, 2019 Jan-Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30657437

RESUMO

Vietnam and Indonesia have rapidly growing and extensive plantation forestry programs, especially of Acacia spp. and Eucalyptus spp. As these plantations expand, the threat from pests and diseases also increases. Calonectria species are among those pathogens causing diseases of trees in plantations and nurseries in these countries. Extensive surveys were conducted across plantations and nurseries of Vietnam and parts of Indonesia, where a large number of Calonectria isolates were retrieved from diseased leaves and soils associated with symptomatic trees. The aim of this study was to identify and resolve the phylogenetic relationships among these isolates using DNA sequence comparisons of four gene regions as well as morphological characters. From a collection of 165 isolates, the study revealed five known and 10 undescribed species. The relatively high diversity of Calonectria species found in this study supports the view that many more species in this genus remain to be discovered in other areas of Southeast Asia.


Assuntos
Variação Genética , Hypocreales/classificação , Hypocreales/genética , Filogenia , Acacia/microbiologia , DNA Fúngico/genética , DNA Espaçador Ribossômico , Eucalyptus/microbiologia , Hypocreales/isolamento & purificação , Indonésia , Doenças das Plantas/microbiologia , Folhas de Planta/microbiologia , Vietnã
10.
Plant Dis ; 103(3): 389-391, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30624143

RESUMO

Corinectria fuckeliana, Neonectria hederae, and N. punicea are fungi in the family Nectriaceae that cause canker diseases of numerous hardwood trees, conifers, and woody perennials, often leading to plant mortality. Here, we report draft genome sequences for these three phytopathogenic fungal species. The genome sizes are consistent with those reported for other members of the Nectriaceae (28 to 43 Mb). These are the first genome resources available for C. fuckeliana, N. hederae, and N. punicea. These genome sequences may provide insights into the mechanisms of virulence and pathogenicity employed by these three destructive plant pathogens, and are resources suitable for the development of molecular markers that could be used for species identification, diagnostic tools and barcodes, and population studies.


Assuntos
Genoma Fúngico , Hypocreales , Árvores , Genoma Fúngico/genética , Hypocreales/genética , Casca de Planta/microbiologia , Doenças das Plantas/microbiologia , Caules de Planta/microbiologia , Árvores/microbiologia , Madeira/microbiologia
11.
Int J Mol Sci ; 19(8)2018 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-30061502

RESUMO

To investigate whether the ech42 gene in Clonostachysrosea can improve the biocontrol efficacy of Bacillus amyloliquefaciens and its molecular mechanism. Compared to the wild type, the B. amyloliquefaciens transformed with the ech42 gene exhibited higher chitinase activity. The B. amyloliquefaciens-ech42 also showed significantly higher biocontrol efficiency compared to Botrytiscinerea when tomato plants were pre-treated with B. amyloliquefaciens-ech42. No significant difference in biocontrol efficiency was observed between the wild type and B.amyloliquefaciens-ech42 when tomato plants were first infected by Botrytiscinerea. In addition, the activity of the defense-related enzyme polyphenol oxidase, but not superoxide dismutase, was significantly higher in B. amyloliquefaciens-ech42 than in the wild type. The ech42 enhances the biocontrol efficiency of B.amyloliquefaciens by increasing the capacity of preventative/curative effects in plants, rather than by killing the pathogens.


Assuntos
Bacillus amyloliquefaciens/genética , Botrytis/fisiologia , Quitinases/genética , Hypocreales/enzimologia , Hypocreales/genética , Lycopersicon esculentum/microbiologia , Doenças das Plantas/microbiologia , Bacillus amyloliquefaciens/enzimologia , Resistência à Doença , Expressão Gênica , Genes Fúngicos , Controle Biológico de Vetores , Proteínas Recombinantes/genética , Transformação Genética
12.
Int J Syst Evol Microbiol ; 68(10): 3255-3259, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30113296

RESUMO

An endophytic strain (designated as strain SYPF 8335T) was isolated from a root of Panax notoginseng in Wenshan district, Yunnan province of China. Strain SYPF 8335T grew very slowly and formed white colonies. Phylogenetic analysis of four loci indicated that strain SYPF 8335T was placed in the Drechmeria clade with Drechmeria campanulata as its closest phylogenetic neighbour. The nucleotide differences between strain SYPF 8335T and D. campanulata are 30 substitutions in the internal transcriber region region. A key morphological feature that differentiates the two fungi is that D. campanulata produces campanulate conidia. Combined with the morphology and molecular analyses, a new species named Drechmeria panacis sp. nov., is proposed.


Assuntos
Hypocreales/classificação , Panax notoginseng/microbiologia , Filogenia , Raízes de Plantas/microbiologia , China , DNA Fúngico , Endófitos/classificação , Endófitos/genética , Endófitos/isolamento & purificação , Hypocreales/genética , Hypocreales/isolamento & purificação , Técnicas de Tipagem Micológica , Análise de Sequência de DNA
13.
J Basic Microbiol ; 58(8): 670-678, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29897133

RESUMO

Ustilaginoidea (U.) virens grows on rice grains and leads to significant rice yield losses in most of the major rice producing areas. Meanwhile, ustiloxins produced by U. virens are a serious hazard to human health and ecological safety of farmlands. The other key point is that ustiloxins have been regarded as a novel resource with their potential in the treatment of cancers. There is no better way to extract ustiloxins than from pure culture of the high ustilotoxin-producing strains. U. virens has become a key research organism. However, due to the presence of some interference components, it is a certain difficulty in the successful isolation of the strain from the false smut balls. We present here a detailed study based on the separation, screening and identification of high ustiloxins-producing strains of U. virens. Through this study, we got a satisfactory success rate of separation and provided a good solution to the problem of separation. At the same time, this study provides quality resources for researchers interested in ustiloxins as anticancer agents.


Assuntos
Hypocreales/isolamento & purificação , Micotoxinas/biossíntese , Oryza , Doenças das Plantas/microbiologia , Antineoplásicos/isolamento & purificação , Cromatografia Líquida , Meios de Cultura , DNA Fúngico/genética , DNA Espaçador Ribossômico/genética , Hypocreales/genética , Hypocreales/crescimento & desenvolvimento , Luz , Micotoxinas/classificação , Micotoxinas/isolamento & purificação , Análise de Sequência de DNA , Espectrometria de Massas em Tandem
14.
J Appl Microbiol ; 125(4): 976-985, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-29856506

RESUMO

AIMS: The objectives of this work were to characterize molecularly the morphologically described endophyte Balansia epichloe symbiotic on three grass species, and to determine the in situ production of ergot alkaloids on these three symbiota. METHODS AND RESULTS: Balansia epichloe symbiotic with smut grass (Sporobolus poiretii), love grass (Eragrostis hirsuta) and lace grass (Eragrostis capillaries, a new host) were characterized using DNA barcoding. Laser ablation electro spray ionization (LAESI)-mass spectrometry was used to detect ergot alkaloids in situ for each symbiotum. CONCLUSIONS: The three morphologically described symbionts on the three host grasses were indicated as belonging to the species B. epichloe, DNA barcoding suggested they were related although a cryptic species was suggested. LAESI-mass spectrometry showed that ergot alkaloids were produced in vivo in two hosts but not the third although this same symbiotum was related to one of the ergot alkaloid producing symbiota as revealed by the DNA-barcoding procedure. SIGNIFICANCE AND IMPACT OF THE STUDY: These results established the accumulation of ergot alkaloids in pot culture by a morpho species although there were variations with each species of grass. Barcoding described divergence among species, but considering its limitation, the suggested existence of cryptic species among this morphospecies requires substantiation by studies that are more rigorous.


Assuntos
Endófitos/metabolismo , Alcaloides de Claviceps/química , Hypocreales/metabolismo , Poaceae/química , Poaceae/microbiologia , Endófitos/química , Endófitos/genética , Endófitos/isolamento & purificação , Alcaloides de Claviceps/metabolismo , Hypocreales/química , Hypocreales/genética , Hypocreales/isolamento & purificação , Espectrometria de Massas , Estrutura Molecular , Filogenia , Simbiose
15.
PLoS Genet ; 14(4): e1007322, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29630596

RESUMO

Unlike most other fungi, molds of the genus Trichoderma (Hypocreales, Ascomycota) are aggressive parasites of other fungi and efficient decomposers of plant biomass. Although nutritional shifts are common among hypocrealean fungi, there are no examples of such broad substrate versatility as that observed in Trichoderma. A phylogenomic analysis of 23 hypocrealean fungi (including nine Trichoderma spp. and the related Escovopsis weberi) revealed that the genus Trichoderma has evolved from an ancestor with limited cellulolytic capability that fed on either fungi or arthropods. The evolutionary analysis of Trichoderma genes encoding plant cell wall-degrading carbohydrate-active enzymes and auxiliary proteins (pcwdCAZome, 122 gene families) based on a gene tree / species tree reconciliation demonstrated that the formation of the genus was accompanied by an unprecedented extent of lateral gene transfer (LGT). Nearly one-half of the genes in Trichoderma pcwdCAZome (41%) were obtained via LGT from plant-associated filamentous fungi belonging to different classes of Ascomycota, while no LGT was observed from other potential donors. In addition to the ability to feed on unrelated fungi (such as Basidiomycota), we also showed that Trichoderma is capable of endoparasitism on a broad range of Ascomycota, including extant LGT donors. This phenomenon was not observed in E. weberi and rarely in other mycoparasitic hypocrealean fungi. Thus, our study suggests that LGT is linked to the ability of Trichoderma to parasitize taxonomically related fungi (up to adelphoparasitism in strict sense). This may have allowed primarily mycotrophic Trichoderma fungi to evolve into decomposers of plant biomass.


Assuntos
Parede Celular/metabolismo , Proteínas Fúngicas/genética , Transferência Genética Horizontal , Plantas/metabolismo , Trichoderma/genética , Basidiomycota/classificação , Basidiomycota/enzimologia , Basidiomycota/genética , Parede Celular/microbiologia , Proteínas Fúngicas/metabolismo , Glicosídeo Hidrolases/genética , Glicosídeo Hidrolases/metabolismo , Interações Hospedeiro-Patógeno , Hifas/enzimologia , Hifas/genética , Hifas/ultraestrutura , Hypocreales/classificação , Hypocreales/enzimologia , Hypocreales/genética , Microscopia Eletrônica de Varredura , Filogenia , Plantas/microbiologia , Trichoderma/enzimologia , Trichoderma/fisiologia
16.
Genome ; 61(5): 329-336, 2018 May.
Artigo em Inglês | MEDLINE | ID: mdl-29514010

RESUMO

Lobster mushroom is a wild edible mushroom with potential commercial value. It is the product resulting of the infection, most commonly of Russula brevipes, by Hypomyces lactifluorum. This study undertook quantitative polymerase chain reaction analysis of tissues sampled at different infection stages to investigate R. brevipes - H. lactifluorum interaction. We followed the colonization of R. brevipes sporocarps by H. lactifluorum that leads to the edible lobster mushrooms. In parallel, metabolomics analysis was performed to detect differences in metabolite profile among non-infected R. brevipes sporocarp and lobster mushroom. The results show that H. lactifluorum DNA is not restricted to the margin but is distributed relatively evenly across the sporocarp of the lobster mushroom. Russula brevipes DNA was also present throughout the sporocarp but was less abundant at the margins and increased inwards. Russula brevipes DNA also declined as the infection progressed. Metabolomics analysis revealed that the flesh of lobster mushroom, which remains identical in appearance to the flesh of the host, undergoes transformation that alters its metabolite profile, most notably of lipids and terpene compounds. These results define a parasitic relationship between the two species that entails a decline of R. brevipes DNA and a modification of its metabolite profile.


Assuntos
Agaricales/metabolismo , DNA Fúngico/genética , Interações Hospedeiro-Patógeno , Hypocreales/metabolismo , Metaboloma , Agaricales/genética , Agaricales/crescimento & desenvolvimento , Alcaloides/metabolismo , Aminoácidos/metabolismo , Cromatografia Líquida de Alta Pressão , Hypocreales/genética , Hypocreales/crescimento & desenvolvimento , Hypocreales/patogenicidade , Metabolismo dos Lipídeos , Lipídeos/química , Fenóis/metabolismo , Reação em Cadeia da Polimerase , Terpenos/metabolismo
17.
Appl Microbiol Biotechnol ; 102(5): 2337-2350, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29396588

RESUMO

We used a temperature differential assay with the opportunistic fungal pathogen Cryptococcus neoformans as a simple screening platform to detect small molecules with antifungal activity in natural product extracts. By screening of a collection extracts from two different strains of the coprophilous fungus, Amphichorda felina, we detected strong, temperature-dependent antifungal activity using a two-plate agar zone of inhibition assay at 25 and 37 °C. Bioassay-guided fractionation of the crude extract followed by liquid chromatography-mass spectrometry (LC-MS) and nuclear magnetic resonance spectroscopy (NMR) identified cyclosporin C (CsC) as the main component of the crude extract responsible for growth inhibition of C. neoformans at 37 °C. The presence of CsC was confirmed by comparison with a commercial standard. We sequenced the genome of A. felina to identify and annotate the CsC biosynthetic gene cluster. The only previously characterized gene cluster for the biosynthesis of similar compounds is that of the related immunosuppressant drug cyclosporine A (CsA). The CsA and CsC gene clusters share a high degree of synteny and sequence similarity. Amino acid changes in the adenylation domain of the CsC nonribosomal peptide synthase's sixth module may be responsible for the substitution of L-threonine compared to L-α-aminobutyric acid in the CsA peptide core. This screening strategy promises to yield additional antifungal natural products with a focused spectrum of antimicrobial activity.


Assuntos
Antifúngicos/farmacologia , Cryptococcus neoformans/efeitos dos fármacos , Ciclosporinas/farmacologia , Hypocreales/química , Antifúngicos/química , Antifúngicos/metabolismo , Cryptococcus neoformans/crescimento & desenvolvimento , Ciclosporinas/química , Ciclosporinas/metabolismo , Hypocreales/genética , Hypocreales/metabolismo , Temperatura Ambiente
18.
Sci Rep ; 8(1): 1123, 2018 01 18.
Artigo em Inglês | MEDLINE | ID: mdl-29348510

RESUMO

Pochonia chlamydosporia infects eggs and females of economically important plant-parasitic nematodes. The fungal isolates parasitizing different nematodes are genetically distinct. To understand their intraspecific genetic differentiation, parasitic mechanisms, and adaptive evolution, we assembled seven putative chromosomes of P. chlamydosporia strain 170 isolated from root-knot nematode eggs (~44 Mb, including 7.19% of transposable elements) and compared them with the genome of the strain 123 (~41 Mb) isolated from cereal cyst nematode. We focus on secretomes of the fungus, which play important roles in pathogenicity and fungus-host/environment interactions, and identified 1,750 secreted proteins, with a high proportion of carboxypeptidases, subtilisins, and chitinases. We analyzed the phylogenies of these genes and predicted new pathogenic molecules. By comparative transcriptome analysis, we found that secreted proteins involved in responses to nutrient stress are mainly comprised of proteases and glycoside hydrolases. Moreover, 32 secreted proteins undergoing positive selection and 71 duplicated gene pairs encoding secreted proteins are identified. Two duplicated pairs encoding secreted glycosyl hydrolases (GH30), which may be related to fungal endophytic process and lost in many insect-pathogenic fungi but exist in nematophagous fungi, are putatively acquired from bacteria by horizontal gene transfer. The results help understanding genetic origins and evolution of parasitism-related genes.


Assuntos
Hypocreales/genética , Hypocreales/metabolismo , Metaboloma , Proteoma , Transcriptoma , Cromossomos Fúngicos , Biologia Computacional/métodos , Duplicação Gênica , Transferência Genética Horizontal , Genômica/métodos , Sequenciamento de Nucleotídeos em Larga Escala , Interações Hospedeiro-Parasita , Interações Hospedeiro-Patógeno , Filogenia , Plantas/microbiologia , Plantas/parasitologia , Seleção Genética
19.
PLoS One ; 13(1): e0185087, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29304036

RESUMO

Thousand Cankers Disease (TCD) of Juglans and Pterocarya (Juglandaceae) involves a fungal pathogen, Geosmithia morbida, and a primary insect vector, Pityophthorus juglandis. TCD was described originally from dying Juglans nigra trees in the western United States (USA), but it was reported subsequently from the eastern USA and northern Italy. The disease is often difficult to diagnose due to the absence of symptoms or signs on the bark surface of the host. Furthermore, disease symptoms can be confused with those caused by other biotic and abiotic agents. Thus, there is a critical need for a method for rapid detection of the pathogen and vector of TCD. Using species-specific microsatellite DNA markers, we developed a molecular protocol for the detection of G. morbida and P. juglandis. To demonstrate the utility of the method for delineating TCD quarantine zones, we tested whether geographical occurrence of symptoms and signs of TCD was correlated with molecular evidence for the presence of the cryptic TCD organisms. A total of 1600 drill cores were taken from branch sections collected from three regions (n = 40 trees for each location): California-J. hindsii (heavy disease incidence); Tennessee-J. nigra (mild disease incidence); and outside the known TCD zone (Missouri-J. nigra, no record of the disease). California samples had the highest incidence of the TCD organisms (85%, 34/40). Tennessee had intermediate incidence (42.5%, 17/40), whereas neither organism was detected in samples from Missouri. The low cost molecular protocol developed here has a high degree of sensitivity and specificity, and it significantly reduces sample-processing time, making the protocol a powerful tool for rapid detection of TCD.


Assuntos
Hypocreales/genética , Hypocreales/patogenicidade , Insetos Vetores/genética , Insetos Vetores/microbiologia , Juglans/microbiologia , Doenças das Plantas/microbiologia , Gorgulhos/genética , Gorgulhos/microbiologia , Animais , California , DNA Fúngico/genética , Técnicas Genéticas , Hypocreales/isolamento & purificação , Repetições de Microssatélites , Missouri , Especificidade da Espécie , Tennessee
20.
Appl Biochem Biotechnol ; 184(3): 1047-1060, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-28942502

RESUMO

Several filamentous fungi are able to concomitantly assimilate both aliphatic and polycyclic aromatic hydrocarbons that are the biogenic by-products of some industrial processes. Cytochrome P450 monooxygenases catalyze the first oxidation reaction for both types of substrate. Among the cytochrome P450 (CYP) genes, the family CYP52 is implicated in the first hydroxylation step in alkane-assimilation processes, while genes belonging to the family CYP53 have been linked with oxidation of aromatic hydrocarbons. Here, we perform a comparative analysis of CYP genes belonging to clans CYP52 and CYP53 in Aspergillus niger, Beauveria bassiana, Metarhizium robertsii (formerly M. anisopliae var. anisopliae), and Penicillium chrysogenum. These species were able to assimilate n-hexadecane, n-octacosane, and phenanthrene, exhibiting a species-dependent modification in pH of the nutrient medium during this process. Modeling of the molecular docking of the hydrocarbons to the cytochrome P450 active site revealed that both phenanthrene and n-octacosane are energetically favored as substrates for the enzymes codified by genes belonging to both CYP52 and CYP53 clans, and thus appear to be involved in this oxidation step. Analyses of gene expression revealed that CYP53 members were significantly induced by phenanthrene in all species studied, but only CYP52X1 and CYP53A11 from B. bassiana were highly induced with n-alkanes. These findings suggest that the set of P450 enzymes involved in hydrocarbon assimilation by fungi is dependent on phylogeny and reveal distinct substrate and expression specificities.


Assuntos
Sistema Enzimático do Citocromo P-450 , Eurotiales , Proteínas Fúngicas , Hidrocarbonetos Cíclicos/metabolismo , Hypocreales , Sistema Enzimático do Citocromo P-450/genética , Sistema Enzimático do Citocromo P-450/metabolismo , Eurotiales/enzimologia , Eurotiales/genética , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Hypocreales/enzimologia , Hypocreales/genética
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