A forward genetic screen in Sclerotinia sclerotiorum revealed the transcriptional regulation of its sclerotial melanisation pathway

Most plant fungal pathogens that cause worldwide crop losses are understudied due to various technical challenges. With the increasing availability of sequenced whole genomes of these non-model fungi, effective genetic analysis methods are highly desirable. Here we describe a newly developed pipeline, which combines forward genetic screening with high-throughput next-generation sequencing to enable quick gene discovery. We applied this pipeline in the notorious soilborne phytopathogen, Sclerotinia sclerotiorum, and identified 32 mutants with various developmental and growth deficiencies. Detailed molecular studies of three melanisation-deficient mutants provide a proof of concept for the effectiveness of our method. A master transcription factor was found to regulate melanisation of sclerotia through the DHN (1,8-dihydroxynaphthalene) melanin biosynthesis pathway. In addition, these mutants revealed that sclerotial melanisation is important for sclerotia survival under abiotic stresses, sclerotial surface structure, and sexual reproduction. Foreseeably, this pipeline can be applied to facilitate efficient in-depth studies of other non-model fungal species in the future.

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Benzyl and Phenyl Glucosinolates Are Metabolized by the Specific Plant Pathogen Alternaria Brassicicola but Not by the Generalist Fungal Pathogens Rhizoctonia Solani or Sclerotinia Sclerotiorum

10.26434/chemrxiv.13681603.v1 ◽

2021 ◽

Author(s):

M. Soledade C. Pedras ◽

Chintamani Thapa ◽

Sajjad Hossain

Keyword(s):

Carboxylic Acid ◽

Rhizoctonia Solani ◽

Sclerotinia Sclerotiorum ◽

Plant Pathogen ◽

Fungal Pathogens ◽

Spectroscopic Characterization ◽

Fungal Species ◽

Alternaria Brassicicola ◽

Metabolic Products

The metabolism of benzyl and phenyl glucosinolates by three phytopathogenic fungal species is investigated and established that <i>A. brassicicola </i>metabolized intact benzyl and phenyl glucosinolates and the corresponding desulfo derivatives. Syntheses and spectroscopic characterization of benzyl and phenyl desulfo-glucosinolates are reported. Phenylacetonitrile and benzylisothiocyanate are the first metabolic products of benzyl glucosinolate; benzyl isothiocyanate is further metabolized to 3-benzyl-2-thioxo-2,3-dihydrothiazolidine-4-carboxylic acid.

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Benzyl and Phenyl Glucosinolates Are Metabolized by the Specific Plant Pathogen Alternaria Brassicicola but Not by the Generalist Fungal Pathogens Rhizoctonia Solani or Sclerotinia Sclerotiorum

10.26434/chemrxiv.13681603 ◽

2021 ◽

Author(s):

M. Soledade C. Pedras ◽

Chintamani Thapa ◽

Sajjad Hossain

Keyword(s):

Carboxylic Acid ◽

Rhizoctonia Solani ◽

Sclerotinia Sclerotiorum ◽

Plant Pathogen ◽

Fungal Pathogens ◽

Spectroscopic Characterization ◽

Fungal Species ◽

Alternaria Brassicicola ◽

Metabolic Products

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Post-translational regulation of autophagy is involved in intra-microbiome suppression of fungal pathogens

Microbiome ◽

10.1186/s40168-021-01077-y ◽

2021 ◽

Vol 9 (1) ◽

Author(s):

Jing Wang ◽

Chaoyun Xu ◽

Qiming Sun ◽

Jinrong Xu ◽

Yunrong Chai ◽

...

Keyword(s):

Translational Regulation ◽

Fungal Pathogens ◽

Fungal Growth ◽

Fungal Species ◽

26S Proteasome ◽

Healthy Plant ◽

Autophagy Induction ◽

Histone Acetyltransferase Gcn5 ◽

Plant Microbiota ◽

Autophagic Process

Abstract Background Microbiome interactions are important determinants for ecosystem functioning, stability, and health. In previous studies, it was often observed that bacteria suppress potentially pathogenic fungal species that are part of the same plant microbiota; however, the underlying microbe-microbe interplay remains mostly elusive. Here, we explored antagonistic interactions of the fungus Fusarium graminearum and bacterium Streptomyces hygroscopicus at the molecular level. Both are ubiquitous members of the healthy wheat microbiota; under dysbiosis, the fungus causes devastating diseases. Results In co-cultures, we found that Streptomyces alters the fungal acetylome leading to substantial induction of fungal autophagy. The bacterium secrets rapamycin to inactivate the target of rapamycin (TOR), which subsequently promotes the degradation of the fungal histone acetyltransferase Gcn5 through the 26S proteasome. Gcn5 negatively regulates fungal autophagy by acetylating the autophagy-related protein Atg8 at the lysine site K13 and blocking cellular relocalization of Atg8. Thus, degradation of Gcn5 triggered by rapamycin was found to reduce Atg8 acetylation, resulting in autophagy induction in F. graminearum. Conclusions Autophagy homeostasis plays an essential role in fungal growth and competition, as well as for virulence. Our work reveals a novel post-translational regulation of autophagy initiated by a bacterial antibiotic. Rapamycin was shown to be a powerful modulator of bacteria–fungi interactions with potential importance in explaining microbial homeostasis in healthy plant microbiomes. The autophagic process provides novel possibilities and targets to biologically control pathogens.

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The role of pH in the melanin biosynthesis pathway.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(18)34190-5 ◽

1982 ◽

Vol 257 (15) ◽

pp. 8738-8744

Author(s):

F G Cánovas ◽

F García-Carmona ◽

J V Sánchez ◽

J L Pastor ◽

J A Teruel

Keyword(s):

Biosynthesis Pathway ◽

Melanin Biosynthesis

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Mitotic Recombination and Adaptive Genomic Changes in Human Pathogenic Fungi

Genes ◽

10.3390/genes10110901 ◽

2019 ◽

Vol 10 (11) ◽

pp. 901 ◽

Cited By ~ 10

Author(s):

Asiya Gusa ◽

Sue Jinks-Robertson

Keyword(s):

Fungal Pathogens ◽

Repetitive Sequences ◽

Pathogenic Fungi ◽

Genetic Alterations ◽

Fungal Species ◽

Chronic Infections ◽

Yeast Saccharomyces Cerevisiae ◽

Genomic Changes ◽

Break Site ◽

Repair Mechanisms

Genome rearrangements and ploidy alterations are important for adaptive change in the pathogenic fungal species Candida and Cryptococcus, which propagate primarily through clonal, asexual reproduction. These changes can occur during mitotic growth and lead to enhanced virulence, drug resistance, and persistence in chronic infections. Examples of microevolution during the course of infection were described in both human infections and mouse models. Recent discoveries defining the role of sexual, parasexual, and unisexual cycles in the evolution of these pathogenic fungi further expanded our understanding of the diversity found in and between species. During mitotic growth, damage to DNA in the form of double-strand breaks (DSBs) is repaired, and genome integrity is restored by the homologous recombination and non-homologous end-joining pathways. In addition to faithful repair, these pathways can introduce minor sequence alterations at the break site or lead to more extensive genetic alterations that include loss of heterozygosity, inversions, duplications, deletions, and translocations. In particular, the prevalence of repetitive sequences in fungal genomes provides opportunities for structural rearrangements to be generated by non-allelic (ectopic) recombination. In this review, we describe DSB repair mechanisms and the types of resulting genome alterations that were documented in the model yeast Saccharomyces cerevisiae. The relevance of similar recombination events to stress- and drug-related adaptations and in generating species diversity are discussed for the human fungal pathogens Candida albicans and Cryptococcus neoformans.

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Melanin biosynthesis pathway in Agaricus bisporus mushrooms

Fungal Genetics and Biology ◽

10.1016/j.fgb.2012.10.004 ◽

2013 ◽

Vol 55 ◽

pp. 42-53 ◽

Cited By ~ 46

Author(s):

A. Weijn ◽

S. Bastiaan-Net ◽

H.J. Wichers ◽

J.J. Mes

Keyword(s):

Agaricus Bisporus ◽

Biosynthesis Pathway ◽

Melanin Biosynthesis

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Modelling Melanin Biosynthesis Pathway with Petri Nets

International Journal of Simulation Systems Science & Technology ◽

10.5013/ijssst.a.22.01.02 ◽

2021 ◽

Author(s):

Mohammed Hathibelagal ◽

Reggie Davidrajuh

Keyword(s):

Petri Nets ◽

Biosynthesis Pathway ◽

Melanin Biosynthesis

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A silencer repressing redundant enhancer activities revealed by deleting endogenouscis-regulatory element ofebonyinDrosophila melanogaster

10.1101/2021.03.25.436947 ◽

2021 ◽

Author(s):

Noriyoshi Akiyama ◽

Shoma Sato ◽

Kentaro M. Tanaka ◽

Takaomi Sakai ◽

Aya Takahashi

Keyword(s):

Drosophila Melanogaster ◽

Genome Editing ◽

Expression Pattern ◽

Regulatory Region ◽

Regulatory Elements ◽

Biosynthesis Pathway ◽

Melanin Biosynthesis ◽

Pigment Synthesis ◽

Endogenous Expression ◽

Spatiotemporal Regulation

AbstractThe spatiotemporal regulation of gene expression is essential to ensure robust phenotypic outcomes. Pigmentation patterns inDrosophilaare formed by the deposition of different pigments synthesized in the developing epidermis and the role ofcis-regulatory elements (CREs) of melanin biosynthesis pathway-related genes is well-characterized. These CREs typically exhibit modular arrangement in the regulatory region of the gene with each enhancer regulating a specific spatiotemporal expression of the gene. However, recent studies have suggested that multiple enhancers of a number of developmental genes as well as those ofyellow(involved in dark pigment synthesis) exhibit redundant activities. Here we report the redundant enhancer activities in thecis-regulatory region of another gene in the melanin biosynthesis pathway,ebony, in the developing epidermis ofDrosophila melanogaster. The evidence was obtained by introducing an approximately 1 kbp deletion at the endogenous primary epidermis enhancer (priEE) by genome editing. The effect of the priEE deletion on pigmentation and on the endogenous expression pattern of amCherry-taggedebonyallele was examined in the thoracic and abdominal segments. The expression level ofebonyin the priEE-deleted strains was similar to that of the control strain, indicating the presence of redundant enhancer activities that drive the broad expression ofebonyin the developing epidermis. Additionally, the priEE fragment contained a silencer that suppressesebonyexpression in the dorsal midline of the abdominal tergites, which is necessary for the development of the subgenusSophophora-specific dark pigmentation patterns along the midline. The endogenous expression pattern ofebonyin the priEE-deleted strains and the reporter assay examining the autonomous activity of the priEE fragment indicated that the silencer is involved in repressing the activities of both proximal and distant enhancers. These results suggest that multiple silencers are dispensable in the regulatory system of a relatively stable taxonomic character. The prevalence of other redundant enhancers and silencers in the genome can be investigated using a similar approach.Author summaryGenes are expressed at the right timing and place to give rise to diverse phenotypes. The spatiotemporal regulation is usually achieved through the coordinated activities of transcription-activating and transcription-repressing proteins that bind to the DNA sequences called enhancers and silencers, respectively, located near the target gene. Most studies identified the locations of enhancers by examining the ability of the sequence fragments to regulate the expression of fused reporters. Various short enhancers have been identified using this approach. This study employed an alternative approach in which the previously identified enhancer that regulates expression ofebony(a gene involved in body color formation) was deleted in a fruitfly,Drosophila melanogaster, using the genome-editing technique. The knockout of this enhancer did not affect the transcription level of the gene to a large extent. This indicated the presence of transcription-activating elements with redundant functions outside the deleted enhancer. Additionally, the transcription ofebonyat the midline of the abdomen, which is repressed in the normal flies, were derepressed in the enhancer-deleted flies, which indicated that the deleted enhancer fragment contained a silencer that negatively regulates multiple enhancer activities in a spatially restricted manner.

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Epidemiology, Clinical Profile and Treatment Outcomes of Bacterial and Fungal Keratitis

10.21203/rs.3.rs-573494/v1 ◽

2021 ◽

Author(s):

Nabila Mabrouk ◽

Mohamed Abdelkader ◽

Mohamed Abdelhakeem ◽

Khaled Mourad ◽

Ahmed Abdelghany

Keyword(s):

Treatment Outcomes ◽

Fungal Pathogens ◽

Fungal Growth ◽

Fungal Species ◽

Clinical Findings ◽

Fungal Keratitis ◽

Clinical Profile ◽

Clinical Series ◽

Vegetative Matter ◽

Department Faculty

Abstract Purpose: The purpose of the study is to determine the microbiological aetiology, epidemiological factors, and clinical profile and treatment outcomes of infective keratitis in Ophthalmology department, Minia University. EgyptMethods:Prospective, non-randomized, observational clinical series of cases, including 150 patients with mean age 30 (range 12 to 85 years), 90 patients (60 %) were males and 60 (40%) were females, clinically diagnosed as infective corneal ulcer, attending the Ophthalmology Department – Faculty of Medicine. Minia University, Minia, Egypt. From 2018 to 2020.Detailed history taking and all clinical findings were collected. Corneal scrapings were obtained from patients and subjected to staining and culture for bacterial and fungal pathogens; Bacterial and fungal growth were identified by standard laboratory procedures.Results:Corneal trauma by a vegetative matter was the commesnest risk factor associated with infective keratitis in 92 cases (61.3%). Smear and Culture was positive in 83 cases (58.4%) of 142 corneal scrapings obtained, of which 60 cases were fungal (72.3%), 21 cases were bacterial (25.3%) and 2 cases were mixed bacterial and fungal (2.4%), Aspergillus species was the commonest fungal species isolated in fungal keratitis. 142 cases (94.67 %) healed completely with scar. Only 6 cases (4%) required evisceration due to aggressive presentation from the start and keratoplasty was performed for 2 cases (1.33%).Conclusions:Fungal keratitis was the commonest type in cases attending to our department. Adequate diagnosis, management and follow up helped in achieving high successful curative outcomes. Clinical Trials.gov ID: NCT04894630. Time of registration 1 December 2018

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