scholarly journals Transcriptome Profiling of Botrytis cinerea Conidial Germination Reveals Upregulation of Infection-Related Genes during the Prepenetration Stage

2013 ◽  
Vol 12 (4) ◽  
pp. 614-626 ◽  
Author(s):  
Michaela Leroch ◽  
Astrid Kleber ◽  
Evelyn Silva ◽  
Tina Coenen ◽  
Dieter Koppenhöfer ◽  
...  
Keyword(s):  
Gene Expression ◽  
Botrytis Cinerea ◽  
Transcriptome Profiling ◽  
Gray Mold ◽  
Secretory Proteins ◽  
Global Changes ◽  
Lytic Enzymes ◽  
Chitin Deacetylase ◽  
Specific Expression ◽  
Content Type

ABSTRACTBotrytis cinereacauses gray mold on a great number of host plants. Infection is initiated by airborne conidia that invade the host tissue, often by penetration of intact epidermal cells. To mimic the surface properties of natural plant surfaces, conidia were incubated on apple wax-coated surfaces, resulting in rapid germination and appressorium formation. Global changes in gene expression were analyzed by microarray hybridization between conidia incubated for 0 h (dormant), 1 h (pregermination), 2.5 h (postgermination), 4 h (appressoria), and 15 h (early mycelium). Considerable changes were observed, in particular between 0 h and 1 h. Genes induced during germination were enriched in those genes encoding secreted proteins, including lytic enzymes. Comparison of wild-type and a nonpathogenic MAP kinase mutant (bmp1) revealed marked differences in germination-related gene expression, in particular related to secretory proteins. Using promoter-GFP reporter strains, we detected a strictly germination-specific expression pattern of a putative chitin deacetylase gene (cda1). In contrast, a cutinase gene (cutB) was found to be expressed only in the presence of plant lipids, in a developmentally less stringent pattern. We also identified a coregulated gene cluster possibly involved in secondary metabolite synthesis which was found to be controlled by a transcription factor also encoded in this cluster. Our data demonstrate that early conidial development inB. cinereais accompanied by rapid shifts in gene expression that prepare the fungus for germ tube outgrowth and host cell invasion.

Streptomyces botrytidirepellens sp. nov., a novel actinomycete with antifungal activity against Botrytis cinerea

2021 ◽  
Vol 71 (9) ◽  
Author(s):  
Mengqi Jiang ◽  
Xi Xu ◽  
Jia Song ◽  
Dongmei Li ◽  
Liyuan Han ◽  
...  
Keyword(s):  
Antifungal Activity ◽  
Botrytis Cinerea ◽  
Gene Clusters ◽  
Gray Mold ◽  
Phytopathogenic Fungi ◽  
Antagonistic Effect ◽  
Economic Losses ◽  
Content Type ◽  
Desferrioxamine B ◽  
Cell Free Filtrate

The fungal pathogen Botrytis cinerea is the causal agent of devastating gray mold diseases in many economically important fruits, vegetables, and flowers, leading to serious economic losses worldwide. In this study, a novel actinomycete NEAU-LD23T exhibiting antifungal activity against B. cinerea was isolated, and its taxonomic position was evaluated using a polyphasic approach. Based on the genotypic, phenotypic and chemotaxonomic data, it is concluded that the strain represents a novel species within the genus Streptomyces , for which the name Streptomyces botrytidirepellens sp. nov. is proposed. The type strain is NEAU-LD23T (=CCTCC AA 2019029T=DSM 109824T). In addition, strain NEAU-LD23T showed a strong antagonistic effect against B. cinerea (82.6±2.5%) and varying degrees of inhibition on nine other phytopathogenic fungi. Both cell-free filtrate and methanol extract of mycelia of strain NEAU-LD23T significantly inhibited mycelial growth of B. cinerea. To preliminarily explore the antifungal mechanisms, the genome of strain NEAU-LD23T was sequenced and analyzed. AntiSMASH analysis led to the identification of several gene clusters responsible for the biosynthesis of bioactive secondary metabolites with antifungal activity, including 9-methylstreptimidone, echosides, anisomycin, coelichelin and desferrioxamine B. Overall, this research provided us an excellent strain with considerable potential to use for biological control of tomato gray mold.

scholarly journals Single-Cell Analysis Reveals Distinct Gene Expression and Heterogeneity in Male and FemalePlasmodium falciparumGametocytes

mSphere ◽  
2018 ◽  
Vol 3 (2) ◽  
Author(s):  
Katelyn A. Walzer ◽  
Danielle M. Kubicki ◽  
Xiaohu Tang ◽  
Jen-Tsan Ashley Chi
Keyword(s):  
Gene Expression ◽  
Plasmodium Falciparum ◽  
Single Cell ◽  
Specific Expression ◽  
Male And Female ◽  
Content Type ◽  
Population Analyses ◽  
Specific Differentiation ◽  
Female Specific ◽  
Gender Specific

ABSTRACTSexual reproduction is an obligate step in thePlasmodium falciparumlife cycle, with mature gametocytes being the only form of the parasite capable of human-to-mosquito transmission. Development of male and female gametocytes takes 9 to 12 days, and although more than 300 genes are thought to be specific to gametocytes, only a few have been postulated to be male or female specific. Because these genes are often expressed during late gametocyte stages and for some, male- or female-specific transcript expression is debated, the separation of male and female populations is technically challenging. To overcome these challenges, we have developed an unbiased single-cell approach to determine which transcripts are expressed in male versus female gametocytes. Using microfluidic technology, we isolated single mid- to late-stage gametocytes to compare the expression of 91 genes, including 87 gametocyte-specific genes, in 90 cells. Such analysis identified distinct gene clusters whose expression was associated with male, female, or all gametocytes. In addition, a small number of male gametocytes clustered separately from female gametocytes based on sex-specific expression independent of stage. Many female-enriched genes also exhibited stage-specific expression. RNA fluorescentin situhybridization of male and female markers validated the mutually exclusive expression pattern of male and female transcripts in gametocytes. These analyses uncovered novel male and female markers that are expressed as early as stage III gametocytogenesis, providing further insight intoPlasmodiumsex-specific differentiation previously masked in population analyses. Our single-cell approach reveals the most robust markers for sex-specific differentiation inPlasmodiumgametocytes. Such single-cell expression assays can be generalized to all eukaryotic pathogens.IMPORTANCEMost human deaths that result from malaria are caused by the eukaryotic parasitePlasmodium falciparum. The only form of this parasite that is transmitted to the mosquito is the sexual form, called the gametocyte. The production of mature gametocytes can take up to 2 weeks and results in phenotypically distinct males and females, although what causes this gender-specific differentiation remains largely unknown. Here, we demonstrate the first use of microfluidic technology to capture single gametocytes and determine their temporal sex-specific gene expression in an unbiased manner. We were able to determine male or female identity of single cells based on the upregulation of gender-specific genes as early as mid-stage gametocytes. This analysis has revealed strong markers for male and female gametocyte differentiation that were previously concealed in population analyses. Similar single-cell analyses in eukaryotic pathogens using this method may uncover rare cell types and heterogeneity previously masked in population studies.

scholarly journals Botrytis fragariae, a New Species Causing Gray Mold on Strawberries, Shows High Frequencies of Specific and Efflux-Based Fungicide Resistance

2017 ◽  
Vol 83 (9) ◽  
Author(s):  
Sabrina Rupp ◽  
Cecilia Plesken ◽  
Sibylle Rumsey ◽  
Madeline Dowling ◽  
Guido Schnabel ◽  
...  
Keyword(s):  
United States ◽  
New Species ◽  
Multidrug Resistance ◽  
Botrytis Cinerea ◽  
Fungicide Resistance ◽  
Southeastern United States ◽  
Gray Mold ◽  
Diagnostic Pcr ◽  
Content Type ◽  
A New Species

ABSTRACT Botrytis cinerea causes pre- and postharvest decay of many fruit and vegetable crops. A survey of German strawberry fields revealed Botrytis strains that differed from B. cinerea in diagnostic PCR markers and growth appearance. Phylogenetic analyses showed that these strains belong to an undescribed species in Botrytis clade 2, named Botrytis fragariae sp. nov. Isolates of B. fragariae were detected in strawberry fields throughout Germany, sometimes at frequencies similar to those of B. cinerea, and in the southeastern United States. B. fragariae was isolated from overwintering strawberry tissue but not from freshly infected fruit. B. fragariae invaded strawberry tissues with an efficiency similar to or lower than that of B. cinerea but showed poor colonization of inoculated nonhost plant tissues. These data and the exclusive occurrence of this fungus on strawberry plants indicate that B. fragariae is host specific and has a tissue preference different from that of B. cinerea. Various fungicide resistance patterns were observed in B. fragariae populations. Many B. fragariae strains showed resistance to one or several chemical classes of fungicides and an efflux-based multidrug resistance (MDR1) phenotype previously described in B. cinerea. Resistance-related mutations in B. fragariae were identical or similar to those of B. cinerea for carbendazim (E198A mutation in tubA), azoxystrobin (G143A in cytB), iprodione (G367A+V368F in bos1), and MDR1 (gain-of-function mutations in the transcription factor mrr1 gene and overexpression of the drug efflux transporter gene atrB). The widespread occurrence of B. fragariae indicates that this species is adapted to fungicide-treated strawberry fields and may be of local importance as a gray mold pathogen alongside B. cinerea. IMPORTANCE Gray mold is the most important fruit rot on strawberries worldwide and requires fungicide treatments for control. For a long time, it was believed to be caused only by Botrytis cinerea, a ubiquitous pathogen with a broad host range that quickly develops fungicide resistance. We report the discovery and description of a new species, named Botrytis fragariae, that is widely distributed in commercial strawberry fields in Germany and the southeastern United States. It was observed on overwintering tissue but not on freshly infected fruit and seems host specific on the basis of its occurrence and artificial infection tests. B. fragariae has also developed resistance to several fungicides that is caused by mutations similar to those known in B. cinerea, including an efflux-based multidrug resistance. Our data indicate that B. fragariae could be of practical importance as a strawberry pathogen in some regions where its abundance is similar to that of B. cinerea.

scholarly journals Novel Hypovirulence-Associated RNA Mycovirus in the Plant-Pathogenic Fungus Botrytis cinerea: Molecular and Biological Characterization

2015 ◽  
Vol 81 (7) ◽  
pp. 2299-2310 ◽  
Author(s):  
Lin Yu ◽  
Wen Sang ◽  
Ming-De Wu ◽  
Jing Zhang ◽  
Long Yang ◽  
...  
Keyword(s):  
Botrytis Cinerea ◽  
Fluorescent Protein ◽  
Rna Virus ◽  
Pathogenic Fungus ◽  
Ornamental Plants ◽  
Gray Mold ◽  
Open Reading Frames ◽  
Content Type ◽  
Molecular Features ◽  
Green Fluorescent

ABSTRACTBotrytis cinereais a pathogenic fungus causing gray mold on numerous economically important crops and ornamental plants. This study was conducted to characterize the biological and molecular features of a novel RNA mycovirus, Botrytis cinerea RNA virus 1 (BcRV1), in the hypovirulent strain BerBc-1 ofB. cinerea. The genome of BcRV1 is 8,952 bp long with two putative overlapped open reading frames (ORFs), ORF1 and ORF2, coding for a hypothetical polypeptide (P1) and RNA-dependent RNA polymerase (RdRp), respectively. A −1 frameshifting region (designated the KNOT element) containing a shifty heptamer, a heptanucleotide spacer, and an H-type pseudoknot was predicted in the junction region of ORF1 and ORF2. The −1 frameshifting role of the KNOT element was experimentally confirmed through determination of the production of the fusion protein red fluorescent protein (RFP)-green fluorescent protein (GFP) by the plasmid containing the constructdsRed-KNOT-eGFPinEscherichia coli. BcRV1 belongs to a taxonomically unassigned double-stranded RNA (dsRNA) mycovirus group. It is closely related to grapevine-associated totivirus 2 and Sclerotinia sclerotiorum nonsegmented virus L. BcRV1 in strain BerBc-1 was found capable of being transmitted vertically through macroconidia and horizontally to otherB. cinereastrains through hyphal contact. The presence of BcRV1 was found to be positively correlated with hypovirulence inB. cinerea, with the attenuation effects of BcRV1 on mycelial growth and pathogenicity being greatly affected by the accumulation level of BcRV1.

scholarly journals The Transcriptional Aftermath in Two Independently Formed Hybrids of the Opportunistic Pathogen Candida orthopsilosis

mSphere ◽  
2020 ◽  
Vol 5 (3) ◽  
Author(s):  
Hrant Hovhannisyan ◽  
Ester Saus ◽  
Ewa Ksiezopolska ◽  
Toni Gabaldón
Keyword(s):  
Gene Expression ◽  
Loss Of Heterozygosity ◽  
Opportunistic Pathogen ◽  
Specific Gene ◽  
Specific Expression ◽  
Allele Specific Expression ◽  
Content Type ◽  
Additional Layer ◽  
Allele Specific ◽  
Candida Orthopsilosis

ABSTRACT Interspecific hybridization can drive evolutionary adaptation to novel environments. The Saccharomycotina clade of budding yeasts includes many hybrid lineages, and hybridization has been proposed as a source for new pathogenic species. Candida orthopsilosis is an emerging opportunistic pathogen for which most clinical isolates are hybrids, each derived from one of at least four independent crosses between the same two parental lineages. To gain insight into the transcriptomic aftermath of hybridization in these pathogens, we analyzed allele-specific gene expression in two independently formed hybrid strains and in a homozygous strain representative of one parental lineage. Our results show that the effect of hybridization on overall gene expression is rather limited, affecting ∼4% of the genes studied. However, we identified a larger effect in terms of imbalanced allelic expression, affecting ∼9.5% of the heterozygous genes in the hybrids. This effect was larger in the hybrid with more extensive loss of heterozygosity, which may indicate a tendency to avoid loss of heterozygosity in these genes. Consistently, the number of shared genes with allele-specific expression in the two independently formed hybrids was higher than random expectation, suggesting selective retention. Some of the imbalanced genes have functions related to pathogenicity, including zinc transport and superoxide dismutase activities. While it remains unclear whether the observed imbalanced genes play a role in virulence, our results suggest that differences in allele-specific expression may add an additional layer of phenotypic plasticity to traits related to virulence in C. orthopsilosis hybrids. IMPORTANCE How new pathogens emerge is an important question that remains largely unanswered. Some emerging yeast pathogens are hybrids originated through the crossing of two different species, but how hybridization contributes to higher virulence is unclear. Here, we show that hybrids selectively retain gene regulation plasticity inherited from the two parents and that this plasticity affects genes involved in virulence.

scholarly journals Global Changes in Mycoplasma gallisepticum Phase-Variable Lipoprotein GenevlhAExpression duringIn VivoInfection of the Natural Chicken Host

2015 ◽  
Vol 84 (1) ◽  
pp. 351-355 ◽  
Author(s):  
K. Pflaum ◽  
E. R. Tulman ◽  
J. Beaudet ◽  
X. Liao ◽  
S. J. Geary
Keyword(s):  
Gene Expression ◽  
Phase Variation ◽  
Mycoplasma Gallisepticum ◽  
Chronic Respiratory Disease ◽  
Etiologic Agent ◽  
Economic Losses ◽  
Global Changes ◽  
Phase Variable ◽  
Content Type ◽  
Host Interaction

Mycoplasma gallisepticumis the primary etiologic agent of chronic respiratory disease in poultry, a disease largely affecting the respiratory tract and causing significant economic losses worldwide. Immunodominant proteins encoded by members of the variable lipoprotein and hemagglutinin (vlhA) gene family are thought to be important for mechanisms ofM. gallisepticum-host interaction, pathogenesis, and immune evasion, but their exact role and the overall nature of their phase variation are unknown. To better understand these mechanisms, we assessed global transcriptomicvlhAgene expression directly fromM. gallisepticumpopulations present on tracheal mucosae during a 7-day experimental infection in the natural chicken host. Here we report differences in both dominant and minorvlhAgene expression levels throughout the first week of infection and starting as early as day 1 postinfection, consistent with a functional role not dependent on adaptive immunity for driving phase variation. Notably, data indicated that, at given time points, specificvlhAgenes were similarly dominant in multiple independent hosts, suggesting a nonstochastic temporal progression of dominantvlhAgene expression in the colonizing bacterial population. The dominant expression of a givenvlhAgene was not dependent on the presence of 12-copy GAA trinucleotide repeats in the promoter region and did not revert to the predominatevlhAgene when no longer faced with host pressures. Overall, these data indicate thatvlhAphase variation is dynamic throughout the earliest stages of infection and that the pattern of dominantvlhAexpression may be nonrandom and regulated by previously unrecognized mechanisms.

scholarly journals Living Colors in the Gray Mold Pathogen Botrytis cinerea: Codon-Optimized Genes Encoding Green Fluorescent Protein and mCherry, Which Exhibit Bright Fluorescence

2011 ◽  
Vol 77 (9) ◽  
pp. 2887-2897 ◽  
Author(s):  
Michaela Leroch ◽  
Dennis Mernke ◽  
Dieter Koppenhoefer ◽  
Prisca Schneider ◽  
Andreas Mosbach ◽  
...  
Keyword(s):  
Green Fluorescent Protein ◽  
Botrytis Cinerea ◽  
Fluorescent Protein ◽  
Gc Content ◽  
Gray Mold ◽  
Mrna Levels ◽  
Content Type ◽  
Mold Fungus ◽  
Encoding Gene ◽  
Green Fluorescent

ABSTRACTThe green fluorescent protein (GFP) and its variants have been widely used in modern biology as reporters that allow a variety of live-cell imaging techniques. So far, GFP has rarely been used in the gray mold fungusBotrytis cinereabecause of low fluorescence intensity. The codon usage ofB. cinereagenes strongly deviates from that of commonly used GFP-encoding genes and reveals a lower GC content than other fungi. In this study, we report the development and use of a codon-optimized version of theB. cinereaenhanced GFP (eGFP)-encoding gene (Bcgfp) for improved expression inB. cinerea. Both the codon optimization and, to a smaller extent, the insertion of an intron resulted in higher mRNA levels and increased fluorescence. Bcgfpwas used for localization of nuclei in germinating spores and for visualizing host penetration. We further demonstrate the use of promoter-Bcgfpfusions for quantitative evaluation of various toxic compounds as inducers of theatrBgene encoding an ABC-type drug efflux transporter ofB. cinerea. In addition, a codon-optimized mCherry-encoding gene was constructed which yielded bright red fluorescence inB. cinerea.

scholarly journals Compartmentalization of Melanin Biosynthetic Enzymes Contributes to Self-Defense against Intermediate Compound Scytalone in Botrytis cinerea

mBio ◽  
2021 ◽  
Vol 12 (2) ◽  
Author(s):  
Xue Chen ◽  
Chuanxi Zhu ◽  
Yantao Na ◽  
Dandan Ren ◽  
Chenghua Zhang ◽  
...  
Keyword(s):  
Chemical Analysis ◽  
Cell Surface ◽  
Botrytis Cinerea ◽  
Filamentous Fungi ◽  
Gray Mold ◽  
Biosynthetic Enzymes ◽  
Melanin Synthesis ◽  
Melanin Biosynthesis ◽  
Content Type ◽  
Sclerotial Germination

ABSTRACT In filamentous fungi, 1,8-dihydroxynaphthalene (DHN) melanin is a major component of the extracellular matrix, endowing fungi with environmental tolerance and some pathogenic species with pathogenicity. However, the subcellular location of the melanin biosynthesis pathway components remains obscure. Using the gray mold pathogen Botrytis cinerea, the DHN melanin intermediate scytalone was characterized via phenotypic and chemical analysis of mutants, and the key enzymes participating in melanin synthesis were fused with fluorescent proteins to observe their subcellular localizations. The Δbcscd1 mutant accumulated scytalone in the culture filtrate rather than in mycelium. Excessive scytalone appears to be self-inhibitory to the fungus, leading to repressed sclerotial germination and sporulation in the Δbcscd1 mutant. The BcBRN1/2 enzymes responsible for synthesizing scytalone were localized in endosomes and found to be trafficked to the cell surface, accompanied by the accumulation of BcSCD1 proteins in the cell wall. In contrast, the early-stage melanin synthesis enzymes BcPKS12/13 and BcYGH1 were localized in peroxisomes. Taken together, the results of this study revealed the subcellular distribution of melanin biosynthetic enzymes in B. cinerea, indicating that the encapsulation and externalization of the melanin synthetic enzymes need to be delicately orchestrated to ensure enzymatic efficiency and protect itself from the adverse effect of the toxic intermediate metabolite. IMPORTANCE The devastating gray mold pathogen Botrytis cinerea propagates via melanized conidia and sclerotia. This study reveals that the sclerotial germination of B. cinerea is differentially affected by different enzymes in the melanin synthesis pathway. Using gene knockout mutants and chemical analysis, we found that excessive accumulation of the melanin intermediate scytalone is inhibitory to B. cinerea. Subcellular localization analysis of the melanin synthesis enzymes of B. cinerea suggested two-stage partitioning of the melanogenesis pathway: the intracellular stage involves the steps until the intermediate scytalone was translocated to the cell surface, whereas the extracellular stage comprises all the steps occurring in the wall from scytalone to final melanin formation. These strategies make the fungus avert self-poisoning during melanin production. This study opens avenues for better understanding the mechanisms of secondary metabolite production in filamentous fungi.

scholarly journals Transcriptomic analysis of Pseudomonas ogarae F113 reveals the antagonistic roles of AmrZ and FleQ during rhizosphere adaption

2022 ◽  
Vol 8 (1) ◽  
Author(s):  
Esther Blanco-Romero ◽  
David Durán ◽  
Daniel Garrido-Sanz ◽  
Rafael Rivilla ◽  
Marta Martín ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transcription Factors ◽  
Biofilm Formation ◽  
Type Species ◽  
Global Changes ◽  
Secretion Systems ◽  
Content Type ◽  
Rhizosphere Colonization ◽  
Link Type ◽  
Metabolic Versatility

Rhizosphere colonization by bacteria involves molecular and cellular mechanisms, such as motility and chemotaxis, biofilm formation, metabolic versatility, or biosynthesis of secondary metabolites, among others. Nonetheless, there is limited knowledge concerning the main regulatory factors that drive the rhizosphere colonization process. Here we show the importance of the AmrZ and FleQ transcription factors for adaption in the plant growth-promoting rhizobacterium (PGPR) and rhizosphere colonization model Pseudomonas ogarae F113. RNA-Seq analyses of P. ogarae F113 grown in liquid cultures either in exponential and stationary growth phase, and rhizosphere conditions, revealed that rhizosphere is a key driver of global changes in gene expression in this bacterium. Regarding the genetic background, this work has revealed that a mutation in fleQ causes considerably more alterations in the gene expression profile of this bacterium than a mutation in amrZ under rhizosphere conditions. The functional analysis has revealed that in P. ogarae F113, the transcription factors AmrZ and FleQ regulate genes involved in diverse bacterial functions. Notably, in the rhizosphere, these transcription factors antagonistically regulate genes related to motility, biofilm formation, nitrogen, sulfur, and amino acid metabolism, transport, signalling, and secretion, especially the type VI secretion systems. These results define the regulon of two important bifunctional transcriptional regulators in pseudomonads during the process of rhizosphere colonization.

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