scholarly journals Sphingolipid C-9 Methyltransferases Are Important for Growth and Virulence but Not for Sensitivity to Antifungal Plant Defensins in Fusarium graminearum

2008 ◽  
Vol 8 (2) ◽  
pp. 217-229 ◽  
Author(s):  
Vellaisamy Ramamoorthy ◽  
Edgar B. Cahoon ◽  
Mercy Thokala ◽  
Jagdeep Kaur ◽  
Jia Li ◽  
...  
Keyword(s):  
Fusarium Graminearum ◽  
Type Strain ◽  
Wild Type Strain ◽  
Fungal Growth ◽  
Wild Type ◽  
Knockout Mutant ◽  
Double Knockout ◽  
Disease Symptoms ◽  
Plant Defensins ◽  
Antifungal Action

ABSTRACT The C-9-methylated glucosylceramides (GlcCers) are sphingolipids unique to fungi. They play important roles in fungal growth and pathogenesis, and they act as receptors for some antifungal plant defensins. We have identified two genes, FgMT1 and FgMT2, that each encode a putative sphingolipid C-9 methyltransferase (C-9-MT) in the fungal pathogen Fusarium graminearum and complement a Pichia pastoris C-9-MT-null mutant. The ΔFgmt1 mutant produced C-9-methylated GlcCer like the wild-type strain, PH-1, whereas the ΔFgmt2 mutant produced 65 to 75% nonmethylated and 25 to 35% methylated GlcCer. No ΔFgmt1ΔFgmt2 double-knockout mutant producing only nonmethylated GlcCer could be recovered, suggesting that perhaps C-9-MTs are essential in this pathogen. This is in contrast to the nonessential nature of this enzyme in the unicellular fungus P. pastoris. The ΔFgmt2 mutant exhibited severe growth defects and produced abnormal conidia, while the ΔFgmt1 mutant grew like the wild-type strain, PH-1, under the conditions tested. The ΔFgmt2 mutant also exhibited drastically reduced disease symptoms in wheat and much-delayed disease symptoms in Arabidopsis thaliana. Surprisingly, the ΔFgmt2 mutant was less virulent on different host plants tested than the previously characterized ΔFggcs1 mutant, which lacks GlcCer synthase activity and produces no GlcCer at all. Moreover, the ΔFgmt1 and ΔFgmt2 mutants, as well as the P. pastoris strain in which the C-9-MT gene was deleted, retained sensitivity to the antifungal plant defensins MsDef1 and RsAFP2, indicating that the C-9 methyl group is not a critical structural feature of the GlcCer receptor required for the antifungal action of plant defensins.

scholarly journals A Natural Mutation Involving both Pathogenicity and Perithecium Formation in the Fusarium graminearum Species Complex

2016 ◽  
Vol 6 (12) ◽  
pp. 3883-3892 ◽  
Author(s):  
Haruhisha Suga ◽  
Koji Kageyama ◽  
Masafumi Shimizu ◽  
Misturo Hyakumachi
Keyword(s):  
Mutant Strain ◽  
Fusarium Graminearum ◽  
Type Strain ◽  
Species Complex ◽  
Wild Type Strain ◽  
Genetic Locus ◽  
Chromosome 2 ◽  
Wild Type ◽  
Head Blight ◽  
Fusarium Graminearum Species Complex

Abstract Members of the Fusarium graminearum species complex (Fg complex or FGSC) are the primary pathogens causing Fusarium head blight in wheat and barley worldwide. A natural pathogenicity mutant (strain 0225022) was found in a sample of the Fg complex collected in Japan. The mutant strain did not induce symptoms in wheat spikes beyond the point of inoculation, and did not form perithecia. No segregation of phenotypic deficiencies occurred in the progenies of a cross between the mutant and a fully pathogenic wild-type strain, which suggested that a single genetic locus controlled both traits. The locus was mapped to chromosome 2 by using sequence-tagged markers; and a deletion of ∼3 kb was detected in the mapped region of the mutant strain. The wild-type strain contains the FGSG_02810 gene, encoding a putative glycosylphosphatidylinositol anchor protein, in this region. The contribution of FGSG_02810 to pathogenicity and perithecium formation was confirmed by complementation in the mutant strain using gene transfer, and by gene disruption in the wild-type strain.

scholarly journals A sterol C-14 reductase encoded by FgERG24B is responsible for the intrinsic resistance of Fusarium graminearum to amine fungicides

Microbiology ◽  
2011 ◽  
Vol 157 (6) ◽  
pp. 1665-1675 ◽  
Author(s):  
Xin Liu ◽  
Jing Fu ◽  
Yingzi Yun ◽  
Yanni Yin ◽  
Zhonghua Ma
Keyword(s):  
Fusarium Graminearum ◽  
Type Strain ◽  
Wild Type Strain ◽  
Ergosterol Biosynthesis ◽  
Deletion Mutants ◽  
Wild Type ◽  
Head Blight ◽  
Intrinsic Resistance ◽  
In The Wild ◽  
Increased Sensitivity

Fusarium graminearum, the causal agent of wheat head blight, shows intrinsic resistance to amine fungicides. It is commonly accepted that the amines target sterol C-14 reductase and sterol Δ8–Δ7 isomerase of ergosterol biosynthesis, encoded by the genes ERG24 and ERG2, respectively. Analysis of the genome sequence of F. graminearum revealed that the fungus contains two paralogous FgERG24 genes (FgERG24A and FgERG24B), which are homologous to the ERG24 of Saccharomyces cerevisiae. In this study, we disrupted FgERG24A and FgERG24B in F. graminearum. Compared to the wild-type strain HN9-1, FgERG24A and FgERG24B deletion mutants did not show recognizable phenotypic changes in mycelial growth on potato dextrose agar or in virulence on wheat heads. HPLC analysis showed that the amount of ergosterol in FgERG24A or FgERG24B deletion mutants was not significantly different from that in the wild-type strain. These results indicate that neither of the two genes is essential for growth, pathogenicity or ergosterol biosynthesis in F. graminearum. FgERG24B deletion mutants exhibited significantly increased sensitivity to amine fungicides, including tridemorph, fenpropidin and spiroxamine, but not to non-amine fungicides. In contrast, FgERG24A deletion mutants did not show changed sensitivity to any amine tested. The resistance of the FgERG24B deletion mutant to amines was restored by genetic complementation of the mutant with wild-type FgERG24B. These results indicate that FgERG24B controls the intrinsic resistance of F. graminearum to amines. The finding of this study provides new insights into amine resistance in filamentous fungi.

scholarly journals Functional Characterization of Calcineurin-Responsive Transcription Factors Fg01341 and Fg01350 in Fusarium graminearum

2020 ◽  
Vol 11 ◽  
Author(s):  
Xiangxiang Zhang ◽  
Shulin Cao ◽  
Wei Li ◽  
Haiyan Sun ◽  
Yuanyu Deng ◽  
...  
Keyword(s):  
Fusarium Graminearum ◽  
Type Strain ◽  
Deletion Mutant ◽  
Wild Type Strain ◽  
Functional Characterization ◽  
Hyphal Growth ◽  
Basic Medium ◽  
Dependent Manner ◽  
Wild Type ◽  
Sexual Production

Ca2 +/calmodulin-dependent phosphatase calcineurin is one of the important regulators of intracellular calcium homeostasis and has been investigated extensively in Saccharomyces cerevisiae. However, only a few reports have explored the function of the Crz1 homolog in filamentous fungi, especially in Fusarium graminearum. In this study, we identified Fg01341 as a potential ortholog of yeast Crz1. Fg01341 could interact with calcineurin and initiate nuclear transport in a calcineurin-dependent manner. The ΔFg01341 mutant exhibited normal hyphal growth on basic medium and conidia formation, but sexual reproduction was partially blocked. Pathogenicity assays showed that the virulence of the ΔFg01341 mutant in flowering wheat heads and corn silks dramatically decreased and was thus consistent with the reduction in deoxynivalenol production. Unexpectedly, the sensitivity to osmotic stress of the deletion mutant and that of the wild-type strain did not present any differences. The deletion mutant showed higher sensitivity to tebuconazole than the wild-type strain. Results also showed that the transcription factor Fg01350 might be the calcineurin target and was independent of Crz1. Furthermore, ΔFg01350 showed defects in hyphal growth, sexual production, virulence, and deoxynivalenol production. Collectively, the results indicate that these two proteins functionally redundant and that the calcineurin–Crz1-independent pathway is particularly important in F. graminearum.

scholarly journals A ClpB Chaperone Knockout Mutant of Mesorhizobium ciceri Shows a Delay in the Root Nodulation of Chickpea Plants

2012 ◽  
Vol 25 (12) ◽  
pp. 1594-1604 ◽  
Author(s):  
Clarisse Brígido ◽  
Marta Robledo ◽  
Esther Menéndez ◽  
Pedro F. Mateos ◽  
Solange Oliveira
Keyword(s):  
Type Strain ◽  
Wild Type Strain ◽  
Fluorescent Protein ◽  
Root Nodules ◽  
Strain Analysis ◽  
Wild Type ◽  
Knockout Mutant ◽  
Mesorhizobium Ciceri ◽  
Acid Shock ◽  
Green Fluorescent

Several molecular chaperones are known to be involved in bacteria stress response. To investigate the role of chaperone ClpB in rhizobia stress tolerance as well as in the rhizobia-plant symbiosis process, the clpB gene from a chickpea microsymbiont, strain Mesorhizobium ciceri LMS-1, was identified and a knockout mutant was obtained. The ClpB knockout mutant was tested to several abiotic stresses, showing that it was unable to grow after a heat shock and it was more sensitive to acid shock than the wild-type strain. A plant-growth assay performed to evaluate the symbiotic performance of the clpB mutant showed a higher proportion of ineffective root nodules obtained with the mutant than with the wild-type strain. Nodulation kinetics analysis showed a 6- to 8-day delay in nodule appearance in plants inoculated with the ΔclpB mutant. Analysis of nodC gene expression showed lower levels of transcript in the ΔclpB mutant strain. Analysis of histological sections of nodules formed by the clpB mutant showed that most of the nodules presented a low number of bacteroids. No differences in the root infection abilities of green fluorescent protein–tagged clpB mutant and wild-type strains were detected. To our knowledge, this is the first study that presents evidence of the involvement of the chaperone ClpB from rhizobia in the symbiotic nodulation process.

scholarly journals 4-Chlorobenzoate Uptake in Comamonas sp. Strain DJ-12 Is Mediated by a Tripartite ATP-Independent Periplasmic Transporter

2006 ◽  
Vol 188 (24) ◽  
pp. 8407-8412 ◽  
Author(s):  
Jong-Chan Chae ◽  
Gerben J. Zylstra
Keyword(s):  
Growth Rate ◽  
Gene Cluster ◽  
Type Strain ◽  
Wild Type Strain ◽  
Proton Motive Force ◽  
Significant Inhibition ◽  
Motive Force ◽  
Wild Type ◽  
Knockout Mutant

ABSTRACT The fcb gene cluster involved in the hydrolytic dehalogenation of 4-chlorobenzoate is organized in the order fcbB-fcbA-fcbT1-fcbT2-fcbT3-fcbC in Comamonas sp. strain DJ-12. The genes are operonic and inducible with 4-chloro-, 4-iodo-, and 4-bromobenzoate. The fcbT1, fcbT2, and fcbT3 genes encode a transporter in the secondary TRAP (tripartite ATP-independent periplasmic) family. An fcbT1T2T3 knockout mutant shows a much slower growth rate on 4-chlorobenzoate compared to the wild type. 4-Chlorobenzoate is transported into the wild-type strain five times faster than into the fcbT1T2T3 knockout mutant. Transport of 4-chlorobenzoate shows significant inhibition by 4-bromo-, 4-iodo-, and 4-fluorobenzoate and mild inhibition by 3-chlorobenzoate, 2-chlorobenzoate, 4-hydroxybenzoate, 3-hydroxybenzoate, and benzoate. Uptake of 4-chlorobenzoate is significantly inhibited by ionophores which collapse the proton motive force.

The groEL2 gene, but not groEL1, is required for biosynthesis of the secondary metabolite myxovirescin in Myxococcus xanthus DK1622

Microbiology ◽  
2014 ◽  
Vol 160 (3) ◽  
pp. 488-495 ◽  
Author(s):  
Yan Wang ◽  
Xi Li ◽  
Wenyan Zhang ◽  
Xiuwen Zhou ◽  
Yue-zhong Li
Keyword(s):  
Secondary Metabolite ◽  
Mutant Strain ◽  
Type Strain ◽  
Wild Type Strain ◽  
Myxococcus Xanthus ◽  
Domain Swapping ◽  
Wild Type ◽  
Zone Of Inhibition ◽  
Knockout Mutant ◽  
E Coli

Myxococcus xanthus DK1622 possesses two copies of the groEL gene: groEL1, which participates in development, and groEL2, which is involved in the predatory ability of cells. In this study, we determined that the groEL2 gene is required for the biosynthesis of the secondary metabolite myxovirescin (TA), which plays essential roles in predation. The groEL2-knockout mutant strain was defective in producing a zone of inhibition and displayed decreased killing ability against Escherichia coli, while the groEL1-knockout mutant strain exhibited little difference from the wild-type strain DK1622. HPLC revealed that deletion of the groEL2 gene blocked the production of TA, which was present in the groEL1-knockout mutant. The addition of exogenous TA rescued the inhibition and killing abilities of the groEL2-knockout mutant against E. coli. Analysis of GroEL domain-swapping mutants indicated that the C-terminal equatorial domain of GroEL2 was essential for TA production, while the N-terminal equatorial or apical domains of GroEL2 were not sufficient to rescue TA production of the groEL2 knockout.

scholarly journals Virulence Traits of a Serogroup C Meningococcus and IsogeniccssAMutant, Defective in Surface-Exposed Sialic Acid, in a Murine Model of Meningitis

2019 ◽  
Vol 87 (4) ◽  
Author(s):  
Roberta Colicchio ◽  
Chiara Pagliuca ◽  
Susanna Ricci ◽  
Elena Scaglione ◽  
Denis Grandgirard ◽  
...  
Keyword(s):  
Sialic Acid ◽  
Corpus Callosum ◽  
Type Strain ◽  
Wild Type Strain ◽  
Lethal Dose ◽  
Sialic Acids ◽  
Wild Type ◽  
Knockout Mutant ◽  
Content Type

ABSTRACTIn serogroup CNeisseria meningitidis, thecssA(siaA) gene codes for an UDP-N-acetylglucosamine 2-epimerase that catalyzes the conversion of UDP-N-acetyl-α-d-glucosamine intoN-acetyl-d-mannosamine and UDP in the first step in sialic acid biosynthesis. This enzyme is required for the biosynthesis of the (α2→9)-linked polysialic acid capsule and for lipooligosaccharide (LOS) sialylation. In this study, we have used a reference serogroup C meningococcal strain and an isogeniccssAknockout mutant to investigate the pathogenetic role of surface-exposed sialic acids in a model of meningitis based on intracisternal inoculation of BALB/c mice. Results confirmed the key role of surface-exposed sialic acids in meningococcal pathogenesis. The 50% lethal dose (LD50) of the wild-type strain 93/4286 was about four orders of magnitude lower than that of thecssAmutant. Compared to the wild-type strain, the ability of this mutant to replicate in brain and spread systemically was severely impaired. Evaluation of brain damage evidenced a significant reduction in cerebral hemorrhages in mice infected with the mutant in comparison with the levels in those challenged with the wild-type strain. Histological analysis showed the typical features of bacterial meningitis, including inflammatory cells in the subarachnoid, perivascular, and ventricular spaces especially in animals infected with the wild type. Noticeably, 80% of mice infected with the wild-type strain presented with massive bacterial localization and accompanying inflammatory infiltrate in thecorpus callosum, indicating high tropism of meningococci exposing sialic acids toward this brain structure and a specific involvement of thecorpus callosumin the mouse model of meningococcal meningitis.

scholarly journals Molecular Monitoring of Wild-Type and Genetically Engineered Colletotrichum coccodes Biocontrol Strains In Planta

Plant Disease ◽  
2006 ◽  
Vol 90 (12) ◽  
pp. 1504-1510 ◽  
Author(s):  
A. L. Dauch ◽  
B. Ahn ◽  
A. K. Watson ◽  
P. Seguin ◽  
S. H. Jabaji-Hare
Keyword(s):  
Type Strain ◽  
Wild Type Strain ◽  
Quantitative Polymerase Chain Reaction ◽  
Fungal Growth ◽  
Genetically Engineered ◽  
Colletotrichum Coccodes ◽  
Wild Type ◽  
In Planta ◽  
Molecular Monitoring ◽  
Dna Amounts

Two strains of Colletotrichum coccodes, the wild type (DAOM 183088) and T-20a, engineered with the necrosis- and ethylene-inducing peptide (NEP1) gene for hypervirulence on velvetleaf (Abutilon theophrasti, Medik.), were monitored in planta for the first 2 weeks after infection. Real-time quantitative polymerase chain reaction (QPCR) was used to assess the extent of colonization of both strains on velvetleaf using SYBR Green chemistry. Quantification of both strains was successful as soon as the conidia were sprayed on the leaves and up to 14 days after infection. The increase in fungal DNA amounts corroborated with the appearance of necrotic lesions on velvetleaf leaves infected with the wild-type strain. The wild-type C. coccodes was more efficient at infecting velvetleaf than the transgenic T-20a strain. In addition, detection of host DNA allowed us to quantitatively monitor the decrease in plant DNA amounts in response to wild-type strain infection. Expression of the NEP1 transgene by conventional retro-transcription (RT)-PCR was absent from T-20a growing on either V8 agar or in planta, suggesting that the gene may be silenced. The application of QPCR to monitor fungal growth was proven to detect the target organisms in planta prior to the appearance of symptoms.

scholarly journals Who Needs Neighbors? PKS8 Is a Stand-Alone Gene in Fusarium graminearum Responsible for Production of Gibepyrones and Prolipyrone B

Molecules ◽  
2018 ◽  
Vol 23 (9) ◽  
pp. 2232 ◽  
Author(s):  
Klaus Westphal ◽  
Asmus Muurmann ◽  
Iben Paulsen ◽  
Kim Nørgaard ◽  
Marie Overgaard ◽  
...  
Keyword(s):  
Fusarium Graminearum ◽  
Type Strain ◽  
Wild Type Strain ◽  
Cytochrome P450 Monooxygenases ◽  
Wild Type ◽  
Great Capacity ◽  
Genus Fusarium ◽  
P450 Monooxygenases ◽  
Genomic Analyses ◽  
In The Wild

Genome sequencing of the genus Fusarium has revealed a great capacity for discovery of new natural products of potential economical and therapeutic importance. Several of these are unknown. In this study, we investigated the product of the PKS8 gene in Fusarium graminearum, which was recently linked to gibepyrones in F. fujikuroi. Genomic analyses showed that PKS8 constitutes a stand-alone gene in F. graminearum and related species. Overexpression of PKS8 resulted in production of gibepyrones A, B, D, G and prolipyrone B, which could not be detected in the wild type strain. Our results suggest that PKS8 produces the entry compound gibepyrone A, which is subsequently oxidized by one or several non-clustering cytochrome P450 monooxygenases ending with prolipyrone B.

scholarly journals F240 of β2-Tubulin Explains why Fusarium graminearum is Less Sensitive to Carbendazim than Botrytis cinerea

2018 ◽  
Vol 108 (3) ◽  
pp. 352-361 ◽  
Author(s):  
Yuanye Zhu ◽  
Xiaoyu Liang ◽  
Yanjun Li ◽  
Yabing Duan ◽  
Zhitian Zheng ◽  
...  
Keyword(s):  
Botrytis Cinerea ◽  
Binding Affinity ◽  
Fusarium Graminearum ◽  
Type Strain ◽  
Wild Type Strain ◽  
Fluorescent Protein ◽  
Amino Acid Sequences ◽  
Wild Type ◽  
Benzimidazole Fungicides ◽  
Β Tubulin

β-Tubulin is the target of benzimidazole fungicides, the most widely used of which is carbendazim (methyl benzimidazol-2-ylcarbamate [MBC]). MBC sensitivity is determined by the differential affinity of MBC for β-tubulins. However, the mechanism of less sensitivity of Fusarium graminearum to MBC compared with other fungi, including Botrytis cinerea, Colletotrichum gloeosporioides, and Sclerotinia sclerotiorum, remains exclusive. Alignment of β-tubulin amino acid sequences showed that position 240 of β-tubulins is leucine (L) in most pathogenic fungi but is phenylalanine (F) in the Fgβ2-tubulin of the F. graminearum wild type. The effective concentration resulting in 50% inhibition (EC50) value of MBC against the Fgβ2F240L mutant of F. graminearum is 0.047 μg/ml, which was 10-fold lower than that of wild-type strain 2021. Moreover, The EC50 value of MBC against the BcβL“240”F (actually position 232) mutant of Botrytis cinerea was 0.44 μg/ml, which was ninefold higher than that of B. cinerea wild-type strain Bt4-1. In response to MBC treatment (0.15 μg/ml), microtubules were clearly visible in Fgβ2-enhanced green fluorescent protein (EGFP) but not in Fgβ2F240L-EGFP. Moreover, a molecular docking assay indicated that F240L mutation created a pi-pi interaction between Fgβ2-tubulin and MBC and increased the binding affinity of Fgβ2-tubulin to MBC. Our results suggest that F240 is responsible for the naturally less MBC sensitivity in F. graminearum compared with B. cinerea, C. gloeosporioides, and S. sclerotiorum by decreasing the binding affinity between Fgβ2-tubulin and MBC.

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