scholarly journals DNA Methylation on N6-Adenine Regulates the Hyphal Development during Dimorphism in the Early-Diverging Fungus Mucor lusitanicus

2021 ◽  
Vol 7 (9) ◽  
pp. 738
Author(s):  
Macario Osorio-Concepción ◽  
Carlos Lax ◽  
Eusebio Navarro ◽  
Francisco E. Nicolás ◽  
Victoriano Garre
Keyword(s):  
Wild Type Strain ◽  
Pathogenic Fungi ◽  
Infection Process ◽  
Deletion Mutants ◽  
Polar Growth ◽  
Wild Type ◽  
C Elegans ◽  
Double Deletion ◽  
Multiple Processes ◽  
Causative Agents

The epigenetic modifications control the pathogenicity of human pathogenic fungi, which have been poorly studied in Mucorales, causative agents of mucormycosis. This order belongs to a group referred to as early-diverging fungi that are characterized by high levels of N6-methyldeoxy adenine (6mA) in their genome with dense 6mA clusters associated with actively expressed genes. AlkB enzymes can act as demethylases of 6mA in DNA, with the most remarkable eukaryotic examples being mammalian ALKBH1 and Caenorhabditis elegans NMAD-1. The Mucor lusitanicus (formerly M. circinelloides f. lusitanicus) genome contains one gene, dmt1, and two genes, dmt2 and dmt3, encoding proteins similar to C. elegans NMAD-1 and ALKBH1, respectively. The function of these three genes was analyzed by the generation of single and double deletion mutants for each gene. Multiple processes were studied in the mutants, but defects were only found in single and double deletion mutants for dmt1. In contrast to the wild-type strain, dmt1 mutants showed an increase in 6mA levels during the dimorphic transition, suggesting that 6mA is associated with dimorphism in M. lusitanicus. Furthermore, the spores of dmt1 mutants challenged with macrophages underwent a reduction in polar growth, suggesting that 6mA also has a role during the spore–macrophage interaction that could be important in the infection process.

scholarly journals DNA Methylation on N6-Adenine Regulates the Hyphal Development During Dimorphism in the Early-Diverging Fungus Mucor lusitanicus

2021 ◽  
Author(s):  
Macario Osorio-Concepción ◽  
Carlos Lax ◽  
Eusebio Navarro ◽  
Francisco E. Nicolás ◽  
Victoriano Garre
Keyword(s):  
Wild Type Strain ◽  
Pathogenic Fungi ◽  
Infection Process ◽  
Deletion Mutants ◽  
Polar Growth ◽  
Wild Type ◽  
C Elegans ◽  
Double Deletion ◽  
Multiple Processes ◽  
Causative Agents

The epigenetic modifications control the pathogenicity of human pathogenic fungi, which have been poorly studied in Mucorales; causative agents of mucormycosis. This order belongs to a group referred to as early-diverging fungi that are characterized by high levels of N6-methyldeoxyadenine (6mA) in their genome with dense 6mA clusters associated with actively expressed genes. AlkB enzymes can act as demethylases of 6mA in DNA, with the most remarkable eukaryotic examples being mammalian ALKBH1 and Caenorhabditis elegans NMAD-1. Mucor lusitanicus (formerly M. circinelloides f. lusitanicus) genome contains one gene, dmt1, and two genes, dmt2 and dmt3, encoding proteins homologs to C. elegans NMAD-1 and ALKBH1, respectively. The function of the three genes was analyzed by the generation of single and double deletion mutants for each gene. Multiple processes were studied in the mutants, but defects were only found in single and double deletion mutants for dmt1. In contrast to the wild-type strain, dmt1 mutants showed an increase of 6mA levels during the dimorphic transition, suggesting that 6mA regulates dimorphism in M. lusitanicus. Furthermore, the spores of dmt1 mutants challenged with macrophages underwent a reduction of polar growth, suggesting that 6mA also has a role during the spore-macrophage interaction that could be important in the infection process.

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 The three NADH dehydrogenases of Pseudomonas aeruginosa: Their roles in energy metabolism and links to virulence

PLoS ONE ◽  
2021 ◽  
Vol 16 (2) ◽  
pp. e0244142
Author(s):  
Teri N. Hreha ◽  
Sara Foreman ◽  
Ana Duran-Pinedo ◽  
Andrew R. Morris ◽  
Patricia Diaz-Rodriguez ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Opportunistic Pathogen ◽  
Lag Phase ◽  
Deletion Mutants ◽  
Wild Type ◽  
Nadh:Quinone Oxidoreductase ◽  
Double Deletion ◽  
Single Deletion ◽  
Nadh Dehydrogenases ◽  
Pyocyanin Production

Pseudomonas aeruginosa is a ubiquitous opportunistic pathogen which relies on a highly adaptable metabolism to achieve broad pathogenesis. In one example of this flexibility, to catalyze the NADH:quinone oxidoreductase step of the respiratory chain, P. aeruginosa has three different enzymes: NUO, NQR and NDH2, all of which carry out the same redox function but have different energy conservation and ion transport properties. In order to better understand the roles of these enzymes, we constructed two series of mutants: (i) three single deletion mutants, each of which lacks one NADH dehydrogenase and (ii) three double deletion mutants, each of which retains only one of the three enzymes. All of the mutants grew approximately as well as wild type, when tested in rich and minimal medium and in a range of pH and [Na+] conditions, except that the strain with only NUO (ΔnqrFΔndh) has an extended lag phase. During exponential phase, the NADH dehydrogenases contribute to total wild-type activity in the following order: NQR > NDH2 > NUO. Some mutants, including the strain without NQR (ΔnqrF) had increased biofilm formation, pyocyanin production, and killed more efficiently in both macrophage and mouse infection models. Consistent with this, ΔnqrF showed increased transcription of genes involved in pyocyanin production.

scholarly journals Role of the Type VI Secretion System in the Pathogenicity of Pseudomonas syringae pv. actinidiae, the Causative Agent of Kiwifruit Bacterial Canker

2021 ◽  
Vol 12 ◽  
Author(s):  
Nana Wang ◽  
Ning Han ◽  
Runze Tian ◽  
Jiliang Chen ◽  
Xiaoning Gao ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Pseudomonas Syringae ◽  
Biofilm Formation ◽  
Wild Type Strain ◽  
Deletion Mutants ◽  
Wild Type ◽  
Type Vi Secretion System ◽  
Bacterial Competition ◽  
Type Vi Secretion

The type VI secretion system (T6SS), a macromolecular machine, plays an important role in the pathogenicity of many Gram-negative bacteria. However, the role of T6SS in the pathogenicity of Pseudomonas syringae pv. actinidiae (Psa), the pathogen of kiwifruit bacterial canker, is yet to be studied. Here, we found a T6SS gene cluster consisting of 13 core genes (A-J) in the genome of Psa M228 based on a genome-wide analysis. To determine whether the T6SS gene cluster affects the pathogenicity of Psa M228, T6SS and its 13 core gene deletion mutants were constructed and their pathogenicity was determined. The deletion mutants showed different degrees of reduction in pathogenicity compared with the wild-type strain M228; in tssM and tssJ mutants, pathogenicity was significantly reduced by 78.7 and 71.3%, respectively. The pathogenicity results were also confirmed by electron microscopy. To further confirm that the reduction in pathogenicity is related to the function of T6SS, we selected the T6SS gene cluster, comprising tssM and tssJ, for further analyses. Western blot results revealed that tssM and tssJ were necessary for hemolytic co-regulatory protein secretion, indicating that they encode a functional T6SS. Further, we explored the mechanism by which T6SS affects the pathogenicity of Psa M228. The ability of bacterial competition, biofilm formation, hydrogen peroxide tolerance, and proteolytic activity were all weakened in the deletion mutants M228ΔT6SS, M228ΔtssM, and M228ΔtssJ. All these properties of the two gene complementation mutants were restored to the same levels as those of the wild-type strain, M228. Quantitative real-time results showed that during the interaction between the deletion mutant M228ΔT6SS and the host, expression levels of T3SS transcriptional regulatory gene hrpR, structural genes hrpZ, hrcC, hopP1, and effector genes hopH1 and hopM1 were down-regulated at different levels. Taken together, our data provide evidence for the first time that the T6SS plays an important role in the pathogenicity of Psa, probably via effects on bacterial competition, biofilm formation, and environmental adaptability. Moreover, a complicated relationship exists between T6SS and T3SS.

scholarly journals Cutinase A of Botrytis cinerea is Expressed, but not Essential, During Penetration of Gerbera and Tomato

1997 ◽  
Vol 10 (1) ◽  
pp. 30-38 ◽  
Author(s):  
J. A. L. van Kan ◽  
J. W. van 't Klooster ◽  
C. A. M. Wagemakers ◽  
D. C. T. Dees ◽  
C. J. B. van der Vlugt-Bergmans
Keyword(s):  
Botrytis Cinerea ◽  
Wild Type Strain ◽  
Gus Expression ◽  
Infection Process ◽  
Gus Activity ◽  
Wild Type ◽  
Tomato Fruits ◽  
Fusion Construct ◽  
Gus Reporter ◽  
Encoding Gene

The plant pathogen Botrytis cinerea can infect undamaged plant tissue directly by penetration of the cuticle. This penetration has been suggested to be enzyme-mediated, and an important role for cutinase in the infection process has been proposed. In this study the expression of the cutinase encoding gene cutA of B. cinerea was analyzed using a cutA promoter-GUS reporter gene fusion. Transformants containing the fusion construct were examined for GUS expression on gerbera flowers and tomato fruits. High GUS activity was detected from the onset of conidial germination and during penetration into epidermal cells, indicating that cutA is expressed during the early stages of infection. To determine the biological relevance of cutinase A for successful penetration, cutinase A-deficient mutants were constructed by gene disruption. Pathogenicity of two transformants lacking a functional cutA gene was studied on gerbera flowers and tomato fruits. Their ability to penetrate and cause symptoms was unaltered compared to the wild-type strain. These results exclude an important role for cutinase A during direct penetration of host tissue by B. cinerea.

scholarly journals Differential Contribution of Plant-Beneficial Functions from Pseudomonas kilonensis F113 to Root System Architecture Alterations in Arabidopsis thaliana and Zea mays

2018 ◽  
Vol 31 (2) ◽  
pp. 212-223 ◽  
Author(s):  
Jordan Vacheron ◽  
Guilhem Desbrosses ◽  
Sébastien Renoud ◽  
Rosa Padilla ◽  
Vincent Walker ◽  
...  
Keyword(s):  
Arabidopsis Thaliana ◽  
Type Strain ◽  
Wild Type Strain ◽  
Phosphate Solubilization ◽  
Glucose Dehydrogenase ◽  
Root System Architecture ◽  
Deletion Mutants ◽  
Wild Type ◽  
Relative Contribution

Fluorescent pseudomonads are playing key roles in plant-bacteria symbiotic interactions due to the multiple plant-beneficial functions (PBFs) they are harboring. The relative contributions of PBFs to plant-stimulatory effects of the well-known plant growth-promoting rhizobacteria Pseudomonas kilonensis F113 (formerly P. fluorescens F113) were investigated using a genetic approach. To this end, several deletion mutants were constructed, simple mutants ΔphlD (impaired in the biosynthesis of 2,4-diacetylphloroglucinol [DAPG]), ΔacdS (deficient in 1-aminocyclopropane-1-carboxylate deaminase activity), Δgcd (glucose dehydrogenase deficient, impaired in phosphate solubilization), and ΔnirS (nitrite reductase deficient), and a quadruple mutant (deficient in the four PBFs mentioned above). Every PBF activity was quantified in the wild-type strain and the five deletion mutants. This approach revealed few functional interactions between PBFs in vitro. In particular, biosynthesis of glucose dehydrogenase severely reduced the production of DAPG. Contrariwise, the DAPG production impacted positively, but to a lesser extent, phosphate solubilization. Inoculation of the F113 wild-type strain on Arabidopsis thaliana Col-0 and maize seedlings modified the root architecture of both plants. Mutant strain inoculations revealed that the relative contribution of each PBF differed according to the measured plant traits and that F113 plant-stimulatory effects did not correspond to the sum of each PBF relative contribution. Indeed, two PBF genes (ΔacdS and ΔnirS) had a significant impact on root-system architecture from both model plants, in in vitro and in vivo conditions. The current work underscored that few F113 PBFs seem to interact between each other in the free-living bacterial cells, whereas they control in concert Arabidopsis thaliana and maize growth and development.

scholarly journals Predatory behaviour of trapping fungi against srf mutants of Caenorhabditis elegans and different plant and animal parasitic nematodes

Parasitology ◽  
1999 ◽  
Vol 119 (1) ◽  
pp. 95-104 ◽  
Author(s):  
P. MENDOZA DE GIVES ◽  
K. G. DAVIES ◽  
S. J. CLARK ◽  
J. M. BEHNKE
Keyword(s):  
Caenorhabditis Elegans ◽  
Infection Process ◽  
Parasitic Nematodes ◽  
Plant Parasitic Nematodes ◽  
Wild Type ◽  
Duddingtonia Flagrans ◽  
Predatory Behaviour ◽  
C Elegans ◽  
Nematode Cuticle ◽  
Predatory Response

The initial infection process of nematode-trapping fungi is based on an interaction between the trapping structure of the fungus and the surface of the nematode cuticle. A bioassay was designed to investigate the predatory response of several isolates of nematode-trapping fungi against 3 mutants of Caenorhabditis elegans (AT6, AT10 and CL261), which have been reported to differ in the reaction of their cuticle to antibodies and lectins. The bioassay was also applied to infective larvae of animal (Haemonchus contortus, Teladorsagia (Ostertagia) circumcincta and Trichostrongylus axei) and plant (Meloidogyne spp.) parasitic nematodes. Differences in trapping ability were most marked in the first 24 h, and were density dependent. Although the isolate of Arthrobotrys responded very rapidly in the first 24 h, Duddingtonia flagrans was generally the most effective isolate and Monacrosporium responded relatively poorly throughout all experiments. All the fungi tested trapped the srf mutants of C. elegans more efficiently than the wild type, and there were differences between the different srf mutants of C. elegans. Differences in trapping ability were also observed between different isolates of D. flagrans; similarly, differences in trapping behaviour were observed not only amongst the different species of plant-parasitic nematodes, but also between the sheathed and exsheathed larvae of the animal-parasitic nematodes.

scholarly journals Exopolyphosphatases PPX1 and PPX2 from Corynebacterium glutamicum

2009 ◽  
Vol 75 (10) ◽  
pp. 3161-3170 ◽  
Author(s):  
Steffen N. Lindner ◽  
Sandra Knebel ◽  
Hendrik Wesseling ◽  
Siegfried M. Schoberth ◽  
Volker F. Wendisch
Keyword(s):  
Corynebacterium Glutamicum ◽  
Type Strain ◽  
Wild Type Strain ◽  
Deletion Mutants ◽  
Gene Products ◽  
Wild Type ◽  
Inorganic Polyphosphate ◽  
Deletion Mutations ◽  
In The Wild ◽  
Growth Experiments

ABSTRACT Corynebacterium glutamicum accumulates up to 300 mM of inorganic polyphosphate (PolyP) in the cytosol or in granules. The gene products of cg0488 (ppx1) and cg1115 (ppx2) were shown to be active as exopolyphosphatases (PPX), as overexpression of either gene resulted in higher exopolyphosphatase activities in crude extracts and deletion of either gene with lower activities than those of the wild-type strain. PPX1 and PPX2 from C. glutamicum share only 25% identical amino acids and belong to different protein groups, which are distinct from enterobacterial, archaeal, and yeast exopolyphosphatases. In comparison to that in the wild type, more intracellular PolyP accumulated in the Δppx1 and Δppx2 deletion mutations but less when either ppx1 or ppx2 was overexpressed. When C. glutamicum was shifted from phosphate-rich to phosphate-limiting conditions, a growth advantage of the deletion mutants and a growth disadvantage of the overexpression strains compared to the wild type were observed. Growth experiments, exopolyphosphatase activities, and intracellular PolyP concentrations revealed PPX2 as being a major exopolyphosphatase from C. glutamicum. PPX2His was purified to homogeneity and shown to be active as a monomer. The enzyme required Mg2+ or Mn2+ cations but was inhibited by millimolar concentrations of Mg2+, Mn2+, and Ca2+. PPX2 from C. glutamicum was active with short-chain polyphosphates, even accepting pyrophosphate, and was inhibited by nucleoside triphosphates.

Isogenic P-fimbrial deletion mutants of pyelonephritogenic Escherichia coli: the role of ? Gal(1?4)? Gal binding in virulence of a wild-type strain

1993 ◽  
Vol 10 (1) ◽  
pp. 143-155 ◽  
Author(s):  
Harry L.T. Mobley ◽  
Karen G. Jarvis ◽  
John P. Elwood ◽  
Donna I. Whittle ◽  
C. Virginia Lockatell ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Type Strain ◽  
Wild Type Strain ◽  
Deletion Mutants ◽  
Wild Type

scholarly journals Mycosubtilin Overproduction by Bacillus subtilis BBG100 Enhances the Organism's Antagonistic and Biocontrol Activities

2005 ◽  
Vol 71 (8) ◽  
pp. 4577-4584 ◽  
Author(s):  
Valérie Leclère ◽  
Max Béchet ◽  
Akram Adam ◽  
Jean-Sébastien Guez ◽  
Bernard Wathelet ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Type Strain ◽  
Wild Type Strain ◽  
Germination Rate ◽  
Pathogenic Fungi ◽  
Strain Atcc ◽  
Wild Type ◽  
Native Promoter ◽  
Replication Gene ◽  
Biocontrol Potential

ABSTRACT A Bacillus subtilis derivative was obtained from strain ATCC 6633 by replacement of the native promoter of the mycosubtilin operon by a constitutive promoter originating from the replication gene repU of the Staphylococcus aureus plasmid pUB110. The recombinant strain, designated BBG100, produced up to 15-fold more mycosubtilin than the wild type produced. The overproducing phenotype was related to enhancement of the antagonistic activities against several yeasts and pathogenic fungi. Hemolytic activities were also clearly increased in the modified strain. Mass spectrometry analyses of enriched mycosubtilin extracts showed similar patterns of lipopeptides for BBG100 and the wild type. Interestingly, these analyses also revealed a new form of mycosubtilin which was more easily detected in the BBG100 sample. When tested for its biocontrol potential, wild-type strain ATCC 6633 was almost ineffective for reducing a Pythium infection of tomato seedlings. However, treatment of seeds with the BBG100 overproducing strain resulted in a marked increase in the germination rate of seeds. This protective effect afforded by mycosubtilin overproduction was also visualized by the significantly greater fresh weight of emerging seedlings treated with BBG100 compared to controls or seedlings inoculated with the wild-type strain.

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