ATP-binding cassette (ABC) transporters mediate active efflux of natural and synthetic toxicants and are considered to be important for drug tolerance in microorganisms. In biological control agents ...(BCA), ABC transporters can play important roles in antagonism by providing protection against toxins derived from the fungal prey and by mediating the secretion of endogenous toxins. In the present study, we generated deletion and complementation strains of the ABC transporter abcG5 in the fungal BCA Clonostachys rosea to study its role in xenobiotic tolerance and antagonism. Gene expression analysis shows induced expression of abcG5 in the presence of the Fusarium mycotoxin zearalenone (ZEA), secreted metabolites of F. graminearum, and different classes of fungicides. Phenotypic analysis of abcG5 deletion and complementation strains showed that the deletion strains were more sensitive towards F. graminearum culture filtrates, ZEA, and iprodione- and mefenoxam-based fungicides, thus suggesting the involvement of abcG5 in cell protection. The ΔabcG5 strains displayed reduced antagonism towards F. graminearum in a plate confrontation assay. Furthermore, the ΔabcG5 strains failed to protect barley seedlings from F. graminearium foot rot disease. These data show that the abcG5 ABC transporter is important for xenobiotic tolerance and biocontrol traits in C. rosea.
Aegerolysins are small secreted pore-forming proteins that are found in both prokaryotes and eukaryotes. The role of aegerolysins in sporulation, fruit body formation, and in lysis of cellular ...membrane is suggested in fungi. The aim of the present study was to characterize the biological function of the aegerolysin gene
agl1
in the mycoparasitic fungus
Trichoderma atroviride
, used for biological control of plant diseases. Gene expression analysis showed higher expression of
agl1
during conidiation and during growth in medium supplemented with cell wall material from the plant pathogenic fungus
Rhizoctonia solani
as the sole carbon source. Expression of
agl1
was supressed under iron-limiting condition, while
agl1
transcript was not detected during
T
.
atroviride
interactions with the prey fungi
Botrytis cinerea
or
R
.
solani
. Phenotypic analysis of
agl1
deletion strains (Δ
agl1
) showed reduced conidiation compared to
T
.
atroviride
wild type, thus suggesting the involvement of AGL1 in conidiation. Furthermore, the Δ
agl1
strains display reduced antagonism towards
B. cinerea
and
R
.
solani
based on a secretion assay, although no difference was detected during direct interactions. These data demonstrate the role of AGL1 in conidiation and antagonism in the mycoparasitic fungus
T
.
atroviride
.
Filamentous fungi produce small cysteine rich surface active amphiphilic hydrophobins on the outer surface of cell walls that mediate interactions between the fungus and the environment. The role of ...hydrophobins in surface hydrophobicity, sporulation, fruit body formation, recognition and adhesion to host surface and virulence have been reported. The aim of the present study was to characterize the biological function of hydrophobins in the fungal biocontrol agent Clonostachys rosea in order to understand their potential roles in biocontrol mechanisms.
Based on the presence of hydrophobin domains, cysteine spacing patterns and hydropathy plots, we identified three class II hydrophobin genes in C. rosea. Gene expression analysis showed basal expression of Hyd1, Hyd2 and Hyd3 in all conditions tested with the exception of induced Hyd1 expression in conidiating mycelium. Interestingly, up-regulation of Hyd1, Hyd2 and Hyd3 was found during C. rosea self interaction compared to interactions with the fungal plant pathogens Botrytis cinerea or Fusarium graminearum in dual culture assays. Phenotypic analysis of C. rosea deletion and complementation strains showed that Hyd1 and Hyd3 are jointly required for conidial hydrophobicity, although no difference in mycelia hydrophobicity was found between wild type (WT) and mutant strains. Interestingly, mutant strains showed increased growth rates, conidiation and enhanced tolerances of conidia to abiotic stresses. Antagonism tests using in vitro dual culture and detached leaf assays showed that the mutant strains were more aggressive towards B. cinerea, F. graminearum or Rhizoctonia solani, and that aggression was partly related to earlier conidial germination and enhanced tolerance of mutant strains to secreted fungal metabolites. Furthermore, in vitro Arabidopsis thaliana root colonization assays revealed reduced root colonization ability of the ΔHyd3 strain, but not for the ΔHyd1 strain. Furthermore, enhanced root colonization ability for the ΔHyd1ΔHyd3 strain was found in comparison to WT.
These results show a role for hydrophobins in conidial hydrophobicity, control of conidial germination under stress conditions, and in root colonization in C. rosea. However, functional studies of Hyd2 remains to be performed in order to fully assess the role of hydrophobins in C. rosea.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Polytetrafluoroethylene (PTFE) is known as a high performance engineering polymer as well as a solid lubricant. The literature contains several papers that report on the effects of nano-sized solid ...lubricant additives, but no systematic studies have been published on the effects of nano-sized PTFE particles. This paper first describes the methodology for preparing nano- and micro-PTFE particles that were blended into a 150 N API Group II base oil. The particle sizes were 50nm, 150nm, 400nm and 12µm, and the particle concentrations were 4, 8 and 12%. Formulations were characterized for physical properties and tribological behavior. Physical properties included density, viscosity, pour point, and flash point. Tribological characteristics were (1) weld load tests (a four-ball EP method to determine load carrying capacity), (2) anti-wear properties (using a different four-ball test method) and (3) friction (using an Optimol-SRV III oscillating friction and wear tester). Results from tribological testing of the experimental oils and bearing steel test specimens showed that PTFE particles can significantly improve the weld load, as well as anti-wear and friction reduction properties. The smaller the size of PTFE particles and the higher their concentration, the greater was the performance improvement. The topography and surface chemistry of the test specimens were examined using scanning electron microscopy, and the improved quality of the tribo-films formed on the counter faces was seen to correlate with performance improvements.
•Nano-PTFE particles were used first time as additive for developing nano-lubricant.•Oils were evaluated to understand their influence on friction, wear and EP performance.•Higher the amount and lower the size, higher was the performance; nano-size being the best.•Different qualities of transferred films on the surface were responsible for the variation in performance.
Methylisocitrate lyase (MCL), a signature enzyme of the methylcitrate cycle, which cleaves methylisocitrate to pyruvate and succinate, is required for propionate metabolism, for secondary metabolite ...production and for virulence in bacteria and fungi. Here we investigate the role of the methylcitrate cycle by generating an mcl deletion mutant in the fungal biocontrol agent Trichoderma atroviride. Gene expression analysis shows that a basal expression of mcl is observed in all growth conditions tested. Phenotypic analysis of an mcl deletion mutant suggests the requirement of MCL in propionate resistance, growth, conidial pigmentation and germination, and abiotic stress tolerance. A plate confrontation assay did not show a difference between the WT and the Δmcl strain in antagonism towards Botrytis cinerea. However, the Δmcl strain displays reduced antagonism towards B. cinerea based on a secretion assay. Furthermore, an in vitro root colonization assay shows that the Δmcl strain had reduced ability to colonize Arabidopsis thaliana roots, which results in reduced induction of systemic resistance towards B. cinerea. These data show that MCL is important not only for growth and development in T. atroviride but also in antagonism, root colonization and induction of defence responses in plants.
Lysin motif (LysM) modules are approximately 50 amino acids long and bind to peptidoglycan, chitin and its derivatives. Certain LysM proteins in plant pathogenic and entomopathogenic fungi are shown ...to scavenge chitin oligosaccharides and thereby dampen host defense reactions. Other LysM proteins can protect the fungal cell wall against hydrolytic enzymes. In this study, we investigated the biological function of LysM proteins in the mycoparasitic fungus
. The
.
genome contained three genes coding for LysM-containing proteins and gene expression analysis revealed that
and
were induced during mycoparasitic interaction with
and during colonization of wheat roots.
was suppressed in germinating conidia, while
was induced during growth in chitin or peptidoglycan-containing medium. Deletion of
and
resulted in mutants with increased levels of conidiation and conidial germination, but reduced ability to control plant diseases caused by
.
and
. The Δ
strain showed a distinct, accelerated mycelial disintegration phenotype accompanied by reduced biomass production and hyphal protection against hydrolytic enzymes including chitinases, suggesting a role of LYSM2 in hyphal protection against chitinases. The Δ
and Δ
Δ
strains displayed reduced ability to colonize wheat roots, while only Δ
Δ
failed to suppress expression of the wheat defense response genes
and
. Based on our data, we propose a role of LYSM1 as a regulator of fungal development and of LYSM2 in cell wall protection against endogenous hydrolytic enzymes, while both are required to suppress plant defense responses. Our findings expand the understanding of the role of LysM proteins in fungal-fungal interactions and biocontrol.
Clonostachys rosea is a mycoparasitic fungus that can control several important plant diseases. Here, we report on the genome sequencing of C. rosea and a comparative genome analysis, in order to ...resolve the phylogenetic placement of C. rosea and to study the evolution of mycoparasitism as a fungal lifestyle. The genome of C. rosea is estimated to 58.3 Mb, and contains 14,268 predicted genes. A phylogenomic analysis shows that C. rosea clusters as sister taxon to plant pathogenic Fusarium species, with mycoparasitic/saprotrophic Trichoderma species in an ancestral position. A comparative analysis of gene family evolution reveals several distinct differences between the included mycoparasites. Clonostachys rosea contains significantly more ATP-binding cassette (ABC) transporters, polyketide synthases, cytochrome P450 monooxygenases, pectin lyases, glucose-methanol-choline oxidoreductases, and lytic polysaccharide monooxygenases compared with other fungi in the Hypocreales. Interestingly, the increase of ABC transporter gene number in C. rosea is associated with phylogenetic subgroups B (multidrug resistance proteins) and G (pleiotropic drug resistance transporters), whereas an increase in subgroup C (multidrug resistance-associated proteins) is evident in Trichoderma virens. In contrast with mycoparasitic Trichoderma species, C. rosea contains very few chitinases. Expression of six group B and group G ABC transporter genes was induced in C. rosea during exposure to the Fusarium mycotoxin zearalenone, the fungicide Boscalid or metabolites from the biocontrol bacterium Pseudomonas chlororaphis. The data suggest that tolerance toward secondary metabolites is a prominent feature in the biology of C. rosea.
Biological control
, or
biocontrol
, is the exploitation of living agents (incl. viruses) to combat pestilential organisms (incl. pathogens, pests, and weeds) for diverse purposes to provide human ...benefits. Thus, during the last century the practices and concepts involved have evolved in separate streams associated with distinct scientific and taxonomic disciplines. In parallel developments, there have been increasing references to biological control in industrial contexts and legislation, resulting in conceptual and terminological disintegration. The aim of this paper is to provide a global conceptual and terminological platform that facilitates future development of the field. We review use of previously suggested terms in key fields (e.g., phytopathology, entomology, and weed science), eliminate redundant terminology, identify three principles that should underpin the concept, and then present a new framework for biological control, rooted in seminal publications. The three principles establish that (1) only living agents can mediate biological control, (2) biological control always targets a pest, directly or indirectly, and (3) all biocontrol methods can be classified in four main categories depending on whether resident agents are utilized, with or without targeted human intervention (
conservation biological control
and
natural biological control
, respectively) or agents are added for permanent or temporary establishment (
classical biological control
and
augmentative biological control
, respectively). Correct identification of what is, and is not, biological control can help efforts to understand and optimize biological pest control for human and environmental benefits. The new conceptual framework may contribute to more uniform and appropriate regulatory approaches to biological control, and more efficient authorization and application of biocontrol products.
The recently identified phylogenetic subgroup B5 of fungal glycoside hydrolase family 18 genes encodes enzymes with mannosyl glycoprotein endo-N-acetyl-β-D-glucosaminidase (ENGase)-type activity. ...Intracellular ENGase activity is associated with the endoplasmic reticulum associated protein degradation pathway (ERAD) of misfolded glycoproteins, although the biological relevance in filamentous fungi is not known. Trichoderma atroviride is a mycoparasitic fungus that is used for biological control of plant pathogenic fungi. The present work is a functional study of the T. atroviride B5-group gene Eng18B, with emphasis on its role in fungal growth and antagonism. A homology model of T. atroviride Eng18B structure predicts a typical glycoside hydrolase family 18 (αβ)(8) barrel architecture. Gene expression analysis shows that Eng18B is induced in dual cultures with the fungal plant pathogens Botrytis cinerea and Rhizoctonia solani, although a basal expression is observed in all growth conditions tested. Eng18B disruption strains had significantly reduced growth rates but higher conidiation rates compared to the wild-type strain. However, growth rates on abiotic stress media were significantly higher in Eng18B disruption strains compared to the wild-type strain. No difference in spore germination, germ-tube morphology or in hyphal branching was detected. Disruption strains produced less biomass in liquid cultures than the wild-type strain when grown with chitin as the sole carbon source. In addition, we determined that Eng18B is required for the antagonistic ability of T. atroviride against the grey mould fungus B. cinerea in dual cultures and that this reduction in antagonistic ability is partly connected to a secreted factor. The phenotypes were recovered by re-introduction of an intact Eng18B gene fragment in mutant strains. A putative role of Eng18B ENGase activity in the endoplasmic reticulum associated protein degradation pathway of endogenous glycoproteins in T. atroviride is discussed in relation to the observed phenotypes.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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