Genetic and Phenotypic Characterization of Cryphonectria hypovirus 1 from Eurasian Georgia

Daniel Rigling; Nora Borst; Carolina Cornejo; Archil Supatashvili; Simone Prospero

doi:10.3390/v10120687

Genetic and Phenotypic Characterization of Cryphonectria hypovirus 1 from Eurasian Georgia

Viruses ◽

10.3390/v10120687 ◽

2018 ◽

Vol 10 (12) ◽

pp. 687 ◽

Cited By ~ 3

Author(s):

Daniel Rigling ◽

Nora Borst ◽

Carolina Cornejo ◽

Archil Supatashvili ◽

Simone Prospero

Keyword(s):

Biological Control ◽

Biological Control Agent ◽

Cryphonectria Parasitica ◽

Control Agent ◽

Phenotypic Characterization ◽

Tree Disease ◽

Caucasus Region ◽

Chestnut Blight Fungus ◽

Part Sequencing ◽

Cryphonectria Hypovirus

Cryphonectria hypovirus 1 (CHV-1) infects the chestnut blight fungus Cryphonectria parasitica and acts as a biological control agent against this harmful tree disease. In this study, we screened the recently characterized C. parasitica population in Eurasian Georgia for the presence of CHV-1. We found 62 CHV-1 infected C. parasitica isolates (9.3%) among a total of 664 isolates sampled in 14 locations across Georgia. The prevalence of CHV-1 at the different locations ranged from 0% in the eastern part of the country to 29% in the western part. Sequencing of two specific regions of the viral genome one each in ORFA and ORFB revealed a unique CHV-1 subtype in Georgia. This subtype has a recombinant pattern combining the ORFA region from the subtype F2 and the ORFB region from subtype D. All 62 viral strains belonged to this Georgian CHV-1 subtype (subtype G). The CHV-1 subtype G strongly reduced the parasitic growth of C. parasitica isolates from Georgia, with a more severe effect on the European genepool compared to the Georgian genepool. The CHV-1 subtype detected in Georgia provides a valuable candidate for biological control applications in the Caucasus region.

Download Full-text

Hypovirus Virulence and Vegetative Incompatibility in Populations of the Chestnut Blight Fungus

Phytopathology ◽

10.1094/phyto-01-12-0013-r ◽

2012 ◽

Vol 102 (12) ◽

pp. 1161-1167 ◽

Cited By ~ 18

Author(s):

Sarah Franziska Bryner ◽

Daniel Rigling

Keyword(s):

Biocontrol Agent ◽

Cryphonectria Parasitica ◽

Vegetative Compatibility ◽

Chestnut Blight ◽

Vegetative Incompatibility ◽

Tree Disease ◽

Chestnut Blight Fungus ◽

Fungal Populations ◽

Cryphonectria Hypovirus ◽

High Diversity

Cryphonectria hypovirus 1 hyperparasitizes the chestnut blight fungus Cryphonectria parasitica and acts as a biocontrol agent for this serious tree disease. The virus is transmitted cytoplasmatically between fungal individuals. However, highly virulent viruses strongly debilitate their host and, thus, reduce their own transmission probability. Furthermore, vegetative incompatibility between fungi is an important transmission barrier. Therefore, virulent viruses are expected to be strongly selected against in fungal populations with high levels of vegetative incompatibility, eventually leading to the erosion of biocontrol. To test this prediction, we assessed the virulence of the virus in four European C. parasitica populations with high diversity of vegetative compatibility types and in four populations with low diversity. We expected the degree of virus virulence to be lower in fungal populations with high levels of vegetative incompatibility. However, our results did not reveal such a trend. No significant differences in virus virulence between populations with low versus high diversity of vegetative compatibility types were observed. There was no evidence for an erosion of disease control due to the presence of these transmission barriers. Thus, the findings of this study are promising for the sustainability of Cryphonectria hypovirus 1 as a biocontrol agent for chestnut blight in Europe.

Download Full-text

Role of Fresh Dead Wood in the Epidemiology and the Biological Control of the Chestnut Blight Fungus

Plant Disease ◽

10.1094/pdis-05-18-0796-re ◽

2019 ◽

Vol 103 (3) ◽

pp. 430-438 ◽

Cited By ~ 2

Author(s):

Joana Beatrice Meyer ◽

Loïc Chalmandrier ◽

Fabio Fässler ◽

Christopher Schefer ◽

Daniel Rigling ◽

...

Keyword(s):

Biological Control ◽

Dead Wood ◽

Biocontrol Agent ◽

Castanea Sativa ◽

Cryphonectria Parasitica ◽

Chestnut Blight ◽

Chestnut Blight Fungus ◽

Study Sites ◽

Cryphonectria Hypovirus

The invasive fungus Cryphonectria parasitica, the causal agent of chestnut blight, is able to survive and sporulate on the bark of fresh dead Castanea sativa wood for at least 2 years. Here, we experimentally investigated the role of fresh dead wood in the epidemiology of chestnut blight, specifically in the spread of the hyperparasitic virus Cryphonectria hypovirus 1, which acts as biocontrol agent of C. parasitica. A total of 152 artificially initiated, virulent bark cankers in four chestnut stands were treated with virus-infected asexual spores originating either from sporulating dead wood or from a spore suspension. Molecular markers for both the virus and the fungal carrier were used to examine the spread of the applied biocontrol virus. Fourteen months after treatment, 42 to 76% of the conidial spray-treated cankers and 50 to 60% of the cankers exposed to a sporulating dead stem had been virus infected by the applied hypovirulent conidia in all four study sites. Virus infection reduced canker expansion and promoted canker healing (callusing). Thus, fresh chestnut dead wood may play an important role in supporting the successful spread of natural hypovirulence in chestnut forests. Further, combined with the application of virus-infected conidial suspensions, it may help promote the establishment of artificially released hypoviruses in chestnut stands to control chestnut blight.

Download Full-text

Generation of Trichoderma harzianum with pyr4 auxotrophic marker by using the CRISPR/Cas9 system

Scientific Reports ◽

10.1038/s41598-020-80186-4 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Amanda A. Vieira ◽

Giovanni R. Vianna ◽

Jessica Carrijo ◽

Francisco J. L. Aragão ◽

Pabline M. Vieira

Keyword(s):

Biological Control ◽

Trichoderma Harzianum ◽

Biological Control Agent ◽

Control Agent ◽

Phenotypic Characterization ◽

Agricultural Pests ◽

Control Activity ◽

Guide Rna ◽

Endogenous Expression ◽

Auxotrophic Marker

AbstractTrichoderma harzianum is a filamentous fungus used as a biological control agent for agricultural pests. Genes of this microorganism have been studied, and their applications are patented for use in biofungicides and plant breeding strategies. Gene editing technologies would be of great importance for genetic characterization of this species, but have not yet been reported. This work describes mutants obtained with an auxotrophic marker in this species using the CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)/ Cas (CRISPR-associated) system. For this, sequences for a guide RNA and Cas9 overexpression were inserted via biolistics, and the sequencing approach confirmed deletions and insertions at the pyr4 gene. Phenotypic characterization demonstrated a reduction in the growth of mutants in the absence of uridine, as well as resistance to 5-fluorotic acid. In addition, the gene disruption did not reduce mycoparasitc activity against phytopathogens. Thus, target disruption of the pyr4 gene in T. harzianum using the CRISPR/Cas9 system was demonstrated, and it was also shown that endogenous expression of the system did not interfere with the biological control activity of pathogens. This work is the first report of CRISPR Cas9-based editing in this biocontrol species, and the mutants expressing Cas9 have potential for the generation of useful technologies in agricultural biotechnology.

Download Full-text