Cytological and electrophoretic karyotyping of the chestnut blight fungus Cryphonectria parasitica

Eusebio-Cope, Ana; Suzuki, Nobuhiro; Garmaroodi, H. Sadeghi; Taga, Masatoki

doi:10.1016/j.fgb.2009.01.005

Cited by 16 publications

(12 citation statements)

References 59 publications

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“…Clumping and co-migration of bands is a common phenomenon in PFG, as shown, for example, by Eusebio-Cope et al . [42]. Resolution of co-migrating bands requires techniques such as Southern blotting [43] and fluorescence in situ hybridization [44] for accurate discrimination.…”

Section: Discussionmentioning

confidence: 99%

A first genome assembly of the barley fungal pathogen Pyrenophora teres f. teres

et al. 2010

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BackgroundPyrenophora teres f. teres is a necrotrophic fungal pathogen and the cause of one of barley's most important diseases, net form of net blotch. Here we report the first genome assembly for this species based solely on short Solexa sequencing reads of isolate 0-1. The assembly was validated by comparison to BAC sequences, ESTs, orthologous genes and by PCR, and complemented by cytogenetic karyotyping and the first genome-wide genetic map for P. teres f. teres.ResultsThe total assembly was 41.95 Mbp and contains 11,799 gene models of 50 amino acids or more. Comparison against two sequenced BACs showed that complex regions with a high GC content assembled effectively. Electrophoretic karyotyping showed distinct chromosomal polymorphisms between isolates 0-1 and 15A, and cytological karyotyping confirmed the presence of at least nine chromosomes. The genetic map spans 2477.7 cM and is composed of 243 markers in 25 linkage groups, and incorporates simple sequence repeat markers developed from the assembly. Among predicted genes, non-ribosomal peptide synthetases and efflux pumps in particular appear to have undergone a P. teres f. teres-specific expansion of non-orthologous gene families.ConclusionsThis study demonstrates that paired-end Solexa sequencing can successfully capture coding regions of a filamentous fungal genome. The assembly contains a plethora of predicted genes that have been implicated in a necrotrophic lifestyle and pathogenicity and presents a significant resource for examining the bases for P. teres f. teres pathogenicity.

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Section: Discussionmentioning

confidence: 99%

A first genome assembly of the barley fungal pathogen Pyrenophora teres f. teres

et al. 2010

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“…The polyethyleneglycol (PEG)-mediated transfection described by Sun et al (2006) was performed. The purified total RNA containing viral RNAs was introduced into the freshly prepared protoplasts (Eusebio-Cope et al, 2009).…”

Section: Methodsmentioning

confidence: 99%

Identification of a Novel Hypovirulence-Inducing Hypovirus From Alternaria alternata

Bian

Liu

et al. 2019

Front. Microbiol.

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Mycoviruses are wide spread throughout almost all groups of fungi but only a small number of mycoviruses can attenuate the growth and virulence of their fungal hosts. Alternaria alternata is an ascomycete fungus that causes leaf spot diseases on various crop plants. In this study, we identified a novel ssRNA mycovirus infecting an A. alternata f. sp. mali strain isolated from an apple orchard in China. Sequence analyses revealed that this virus is related to hypoviruses, in particular to Wuhan insect virus 14, an unclassified hypovirus identified from insect meta-transcriptomics, as well as other hypoviruses belonging to the genus Hypovirus , and therefore this virus is designed as Alternaria alternata hypovirus 1 (AaHV1). The genome of AaHV1 contains a single large open-reading frame encoding a putative polyprotein (∼479 kDa) with a cysteine proteinase-like and replication-associated domains. Curing AaHV1 from the fungal host strain indicated that the virus is responsible for the slow growth and reduced virulence of the host. AaHV1 defective RNA (D-RNA) with internal deletions emerging during fungal subcultures but the presence of D-RNA does not affect AaHV1 accumulation and pathogenicities. Moreover, AaHV1 could replicate and confer hypovirulence in Botryosphaeria dothidea , a fungal pathogen of apple white rot disease. This finding could facilitate better understanding of A. alternata pathogenicity and is relevant for development of biocontrol methods of fungal diseases.

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“…Transfection of C. parasitica was performed as described by Hillman et al (2004). Spheroplasts of C. parasitica EP155 and Δ dcl- 2 strains were prepared using the method of Eusebio-Cope et al (2009). RnMBV1 virions were purified according to the method of Chiba et al (2009) by differential and sucrose or caesium sulphate gradient centrifugation.…”

Section: Methodsmentioning

confidence: 99%

Biological properties and expression strategy of rosellinia necatrix megabirnavirus 1 analysed in an experimental host, Cryphonectria parasitica

Salaipeth

Chiba

Eusebio-Cope

et al. 2014

Journal of General Virology

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Rosellinia necatrix megabirnavirus 1 (RnMBV1) with a bipartite dsRNA genome (dsRNA1 and dsRNA2) confers hypovirulence to its natural host, the white root rot fungus, and is thus regarded as a potential virocontrol (biocontrol) agent. Each segment has two large ORFs: ORF1 and partially overlapping ORF2 on dsRNA1 encode the major capsid protein (CP) and RNAdependent RNA polymerase (RdRp), whilst ORF3 and ORF4 on dsRNA2 encode polypeptides with unknown functions. Here, we report the biological and molecular characterization of this virus in the chestnut blight fungus, Cryphonectria parasitica, a filamentous fungus that has been used as a model for mycovirus research. Transfection with purified RnMBV1 particles into an RNAsilencing-defective strain (Ddcl-2) of C. parasitica and subsequent anastomosis with the WT strain (EP155) resulted in stable persistent infection in both host strains. However, accumulation levels in the two strains were different, being~20-fold higher in Ddcl-2 than in EP155. Intriguingly, whilst RnMBV1 reduced both virulence and growth rate in Ddcl-2, it attenuated virulence without affecting significantly other traits in EP155. Western blot analysis using antiserum against recombinant proteins encoded by either ORF1 or ORF2 demonstrated the presence of a 250 kDa protein in purified virion preparations, suggesting that RdRp is expressed as a CP fusion product via a "1 frameshift. Antiserum against the ORF3-encoded protein allowed the detection of 150, 30 and 23 kDa polypeptides specifically in RnMBV1-infected mycelia. Some properties of an RnMBV1 mutant with genome rearrangements, which occurred after transfection of Ddcl-2 and EP155, were also presented. This study provides an additional example of C. parasitica serving as a versatile, heterologous fungus for exploring virus-host interactions and virus gene expression strategies.

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Cytological and electrophoretic karyotyping of the chestnut blight fungus Cryphonectria parasitica

Cited by 16 publications

References 59 publications

A first genome assembly of the barley fungal pathogen Pyrenophora teres f. teres

A first genome assembly of the barley fungal pathogen Pyrenophora teres f. teres

Identification of a Novel Hypovirulence-Inducing Hypovirus From Alternaria alternata

Biological properties and expression strategy of rosellinia necatrix megabirnavirus 1 analysed in an experimental host, Cryphonectria parasitica

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