Different approaches to search for mycoviruses in a collection of fungal endophytes isolated from the marine plant Posidonia oceanica
| Autor: | Turina M., Nerva L., Ciuffo M., Frola M., Gnavi G., Vallino M., Varese G.C. |
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| Rok vydání: | 2014 |
| Předmět: | |
| Zdroj: | The Third International Mycovirus Symposium, August 2-5, 2014 Burlington, VT 05405, Burlington, VT USA, August 2-5, 2014 info:cnr-pdr/source/autori:Turina M., Nerva L., Ciuffo M., Frola M., Gnavi G., Vallino M., Varese G.C./congresso_nome:The Third International Mycovirus Symposium, August 2-5, 2014 Burlington, VT 05405/congresso_luogo:Burlington, VT USA/congresso_data:August 2-5, 2014/anno:2014/pagina_da:/pagina_a:/intervallo_pagine |
| Popis: | The Mycotheca Universitatis Taurinensis (MUT) preserves a wide collection of endophytic fungal species isolated from the sea grass Posidonia oceanica, a key component in the ecology of the Mediterranean sea. We have selected 91 isolates representative of the fungal taxa isolated from P. oceanica and assessed their halophylic properties growing them on solid media in presence or absence of marine salts (3%). Growth data were used to select the most appropriate media for liquid culture. Lyophilized mycelia were used for total nucleic acids and dsRNA purification. Total nucleic acid was digested with RNAse and used as template in a Rolling Cycle Amplification (RCA) protocol in order to determine the possible presence of circular ssDNA (possibly viruses). In parallel, samples positive for dsRNA presence were instead subjected to DNAse and prepared for traditional cloning and sequencing; the same dsRNA samples were also prepared for direct sequencing using a next generation sequencing (NGS) platform. We found 16 samples positive for dsRNA presence. So far we have confirmed the presence of mycovirus sequences in 7 isolates, and each isolate contained virus sequences sufficiently distant from those deposited in the databases to warrant a "new species" status; nevertheless, all the viruses so far characterized molecularly belong to known taxonomic groups. Six isolates were shown to contain abundant accumulation of dsRNA with homology to known fungal transposon/retro-transposon sequences. The three remaining dsRNAs are still under characterization. So far only one sample was positive in RCA analysis (out of 91 tested) and it carried sequence homology to Neurospora and Cryphonectria mitochondrial plasmids present in the database. All dsRNA positive samples were also subjected to a partial virus purification protocol and electron microscopic observation and presence of particles was confirmed in almost all the samples. Penicillium aurantiogriseum var. viridicatum MUT 4330 contained isodiametric particles of at least two different diameters. We screened colonies from single conidia progeny for presence/absence of each of the dsRNA bands present in the original 4330 isolate: so far we have derived at least two different dsRNA patterns that are isogenic for nuclear background and show different phenotype in presence or absence of marine salts in the media. |
| Databáze: | OpenAIRE |
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