Identification And Characterization Of A Novel SsDNA Mycovirus In The Phytopathogenic Fungus Diaporthe Pseudophoenicicola

Areca catechu L is one of the second largest tropical cash crops in Hainan.Due to the shallow and small root system of areca and improper production management,the occurrence of areca diseases has been aggravated in recent years,and the areca leaf spot disease has seriously affected its planting and yield.In this study,a novel DNA mycovirus were extracted from Diaporthe pseudophoenicicola strain HNWC-3,which was isolated from areca plantation in Wenchang city,Hainan province.The whole genome of the virus was cloned and sequenced.We also further study the effect of the virus on the biological properties of host strain HNWC-3.All the results are as follows:Here,we used Illumina and Sanger sequencing to determine the genome sequence of a novel circular DNA virus from the strain HNWC-3.The full-length genome sequence of Dp CV1 was obtained by both polymerase chain reaction(PCR)and rolling circle amplification(RCA)methods.The complete genome of the virus is 3143nucleotides(nts)in length and contains two ORFs encoding a large putative replication initiation protein(Rep)and a capsid protein(CP).The first ORF(sense)is 939-nt in length,encoding a 313-aa putative capsid protein(CP),and the second ORF(antisense)encodes a 434-aa replication-associated protein(Rep),which was spliced and formed two small Reps by a 179-nt putative intron.The 5‘-and 3‘-terminal sequence of the intron indicated that it has the characteristic of U2 plant introns.The Rep and the CP ORFs were separated by two intergenic regions: the large intergenic region(LIR)and the small intergenic region(SIR)separately contained 708 and 9 nts,respectively.A non-nucleotide (?)TAATATTAC‘ was analyzed and possibly become the initiation site for viral DNA replication.Phylogenetic analysis based on the amino acid(aa)sequence of the Rep protein did not cluster Dp DV1 into any established genus of the family Genomoviridae.Therefore,we suggest that Dp DV1 may belong to a new genus within the family Genomoviridae.We temporarily named it Diaporthe pseudophoenicicola DNA virus 1(Dp DV1).To identify the effect of Dp DV1 on the D.pseudophoenicicola populations,a virus-free stain HNWV-P7 was obtained by the protoplast regeneration method.Comparing to the virus-free strains,the results indicated that Dp DV1 could enhance aerial hyphae and decrease the pathogenicity of D.pseudophoenicicola strain HNWC-3.The results indicated that the virus Dp DV1 had the potential to biological control the leaf spot of areca.In this study,we report a novel genomovirus,Dp DV1,which resulted in an increase in the aerial mycelium and attenuated virulences of D.pseudophoenicicol.This is the first report of a DNA mycovirus infecting D.pseudophoenicicola,which can significantly diminish the host virulence and is expected for further application in biocontrol of the fungal disease.