The Cloning Of Three 5'-Proximal Genes Of CHV1 (Cryphonectria Hypovirus 1) And Biological Control Of Chestnut Blight

Hypoviruses are a group of unencapsidated, cytoplasmically replicating double-stranded RNA viruses which can alter host biological processes, resulting in modified colony morphology, suppressed pigmentation, reduced asexual production, loss of female fertility, and reduced laccase production and activity. But most of all, hypovirus will reduce virulence of Ciyphonectria parasitica, of which the discovery presented a new avenue for biological control of chestnut blight.To date three European isolates of Cryphonectria hypovirus 1 (CHV1) which have been submitted to GenBank share very high homology, compared to various divergence of Chinese CHV1 populations according to a group of short sequences reported by co-workers of our laboratory. To further look into the biological diversity of CHV1 in China, we have analyzed a series of longer sequences. Taking the usage of CHV1-CN280 which has been sequenced throughout the genome in our lab, and the other three European isolates, we designed specific primers within the conserved region. And then 14 isolates from China, Japan and Italy were cloned and sequenced in the 5'-proximal region about 3200bp by four split segments, based on the technique of reverse transcript PCR (RT-PCR). Following which the variation and phylogenesis of 5'-UTR, p29, p40, p48 and their orderly spliced sequences were analyzed.We now report the similar diversity of 5'-URT (untranslated region) and protein p29 compared to the more conserved p40, of which the phylogenetic tree shows indistinct branches, while p48 shows great variation in the N-terminal portion and tend to be conserved in the C-terminal region. It is intriguing that a deletion-mutated p29 is discovered in the CHV1-Jal9 isolate. The overlapping codons between the two open reading frames (ORF) of CHV1 are also found to be varied, with that all of North Chinese isolates contain octanucleotide AUGUAUAA, yet South Chinese and European CHV1 isolates incline to share pentanucleotide UAAUG. Besides. three CHV1 isolates from Europe comprising octanucleotide are also discovered with the help of blast in GenBank. In the work of phylogenetic analysis, we have revealed extensive occurrence of RNA recombination among CHV1 populations. For example, the N-terminal region of 5'-UTR and p29 of CHV1-CN431, CHV1-CN238 and CHV1-CHR011 are genetically closer, but CHV1-A 130 in opposite result; and the closer genetic relationship between two Japanese isolates and South Chinese isolates within 5'-UTR and p29 is observed, yet deviate from each other in p48 segment, implying unknown genomic resources for Japanese CHV1. Moreover. European CHV1 are proposed closer relationship with North Chinese CHV1.It is reported in addition about recent achievement in biological control of chestnut blight in Suichang County, Zhejiang province of China. Hypovirulent strains of C. parasitica will encounter challenge for colonization and spread in the field. However, when inoculated in the edge of chestnut blight cankers. CHV1 can be transmitted into natural populations in the field, regardless of vegetative incompatibility. Given to genetic diversity of Chinese C. parasitica, it is good news for biological control of chestnut blight in China, and other more associated factors need to be researched.