Molecular Genetic Variation And Evolution Mechanism Of Sugarcane Bacilliform Virus Populations

Sugarcane bacilliform virus (SCBV; genus Badnavirus; family Caulimoviridae) is considered an important pathogen of sugarcane disease and causes a serious threat to the yield and quality of sugarcane. To exploring the genetic diversity, molecular evolution mechanism and population genetic structure of SCBV isolates in sugarcane planting areas in China,280 sugarcane leaf tissue samples from six provinces in China,25 from three states in USA and 5 from Queensland Australia were tested for the occurrence of SCBV using PCR with a newly degenerate primer (SCBV-F/SCBV-R) designed targeting SCBV RT/RNase H genomic region. PCR results showed that 30.3%(94/310) samples were detected as SCBV-positive. The results further revealed that the SCBV was prevalent among the 6 sugarcane production areas of China. All the PCR amplified fragments were then cloned and sequenced. A total of 122 sequence haplotypes were obtained from the SCBV positive samples, indicating considerable haplotype diversity within individual SCBV isolates. Combined with 45 SCBV sequences from GenBank database, the analysis of Neighbor-joining phylogenetic tree revealed the segregation of global SCBV isolates into three major monophyletic clades encompassing 18 subgroups, including five previously undescribed subgroups named as SCBV-N to-R. Eight SCBV strains (genotypes) were observed in the sugarcane-growing areas of China, and SCBV-Q and SCBV-R were mainly prevalent genotypes. Furthermore, some SCBV variants were found in mixed infection. Genetic differentiation (Snn and Z*) and gene flow (Fst) analysis data indicated that relatively low levels of genetic exchange have occurred within SCBV populations from different countries and the geographic regions of China. The results also indicated that there were a distinct genetic differentiation and a higher level of genetic diversity among SCBV populations.The complete genomes of SCBV-CHN1 and SCBV-CHN2 from China were determined to be 7764 and 7629 base pairs (bp) in size, respectively. Both isolates displayed a typical badnavirus genome organization with three open reading frames (ORFs) but differed in their putative scanning model for P2 (ORF2-encoded) and P3 (ORF3-encoded) protein translation. Phylogenetic analysis revealed the segregation of SCBV-CHN1, SCBV-CHN2 and 10 SCBV isolates from GenBank into nine phylogroups based on complete genomes, individual ORFs, and the RT/RNase H region. SCBV-CHN1 and SCBV-BB were clustered in SCBV-H clade, and SCBV-CHN2 was clustered into SCBV-G clade. SCBV-CHN1 and SCBV-CHN2 shared 70.2% and 80.8% nt sequence identity and genetic distances of 0.32 and 0.22 based on analysis of genome and RT/RNase H region, respectively. Recombination analysis revealed that SCBV-CHN1 and SCBV-CHN2 were putative new recombinant variants arising from putative inter-and intra-species recombination events. The recombination hot-spots of SCBV-CHN1 and SCBV-CHN2 were found in the C-terminal & N-terminal of ORF3 and the IGR region & ORF1, respectively. Recombination was a major driving factor for SCBV evolution.In conclusion, we firstly reported the genetic avariation, molecular evolution driving forces and population genetic structure of Chinese SCBV isolates at RT/RNase H gene and genome levels. These findings further enrich the molecular biological information of SCBV genetic diversity in the world and provide vital support for ecological controls of sugarcane diseases caused by SCBV and molecular breeding of SCBV resistance.