Identification And Variability Analysis Of Geminiviruses On Malva Parviflora In Sichuan Province

Geminiviruses are the only plant viruses characterized by having a circular single-stranded ssDNA genome packaged within twinned icosahedral particles. Most of geminiviruses have caused significant yield losses to many crops in tropical and subtropical regions worldwide. Based on insect vectors, genome characterizations and hosts range, Geminiviridae is divided into four genera(Mastrevirus, Curtovirus, Topocuvirus and Begomovirus). Begomoviruses, which are transmitted permanently by whiteflies in nature, are also called as WTG (Whitefly-transmitted geminivirus). Begomoviruses are widely spreaded and have caused severe losses on crop production in 39 countries around the world. Recently, begomoviruses have been found in many crops such as tobacco, tomato, papaya and weeds in the regions like Yunnan, Hainan, Guangdong, Guangxi and Fujian province, etc. In addition, it was also reported that begomoviruses were found in tomatoes, peppers and other crops and weeds such as Malvastrum coromandelianum in Sichuan province. Weeds are considered to be an important intermediate host (between insects and crops) and initial sources of infection. In order to identify begomoviruses infecting Malva parviflora in Sichuan province, the molecular identification of pathogens that caused yellow vein in Malva parviflora samples collected from Sichuan were conducted, as well as genome organization and mutation were analyzed. Using degenerate primers PA and PB, ten virus samples, collected from Malva parviflora showing yellow vein symptom from Panzhihua of Sichuan province in 2010, were tested by PCR. The results showed that a specific band about 500bp was amplified from nine of ten samples (expect SC193), which revealed nine samples were infected by geminivirus. Partial samples were chosed to be tested by RCA-PCR and an expected size product of 500bp was obtained from SC193, which showed ten samples were all infected by geminivirus. The 500bp fragments from Malva parviflora samples were cloned and sequenced, sequence analysis of PCR products showed that these samples are probably infected by two kinds of geminiviruses which include Tomato yellow leaf curl China virus (TYLCCNV) and Malvastrum yellow vein Yunnan virus (MYVYNV).Complete genomic structure of geminiviruses infecting samples of SC176, SC224 and SC226 chosed optionally were studied. The complete nucleotide sequence of DNA-A of SC176 and SC226-1 were analyzed. The results showed that DNA-A of SC176 and SC226-1 contain 2737 and 2738 nucleotides respectively, and have typical genomic organization of begomovirus. They showed relatively high sequence identity (96.9%) with each other, and are most related to that of TYLCCNV with 98.2% and 97.1% nucleotide sequence identity respectively, which indicated they are different isolates of TYLCCNV. The complete nucleotide sequence of DNA-A of SC224 and SC226-2 were analyzed. The results showed that DNA-A of SC224 and SC226-2 contain 2747 nucleotides and 2748 nucleotides respectively, and have typical genomic organization of begomovirus. They shared relatively high sequence identity (99.2%) with each other, and 99.0% and 99.3% with that of MYVYNV respectively, which indicated they are different isolates of MYVYNV. Using special primers for satellite DNA(3 molecules associated with geminiviruses, the full length of DNAβwere obtained from SC176, SC224 and SC226. Sequence comparison showed that SC224βand SC226βwere 1356bp and 1350bp in length respectively. They showed relatively high sequence identity (99.0%) with each other, and are most related to that of DNAβassociated with MYVYNV (MYVYNB) with 99.0% and 99.1% nucleotide sequence identity respectively. The SC176βis 1335 nucleotides long and shared 85.8% nucleotide sequence identity with that of DNAβassociated with TYLCCNV (TYLCCNB). These resultes showed that SC176 and SC224 are infected by TYLCCNV and MYVYNV respectively, and SC226 is co-infected by TYLCCNV and MYVYNV. Co-infection of TYLCCNV and MYVYNV was tested with the specific primers among the ten samples. The results showed that the co-infections of TYLCCNV and MYVYNV were found in seven of ten samples. The ratio of co-infection is 70%. While the single infection of TYLCCNV or MYVYNV was found in other three samples. These resultes indicated that co-infections of begomoviruses occur commonly in fields.