Table 1.
Primers used for amplification of the MVBaV genome.
Table 2.
List of accession number and abbreviation of Tospoviruses used in this study.
Fig 1.
The symptoms of MVBaV-infected mulberry leaves.
(A) vein banding, (B) mosaic, (C) chlorotic spots, (D) leaf deformation and (E) vein necrosis. The photos were taken over a period of eight months from April to November in 2013.
Fig 2.
Electron micrograph of a MVBaV-infected mulberry plant leaves showing typical tospovirus-like particle morphology.
Typical spherical, enveloped virions are shown accumulating in the endoplasmic reticulum (A) or dispersing in the cytoplasm as single particles in leaf cells (B). The scale bar represents 200 nm.
Fig 3.
Cloning strategies for L, M, and S RNAs.
Arrows indicate the annealing positions of each primer. To determine the 5′-terminal sequences, total RNAs were denatured by heating at 65°C for 5 min and then mixed with the first primer, LRNA 5–1 for L RNA, MRNA 5–1 for M RNA, and SRNA 5–1 for S RNA, respectively. After removal of template RNAs by RNaseH digestion, PCR amplification of the 5′-cDNAs was performed with Ex Taq DNA polymerase (Takara Bio, Dalian, China) using UPM primer (provided with the kits) and a nested primer, LRNA 5–2 for L RNA, MRNA 5–2 for M RNA, and SRNA 5–2 for S RNA, respectively. To determine the 3′-terminal sequence, first strand cDNA was synthesized by using the primer Tos-UPA. PCR amplification of the 3′-cDNAs was performed with Ex Taq DNA polymerase using the primers Tos-UPA and the LRNA 3 primer for L RNA, MRNA 3 for M RNA, and SRNA 3 for S RNA, respectively. The primers used for RACE and amplifying the MVBaV Genome are shown in Table 1.
Fig 4.
The RdRp conserved motifs in viruses of the genus Tospovirus.
The consensus amino acid residues in each RdRp motif of the family Bunyaviridae are shown in bold and underlined. Identical amino acid (aa) residues are indicated with dots and deficient aa residues with hyphens. The positions of the conserved motifs in RdRp are indicated. Abbreviations and accession numbers of the analyzed sequences of tospoviruses are listed in Table 2.
Fig 5.
The NSm conserved motifs in viruses of the genus Tospovirus.
The positions of the conserved motifs in NSm are indicated. Abbreviations and accession numbers of the analyzed sequences of tospoviruses are listed in Table 2.
Fig 6.
The NSs conserved motifs in viruses of the genus Tospovirus.
The positions of the conserved motifs in NSs are indicated. Abbreviations and accession numbers of the analyzed sequences of tospoviruses are listed in Table 2.
Table 3.
Comparison of the RNAs and deduced proteins of MVBaV with those of other Tospoviruses.
Fig 7.
Phylogenetic trees of tospoviruses.
Amino acid sequences of the RNA-dependent RNA Polymerase (RdRp), GP protein (GP),NSm protein (NSm),NSs protein (NSs), and N protein (N) were used to constructed the trees. The dendrograms were produced using the Neighbour—Joining algorithm with 1000 bootstrap replicates. The tospovirus species belonging to WSMoV serogroup are circled. The scale bar represents a relative genetic distance. Numbers above critical branches are significant bootstrap values (>70%). Abbreviations and accession numbers of the analyzed sequences of tospoviruses are listed in Table 2.
Table 4.
Comparison of the S RNA from isolate XCSY-3 to those of nine additional MVBaV isolates.