First Report of Bidens mottle virus Infecting Calendula in Taiwan

In March of 2010, calendula (Calendula officinalis L.), a perennial herb known as the pot marigold, showing chlorotic spots on leaves, chlorosis, and stunting were collected from Puli Township, Nantou County, Taiwan. The disorder occurred in more than 50% of the calendula plants in the field. A virus culture isolated from one of the symptomatic calendulas was established in Chenopodium quinoa through triple single-lesion isolation and designated as TwCa1. With transmission electron microscopy (TEM), negatively stained flexuous filamentous virions approximately 12 × 720 nm were observed in the crude sap of TwCa1-infected C. quinoa leaves and pinwheel inclusion bodies were found in the infected cells. On the basis of the sizes of the viral particles and inclusion bodies, isolate TwCa1 was a suspected potyvirus. By reverse transcription (RT)-PCR and potyvirus degenerate primers (Hrp5/Pot1) (1,2), a 0.65-kb DNA fragment, which included the 3′-end of the NIb gene and the 5′-end of coat protein (CP) gene of the virus, was amplified from total RNA isolated from TwCa1-infected plants. The amplified DNA fragment was cloned and sequenced. A homology search indicated that the new calendula-infecting virus in Taiwan might belong to Bidens mottle virus (BiMoV) because its partial genomic sequence shared 94.9 to 97.3% nucleotide and 96.6 to 98.1% amino acid identity with 11 BiMoV isolates available in NCBI GenBank. Primer pairs Hrp5/oligo d(T) were used to amplify the 3′-end genome of BioMV TwCa1 including the 3′-end of the NIb gene, the full-length CP gene, and the 3′-nontranslatable region of the virus. The 807-nt CP gene of TwCa1 (Accession No. HQ117871) shared 97.3 to 98.6% nucleotide and 98.5 to 98.9% amino acid identity with those of 11 BiMoV isolates available in GenBank. Results from TEM observations and CP gene sequence analysis indicated that TwCa1 is an isolate of BiMoV. BiMoV was later detected by RT-PCR in eight symptomatic calendulas collected from the same field. To our knowledge, this is the first report of BiMoV infecting calendula in Taiwan. This newly identified calendula-infecting BiMoV could have a direct impact on the economically important vegetable and floral industry in Taiwan.