The coat protein sequences of the two isolates are identical and share 67.3% aa identity to HoCSV. Pairwise alignment using Clustal Omega (1.2.2) showed that the amino acid (aa) sequences of the movement protein (MP) of the two isolates of HoToV2 share 99.7% aa identity to each other and 52.3% identity to HoCSV. In sample JKI2001336, HoToV2 was detected whilst in sample JKI2001337, HoToV2 and Tomato spotted wilt virus were detected in a mixed infection. Two of the samples (JKI2001336 and JKI2001337) were sequenced using MinION sequencing as described by Gaafar et al. The sequences showed 69.4% nucleotide (nt) identity to Hoya chlorotic spot virus (HoCSV KX434725) (Adkins et al., 2018) indicating the presence of a novel tobamovirus, provisionally named Hoya tobamovirus 2 (HoToV2).

800 bp were Sanger sequenced in both directions (Microsynth Seqlab, Germany). Total RNA was extracted from infected leaf material and a generic tobamovirus RT-PCR was performed using the Tob-Uni 1 and Tob-Uni 2 primers of Letschert et al. tabacum developed chlorotic lesions four weeks post inoculation. Following mechanical transmission, Nicotiana benthamiana and N. 2B) but there was no reaction with other tobamovirus antisera. Virions showed a cross-reaction with a Youcai mosaic virus antiserum (DSMZ, Germany) in immunosorbent electron microscopy (Fig. The plants were analysed by electron microscopy and non-enveloped rod-shaped particles were observed, suggesting infection with a tobamovirus (Fig. ringspots, necrotic lesions, and irregular chlorosis or chlorotic leaf spots (Fig. The plants had been originally imported from Thailand and a large proportion developed symptoms consistent with viral infection, e.g. (family: Asclepiadaceae) was grown in a German nursery. During 2019/20, a batch of 6,000 young plants of different Hoya spp.

Microsynth seqlab