Natural mixed infections on Phalaenopsis species by groundnut bud necrosis orthotospovirus, cymbidium mosaic virus and odontoglossum ringspot virus in India

Three viruses belonging to different plant virus genera were found associated with Phalaenopsis plants showing symptoms of mild chlorotic spots, mild mosaic and dark green islands with light green depressions. Symptomatic and non-symptomatic plant samples were collected from the experimental farm of National Research Centre of Orchids, Pakyong Sikkim, India. Electron microscopy revealed the presence of virus particles with enveloped quasi-spherical, flexuous and rod-shaped particles morphologies measuring 80–110 nm, 480 × 13 nm and 300 × 18 nm respectively. In DAC-ELISA the symptomatic Phalaenopsis sample reacted with the antibodies raised previously to groundnut bud necrosis orthotospovirus (GBNV), cymbidium mosaic virus (CymMV) and odontoglossum ringspot virus (ORSV). Sequence information generated from the cloned RT-PCR amplified products, followed by sequence identity and phylogeny analyses, further confirmed the identity of the three different viruses as GBNV, CymMV and ORSV. Upon mechanical sap transmission on the N. benthamiana test plants, the phalaenopsis sample with mixed infection could induce a range of mixed symptoms such as systemic symptoms with blotching, mosaic, mottling and green-islands on the leaves, unlike those indicative of the symptoms known for the single infection by CymMV or GBNV or ORSV. The virus transmission and symptomatology revealed that the mixed infection by these three viruses masked the known symptoms of respective viruses induced individually. The present investigation describes the mixed infection by GBNV, CymMV and ORSV on Phalaenopsis for the first time in the world.