Alstroemeria-Infecting Cucumber Mosaic Virus Isolates Contain Additional Sequences in the RNA 3 Segment

The coat protein (CP) genes and flanking regions of three alstroemeriainfecting cucumber mosaic virus isolates (CMV-ALS), denoted ALS-LBO, ALS-IPO, and ALS-NAK, were cloned and their nucleotide sequence determined and compared at both nucleic acid and deduced protein level with the published sequences of CMV RNA 3. The sequences of these isolates showed more than 95% nucleotide and peptide sequence homology to each other and to other members of subgroup II CMV. Strikingly, an additional sequence of 218 nt was found in the central region of the 3’-non-translated region (NTR) of RNA 3 of two out of three isolates. The additional sequence appeared to have arisen from RNAs 1 or 2. A subgroup-specific DIG-labeled probe has been developed from this additional sequence and applied to detect subgroup II CMV strains in dot blot hybridization and to differentiate the Alstroemeria isolates containing additional sequences in Northern blot hybridization. INTRODUCTION Alstroemeria has become an important ornamental crop during the past decade in the Netherlands and all over the world (van Zaayen, 1995). Several viruses infecting alstroemeria have been described, including alstroemeria mosaic potyvirus (AlMV), alstroemeria carla virus (AlCV) (Phillips et al., 1986), alstroemeria streak potyvirus (AlSV) (Wang et al., 1992; van Zaayen et al., 1994), tomato spotted wilt tospovirus (TSWV) (Hakkaart and Versluijs, 1985). Also cucumber mosaic cucumovirus (CMV) (Hakkaart, 1986) has been reported in Alstroemeria. However, very little was reported on the interaction between CMV and alstroemeria plants because of its sporadic occurrence. Since CMV in alstroemeria usually was reported together with other viruses (van Zaayen, 1995), the symptoms caused by CMV-infection were not clear and not described until 1996. Bellardi and Bertaccini (1996) reported that CMV-infected alstroemeria showed necrosis on the leaves of Alstroemeria cv. Red Sunset and malformed flowers on Alstroemeria cv. Rossella. In addition, chlorotic mosaic on the leaves and stunting also were observed in Alstroemeria Red Sunset. The combination of CMV with other alstroemeria viruses can induce different symptoms (necrosis. red stripes, yellowing) which could depend upon environmental conditions, alstroemeria cultivars and growth stage when infection occurred (Bellardi and Bertaccini, 1997). Seed transmission of CMV in alstroemeria was reported as well (Bellardi and Bertaccini, 1997). Antisera against alstroemeria CMV strains have been developed and have been applied to routine inspection. Cucumber mosaic virus (CMV) is the type species of the genus Cucumovirus in the family of Bromoviridae (Rybicki, 1995) and consists of isometric particles with a diameter of about 28 nm. Its genome consists of three functional single-stranded RNA molecules designated as RNA 1, 2, and 3, which are required for infection. RNA 1 and 2 encode components of the viral RNA-dependent RNA polymerase (Nitta et al., 1988), while RNA 3 encodes the movement protein (MP) (Suzuki et al., 1991) and coat protein (CP) (Schwinghamer and Symons, 1977). The CP is translated from the subgenomic RNA 4 which is encoded by the 3’-half of the RNA 3 (Gould and Symons, 1982). A small Proc. 10 IS Virus Diseases Ornamentals