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An isolate of hippeastrum chlorotic ringspot virus (HCRV) named HLS1-2, causing necrotic spots on leaves of spider lily, was obtained and characterized in China. The complete S RNA and M RNAs of the HLS1-2 isolate are 2724 nt and 4741 nt in length, respectively. The HLS1-2 S RNA sequence is most closely related to that of HCRV, with 99 % identity. Viral proteins encoded by the M RNA are closely related to those of tomato yellow ring virus (TYRV), polygonum ringspot virus (PolRSV) and iris yellow spot virus (IYSV). Phylogenetic trees for the four viral proteins encoded by the S and M RNAs placed HCRV-HLS1-2 in a distinct cluster with IYSV, TYRV and PolRSV and provided further support for the subdivision of tospoviruses into American and Eurasian groups.

A new tospovirus, HCRV 2007-ZDH, was isolated from a Hippeastrum sp. plant displaying necrotic and chlorotic ringspot symptoms in Yunnan province. This virus isolate was characterized based on particle morphology and RNA sequences analyses. Quasi-spherical, enveloped particles measuring about 70-100 nm, typical of tospoviruses, were observed in sap and cells of the infected plants. Transmission studies by inoculating this isolate mechanically to Hippeastrum sp. confirmed that 2007-ZDH is the causal agent of the chlorotic ringspot disease of Hippeastrum sp. The complete sequence of S RNA of 2007-ZDH was 2,744 nucleotides in length, sharing 74.4 % nucleotide identity with Tomato yellow ring virus (TYRV) isolate tomato (AY686718). The S RNA encoded a non-structural protein (NSs) (444 aa, 50.4 kDa) and the nucleocapsid (N) protein (273 aa, 30.1 kDa).The deduced NSs protein shared amino acid identities of 78.6, 76.3, and 74.9 % with that of TYRV, IYSV, and PolRSV, respectively. The deduced N protein shared amino acid identities of 86.1, 84.7, and 70.0 % with that of PolRSV, TYRV, and IYSV, respectively. These results suggest that the chlorotic ringspot virus belongs to a new tospovirus species, for which the name Hippeastrum chlorotic ringspot virus (HCRV) is proposed.

Complete genome sequences were obtained from two isolates of the carlavirus nerine latent virus from hippeastrum and narcissus plants, two isolates of the potyvirus hippeastrum mosaic virus from a hippeastrum plant, and one isolate each of the potyviruses narcissus degeneration virus, narcissus yellow stripe virus and Vallota speciosa virus from narcissus plants. Proposals are made to clarify the current confusion surrounding the naming of some of these viruses.

Plants of Triteleia hyacinthina, Triteleia ixioides Starlight, and Triteleia laxa Corina with severe mosaic and yellow vein-banding were found to be infected with a potyvirus. The 3'-terminal region of the virus was amplified by RT-PCR from total RNA using a potyvirus-specific degenerate primer (poty5P: 5' GGN AAY AAY AGY GGN CAR CC 3') and an oligo-dTprimer. The sequence generated included the 3'-NIb protein coding region (680 nucleotides), the entire coat protein coding region (840 nucleotides), and 3'-untranslated region (UTR) (253 nucleotides). Amino acid identity of the whole CP between the triteleia virus and potyvirus member ranged from 54% Apium virus Y (ApVY) to 67% Auraujia mosaic virus (ArjMV) and Twisted-stalk chlorotic streak virus (TSCSV) and the core ranged from 59% (ApVY) to 75% (ArjMV). The 3-UTR showed no significant homology with other known potyviruses. Phylogenetic relationships suggest this triteleia virus is a new member of the Potyvirus genus and the name of "Triteleia mosaic virus" (TriMV) is proposed. This is the first report of a potyvirus infecting triteleia.

Lily symptomless virus (LSV) occurs frequently in many Lilium species worldwide and often causes developmental abnormalities such as a smaller flower and lower bulb yield. In this study, two moderate and efficient gel filtration chromatography (GFC) methods was compared, these two techniques were, respectively, based on Superdex-200 HR and Sephacryl S-1000 SF. The products purified by the two methods were then characterized by measurements with UV-spectrophotometer, reverse transcriptase (RT)-PCR, transmission electron microscope (TEM), polyacrylamide gel electrophoresis (PAGE), Western blotting and matrix assisted laser desorption-ionisation/time-of-flight mass spectrometry (MALDI-TOF-MS). The final yield of purified LSV by the Superdex-200 HR GFC was 9.4 mg from 50 g of fresh infected tissues of Lanzhou lily. However, from the same amount tissues, only 5.6 mg of LSV were obtained by using Sephacryl S-1000 SF GFC. The Superdex-200 HR method was thus shown to be more suitable for the purification of LSV than the Sephacryl S-1000 SF GFC. The Superdex-200 HR method does not require costly equipment for density centrifugation and ultracentrifugation. Furthermore, it can provide an economical and efficient way to obtain purified products for the preparation of antibodies for serological diagnosis or LSV infection and related investigations.

In four distinct alstroemeria-infecting cucumber mosaic virus (CMV) isolates, additional sequences of various lengths were present in the 3' nontranslated regions of their RNAs 2 and 3, apparently the result of intra- and intermolecular recombination events. Competition experiments revealed that these recombined RNA 2 and 3 segments increased the biological fitness of CMV in alstroemeria.

An overview is presented on the management of viruses of Lilium crops in the Netherlands since the 1960s. This mainly concerns Lily symptomless virus (LSV) and Lily mottle virus (LMoV). Various factors which affect the efficiency of control are considered. The variable symptoms of LMoV in the many vegetatively propagated cultivars grown (c. 340) pose problems in the efficient roguing of diseased plants of some cultivars. Additionally the acceptable incidence of viruses such as LSV which are generally symptomless in field-grown plants may cause problems under the unfavourable light and growing conditions of year-round cut-flower production in greenhouses. The reduction in bulb yield and quality caused by viruses of lilies necessitates a further decrease in the already low virus levels still tolerated in lily bulb stocks. The routine detection of LSV and LMoV by ELISA has been developed extensively over the years. The impact of testing bulbs ('bulb test') during storage has been important in achieving an overall decrease in virus incidence in many stocks of lily cultivars, e.g. in Asiatic hybrids. The 'leaf test' used to assess many other cultivars, e.g. oriental hybrids, in which the bulb test for LMoV is not applicable, will be developed so as to eliminate at an early stage severely infected stocks that were initially intended for further propagation. The spread of the aphid-borne LSV and LMoV generally occurs very rapidly. The low virus incidence in the initially virus-tested stocks obtained by tissue culture procedures is effective in decreasing the access of vectors to virus-infected lilies. The viruses spread mainly in June and July, considerably less in May and least in August/September. Consequently the routine spraying of mixtures of mineral oil and pyrethroid insecticide is generally done weekly in May, June and July and fortnightly in August and September. There are differences in efficacy of the different brands of mineral oil. The rapid propagation of lilies, as done routinely by the scaling of bulbs and by tissue culture procedures, enables the rapid bulking of virus-tested and other stocks of high quality which have the health status required. The impact of the different factors in the management and control of viruses in Lilium crops in the Netherlands is discussed.

Molecular detection of Yam mosaic virus (YMV), the most important potyvirus for yam plants (Dioscorea spp.) has been limited by its high genetic diversity. According to recent phylogenetic data, suitable molecular tools were developed for detection and genotyping. A reliable and rapid molecular test is described using single-tube immunocapture (IC)-RT-PCR combining the use of a monoclonal antibody able to recognise all YMV serotypes and primers designed to take into account the whole variability of YMV. Phylogenetic data were then used to develop an uncoupled IC-RT-PCR procedure adapted to the genotyping of the three major phylogenetic groups and the two recombinants implicated in the YMV epidemics in the Caribbean and French Guiana. A third exploitation of phylogenetic data was carried out with the development of a typing test based on the analysis of short nucleotide sequences. The direct sequencing of a 450 nts fragment constituted a reliable complementary typing tool as the resulting information was quite similar to that obtained with sequences of the complete coat protein gene. The approach, consisting in a robust study of YMV variability followed by the use of phylogenetic data to develop reliable detection and genotyping tools, offers new perspectives for powerful molecular epidemiological studies.

An attempt was made to distinguish between celosia mosaic virus (CIMV) and asparagus virus 1 (AV-1) based on biological properties, which hitherto was obscured from serological data from previous work. The host range of AV-1 was found to be a subset of that of CIMV and AV-1 was transmitted by the aphid Myzus persicae which, on the other hand, did not transmit CIMV. No evidence of cross-protection was obtained between these two viruses.

To evaluate the genetic diversity and understand the evolution of Yam mosaic virus (YMV), a highly destructive pathogen of yam (Dioscorea sp.), sequencing was carried out of the C-terminal part of the replicase (NIb), the coat protein (CP) and the 3'-untranslated region (3'-UTR) of 27 YMV isolates collected from the three main cultivated species (Dioscorea alata, the complex Dioscorea cayenensis-Dioscorea rotundata and Dioscorea trifida). YMV showed the most variable CP relative to eight other potyviruses. This high variability was structured into nine distant molecular groups, as revealed by phylogenetic analyses and validated by assessment of the molecular evolutionary noise. No correlation was observed between the CP and 3'-UTR diversities and phylogenies. The most diversified and divergent groups included isolates from Africa. The remaining groups clustered in a single clade and a geographical distinction between isolates from the Caribbean, South America and Africa was observed. The role of the host in the selection of particular isolates was illustrated by the case of a divergent cultivar from Burkina Faso. Phylogenetic topological incongruence and complementary statistical tests highlighted the fact that recombination events, with single and multiple crossover sites, largely contributed to the evolution of YMV. We hypothesise an African origin of YMV from the yam complex D. cayenensis-D. rotundata, followed by independent transfers to D. alata and D. trifida during virus evolution.

We determined the complete nucleotide sequence of a mild strain of Japanese yam mosaic potyvirus (JYMV-M) and compared it with the published sequence of severe strain of JYMV (JYMV-J1). The genomic RNA of JYMV-M is 9,760 nucleotides (nts) in length, excluding the poly (A) tail, and encodes a polyprotein of 3,132 amino acids. Among nine potential cleavage sites, only the P1 and NIa recognition sites (between 6K1 and CI) had different sequences from those of JYMV-J1. The data confirm the strain status of these two viruses with 91.1% sequence identity for the polyprotein and approximately 94-97% identities for HC, CI, NIa, NIb and CP. The most divergent products P1 and P3 had 62% and 90% sequence identities respectively.

The 3'-terminal regions of the genomic RNAs of two Korean isolates of the lily symptomless Carlavirus (LSV), LSV-Ko and LSV-KII, were cloned and their nucleotide sequences were determined. The nucleotide sequence analysis and protein analysis by the Western blot revealed that E. coli expressed a 32-kDa protein that is the viral coat protein (CP) for the LSV. The two Korean strains share 98.4% and 98.3% sequence identities at the nucleotide and amino acid levels, respectively. The CP gene of LSV-Ko showed 99.1% and 87.0% nucleotide sequence identities, and 99.0% and 96.6% amino acid sequence identities with those of the Netherlands and the Japanese LSV strains, respectively. A pairwise amino acid sequence comparison revealed a sequence similarity of 29.6% to 69.8% between LSV-Ko and other species of the carlavirus. The 16 kDa protein of LSV-Ko shares 17.6% to 42.7% amino acid similarity with those of 8 other the carlaviruses, and they are variable in the N-terminal region. The Cys repeated zinc finger nucleic acid binding domain was found in the 16 kDa protein for all of the LSV strains. Sequence comparisons of the 7 kDa protein of LSV in the strain level showed significant identities from 100.0% to 98.4%. LSV-Ko shares 21.9% to 42.2% amino acid similarity with those of 8 other carlaviruses, 4 members of the potexviruses, and a closterovirus. LSV is closely related to blueberry scorch virus (BISV) based upon the phylogenetic tree analyses of the three proteins, indicating LSV to be a quite distinct member of the genus Carlavirus.

We determined the 3'-termimus 1353 nucleotides (nts) in length excluding the poly (A) tail of yam mild mosaic potyvirus (YMMV) RNA. The sequence starts within a long open reading frame (ORF) 1209 nts and is followed by untranslated region (3'-UTR) of 144 nts. The coat protein (CP) contains 266 amino acids (aa) with molecular ratio (Mr) of approximately 30 kDa. The CP of YMMV differs substantially from yam mosaic virus (YMV), Japanese yam mosaic virus (JYMV) (57 and 61% of amino acid sequence identity) and other potyvirus species. This result suggests that YMMV should be classified as a new yam potyvirus.

Asparagus virus 2 (AV 2) has been detected in all major asparagus growing areas around the world and it has been suggested that, in association with other pathogens, it might be an important factor for the condition called asparagus decline. Although AV 2 was first reported in the 70's, only few of its biochemical characteristics have been actually studied. Some properties of an AV 2 isolate from Guanajuato, Mexico were characterized and its RNA3 cloned and sequenced. Electrophoretic analysis of AV 2 RNA showed 3 genomic RNAs (RNA1. 3.0; RNA2 2.7; and RNA3 2.3 kb) and a fourth, subgenomic, RNA4 of 1.1 kb. In vitro translation data confirmed that coat protein is produced by translation of the subgenomic RNA4, as reported for other bromoviruses. Sequence analysis of AV 2 RNA3 showed two open reading frames identified by sequence comparisons and immunological assays as coding the movement and coat proteins (MP and CP, respectively). AV 2 CP showed a high percentage of identity with the CPs from other members of the subgroup II of the ilarvirus genus. The percentage of identity with other ilarviruses decreased dramatically and in some cases was lower than the values obtained with members of different genera.