Sweet potato chlorotic fleck virus (SPCFV) has recently been classified as a putative new member of the genus Carlavirus (family Flexiviridae) on the basis of its molecular properties. In this study, SPCFV was characterized in terms of host range, physical and biological characteristics, and genetic variability. In addition to sweet potato, SPCFV infected some plant species in the families Convolvulaceae, Chenopodiaceae, and Solanaceae. Limited numbers of virus particles were observed in the assimilation parenchyma cells of infected plant tissues; some cells had a distorted and enlarged endoplasmic reticulum though without any cytoplasmic and amorphous inclusions. The normal length of SPCFV particles was determined to be approximately 800 nm. In enzyme-linked immunosorbent assays, polyclonal antibodies raised against purified SPCFV virions were able to detect the virus in infected sweet potato and indicator plant tissues. In immunoelectron microscopy, SPCFV particles were all strongly decorated when reacted with homologous antiserum. Comparison of the 3' terminal part of the genome of a range of geographically diverse isolates revealed a high level of genetic diversity. The amino acid sequence identity in the coat protein and the nucleic acid binding protein ranged from 89 to 99.7% and from 75.9 to 99.2%, respectively. Phylogenetic analysis of both proteins showed a geographically associated clustering into two genogroups.

In Africa, the crinivirus Sweet potato chlorotic stunt virus (SPCSV) exists in two serologically and genetically distinct strains, geographically distinguished as a West African (SPCSVWA) and an East African (SPCSVEA) strain. To obtain a better understanding of the genetic diversity among SPCSVEA isolates, the major coat protein (CP) and heat shock protein 70 homologue (Hsp70h) gene sequences of 24 further isolates of SPCSVEA were determined and compared. SPCSVEA diversity was also examined using available monoclonal antibodies (mAbs) to SPCSVEA but there was no apparent coincidence between CP and partial Hsp70h gene nucleotide sequences and the subdivision of SPCSVEA isolates by the mAbs into two serotypes, suggesting this latter may not be of great biological significance. The nucleotide (nt) sequences of isolates of SPCSVEA displayed a high degree of conservation and the only variation observed consisted of a few base exchanges. Pairwise alignments of CP nucleotide sequences revealed differences of <4% between SPCSVEA isolates. Comparisons with published SPCSV sequences confirmed a more distant relationship (up to 34.6% nt; 12% amino acid divergence) between the Hsp70h sequences of isolates of SPCSVEA and SPCSVWA and indicated that SPCSVEA in East and Southern Africa is the more homogeneous than SPCSVWA isolates from West Africa, North and South America, which were up to 12.4% nt divergent among themselves.

In a survey of most sweetpotato‐growing areas of Uganda, virus‐like diseases were observed in all districts surveyed. Out of 338 fields sampled in 35 of the then 42 districts, 219 (65%) had some plants with symptoms. The most common symptoms included vein clearing, mottling, leaf distortion, yellowing, stunting and leaf strapping. Particularly high virus‐like disease incidences (means of 34–86%) were encountered in districts around Lake Victoria and in the Rift Valley in southern and western parts of Uganda; particularly low incidences were encountered in the east and north of Uganda. Using four formats of enzyme‐linked immunosorbent assay in combination with immunoelectron microscopy and polymerase chain reaction assays, five viruses were identified. Sweet potato feathery mottle virus (SPFMV) and Sweet potato chlorotic stunt virus (SPCSV) were most commonly detected, being found in about 90% of samples. Sweet potato mild mottle virus at 10%, Sweet potato chlorotic fleck virus (SPCFV) at 8% and Sweet potato caulimo‐like virus (SPCaLV) at 0·07% were more rarely detected. Most infections were multiple, SPCSV + SPFMV constituting > 90% of all double infections. Triple infections, involving mainly SPFMV, SPCSV and either SPMMV or SPCFV, and quadruple infections of SPFMV + SPCSV + SPMMV + SPCFV were observed in < 10% of the diseased samples. The identification of SPCaLV is the first evidence of its occurrence in Africa.

Since the paucity of information on sweet potato chlorotic fleck virus (SPCFV) had precluded its classification, we have determined the complete nucleotide sequence of the single-stranded RNA genome of a Ugandan isolate of SPCFV. The genome is 9104 nucleotides long (excluding the poly(A) tail) and potentially includes six open reading frames (ORFs). Based on genomic organisation and sequence similarity, SPCFV appears to be a member of the genus Carlavirus (family Flexiviridae). However, SPCFV is distantly related to typical carlaviruses, as most of its putative gene products share amino acid sequence identities of < 40% with those of typical carlaviruses. Its closest relative is melon yellowing-associated virus, a proposed carlavirus from Brazil, with which it shares ORF5 and ORF6 amino acid sequence identities of 61 and 46%, respectively.

Sweet potato virus 2 (SPV2) is a tentative member of the genus Potyvirus, family Potyviridae. In addition to the type isolate of SPV2 recently characterised in greater detail, twelve additional isolates of this virus were obtained from sweet potato clones originating from China, Portugal, South Africa and Zambia. Sequences of the coat protein (CP) gene and 3' non-translated region (NTR) were determined. Comparisons of the CP gene sequences of these isolates revealed nucleotide and amino acid sequence identities ranging from 81 to 99% and from 86 to 99%, respectively. Phylogenetic analysis of sequences distinguished several groups, which partially correlated with the geographic origin of the isolates, and indicated that some isolates from South Africa and a Zambian isolate are most distinct both in CP and 3'NTR sequences. Host range studies of a selected number of isolates revealed some differences in test plant reactions, which appeared to correlate to some extent with the geographic origin and molecular distinctness of the SPV2 isolates. The results strongly suggest the occurrence of biologically and genetically diverse strains of SPV2.

Elimination of diseases particularly viruses is an important concern in the production of planting materials of garlic. In the Philippines, cloves used for planting come from the previous crop or from imported bulbs which have not been certified s disease-free. These poor quality-planting materials result in very low average yield (2.78 t/ha) which is attributed to accumulated diseases through generations of asexual propagation. To solve this problem and to assist the Philippines and other garlic-producing countries in producing certified virus-free planting materials, we developed a technique using sequential shoot tip-meristem culture coupled with indexing. Results of virus-indexing using ELISA, PCR and electron microscopy, however, showed that not all plants were cleaned of the viruses. Hence, an improved technique was developed whereby all the plants were freed of the viruses. The technique consists of cold pre-treatment (5 degrees C) of initial planting materials (bulbs) for 3-4 weeks coupled with thermotherapy (50 degrees C for 2 hrs) and sequential shoot tip-meristem culture followed by multiple shoot production and in vitro bulblet (G(0)) formation prior to transplanting. In vitro bulblets survived better than plantlets when transplanted to soil. Indexing was done using ELISA. This improved technique is now routinely used for production of certified virus-free garlic. Increase in bulb weight (up to 14X, i.e. from G(0) to G(2)) was achieved when G(0) bulblets were transplanted to potting media under greenhouse (3-4 weeks) then field condition to produce 1(st) generation (G(1)) bulbs, then planting the G(1) bulbs in the field to produce 2(nd) generation (G(2)) bulbs, in which the normal size of bulbs was achieved from in vitro bulblets. Tissue-cultured materials had higher yield, in terms of the rate of increase in number bulbs produced per bulb planted, compared to conventionally propagated bulbs (1:4-8 vs 1:3-5).

An incompletely described potyvirus isolate from sweet potato in Taiwan, referred to as 'sweet potato virus 2' (SPV2), was further characterised. Electron microscopy revealed that SPV2 has filamentous particles of 850 nm in length and induces cytoplasmic cylindrical inclusions consisting of pinwheels and scrolls. The virus was mechanically transmitted to several species of the genera Chenopodium, Datura, Nicotiana, and Ipomoea. Two biotypes of Myzus persicae transmitted SPV2 in a non-persistent manner. Decoration titer experiments revealed a distant serological relationship between SPV2 and other potyviruses infecting sweet potato. The 3'-terminal 2006 nucleotides of the viral RNA were determined and shown to be a potyviral genome fragment comprising the coding region for the C-terminal half of the NIb protein, the entire coat protein cistron, and the 3' untranslated region (UTR). Comparison of the capsid protein and 3' UTR sequences of SPV2 with those of other potyviruses demonstrated that it is a distinct member of the genus Potyvirus (family Potyviridae). We propose that SPV2 is named Sweet potato virus Y.

Four hundred and forty-eight symptomatic and 638 asymptomatic samples were collected from sweet potato fields throughout Kenya and analysed serologically using antibodies to Sweetpotato feathery mottle virus (SPFMV), Sweetpotato chlorotic stunt virus (SPCSV), Sweetpotato mild mottle virus (SPMMV), Cucumber mosaic virus (CMV), Sweet potato chlorotic fleck virus (SPCFV), Sweet potato latent virus (SwPLV), Sweet potato caulimo-like virus (SPCaLV), Sweet potato mild speckling virus (SPMSV) and C-6 virus in enzyme-linked immunosorbent assays (ELISA). Only SPFMV, SPMMV, SPCSV, and SPCFV were detected. Ninety-two percent and 25% of the symptomatic and asymptomatic plants respectively tested positive for at least one of these viruses. Virus-infected plants were collected from 89% of the fields. SPFMV was the most common and the most widespread, detected in 74% of the symptomatic plants and 86% of fields surveyed. SPCSV was also very common, being detected in 38% of the symptomatic plants and in 50% of the fields surveyed. SPMMV and SPCFV were detected in only 11% and 3% of the symptomatic plant samples respectively. Eight different combinations of these four viruses were found in individual plants. The combination SPFMV and SPCSV was the most common, observed in 22% of symptomatic plants. Virus combinations were rare in the asymptomatic plants tested. Incidence of virus infection was highest (18%) in Kisii district of Nyanza province and lowest (1%) in Kilifi and Malindi districts of Coast province.

Reverse transcriptase (RT)‐polymerase chain reaction (PCR) and immunocapture (IC)‐RT‐PCR protocols were developed and optimized for the sensitive detection of Onion yellow dwarf virus, Leek yellow stripevirus and allexiviruses infecting Allium species. Polyvalence of the designed primers was successfully demonstrated, using samples of different plant species and geographic origins. Different sample preparation procedures were evaluated for their suitability to provide appropriate PCR templates. IC‐PCR, RT‐PCR with plant tissue extracts, and RT‐PCR with total RNA, proved to be 102–104 times more sensitive than double‐antibody sandwich‐enzyme‐linked immunosorbent assay (ELISA). Furthermore, a ‘one step’ IC‐RT‐PCR assay was developed using plant leaf extract as template source, which proved to be 102 times more sensitive than ELISA, and convenient for testing large numbers of leaf samples.

ABSTRACT An antiserum to shallot yellow stripe virus (SYSV) was raised and used in combination with a range of other antisera to potyviruses of Allium spp. in electron microscopic decoration experiments. The serological results corroborated an earlier finding that the type isolates of SYSV and Welsh onion yellow stripe virus (WoYSV) are closely related to each other and only distantly related to onion yellow dwarf (OYDV) and leek yellow stripe (LYSV) viruses, the two other major potyviruses infecting Allium spp. Moreover, the decoration results indicated that Japanese potyviruses named OYDV and Wakegi yellow dwarf virus are isolates of SYSV. Sequence analysis of the 3'-terminal regions of the SYSV and WoYSV ge-nomes revealed coat protein (CP) amino acid and 3'-nontranslated region (3'-NTR) nucleotide sequence identities of 95 and 89%, respectively. The CP amino acid and 3'-NTR nucleotide sequences of these viruses differed from those of OYDV and LYSV by >25 and >67%, respectively. The serological and molecular studies showed that SYSV and WoYSV are different strains of a potyvirus distinct from OYDV and LYSV. For priority reasons, we propose that these strains together with the Wakegi-type isolates of OYDV described in Japan be referred to as SYSV and that SYSV isolates from Allium spp. other than shallot be designated as the Welsh onion strain of SYSV (SYSV-Wo).

At least nine distinct viruses are currently known to prevalently infect cultivated Allium spp.: two aphidtransmitted Carlaviruses, shallot latent virus (SLV) and garlic common latent virus (GCLV), four aphid-transmitted Potyviruses, leek yellow stripe virus (LYSV), onion yellow dwarf virus (OYDV), shallot yellow stripe (SYSV) virus and the incompletely characterized welsh onion yellow stripe virus (WOYSV), and three mite-borne filamentous viruses whose taxonomic position is still unclear, onion mite-borne latent virus (OMbLV), shallot mite-borne latent virus (SMbLV) and a mite-borne filamentous virus from garlic (MbFV-G). Several host-specific strains or distinct viruses are currently being characterized e.g. garlic-specific strains of LYSV and OYDV. Work at BBA aims at the production of antisera and monoclonal antibodies specific to these viruses in order to facilitate their specific identification. These diagnostic tools are needed for virus indexing during in vitro virus elimination schemes for gene bank material produced at AVRDC. Based upon our results we can define serologically distinct strains of SLV, LYSV and OYDV. We present the current status of knowledge, especially on the serological differentiation of the Allium viruses, and preliminary results of surveys which aimed at determining the identity and distribution of Allium viruses in various Allium crops of different geographical regions. Our observation that different viruses infect Allium in different regions should be taken into account when exchanging vegetatively propagated Allium germplasm between genebanks.