Species: Lettuce infectious yellows virus
Bryce W. Falk
Department of Plant Pathology, University of California, 1 Shields Ave., Davis, CA 95616, USA
Department of Plant Pathology, University of California, 1 Shields Ave., Davis, CA 95616, USA
Host Range and Symptomatology
Transmission by Vectors
Transmission through Seed
Transmission by Grafting
Transmission by Dodder
Nucleic Acid Hybridization
Stability in Sap
Properties of Particles
Properties of Infective Nucleic Acid
Relations with Cells and Tissues
Ecology and Control
LIYV virions are morphologically polar long flexuous filaments, about 12 × 800-900 nm. They are probably of two types, one for each of the two genomic ssRNA species, RNA-1 (8,118 nt) and RNA-2 (7,193 nt). LIYV infections are phloem-limited within the plant host, and the virus is not mechanically transmissible. It is transmitted from plant to plant by the sweet potato whitefly, Bemisia tabaci, in a semi-persistent manner, the insects remaining viruliferous for only 3 days or less. LIYV has a wide host range among dicotyledonous plants and has caused significant economic losses in important food crops, including cucurbits and lettuce. The geographical incidence is limited primarily to southwestern USA, where incidence in recent years has declined dramatically.
LIYV was originally discovered in 1981 when commercial lettuce plantings in southwestern USA exhibited sudden and severe yellowing and stunting. Coincident with the discovery of LIYV was the appearance of the sweet potato whitefly, Bemisia tabaci, which built up high populations during the summer. Many autumn-planted vegetable crops, including lettuce, sugarbeets, crucifers and cucurbits, were subsequently affected by LIYV. In the early 1980's, 100% of susceptible crop plants were affected and LIYV caused economic agricultural losses of $20 million in a single growing season (Duffus et al., 1986; Wisler et al., 1998). In recent years the incidence of LIYV has declined dramatically.
All commercial cultivars of lettuce, and of various cucurbits, including squash, cantaloupes, watermelons and other melons, are susceptible to LIYV. There is no good genetic resistance developed for these crop hosts.
Lactuca sativa (lettuce, cv. Summer Bibb) plants are very good for diagnosis. Young plants (2-3 cm) are very susceptible to LIYV and are also favoured by the B. tabaci vector. Symptom development is very consistent: the plants show initial interveinal chlorotic spots and distinct yellowing symptoms on older leaves 14-20 days post-inoculation. By 25-30 days post-inoculation, the interveinal areas of older leaves become completely yellow-white while the veins remain green (Fig.1). These symptoms can sometimes be confused with those caused by nutritional disorders, or by other whitefly-transmitted viruses also within the family Closteroviridae, or by BWYV.
Chenopodium murale. Yellowing of older leaves 20-24 days post-inoculation (Fig.2).
Nicotiana benthamiana. Yellowing on older leaves, but often only 4-6 weeks post-inoculation.
Nicotiana clevelandii. Characteristic and distinct yellowing on older leaves starting about 3 weeks post-inoculation.
Symptoms on the above plants are helpful for diagnosis but are not sufficient on their own; they must be supported by evidence of transmission by B. tabaci, and/or by the results of molecular diagnostic tests.
Propagation of LIYV requires the use of plants that are good hosts for both LIYV and its vector, B. tabaci. Both L. sativa and C. murale are good propagation hosts, although LIYV-infected lettuce plants cannot be easily maintained for longer than about 2.5 months, whereas infected C. murale plants can be kept for 4-5 months. C. murale and N. clevelandii (but not lettuce) are useful for producing quantities of plant material sufficient for virion and/or ds-RNA purification. Additional plants suitable for propagating and maintaining LIYV for longer periods of time include sugarbeets (Beta vulgaris) and Malva parviflora. B. tabaci readily acquires LIYV from these species.
Lactuca sativa cv. Summer Bibb. A good host for studies with the whitefly vector.
At the present time the genus Crinivirus includes LIYV and six other definitive members as well as several tentative members (Martelli et al., 2000). All viruses in this genus are whitefly-transmitted. LIYV is the only member for which the complete nucleotide sequences of the genomic RNAs have been determined (Klaassen et al., 1995). However, sequence data obtained so far for other members of this genus suggest that they have genomic organizations similar to that of LIYV.
Nucleic acid: The genome is composed of two species of positive-sense ssRNA: RNA-1 (8,118 nucleotides) and RNA-2 (7,193 nucleotides) (accession numbers U15440 and U15441 respectively). The 5' terminus of each genomic RNA is probably capped but the 3' termini contain no obvious structural components. There are two regions of homology between the two genomic RNAs: (i) the first 5 nucleotides at the 5' terminus of each genomic segment are the same (5'-GGUAA); (ii) each RNA has an identical 23-nucleotide sequence which occurs at about 97 (RNA-1) or 136 (RNA-2) nucleotides from the 5' terminus (Klaassen et al., 1995). The virion RNAs are infective when inoculated to Nicotiana spp. protoplasts (Klaassen et al., 1996).
Protein: The virions are composed of two structural proteins: the major capsid protein (CP; Mr 27,800) and the diverged duplicate capsid protein or minor capsid protein (CPm; Mr 52,300). The CP constitutes approximately 90% of the total virion protein, the remaining 10% consisting of the CPm, which is localized to only one end of the virion (Fig.4). Current evidence suggests that CPm is a whitefly transmission determinant (Tian et al., 1999). Purified virions also have associated with them at least two other LIYV-encoded proteins, the HSP70 homologue and the p59 (Tian et al., 1999). The significance of the association of these proteins with LIYV virions is unknown.
The LIYV bipartite genome is large for plant viruses, having a total size of 15,311 nucleotides. The complete nucleotide sequences of the two LIYV genomic RNAs have been determined and sequence analyses have allowed identification of ORFs as well as predictions for functions of some of the encoded proteins.
RNA-1 encodes for only three ORFs (Fig.5; Klaassen et al., 1995). The first, ORF 1A, begins at nucleotide 98 and can encode a protein of Mr 217,254. This protein contains a predicted papain-like protease as well as characteristic methyl transferase (MTR) and helicase (HEL) motifs. ORF 1B overlaps ORF 1A, and its encoded protein contains characteristic RNA-dependent RNA polymerase (RdRp) motifs. ORFs 1A and 1B are probably translated together via a +1 ribosomal frameshift, which occurs in the overlapping region. RNA-1 ORF 2 overlaps slightly ORF 1B and encodes a protein of Mr ca. 32,000. Computer sequence comparisons showed no similar proteins in existing databases and the function of this protein is unknown.
RNA-2 contains seven ORFs (Fig.5; Klaassen et al., 1995). After a 326-nucleotide untranslated region, RNA-2 ORF 1 encodes a small, hydrophobic protein (Mr 4,600) of unknown function. RNA-2 ORF 2 encodes for the heat-shock protein 70 homologue (Mr 62,300). ORF 3 encodes for a protein of Mr 59,200 of unknown function. ORF 4 encodes for a small Mr 9,000 protein, and ORF 5 encodes for the Mr 27,800 CP. ORF 6 encodes for the Mr 52,300 CPm and ORF 7 encodes for a Mr 26,000 protein of unknown function. The RNA-2 ORFs 1, 2, 3, 5 and 6 compose the five ORF hallmark gene array which is characteristic of all viruses found in the family Closteroviridae (Dolja et al., 1994; Martelli et al., 2000).
The above characteristics suggest that RNA-1 encodes replication-associated proteins whereas RNA-2 encodes for proteins involved in other functions (i.e. movement in planta, encapsidation, vector transmission, etc.). Indeed, LIYV RNA-1 is competent alone for replication in protoplasts whereas RNA-2 is dependent on co-infection with RNA-1 (Klaassen et al., 1996).
With the exception of RNA-1 ORF 1B (described above), other internal ORFs appear to be expressed during infection via subgenomic RNAs. Double-stranded RNAs also can be readily isolated from LIYV-infected plants, and northern hybridization analysis using specific cloned cDNA probes corresponding to each LIYV ORF has allowed mapping of the ds- and subgenomic RNAs. However, these analyses also showed that LIYV-infected plants contain abundant defective RNAs (D-RNAs). So far D-RNAs have been identified only for LIYV RNA-2.
LIYV infections are phloem-limited within plant hosts, and phloem cells contain virion aggregates, typical closterovirus vesiculated inclusion bodies and plasmalemma deposits (Hoeffert et al., 1988; Pinto et al., 1988). All these types of inclusion body are formed when mesophyll protoplasts are inoculated with LIYV (Medina et al., 1998). However, when protoplasts are inoculated with LIYV RNA-1 alone, only the characteristic vesiculated inclusion bodies are formed. The virion aggregates and plasmalemma deposits are found only in protoplasts infected by both RNA-1 and RNA-2 (Medina et al., 1998). Plasmalemma deposits are not common for other plant viruses in the family Closteroviridae and their functions in LIYV are unknown. Within plants, plasmalemma deposits are often found associated with pit fields. In plants and protoplasts, plasmalemma deposits are often associated with masses of virions oriented perpendicular to the plasmalemma; this has led to speculation (Pinto et al., 1988) that these structures may be involved in trafficking LIYV virions within plant cells.
Although epidemic and of considerable importance in southwestern USA in the early 1980's, LIYV is no longer economically important in the USA. During the early 1980's LIYV was associated with high populations of B. tabaci. B. tabaci populations increased to high densities on summer crops such as cotton (a non-host for LIYV), and on indigenous weeds. As cotton was harvested in late summer, B. tabaci moved from cotton to newly emerging crop hosts (including autumn-planted melons and lettuce) and weed hosts. Whitefly populations continued to increase on melons, which also were susceptible to LIYV. This led to tremendous populations of B. tabaci, and LIYV infections were extremely high in susceptible crops such as lettuce. In theory, control could be achieved by elimination of late summer cotton or possibly autumn melons so as to prevent the large population increases of B. tabaci and subsequent epidemic spread of LIYV. This was not done but in the early 1990's a shift occurred in the whitefly population. B. tabaci was largely displaced by B. argentifolii, the latter being a relatively poor vector of LIYV. LIYV is no longer a significant economic problem in southwestern USA.
Differentiation of LIYV from other viruses in the genus Crinivirus can be difficult if symptoms on affected plants are the primary diagnostic approach. The best way of identifying LIYV is to use nucleic acid-based diagnostic methods such as RNA hybridization or RT-PCR. Specific probes and nucleotide sequences for designing RT-PCR primers are available for LIYV and many of the viruses in the genus Crinivirus (e.g. Tian et al., 1996). These are making it much easier to work with LIYV and related viruses.
Leaves from lettuce (Lactuca sativa) plants showing typical symptoms of LIYV infection. Note interveinal yellowing and green veins.
Leaves from Chenopodium murale plants showing typical symptoms of LIYV infection. Note interveinal yellowing and green veins.
The LIYV whitefly vector, Bemisia tabaci. Note winged adults and nymphs on leaf surface.
Immunogold labelling of LIYV virions, treated with (A), antiserum to the CP; (B) and (C), antiserum to the CPm; (D) pre-immune antiserum. Labelling was done with 10-nm gold particles conjugated with goat anti-rabbit antibodies to (A), CP; (B) and (C), CPm. Arrow in (A) indicates a virion terminal region not coated with CP antiserum. Arrows in (B) and (C) indicate virion termini coated with CPm antiserum. Bars represent 224 nm. Reprinted with the permission of the Society for General Microbiology from Tian et al. (1999).
Schematic representation of the LIYV genome. Rectangles represent ORFs. P-PRO, papain-like protease; MTR, methyltransferase; HEL, RNA helicase; RDRP, RNA-dependent RNA polymerase; HSP70, homologue of HSP70 proteins; CP, capsid protein; and CPm, diverged duplicate of the capsid protein (minor capsid protein). Proteins of unknown function are indicated by their respective molecular weight × 10-3 (i.e. P32 = a protein of unknown function of Mr 32,000).