Virus Diseases of Lettuce in Brazil

® Plant Viruses ©2008 Global Science Books Virus Diseases of Lettuce in Brazil Marcelo Agenor Pavan1 • Renate Krause-Sakate1 • Norberto da Silva1 • Francisco Murilo Zerbini2 • Olivier Le Gall3* 1 Departamento de Produção Vegetal, FCA-UNESP, Rua José Barbosa de Barros, 1780, CEP: 18610-307, Botucatu, SP, Brazil 2 Departamento de Fitopatologia/BIOAGRO, Universidade Federal de Viçosa, Viçosa, Brazil 3 Virologie Végétale, UMR GDPP INRA-Bordeaux 2, IBVM, INRA, BP81, F-33883 Villenave d’Ornon Cedex, France Corresponding author: * mapavan@fca.unesp.br ABSTRACT Several viruses have been reported to infect lettuce. The most important is Lettuce mosaic virus (LMV), a potyvirus found worldwide transmitted by seeds and aphids, in a non-persistent manner. LMV causes quite variable symptoms, including mosaic, dwarfing, failure to form proper heads, and sometimes necrotic reactions. Cultivars carrying the mo11 and mo12 genes have resistance to the common strains, although most (mo breaking seed transmitted) strains can overcome this resistance. At least three species of tospovirus, including Tomato spotted wilt virus (TSWV), Tomato chlorotic spot virus (TCSV) and Groundnut ringspot virus (GRSV) causes significant losses, especially during summer, in which high populations of thrips vectors can be found in the field. Tospoviruses causes systemic necrosis and plant death. During the cooler season (May to September), two viruses have been found associated with big-vein disease, Mirafiori lettuce big-vein virus (MLBVV) which belongs to the genus Ophiovirus and Lettuce big-vein associated virus (LBVaV; genus Varicosavirus). LBVaV and MLBVV are both transmitted by the soil-borne fungus Olpidium brassicae. Lettuce mottle virus (LeMoV, genus Sequivirus); Cucumber mosaic virus (CMV, genus Cucumovirus), and Bidens mosaic virus (BiMV, genus Potyvirus) are also found to cause mosaic symptoms on lettuce, although their incidence in lettuce fields is low throughout the year. Epidemiological aspects, variability of viruses, methods of control, genetic variability for lettuce resistance and breeding programs will be discussed. _____________________________________________________________________________________________________________ Keywords: Bidens mosaic virus, Cucumber mosaic virus, Lettuce big-vein associated virus, Lettuce mosaic virus, Lettuce mottle virus, Mirafiori lettuce big-vein virus, tospovirus CONTENTS INTRODUCTION........................................................................................................................................................................................ 35 LETTUCE VIRUSES .................................................................................................................................................................................. 35 Lettuce mosaic virus – LMV ................................................................................................................................................................... 35 Big-vein disease....................................................................................................................................................................................... 37 Lettuce-infecting Tospovirus species ....................................................................................................................................................... 37 Lettuce viruses of minor importance in Brazil......................................................................................................................................... 38 CONCLUSIONS.......................................................................................................................................................................................... 40 REFERENCES............................................................................................................................................................................................. 40 _____________________________________________________________________________________________________________ INTRODUCTION Lettuce (Lactuca sativa L) was domesticated in the Mediterranean area of Europe and was introduced in Brazil by the Portuguese by 1650. Lettuce is cultivated worldwide and is one of the most important vegetables consumed in the world (FAOSTAT, http:www.fao.org). Nowadays it is the most consumed leaf plant in Brazil and cultivated over about 30,000 hectares, the largest producer being the state of São Paulo (Agrianual 2004). In Brazil lettuce is cultivated in open fields, in hydroponic systems, in greenhouses or as organic cultures, typically on small family-owned periurban areas in green belts. Brazilian lettuce cultivation occurs all year long and plants are therefore permanently exposed to phytopathogen attack. Of these the viruses are of particular importance because they are difficult to control and their vectors are present year around. Furthermore, viruses directly affect the quality of the leaves, preventing sale of affected plants. Depending on the environmental conditions and on the degree of care given to the culture, viruses can be responsible for the loss of up to 100% of lettuce crops (Resende and Cupertino 1995). Even under hydroponic cultivation viruses have Received: 22 July, 2008. Accepted: 25 November, 2008. been identified in different producing regions. In this article the main types of virus occurring in Brazil, as well as issues related to diagnosis, control and strain variability are described. LETTUCE VIRUSES Lettuce mosaic virus – LMV Lettuce mosaic virus (LMV) is one of the most important pathogens of lettuce (Lactuca sativa) worldwide (Dinant and Lot 1992). LMV belongs to the genus Potyvirus within the family Potyviridae. The genomic organization of LMV is typical of potyviruses, with a single positive-sense genomic RNA of 10,080 nucleotides (nt) encapsidated as flexuous rods (Revers et al. 1997). The genome organization of LMV is typical of potyviruses, with a single positive-sense, single-stranded genomic RNA of 10,080 nucleotides (nt) encapsidated by coat protein subunits as flexuous rods (Revers et al. 1997). The viral genomic RNA has a virus encoded protein linked covalently at its 5 end, a poly-A tail at its 3 end, and contains a single open reading frame (ORF) which encodes a large polyprotein with 3255 amino acids Invited Mini-Review Plant Viruses 2 (1), 35-41 ©2008 Global Science Books (Costa and Duffus 1958; McLean and Kinsey 1963; Zerbini et al. 1995, 1997). Detection of LMV in infected plants or in seed lots is routinely carried out using immunological techniques such as ELISA (Clark and Adams 1977; Falk and Purcifull 1983) or radioimmunosorbent assay (Ghabrial and Sheperd 1982). More recently, efforts have been made to develop more sensitive techniques for the detection of LMV based on the polymerase chain reaction (PCR) (Revers et al. 1999; Peypelut et al. 2004). Degenerate primer pair 08894p (5-CCG TACATAGCIGARTGTGCT-3) and 09171m (5-GCGTTG ATGTCGTCATCYTT-3) that amplifies a fragment of 278 nucleotide size covering part of the CP region of LMV can be used for general detection of LMV isolates (Revers et al. 1997a; Krause-Sakate et al. 2002; Peypelut et al. 2004) and primer pair Most5930p (5-GATGGGGGTATTTTCGAT-3) and Most6544m (5-GACAAGATAAGCTCTAATTCCAC3) can be used for the specific detection of Most isolates (Peypelut et al. 2004). The control of lettuce mosaic relies on prophylactic measures such as the elimination of contaminated commercial seed lots and on genetic resistance (Dinand and Lot 1992; Ryder 1970). Two alleles of the recessive gene mo1 (mo1¹, formerly named g, and mo1², formerly named mo) were introgressed into different lettuce cultivars, conferring either tolerance (systemic virus accumulation but no symptoms) or resistance (no systemic virus accumulation), depending on the virus isolate considered. The mo1 alleles from resistant and susceptible lettuce cultivars were isolated recently and shown to encode the cap-binding protein, eIF4E (Nicaise et al. 2003). LMV isolates capable of overcoming the resistance afforded by mo1 have been described in various parts of the world, including Europe (Dinant and Lot 1992; Pink et al. 1992a, 1992b; Revers et al. 1997a; Varveri et al. 2002), South America (Stangarlin et al. 2000) and North Africa (Fakhfakh et al. 2001). While generally resistance-breaking isolates generally are not seed-borne, limiting their economic significance to local outbreaks, these newly observed LMV isolates combine resistance breaking and efficient seed transmission in resistant hosts. In a study of the genetic diversity within LMV isolates collected on a worldwide scale, such isolates clustered separately, suggesting a monophyletic origin of this group of isolates for which the name LMV-Most (for mo1-breaking, seed transmitted) was proposed (Krause-Sakate et al. 2002). Similarly, the name LMV-Common was proposed for another monophyletic group corresponding to the seed-borne isolates that are unable to causes symptoms on mo1 plants. The mo1 alleles also provide control of seed transmission for the LMVCommon isolates because, even in the tolerance cases, these isolates accumulate in the mother plants containing mo1¹ or mo1², but do not access the embryo. Most-type isolates should be considered an increasing threat to lettuce production worldwide, because of their ability to spread in seed lots even in the presence of the two available LMV resistance genes. To evaluate the occurrence of these two types of LMV isolates, a survey was carried out during 2002-2005 in three lettuce production areas of São Paulo State, on susceptible cultivars, LMV-Common isolates were prevalent (77.3% of the plants evaluated) and LMV-Most isolates were found frequently associated with tolerant (mo11) lettuce cultivars. Susceptible cultivars are grown today in most of the lettuce production areas in São Paulo State. So, despite the ability of LMV-Most isolates to overcome the resistance provided by the recessive mo11 gene, they are not prevalent in our conditions (Firmino et al. 2008). The comparison of LMV-AF-199 (Most) and LMVAF198 (Common) on susceptible cultivars reveals that the Most strain reduces drastically the fresh weight, leaf area and chlorophyll content, on the White Boston (susceptible) and Elisa (mo11) cultivars (Jadão et al. 2003). At least one naturally recombinant isolate between LMV-Most and LMV-Common was identified in Tunisia (Krause-Sakate et al. 2004). Fig. 1 Elisa lettuce cultivar infected by Lettuce mosaic virus (right). Healthy plant (left). (Revers et al. 1997). Disease symptoms are quite variable and depend on the particular isolate–host plant combination but frequently include dwarfing, failure to form proper heads, leaf distortion, leaf mosaic or mottling, vein clearing and sometimes necrosis (Candresse et al. 2007). Lettuce plants may fail to ‘heart’ and inner leaves remain dwarfed and rosetted (Fig. 1). Infected plants like Chenopodium amaranticolor develop pale green or chlorotic local lesions (usually with reddish margins) after 8-10 days and systemic yellow veinal flecks or yellow netting of the younger leaves especially in winter. C. quinoa is more sensitive than C. amaranticolor and local lesions more numerous but without reddish margins followed by systemic yellow vein-net symptoms with twisting and stunting of apical leaves can be observed. Gomphrena globosa develop whitish, local necrotic dots (47 days) enlarging into red-rimmed lesions (http://www. dpvweb.net/dpv/showdpv.php?dpvno=399). As with other potyviruses, LMV is transmitted efficiently by aphids in a nonpersistent manner (Tomlinson 1970), notably by Myzus persicae, Macrosiphum euphorbiae and Acyrthosiphon scariolae barri. Non-vectors include Nasonovia ribisnigri. All instars of M. persicae transmit but alates are less efficient than apterae. Transmission efficiency increases with increasing periods of fasting (5-240 min), but decreases with increasing acquisition access time from 5 to 120 min (Sylvester 1955). Rapid epidemics can develop in susceptible lettuce cultivars, leading to losses of up to 80-100% (Dinant and Lot 1992). In addition, a number of LMV isolates can also be transmitted through infected seeds (Grogan et al. 1952; Tomlinson 1970). Seed transmission is probably the major factor in the spread of the disease (Broadbent et al. 1951; Grogan et al. 1952; Tomlinson 1962). Spread occurs (a) from seedlings infected through the seed and (b) from neighboring infected lettuce. The disease can be controlled by ensuring that the crop is isolated from external sources of virus and that less than 0.1% of the seed carries the virus (Zink et al. 1956; Tomlinson 1962; Grogan 1980). Even where adjacent crops are infected, use of mosaic-free seed provides some control (Tomlinson 1962). LMV transmission rate varies among genotypes as a function of LMV variability. This rate depends on the time the receptor plant has been infected as well as on the genotype and on environmental conditions. The highest levels are observed when the plant grows in mild temperatures. Studies related to the seed transmission of LMV showed transmission rates of 1.9 and 16.5%, respectively in tolerant and susceptible lettuce cultivars (Jadão et al. 2002). LMV-susceptible species occur in 20 genera (9 genera of Compositae) in 10 families. The virus is transmissible by inoculation with sap from young infected plants, but transmission with sap from old leaves may be difficult. Weeds and ornamental plants can serve as reservoirs of LMV 36 Virus diseases of lettuce in Brazil. Pavan et al. ting spores of the fungus can persist for over 20 years in soil and can retain the ability to transmit the virus for over 15 years (Campbell 1996). Both LBVaV and MLBVV are sap transmitted, but with low efficiency (Lot et al. 2002; Sanches et al. 2008). LBVaV are transmitted to C. quinoa, C. amaranthicolor, Nicotiana benthamiana, N. clevelandii and N. occidentalis (Hujberts 1990). MLBVV is transmitted to C. quinoa, N. benthamiana, N. tabacum White Burley, N. occidentalis (Roggero et al. 2000), N. clevelandii (Lot et al. 2002) and N. hesperis (van der Wilk et al. 2002). The natural infection of Sonchus oleraceus by both viruses was also observed by Navarro et al. (2005) indicating that this weed acts as a reservoir for the viruses and for O. brassicae. LBVaV and MLBVV detection can be carried out with virus-specific antiserum (Lot et al. 2002; Colariccio et al. 2003; Roggero et al. 2003; Colariccio et al. 2005), as well as by RT-PCR, where the primers described by Rosales et al. (2004) or Navarro et al. (2004) can be used. Navarro et al. (2004) observed that both LBVaV and MLBVV show greatest concentration on the older roots and leaves of lettuce plants and that symptoms begin 40-50 days after transplanting, coinciding with the peak of spore production of vector Olpidium brassicae. Previous studies have found that plants exhibiting big vein symptoms were frequently co infected with both viruses, suggesting that LBVaV may also contribute to the disease symptoms (Roggero et al. 2003; Navarro et al. 2004, 2005b). The presence of symptomatic lettuce plants, MLBVV-negative, but LBVaV-positive by ELISA, also has been observed in a field survey (Roggero et al. 2003). LBVaV and MLBVV are widespread in the important lettuce-producing areas of São Paulo State and most plants are co-infected with LBVaV and MLBVV (Sanches et al. 2008). Similar results were reported by Roggero et al. (2003), Navarro et al. (2004) and Hayes et al. (2006), showing a strong correlation between big vein symptom expression and MLBVV presence, and indicating that coinfection with both MLBVV and LBVaV could be due to the shared vector (Lot et al. 2002). In Brazil during summer plants were found infected alone or in combination by both MLBVV and LBVaV, but the plants did not show any symptoms of the disease (Sanches et al. 2007). The amino acid identities in the coat protein (CP) gene between brazilian LBVaV isolates and other sequences deposited in the Genbank is higher than 93% and no correlation of the geographic origin can be made. For the MLBVV isolates, the aminoacid identities is higher tham 91%, and MLBVaV brazilian isolates belong to the Subgroup A (Sanches et al. 2008) according to Navarro et al. (2005) classification, that implies the presence of an RsaI restriction site in the CP gene. The subgroup B consists only of Spanish isolates (ALM1, ALM4, ALM5 and SON3) previously described by Navarro et al. (2005). These results supports the suggestion of Hayes et al. (2006) that research on big vein disease in the United States, Europe and Japan will likely be relevant for lettuce production in others areas as well, due to nucleotide sequence conservation among lettuce isolates of these pathogens, including countries in South America. For an effective disease control several cultural measures like irrigation, to avoid fungal zoospore dissemination, the use of healthy shoots, elimination of alternative hosts, soil solarization to reduce fungal population and sowing with plastic films to reduce soil humidity and increase temperature therefore reducing zoospore activity, are recommended (Jones 2003; Lathan and Jones 2004). Fletcher et al. (2005) recommends the use of products like carbendazim, propamocarb or thiabendazole to improve lettuce production in high incidence areas. Culture rotation is indicated specially during winter, the season when the disease is most prevalent in lettuce. No tolerant or resistant varieties exist in Brazil. Among the many control measures that should be adopted one may mention the use of virus-free seeds, vector aphid control in order to keep insect population low in the fields and elimination of alternative hosts. Also, should the virus incidence be low, it is recommended to remove and burn infected plants as well as to use quality shoots produced under screen-surrounded by aphid-free environments. Big-vein disease Lettuce big vein disease was first described in California (Jagger and Chandler 1934), and is a soil-borne disease found worldwide (Roggero et al. 2003). In Brazil the disease was reported in 2003 and observed during the cooler season (Colariccio et al. 2003; Lima Neto et al. 2004). The name of the disease refers to the appearance of chlorotic areas surrounding the vascular tissue that confers the aspect of an anomalous vein enlargement (Fig. 2), commonly accompanied by severe leaf deformations and growth reducetions. The economic importance of the disease is a result of the unsightliness of the lettuce foliage, which reduces market value, due to delayed head formation, decreased head size and a reduced proportion of harvestable plants (Zink and Grogan 1954). The symptom expression of big-vein disease is dependent on local factors such as low temperature, luminosity and soil condition (Walsh 1994). This disease is more common when temperatures remain below 20°C. In Brazil symptoms are observed mainly in winter when daylight temperatures range from 18 to 22°C and night temperatures from 10 to 16°C (Colariccio et al. 2003). The big vein symptoms were historically attributed to Lettuce big-vein associated virus (LBVaV; genus Varicosavirus), formerly known as Lettuce big-vein virus (LBVV), but a causative relationship was never confirmed (Kuwata et al. 1983; Vetten et al. 1987). According to Roggero et al. (2000), the Mirafiori lettuce big-vein virus (MLBVV), formerly known as Mirafiori lettuce virus (MiLV), which belongs to the genus Ophiovirus, was reported to be the causal agent of big-vein disease. Symptoms occur in plants after they have been inoculated mechanically with MLBVV or by the vector (Lot et al. 2002). LBVaV and MLBVV have a segmented genome of ssRNA and virus particles contain RNA molecules of both polarities. Negative-sense RNAs predominate for LBVaV while MLBVV contain nearly equimolar amounts of RNA molecules of both polarities (Sasaya et al. 2001, 2002; van der Wilk et al. 2002; Sasaya et al. 2004) LBVaV and MLBVV are both transmitted by the soilborne fungus Olpidium brassicae (Lot et al. 2002). The res- Lettuce-infecting Tospovirus species Fig. 2 Symptoms exhibited by Lettuce big-vein associated virus and Mirafiori lettuce big-vein virus. Tospovirus is the only plant-infecting genus in the family 37 Plant Viruses 2 (1), 35-41 ©2008 Global Science Books ral vegetable crops. Assay plants like Nicotiana glutinosa, N. rustica, Gomphrena globosa and Tropaeolum majus can be used to propagate the virus. Local lesions in Petunia hybrida, chlorotic and necrotics lesion in N. glutinosa and necrotic lesions in Datura stramonium can indicate the presence of tospoviruses (Costa and Forster 1942). TSWV can infect 550 species of plants, including monocots and dicots (de Ávila 1992). The following tospovirus species have been reported in Brazil: Tomato spotted wilt virus (TSWV), Tomato chlorotic spot virus (TCSV), Groundnut ringspot virus (GRSV), Chrysanthemum stem necrosis virus (CSNV), Zucchini lethal chlorotic virus (ZLCV), and Iris yellow spot virus (IYSV) (Pozzer et al. 1999). In lettuce, only TSWV, TCSV and GRSV were observed (Chaves et al. 2001). TCSV is the main tospovirus in the state of São Paulo where it is harmful to different crops, but especially to vegetables, while GRSV is prevalent in lettuce in the São Francisco River Valley, in the state of Pernambuco (Colariccio et al. 2001). TCSV is also found in hydroponically-grown lettuce in São Paulo State (Colariccio et al. 2004). Both TCSV and GRSV are efficiently transmitted by the thrips species F. occidentalis Pergande and F. schultzei (Wijkamp et al. 1995; Borbon and Garcia 1996), which are prevalent in tropical and subtropical regions (Wijkamp et al. 1995). Breeding cultivars for high resistance levels seems to be the best control strategy, since thrips control has not been efficient. In this context, cultures susceptible to tospoviruses, including vegetables and ornamentals, represent important sources for spreading these viruses. Eradication of weeds and volunteer growth close to lettuce fields, in association with other cultural practices, could minimize and prevent infection by tospoviruses. Before sowing the following measures should be adopted: culture rotation with non-susceptible plants, sowing in sites without adjacent susceptible crops and alternate vector and virus host control. During sowing, virus-free shoots should be used, insecticide should be regularly applied (both in the field and in shoot storage areas), trap-plants like broccoli, cauliflower and tolerant wild tomato species, which blossom intensely attracting thrips, be set up and corn barriers around the crop area to hinder vector insect migration and reduce cultivation operations that avoid the motion of thrips from infected to healthy plants, be erected. After harvesting, it is recommended to let the areas with high disease incidence rest for 3 to 4 weeks, and to treat soil (fumigation) to eliminate thrips associated with harvest left-overs. Soil handling is not totally effective if virus and vector have high incidence all over the area. In those conditions sowing should be avoided. Community cooperation is important for thrips control. TSWV resistance has been observed in Tinto and PI 342517 (“Ancora”) cultivars and such resistance is of partial dominance. Research towards resistance transfer and selection of plants more adequate to the local conditions are being carried out by Norberto da Silva (unpublished). Fig. 3 Tospovirus disease observed on Elisa lettuce cultivar. Bunyaviridae, a large group of enveloped, mostly arthropod-transmitted, animal-infecting viruses with tripartite negative-stranded ssRNA genomes (Nichol et al. 2005). TSWV is the type species and has a genome consisting of three negative or ambisense ssRNAs designated S (2.9 kb), M (4.8 kb), and L (8.9 kb) (Nichol et al. 2005). The RNAs may form a panhandle conformation created by base pairing of about 60 complementary nucleotides at the 3 and 5 ends of each strand (De Haan 1989). The core of the virion contains ribonucleoproteins (RNPs) composed of the ssRNA components encapsidated by the nucleoprotein (N) and a few copies of the viral RNA-dependent RNA polymerase (RdRp or L protein). The 80-120 nm pleiomorphic virus particles are formed by enclosure of the RNPs in a host derived lipid membrane studded with surface projections composed of two viral glycoproteins, GN and GC (Nichol et al. 2005). At least 10 species of insects in the order Thysanoptera (commonly known as thrips) transmit viruses in the genus Tospovirus (Whitfield et al. 2005). Tospoviruses are transmitted in a persistent propagative fashion and are transstadially passed on their insect vector. Thrips eggs are oviposited into plant tissue and within a few days the first instar larvae emerge. Virus acquisition occurs solely during the larval stages after which the virus is passed transstadially to the adult. The pupal stages are non-feeding and do not move, although they do maintain virus infection. In nature, Frankliniella occidentalis pupates in the soil. Many other vector species, e.g., Thrips tabaci, pupate in the foliage. Adults emerge and have a tendency to disperse widely. Only adult thrips (male and female) that acquired the virus during their larval stages can transmit tospoviruses (Whitfield et al. 2005). F. occidentalis, F. schultzei, T.s tabaci and T. palmi are the main vetors species of tospovirus in Brazil (Nagata and Inoue-Nagata 2003). In Brazil the first report of a disease caused by a virus of the genus Tospovirus was made by Costa and Forster (1938). The lettuce is a natural host of tospoviruses in the field (Costa and Forster 1942). Since 1986 severe losses are related to tospoviruses in lettuce. In the summer, (December to March) losses may occur from 30 to 100% in field conditions (Moraes et al. 1986) or 40% under hydroponic cultivation (Colariccio et al. 2004). Symptoms observed in lettuce are circular necrotic stains and browning of the leaves. With systemic infection the plant usually falls to one side. Eventually it becomes completely necrotic and death of the plant occurs (Fig. 3). The identification of species in the genus Tospovirus is made by examining host range, serology, and according to the divergence of amino acids in the nucleoprotein (N Protein) (de Ávila et al. 1993a, 1993b; Pozzer et al. 1999). Such species infect plants in 92 botanical families (van Regenmortel et al. 2000), causing significant losses in seve- Lettuce viruses of minor importance in Brazil Bidens mosaic virus (BiMV) is a tentative species of the genus Potyvirus. It has flexuous rod shaped particles around 720 nm length by 12-13 nm diameter (Kitajima et al. 1961). It was verified naturally infecting lettuce in Brazil by Costa and Kitajima (1966). So far this virus has been described only in Brazil. Besides lettuce, some BiMV isolates also infect tobacco (Nicotiana tabacum Turkish), sunflower (Helianthus annus), Physalis floridana, Chenopodium amaranticolor, C. ambrosioides, broom stick (Bidens pilosa), Cassia occidentalis, Leonotis nepaetifolia (Kitajima et al. 1961), Pisum sativum (Nagata et al. 1995), Zinia elegans, N. tabacum TNN (Hasegawa 2004), Emilia sonchifolia, Acanthospermum hispidum, Amaranthus sp., Solanum nigrum (Kuhn et al. 1980) and Coreopsis lanceolata (Rodrigues et al. 1991). Hasegawa 2006 observed that a BiMV isolate from lettuce was not able to infect sunflowers, B. pilosa or N. tabacum TNN, 38 Virus diseases of lettuce in Brazil. Pavan et al. Fig. 4 Symptoms exhibited by Bidens mosaic virus. hosts therefore considered susceptible for the virus. Symptoms observed in lettuce include mosaic and foliar deformation very similar to those caused by LMV (Fig. 4). The diagnosis of the virus is hindered by the absence of good antisera and of RT-PCR-specific oligonucleotides. The virus is sap transmitted and also by the aphids Myzus persicae, Aphis coreopsidis and Dactynotus sp. In lettuce, transmission tests with Myzus persicae showed results from 20 to 80% depending on the cultivar. The virus is not transmitted by the seed (Kuhn et al. 1980). There is no study in Brazil in the incidence of this virus in field conditions. Economic losses have been observed in sunflowers and peas (Nagata et al. 1995). However the lettuce cultivars in São Paulo appear to be susceptible to at least one BiMV isolate naturally collected from lettuce. Of the tested cultivars only ‘Gizele’ has proved tolerant to the virus (Krause-Sakate, pers. comm.). Control measures for this virus include handling to keep vector insect population low, elimination of infected plants and alternative hosts as well as the use of tolerant varieties. Lettuce mottle virus (LeMoV) is a possible member of the genus Sequivirus, family Sequiviridae (Jadão et al. 2007) infecting lettuce in Brazil (Marinho et al. 1982) and Chile (Krause-Sakate et al. 2005). It is closely related to Dandelion yellow mosaic virus (DaYMV) a sequivirus that infects lettuce in Europe. LeMoV possesses isometric particles 30 nm in diameter, occurs at low concentration in plants and is sap-transmitted but has a narrow host range (Marinho et al. 1982; Jadão et al. 2007). C. quinoa is systemically infected by the virus and some lettuce cultivars like ‘Vanguard-75’ and ‘Elisa’ are tolerant (Jadão et al. 2007). LeMoV is not seed-borne in lettuce and causes mosaic symptoms very similar to LMV (Fig. 5) (Jadão et al. 2007). Transmission is in a semi-persistent manner by Hyperomyzus lactucae (Marinho et al. 1982). Specifics primers for LeMoV (Lmo3 5-ACATGAGCACTAGTGAGG-3 and Lmo4 5-AGATAGAGCCGTCTGGCG-3) can be used for diagnosis by RT-PCR. The incidence of LeMoV in the field is low. A survey was carried out in São Paulo State and in 1,362 samples of lettuce showing mosaic symptoms tested, only 137 (10.05%) were positive for LeMoV. LeMoV can also be found in mixed infections with LMV (KrauseSakate et al. 2008). Lettuce mottle virus (LeMoV) is a possible member of the genus Sequivirus, family Sequiviridae (Jadão et al. 2007) reported infecting lettuce in Brazil (Marinho et al. 1982) and Chili (Krause-Sakate et al. 2005) and closely related to Dandelion yellow mosaic virus (DaYMV) a sequivirus that infects lettuce in Europe. LeMoV possesses iso- Fig. 5 Symptoms exhibited by Lettuce mottle virus. metric particles 30 nm in diameter, occurs at low concentration in plants and is sap transmitted but have a narrow host range (Marinho et al. 1982; Jadão et al. 2007). C. quinoa is systemically infected by the virus and some lettuce cultivars like ‘Vanguard-75’ and ‘Elisa’ are tolerant to the virus (Jadão et al. 2007). LeMoV is not seed-borne in lettuce and causes mosaic symptoms very similar to LMV (Fig. 5) (Jadão et al. 2007). Transmission is in a semi-persistant manner by Hyperomyzus lactucae (Marinho et al. 1982). Specifics primers for LeMoV (Lmo3 5-ACATGAGCACTAGTG AGG-3 and Lmo4 5-AGATAGAGCCGTCTGGCG-3) can be used for diagnosis by RT-PCR. The incidence of LeMoV in the champ is low. A survey was carried out in São Paulo State and for 1,362 samples of lettuce showing mosaic symptoms tested, only 137 (10.05%) were positive for LeMoV. The incidence of LeMoV in the field is low, but its symptoms are very similar to those caused by LMV, while LeMoV is not controlled by the mo1 LMV resistance alleles. A survey was carried out in São Paulo State and out of 1,362 samples of lettuce showing mosaic symptoms tested, only 137 (10.05%) were positive for LeMoV. LeMoV can also be found in mixed infections with LMV (KrauseSakate et al. 2008). Turnip mosaic virus (TuMV, genus Potyvirus) was first reported infecting lettuce in California. In susceptible lettuce cultivars the initial symptoms include abundant, small, light-green lesions, circular to irregular lesions distributed randomly on the leaves (Duffus 1997). Non-persistent transmission can be performed by aphids Myzus persicae and 39 Plant Viruses 2 (1), 35-41 ©2008 Global Science Books Brevicoryne brassicae (Brunt et al. 1996). No data on its incidence in the field exist in Brazil. Cucumber mosaic virus (CMV, genus Cucumovirus) also cause symptoms similar to LMV but is not seed borne like LMV so distribution within the field is usually along margins. The virus is transmitted by several species of aphids including in a non-persistent manner. There is no data of incidence of CMV on lettuce in Brazil, and the disease can be prevented by controlling the vector sources. There is no information of resistance cultivars to CMV. logical and molecular properties. Journal of Plant Pathology 83, 3-11 Falk BW, Purcifull DE (1983) Development and application of an enzymelinked immunosorbent assay (ELISA) test to index lettuce seeds for lettuce mosaic virus in Florida. Plant Disease 67, 413-416 Firmino AC, Krause-Sakate R, Pavan MA, da Silva N, Hanai SM, Anbo RH, Nietzshe T, Le Gall O (2008) Prevalence of Lettuce mosaic virus – common strain on three lettuce producing areas from São Paulo State. Summa Phytopathologica 34, 161-163 Fletcher JD, France CM, Butler RC (2005) Virus surveys of lettuce crops and management of lettuce big-vein disease in New Zealand. New Zealand Plant Protection 58, 239-244 Ghabrial SA, Li D, Sheperd RJ (1982) Radioimmunosorbent assay for detection of lettuce mosaic virus in lettuce seeds. Plant Disease 66, 1037-1040 Grogan RG (1980) Control of lettuce mosaic with virus-free seed. Plant Disease 64, 446-449 Grogan RG, Welch JE, Bardin R (1952) Common lettuce mosaic and its control by the use of mosaic-free seed. Phytopathology 42, 573-578 Hasegawa JM (2006) Identificação e caracterização de isolado de Bidens mosaic virus e triagem de alface para resistência. MSc thesis, Universidade Estadual Paulista Júlio de Mesquita Filho, Botucatu, Brasil, 44 pp Hayes RJ, Wintermantel WM, Nicely PA, Ryder EJ (2006) Host resistance to Mirafiori lettuce big-vein virus and Lettuce big-vein associated virus and virus sequence diversity and frequency in California. Plant Disease 90, 233239 Huijberts N, Blystad DR, Bos L (1990) Lettuce big-bein virus: Mechanical transmission and relationships to tobacco stunt virus. Annals of Applied Biology 116, 463-475 Jadão A S, Pavan MA, da Silva N, Zerbini FM (2002) Transmissão via semente do Lettuce mosaic virus (LMV) patótipos II e IV em diferentes genótipos de alface. Summa Phytopathologica 281, 58-61 Jadão AS, Krause-Sakate R, Liberti D, Pavan MA, Echer MM, SvanellaDumas L, Zerbini FM, Candresse T, LeGall O (2007) Further characterization of two sequiviruses infecting lettuce and development of specific RTPCR primers. Archives of Virology 152, 999-1007 Jadão AS, Pavan MA, Krause-Sakate R, Zerbini FM (2003) Efeitos na fotossíntese e área foliar de cultivares de alface inoculadas mecanicamente com patótipos do Lettuce mosaic virus (LMV) e Lettuce mottle virus (LeMoV). Fitopatologia Brasileira 28, 12-16 Jagger IC, Chandler N (1934) Big Vein, a disease of lettuce. Phytopathology 24, 1253-1256 Jones RAC (2003) Using epidemiological information to develop effective integrated virus disease management strategies. Virus Research 100, 5-30 Kitajima EW, Carvalho AMB, Costa AS (1961) Morfologia do vírus do mosaico do picão. Bragantia 20, 503-512 Krause-Sakate R, Fakhfakh, H, Peypelut M, Pavan MA, Zerbini FM, Marrakchi M, Candresse T, Le Gall (2004) A naturally occurring recombinant isolate of Lettuce mosaic virus. Archives of Virology 149, 191-197 Krause-Sakate R, Firmino AC, Jadão AS, Pavan MA, da Silva N, Hanai SM, Anbo RH, Nietzsche T (2008) Ocorrência generalizada do Lettuce mottle virus em três regiões produtoras de alface do Estado de São Paulo. Summa Phytopathologica 34, 88-89 Krause-Sakate R, Jadão AS, Firmino A C, Pavan MA, Zerbini FM, Rosales MI, Bustamente P, Le Gall O (2005) First report of a lettuce infecting sequivirus in Chile. Plant Disease 89, 1129 Krause-Sakate R, Le Gall O, Fakhfakh H, Peypelut M, Marrakchi M, Varveri C, Pavan MA, Souche S, Lot H, Zerbini FM, Candresse T (2002) Molecular characterization of Lettuce mosaic virus field isolates reveals a distinct and widespread type of resistance-breaking isolate: LMV-Most. Phytopathology 92, 563-572 Kuhn CB, Lin MT, Costa CL (1980) Transmissão, círculo de hospedeiros e sintomatologia do vírus do mosaico do picão. Fitopatologia Brasileira 5, 3950 Kuwata S, Kubo S, Yamashita S, Doi Y (1983) Rod-shaped particles, a probable entity of Lettuce big vein virus. Annals of the Phytopathological Society of Japan 49, 246-251 Latham LJ, Jones RAC (2004) Deploying partially resistant genotypes and plastic mulch on the soil surface to suppress spread of lettuce big-vein disease in lettuce. Australian Journal of Agricultural Research 55, 131-138 Lima Neto VC, Colariccio A, Chaves ALR, Strapasson M, Chagas CM, Roggero P (2004) Ocorrência de Varicosavirus e Ophiovirus associados ao espessamento clorótico das nervuras da alface no estado do Paraná. XXVII Congresso Paulista de Fitopatologia, Brasil, p 30 Lot H, Campbell RN, Souche S, Milne RG, Roggero P (2002) Transmission by Olpidium brassicae of Mirafiori lettuce virus and Lettuce big-vein virus, and their roles in Lettuce Big-Vein etiology. Phytopathology 92, 288-293 Marinho VL, Kitajima EW, Lin MT, Costa CL (1982) Caracterização do vírus do mosqueado da alface. Fitopatologia Brasileira 7, 543 Moraes GJ, Wanderly LJ, Costa AS (1986) Surto de viracabeça na cultura de alface em Pernambuco. Horticultura Brasileira 6, 24-25 Nagata T, Inoe-Nagata AK (2003) Vetores de vírus. Revista Cultivar Hortaliças e Frutas 17, 34-36 Nagata T, Inoue-Nagata AK, Dusi A, Kitajima EW (1995) Bidens Mosaic potyvirus newly isolated from pea, its characteristics and serological relation- CONCLUSIONS Lettuce viruses are one of the great current challenges to its culture since no tolerant or resistant varieties are available commercially for most viruses herein described in this review. Improvement programs towards multiple viral resistance cultivars are in progress at FCA/UNESP/Botucatu, using sources tolerant to tospoviruses, Lettuce mosaic virus, LMV common and most strains, LeMoVLettuce mottle virus and Bidens mosaic virusBiMV in different lettuce groups. This genetic research has made it possible in 2007 the introduction of the cultivar ‘Cuesta’ with that carries multiple resistances resistance genes to tospoviruses, LMV and LeMoV. REFERENCES Agrianual (2004) Anuário da Agricultura Brasileira. Argos Comunicações, São Paulo, 496 pp Berger PH, Adams MJ, Barnett OW, Brunt AA, Hammond J, Hill JH, Jordan RL, Kashiwazaki S, Rybicki E, Spence N, Stenger DC, Ohki ST, Uyeda I, van Zaayen A, Valkonen J, Vetten HJ (2005) Potyviridae In: Fauquet CM, Mayo MA, Maniloff J, Desselberger U, Ball LA (Eds) Virus Taxonomy, Academic Press, San Diego, California, pp 819-841 Borbon CM, Gracia O (1996) Frankliniella schultzei (Tribom), eficiente vector de Groundnut ringspot virus (GRSV) en cultivo de lechuga en Mendoza, Argentina. XXIX Congresso Brasileiro de Fitopatologia, Brasil, p 423 Brunt AA, Crabtree K, Dallwitz MJ, Gibbs AJ, Watson L, Zurcher EJ (1996) Plant Viruses Online. Aailable online: http://biology.anu.edu.au/ Groups/MES/vide/ Campbell RN (1996) Fungal transmission of plant viruses. Annual Review of Phytopathology 34, 87-108 Candresse T, Lot H, German-Retana S, Krause-Sakate R, Thomas J, Souche S, Delaunay T, Lanneau M, Le Gall O (2007) Analysis of the serological variability of Lettuce mosaic virus using monoclonal antibodies and surface plasmon resonance technology. Journal of General Virology 88, 26052610 Clark MF, Adams AN (1977) Characteristics of the microplate method of enzyme-linked immunosorbent assay for the detection of plant viruses. Journal of General Virology 34, 475-483 Colariccio A, Chaves ALR, Chagas C, Lenzi R, Roggero P (2003) Presence of lettuce big-vein disease and associated viruses in a subtropical area of Brazil. Plant Pathology 52, 792 Colariccio A, Chaves ALR, Eiras M, Chagas CM, Roggero P (2005) Detection of Varicosavirus and Ophiovirus in lettuce associated with lettuce bigvein symptoms in Brazil. Fitopatologia Brasileira 30, 416-419 Colariccio A, Eiras M, Chaves ALR, Harakawa R, Chagas CM (2004) Tomato chlorotic spot virus in hydroponic-grown lettuce in São Paulo state, Brazil. Fitopatologia Brasileira 29, 307-311 Colariccio A, Eiras M, Chaves ALR, Roggero P, Chagas CM (2001) Diversidade de tospovírus em olerícolas no Estado de São Paulo. Summa Phytopathologica 27, 177-182 Costa AS, Kitajima EW (1966) Vírus do mosaico do picão ataca girassol. O Estado de São Paulo. Suplemento Agrícola 13, 3 de Ávila AC, de Haan P, Kormelink R, Resende R de O, Goldbach RW, Peters D (1993a) Classification of tospoviruses based on the phylogeny of nucleoprotein gene sequences. Journal of General Virology 74, 153-159 de Ávila AC, de Haan P, Smeets MLL, Resende R de O, Kitajima EW, Golbach RW, Peters D (1993b) Distinct levels of relationships between tospovirus isolates. Archives of Virology 128, 211-227 De Haan P, Wagemakers L, Peters D, Goldbach R (1989) Molecular cloning and terminal sequence determination of the S and M RNAs of tomato spotted wilt virus. Journal of General Virology 70, 3469-3473 Dewey RA, Semorile LC, Grau O (1996) Detection of Tospovirus species by RT-PCR of the N-gene and restriction enzyme digestions of the products. Journal of Virological Methods 56, 19-26 Dinant S, Lot H (1992) Lettuce mosaic virus. Plant Pathology 41, 528-542 Fakhfakh H, Le Gall O, Candresse T, Marrakchi M (2001) Analysis of the variability of Tunisian isolates of Lettuce mosaic potyvirus (LMV) using bio- 40 Virus diseases of lettuce in Brazil. Pavan et al. Sanches MM, Krause-Sakate R, Pavan MA (2008) Sequence diversity in the coat protein gene of Lettuce big-vein associated virus and Mirafiori lettuce big-vein virus infecting lettuce in Brazil. Summa Phytopathologica 34, 175177 Sanches MM, Krause-Sakate R, Zerbini FM, Rosales M, Le Gall O, Pavan MA (2007) Frequência do Lettuce big-vein associated virus e Mirafiori lettuce big-vein virus em plantas de alface no Estado de São Paulo e transmissão através de extrato vegetal e solo. Summa Phytophatologica 33, 378-382 Sasaya T, Ishikawa K, Koganezawa H (2001) Nucleotide sequence of the coat protein gene of Lettuce big-vein virus. Journal of General Virology 82, 15091515 Sasaya T, Ishikawa K, Kogenazawa H (2002) The nucleotide sequence of RNA1 of Lettuce big-vein virus, genus Varicosavirus, reveals its relation to nonsegmented negative-strand RNA viruses. Virology 297, 289-297 Sasaya T, Kusaba S, Ishikawa K, Koganezawa H (2004) Nucleotide sequence of RNA2 of Lettuce big-vein virus and evidence for a possible transcription termination/initiation strategy similar to that of rhabdoviruses. Journal of General Virology 85, 2709-2717 Stangarlin OS, Pavan MA, da Silva N (2000) Occurrence of a new pathotype of Lettuce mosaic virus on Lettuce in Brazil. Plant Disease 8, 490 Tomlinson JA (1970) Lettuce mosaic virus. No. 9 in: Descriptions of Plant Viruses. Commonwealth Mycological Institute of Associated and Applied Biology, Kew, England Tomlinson JA (1962) Control of lettuce mosaic by the use of healthy seed. Plant Pathology 11, 61-64 Van Der Wilk F, Dullemans AM, Verbeek M, Van Den Heuvel JFJM (2002) Nucleotide sequence and genomic organization of an ophiovirus associated with lettuce big-vein disease. Journal of General Virology 83, 2869-2877 Van Regenmortel MHV, Fauquet CM, Bishop DHL, Carstens EB, Estes MK, Lemon SM, Maniloff J, Mayo MA, McGeoch DJ, Pringle CR, Wickner RB (2000) Virus Taxonomy, Academic Press, San Diego, California, USA, 1162 pp Varveri C, Peypelut M, Le Gall O, Candresse T (2002) Molecular characterization of Greek Lettuce mosaic virus isolates [in Greek]. Hellenic Virology 7, 50-57 Vetten HJ, Lesemann DE, Dalchow J (1987) Electron microscopial and serological detection of virus-like particles associated with lettuce big-vein disease. Journal of Phytopathology 120, 53-59 Walsh JA (1994) Effects of some biotic and abiotic factors on symptom expression of Lettuce big-vein virus in lettuce (Lactuca sativa). Journal of Horticultural Science 69, 21-28 Whitfield AE, Ullman DE, German TL (2005) Tospovirus-thrips interactions. Annual Review of Phytopathology 43, 459-489 Wijkamp I, Almarza N, Golbach R, Peters D (1995) Distinct levels of specificity in thrips transmission of tospoviruses. Phytopathology 85, 1069-1074 Zerbini FM, Koike ST, Gilbertson RL (1997) Gazania spp.: A new host of lettuce mosaic potyvirus, and a potential inoculum source for recent lettuce mosaic outbreaks in the Salinas Valley of California. Plant Disease 81, 641646 Zerbini FM, Koike ST, Gilbertson RL (1995) Biological and molecular characterization of lettuce mosaic potyvirus isolates from the Salinas Valley of California. Phytopathology 85, 746-752 Zink FW, Grogan RG (1954) The interrelated effects of big vein and market price on the yield of head lettuce. Plant Disease Reporter 38, 844-846 Zink FW, Grogan RG, Welch JE (1956) The effect of the percentage of seed transmission upon subsequent spread of lettuce mosaic. Phytopathology 46, 662-664 ship with other potyviruses. Fitopatologia Brasileira 20, 473-478 Navarro JA, Botella F, Maruhenda A, Sastre P, Sánchez-Pina MA, Pallas V (2004) Comparative infection progress analysis of Lettuce big-vein virus and Mirafiori lettuce virus in lettuce crops by developed molecular diagnosis techniques. Phytopathology 94, 470-477 Navarro JA, Torok VA, Vetten HJ, Pallas V (2005) Genetic variability in the coat protein genes of Lettuce big-vein associated virus and Mirafiori lettuce big-vein virus. Archives of Virology 150, 681-694 Navarro JA, Torok VA, Vetten HJ, Pallas V (2005) Genetic variability in the coat protein genes of Lettuce big-vein associated virus and Mirafiori lettuce big-vein virus. Archives of Virology 150, 681-694 Nicaise V, German-Retana S, Sanjuan R, Dubrana MP, Mazier M, Maisonneuve B, Candresse T, Caranta C, Le Gall O (2003) The eukaryotic translation initiation factor 4E controls lettuce susceptibility to the potyvirus Lettuce mosaic virus. Plant Physiology 132, 1272-1282 Nichol ST, Beaty BJ, Elliot RM, Goldbach R, Plyusnin A, Schmaljohn CS, Tesh RB (2005) Bunyaviridae In: Fauquet CM, Mayo MA, Maniloff J, Desselberger U, Ball LA (Eds) Virus Taxonomy, Academic Press, California, San Diego, USA, pp 695-716 Peypelut M, Krause-Sakate R, Guiraud T, Pavan MA, Candresse T, Zerbini FM, Le Gall O (2004) Specific detection of Lettuce mosaic virus isolates belonging to the ‘‘Most’’ type. Journal of Virological Methods 121, 119124 Pink DAC, Kostova D, Walkey DGA (1992a) Differentiation of pathotypes of lettuce mosaic virus. Plant Pathology 41, 5-12 Pink DAC, Lot H, Johnson R (1992b) Novel pathotypes of lettuce mosaic virus – breakdown of a durable resistance? Euphytica 63, 169-174 Pozzer L, Bezerra IC, Kormelink R, Prins M, Peters D, Resende RO, de Ávila AC (1999) Characterization of a Tospovirus isolate of Iris yellow spot virus associated with a disease in onion fields in Brazil. Plant Disease 83, 345-350 Resende JAM, Cupertino EP (1995) Doenças em hortaliças. Informe Agropecuário 18, 18-27 Revers F, Lot H, Souche S, Le Gall O, Candresse T, Dunez J (1997a) Biological and molecular variability of lettuce mosaic virus isolates. Phytopathology 87,397-403 Revers F, van der Vlugt RAA, Souche S, Lanneau M, Lot H, Candresse T, Le Gall O (1999) Nucleotide sequence of the 3' terminal region of the genome of four Lettuce mosaic virus isolates from Greece and Yemen. Archives of Virology 144, 1619-1626 Revers F, Yang SJ, Walter J, Souche S, Lot H, Le Gall O, Candresse T, Dunez J (1997b) Comparison of the complete nucleotide sequences of two isolates of lettuce mosaic virus differing in their biological properties. Virus Research 47, 167-177 Rodrigues MGR, Marinho, VLA, Ribeiro SG, Kitajima EW (1991) Mosaico em margarida amarela Coreopsis lanceolata causado pelo vírus do mosaico do picão. Fitopatologia Brasileira 16, 114-117 Roggero P, Ciuffo M, Vaira AM, Accotto GP, Masenga V, Milne RG (2000) An Ophiovirus isolated from lettuce with big-vein symptons. Archives of Virology 145, 2629-2642 Roggero P, Lot H, Souche S, Lenzi R, Milne RG (2003) Occurrence of Mirafiori lettuce virus and Lettuce big-vein virus in relation to development of big-vein symptoms in lettuce crops. European Journal of Plant Pathology 109, 261-267 Rosales M, Sepulveda P, Bruna A (2004) First report of Lettuce big-vein virus and Mirafiori lettuce virus in Chile. Plant Disease 88, 1286 Ryder EJ (1970) Inheritance of resistance to common lettuce mosaic. Journal of the American Society for Horticultural Sciences 95, 378-379 41