Chinese tomato yellow leaf curl virus--a new species of geminivirus

ＧｅｍｉｎｉｖｉｒｕｓｅｓａｒｅａｇｒｏｕｐｏｆｐｌａｎｔｖｉｒｕｓｅｓｃｈａｒａｃｔｅｒｉｚｅｄｂｙｔｈｅｉｒｃｉｒｃｕｌａｒｓｉｎｇｌｅｓｔｒａｎｄｅｄＤＮＡ(ｓｓＤＮＡ)ｇｅｎｏｍｅａｎｄａｕｎｉｑｕｅｇｅｍｉｎａｔｅｐａｒｔｉｃｌｅｍｏｒｐｈｏｌｏｇｙ[1].Ｇｅｍｉｎｉｖｉｒｕｓｅｓａｒｅｄｉｖｉｄｅｄｉｎｔｏｔｈｒｅｅｓｕｂｇｒｏｕｐｓｏｎｔｈｅｂａｓｉｓｏｆｇｅｎｏｍｅｏｒｇａｎｉｚａｔｉｏｎａｎｄｉｎｓｅｃｔｖｅｃｔｏｒ:ＡｌｌｓｕｂｇｒｏｕｐＩｇｅｍｉｎｉｖｉｒｕｓｅｓａｒｅｌｅａｆ…     

Serological studies on the accumulation and localisation of three tomato leaf curl geminiviruses in resistant and susceptible Lycopersicon species and tomato cultivars

Accessions of wild Lycopersicon spp. and selected Fl hybrid tomato cultivars were compared for their resistance to three whitefly-transmissible geminiviruses: Indian tomato leaf curl virus (ITmLCV) and tomato yellow leaf curl viruses from Sardinia (TYLCV-Sar) and Senegal (TYLCV-Sen). The resistance of different plant lines was expressed in different ways but in most instances a given line reacted similarly to graft inoculation with the three viruses. L. pimpinellifolium LA1478 produced as much virus antigen, assessed by triple antibody sandwich-ELISA, as the susceptible cv. Moneymaker but developed only very mild symptoms and is therefore tolerant of infection. In L. hirsutum LA1777 and L. peruvianum CMV-INRA, very mild or no symptoms developed but antigen concentrations were substantially less than in Moneymaker. L. chilense LA1969 remained symptomless and its antigen concentration was < 1% of that in Moneymaker. Symptoms were mild or barely evident in the Fl hybrid cultivars. Cultivars Tyking and Fiona had antigen concentrations about 5–10% of those of Moneymaker, whereas TY20, Top 21 and Tyger had intermediate antigen concentrations. In a few instances, the extent to which virus accumulation was restricted depended on the challenge virus. Accumulation of TYLCV-Sen in TY20, Top 21 and Tyger was less affected than that of the other two viruses, and accumulation of TYLCV-Sar in accessions LA1777 and CMV-INRA was less affected than that of TYLCV-Sen or ITmLCV. Tissue-printing tests showed that ITmLCV and TYLCV-Sen antigens were confined to phloem tissue. In Tyking, the number of virus antigen-containing phloem traces and the antigen content of individual traces were less than in Moneymaker but the partitioning of antigen between internal and external phloem was unaffected.     

Particle purification, properties and epitope variability of Indian tomato leaf curl geminivirus

A whitefly-transmissible stock isolate of Indian tomato leaf curl geminivirus (ITmLCV) was cultured in graft-inoculated tomato plants and its particles purified from chloroform-clarified extracts in citrate buffer by precipitation with 70 g/litre polyethylene glycol, ultracentrifugation and sucrose density gradient centrifugation. Contaminating helical filaments were eliminated by banding in caesium sulphate gradients. ITmLCV particles had the shape typical for geminiviruses, measured c. 30 × 20 nm and contained a single major protein of estimated mol. wt c. 32 000. They reacted in immunosorbent electron microscopy with antisera to four other whitefly-transmitted geminiviruses. ITmLCV reacted with one out of 17 monoclonal antibodies specific for different epitopes in the particle protein of African cassava mosaic geminivirus and five or six out of 10 monoclonal antibodies to the particle protein of Indian cassava mosaic geminivirus. Virus isolates from tomato at nine locations in Karnataka State showed only slight differences in epitope profile, and isolates from four weed species in tomato fields were similar or identical to those from tomato.     

烟粉虱传播双生病毒研究进展

综述了烟粉虱Bemisia tabaci对双生病毒的获取、传播及存留等方面的特性。烟粉虱最短的获毒和接种时间为15～30 min;双生病毒在烟粉虱体内可存留1至数周,有的终身存在。烟粉虱对双生病毒的传毒效率除了随其获毒及传毒时间的延长、传毒烟粉虱个体数量的增加以及病毒体浓度的增加而提高外,还与烟粉虱的龄期及性别有关。双生病毒除了在植物与粉虱之间直接传播外,还可通过烟粉虱交配及经卵携带的途径在烟粉虱个体和代别间进行传播。寄主植物、双生病毒的一些特殊蛋白以及烟粉虱内共生菌产生的GroEL蛋白,都可影响烟粉虱携带的双生病毒种类及传毒的可能性。双生病毒可对烟粉虱的发育、存活和生殖产生不利或有利的影响。雌成虫携带番茄黄化曲叶病毒（tomato yellow leaf curl virus, TYLCV）后,存活力和生殖力均下降; 而携带番茄斑驳病毒（tomato mottle virus, ToMoV）后,生殖力提高。此外,植物感染双生病毒后,其对烟粉虱的适合性可能提高。     

A shift of vector specificity acquired by a begomovirus through natural homologous recombination

Recombination is common in plant viruses such as geminiviruses, but the ecological and pathogenic consequences have been explored only in a few cases. Here, we found that a new begomovirus, tomato yellow leaf curl Shuangbai virus (TYLCSbV), probably originated from the recombination of Ageratum yellow vein China virus (AYVCNV) and tobacco curl shoot virus (TbCSV). Agrobacterium-mediated inoculation showed that TYLCSbV and AYVCNV have similar levels of infectivity on tomato and tobacco plants. However, the two viruses exhibit contrasting specificities for vector transmission, that is, TYLCSbV was efficiently transmitted by the whitefly Bemisia tabaci Mediterranean (MED) rather than by the whitefly B. tabaci Middle East-Asia Minor 1 (MEAM1), whereas AYVCNV was more efficiently transmitted by MEAM1. We also showed that the transmission efficiencies of TYLCSbV and AYVCNV are positively correlated with the accumulation of the viruses in whitefly whole bodies and organs/tissues. The key coat protein amino acids that determine their accumulation are between positions 147 and 256. Moreover, field surveys suggest that MED has displaced MEAM1 in some regions where TYLCSbV was collected. Viral competition assays indicated that TYLCSbV outcompeted AYVCNV when transmitted by MED, while the outcome was the opposite when transmitted by MEAM1. Our findings suggest that recombination has resulted in a shift of vector specificity that could provide TYLCSbV with a potential selective transmission advantage, and the population shift of whitefly cryptic species could have influenced virus evolution towards an extended trajectory of transmission.     

Detection and relationships of cotton leaf curl virus and allied whitefly-transmitted geminiviruses occurring in Pakistan

A stock culture of cotton leaf curl virus from Pakistan (CLCuV-PK), was transmitted by whiteflies (Bemisia tabaci) to seven plant species, including French bean, okra, tobacco and tomato, and caused vein thickening and leaf curl symptoms. It was readily detected in triple antibody sandwich ELISA (TAS-ELIS A) by 11 out of 31 monoclonal antibodies raised against the particles of three other geminiviruses: African cassava mosaic, Indian cassava mosaic and okra leaf curl viruses. Reaction strength was enhanced when the tissue extraction fluid contained sodium sulphite. Minor variations in epitope profile were found among virus isolates from cotton (Gossypium hirsutum) collected from different districts in Pakistan over a 5-year period. These epitope profiles were distinguishable from that of cotton leaf curl virus from G. barbadense in southern India but indistinguishable from the profiles of viruses causing yellow vein disease of okra in India or Pakistan, or leaf curl of okra {Abelmoschus esculentus), Hibiscus tiliaceus, radish or sunflower in Pakistan, suggesting that these plants are putative natural hosts of CLCuV-PK. The viruses in cotton, and in okra with leaf curl or yellow vein symptoms, were also detected by PCR with three pairs of CLCuV-PK-specific primers. Five additional whitefly-transmitted geminiviruses were found among isolates from 11 other naturally-infected species in Pakistan, and were distinguished by their epitope profiles. These viruses were associated, respectively, with tobacco leaf curl, squash yellow blotch, tomato yellow leaf curl, watermelon leaf crinkle and soybean yellow mosaic diseases. The first four of these viruses were detected readily by PCR with geminivirus general primers but only weakly, if at all, with two pairs of CLCuV-PK-specific primers. Pakistani crops are infected with a range of distinguishable but relatively closely related whitefly-transmitted geminiviruses, some of which resemble those found in India.     

番茄曲叶病及其血清学和PCR测定

我国曾报道的番茄病毒病有多种,其中最常见的是黄瓜花叶病毒(CMV)和烟草花叶病毒(TMV)引起的花叶病。柯冲等(1964)在大陆首次报道烟粉虱(Bemisia tabaci)传播的番茄病毒病——番茄黄顶病,此病在50～60年代曾在广州市郊流行,造成大面积减产。Green等(1984)报道台湾发生番茄黄曲叶病,此病与日本的番茄黄矮病(Tomato yellow dwarf)相似,并且与烟草曲叶病毒(TLCV)有血清学关系。印度、委内瑞拉等国也曾报道发生由烟粉虱传播的番茄曲叶病和番茄黄曲叶病。1991和1992年秋,在广西南宁市郊发现一种症状表现为植株矮缩,叶片向上向内卷曲,叶背面产生耳状或杯状增生物,对光看有时可见叶脉呈墨绿色,不结果或少结果的番茄病害。1992年秋广西农业科学院的番茄试验地发病率高达6.8％,对当地秋番茄生产构成了威胁。作者对病害症状、传播、血清学反应及PCR分析等方面与烟草曲叶病毒进行了比较研究,证实了该病的病原与烟草曲叶病毒有很高的同源性。现将研究结果简报如下。     

Leaf curl disease of tomato in Haldwani (Uttarakhand), India region is caused by a begomovirus with satellite molecule DNA β

The leaf curl disease of tomato was observed in the Haldwani region of Uttarakhand, India during 2004–2007 with an average disease incidence of 49.8 and 73.7% during the month of October and February, respectively. The virus isolate from the infected tomato plants was transmissible to healthy tomato plants by whiteflies (Bemisia tabaci), and the inoculated plants showed typical leaf curl symptoms with a latent period of 16–18 days. The total DNA was extracted from the infected plants and subjected to polymerase chain reaction to amplify the genomic components. The coat protein (CP) gene of ～750 nt was amplified using a set of CP gene specific primer and sequenced (EU847240). Sequence analysis of 701 nt from the N′ terminal region revealed that it had a sequence identity of more than 90% with other isolates/strains of Tomato leaf curl New Delhi virus. A satellite molecule, DNA β of ～1.4 kb was also amplified using universal DNA β-specific primers, cloned and sequenced (EU847239). The isolated DNA β was 1370 nt in length and had a nucleotide sequence identity of 91–93% with DNA β associated with cowpea severe leaf curl and tomato leaf curl disease (TomLCD) reported from India and Pakistan, respectively, and followed by 79% with DNA β associated with TomLCDs reported from Rajasthan. This result showed that the satellite DNA β was associated with TomLCD in Haldwani.     

Host range, vector relations and serological relationships of cotton leaf curl virus from southern India

In 1989 to 1991, leaf curl disease was observed in cotton (Gossypium bar-badense cv. Local) grown in kitchen gardens in five districts in Karnataka State, India, and in 1994 it was recorded in G. hirsutum cv. Sharada in two districts. Symptoms consist of leaf curling, vein thickening, leaf enations, and stunting and distortion of plants. The disease is caused by cotton leaf curl virus (CLCuV-K), which was transmitted by the whitefly Bemisia tabaci to 24 plant species in six families. Hosts include bean (Phaseolus vulgaris), pepper, tobacco, tomato and several weeds, almost all of which developed leaf curl, with or without vein thickening. CLCuV-K was transmitted from cotton to cotton by adult B. tabaci after an acquisition access period of 1 h, could be inoculated in 5 min, had a minimum latent period of 8 h and was retained by viruliferous insects for up to 9 days. Female B. tabaci transmitted more frequently than males. CLCuV-K is a whitefly-transmitted geminivirus. It reacted with two out of 17 monoclonal antibodies (MAbs) raised to African cassava mosaic virus and five out of 10 MAbs raised to Indian cassava mosaic virus. CLCuV-K isolates from different locations in Karnataka had similar epitope profiles. As judged by these profiles, CLCuV-K is closely related to Indian tomato leaf curl virus from Karnataka, is distinguishable from several other whitefly-transmitted geminiviruses found in India and is still more distantly related to those, including cotton leaf crumple virus from the USA, found in other continents. CLCuV-K infected all cultivars tested of G. barbadense and one of six cultivars of G. hirsutum but none of G. arboreum or G. herbaceum.     

Different transmission efficiencies may drive displacement of tomato begomoviruses in the fields in Taiwan

A progressive displacement of Tomato leaf curl Taiwan virus (ToLCTWV) by Tomato yellow leaf curl Thailand virus (TYLCTHV) from 2005 to 2009 has been recorded in tomato fields in Taiwan. Begomoviruses are exclusively transmitted by Bemisia tabaci complex, so we hypothesised that the displacement of tomato begomoviruses in the fields may be due to the invasion of a new virus/vector and the different transmission efficiencies of the viruses by the vectors. The objective of this research was to compare the transmission efficiency of TYLCTHV and ToLCTWV by the B and Q biotypes of B. tabaci complex. When transmission efficiency, virus retention in vector, and latent period for vector transmission were compared, the B biotype transmitted TYLCTHV and ToLCTWV more efficiently than did the Q biotype, and transmitted TYLCTHV more efficiently than ToLCTWV. The B biotype retained both viruses and remained infective throughout adulthood, but the Q biotype did not keep its infectivity, although it did retain both viruses lifelong. The B biotype transmitted TYLCTHV and ToLCTWV with the shortest latent period. In summary, B. tabaci B biotype and TYLCTHV is the best alliance for disease transmission, so we conclude that this may be one of drivers responsible for the displacement of ToLCTWV by TYLCTHV in tomato fields in Taiwan.     

Differential indirect effects of two plant viruses on an invasive and an indigenous whitefly vector: implications for competitive displacement

The role of vector–begomovirus–plant interactions in the widespread invasion by some members of the whitefly species complex Bemisia tabaci is poorly understood. The invasive B biotype of B. tabaci entered China in the late 1990s and had become the predominant or only biotype of the whitefly in many regions of the country by 2005–2006. Meanwhile epidemics of begomoviruses have been observed in many crops including tomato for which Tomato yellow leaf curl China virus (TYLCCNV) and Tomato yellow leaf curl virus (TYLCV) have been identified as two major disease-causing agents. Here, we conducted laboratory experiments to compare the performance of the invasive B and indigenous ZHJ1 whitefly biotypes on uninfected, TYLCCNV-infected and TYLCV-infected plants of tomato cv. Hezuo903, a cultivar that has been widely cultivated in many regions of China. The infection of tomato plants by either of the viruses had no or only marginal effects on the development, survival and fecundity of the B biotype. In contrast, survival and fecundity of the ZHJ1 biotype were significantly reduced on virus-infected plants compared to those on uninfected plants. Populations of the B biotype on uninfected and TYLCCNV-infected plants increased at similar rates, whereas population increase of the ZHJ1 biotype on TYLCCNV-infected plants was affected adversely. These asymmetric responses to virus infection of tomato plants between the B and ZHJ1 biotypes are likely to offer advantages to the B biotype in its invasion and displacement of the indigenous biotype.     

Virus–vector Relationships,Host Range,Detection and Sequence Comparison of Chilli leaf curl virus Associated with an Epidemic of Leaf Curl Disease of Chilli in Jodhpur,India

An epidemic of chilli leaf curl disease was recorded in 2004 in Jodhpur, a major chilli‐growing area in Rajasthan, India. Several isolates were efficiently transmitted by the whitefly (Bemisia tabaci), all of which induced severe leaf curl symptoms in chilli. A single whitefly was capable of transmitting the virus, and eight or more whiteflies per plant resulted in 100% transmission. The minimum acquisition access period (AAP) and inoculation access period (IAP) were 180 and 60 min, respectively. The virus persisted in whiteflies for up to 5 days postacquisition. Of 25 species tested, the virus infected only five (Capsicum annuum, Carica papaya, Solanum lycopersicum, Nicotiana tabacum and N. benthamiana). The virus was identified as Chilli leaf curl virus (ChiLCV), which shared the closest sequence identity (96.1%) with an isolate of ChiLCV from potato in Pakistan and showed sequence diversity up to 12.3% among the ChiLCV isolates reported from India and Pakistan. A betasatellite was identified, which resembled most closely (97.3%) that of Tomato leaf curl Bangladesh betasatellite previously reported from chilli and tomato leaf curl in India. The betasatellite was very different from that reported from chilli leaf curl in Pakistan, indicating that different betasatellites are associated with chilli leaf curl in India and Pakistan. We describe here for the first time the virus–vector relationships and host range of ChiLCV.     

Engineering resistance against tomato yellow leaf curl virus (TYLCV) using antisense RNA

One of the most severe diseases of cultivated tomato worldwide is caused by tomato yellow leaf curl virus (TYLCV), a geminivirus transmitted by the whitefly Bemisia tabaci. Here we describe the application of antisense RNAs to interfere with the disease caused by TYLCV. The target of the antisense RNA is the rare messenger RNA of the Rep protein, encoded by the C1 gene. Transgenic Nicotiana benthamiana plants expressing C1 antisense RNA were obtained and shown to resist infection by TYLCV. Some of the resistant lines are symptomless, and the replication of challenge TYLCV almost completely suppressed. The transgenes mediating resistance were shown to be effective through at least two generations of progeny.     

Tomato yellow leaf curl virus and Tomato leaf curl Taiwan virus Invade South-east Coast of China

An epidemic outbreak of severe yellow leaf curl disease was reported in field grown tomato within Zhejiang Province of China in the autumn–winter cropping season of 2006. A molecular diagnostic survey was carried out based on comparisons of partial and complete viral DNA sequences. Comparison of partial DNA‐A sequences amplified with degenerate primers specific for begomoviruses confirmed the presence of two types of begomoviruses. The complete DNA sequences of five isolates, corresponding to the two types, were determined. Sequence comparisons and phylogenetic analysis revealed that they correspond to two previously identified begomoviruses, Tomato yellow leaf curl virus and Tomato leaf curl Taiwan virus. The satellite DNAβ molecule was not detected in these samples by either PCR or Southern blot hybridization analysis. There has been no previous report of geminivirus disease incidence in Zhejiang Province, indicating that the introduction of these two tomato infecting geminiviruses into the agro‐ecological zone of South‐eastern China is a fairly recent event. The implications for disease control are discussed.     

Tomato leaf curl geminivirus associated with cantaloupe yellow leaf disease in Thailand

Geminivirus associated with yellow leaf disease of cantaloupe plants was detected using polymerase chain reaction (PCR) with geminivirus-specific degenerate primers which anneal within the AC1 ORF (replication initiator protein gene) and the AV1 ORF (coat protein gene). A DNA fragment of 1.2 kbp was amplified, cloned and sequenced. The 32-base stem loop region was found in the amplified fragment. This included the conserved nonanucleotide sequence TAATATTAC present in all geminiviruses. The nucleotide sequence of the intergenic region (IR) was compared with 28 whitefly-transmitted geminiviruses. The geminivirus associated with yellow leaf disease of cantaloupe plants showed 96.2% sequence identity with DNA A of tomato leaf curl geminivirus from India (ToLCV-In2). These data suggest that cantaloupe yellow leaf disease was caused by ToLCV.     

Occurrence of whitefly-transmitted geminiviruses in crops in Burkina Faso, and their serological detection and differentiation

Whitefly-transmitted geminiviruses were found to be associated with four diseases of crop plants in Burkina Faso: cassava mosaic, okra leaf curl, tobacco leaf curl and tomato yellow leaf curl. Tomato yellow leaf curl is an economically serious disease, reaching a high incidence in March, following a peak population of the vector whitefly, Bemisia tabaci, in December. Okra leaf curl is also a problem in the small area of okra grown in the dry season but is not important in the main period of okra production in the rainy season. The geminiviruses causing these four diseases, African cassava mosaic (ACMV), okra leaf curl (OLCV), tobacco leaf curl (TobLCV) and tomato yellow leaf curl (TYLCV) viruses, were each detected in field-collected samples by triple antibody sand-wich-ELISA with cross-reacting monoclonal antibodies (MAbs) to ACMV. Epitope profiles obtained by testing each virus isolate with panels of MAbs to ACMV, OLCV and Indian cassava mosaic virus enabled four viruses to be distinguished. ACMV and OLCV had similar but distinguishable profiles. The epitope profile of TobLCV was the same as that of one form of TYLCV (which may be the same virus) and was close to the profile of TYLCV from Sardinia. The other form of TYLCV reacted with several additional MAbs and had an epitope profile close to that of TYLCV from Senegal. Only minor variations within each of these four types of epitope profile were found among geminivirus isolates from Burkina Faso. Sida acuta is a wild host of OLCV.     

The autophagy pathway participates in resistance to tomato yellow leaf curl virus infection in whiteflies

Macroautophagy/autophagy plays an important role against pathogen infection in mammals and plants. However, little has been known about the role of autophagy in the interactions of insect vectors with the plant viruses, which they transmit. Begomoviruses are a group of single-stranded DNA viruses and are exclusively transmitted by the whitefly Bemisia tabaci in a circulative manner. In this study, we found that the infection of a begomovirus, tomato yellow leaf curl virus (TYLCV) could activate the autophagy pathway in the Middle East Asia Minor 1 (MEAM1) species of the B. tabaci complex as evidenced by the formation of autophagosomes and ATG8-II. Interestingly, the activation of autophagy led to the subsequent degradation of TYLCV coat protein (CP) and genomic DNA. While feeding the whitefly with 2 autophagy inhibitors (3-methyladenine and bafilomycin A₁) and silencing the expression of Atg3 and Atg9 increased the viral load; autophagy activation via feeding of rapamycin notably decreased the amount of viral CP and DNA in the whitefly. Furthermore, we found that activation of whitefly autophagy could inhibit the efficiency of virus transmission; whereas inhibiting autophagy facilitated virus transmission. Taken together, these results indicate that TYLCV infection can activate the whitefly autophagy pathway, which leads to the subsequent degradation of virus. Furthermore, our report proves that an insect vector uses autophagy as an intrinsic antiviral program to repress the infection of a circulative-transmitted plant virus. Our data also demonstrate that TYLCV may replicate and trigger complex interactions with the insect vector.     

Expression of the Full‐length Coat Protein Gene of Tomato leaf curl Taiwan virus is Not Necessary for Recovery Phenotype in Transgenic Tomato

Transgenic tomato plants expressing full‐length (CPV1) and truncated coat protein (CP) gene (CPV2) of Tomato leaf curl Taiwan virus (ToLCTWV) were generated by Agrobacterium‐mediated transformation. Transgene integration and expression was confirmed by PCR and Southern blotting and Northern analysis, respectively. Resistance was evaluated both in plants of T0 and T1 progenies using viruliferous whiteflies under two different inoculum pressures (10–15 and 40–50 whiteflies/plant). Upon inoculation with ToLCTWV using viruliferous whiteflies, various levels of phenotypic reaction were observed. No complete resistance was observed in any of the plants tested. The reaction of the transgenic tomato lines carrying full‐length and truncated CP gene to ToLCTWV phenotype was (i) susceptible as non‐transgenic control, (ii) delayed symptom expression, (iii) complete susceptible (from delayed symptom expression phenotype) and (iv) recovered phenotype (either plants from symptom expression as non‐transgenic plants or delayed symptom expression phenotype). Dot blot quantification of the ToLCTWV using the replicase gene as a probe revealed that the recovered phenotypes accumulated a low level of ToLCTWV, and virus concentration was gradually reduced from 10 to 14 weeks postinoculation. The possible mechanisms of CP‐mediated resistance are discussed.     

Detection of three whitefly-transmitted geminiviruses occurring in Europe by tests with heterologous monoclonal antibodies

Selected monoclonal antibodies (MAbs), prepared to particles of African cassava mosaic or Indian cassava mosaic geminiviruses, detected three geminiviruses that occur in Europe: abutilon mosaic virus in Abutilon pictum ‘Thompsonii’, tobacco leaf curl virus in Lonicera japonica var. aureo-reticulata and tomato yellow leaf curl virus in Lycopersicon esculentum. All three viruses were detected in indirect ELISA by MAbs SCR 17 and SCR 20 but they were differentiated by their reactions with SCR 18 and SCR 23. Tobacco leaf curl virus was detected only when reducing agents were included in the leaf extraction medium. Inclusion of sodium sulphite slightly improved detection of tomato yellow leaf curl virus but reducing agents were not needed for detection of abutilon mosaic virus.     

Reaction of some tomato cultivars to tomato leaf curl virus and evaluation of the endophytic colonisation with Beauveria bassiana on the disease incidence and its vector,Bemisia tabaci

Tomato line LA1478 and Pusa Ruby were resistant to tomato leaf curl virus (TLCV) disease. They registered higher plant height, number of branches, total phenol content and yield per plant than the other cultivars. Variety Peto 86 was tolerant to the disease while the other popular tomato cultivars, i.e. Ace, Early Pack, Money Maker, Prichard and Strain B were highly susceptible to the disease. Plant height and number of branches per plant revealed significantly positive association with fruit yield per plant. The disease index of TLCV exhibited significant negative correlations with plant height, total phenol content and fruit yield per plant – 0–4 and 5–25 adult whiteflies were observed on resistant susceptible cultivars. In the case of epiphytically colonisation by Beauveria bassiana conidia, not all developing hyphae on the leaf surface penetrated the whitefly cuticle. Many of the germ tubes elongated to a short distance before terminating its growth. On the other hand, the rapid staining of tomato tissues injected with B. bassiana conidial suspension indicates that the entomopathogenic fungus was established inside tomato tissues until the end time of the trial. The direct injection with the spore suspension yielded high post-colonisation, where the fungus was recovered from sites distant from the point of inoculation. This indicates that the fungus has the potential to move throughout the plant tissues. Laboratory bioassay of tomato whitefly feeding on tomato tissues containing B. bassiana conidial spores indicates that plant endophytic colonisation with entomopathogenic fungi may reduce insect survival on these plants. LT₅₀ values of the test diet were between three and four days. The mortality of Bemisia tabaci was high in the case of endophytically colonisation compared to epiphytically one (90.0% compared to 10.0% during three days) for whiteflies fed tomato tissues containing 1.5 × 10⁷ B. bassiana spores/ml. Application of B. bassiana as an artificial endophyte inside tomato plants can be an important component in the integrated control of tomato whiteflies. The endophytic colonising can achieve biocontrol effect based on induced disease resistance in plant tissues. According the available references, this is the first report on B. tabaci controlling by plant endophytic treatment.