Viruliferous Insects Research Articles

Alteraciones fisiológicas asociadas a la infección con Mal de Río Cuarto virus (MRCV) y a fitotoxicidad provocada por su insecto vector (Delphacodes kuscheli Fennah) en trigo

Mal de Rio Cuarto virus produces a wheat disease, resulting from the combined action of this pathogen and its main vector, Delphacodes kuscheli. Knowing that viruses and insects cause a severe symptomatology in different genotypes, the objective of this work was to determine the possible physiological alterations in three phenological stages of wheat (cv ProINTA Federal) inoculated by two geographically distant MRCV isolates and exposed to non viruliferous insects. Treatments were run at three plant stages: first leaf emergence through coleoptile, four and five unfolded leaves. The health status of these plants was confirmed by ELISA. Physiological alterations were observed in both infected plants and plants exposed to non viruliferous insects (PS). The total soluble sugar, starch and soluble protein contents increased remarkably in the plants infected by MRCV. While chlorophyll content decreased considerably in PS and MRCV infected plants, malondialdehyde levels increased in both cases. These variations could indicate oxidative damage associated to biotic stress in these plants. The physiological alterations were similar on the different phenological stages. Only the starch increase was significantly higher in plants infected at the first leaf emergence from the coleoptile.

벼 바이러스(RSV, RBSDV)와 애멸구의 간편한 VC/RT-PCR 유전자 진단기술

우리나라 벼에 발생하는 주요 바이러스 중 애멸구에 의하여 전염하는 벼줄무의잎마름병(RSV)과 벼검은줄오갈병(RBSDV)에 대한 간편한 유전자 진단법인 VCHT-PCR 방법을 개발하였다. 벼 잎의 경우 즙액 추출 완충액은 0.5% sodium sulfite를 첨가한 0.01 M 인산완충액(pH 7.0)을 기본 완충액으로 이용하고 애멸구를 진단할 경우에는 기본 완충액에 2% PVP을 첨가하였을 때 VC/RT-PCR 진단이 잘 되었다. VC/RT-PCR을 이용한 진단에 적합한 RSV와 RBSDV 프라이머를 선발하였고 이것을 이용하여 동시진단으로 경기도 김포, 평택, 시흥지역에서 채집한 애멸구의 RSV와 RBSDV의 보독충을 쉽고 경제적으로 진단 할 수 있었다. ELISA에 의한 진단결과와 비교할 때 세 지역을 합하여 RSV에 대한 평균 보독충률은 9.2%로 동일하였으나 보독충 중 일부가 ELISA와 VC/RT-PCR 두 방법에 의한 진단결과가 다르게 나온 것은 RSV의 혈청학적, 유전적 계통 존재 가능성을 제시하고 있다. 포장에서 채집한 애멸구의 VC/RT-PCR 진단효율은 RSV와 RBSDV를 동시에 진단하므로 써 RSV만 진단이 가능한 ELISA 결과 보다 3.2% 높았다. Genetic diagnosis method of Virion Captured (VC)/RT-PCR for Rice stripe virus (RSV) and Rice black-streaked dwarf virus (RBSDV), Korean major rice viruses transmitted by small brown plant hopper, Laodelphax striatellus, was developed. Virion extraction buffer for rice plant was 0.01M potassium phosphate buffer, pH 7.0, containing 0.5% sodium sulfite. However, the extraction buffer for L. striatellus was 0.01M potassium phosphate buffer, pH 7.0, containing 0.5% sodium sulfite and 2% polyvinylpyrrolidone wt 40,000 (PVP-40). Specific primers for detection of RSV and RBSDV were selected for VC/RT-PCR method. The specific primers were used as a duplex primer to detect viruliferous small brown plant hopper collected from Gimpo, Pyeongtaek and Siheung areas in Gyeonggi province. The genetic diagnosis methods of single and duplex VC/RT-PCR for RSV and RBSDV could be used easily and economically, especially on the diagnosis of L. striatellus. The rate of viruliferous insect (RVI) for RSV was compared with ELISA and VC/RT-PCR for L. striatellus collected from fields. RVI by ELISA was same as 9.2% with RVI by VC/RT-PCR. However, there were some different detection results between the methods. It could be suggested that there is a possibility of serological and/or genomic differences among RSV isolates. The portion of RVI detected simultaneously by ELISA and VC/RT-PCR was 71.0%, and the detection rate from VC/RT-PCR was higher as 3.2% than that from ELISA, which had a reason of simultaneous detection ability both RSV and RBSDV of VC/RT-PCR.

옥수수 재배지의 벼검은줄오갈병 매개충인 애멸구의 보독충율

2007년 경북문경의 옥수수 재배지 약 <TEX>$1000\;m^2$</TEX>에 RBSDV가 약 <TEX>$99{\sim}100%$</TEX> 발생되었다. 이 포장은 바로 인접지역에 감나무가 재배되고 있었으며 이 나무 아래에 녹비작물로서 보리를 재배하였다. 보리는 RBSDV의 매개충인 애멸구의 겨울철 중간기주로 작용하여 애멸구의 월동에 유리한 환경으로 작용한 것으로 생각된다. 진단은 S9 특이 primer를 사용한 RT-PCR 방법으로 확인하였다. 이 포장에서 매개충인 애멸구를 채집하여 RT-PCR에 의한 보독충율을 조사하였다. 5월 3일, 6월 7일, 8월 4일 3회 애멸구를 채집하여 검정한 결과 각각 2.9%, 4.8%, 4.4%의 보독충율을 나타내었다. 이것은 RBSDV에 이병된 벼 포장에서 나타나는 보독충율과 비슷한 결과를 나타내었다. The Rice black-streaked dwarf virus (RBSDV) infected 99-100% of a <TEX>$1000\;m^2$</TEX>-maize field in Mungyeong City in 2007. Adjacent to the area is a Persimmon orchard where barley crops were grown under the trees as green manure crops and for soil amendments. The barley acted as winter host to the small brown plant hoppers (SBPH) enabling the insects to survive and pass the winter season. The existence of RBSDV was detected and confirmed by RT-PCR using S9 specific primer. Samples of the insect vector SBPH were collected in the area on May 3, June 7 and, August 4 and the results of the RT-PCR analysis revealed viruliferous insect rates of 2.9, 4.8, and 4.4%, respectively. These observed viruliferous insect rates were similar with those detected in RBSDV infected rice fields.

Expression of Stress-Response Proteins Upon Whitefly-Mediated Inoculation of Tomato yellow leaf curl virus in Susceptible and Resistant Tomato Plants

To better understand the nature of resistance of tomato to the whitefly (Bemisia tabaci, B biotype)-transmitted Tomato yellow leaf curl virus (TYLCV), whiteflies and TYLCV were considered as particular cases of biotic stresses and virus resistance as a particular case of successful response to these stresses. Two inbred tomato lines issued from the same breeding program that used Solanum habrochaites as a TYLCV resistance source, one susceptible and the other resistant, were used to compare the expression of key proteins involved at different stages of the plant response with stresses: mitogen-activated protein kinases (MAPKs), cellular heat shock proteins (HSPs, proteases), and pathogenesis-related (PR) proteins. The two biotic stresses-non-viruliferous whitefly feeding and virus infection with viruliferous insects--led to a slow decline in abundance of MAPKs, HSPs, and chloroplast protease FtsH (but not chloroplast protease ClpC), and induced the activities of the PR proteins, beta-1,3-glucanase, and peroxidase. This decline was less pronounced in virus-resistant than in virus-susceptible lines. Contrary to whitefly infestation and virus infection, inoculation with the fungus Sclerotinia sclerotiorum induced a rapid accumulation of the stress proteins studied, followed by a decline; the virus-susceptible and -resistant tomato lines behaved similarly in response to the fungus.

Retention of Rice dwarf virus by Descendants of Pairs of Viruliferous Vector Insects After Rearing for 6 Years

ABSTRACT Rice dwarf virus (RDV) is characterized by its unusual ability to multiply in both plants and leafhopper vector insects and by its transovarial mode of transmission. Colonies of Nephotettix cincticeps, derived originally from pairs of leafhoppers infected with an ordinary strain of RDV, were maintained for 6 years in the laboratory and were found, at the end of this time, still to harbor RDV. Moreover, the isolate of RDV, designated RDV-I, obtained from these colonies retained the ability to infect rice plants. When we raised leafhoppers separately from eggs that had been placed individually on pieces of water-soaked filter paper and reared them in the presence of healthy rice seedlings, we found that all of these leafhoppers harbored RDV. This observation suggested that RDV-I had been maintained in the leafhoppers by transovarial transmission. Two further observations, namely, the low rate of acquisition of RDV by virus-free insect nymphs on symptomless plants on which viruliferous insects had been reared, and the fact that only 2 to 5% of plants had symptoms when rice seedlings were inoculated via RDV-I-viruliferous insects, confirmed that the maintenance of RDV-I by any other mode of transmission through plants and insects was unlikely. This efficient and long-term maintenance of RDV in a population of viruliferous insects might explain the prolonged duration of rice dwarf disease in the field, once there has been a serious outbreak.

시설 및 노지재배 고추의 바이러스병 발생과 분포

2002년 노지 및 시설재배 고추를 대상으로 전국적인 규모의 바이러스병 발생실태를 조사하였다. 시설(155시료) 및 노지포장(227시료)에서 바이러스 감염주를 채집하여 전자현미경과 RT-PCR로 분석하였다. 시설 및 노지재배 고추에서 수집한 시료에서 TMV, RMV, PVY, AMV, TSWV에 감염된 것은 없었다. 그러나 CMV, BBWV, PepMoV, PMMoV, ToMV, TMGMV는 RT-PCR에 의해 검출되었다. 시설 및 노지재배 포장에서 바이러스 발병율은 상당히 다르게 나타났다. 시설재배에서는 10%의 발병율을 보였으나, 노지재배에서는 약 30%의 발병율을 나타냈다. 복합감염율은 시설 및 노지재배에서 각각 16%와 61%로 나타났다. 시설재배에서는 PMMoV가 우점하는 것으로 보아, 바이러스에 오염된 종자와 재배지에서의 바이러스 감염잔재물이 중요한 전염원이 된 것으로 보인다. 반대로 노지재배에서는 CMV와 BBWV가 우점하였다. 이것으로 보아 바이러스를 보독한 매개충의 이동이 가장 중요한 전염원으로 나타났다. 또한 이 연구에서 BBWV2는 우리나라에서 방제해야 할 새로운 바이러스로 대두되었다. 노지 및 시설재배에서 바이러스병의 발병율과 분포의 차이로 볼 때 연작에 따른 전염원의 축적 및 분포상과 재배지로의 바이러스 보독매개충의 이동이 바이러스 발생을 결정하는 중요한 요소로 보인다. 그러므로 식물병 역학과 고추의 새로운 육종 프로그램에 대한 진전된 연구가 바이러스 병을 줄이는데 필수적일 것이다. In the year of 2002 annual nationwide survey of virus diseases occurring in the pepper fields and greenhouses in Korea, the distribution and the incidence of viral diseases was investigated. The pepper samples from both greenhouses (155 samples) and open fields (227 samples) were collected and further analyzed to detect eleven different viruses by RT-PCR. The results indicate that no sample collected from both greenhouse and open field seems to be infected by TMV, RMV, PVY, AMV, and TSWV. On the other hand, CMV, BBWV2, PepMoV, PMMoV, TMGMV and ToMV are readily identified from greenhouse and open field samples by RT-PCR. The infection rates of the collected samples between greenhouse and open field are largely different. Comparing with 10% of virus-infected pepper samples grown in greenhouse, approximately one third of pepper samples collected from open field are infected. The mixed-infection rates in the virus-infected greenhouse and open field samples are 16% and 61%, respectively. The dominant virus occuring in greenhouse is PMMoV, indicating that virus-infected seed stocks and infected plant debris in the growing area may be important sources of inocula. On the other hand, both CMV and BBWV2 are dominant viruses in open field. This may indicate that the migration of viruliferous insect vectors into pepper fields may be the most important source of inoculum. Also, the survey shows that BBWV2 is newly immerging virus to be controlled in Korea. The discrepancies on the distribution and the occurrence of viral diseases between field and greenhouse may provide a fact that the accumulation and distribution of inoculum by successive cultivation and the migration of viruliferous vectors into growing areas are likely to be important factors to determine the incidence of viral diseases. Therefore, the further studies on epidemiology and the consideration of new breeding program of pepper are essential to minimize virus diseases.

Tomato yellow leaf curl geminivirus (TYLCV-Is) is transmitted among whiteflies (Bemisia tabaci) in a sex-related manner.

Tomato yellow leaf curl virus (TYLCV) is the name given to a complex of geminiviruses infecting tomato cultures worldwide. TYLCV is transmitted by a single insect species, the whitefly Bemisia tabaci. Herein we show that a TYLCV isolate from Israel (TYLCV-Is) can be transmitted among whiteflies in a sex-dependent manner, in the absence of any other source of virus. TYLCV was transmitted from viruliferous males to females and from viruliferous females to males but not among insects of the same sex. Transmission took place when insects were caged in groups or in couples, in a feeding chamber or on cotton plants, a TYLCV nonhost. The recipient insects were able to efficiently inoculate tomato test plants. Insect-to-insect virus transmission was instrumental in increasing the number of whiteflies capable of infecting tomato test plants in a whitefly population. TYLCV was present in the hemolymph of whiteflies caged with viruliferous insects of the other sex; therefore, the virus follows, at least in part, the circulative pathway associated with acquisition from infected plants. Taken as a whole, these results imply that a plant virus can be sexually transmitted from insect to insect.

Sequence analysis of Pns11, a nonstructural protein of rice gall dwarf virus, and its expression and detection in infected rice plants and vector insects.

The nucleotide sequence of genome segment S11 of rice gall dwarf virus (RGDV), a member of Phytoreovirus, was determined. The segment encodes a putative protein of 40 kDa that exhibits approximately 37% homology at the amino acid level to the nonstructural proteins Pns10 of rice dwarf and wound tumor viruses, which are other members of Phytoreovirus. A band of a protein with an apparent molecular mass of 40 kDa was specifically detected in an analysis of cells transfected with S11 cDNA. An antiserum raised against this protein reacted with a protein of approximately 40kDa after fractionation by SDS-PAGE of materials prepared from infected plants and from viruliferous vector insects. However, the antiserum did not react with purified viral proteins. These results suggest that S11 encodes a nonstructural protein of RGDV. This protein was named Pns11.

Transmission of Tomato Yellow Leaf Curl Geminivirus to Imidacloprid Treated Tomato Plants by the Whitefly Bemisia tabaci(Homoptera: Aleyrodidae)

The ability of whiteflies [ Bemisia tabaci (Gennadius), B biotype] to transmit tomato yellow leaf curl geminivirus (TYLCV, Israeli isolate) to imidacloprid-treated and untreated tomato plants was investigated. Viruliferous whiteflies were caged with treated tomato plants held in a nethouse. Insect mortality and tomato infection was assessed during the summer and the winter seasons, after insecticide application. In summer, insects that were given access to tomato plants 3 and 11 d after insecticide treatment died within 80 min. This period increased to 150 min 18 d after treatment. The insecticide lost its potency 25 d after application. In winter the lethal effect of the insecticide lasted longer than in summer. In summer as well as in winter, 70% of the tomato plants caged with viruliferous whiteflies 3 d after insecticide treatment became infected. These figures increased to 80% 11 d after treatment and to nearly 100% 18 d after treatment. Insecticide-treated plants were as prone to infection as nontreated plants. Approximately 48 min of access to a treated tomato plant was sufficient for a single viruliferous insect to inoculate the virus with an efficiency rate of 20% or more. Transmission efficiency was similar to that achieved on nontreated plants. Therefore, viruliferous insects had enough time to inoculate TYLCV to imidacloprid-treated plants before they died.

Detecting the Protein (Pns10) Encoded by Genome Segment 10 of Rice Ragged Stunt Oryzavirus in Diseased Rice and Viruliferous Vector Insects.

The rice ragged stunt virus genome contains 10 dsRNA segments. In segment 10 (S10), a minicistron (nt 20-55) precedes the major ORF (nt 142-1032). The initiation codon of the minicistron was recognized by in vitro translation of wheat germ extracts, while the product of the major downstream ORF was not detected. In order to investigate the expression of the major ORF in vivo, Pns10 was expressed in bacteria and antiserum was produced for immunodetection. Although the detected protein migrated slightly slower on SDS-PAGE than that expressed by bacteria, the major S10 ORF was expressed as a 34kDa protein in both diseased rice plants and viruliferous vector insects.

Evidence for Transovarial Transmission of Tomato Yellow Leaf Curl Virus by Its Vector, the WhiteflyBemisia tabaci

The whiteflyBemisia tabaciis the only vector of the tomato yellow leaf curl geminivirus (TYLCV). The insect transmits the virus in a persistent-circulative manner. TYLCV DNA was detected by polymerase chain reaction and by Southern blot hybridization in progeny (eggs, first and second instars, adults) of single viruliferous whiteflies that developed on eggplant or on cotton (two TYLCV nonhost plants). Furthermore, TYLCV DNA was present in the progeny of insects that had acquired the virus through the egg. The adult progeny of the viruliferous insects and their own progeny were able to infect tomato test plants, producing typical disease symptoms. Ovaries and maturing eggs of viruliferous insects contained viral DNA, as did eggs laid by viruliferous insects maintained on an artificial diet. Eggs laid by nonviruliferous whiteflies on cotton plants previously caged with viruliferous insects did not acquire viral DNA from the plant. Hence, TYLCV can be transmitted through the egg for at least two generations. In the absence of an available plant host, the whitefly may serve as a reservoir of the virus between growing seasons.

Pcr-amplification of tomato yellow leaf curl virus (TYLCV) DNA from squashes of plants and whitefly vectors: Application to the study of TYLCV acquisition and transmission

DNA of tomato yellow leaf curl virus (TYLCV), a geminivirus transmitted by the whitefly Bemisia tabaci, was amplified from squashes of infected tomato plants and of viruliferous vectors using the polymerase chain reaction (PCR). Samples of infected tissues as small as 1 mm2 were squashed onto a nylon membrane. A 1 × 2 mm strip containing the squash was introduced into a 25 µl PCR reaction mix. The reaction products were subjected to gel electrophoresis, blotted and hybridized with a radiolabeled virus-specific DNA probe. TYLCV DNA was amplified from squashes of leaves, roots, and stem of infected tomato and from individual viruliferous whiteflies. The same squash could be used several times to amplify different virus DNA fragments with various sets of primers. Thus plant and insect squashes can be used as templates for the amplification of geminiviral DNA with no need to prepare tissue extracts or purify nucleic acids. The squash-PCR procedure was applied to study whitefly transmission of TYLCV. Tomato plants were inoculated by placing a single viruliferous insect in the center of a young leaflet. In some plants TYLCV DNA was detected at the site of inoculation as early as 5 min after the beginning of the access feeding and in all plants after 30 min. The squash-PCR procedure also was applied to the study of TYLCV acquisition by the insect vector. TYLCV DNA was detected in the head of whiteflies as early as 5 min after the beginning of the access feeding on infected tomato plants. Viral DNA was detected in the thorax after 10 min and in the abdomen after 25 min.

Long-term association of tomato yellow leaf curl virus with its whitefly vector Bemisia tabaci: effect on the insect transmission capacity, longevity and fecundity.

The association between tomato yellow leaf curl geminivirus (TYLCV, Israeli isolate) and its insect vector, the whitefly Bemisia tabaci, was investigated. Insects that emerged during a 24 h period were caged with TYLCV-infected plants for a 48 h acquisition access period, then with egg-plants--a TYLCV non-host--for the rest of their lives. While TYLCV DNA was associated with the whiteflies during their entire adult life, the amount of capsid protein rapidly decreased and was not detectable in the insect after approximately 12 days of age. The ability of the infected whiteflies to transmit TYLCV to tomato test plants steadily decreased with age but did not disappear completely. Transmission by viruliferous insects decreased from 100% to 10-20% during their adult lifetime, compared with a decrease from 100% to 50% for non-viruliferous insects. The association of TYLCV with adult B. tabaci led to a reduction of 17-23% in their life expectancy compared with insects that had not acquired the virus, and to a 40-50% decrease in the mean number of eggs laid. These results suggest that TYLCV has some features reminiscent of an insect pathogen.

高比重ラテックスによる植物ウイルスの簡易検定

The flocculation test using high density latex (HDLF) which gives a reaction figure very similar to that of sensitized sheep red blood cells was established for simplified detection of rice stripe virus from infected rice plants and viruliferous insect vectors. Furthermore, six other plant viruses such as rice dwarf virus, odontoglossum ringspot virus, cymbidium mosaic virus, turnip mosaic virus, carnation mottle virus and cucumber mosaic virus were detected by HDLF of each infected host. These dilution end points for the positive reaction of sap were 1:8×103 to 1:16×103. Though weak non-specific reactions of HDLF were observed in sap from some healthy plants, they could be prevented by diluting sap more than 1:100.

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

SummaryIn 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.

Virus‐vector relationships of a planthopper(Peregrinus maidis)‐bornesorghum stripe tenuivirus∗

Abstract Sorghum stripe virus (SStV), an isolate of maize stripe tenuivirus on sorghum, is transmitted by a delphacid planthopper (Peregrinus maidis) in a persistent manner throughout its life with intermittent transmission in some individuals. The efficiency of virus transmission by the vector was 42% when individual viruliferous vectors were used per test seedling and 100% with five viruliferous insects. The minimum acquisition and inoculation feeding periods were 4 h and 1 h respectively. Vectors became viruliferous and able to transmit the virus within an 8 days’ incubation period after acquisition of virus from the source plant. The first instar nymphs were found to be the most efficient transmitters (64%) compared with the second to fourth instars (50%) and adults (33%). Fifty‐three percent and 35% of macropterous females and males respectively transmitted the virus, whereas in brachypterous females and males the transmission efficiency was 47% and 27% respectively. Of 595 nymphs obtained from the eggs laid by 10 viruliferous females, only 3% of nymphs transmitted the virus to healthy seedlings. The seedlings Inoculated at the one‐leaf and 10‐leaf stage by viruliferous P. maidis showed 62% and 47% infection and the latent period of virus in the host was 8 and 18 days respectively.

Nonpathogenic Nilaparvata lugens Reovirus Is Transmitted to the Brown Planthopper through Rice Plant

Nilaparvata lugens reovirus (NLRV) was found in a colony of the brown planthopper, N. lugens, an important plant virus vector. Unlike with usual phytopathogenic reoviruses, there are no visible symptoms on a rice plant which has been attacked by NLRV-infected planthoppers. Manner of transmission, host range, and multiplication of NLRV in rice plant were studied by enzyme-linked immunosorbent assay or polymerase chain reaction using reverse transcriptase. NLRV was transovumly transmitted to about 15% of nymphs. NLRV could further spread to planthoppers via rice plants through sucking by viruliferous insects. This horizontal transmission was apparently of primary importance for continuous NLRV infection to a colony of N. lugens. By corearing with viruliferous N. lugens on rice seedlings, Laodelphax striatellus acquired NLRV but Sogatella furcifera, Nephotettix cincticeps, and N. malayanus did not during the period studied. NLRV inoculated onto rice plant by viruliferous N. lugens failed to multiply, which accounts for the lack of symptoms on the plant even after an attack by viruliferous planthoppers. These biological properties and our previous data, which revealed a molecular similarity between NLRV and fijiviruses, suggest that NLRV is an ancestral virus of fijiviruses.

Cytopathology and Isolation of Reovirus-like Particles from the Leafhopper Graminella nigrifrons (Homoptera, Cicadellidae)

Nilaparvata lugens reovirus (NLRV) was found in a colony of the brown planthopper, N. lugens, an important plant virus vector. Unlike with usual phytopathogenic reoviruses, there are no visible symptoms on a rice plant which has been attacked by NLRV-infected planthoppers. Manner of transmission, host range, and multiplication of NLRV in rice plant were studied by enzyme-linked immunosorbent assay or polymerase chain reaction using reverse transcriptase. NLRV was transovumly transmitted to about 15% of nymphs. NLRV could further spread to planthoppers via rice plants through sucking by viruliferous insects. This horizontal transmission was apparently of primary importance for continuous NLRV infection to a colony of N. lugens. By corearing with viruliferous N. lugens on rice seedlings, Laodelphax striatellus acquired NLRV but Sogatella furcifera, Nephotettix cincticeps, and N. malayanus did not during the period studied. NLRV inoculated onto rice plant by viruliferous N. lugens failed to multiply, which accounts for the lack of symptoms on the plant even after an attack by viruliferous planthoppers. These biological properties and our previous data, which revealed a molecular similarity between NLRV and fijiviruses, suggest that NLRV is an ancestral virus of fijiviruses.

RICE CULTURES AND VARIETIES WITH RESISTANCE TO RICE TUNGRO VIRUS (RTV) DISEASE

Investigations were carried out to identify rice cultures resistant to rice tungro virus (RTV) disease. Sixty two rice cultures/varieties were screened for resistance to RTV by exposing to one, two and five viruliferous vector insects [green leafhoppers (GLH), Nephotettix virescens (Distant)] per seedling. Seven cultures and one variety were identified as resistant, when exposed to one viruliferous GLH per seedling. Among these only four were found resistant when the inoculum pressure was increased as two viniliferous GLH per seedling and remaining cultures were moderately resistant. All the eight changed from resistant to moderately resistant, when exposed to five viruliferous GLH per seedling.

Acquisition of tomato yellow leaf curl virus by the whitefly Bemisia tabaci

Tomato yellow leaf curl virus (TYLCV) genomic DNA can be detected by Southern blot analysis in nucleic acid extracted from a single whitefly. Acquisition of TYLCV by individual whiteflies in relation to the length of the access period, the virus concentration in, and the developmental stage of plant tissues was studied. The frequency of TYLCV detection increased with the length of the access-period; DNA was detected in 15% of whiteflies tested after a period of access to infected tissue of 30 min, regardless of whether it had a high or a low virus content (5 ng or 0.05 ng TYLCV DNA/micrograms plant chromosomal DNA), and in all insects tested after an 8 h period of access to all the plants. Those insects which had access to the youngest leaves of source plants, which have a high virus content, acquired detectable TYLCV DNA within 2 h. Insects which had access to a tissue for the same period acquired variable amounts of TYLCV DNA; insects feeding on plants with a low virus concentration acquired amounts of viral DNA comparable to those acquired by insects feeding on plants containing a 100-fold greater concentration of virus. Viruliferous insects retained TYLCV DNA for at least 13 days when placed on uninfected tomato plants. In these tests, whitefly could not acquire more than 600 million virus genomes (1 ng viral DNA), suggesting the existence of factors controlling the number of virions present in an insect.