Analysis of the occurrence and distribution of tobacco rattle virus in field soil and disease in a subsequent tulip crop

The occurrence and distribution of tobacco rattle virus (TRV) in field plots was determined by soil bait-testing and disease incidence in tulips subsequently grown on these plots was studied. The virus occurred in patches, calculated as 1.5 m × 3.6 m. The presence of virus was not correlated with numbers of potential vector trichodorid nematodes. Of three trichodorid nematode species present, only Paratrichodorus teres transmitted TRV which, as with virus isolates obtained in bait-tests and from infected tulips, reacted in serological tests with an antiserum prepared against a Dutch isolate of pea-early browning virus (PEBV). Virus prevalence in a subsequent tulip crop was 0.8% and in a sample of tulip plants, virus was recovered only from plants showing virus symptoms. Plots from which TRV was recovered in bait-tests yielded significantly more virus diseased tulips than plots which tested negative for virus. Growing bait-plants in field-plots, as compared with greenhouse tests using soil collected as a series of sub-samples, resulted in an underestimate of the occurrence of TRV.     

Delivery of macromolecules to plant parasitic nematodes using a tobacco rattle virus vector

Plant parasitic nematodes cause significant damage to crops on a worldwide scale. These nematodes are often soil dwelling but rely on plants for food and to sustain them during reproduction. Complex interactions occur between plants and nematodes during the nematode life cycle with plant roots developing specialized feeding structures through which nematodes withdraw nutrients. Here we describe a novel method for delivering macromolecules to feeding nematodes using a virus-based vector [tobacco rattle virus (TRV)]. We show that the parasitic nematode Heterodera schachtii will ingest fluorescent proteins transiently expressed in plant roots infected with a TRV construct carrying the appropriate protein sequence. A prerequisite for this delivery is the presence of replicating virus in root tips prior to the formation of nematode-induced syncytia. We show also that TRV vectors expressing nematode gene sequences can be used to induce RNAi in the feeding nematodes.     

Identification and Nucleotide Sequence of a Tobacco rattle virus RNA-1 Variant that Causes Spraing Disease in Potato cv. Bintje

An isolate of Tobacco rattle virus (TRV) obtained from a site in the Netherlands induced symptoms of spraing disease in tubers of the potato variety Bintje, which is generally considered to be resistant to infection by TRV. The isolate contained two phenotypically distinguishable RNA‐1 variants, one of which was shown to carry the determinant for the ability to cause spraing in cv. Bintje. The nucleotide sequence of the coding region of this RNA‐1 was determined and found to differ at 5.2–5.4% of positions from other TRV RNA‐1 sequences in the database. The amino acid sequences of the predicted translation products were between 92 and 99% identical to those of a TRV RNA‐1 that did not cause spraing in cv. Bintje, with P1b being the most divergent and the replicase read‐through domain the least.     

Virus‐induced gene silencing in Catharanthus roseus by biolistic inoculation of tobacco rattle virus vectors

Catharanthus roseus constitutes the unique source of several valuable monoterpenoid indole alkaloids, including the antineoplastics vinblastine and vincristine. These alkaloids result from a complex biosynthetic pathway encompassing between 30 and 50 enzymatic steps whose characterisation is still underway. The most recent identifications of genes from this pathway relied on a tobacco rattle virus‐based virus‐induced gene silencing (VIGS) approach, involving an Agrobacterium‐mediated inoculation of plasmids encoding the two genomic components of the virus. As an alternative, we developed a biolistic‐mediated approach of inoculation of virus‐encoding plasmids that can be easily performed by a simple bombardment of young C. roseus plants. After optimisation of the transformation conditions, we showed that this approach efficiently silenced the phytoene desaturase gene, leading to strong and reproducible photobleaching of leaves. This biolistic transformation was also used to silence a previously characterised gene from the alkaloid biosynthetic pathway, encoding iridoid oxidase. Plant bombardment caused down‐regulation of the targeted gene (70%), accompanied by a correlated decreased in MIA biosynthesis (45–90%), similar to results obtained via agro‐transformation. Thus, the biolistic‐based VIGS approach developed for C. roseus appears suitable for gene function elucidation and can readily be used instead of the Agrobacterium‐based approach, e.g. when difficulties arise with agro‐inoculations or when Agrobacterium‐free procedures are required to avoid plant defence responses.     

Differential responses between a vector species Bemisia tabaci and a nonvector species Trialeurodes vaporariorum following ingestion of tomato yellow leaf curl virus

In transmitting plant viruses, insect vectors undergo physiological and behavioral alterations. The whitefly Bemisia tabaci is a vector of tomato yellow leaf curl virus (TYLCV), causing severe damages to various horticultural crop plants. To determine whether whitefly alteration is specific to vector species, the responses to TYLCV ingestion were compared between B. tabaci and Trialeurodes vaporariorum, a nonvector for TYLCV. The two species were reared on TYLCV‐infected and noninfected tomato, a host of TYLCV, and their longevity and fecundity were determined while rearing in either tomato or eggplant, a nonhost of TYLCV. TYLCV‐ingested B. tabaci increased their developmental rates but reduced fecundity when they were reared in either tomato or eggplant compared with those of TYLCV‐free ones. In contrast, TYLCV‐ingested T. vaporariorum did not show any of the aforementioned changes when reared on both plant species. In addition, TYLCV‐ingested B. tabaci increased their levels of three heat shock protein genes ( hsp20, hsp70, and hsp90) against thermal stress, whereas TYLCV‐ingested T. vaporariorum did not. The presence of TYLCV virions was identified in two colonies of both species via polymerase chain reaction analysis. TYLCV was detected in the whole body, saliva, and eggs of B. tabaci, while TYLCV was detected only in the whole body but not in the saliva and eggs of T. vaporariorum. The present results strongly indicated that TYLCV specifically manipulate physiological processes of the vector species, B. tabaci.     

The role of weed hosts and the distribution and activity of vector nematodes in the ecology of tobacco rattle virus

At a site in eastern Scotland, nine common species of arable weeds were infected with tobacco rattle virus (TRV), and some of these, notably Viola arvensis and Stellaria media, comprised an overwintering reservoir of the virus. TRV was seed-borne both in naturally and in experimentally infected V. arvensis (2–10%), and occasionally in other weed species. In the glasshouse at 20 ^oC a naturally infective population of vector nematodes (Tricho-dorus spp.) kept in soil free of plants retained its infectivity for 20 wk, although few Trichodorus survived for this period. In the field, the incidence of TRV infection in potato (spraing disease) in plots kept free of weeds for 1–5 years was 3–4 times that in weed-infested plots but Trichodorus numbers did not differ appreciably between the two treatments. Presumably the virus is retained for long periods in its vectors and these feed on potato more frequently when other hosts are not available. Weeds are probably important in the long term as hosts of both TRV and its vectors, but in the short term weed control seems unlikely to prevent potato spraing because of the long persistence of TRV in vector populations. In the field, Trichodorus accumulated near the interface between topsoil and subsoil, and the incidence of spraing was greatest where the topsoil was shallowest. When cucumber seedlings were exposed to virus-carrying Trichodorus, TRV reached a greater concentration in roots at 20 ^oC than at 24 ^oC, and the virus was not detected in roots at 29 ^oC. In a sandy soil, TRV was transmitted only when the water content exceeded 15%, and at least 30 % water was needed for maximum transmission. Annual records of rainfall and spraing disease suggest that spraing is most prevalent when the summer is wettest. TRV is not confined to cultivated land. Stabilized sand dunes supporting a pure stand of Ammophila armaria were colonized by Trichodorus pachyder-mus, but TRV was detected only where the plant community had enlarged to include V. arvensis and other dicotyledons. In such situations, TRV may be introduced in the seed of V. arvensis, and the movement of soil by wind probably contributes to the dispersal of Trichodorus.     

昆虫介体行为与植物病毒的传播

大多数植物病毒都是依赖昆虫介体进行传播,其中超过80%的传毒介体昆虫都是属于半翅目同翅亚目。昆虫介体识别寄主植物和取食的过程与病毒的传播密切相关,本文主要综述了同翅亚目昆虫、蓟马等介体昆虫取食行为与植物病毒的相互作用方面的研究进展,着重于介绍昆虫不同取食阶段的行为对植物病毒传播的影响,病毒侵染对介体取食和识别寄主行为的影响。     

Agrodrench: a novel and effective agroinoculation method for virus-induced gene silencing in roots and diverse Solanaceous species

Summary Virus-induced gene silencing (VIGS) is an extremely powerful tool for plant functional genomics. We used Tobacco rattle virus (TRV)-derived VIGS vectors expressed from binary vectors within Agrobacterium to induce RNA silencing in plants. Leaf infiltration is the most common method of agroinoculation used for VIGS but this method has limitations as it is laborious for large-scale screening and some plants are difficult to infiltrate. Here we have developed a novel and simple method of agroinoculation, called 'agrodrench', where soil adjacent to the plant root is drenched with an Agrobacterium suspension carrying the TRV-derived VIGS vectors. By agrodrench we successfully silenced the expression of phytoene desaturase (PDS), a 20S proteasome subunit (PB7) or Mg-protoporphyrin chelatase (Chl H) encoding genes in Nicotiana benthamiana and in economically important crops such as tomato, pepper, tobacco, potato, and Petunia, all belonging to the Solanaceae family. An important aspect of agrodrench is that it can be used for VIGS in very young seedlings, something not possible by the leaf infiltration method, which usually requires multiple fully expanded leaves for infiltration. We also demonstrated that VIGS functioned to silence target genes in plant roots. The agrodrench method of agroinoculation was more efficient than the leaf infiltration method for VIGS in roots. Agrodrench will facilitate rapid large-scale functional analysis of cDNA libraries and can also be applied to plants that are not currently amenable to VIGS technology by conventional inoculation methods.     

Occurrence and localization of spherical viruslike particles in tissues of apparently healthy tobacco thrips,Frankliniella fusca,a vector of tomato spotted wilt virus

Spherical viruslike particles (VLP) were found in the tissues of apparently healthy tobacco thrips, Frankliniella fusca. The particles occurred in abundance in thrips from Ontario but were absent in thrips from Oklahoma reared under identical conditions. The VLP were not transmissible to any of the seven plant hosts (in four families) of F. fusca suggesting that they may be an insect virus. Transmission of tomato spotted wilt virus (TSWV) by F. fusca, a known vector, was not affected by the presence or absence of the VLP. No TSWV particles were detected in tissues of F. fusca that transmitted TSWV to test plants. The VLP occurred in several internal organs and hemocoele of the thrips and were isolated in vitro by preparing homogenates of gut tissues. Infection of oocytes and presence of VLP in young nymphs suggested transovarial transmission of the particles. The VLP measured 62 ± 4 nm in diameter and usually occurred in dense viroplasms in the cell cytoplasm. Development of the particles within the viroplasms is discussed.     

The association between smokeless tobacco use and pancreatic adenocarcinoma: A systematic review

BackgroundSmokeless tobacco is a possible risk factor for developing pancreatic adenocarcinoma. This systematic review addressed the question: Is there an association between smokeless tobacco use and pancreatic adenocarcinoma diagnosis?MethodsFive electronic databases, grey literature, and citations of relevant articles were searched to identify studies. Six researchers double-reviewed records for inclusion in the review. The information extracted from these studies was selected using criteria outlined in the Newcastle–Ottawa Quality Assessment Scale for observational studies. A qualitative synthesis of included studies was performed.ResultsThe search of electronic databases resulted in a total of 1747 citations. Eleven studies met the inclusion criteria for this review, including three cohort studies, seven case control studies and one study that pooled data from multiple case-control studies. Studies were heterogeneous in their assessment of exposure intensity and ascertainment of outcomes. Quality of the studies varied. Existing investigations of the association of interest appear to exhibit several types of biases including selection bias, information bias and bias in the analysis.ConclusionThe association between smokeless tobacco use and pancreatic adenocarcinoma is inconclusive. More definitive conclusions regarding this relationship await the results of more methodologically rigorous epidemiologic studies.     

Experimental parasite community perturbation reveals associations between Sin Nombre virus and gastrointestinal nematodes in a rodent reservoir host

Individuals are often co-infected with several parasite species, yet measuring within-host interactions remains difficult in the wild. Consequently, the impacts of such interactions on host fitness and epidemiology are often unknown. We used anthelmintic drugs to experimentally reduce nematode infection and measured the effects on both nematodes and the important zoonosis Sin Nombre virus (SNV) in its primary reservoir (Peromyscus spp.). Treatment significantly reduced nematode infection, but increased SNV seroprevalence. Furthermore, mice that were co-infected with both nematodes and SNV were in better condition and survived up to four times longer than uninfected or singly infected mice. These results highlight the importance of investigating multiple parasites for understanding interindividual variation and epidemiological dynamics in reservoir populations with zoonotic transmission potential.     

HIV-1载体的研发沿革

自从科学家于1983年发现了人类免疫性缺陷病毒1(human immunodeficiency virus1,HIV-1)以来,随着对它的研究不断深入,其表达载体的开发也有了长足的进步。与其他逆转录病毒载体相比,如莫罗尼小鼠白血病病毒(murine leukemiavirus,MLV)载体和泡沫病毒(foamyvirus,FV)载体等,HIV-1载体具有诸多独特的优点,因而有着更广泛的应用于临床基因治疗的前景。该文对HIV-1载体的研发过程及其优缺点进行综述。     

Effects of In-furrow and Water-run Oxamyl on Paratrichodorus allius and Corky Ringspot Disease of Potato in the Klamath Basin

Corky ringspot disease (CRS) of potato (Solanum tuberosum) is caused by the tobacco rattle virus (TRV), which is vectored by stubby-root nematodes, Paratrichodorus spp. and Trichodorus spp., and is a significant threat to potato quality and production in many areas of the western United States. Between 2002 and 2005, fields with a history of CRS were planted to potato and treated with various combinations of in-furrow (IF) and chemigated (water run, WR) oxamyl [Methyl N'N'-dimethyl-N-[(methyl carbamoyl)oxy]-1-thiooxamimidate] applications. Soil samples were collected to determine how Paratrichodorus allius populations responded to the various treatment regimes (2002-2004); potato tubers were evaluated for symptoms of CRS in 2004-2005. Applications of oxamyl to potato (1.1 kg a.i./ha) did not cause significant mortality of P. allius but did prevent the populations from increasing. Oxamyl applications that began at 55 days after planting (DAP) or later did not control CRS and were not different from the untreated control. However, application schedules that began early-season, either IF at planting, early WR (33 - 41 DAP), or both, significantly reduced CRS expression in cv. Yukon Gold. Therefore, oxamyl applications must be made early in the growing season to be effective in controlling CRS. Effects of oxamyl on CRS may be due to nematostatic action that suppresses feeding activity during early field season when most virus transmission probably occurs.     

Cauliflower mosaic virus protein P6-TAV plays a major role in alteration of aphid vector feeding behaviour but not performance on infected Arabidopsis

Emerging evidence suggests that viral infection modifies host plant traits that in turn alter behaviour and performance of vectors colonizing the plants in a way conducive for transmission of both nonpersistent and persistent viruses. Similar evidence for semipersistent viruses like cauliflower mosaic virus (CaMV) is scarce. Here we compared the effects of Arabidopsis infection with mild (CM) and severe (JI) CaMV isolates on the feeding behaviour (recorded by the electrical penetration graph technique) and fecundity of the aphid vector Myzus persicae. Compared to mock-inoculated plants, feeding behaviour was altered similarly on CM- and JI-infected plants, but only aphids on JI-infected plants had reduced fecundity. To evaluate the role of the multifunctional CaMV protein P6-TAV, aphid feeding behaviour and fecundity were tested on transgenic Arabidopsis plants expressing wild-type (wt) and mutant versions of P6-TAV. In contrast to viral infection, aphid fecundity was unchanged on all transgenic lines, suggesting that other viral factors compromise fecundity. Aphid feeding behaviour was modified on wt P6-CM-, but not on wt P6-JI-expressing plants. Analysis of plants expressing P6 mutants identified N-terminal P6 domains contributing to modification of feeding behaviour. Taken together, we show that CaMV infection can modify both aphid fecundity and feeding behaviour and that P6 is only involved in the latter.     

Distribution of velvet tobacco mottle virus in its mirid vector and its relationship to transmissibility

Following acquisition by feeding, velvet tobacco mottle virus (VTMoV) was detected in the gut, haemolymph and faeces of the mirid vector, Cyrtopeltis nicotianae, but not in the salivary glands. Virus antigen was detected in the gut and haemolymph for up to nine days following acquisition. Infective virus was detected in the secretions and excretions of the mirids immediately after acquisition and was also detected in the faeces of nymphs after six days. Insoluble nigrosin dye was eliminated intermittently from the gut up to six days after ingestion, in a manner similar to the loss of virus infectivity. Non-infective mirids were able to inoculate plants from infectious sap deposits on the upper epidermis. An ingestion-defecation model of insect transmission in which the salivary glands are not implicated is proposed as one explanation for the persistence of transmission in this mirid-virus association.