Production of nitric oxide in host-virus interaction: A case study with a compatible begomovirus-kenaf host-pathosystem

Nitric oxide (NO) plays a key role in plant diseases resistance. Here we have first time demonstrated that begomovirus infection in susceptible H. cannabinus plants, results in elevated NO and reactive nitrogen species production during early infection stage not only in infected leaf but also in root and shoot. Production of NO was further confirmed by oxyhemoglobin assay. Furthermore, we used Phenyl alanine ammonia lyase as marker of pathogenesis related enzyme. In addition evidence for protein tyrosine nitration during the early stage of viral infection clearly showed the involvement of nitrosative stress.Key words: nitric oxide, mesta yellow vein mosaic virus, protein nitration     

Purification and Characterization of Indian Cassava Mosaic Virus

The Indian cassava mosaic virus (ICMV) was transmitted by the whitefly Bemisia tabaci and sap inoculation. ICMV was purified from cassava and from systemically infected Nicotiana benthamiana leaves. Geminate particles of 16–18 × 30 nm in size were observed by electron microscopy. The particles contained a single major protein of an estimated molecular weight of 34,000. Specific antiserum trapped geminate particles from the extracts of infected cassava and N. benthamiana plants in ISEM test. The virus was detected in crude extracts of infected cassava, ceara rubber, TV. benthamiana and N. tabacum cv. Jayasri plants by ELISA. ICMV appeared serologically related to the gemini viruses of Acalypha yellow mosaic, bhendi yellow vein mosaic, Croton yellow vein mosaic, Dolichos yellow mosaic, horsegram yellow mosaic, Malvastrum yellow vein mosaic and tobacco leaf curl.     

Detection of new viruses in alfalfa,weeds and cultivated plants growing adjacent to alfalfa fields in Saudi Arabia

A total of 1368 symptomatic plant samples showing different virus-like symptoms such as mottling, chlorosis, mosaic, yellow mosaic, vein clearing and stunting were collected from alfalfa, weed and cultivated plant species growing in vicinity of alfalfa fields in five principal regions of alfalfa production in Saudi Arabia. DAS-ELISA test indicated occurrence of 11 different viruses in these samples, 10 of which were detected for the first time in Saudi Arabia. Eighty percent of the alfalfa samples and 97.5% of the weed and cultivated plants samples were found to be infected with one or more of these viruses. Nine weed plant species were found to harbor these viruses namely, Sonchus oleraceus, Chenopodium spp., Hibiscus spp., Cichorium intybus, Convolvulus arvensis, Malva parviflora, Rubus fruticosus, Hippuris vulgaris, and Flaveria trinervia. These viruses were also detected in seven cultivated crop plants growing adjacent to the alfalfa fields including Vigna unguiculata, Solanum tuberosum, Solanum melongena, Phaseolus vulgaris, Cucurbita maxima, Capsicum annuum, and Vicia faba. The newly reported viruses together with their respective percent of detection in alfalfa, and in both weeds and cultivated crop plant species together were as follows: Bean leaf roll virus (BLRV) {12.5 and 4.5%}, Lucerne transient streak virus (LTSV) {2.9 and 3.5%}, Bean yellow mosaic virus (BYMV) {1.4 and 4.5%}, Bean common mosaic virus (BCMV) {1.2 and 4.5%}, Red clover vein mosaic virus (RCVMV) {1.2 and 4%}, White clover mosaic virus (WCIMV) {1.0 and 5%}, Cucumber mosaic virus (CMV) {0.8 and 3%}, Pea streak virus (PeSV) {0.4 and 4.5%} and Tobacco streak virus (TSV) {0.3 and 2.5%}. Alfalfa mosaic virus (AMV), the previously reported virus in alfalfa, had the highest percentage of detection in alfalfa accounting for 58.4% and 62.8% in the weeds and cultivated plants. Peanut stunt virus (PSV) was also detected for the first time in Saudi Arabia with a 66.7% of infection in 90 alfalfa samples collected from the surveyed regions during the last visit that tested negative to all the previously detected viruses.     

Yellow Vein Mosaic Disease in Okra (<i>Abelmoschus esculentus</i> L.): An Overview on Causal Agent,Vector and Management

Okra (Abelmoschus esculentus L.) belongs to the Malvaceae family and is one of the most essential and popular vegetables globally. It is rich in proteins, carbohydrates, and vitamins. Abiotic and biotic factors threaten okra productivity. Okra yellow vein mosaic disease (OYVMD) is the most destructive disease of okra. The causal agent, [(i.e., Okra yellow vein mosaic virus (OYVMV)] of this disease belongs to the family Geminiviridae and genus Begomovirus. OYVMV is a monopartite with additional ssDNA molecule. This virus has two components DNA-A for protein coding and DNA-B for symptoms induction. Whitefly transmits OYVMV in persistent manner. Characteristic symptoms of OYVMV infected okra plants are chlorosis, dwarfing, and yellowing of veins and fruits. High temperatures with moderate rainfall enhance the development of OYVMV disease and the whitefly population. However, high humidity with low temperature and rainfall has no significant role in developing the OYVMD and whitefly population. Moreover, the virus also affects the secondary metabolites in the infected okra plants. The virus can be managed through various strategies including the application of plant defense activators, the development of resistant varieties and by controlling its vector via pesticides and plant extracts. Various plant defense activators such as monopotassium phosphate (KH₂PO), salicylic acid, benzoic acid, and citric acid enhance resistance in okra against OYMVD. In addition, the resistance to OYMVD can also be achieved by successfully incorporating high yielding but resistant cultivars of acceptable quality. In this review, we have discussed history, economic impact, symptomology, disease development under a natural environment, genetics and management of OYVMV.     

Viruses infecting winter oilseed rape (Brassica napus ssp. oleifera)

Turnip mosaic virus (TuMV) and cauliflower mosaic virus (CaMV) have been found infecting field crops of winter oilseed rape (Brassica napus ssp. oleifera) in South Warwickshire. Other viruses found include broccoli necrotic yellows virus (BNYV) and a member of the beet western yellows virus group. Systemic leaf symptoms caused by TuMV varied within and between cultivars; the three predominant reaction types were classified as necrotic, mosaic and immune. Some recently introduced cultivars of oilseed rape were more severely affected by TuMV infection than older cultivars. Reactions to CaMV were less varied and immunity was not found. The seed yield from TuMV and CaMV-infected plants was less than that of healthy control plants. This effect was due to infected plants producing either fewer seeds, smaller seeds or both. Germination of seeds from infected plants was unaffected if sown soon after harvest. After storage for one year the germination of seed from a virus infected plant was significantly less than that of seed from a virus-free plant. All commercial cultivars tested were experimentally susceptible to turnip yellow mosaic virus (TYMV) and some American strains of cucumber mosaic virus (CMV).     

Molecular and biochemical characterization of mungbean yellow mosaic India virus resistance in leguminous host <Emphasis Type="Italic">Vigna mungo</Emphasis>

Mungbean yellow mosaic India virus (MYMIV)—the causal agent of the yellow mosaic disease is responsible for severe damage of crops that are of great economic importance. In the current study, we explored the process of MYMIV infection and its natural resistance by analysing the expression of early and late viral genes at different time points in the leaves of resistant and susceptible Vigna mungo plants. Accordingly, we have periodically evaluated several biochemical parameters commonly associated with oxidative status of resistant and susceptible V. mungo plants during MYMIV infection. Our study revealed that accumulation levels of the early as well as late expressed genes of MYMIV were low and high in the resistant and susceptible plants, respectively; whereas membrane stability index (MSI) exhibited an opposite response. Moreover, a decrease in the malondialdehyde levels along with an increase in the activities/levels of different antioxidant enzymes, total phenol and H₂O₂ was noted during the early stages of infection in the resistant plants. Such observations argue in favour of strong defensive capability of the resistant plants in restricting the accumulation of viral RNA and generation of harmful free radicals within the studied tissue. Collectively, it appears that obstruction of viral invasion in plant cell wall, restriction in viral DNA replication, and early onset of antioxidant defense responses altogether might be responsible for MYMIV natural resistance. Such information is helpful in understanding the pathogenesis of MYMIV infection and its resistance in V. mungo and other economically important crops.     

Recognition and differentiation of seven whitefly-transmitted geminiviruses from India, and their relationships to African cassava mosaic and Thailand mung bean yellow mosaic viruses

Particles resembling those of geminiviruses were found by immunosorbent electron microscopy in extracts of plants infected in India with bhendi yellow vein mosaic, croton yellow vein mosaic, dolichos yellow mosaic, horsegram yellow mosaic, Indian cassava mosaic and tomato leaf curl viruses. All these viruses were transmitted by Bemisia tabaci whiteflies, all reacted with at least one out of ten monoclonal antibodies to African cassava mosaic virus (ACMV), and all reacted with a probe for ACMV DNA-1, but scarcely or not at all with a full-length probe for ACMV DNA-2. Most of the viruses were distinguished by their host ranges when transmitted by whiteflies, and the rest could be distinguished by their pattern of reactions with the panel of monoclonal antibodies. Horsegram yellow mosaic virus was distinguished from Thailand mung bean yellow mosaic virus by its lack of sap transmissibility, ability to infect Arachis hypogaea, failure to react strongly with the probe for ACMV DNA-2 and its pattern of reactions with the monoclonal antibodies. Structures resembling a ‘string of pearls’, but not geminate particles, were found in leaf extracts containing malvastrum yellow vein mosaic virus. Such extracts reacted with two of the monoclonal antibodies, suggesting that this whitefly-transmitted virus too is a geminivirus. All seven viruses from India can therefore be considered whitefly-transmitted geminiviruses.     

Diagnosis of Symptomless Yellow mosaic begomovirus Infection in Pigeonpea by Using Cloned Mungbean yellow mosaic India virus as Probe

One isolate of Mungbean yellow mosaic India virus (MYMIV) of mungbean plants from Sri Ganganagar, Rajasthan, designated as MYMIV-Mg was isolated and DNA-A and DNA-B, the two full length bipartite genomic components of this virus, were cloned. The [α-³²P] labeled diagnostic probes specific to these cloned DNA-A and -B of MYMIV-Mg were used to detect the virus infection in infected plants by nucleic acid spot hybridization (NASH) test. The NASH tests detected the MYMIV infection and concentration of viral titre in susceptible, moderately susceptible, resistant and symptomless genotypes of pigeonpea (Cajanus cajan) plants. Fourteen genotypes of pigeonpea were tested against five naturally occurring MYMIV variants viz.,.MYMIV Bg, -MgD, -MoL, -Mg and -Pp1 through viruliferous whitefly (Bemisia tabaci) transmission in greenhouse condition. Disease incidence and severity of MYMIV in different pigeonpea genotypes varied with the variants of MYMIV. Many genotypes of pigeonpea did not produce visible yellow mosaic symptoms after inoculation with MYMIV variants MYMIV-Bg, -MbD and -MoL, although, majority of the symptomless genotypes were found to be infected by MYMIV, as viral DNA was detected by NASH test.     

Small-RNA Deep Sequencing Reveals Arctium tomentosum as a Natural Host of Alstroemeria virus X and a New Putative Emaravirus

Background

Arctium species (Asteraceae) are distributed worldwide and are used as food and rich sources of secondary metabolites for the pharmaceutical industry, e.g., against avian influenza virus. RNA silencing is an antiviral defense mechanism that detects and destroys virus-derived double-stranded RNA, resulting in accumulation of virus-derived small RNAs (21–24 nucleotides) that can be used for generic detection of viruses by small-RNA deep sequencing (SRDS).

Methodology/Principal Findings

SRDS was used to detect viruses in the biennial wild plant species Arctium tomentosum (woolly burdock; family Asteraceae) displaying virus-like symptoms of vein yellowing and leaf mosaic in southern Finland. Assembly of the small-RNA reads resulted in contigs homologous to Alstroemeria virus X (AlsVX), a positive/single-stranded RNA virus of genus Potexvirus (family Alphaflexiviridae), or related to negative/single-stranded RNA viruses of the genus Emaravirus. The coat protein gene of AlsVX was 81% and 89% identical to the two AlsVX isolates from Japan and Norway, respectively. The deduced, partial nucleocapsid protein amino acid sequence of the emara-like virus was only 78% or less identical to reported emaraviruses and showed no variability among the virus isolates characterized. This virus—tentatively named as Woolly burdock yellow vein virus—was exclusively associated with yellow vein and leaf mosaic symptoms in woolly burdock, whereas AlsVX was detected in only one of the 52 plants tested.

Conclusions/Significance

These results provide novel information about natural virus infections in Acrtium species and reveal woolly burdock as the first natural host of AlsVX besides Alstroemeria (family Alstroemeriaceae). Results also revealed a new virus related to the recently emerged Emaravirus genus and demonstrated applicability of SRDS to detect negative-strand RNA viruses. SRDS potentiates virus surveys of wild plants, a research area underrepresented in plant virology, and helps reveal natural reservoirs of viruses that cause yield losses in cultivated plants.     

Tomato yellow leaf curl virus infection of a resistant tomato line with a silenced sucrose transporter gene LeHT1 results in inhibition of growth, enhanced virus spread, and necrosis

To identify genes involved in resistance of tomato to Tomato yellow leaf curl virus (TYLCV), cDNA libraries from lines resistant (R) and susceptible (S) to the virus were compared. The hexose transporter LeHT1 was found to be expressed preferentially in R tomato plants. The role of LeHT1 in the establishment of TYLCV resistance was studied in R plants where LeHT1 has been silenced using Tobacco rattle virus-induced gene silencing (TRV VIGS). Following TYLCV inoculation, LeHT1-silenced R plants showed inhibition of growth and enhanced virus accumulation and spread. In addition, a necrotic response was observed along the stem and petioles of infected LeHT1-silenced R plants, but not on infected not-silenced R plants. This response was specific of R plants since it was absent in infected LeHT1-silenced S plants. Necrosis had several characteristics of programmed cell death (PCD): DNA from necrotic tissues presented a PCD-characteristic ladder pattern, the amount of a JNK analogue increased, and production of reactive oxygen was identified by DAB staining. A similar necrotic reaction along stem and petioles was observed in LeHT1-silenced R plants infected with the DNA virus Bean dwarf mosaic virus and the RNA viruses Cucumber mosaic virus and Tobacco mosaic virus. These results constitute the first evidence for a necrotic response backing natural resistance to TYLCV in tomato, confirming that plant defense is organized in multiple layers. They demonstrate that the hexose transporter LeHT1 is essential for the expression of natural resistance against TYLCV and its expression correlates with inhibition of virus replication and movement.     

A yellow mosaic disease of horse chestnut (Aesculus spp.) caused by apple mosaic virus

A severe foliar yellow mosaic disease was observed in horse chestnut trees (Aesculus carnea and A. hippocastanum). Reactions in woody indicator plants grafted with diseased horse chestnut suggested the presence of an ilarvirus. Virus isolates obtained by mechanical inoculation of herbaceous test plants reacted with antisera to apple mosaic virus but not with antisera to its serotype prunus necrotic ringspot virus, or to prune dwarf virus. Yellow mosaic was induced in horse chestnut seedlings grafted with tissues from herbaceous hosts infected with horse chestnut isolates or with the European plum line pattern isolate of apple mosaic virus. Virus was detected by enzyme-linked immunosorbent assay (ELISA) in embryo and endosperm of immature seed from infected trees but not in mature seed, or progeny seedlings. Strawberry latent ringspot virus was detected in one of six A. hippocastanum trees with a leaf vein yellows disease.     

Effects of turnip mosaic virus and turnip yellow mosaic virus on the survival, growth and reproduction of wild cabbage (Brassica oleracea)

Wild plants of Brassica oleracea (wild cabbage) are commonly infected with turnip mosaic poty virus (TuMV), turnip yellow mosaic tymovirus (TYMV) and several other viruses. A field experiment in which plants were inoculated either with TuMV or TYMV showed that virus infection significantly reduced survival, growth and reproduction. Relative to water inoculated-controls, plants infected with TYMV had greater mortality, were shorter, had a smaller leaf area and number, showed a greater amount of damage from herbivory and chlorosis, were less likely to flower and produced fewer pods and lower total seed output. Plants infected with TuMV did not appear to be adversely affected at first; however, mortality after 18 months was higher than control plants. Although TuMV infection had no effect on the number of plants flowering, the infected plants did produce fewer pods and a lower total seed output. We conclude that both viruses can significantly affect vegetative and reproductive performance of wild cabbage and hence that introgression of virus resistance (particularly when conferred by a major gene or a transgene) from a crop might increase plant fitness in natural populations of this species. Ecological risk assessments of virus resistance transgenes must do more than survey adult plants in natural populations for the presence of the target virus. Failure to detect the virus could be due to high mortality on infection with the virus.     

Involvement of Volatile Substances in Systemic Resistance of Barley Against Erysiphe graminis f. sp. hordei Induced by Pruning of Leaves

Horsegram yellow mosaic disease was shown to be caused by a geminivirus; horsegram yellow mosaic virus (HYMV). The virus could not be transmitted by mechanical sap inoculation. Leaf dip and purified virus preparations showed geminate virus particles, measuring 15-18 * 30 nm. An antiserum for HYMV was produced and in enzyme-linked immunosorbent assay (ELISA) and immunosorbent electron microscopy (ISEM) tests HYMV was detected in leaf extracts of fieldinfected bambara groundnut, french bean, groundnut, limabean, mungbean, pigeonpea and soybean showing yellow mosaic symptoms. Bemisia tabaci fed on purified HYMV through a parafilm membrane transmitted the virus to all the hosts listed above but not to Ageratum conyzoides, okra, cassava, cowpea, Croton bonplandianus, Lab-lab purpureus, Malvastrum coromandalianum and tomato. No reaction was obtained in ELISA and ISEM tests between HYMV antibodies and extracts of plants diseased by whitefly-transmitted agents in India such as A. conyzoides yellow mosaic, okra yellow vein mosaic, C. bonplandianus, yellow vein mosaic, M. coromandalianum yellow vein mosaic, tomato leaf curl and cassava mosaic. HYMV was also not found to be related serologically to bean golden mosaic, virus.     

Detection of begomovirus associated with okra leaf curl disease

Leaf curl and yellow vein mosaic viral disease is the major constraint on okra (Abelmoschus esculentus L.) production in India. Amplified fragment sequence of DNA-β showed highest similarity of 91.7% with Bhendi yellow vein mosaic virus-Tamil Nadu (AJ308425, NC_003405) and lowest similarity of 48.5% with OKLCV (NC_004093), whereas coat protein specific amplified sequence showed highest homology with isolate of Madurai, Haryana, Ludhiana and lowest homology of 92% with Mesta yellow vein mosaic Bahraich virus (MYVMBV) (EU360303). The results obtained in the present study confirm that both the viral diseases of okra reported in southern India are caused by a begomovirus associated with DNA-β in which the plants show leaf curl symptoms and never develops yellow vein mosaic and those plants which show yellow vein mosaic, never develops leaf curl symptoms even in the same rows and field. The okra leaf curl is an emerging virus disease in India.     

Antibody-mediated improved resistance to ClYVV and PVY infections in transgenic tobacco plants expressing a single-chain variable region antibody

Transgenic Nicotiana tabacum plants expressing a single-chain variable region antibody fragment derived from a broad-spectrum monoclonal antibody 3-17 showed suppression of virus infection following challenge by two distinct potyviruses: potato virus Y strain D, and clover yellow vein virus strain 300. Monoclonal antibody 3-17, which was raised against the potyvirus Johnsongrass mosaic virus, was shown to react strongly with 14 potyvirus species. Two different single-chain antibody constructs were used to produce chimeric genes encoding recombinant proteins designed to be targeted either to the apoplasm or to the cytoplasm. Transgenic plant lines showed reduced numbers of local lesions and systemic symptoms when challenged with potato virus Y, strain D and reduced local lesions following challenge with clover yellow vein virus, strain 300. The level of suppression conferred by the transgene when plants were challenged under laboratory conditions with high concentrations of virus, together with the ability of the transgene to partially protect plants against distinct viruses suggest that one single-chain gene construct might be used to protect plants from distinct potyviruses.     

Aphid behavioral responses to virus‐infected plants are similar despite divergent fitness effects

We compared the settling preferences and reproductive potential of an oligophagous herbivore, the pea aphid, Acyrthosiphon pisum Harris (Hemiptera: Aphididae), in response to pea plants, Pisum sativum L. cv. ‘Aragorn’ (Fabaceae), infected with two persistently transmitted viruses, Pea enation mosaic virus (PEMV) and Bean leaf roll virus (BLRV), that differ in their distribution within an infected plant. Aphids preferentially oriented toward and settled on plants infected with PEMV or BLRV in comparison with sham‐inoculated plants (plants exposed to herbivory by uninfected aphids), but aphids did not discriminate between plants infected with the two viruses. Analysis of plant volatiles indicated that plants inoculated with either virus had significantly higher green leaf volatile‐to‐monoterpene ratios. Time until reproductive maturity was marginally influenced by plant infection status, with a trend toward earlier nymph production on infected plants. There were consistent age‐specific effects of plant infection status on aphid fecundity: reproduction was significantly enhanced for aphids on BLRV‐infected plants across most time intervals, though mean aphid fecundity did not differ between sham and PEMV‐infected plants. There was no clear pattern of age‐specific survivorship; however, mean aphid lifespan was reduced on plants infected with PEMV. Our results are consistent with predictions of the host manipulation hypothesis, extended to include plant viruses: non‐viruliferous A. pisum preferentially orient to virus‐infected host plants, potentially facilitating pathogen transmission. These studies extend the scope of the host manipulation hypothesis by demonstrating that divergent fitness effects on vectors arise relative to the mode of virus transmission.     

Developing reverse genetics systems of northern cereal mosaic virus to reveal superinfection exclusion of two cytorhabdoviruses in barley plants

Recently, reverse genetics systems of plant negative‐stranded RNA (NSR) viruses have been developed to study virus–host interactions. Nonetheless, genetic rescue of plant NSR viruses in both insect vectors and monocot plants is very limited. Northern cereal mosaic virus (NCMV), a plant cytorhabdovirus, causes severe diseases in cereal plants through transmission by the small brown planthopper (SBPH, Laodelphax striatellus) in a propagative manner. In this study, we first developed a minireplicon system of NCMV in Nicotiana benthamiana plants, and then recovered a recombinant NCMV virus (rNCMV‐RFP), with a red fluorescent protein (RFP) insertion, in SBPHs and barley plants. We further used rNCMV‐RFP and green fluorescent protein (GFP)‐tagged barley yellow striate mosaic virus (rBYSMV‐GFP), a closely related cytorhabdovirus, to study superinfection exclusion, a widely observed phenomenon in dicot plants rarely studied in monocot plants. Interestingly, cellular superinfection exclusion of rBYSMV‐GFP and rNCMV‐RFP was observed in barley leaves. Our results demonstrate that two insect‐transmitted cytorhabdoviruses are enemies rather than friends at the cellular level during coinfections in plants.