Identification and Functional Analysis of Four RNA Silencing Suppressors in Begomovirus Croton Yellow Vein Mosaic Virus

Croton yellow vein mosaic virus (CYVMV), a species in the genus Begomovirus, is a prolific monopartite begomovirus in the Indian sub-continent. CYVMV infects multiple crop plants to cause leaf curl disease. Plants have developed host RNA silencing mechanisms to defend the threat of viruses, including CYVMV. We characterized four RNA silencing suppressors, namely, V2, C2, and C4 encoded by CYVMV and betasatellite-encoded C1 protein (βC1) encoded by the cognate betasatellite, croton yellow vein betasatellite (CroYVMB). Their silencing suppressor functions were verified by the ability of restoring the β-glucuronidase (GUS) activity suppressed by RNA silencing. We showed here for the first time that V2 was capable of self-interacting, as well as interacting with the V1 protein, and could be translocalized to the plasmodesmata in the presence of CYVMV. The knockout of either V2 or V1 impaired the intercellular mobility of CYVMV, indicating their novel coordinated roles in the cell-to-cell movement of the virus. As pathogenicity determinants, each of V2, C2, and C4 could induce typical leaf curl symptoms in Nicotiana benthamiana plants even under transient expression. Interestingly, the transcripts and proteins of all four suppressors could be detected in the systemically infected leaves with no correlation to symptom induction. Overall, our work identifies four silencing suppressors encoded by CYVMV and its cognate betasatellite and reveals their subcellular localizations, interaction behavior, and roles in symptom induction and intercellular virus movement.

Molecular evolution and genetic analysis of Mesta yellow vein mosaic virus and associated betasatellites

Mesta yellow vein mosaic disease (MYVMD), one of the major diseases circulating mesta growing regions of Indian sub-continent, is responsible for serious yield loss in mesta crops. A complex of monopartite begomovirus, Mesta yellow vein mosaic virus (MYVMV) and associated betasatellite, is reported in several studies as the causal agent of MYVMD. However, all-inclusive molecular evolutionary analysis of so far available MYVMVs and associated betasatellites disseminating in this region is still lacking. In this study, by estimating and analyzing various indexes of population genetics and evolutionary parameters, we discussed the sources of genetic variations, population dynamics and different forces acting on the evolution of MYVMVs and associated betasatellites. The study finds recombination as a vital force in the evolution and diversification of begomovirus complexes in different geographic locations however, betasatellites were found to be exposed to more diverse recombination events compared to MYVMVs. Indian isolates are reported to have high frequency of polymorphism in this study which suggests a balancing selection or expansion occurring in Indian populations of begomoviruses. Higher degree of genetic differentiation and lower rate of gene flow calculated between the viral populations of Bangladesh and Pakistan is justified by the relatively far geographical distance between these two countries. Although the study detects overall purifying selection, the degrees of constraints acting on individual gene tested are found different. Coat protein (AV1) is estimated with very high nucleotide substitution rate which is very likely to result from the strongest purifying selection pressure (dN/dS = 0.131) calculated in this study on coat protein. The findings of this study on different evolutionary forces that shape the emergence and diversification of MYVMVs and associated betasatellites may provide directions towards future evolutionary trend analysis and development of comprehensive disease control strategies for begomoviruses.

Management of Okra Yellow Vein Mosaic Virus (OYVMV) Through Selected Insecticides and Light Reflecting Colored Mulches

A field experiment was conducted to evaluate the effect of light reflecting colored mulches and only one time spraying with selected insecticides to control the insect vector of Okra Yellow Vein Mosaic Virus (OYVMV) whitefly (Bemisia tabaci) at the central farm of Sher-e-Bangla Agricultural University, Dhaka-1207. The experiment was carried out in three blocks layouted with RCBD comprising eight treatments viz. T0 (control/no spray), T1 (1 time spray with imitaf), T2 (1 time spray with protect), T3 (1 time spray with tiddo plus), T4 (1 time spray with terbine), T5 (light reflecting silver color mulch), T6 (light reflecting red color mulch) and T7 (light reflecting black color mulch) with three replications. The mulches were used before sowing and insecticides were sprayed at 30 DAS. It was observed that among the light reflecting colored mulches, the lowest disease incidence (11.48%) was recorded in the plots which mulched with red color mulch (T6). Among the insecticides, the lowest disease incidence (30.17%) was recorded in plots that was sprayed with terbine (one time). Yield and yield contributing characters, morphological and physiological features of okra plant that changes due to disease infection which cause damages in okra production and reduce the fruit quality as well as market value was also the part of this study. Yield and yield contributing characters showed significant variance among the selected treatments. In the relationship study, it was noticed that the yield and plant height was showed negative relationship with disease incidence. However, considering all measuring parameters, spraying with terbine (one time) and used red color mulch may be recommended as good management approaches that will give higher okra production and lower disease incidence of Okra Yellow Vein Mosaic Virus (OYVMV).

Bhendi yellow vein mosaic virus and bhendi yellow vein mosaic betasatellite cause enation leaf curl disease and alter host phytochemical contents in okra

Whitefly-transmitted begomoviruses cause severe diseases in numerous economically important dicotyledonous plants. In recent years, okra enation leaf curl disease (OELCuD) emerged as a serious threat to okra (Abelmoschus esculentus L. Moench) cultivation in the Indian subcontinent. The present study reports the association of a monopartite begomovirus (bhendi yellow vein mosaic virus - BYVMV) and betasatellite (bhendi yellow vein mosaic betasatellite - BYVB) with OELCuD in the Mau region of Uttar Pradesh, India. The BYVMV alone inoculated N benthamiana and A esculentus cv. Pusa Sawani plants developed mild symptoms. Co-inoculation of BYVMV and BYVB resulted in a reduced incubation period, an increased symptom severity and an enhanced BYVMV accumulation (by Southern hybridization and qPCR). This is the first study which satisfies Koch’s postulates for OELCuD in its natural host. Activities of various antioxidative enzymes were significantly increased in the virus inoculated okra plants. Differential responses in various biochemical components (such as photosynthetic pigments, phenol, proline, sugar) in diseased okra plants were observed. This change in phytochemical responses is of significant importance in understanding its impact on virus pathogenesis and disease development.

Determining genetic combining ability, heterotic potential and gene action for yield contributing traits and Yellow Vein Mosaic Virus (YVMV) resistance in Okra [Abelmoschus esculentus (L.) Monech.]

Breeding for resistance to biotic stress and higher yield is a continuous process. Thus, the identification of desirable parents with good combining ability and nature of gene action for the target trait is of utmost importance. Hence, in this present investigation, 10 lines and three testers of Okra were crossed in line × tester mating design to generate 30 testcross progenies and their evaluation along with parents and check in a randomized complete block design with three replications. To depict the true picture of genetic variation among the parental genotypes, molecular diversity analysis was also carried out using genomic-simple sequence repeats before crossing to ascertain that sufficient variability is present among the parents. The molecular analysis grouped the parental genotypes into four clusters (I–IV). The analysis of variance revealed that all the treatments were significant for most of the traits. The combining ability analysis suggested Pusa A-4 as the best general combiner for earliness, Pusa Bhindi-5 for high yield, and DOV-92 for fruit length, plant height, yield per plant, and coefficient of infection for Yellow Vein Mosaic Virus Disease resistance. Similarly, the specific combining ability analysis suggested that the cross combinations DOV-92 × Pusa Bhindi-5 followed by DOV-92 × Pusa A-4 and DOV-92 × Pusa Sawani exhibit high economic heterosis for yield per plant as well as for disease resistance. Finally, estimation of the degree of dominance and predictability ratio was also worked out which indicated the prevalence of non-additive gene action for most of the traits pointing towards sufficient scope for heterosis breeding in Okra.

EVALUATION OF ORGANIC AMENDMENTS AND INSECTICIDES AGAINST OKRA YELLOW VEIN MOSAIC VIRUS AND ITS VECTOR

Okra (Abelmoschus esculentus L.) is susceptible to number of diseases like Yellow Vein Mosaic Virus disease, Damping Off, Fusarium wilt, Powdery Mildew and Enation Leaf Curl. The Okra is favorite host of whitefly which transmits the okra Yellow Vein Mosaic Virus (OYVMV). OYVMV disease causes considerable yield losses in Okra. Experiment was conducted to check the efficacy of organic matter (poultry manure, leaves) and insecticides (Acetamiprid, Diafenthiuron, Pyridine Carboxamide) against OYVMV and its whitefly vector. In this experiment, ten okra varieties were sown in randomized complete block design (RCBD). Organic matter and insecticides were applied at recommended doses (@5ml/L) on weekly intervals. Among insecticides, Acetamiprid showed the best result to manage whitefly infestation with 60.01% mortality and OYVMV disease severity with 53.98% efficiency. Polo (Diafenthiuron) also showed good results to manage whitefly (56.36%) and OYVMVD (46.34%), but Ulala (Pyridine Carboxamide) insecticide was least effective to control whitefly population (47.27%) and to manage the infection of OYVMV disease (43.91%). Leaf manure was more effective with 61.16% efficiency as compared to poultry manure (56.01%) against OYVMV disease severity. New chemistry insecticide (acetamiprid) gave the most effective control of whitefly and transmission of OYVMV and could be used in place of conventional insecticides. Leaf manure proved significant defense activator in okra plants with effective control of disease and could be used as eco-friendly management approach.