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Begomoviruses (family Geminiviridae) are known for causing devastating diseases in fruit, fibre, pulse, and vegetable crops throughout the world. Begomoviruses are transmitted in the field exclusively through insect vector whitefly (Bemisia tabaci), and the frequent outbreaks of begomoviruses are attributed largely due to the abundance of whitefly in the agri-ecosystem. Begomoviruses being phloem-borne were known not be transmitted through seeds of the infected plants. The recent findings of seed transmission of begomoviruses brought out a new dimension of begomovirus perpetuation and dissemination. The first convincing evidence of seed transmission of begomoviruses was known in 2015 for sweet potato leaf curl virus followed by several begomoviruses, like bhendi yellow vein mosaic virus, bitter gourd yellow mosaic virus, dolichos yellow mosaic virus, mungbean yellow mosaic virus, mungbean yellow mosaic India virus, pepper yellow leaf curl Indonesia virus, tomato leaf curl New Delhi virus, tomato yellow leaf curl virus, tomato yellow leaf curl Sardinia virus, and okra yellow mosaic Mexico virus. These studies brought out two perspectives of seed-borne nature of begomoviruses: (i) the presence of begomovirus in the seed tissues derived from the infected plants but no expression of disease symptoms in the progeny seedlings and (ii) the seed infection successfully transmitted the virus to cause disease to the progeny seedlings. It seems that the seed transmission of begomovirus is a feature of a specific combination of host-genotype and virus strain, rather than a universal phenomenon. This review comprehensively describes the seed transmitted begomoviruses reported in the last 9 years and the possible mechanism of seed transmission. An emphasis is placed on the experimental results that proved the seed transmission of various begomoviruses, factors affecting seed transmission and impact of begomovirus seed transmission on virus circulation, outbreak of the disease, and management strategies.

Aluminium is preferred over iron for various uses as it is lighter, rust-proof and has an exceptional strength-to-weight ratio. Aluminium is more reactive than iron because of its higher standard reduction potential but it forms a self-protective layer of Al2O3 due to its higher reducing potential. Still, aluminium is corroded in acidic and alkaline media. The present research paper reports the corrosion behaviour of aluminium in 0.1 M NaOH solution with Alizarin Red S (ARS) or Xylenol Orange (XO) as an inhibitor. Corrosion potential, Corrosion current, Tafel slopes (βc) and (βa) for aluminium corrosion have been potentiodynamically investigated. The weight loss method has also been used to calculate the inhibition efficiencies of these additives. The corroded surfaces have been characterized by SEM images. It is found that corrosion potential values change significantly towards the anodic direction in the presence of ARS but they are sifted towards a cathodic direction at a high concentration of XO. While ARS shows a corrosion inhibition effect, XO shows a reverse effect. The inhibition efficiency values of ARS and XO calculated from corrosion current in potentiodynamic polarization studies tally with those calculated from weight loss studies. Studies on a laboratory model of aluminium-air batteries in 0.1 M NaOH containing these organic additives show that the discharge curve of the cell is improved in the presence of ARS and XO in combination. The results are explained in terms of the formation of well-known Al+3.3ARS− and Al+3-XO6− complexes. At room temperature, 1.0 × 10−3 M solution of ARS is found effective in inhibiting the corrosion of aluminium while the same concentration of XO is acting as a corrosion accelerator. This interesting study can be very vital in the field of corrosion science.

In tropical countries, combating leaf curl disease in hot peppers has become important in improvement programs. Leaf curl disease is caused by whitefly (Bemisia tabaci) transmitted begomoviruses, which mainly include chilli leaf curl virus (ChiLCV). However, multiple begomoviruses have also been found to be associated with this disease. The Capsicum annuum line, DLS-Sel-10, was found to be a tolerant source against this disease during field screening. In this study, we characterized the resistance of DLS-sel-10 against chilli leaf curl virus (ChiLCV) in comparison to the susceptible cultivar Phule Mukta (PM), focusing on the level, stage, and nature of resistance. Comprehensive investigations involved screening of DLS-Sel-10 against the whitefly vector ChiLCV. The putative tolerant line displayed reduced virus infection at the seedling stage, with increasing resistance during vegetative, flowering, and fruiting stages. Both DLS-Sel-10 and PM could be infected with ChiLCV, although DLS-Sel-10 remained symptomless. Insect feeding assays revealed DLS-Sel-10 as a less preferred host for whiteflies compared to PM. In conclusion, DLS-Sel-10 demonstrated tolerance not only to ChiLCV but also served as an unfavorable host for the whitefly vector. The study highlighted an age-dependent increase in tolerance within DLS-Sel-10, showcasing its potential for effective leaf curl disease management in chilli.

Expanding possibilities for foreign gene expression in cucurbits, we present a novel approach utilising a bipartite vector system based on the cucumber green mottle mosaic virus (CGMMV) genome. Traditional full-length CGMMV vectors face limitations such as a restricted cargo capacity and unstable foreign gene expression. To address these challenges, we developed two ‘deconstructed’ CGMMV genomes, DG-1 and DG-2. DG-1 features a major internal deletion, resulting in the loss of crucial replicase enzyme domains, rendering it incapable of self-replication. However, a staggered infiltration of DG-1 in CGMMV-infected plants enabled successful replication and movement, facilitating gene-silencing experiments. Conversely, DG-2 was engineered to enhance replication rates and provide multiple cloning sites. Although it exhibited higher replication rates, DG-2 remained localised within infiltrated tissue, displaying trans-replication and restricted movement. Notably, DG-2 demonstrated utility in expressing GFP, with a peak expression observed between 6 and 10 days post-infiltration. Overall, our bipartite system represents a significant advancement in functional genomics, offering a robust tool for foreign gene expression in Nicotiana benthamiana.

Potato virus M (PVM) is one of the most prevalent viruses infecting potatoes worldwide, showing a wide range of diversity in their populations; however, the diversity and genome information of PVM occurring in India is hardly known. The present study serologically detected the PVM in 22.8% of leaf samples collected from the potato fields, generated 13 coat protein (CP) genes and one complete genome sequence for the isolates from India, and identified four differential hosts confirming PVM-Del-144 as a distinct strain of PVM occurring in India. The phylogenetic analyses conducted based on the CP gene sequences (14 from India and 176 from other countries) suggested the existence of three evolutionary divergent lineages (PVM-o, PVM-d, and a new divergent group) in the PVM population, where isolates from India belong to only two clusters (PVM-o and PVM-d) within four sub-clusters. High levels of nucleotide diversity (0.124) and genetic distance (0.142) recorded among the isolates from India may be due to the deviation from the neutral evolution and experiencing population expansion in the past. The complete genome of the isolate Del-144 (KJ194171; 8,526 nucleotides) shared 92.2–93.9% nt sequence identity with the population of PVM-o, whereas it shared only 70.2–72.1% identity with PVM-d. In the phylogenetic analyses, Del-144 clustered with the isolates of PVM-o; however, it formed a separate branch away from all other isolates, indicating the diversity of the strain. Overall, this study revealed the diversity of the isolates of PVM from India and reported the first complete genome sequence of a distinct strain of PVM occurring in India.

BACKGROUND: Ventral hernia (primary and incisional) is one of the most frequently encountered problems by the General Surgeon. The enhanced view totally extraperitoneal (eTEP) technique for inguinal hernia was first described by J Daes, which was later used in ventral hernia repair for the first time by Belyansky et al. We applied the same technique for ventral hernia repair to see its feasibility in the context of our country. MATERIALS AND METHODS: It is a single-center prospective observational study of a patient who underwent eTEP (with or without TAR) by a single surgeon between September 2019 and May 2020 and followed up for 12 months. Patient demographics, hernia characteristics, operative details, perioperative complications, and satisfaction scores using the Likert scale were collected for data analysis. RESULTS: The study included 42 patients with 66% of female among them. The mean age was 51 ± 12.97 years, the mean BMI was 29.88 ± 2.16 kg/m2, and the mean ASA of patients was 1.5. Incisional hernia (74%) was the most common diagnosis. The mean operative time was less for the primary hernia (125.45 min) than for the incisional hernia (138.35 min). Intraoperative complications were seen in three (7%) of all the patients. Postoperative length of hospital stay was 1.8 days on average. Postoperative pain was low with only one of the patients having chronic pain while none had a recurrence on follow-up. All the patients were satisfied. CONCLUSION: eTEP is a feasible, cheap, and safe alternative MIS option as it has less morbidity and does not carry an extra cost of mesh in comparison to conventional procedures.

Molecular cloning, a crucial prerequisite for engineering plasmid constructs intended for functional genomic studies, relies on successful restriction and ligation processes. However, the lack of unique restriction sites often hinders construct preparation, necessitating multiple modifications. Moreover, achieving the successful ligation of large plasmid constructs is frequently challenging. To address these limitations, we present a novel PCR strategy in this study, termed ‘long-fragment circular-efficient PCR’ (LC-PCR). This technique involves one or two rounds of PCR with an additional third-long primer that complements both ends of the newly synthesized strand of a plasmid construct. This results in self-circularization with a nick-gap in each newly formed strand. The LC-PCR technique was successfully employed to insert a partial sequence (210 nucleotides) of the phytoene desaturase gene from Nicotiana benthamiana and a full capsid protein gene (770 nucleotides) of a begomovirus (tomato leaf curl New Delhi virus) into a 16.4 kb infectious construct of a tobamovirus, cucumber green mottle mosaic virus (CGMMV), cloned in pCambia. This was done to develop the virus-induced gene silencing vector (VIGS) and an expression vector for a foreign protein in plants, respectively. Furthermore, the LC-PCR could be applied for the deletion of a large region (replicase enzyme) and the substitution of a single amino acid in the CGMMV genome. Various in planta assays of these constructs validate their biological functionality, highlighting the utility of the LC-PCR technique in deciphering plant-virus functional genomics. The LC-PCR is not only suitable for modifying plant viral genomes but also applicable to a wide range of plant, animal, and human gene engineering under in-vitro conditions. Additionally, the LC-PCR technique provides an alternative to expensive kits, enabling quick introduction of modifications in any part of the nucleotide within a couple of days. Thus, the LC-PCR proves to be a suitable ‘all in one’ technique for modifying large plasmid constructs through site-directed gene insertion, deletion, and mutation, eliminating the need for restriction and ligation.

After two years since the declaration of COVID-19 as a pandemic by the World Health Organization (WHO), more than six million deaths have occurred due to SARS-CoV-2, leading to an unprecedented disruption of the global economy. Fortunately, within a year, a wide range of vaccines, including pathogen-based inactivated and live-attenuated vaccines, replicating and non-replicating vector-based vaccines, nucleic acid (DNA and mRNA)-based vaccines, and protein-based subunit and virus-like particle (VLP)-based vaccines, have been developed to mitigate the severe impacts of the COVID-19 pandemic. These vaccines have proven highly effective in reducing the severity of illness and preventing deaths. However, the availability and supply of COVID-19 vaccines have become an issue due to the prioritization of vaccine distribution in most countries. Additionally, as the virus continues to mutate and spread, questions have arisen regarding the effectiveness of vaccines against new strains of SARS-CoV-2 that can evade host immunity. The urgent need for booster doses to enhance immunity has been recognized. The scarcity of “safe and effective” vaccines has exacerbated global inequalities in terms of vaccine coverage. The development of COVID-19 vaccines has fallen short of the expectations set forth in 2020 and 2021. Furthermore, the equitable distribution of vaccines at the global and national levels remains a challenge, particularly in developing countries. In such circumstances, the exigency of plant virus-based vaccines has become apparent as a means to overcome supply shortages through fast manufacturing processes and to enable quick and convenient distribution to millions of people without the reliance on a cold chain system. Moreover, plant virus-based vaccines have demonstrated both safety and efficacy in eliciting robust cellular immunogenicity against COVID-19 pathogens. This review aims to shed light on the advantages and disadvantages of different types of vaccines developed against SARS-CoV-2 and provide an update on the current status of plant-based vaccines in the fight against the COVID-19 pandemic.

Temple tree (Plumeria rubra f. acutifolia), an important fragrant-flower tree extensively used in the urban landscaping is known to be infected with a tobamovirus, frangipani mosaic virus (FrMV). In this study, we describe another tobamovirus, Plumeria mosaic virus (PluMV) infecting temple tree in India. PluMV was isolated from an old temple tree co-infected with FrMV. The presence of another tobamovirus was initially realized based on the distinct symptoms on Gomphrena globosa (globe amaranth), a non-host of FrMV. PluMV was highly transmissible through simple rub-inoculation. In host-range study, brinjal (Solanum melongena), chilli (Capsicum annuum), datura (Datura stramonium), globe amaranth and tobacco (Nicotiana benthamiana, N. glutinosa, N. tabacum cv. Xanthi) could differentiate PluMV from FrMV. The complete genome sequence of PluMV was determined (6,688 nucleotides [nt], GenBank KJ395757), which showed the genome structure typical of tobamovirus encoding four proteins: small replicase (3,549 nt/130 kDa), large replicase (5,061 nt/188 kDa), movement protein (770 nt/29 kDa) and coat protein (527 nt/19 kDa). The 5′ and 3′ UTR of PluMV contained 91 and 284 nt, respectively. The PluMV genome was 45 nts longer than that of FrMV and shared only 71.4–71.6% sequence identity with FrMV and

Abstract Leaf curl, a whitefly-borne begomovirus disease, is the cause of frequent epidemic in chili. In the present study, transmission parameters involved in tripartite interaction are estimated to simulate disease dynamics in a population dynamics model framework. Epidemic is characterized by a rapid conversion rate of healthy host population into infectious type. Infection rate as basic reproduction number, R 0 = 13.54, has indicated a high rate of virus transmission. Equilibrium population of infectious host and viruliferous vector are observed to be sensitive to the immigration parameter. A small increase in immigration rate of viruliferous vector increased the population of both infectious host and viruliferous vector. Migrant viruliferous vectors, acquisition, and transmission rates as major parameters in the model indicate leaf curl epidemic is predominantly a vector -mediated process. Based on underlying principles of temperature influence on vector population abundance and transmission parameters, spatio-temporal pattern of disease risk predicted is noted to correspond with leaf curl distribution pattern in India. Temperature in the range of 15–35 °C plays an important role in epidemic as both vector population and virus transmission are influenced by temperature. Assessment of leaf curl dynamics would be a useful guide to crop planning and evolution of efficient management strategies.

Chilli is an important commercial crop grown in tropical and subtropical climates. The whitefly-transmitted chilli leaf curl virus (ChiLCV) is a serious threat to chilli cultivation. Vector migration rate and host–vector contact rate, the major drivers involved in the epidemic process, have been pinpointed to link management. The complete interception of migrant vectors immediately after transplantation has been noted to increase the survival time (to remain infection free) of the plants (80%) and thereby delay the epidemic process. The survival time under interception (30 days) has been noted to be nine weeks (p < 0.05), as compared to five weeks, which received a shorter period of interception (14–21 days). Non-significant differences in hazard ratios between 21- and 30-day interceptions helped optimize the cover period to 26 days. Vector feeding rate, estimated as a component of contact rate, is noted to increase until the sixth week with host density and decline subsequently due to plant succulence factor. Correspondence between the peak time of virus transmission or inoculation rate (at 8 weeks) and contact rate (at 6 weeks) suggests that host succulence is of critical importance in host–vector interactions. Infection proportion estimates in inoculated plants at different leaf stages have supported the view that virus transmission potential with plant age decreases, presumably due to modification in contact rate. The hypothesis that migrant vectors and contact rate dynamics are the primary drivers of the epidemic has been proved and translated into rules to guide management strategies.

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.

Groundnut bud necrosis virus (GBNV) is the most significant member of the genus Orthotospovirus occurring in the Indian subcontinent. There is hardly any effective measure to prevent GBNV in crop plants. In order to develop GBNV infection prevention procedure, we examined the effect of the direct foliar application of double-stranded RNA (dsRNA) derived from the full-length NSs gene (1,320 nucleotides) of GBNV. The bacterially expressed dsRNA to the non-structural (dsNSs) gene of GBNV was purified and delivered to plants as an aqueous suspension containing 0.01% Celite for evaluating its efficacy in preventing GBNV infection in systemic host, Nicotiana benthamiana as well as in local lesion and systemic host, cowpea cv. Pusa Komal (Vigna unguiculata). The dsNSs application and challenge-inoculation were conducted in three different combinations, where plants were challenge-inoculated with GBNV a day after, immediately, and a day before the application of dsNSs. N. benthamiana plants, which were not treated with dsRNA showed severe systemic wilting and death by 9–16 days post-inoculation (dpi). The non-treated cowpea plants exhibited many chlorotic and necrotic lesions on the cotyledonary leaves followed by systemic necrosis and death of the plants by 14–16 dpi. The dsNSs treated plants in all the combinations showed significant reduction of disease severity index in both N. benthamiana and cowpea. The treatment combination where the GBNV inoculation was conducted immediately after the dsNSs treatment was found to be the most effective treatment in preventing symptom expression. The viral RNA analysis by real time PCR also showed 20 and 12.5 fold reduction of GBNV in cowpea and N. benthamiana, respectively. Our results suggest that the foliar application of dsRNA derived from the full-length NSs gene of GBNV through Celite is successful in delivering long dsRNA leading to effective prevention of GBNV infection.

The major components of RNA silencing include both transitive and systemic small RNAs, which are technically called secondary sRNAs. Double-stranded RNAs trigger systemic silencing pathways to negatively regulate gene expression. The secondary siRNAs generated as a result of transitive silencing also play a substantial role in gene silencing especially in antiviral defense. In this review, we first describe the discovery and pathways of transitivity with emphasis on RNA-dependent RNA polymerases followed by description on the short range and systemic spread of silencing. We also provide an in-depth view on the various size classes of secondary siRNAs and their different roles in RNA silencing including their categorization based on their biogenesis. The other regulatory roles of secondary siRNAs in transgene silencing, virus-induced gene silencing, transitivity, and trans-species transfer have also been detailed. The possible implications and applications of systemic silencing and the different gene silencing tools developed are also described. The details on mobility and roles of secondary siRNAs derived from viral genome in plant defense against the respective viruses are presented. This entails the description of other compatible plant–virus interactions and the corresponding small RNAs that determine recovery from disease symptoms, exclusion of viruses from shoot meristems, and natural resistance. The last section presents an overview on the usefulness of RNA silencing for management of viral infections in crop plants.

Whitefly-transmitted begomoviruses cause serious damage to many economically important food, feed, and fiber crops. Numerous vegetable crops are severely affected and chilli leaf curl virus (ChiLCV) is the most dominant and widely distributed begomovirus in chilli (Capsicum annuum) throughout the Indian subcontinent. Recently, CRISPR-Cas9 technology was used as a means to reduce geminivirus replication in infected plants. However, this approach was shown to have certain limitations such as the evolution of escape mutants. In this study, we used a novel, multiplexed guide RNA (gRNA) based CRISPR-Cas9 approach that targets the viral genome at two or more sites simultaneously. This tactic was effective in eliminating the ChiLCV genome without recurrence of functional escape mutants. Six individual gRNA spacer sequences were designed from the ChiLCV genome and in vitro assays confirmed the cleavage behaviour of these spacer sequences. Multiplexed gRNA expression clones, based on combinations of the above-mentioned spacer sequences, were developed. A total of nine-duplex and two-triplex CRISPR-Cas9 constructs were made. The efficacy of these constructs was tested for inhibition of ChiLCV infection in Nicotiana benthamiana. Results indicated that all the constructs caused a significant reduction in viral DNA accumulation. In particular, three constructs (gRNA5+4, gRNA5+2 and gRNA1+2) were most effective in reducing the viral titer and symptoms. T7E1 assay and sequencing of the targeted viral genome did not detect any escape mutants. The multiplexed genome-editing technique could be an effective way to trigger a high level of resistance against begemoviruses. To our knowledge, this is the first report of demonstrating the effectiveness of a multiplexed gRNA-based plant virus genome editing to minimize and eliminate escape mutant formation.

Thrips are important pests of agricultural, horticultural, and forest crops worldwide. In addition to direct damages caused by feeding, several thrips species can transmit diverse tospoviruses. The present understanding of thrips–tospovirus relationships is largely based on studies of tomato spotted wilt virus (TSWV) and Western flower thrips (Frankliniella occidentalis). Little is known about other predominant tospoviruses and their thrips vectors. In this study, we report the progression of watermelon bud necrosis virus (WBNV) infection in its vector, melon thrips (Thrips palmi). Virus infection was visualized in different life stages of thrips using WBNV-nucleocapsid protein antibodies detected with FITC-conjugated secondary antibodies. The anterior midgut was the first to be infected with WBNV in the first instar larvae. The midgut of T. palmi was connected to the principal salivary glands (PSG) via ligaments and the tubular salivary glands (TSG). The infection progressed to the PSG primarily through the connecting ligaments during early larval instars. The TSG may also have an ancillary role in disseminating WBNV from the midgut to PSG in older instars of T. palmi. Infection of WBNV was also spread to the Malpighian tubules, hindgut, and posterior portion of the foregut during the adult stage. Maximum virus-specific fluorescence in the anterior midgut and PSG indicated the primary sites for WBNV replication. These findings will help to better understand the thrips–tospovirus molecular relationships and identify novel potential targets for their management. To our knowledge, this is the first report of the WBNV dissemination path in its vector, T. palmi.