Leaf Curl Begomovirus Research Articles

A multiplex polymerase chain reaction for the simultaneous detection of the virus and satellite components associated with cotton leaf curl begomovirus disease complex

Cotton leaf curl disease (CLCuD) is caused by a complex of several whiteflies (Bemisia tabaci Genn.)-transmitted begomovirus species, Cotton leaf curl Multan virus (CLCuMuV), Cotton leaf curl Kokhran virus (CLCuKoV) and Cotton leaf curl Alabad virus (CLCuAlV) by individual of mixed infection, associated with Cotton leaf curl Multan betasatellite (CLCuMB) and several alphasatellites. The disease causes major economic losses in cotton in the Indian subcontinent. For monitoring of epidemiology and development of management strategies of CLCuD, a quick, sensitive and effective method capable of detecting all the begomovirus, betasatellite and alphasatellite components associated with CLCuD is required. With this objective, a multiplex polymerase chain reaction (mPCR) assay was developed for the simultaneous detection of these three viral components associated with CLCuD of cotton. Primers for each component were designed based on the retrieved reference sequences from the GenBank. Each pair of primers, designed for each of the respective component, was evaluated for its sensitivity and specificity in both the component-specific simplex polymerase chain reaction (sPCR) and mPCR assay. This report identified three viral component-specific pairs of primers which, in all combinations, amplified simultaneously the CP gene (780 nts) of the begomovirus, the βC1gene (375 nts) of the betasatellite and the Rep gene (452 nts) of the alphasatellite associated with CLCuD in the mPCR assays. The amplified products specific to each component produced by these assays were identified based on their amplicon sizes, and the identities of the viral components amplified were confirmed by cloning and sequencing the amplicons obtained in the mPCR. The mPCR assay was validated using naturally CLCuD-affected cotton plants of the fields. This assay will be useful for rapid detection of CLCuD-associated begomovirus, betasatellite and alphasatellite DNA in field samples, extensive resistance screening in resistance breeding programme, and also monitoring epidemiology for detection of virus and its components when symptoms are mild or absent in the plant.

Complete genome sequence of hollyhock vein yellowing virus, a novel monopartite begomovirus infecting hollyhock in Pakistan.

Hollyhock (Alcea rosea, family Malvaceae) is an ornamental plant grown widely in gardens across South Asia. In a bed of ornamental plants near the village of Chakri (Punjab Province, Pakistan) in 2014, hollyhock plants showing two distinct symptom types were identified: yellow vein mosaic and leaf crumple. PCR amplification with universal primers amplified a begomovirus from separate nucleic acid extracts of single plants of each type but amplified a betasatellite only from the plant with the yellow vein mosaic symptoms. No potential begomovirus DNA B component or alphasatellite could be identified in either sample. After cloning, the genome sequences of two viruses, one from a plant of each symptom type, were determined and shown to share 99.9% nucleotide sequence identity with each other but less than 91% nucleotide sequence identity with all previously characterized begomoviruses, with the highest identity (90%) to an isolate of pedilanthus leaf curl virus (PeLCV). This indicates that the two hollyhock plants were infected with a newly identified begomovirus for which the name "hollyhock vein yellowing virus" (HoVYV) is proposed. HoVYV likely has a recombinant origin. The betasatellite showed the highest nucleotide sequence identity to an isolate of cotton leaf curl Multan betasatellite (CLCuMuB), a betasatellite associated with cotton leaf curl disease across Pakistan and northwestern India. These findings add to the diversity of known begomoviruses in South Asia and again highlight the role of hollyhock as a reservoir of the cotton leaf curl begomovirus betasatellite complex. The results also suggest that the yellow vein mosaic symptoms in hollyhock are due to the betasatellite rather than the virus.

Amplicon-based RNAi construct targeting beta-C1 gene gives enhanced resistance against cotton leaf curl disease.

The online version contains supplementary material available at 10.1007/s13205-021-02816-6.

Interplay of stress responses to carbamazepine treatment, whitefly infestation and virus infection in tomato plants

Reclaimed wastewater is increasingly used to irrigate agriculture crops. We have previously shown that carbamazepine (CBZ), an anticonvulsant human medication, not entirely discarded during wastewater purification, induces a stress response in tomatoes grown with roots bathing in CBZ-containing water. Induction of stress-related osmoprotectants (sugars, amino acids, proteins) was conspicuous in CBZ-treated seedlings. Here, tomato seedlings were grown in pots watered with increasing concentrations of CBZ. Soluble sugars effectively reacted to CBZ in both leaves and roots. However, the induction of stress-related amino acids and proteins was relevant in roots, but insignificant in leaves. Therefore, roots may be the site where CBZ stress is exerted. Moreover, roots may protect the whole plant from the pharmaceutical. Tomato crops endure biotic stresses, caused by whitefly (Bemisia tabaci) infestation and by the tomato yellow leaf curl begomovirus (TYLCV) they vector. The interplay between CBZ, TYLCV infection and B. tabaci was studied in tomato. Whiteflies preferred CBZ-treated plantlets than control plants, which may be due to increased amount of sugars in leaves. The increased amount of viruliferous whiteflies on CBZ-treated plants is expected to be accompanied by a rise in virus amounts. In fact, CBZ caused a reduction of TYLCV amounts. CBZ-dependent activation of autophagy degradation may explain this decrease in virus amounts. TYLCV infection mitigates the activation of stress markers associated with CBZ treatment. Altogether, CBZ in the water used to irrigate tomatoes grown in pots causes a relatively weak plant stress response, but is definitively sensed by insect and by virus.

Diversity and recombination analysis of Cotton leaf curl Multan betasatellite associated with cotton leaf curl begomovirus disease complex

Betasatellites are single-stranded DNA molecules associated with most monopartite begomoviruses of family Geminiviridae. They are symptoms modulating molecules and depend on their helper viruses for replication, encapsidation and movement. To investigate the genetic diversity of cotton leaf curl Multan betasatellite (CLCuMuB), different Pakistani isolates of betasatellites were retrieved from the NCBI data bank. The sequences were compared to all available sequences of CLCuMuB in the database, identified from cotton in Pakistan, India and China. Phylogenetic and recombination analysis showed the sequences are separated into five major groups. This is primarily based on recombination within the satellite conserved region (SCR) of the betasatellite genome. The first group consists of sequences of the original betasatellite associated with the first epidemic of cotton leaf curl disease (CLCuD) in the 1990s, while the second group consists of recombinant betasatellites associated with the second epidemic of CLCuD, containing a short fragment of tomato leaf curl betasatellite (ToLCB) DNA in the “satellite conserved region (SCR)”. The third group contains a short fragment of papaya leaf curl betasatellite (PaLCuB), while the fourth group contains a short fragment of tobacco curly shoot betasatellite (TbCSB) in the SCR region. The fifth group consists of sequences of betasatellite associated with the recent begomovirus disease complex identified from Vehari. This betasatellite contains a short fragment of sequences in the SCR region of a defective betasatellite (KT228331) from India. This new strain of CLCuMuB is named CLCuMuBVeh. Here, the significance of the evolution of CLCuMuB with respect to the CLCuD complex is discussed.

In silico analysis of chili encoded miRNAs targeting Chili leaf curl begomovirus and its associated satellite

Chilli leaf curl virus (ChiLCV), a member of Geminiviridae family (Genus: Begomovirus), is one of the most destructive plant virus. Micro (mi) RNAs, are the endogenous non-coding small RNAs that play significant roles in plant growth and stress resistance by degrading targeted mRNA or repressing mRNA translation. Computational methods have identified numerous miRNAs in many plant species, whereas there is no report of chilli miRNAs targeting essential genes of ChiLCV genome and associated satellites. In this study, we have predicted chilli encoded miRNAs that could be used for silencing against chilli leaf curl virus (Accession no. MF737343) infection. We predicted several potential mir miRNAs, exhibited high complementarities with V1 (CP) and C1 (Rep) genes of ChiLCV. Other overlapping genes V2, C2 and C3 were also targeted by mir-miRNAs.

Spatial and Temporal Physiognomies of Whitefly and Tomato Yellow Leaf Curl Virus Epidemics in Southwestern Florida Tomato Fields.

Epidemics of tomato yellow leaf curl virus (TYLCV; species Tomato yellow leaf curl begomovirus) have been problematic to tomato production in the southeastern United States since the first detection of the virus in Florida in the late 1990s. Current strategies for management focus on farm-centric tactics that have had limited success for controlling either TYLCV or its whitefly vector. Areawide pest management (AWPM)-loosely defined as a coordinated effort to implement management strategies on a regional scale-may be a viable management alternative. A prerequisite for development of an AWPM program is an understanding of the spatial and temporal dynamics of the target pathogen and pest populations. The objective of this study was to characterize populations of whitefly and TYLCV in commercial tomato production fields in southwestern Florida and utilize this information to develop predictors of whitefly density and TYLCV disease incidence as a function of environmental and geographical factors. Scouting reports were submitted by cooperating growers located across approximately 20,000 acres in southwestern Florida from 2006 to 2012. Daily weather data were obtained from several local weather stations. Moran's I was used to assess spatial relationships and polynomial distributed lag regression was used to determine the relationship between weather variables, whitefly, and TYLCV. Analyses showed that the incidence of TYLCV increased proportionally with mean whitefly density as the season progressed. Nearest-neighbor analyses showed a strong linear relationship between the logarithms of whitefly densities in neighboring fields. A similar relationship was found with TYLCV incidences. Correlograms based on Moran's I showed that these relationships extended beyond neighboring fields and out to approximately 2.5 km for TYLCV and up to 5 km for whitefly, and that values of I were generally higher during the latter half of the production season for TYLCV. Weather was better at predicting whitefly density than at predicting TYLCV incidence. Whitefly density was best predicted by the number of days with an average temperature between 16 and 24°C (T16to24), relative humidity (RH) over the previous 31 days, and vapor pressure deficit over the last 8 days. TYLCV incidence was best predicted by T16to24, RH, and maximum wind speed over the previous 31 days. Results of this study helped to identify the extent to which populations of whitefly and TYLCV exist over the agricultural landscape of southwestern Florida, and the environmental conditions that favor epidemic growth. This information was used to propose an approach to AWPM for timing control measures for managing TYLCV epidemics.

The benefit of combining different Ty-genes for resistance to tomato leaf curl begomoviruses

The benefit of combining different <i>Ty</i>-genes for resistance to tomato leaf curl begomoviruses

Non-cultivated Cotton Species (Gossypium spp.) Act as a Reservoir for Cotton Leaf Curl Begomoviruses and Associated Satellites.

A collection of cultivated and non-cultivated species of cotton (Gossypium spp.) has been maintained for the last four decades in Multan, Pakistan. This geographical location has been observed as a hotspot for the evolution of begomoviruses and satellites associated with cotton leaf curl disease (CLCuD). Recent studies showed that begomoviruses responsible for the CLCuD epidemic in the 1990s, and that almost disappeared from the CLCuD complex in 2000s, have been observed again in CLCuD-infected cotton fields. To identify host species that acted as probable reservoirs for these viruses, we characterized begomoviruses and satellites in non-cultivated cotton species G. raimondii, G. thurberi and G. mustelinum and identified several species of CLCuD associated begomoviruses and satellites. Further, phylogenetic analysis indicated that the identified begomoviruses and beta/alphasatellites are closely related to the ones associated with the most recent CLCuD complex. qPCR indicated that the comparative level of virus significantly decreased in the presence of alphasatellites. Our results indicated that non-cultivated cotton species have been continuously challenged by diverse begomoviruses and associated satellites and act as reservoirs for CLCuD associated begomoviruses. These results provide novel insights into understanding the spread of begomoviruses and associated satellites in New World cotton species introduced into the Old World.

Detection of Cotton leaf curl-begomovirus and betasatellite DNA in Weed and other hosts in and around Cotton fields

In the present study, effort was made to identify alternative host of CLCuD-begomovirus and betasatellite based on PCR, using specific primers targeting partial C1 and CP gene of CLCuDbegomovirus and #C1 gene of betasatellite. A total number of 30 weeds and other hosts under 15 host species under 10 plant families were collected from cotton field and its surrounding of Sri Ganganagar and Hanumangarh districts of Rajasthan, and cotton fields of ICAR-IARI, New Delhi. The CP gene was detected in Blumea sp. and brinjal; #C1 gene in Abutilon theophrasti, Croton sp., Ipomoea triloba and brinjal; and partial C1 gene in Abutilon theophrasti, Croton sp., Ipomoea triloba, Ageratum conyzoides, wild cucurbit and brinjal. These weed and other hosts can act as reservoirs of CLCuD and its associated satellite molecules.

Three variants of cotton leaf curl begomoviruses with their satellite molecules are associated with cotton leaf curl disease aggravation in New Delhi

Cotton leaf curl disease (CLCuD), caused by monopartite begomoviruses and its satellite molecules, is one of the serious constrains in cultivation of cotton in India. In the present study, five CLCuD-begomovirus and its associated satellite molecules were characterized based on rolling circle amplification and sequencing of complete genome. Sequence analysis showed 82–99 % nucleotide identity among them. The phylogenetic analysis and nt identity matrix determined that of the five CLCuD-begomovirus isolates, three IARI-34, IARI-42 and IARI-50 were members of Cotton leaf curl Multan virus (CLCuMuV)-Rajasthan isolates, designated as CLCuMuV-Rajasthan-34 and two, IARI-30 and IARI-45 of Cotton leaf curl Kokhran virus (CLCuKoV)-Burewala isolates, designated as CLCuKoV-Burewala-45. The present CLCuMuV-Rajasthan-34 is recombinant isolate showing recombination events in IR, C1 and C4 regions of its genome with high probality (P = 9.9 × 10−10–3.2 × 10−6). Same species of betasatellite (1371 nt) molecules obtained from both the present isolates was related with cotton leaf curl Multan betasatellite by 89–97 % nt identity. Three alphasatellites (1366–1396 nt) related to Cotton leaf curl Burewala alphasatellite and Gossypium darwinii symptomless alphasatellite by 86 % nt identity were also obtained. This is the first report of appearance of CLCuKoV-Burewala isolate and CLCuD associated alphasatellites in New Delhi. The present study demonstrated that CLCuD in New Delhi is caused by three kinds of variants, two are strains of CLCuMuV and one of CLCuKoV, either by single or mixed infection along with beta- and alpha-satellite molecues.

Global positioning system based spatial and temporal distribution of new leaf curl begomovirus disease on sunflower in Northern Karnataka

Leaf curl disease on sunflower caused by begomovirus genus of the family geminiviridae. Present investigations on field survey for disease incidence, field diagnostic symptoms and its spatial and temporal distribution in major sunflower growing parts of North Eastern Karnataka through GPS system during 2013-14, revealed that the disease was found to occur at all the stages of sunflower under field condition and exhibited symptoms such as vein thickening (enations) on abaxial surface of the leaves, upward curling and reduction in leaf size and severe discoluration of capitulum (Head) followed by bushy appearance. GPS based survey indicated that the % disease incidence varied from location to location (spatial variation) and also from season to season (temporal variation). The low incidence was noticed during Kharif condition which is ranged between 6.34-11.16, with the average incidence of 11.2%, 7.4% and 6.3% in Koppal, Raichur and Ballari districts repectively. Whereas during Rabi/summer season, high magnitude of disease noticed in many of the locations surveyed and is recorded upto 92.9 %. The GPS maps plotted based on PDI scale (0-3) represents high risk areas of the disease in Raichur and adjacent areas of Nort Eastern Karnataka and the result shows that the disease occurrence was more in rabi as compared to Kharif situations irrespective of locations. GPS survey map is an indicator to locate the nature of disease spread so as to conclude the hotspot areas.

Biological and molecular evidences on host range of leaf curl begomovirus disease of sunflower (Helianthus annuus L.)

The present study was conducted to identify the alternate hosts of new leaf curl virus disease of sunflower. In the present study several crops and weed hosts were cross inoculated with leaf curl virus of sunflower under laboratory through insect vector whitefly (Bemisia tabaci), further all inoculated samples were retested (3-4 weeks after inoculation) by molecular based Polymerse chain reaction diagnosis for the presence of virus. The results revealed that the causal virus of the disease was successfully transmitted from sunflower to sunflower (Helianthus annuus), tomato (Solanum lycopersicum) and tobacco (Nicotiana tabacum L) and weed hosts such as Acanthospermum hispidum, Amaranthus viridis and Parthenium hysterophorus in a short incubation period (2-3 weeks after inoculation), while on other hosts Chilli (Capsicum annuum L) and Datura stramonium, infection occurs in delayed incubation period. Further molecular analysis thorough polymerase chain reaction (PCR) diagnostic technique using virus specific primers also confirmed the presence of coat protein (CP) of leaf curl begomovirus invirus inoculated hosts viz., chilli, sunflower, tomato, and tobacco and weed hosts such as Acanthospermum hispidum, Amaranthus viridis, Datura stramonium and Parthenium hysterophorus. Thus, findings substantiate that the above hosts are major sources of the virus inoculum and served as potential alternate hosts of the disease during the off season.

Ketahanan Biokimia Tanaman Cabai terhadap Begomovirus Penyebab Penyakit Daun Keriting Kuning

Plant biochemical resistance towards virus infection can be evaluated through measurement of salicylic acid accumulation and peroxidase concentration. A research was conducted to study biochemical resistance of chillipepper to infection of Pepper yellow leaf curl Begomovirus (PYLCV) the causal agent of pepper yellow leaf curl disease. Virus inoculation was done using insect vector, Bemisia tabaci . Salicylic acid accumulation and peroxidase concentration on 6 chillipepper genotypes (35C2, IPB C15, IPB C26, IPB C10, IPB C14, dan IPB C12) was measured 120 h after inoculation. The results showed that salicylic acid accumulation and peroxidase concentration were higher on plants after virus infection. Susceptible genotypes (35C2, IPB C15, IPB C26) tend to have higher salicylic acid accumulation and peroxidase concentration than those of resistant genotype (IPB C12). Positive correlation was observed between biochemical resistance (salicylic acid accumulation) and structural resistance (length of palisade meshophyll cell). Key words: Begomovirus , Pepper yellow leaf curl virus , peroxide enzyme, resistance mechanism, salicylic acid

A novel cassava-infecting begomovirus from Madagascar: cassava mosaic Madagascar virus

Cassava mosaic geminiviruses (CMGs) are implicated in cassava mosaic disease (CMD), the main constraint to cassava production in Africa. Here, we report the complete nucleotide sequences of the DNA-A and DNA-B of a newly characterized CMG found infecting cassava in Madagascar, for which we propose the tentative name cassava mosaic Madagascar virus. With the exception of two recombinant regions that resembled a CMG, we determined that the non-recombinant part of the DNA-A component is distantly related to the other CMGs. Whereas the DNA-B component possesses one recombinant region originating from an unidentified virus, the rest of the genome was seen to be closely related to members of the species East African cassava mosaic Zanzibar virus (EACMZV). Phylogenetic analysis based on complete genome sequences demonstrated that DNA-A and DNA-B components are outliers related to the clade of EACMV-like viruses and that DNA-A is related to the monopartite tomato leaf curl begomoviruses described in islands in the south-west Indian Ocean.

Differential disease phenotype of begomoviruses associated with tobacco leaf curl disease in Comoros

In the 2000s, tobacco plantations on the Comoros Islands were afflicted with a previously unobserved tobacco leaf curl disease characterised by symptoms of severe leaf curling and deformation. Previous molecular characterization of potential viral pathogens revealed a complex of African monopartite tobacco leaf curl begomovirus (TbLCVs). Our molecular investigation allowed the characterization of a new monopartite virus involved in the disease: tomato leaf curl Namakely virus (ToLCNamV). Agroinoculation experiments indicated that TbLCVs and tomato leaf curl viruses (ToLCVs) can infect both tomato and tobacco but that infectivity and symptom expression fluctuate depending on the virus and the plant cultivar combination.

Tomato yellow leaf curl virus (TYLCV), identification, virus vector relationship, strains characterization and a suggestion for its control with plant extracts in Iraq

Tomato yellow leaf curl begomovirus (TYLCV) was identified on the basis of symptoms on test plants, transmission by whiteflies Bemisia tabaci Gennadius (Hemiptera: Aleyrodidae) biotype B and its incubation period in the insect. Symptomatic tomato plants showing leaf curling, leaf blade reduction, distortion and general stunting, indicative of TYLCV in greenhouses, were transplanted into pots (20 × 20 cm) containing soil mix and were maintained in growth room for identification. The virus was found to be easily acquired and transmitted by B. tabacibiotype B when given an acquisition access period on tomato infected plants for 30 min with incubation period of 24 h. Leaf curl and distortion associated with severe stunting and yellowing were developed on inoculated Datura stramonium, Nicotiana tabacum cv. Samsun NN, Nicotiana glutinosa and Phaseolus vulgaris var. Battle and were found as susceptible to the virus without visible symptoms. The virus caused vein clearing on inoculated Solanum nigrum leaves. No infections were observed on Gomphrena globosa, N. tabacum cv. White burly, Physalis floridana, Solanum melongena, Cucumis sativus, Chenopodium amaranlicolor and Chenopodium murale inoculated by the virus. Two strains for TYLCV were characterized by symptoms on tomato and immunu-double diffusion test. A spur was formed between the polyclonal antibodies of TYLCV isolate and the extracts from infected tomato plants showing different symptoms. Extracts from Thuja, Tamarix and Henna plants exhibited inhibitory effects on TYLCV multiplication in tomato treated plants with protection periods of 10 to 12 days. Key words: Bemisia tabaci, control, plant extracts, strains, Tomato yellow leaf curl begomovirus.

Gossypium robinsonii, an Australian wild cotton species is an asymptomatic host of the cotton leaf curl disease pathogen complex

Three wild species of cotton (Gossypium sp.) of Australian origin being maintained in a living herbarium at Central Cotton Research Institute, Multan, Pakistan were screened for the presence of cotton leaf curl begomovirus components. The screening by PCR, rolling circle amplification and Southern hybridisation, showed that G. robinsonii is an asymptomatic carrier of components of the disease complex prevalent in Pakistan.). Only betasatellites associated with CLCuD was identified by PCR and RCA in G. nelsonii, and G. bickii and suggests that begomovirus levels were below detection limits.

Normes OEPP/EPPO Standards Diagnostics Diagnostic

Normes OEPP/EPPO Standards Diagnostics Diagnostic

Normes OEPP/EPPO Standards Diagnostics Diagnostic

Normes OEPP/EPPO Standards Diagnostics Diagnostic