Dicot-infecting Mastreviruses Research Articles

Genome characterization of sweet potato symptomless virus 1: a mastrevirus with an unusual nonanucleotide sequence.

Complete genomic sequences of nine isolates of sweet potato symptomless virus 1 (SPSMV-1), a virus of the genus Mastrevirus in the family Geminiviridae, were determined from sweet potato accessions from different countries and found to be 2,559-2,602 nucleotides in length. These isolates shared 97-100% genome sequence identity and had an unusual nonanucleotide sequence (TAAGATTCC) in a large intergenic region as well as an additional open reading frame, C3, which is conserved in dicot-infecting mastreviruses.

New strains of chickpea chlorotic dwarf virus discovered on diseased papaya and tomato plants in Burkina Faso

This is the first description of full genome sequences of chickpea chlorotic dwarf virus (CpCDV; genus Mastrevirus; family Geminiviridae) identified in papaya and tomato plants sampled in Burkina Faso. The CpCDV full genome sequences from papaya and tomato share the highest pairwise sequence identity (84% and 93.5%) with Sudanese isolates of the CpCDV-K and CpCDV-M strains, respectively. Based on the strain demarcation threshold (>94% identity) for mastreviruses, we propose two new strains, CpCDV-Q and CpCDV-R, identified in papaya and tomato, respectively. Phylogenetic analysis confirmed that the sequences belong to a distinct clade of the highly diverse population of CpCDVs. Evidence of inter-strain recombination provided more support for the important role of recombination in CpCDV evolution. The discovery of CpCDV on papaya, a previously unsuspected host, raises many questions about the natural and potential host range of this dicot-infecting mastrevirus species that is reported to be emerging worldwide.

Identification of an Australian-like dicot-infecting mastrevirus in Pakistan.

Although members of five distinct viral species in the genus Mastrevirus (family Geminiviridae) infect dicotyledonous plants in Australia, in the remainder of the world, only a single dicot-infecting mastrevirus, chickpea chlorotic dwarf virus (CpCDV) has ever been identified. This virus has been found infecting leguminous hosts in Africa, the Middle East and the Indian subcontinent. To further explore the diversity of CpCDV in Pakistan, ten full mastrevirus genome sequences from chickpea and lentil plants were determined. Eight of these genomes were from previously described CpCDV strains and included the first reported strain D and H isolates in Pakistan. Two other genomes derived from infected chickpea plants are more closely related to dicot-infecting mastreviruses found in Australia than they are to CpCDV. These two divergent genomes shared less than 75% genome-wide nucleotide sequence identity with other characterised mastreviruses and therefore are likely to belong to a second species of dicot-infecting mastreviruses outside of Australia. We propose naming this species Chickpea yellow dwarf virus. We discuss how the presence of chickpea yellow dwarf virus (CpYDV) in Pakistan weakens the hypothesis that Australia is the geographical origin of the dicot-infecting mastreviruses.

Molecular diversity of Chickpea chlorotic dwarf virus in Sudan: High rates of intra-species recombination – a driving force in the emergence of new strains

In Sudan Chickpea chlorotic dwarf virus (CpCDV, genus Mastrevirus, family Geminiviridae) is an important pathogen of pulses that are grown both for local consumption, and for export. Although a few studies have characterised CpCDV genomes from countries in the Middle East, Africa and the Indian subcontinent, little is known about CpCDV diversity in any of the major chickpea production areas in these regions. Here we analyse the diversity of 146 CpCDV isolates characterised from pulses collected across the chickpea growing regions of Sudan. Although we find that seven of the twelve known CpCDV strains are present within the country, strain CpCDV-H alone accounted for ∼73% of the infections analysed. Additionally we identified four new strains (CpCDV-M, -N, -O and -P) and show that recombination has played a significant role in the diversification of CpCDV, at least in this region. Accounting for observed recombination events, we use the large amounts of data generated here to compare patterns of natural selection within protein coding regions of CpCDV and other dicot-infecting mastrevirus species.

A Distinct Strain of Chickpea chlorotic dwarf virus Infecting Pepper in Oman.

During a field survey in 2011, pepper (Capsicum annum) plants showing symptoms suggestive of geminivirus infection were observed in three fields in the Al-Sharqiya region of Oman. Symptoms observed included upward leaf curling leading to cupping and stunting with 15 to 25% disease incidence in surveyed fields. Total DNA was extracted from the leaves of seven symptomatic plants and subjected to rolling circle amplification (RCA). The RCA product was digested with several restriction endonucleases to obtain unit length of ~2.6 to 2.8, typical of geminivirus. Out of seven samples, only four yielded a product of ~2.6 kb in size by KpnI digestion. The fragments were cloned in pUC19 and sequenced. The partial sequences of four isolates were >95% identical to each other at the nucleotide (nt) level and thus only one isolate (P-25) was fully sequenced, determined to be 2,572 nt in length, and its sequence deposited in GenBank (KF111683). The P-25 sequence showed a genome organization typical of a mastrevirus, with four open reading frames (ORFs), two in virion-sense and two in complementary-sense. The virion and complementary-sense ORFs were separated by a long intergenic region, containing a predicted hairpin structure with the nonanucleotide sequence (TAATATTAC) in the loop, and a short intergenic region. An initial comparison to all sequences in the NCBI database using BlastN showed the isolate to have the highest level of sequence identity with isolates of the dicot-infecting mastrevirus Chickpea chlorotic dwarf virus (CpCDV). Subsequent alignments of all available CpCDV isolates using the species demarcation tool (2) showed the isolate P-25 to share between 83.6 and 90.3% identity to isolates of CpCDV available in databases, with the highest (90.3%) to CpCDV strain A originating from Syria (FR687959) (3). Amino acid sequence comparison showed that the predicted proteins encoded by the four ORFs of P-25 (coat protein [CP], movement protein [MP], replication associated protein A [RepA], and RepB) share 91.5, 88.2, 89.1, and 90.8% amino acid sequence identity, respectively, with the homologous proteins of the CpCDV isolate from Syria. Based on the recently revised mastreviruses species and strain demarcation criteria (78 and 94% whole genome nt identity, respectively) proposed by Muhire et al. (2), the results indicate that isolate P-25 represents a newly identified strain (strain F) of CpCDV. The presence of CpCDV in symptomatic pepper plants was further confirmed by Southern blot hybridization technique using digoxygenin (DIG) labeled probe prepared from CpCDV isolate P-25. The genus Mastrevirus consists of geminiviruses with single component genomes that are transmitted by leafhoppers. Mastreviruses have so far only been identified in the Old World and infect either monocotyledonous or dicotyledonous plants (1). To our knowledge, this is the first report of a mastrevirus on the Arabian Peninsula and the first record of pepper as host of CpCDV. Recently, several begomoviruses of diverse geographic origins have been found infecting vegetable crops in Oman. The propensity of geminiviruses to evolve through recombination may lead to evolution of recombinant CpCDV with new host adaptability. Due to extensive agricultural/travel links of Oman with rest of the world, there exists high probability for the spread of this virus.

Evidence that dicot-infecting mastreviruses are particularly prone to inter-species recombination and have likely been circulating in Australia for longer than in Africa and the Middle East

Viruses of the genus Mastrevirus (family Geminiviridae) are transmitted by leafhoppers and infect either mono- or dicotyledonous plants. Here we have determined the full length sequences of 49 dicot-infecting mastrevirus isolates sampled in Australia, Eritrea, India, Iran, Pakistan, Syria, Turkey and Yemen. Comprehensive analysis of all available dicot-infecting mastrevirus sequences showed the diversity of these viruses in Australia to be greater than in the rest of their known range, consistent with earlier studies, and that, in contrast with the situation in monocot-infecting mastreviruses, detected inter-species recombination events outnumbered intra-species recombination events. Consistent with Australia having the greatest diversity of known dicot-infecting mastreviruses phylogeographic analyses indicating the most plausible scheme for the spread of these viruses to their present locations, suggest that most recent common ancestor of these viruses is likely nearer Australia than it is to the other regions investigated.

Infectivity and the phylogenetic relationship of a mastrevirus causing chickpea stunt disease in India

Dicot-infecting mastreviruses affect chickpea (Cicer arietinum) plants by causing extreme stunting of the plant and leaf lamina reduction. Chickpea stunt disease (CSD) is presently known to occur in Africa, the Middle East and Asia. Although the disease was first recorded in India, little was known about the pathogen causing the disease. In this study we determined the complete nucleotide sequence of the mastrevirus associated with CSD in the region of Delhi. The genomic component of the virus was cloned using a rolling circle amplification (RCA) method. The virus isolate was found to show 99% sequence identity with the Chickpea chlorotic dwarf Pakistan virus. The complete tandem dimeric construct of the virus was found to be highly infectious to chickpea, and induced severe stunting of the plant, leaf smalling, drying, and the eventual death of the plant. Phylogenetic analysis of all the chickpea-infecting mastreviruses helped to distinguish the current differences between viruses originating from Africa, the Middle East, Asia and Australia.

Australian monocot-infecting mastrevirus diversity rivals that in Africa

Monocotyledonous plant infecting mastreviruses (family Geminiviridae) have been found in the Old World. The greatest diversity of these viruses has been found in Africa but this may simply reflect the more extensive sampling that has been done there. To provide a better understanding of mastrevirus diversity in Australia, we have sequenced the genomes of 41 virus isolates found in naturalised and native grasses and identified four new species in addition to the four previously characterised species. Two of these species, which were recovered from a single Sporobolus plant, are highly divergent and are most closely related to the African streak viruses. This, coupled with the discovery of divergent dicotyledonous plant infecting mastreviruses in Australia brings into question the hypothesis that mastreviruses may have originated in Africa. We found that the patterns of inter- and intra-species recombination and the recombination hotspots mirror those found in both their African monocot-infecting counterparts and dicot-infecting mastrevirus.

Molecular characterisation of dicot-infecting mastreviruses from Australia

Monocotyledonous and dicotyledonous plant infecting mastreviruses threaten various agricultural systems throughout Africa, Eurasia and Australasia. In Australia three distinct mastrevirus species are known to infect dicotyledonous hosts such as chickpea, bean and tobacco. Amongst 34 new “dicot-infecting” mastrevirus full genome sequences obtained from these hosts we discovered one new species, four new strains, and various variants of previously described mastrevirus species. Besides providing additional support for the hypothesis that evolutionary processes operating during dicot-infecting mastrevirus evolution (such as patterns of pervasive homologous and non-homologous recombination, and strong purifying selection acting on all genes) have mostly mirrored those found in their monocot-infecting counterparts, we find that the Australian dicot-infecting viruses display patterns of phylogeographic clustering reminiscent of those displayed by monocot infecting mastrevirus species such as Panicum streak virus and Maize streak virus.

RNA interference-based resistance against a legume mastrevirus

BackgroundRNA interference (RNAi) is a homology-dependant gene silencing mechanism and has been widely used to engineer resistance in plants against RNA viruses. However, its usefulness in delivering resistance against plant DNA viruses belonging to family Geminiviridae is still being debated. Although the RNAi approach has been shown, using a transient assay, to be useful in countering monocotyledonous plant-infecting geminiviruses of the genus Mastrevirus, it has yet to be investigated as a means of delivering resistance to dicot-infecting mastreviruses. Chickpea chlorotic dwarf Pakistan virus (CpCDPKV) is a legume-infecting mastrevirus that affects chickpea and other leguminous crops in Pakistan.ResultsHere a hairpin (hp)RNAi construct containing sequences encompassing part of replication-associated protein gene, intergenic region and part of the movement protein gene of CpCDPKV under the control of the Cauliflower mosaic virus 35S promoter has been produced and stably transformed into Nicotiana benthamiana. Plants harboring the hairpin construct were challenged with CpCDPKV. All non-transgenic N. benthamiana plants developed symptoms of CpCDPKV infection within two weeks post-inoculation. In contrast, none of the inoculated transgenic plants showed symptoms of infection and no viral DNA could be detected by Southern hybridization. A real-time quantitative PCR analysis identified very low-level accumulation of viral DNA in the inoculated transgenic plants.ConclusionsThe results presented show that the RNAi-based resistance strategy is useful in protecting plants from a dicot-infecting mastrevirus. The very low levels of virus detected in plant tissue of transgenic plants distal to the inoculation site suggest that virus movement and/or viral replication was impaired leading to plants that showed no discernible signs of virus infection.

Recombination patterns in dicot-infecting mastreviruses mirror those found in monocot-infecting mastreviruses

Recombination has profoundly shaped the evolution of viruses in the family Geminiviridae and has been studied extensively in the two best characterised geminivirus lineages: the dicotyledonous plant infecting begomoviruses and the monocotyledonous plant infecting mastreviruses. Here, we demonstrate that the sizes and distributions of recombination events detectable within the members of a third major geminivirus lineage--the dicotyledonous plant infecting mastreviruses--are very similar to those of the monocot-infecting mastreviruses. This suggests that, despite host range differences, very similar biochemical, ecological and evolutionary factors must underlie recombination patterns in the dicot- and monocot-infecting mastreviruses.

Analysis of the sequence of a dicot-infecting mastrevirus (family Geminiviridae) originating from Syria

Chickpea stunt disease (CSD) across southern Asia, the Middle East and North Africa is caused by a number of viruses that include single-stranded DNA viruses of the genus Mastrevirus (family Geminiviridae). Despite the importance of CSD in reducing chickpea and lentil production, until recently little was known of the nature of the pathogens causing the disease. Sequence characterisation of virus isolates from Sudan and Pakistan showed the viruses concerned to potentially be new mastrevirus species related to Bean yellow dwarf virus (BeYDV), a virus known to occur in both southern Africa and southern Asia. Here we have determined the complete nucleotide sequence of a mastrevirus associated with CSD in Syria. This virus represents a proposed new species, closely related to the recently characterised Chickpea chlorotic dwarf Sudan virus and Chickpea chlorotic dwarf Pakistan virus but with the highest sequence identity to BeYDV, for which we propose the name Chickpea chlorotic dwarf Syria virus. In addition the biological integrity of the clone was confirmed by infection of Nicotiana benthamiana plants using Agrobacterium-mediated inoculation.

Two novel mastreviruses from chickpea (Cicer arietinum) in Australia

Two novel mastreviruses (genus Mastrevirus; family Geminiviridae), with proposed names chickpea chlorosis virus (CpCV) and chickpea redleaf virus, are described from chickpea (Cicer arietinum) from eastern Australia. The viruses have genomes of 2,582 and 2,605 nucleotides, respectively, and share similar features and organisation with typical dicot-infecting mastreviruses. Two distinct strains of CpCV were suggested by phylogenetic analysis. Additionally, a partial mastrevirus Rep sequence from turnip weed (Rapistrum rugosum) indicated the presence of a distinct strain of Tobacco yellow dwarf virus (TYDV). In phylogenetic analyses, isolates of Bean yellow dwarf virus, Chickpea chlorotic dwarf Pakistan virus and Chickpea chlorotic dwarf Sudan virus from southern and northern Africa and south-central and western Asia clustered separately from these three viruses from Australia. An Australian, eastern Asian, or south-eastern Asian origin for the novel mastreviruses and TYDV is discussed.

Mastreviruses in chickpea (Cicer arietinum) and other dicotyledonous crops and weeds in Queensland and northern New South Wales, Australia

Natural infection by mastreviruses was investigated in chickpea (Cicer arietinum) and other dicotyledonous crops and weeds in grain production areas of Queensland and northern New South Wales, Australia, from 2000 to 2005. Altogether, 33 639 plants comprising 31 species and 10 dicot families were screened for infection by a tissue-blot immunoassay that did not distinguish between mastrevirus strains or species. Nine plant species in three families were identified as natural hosts. Chickpea was infected throughout the region although infection incidence did not exceed 5%. Infection was rare in faba bean (Vicia faba), canola (Brassica napus), and mustard (B. juncea) and not detected in field pea (Pisum sativum). Infection of chickpea and turnip weed (Rapistrum rugosum) was confirmed by immunocapture polymerase chain reaction (IC-PCR) with primers generic for dicot-infecting mastreviruses, and also immunosorbent electron microscopy and graft transmission in the case of chickpea. Individual mastreviruses were identified by comparing their IC-PCR amplicons by a combination of methods. Among 42 isolates from 41 chickpea plants, one was typical Tobacco yellow dwarf virus (TYDV) and the others were three recently distinguished strains including two proposed novel species: 34 Chickpea chlorosis virus strain A, six Chickpea chlorosis virus strain B, and one Chickpea redleaf virus. All of 10 isolates from 10 turnip weed plants were TYDV-B, a strain distinct from typical TYDV. The symptoms associated with mastrevirus infection in chickpea included foliar chlorosis or reddening, stunting, and usually phloem browning. The potential for losses in winter and summer grown field crops is discussed.

Two dicot-infecting mastreviruses (family Geminiviridae) occur in Pakistan

Most mastreviruses (family Geminiviridae) infect monocotyledonous hosts and are transmitted by leafhopper vectors. Only two mastrevirus species, Tobacco yellow dwarf virus from Australia and Bean yellow dwarf virus (BeYDV) from South Africa, have been identified whose members infect dicotyledonous plants. We have identified two distinct mastreviruses in chickpea stunt disease (CSD)-affected chickpea originating from Pakistan. The first is an isolate of BeYDV, previously only known to occur in South Africa. The second is a member of a new species with the BeYDV isolates as its closest relatives. A PCR-based diagnostic test was developed to differentiate these two virus species. Our results show that BeYDV plays no role in the etiology of CSD in Pakistan, while the second virus occurs widely in chickpea across Pakistan. A genomic clone of the new virus was infectious to chickpea (Cicer arietinum L.) and induced symptoms typical of CSD. We propose the use of the name Chickpea chlorotic dwarf Pakistan virus for the new species. The significance of these findings with respect to our understanding of the evolution, origin and geographic spread of dicot-infecting mastreviruses is discussed.