Possible Host Adaptation as an Evolution Factor of Cowpea aphid‐borne mosaic virus Deduced by Coat Protein Gene Analysis

Cowpea aphid‐borne mosaic virus (CABMV) causes major diseases in cowpea and passion flower plants in Brazil and also in other countries. CABMV has also been isolated from leguminous species including, Cassia hoffmannseggii, Canavalia rosea, Crotalaria juncea and Arachis hypogaea in Brazil. The virus seems to be adapted to two distinct families, the Passifloraceae and Fabaceae. Aiming to identify CABMV and elucidate a possible host adaptation of this virus species, isolates from cowpea, passion flower and C. hoffmannseggii collected in the states of Pernambuco and Rio Grande do Norte were analysed by sequencing the complete coat protein genes. A phylogenetic tree was constructed based on the obtained sequences and those available in public databases. Major Brazilian isolates from passion flower, independently of the geographical distances among them, were grouped in three different clusters. The possible host adaptation was also observed in fabaceous‐infecting CABMV Brazilian isolates. These host adaptations possibly occurred independently within Brazil, so all these clusters belong to a bigger Brazilian cluster. Nevertheless, African passion flower or cowpea‐infecting isolates formed totally different clusters. These results showed that host adaptation could be one factor for CABMV evolution, although geographical isolation is a stronger factor.     

Recombination Among Begomoviruses on Malvaceous Plants Leads to the Evolution of Okra Enation Leaf Curl Virus in Pakistan

Whitefly transmitted begomoviruses (family Geminiviridae) are the major reason for significant yield losses of dicotyledonous crops in tropics and subtropics. Okra (Abelmoschus esculentus) is one of the important vegetable crops, and leaf curl disease caused by geminiviruses is the most important limiting factor for its production in Pakistan. Here, we report a new species of okra‐infecting begomovirus in south‐eastern region of Pakistan and the name Okra enation leaf curl virus (OELCuV) complex is proposed. This okra enation leaf curl disease complex (OELCuD) in Pakistan is found to be associated with Ageratum conyzoides symptomless alphasatellite (AConSLA). All efforts to clone the betasatellite were unsuccessful. Comprehensive sequence analyses suggest that intermalvaceous recombination between okra and cotton‐infecting begomoviruses resulted in the evolution of the new species. Surprisingly, Bhendi yellow vein mosaic virus (BYVMV) which has not been reported previously from Pakistan is the major parent while Cotton leaf curl Multan virus (CLCuMV) acts as a distant parent of the virus. Comparative recombination analysis also reveals that okra‐infecting begomoviruses from south and north‐western India is causing OELCuD in the Pakistan by recombining with CLCuMV at the Rep (1964–1513 nts). Recombination is common among geminiviruses and recombining of BYVMV and CLCuMV resulted in a new species: OELCuV. To the best of our knowledge, this evolution of a new species of okra‐infecting begomovirus is the first report of intermalvaceous recombination where Rep acts as the target region.     

A survey of mixed Begomovirus infection in solanaceae and fabaceae at different altitudes in East Java,Indonesia

A multiplex primer set was developed to detect four Begomoviruses in East Java, Indonesia, i.e. Tomato leaf curl New Delhi virus (ToLCNDV), Tomato yellow leaf curl Kanchanaburi virus (TYLCKaV), Pepper yellow leaf curl Indonesia virus (PepYLCIV) and Mungbean yellow mosaic India virus (MYMIV). Survey at different altitudes found that begomoviruses infecting pepper, tomato and long bean were more variable, while in eggplant and string bean were more uniform. As a single virus, TYLCKaV infected eggplant, and sometimes tomato and pepper; PepYLCIV infected pepper, tomato and long bean; ToLCNDV only infected long bean and tomato at low frequency; and MYMIV infected beans. Mixed infection occurred more frequently in the low altitude areas. Subsequent examination indicated that Cucumber mosaic virus (CMV) and potyviruses were also responsible for diseased fabaceous. Our data suggest a relationship between altitudes and virus species occurrence. However, which viral species infects a crop is mainly influenced by the crop rather than by altitude.     

Identification and Characterization of a Potyvirus Isolated from Siratro Plants

The present work describes the identification and characterization of a potyvirus isolated from siratro (Macroptilium atropurpureum Urb.) in the north‐west region of the State of São Paulo, Brazil. The virus was transmitted by mechanical inoculation. Its host range was restricted mainly to members of the Fabaceae. A cDNA fragment of about 930 bp was amplified by RT/PCR, cloned and sequenced. The fragment, which included the coat protein gene, had amino acid identity percentages between 88 and 98% with isolates of Bean common mosaic virus (BCMV). Phylogenetic analysis grouped the siratro potyvirus and BCMV isolates in 99% of the replicates, including Azuki mosaic virus, Dendrobium mosaic virus, Blackeye cowpea mosaic virus and Peanut stripe virus, which have been classified as BCMV strains. This is the first citation on the presence of BCMV in siratro plants in Brazil.     

Tomato yellow leaf curl virus and Tomato leaf curl Taiwan virus Invade South-east Coast of China

An epidemic outbreak of severe yellow leaf curl disease was reported in field grown tomato within Zhejiang Province of China in the autumn–winter cropping season of 2006. A molecular diagnostic survey was carried out based on comparisons of partial and complete viral DNA sequences. Comparison of partial DNA‐A sequences amplified with degenerate primers specific for begomoviruses confirmed the presence of two types of begomoviruses. The complete DNA sequences of five isolates, corresponding to the two types, were determined. Sequence comparisons and phylogenetic analysis revealed that they correspond to two previously identified begomoviruses, Tomato yellow leaf curl virus and Tomato leaf curl Taiwan virus. The satellite DNAβ molecule was not detected in these samples by either PCR or Southern blot hybridization analysis. There has been no previous report of geminivirus disease incidence in Zhejiang Province, indicating that the introduction of these two tomato infecting geminiviruses into the agro‐ecological zone of South‐eastern China is a fairly recent event. The implications for disease control are discussed.     

Natural Infection by Tospoviruses of Cucurbitaceous and Fabaceous Vegetable Crops in India

Natural infection of tospoviruses on three cucurbitaceous (Cucumis sativus, cucumber; Luffa acutangula, ridge gourd; Citrullus lanatus, watermelon) and three fabaceous (Vigna unguiculata, cowpea; Phaseolus vulgaris, French bean; Dolichos lablab, sem) vegetable crops in India was identified on the basis of nucleocapsid protein (NP) gene characteristics. The complete NP gene of the cowpea isolate from Kerala and the sem isolate from Tamil Nadu was 831 nucleotides long, encoding a protein of 276 amino acids. For other Tospovirus isolates from cucumber, French bean, ridge gourd and watermelon, the partial NP gene (291 nt) was sequenced. Comparative NP gene sequence analyses revealed that fabaceous isolates shared maximum identity both at the nucleotide (92–97%) and amino acid (93–97%) levels with the corresponding region of Groundnut bud necrosis virus (GBNV), whereas cucurbitaceous isolates shared maximum identity both at nucleotide (93–99%) and amino acid (95–98%) levels with the corresponding region of Watermelon bud necrosis virus (WBNV), results suggesting that the Tospovirus isolates infecting fabaceous hosts should be regarded as strain of GBNV, whereas those infecting cucurbitaceous hosts as a strain of WBNV. Nucleocapsid protein gene was conserved both in GBNV and WBNV isolates originating from different hosts and locations.     

The ever increasing diversity of begomoviruses infecting non‐cultivated hosts: new species from Sida spp. and Leonurus sibiricus,plus two New World alphasatellites

Begomoviruses (whitefly‐transmitted, single‐stranded DNA plant viruses) are among the most damaging pathogens causing epidemics in economically important crops worldwide. Besides cultivated plants, many weed and wild hosts act as virus reservoirs where recombination may occur, resulting in new species. The aim of this study was to further characterise the diversity of begomoviruses infecting two major weed genera, Sida and Leonurus. Total DNA was extracted from samples collected in the states of Rio Grande do Sul, Paraná and Mato Grosso do Sul during the years 2009–2011. Viral genomes were enriched by rolling circle amplification (RCA), linearised into unit length genomes using various restriction enzymes, cloned and sequenced. A total of 78 clones were obtained: 37 clones from Sida spp. plants and 41 clones from Leonurus sibiricus plants. Sequence analysis indicated the presence of six bipartite begomovirus species and two alphasatellites. In Sida spp. plants we found Sida micrantha mosaic virus (SiMMV), Euphorbia yellow mosaic virus (EuYMV), and three isolates that represent new species, for which the following names are proposed: Sida chlorotic mottle virus (SiCMoV), Sida bright yellow mosaic virus (SiBYMV) and Sida golden yellow spot virus (SiGYSV), an Old World‐like begomovirus. L. sibiricus plants had a lower diversity of begomoviruses compared to Sida spp., with only Tomato yellow spot virus (ToYSV) and EuYMV (for the first time detected infecting plants of the genus Leonurus) detected. Two satellite DNA molecules were found: Euphorbia yellow mosaic alphasatellite, for the first time detected infecting plants of the genus Sida, and a new alphasatellite associated with ToYSV in L. sibiricus. These results constitute further evidence of the high species diversity of begomoviruses in non‐cultivated hosts, particularly Sida spp.     

A new chrysanthemum potyvirus: molecular evidence

Abstract

A number of viruses are known to infect chrysanthemum plants, however in the present study a previously unknown potyvirus was detected using techniques such as ELISA, RT-PCR and hybridization. The ELISA-positive samples were amplified using a potyvirus group-specific primer which gave an amplification of ～850 bp. The amplified product was cloned and sequenced, and shows 72 – 73% homology with known potyviruses that infect chrysanthemums such as Potato virus Y potyvirus, Soyabean mosaic virus and Turnip mosaic potyvirus when compared to the sequence available in the database. However, present potyvirus isolates show 93% homology with Chilli veinal mottle virus and Pepper vein banding virus. The results were further confirmed by Northern hybridization. This is the first report of a potyvirus similar to Chilli veinal mottle virus, and Pepper vein banding virus infecting chrysanthemums.     

Complete Nucleotide Sequence of Ageratum enation virus and an Alphasatellite Infecting a New Host Glycine max in India

During 2011, leaf crumpling, yellowing and stunting were observed on soya bean (Glycine max) in Himachal Pradesh, India. PCR‐based detection confirmed the presence of a begomovirus. The viral genome was amplified by rolling circle amplification, cloned and sequenced. The complete nucleotide sequence of DNA‐A showed highest nucleotide identity to an isolate of Ageratum enation virus infecting a weed Ageratum conyzoides. In addition, a DNA molecule was found which shared 95% nucleotide identity with an alphasatellite infecting ageratum. Neither beta satellite nor DNA‐B was detected in the infected samples.     

Low Genetic Diversity Suggests a Single Introduction and Recent Spread of Tomato chlorosis virus in Brazil

By comparing the partial nucleotide sequences of the heat shock protein HSP70 homologue gene, we assessed the genetic diversity of Brazilian tomato isolates of Tomato chlorosis virus (ToCV), as well as their relationship with other ToCV isolates found worldwide. The Brazilian ToCV isolates shared 99.9–100% nucleotide identity, which indicates low genetic diversity. Brazilian ToCV isolates showed a closer evolutionary relationship to those from Mediterranean countries. Based on these results, the origin of Brazilian ToCV isolates and the possible number of introductions of the virus into Brazil are discussed.     

Occurrence,distribution and seasonal changes of viruses infecting common bean in northwestern Iran

Many surveys were conducted during 2003–2005 to study the identity, prevalence and fluctuation of bean infecting viruses in northwestern Iran. In total, 649 bean samples with virus- like symptoms were collected and analysed by double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) and tissue-print immunoassay to detect infectious viruses. Serological tests revealed the presence of Bean common mosaic virus (BCMV), Bean common mosaic necrosis virus (BCMNV), Bean yellow mosaic virus (BYMV), Cucumber mosaic virus (CMV), Alfalfa mosaic virus (AMV), Bean leaf roll virus (BLRV), Bean pod mottle virus (BPMV) and Southern bean mosaic virus (SBMV), with some co-infection occurred, with prevalence of BCMV, BCMNV and BYMV (17–29% infection rate). The incidence of viruses showed variation in over 3 years of research including more than double increase in CMV from 2004 to 2005 and obvious one-third decrease in AMV from 2003 to 2005. SBMV and BPMV were detected sporadically in the fields and the response of some differential test plants was analysed by mechanical inoculation. Western immunoblotting analysis of SBMV infected bean leaf total proteins using SBMV-specific polyclonal antibody revealed viral CP with molecular mass of 28.5 kDa which confirmed the presence of SBMV as a new threat for bean production.     

Occurrence of Six Begomoviruses Infecting Tomato Fields in Venezuela and Genetic Characterization of Potato Yellow Mosaic Virus Isolates

Begomoviruses are one of the major pathogens in tomato crops worldwide. In Venezuela, six begomovirus species have been described infecting tomato: Potato yellow mosaic virus (PYMV), Euphorbia mosaic Venezuela virus (EuMVV), Merremia mosaic virus (MeMV), Tomato chlorotic leaf distortion virus (ToCLDV), Tomato yellow margin leaf curl virus (TYMLCV) and Tomato yellow leaf curl virus (TYLCV). In this study, the occurrence of these viruses was analysed by PCR in 338 tomato plants exhibiting virus‐like symptoms. Sixty‐three per cent of the plants were positive at least to one of the begomoviruses tested. PYMV and TYLCV were the most frequent viruses showing 39.6 and 23.7% occurrence, respectively. Phylogenetic analyses revealed two groups of PYMV isolates from several Caribbean Basin countries. The first group clustered isolates from several countries, including Venezuela, and the second group clustered only Colombian isolates. Due to the high prevalence of PYMV and TYLCV in Venezuela, it is suggested that the surveillance and control strategies currently applied in the country should be focused on these two begomoviruses.     

Begomoviruses Associated with Leaf Curl Disease of Tomato in Java,Indonesia*

We report that several begomoviruses are associated with tomato leaf curl disease in Java, Indonesia. Tomato plants with leaf curl symptoms were collected from Bandung (west Java), Purwokerto (central Java), Magelang (central Java) and Malang (east Java) of Indonesia, the major tomato‐growing areas of the country. Viruses were detected using the polymerase chain reaction (PCR), with universal primers for the genus Begomovirus. PCR‐amplified fragments were cloned and sequenced. Based on sequence comparisons and phylogenetic analyses, the viruses were divided into three groups. With respect to amino acid (aa) identities of the N‐terminal halves of the coat proteins compared in this study, group I was most closely related to Ageratum yellow vein virus (AYVV) (97%), Ageratum yellow vein China virus‐[Hn2] (AYVCNV‐[Hn2]) (96%) and Ageratum yellow vein virus‐[Taiwan] (AYVV‐[Tai]) (95%), and ageratum‐infecting begomovirus from Java (99%). Group II had high sequence identity with a tentative species of tomato leaf curl Java virus (ToLCJAV) (96% aa) for the CP. Group III was most closely related to a proposed species of Pepper yellow leaf curl Indonesia virus (PepYLCIDV) (90% aa identity) by its partial CP sequence.     

Rust disease of eucalypts,caused by Puccinia psidii,did not originate via host jump from guava in Brazil

The rust fungus, Puccinia psidii, is a devastating pathogen of introduced eucalypts (Eucalyptus spp.) in Brazil where it was first observed in 1912. This pathogen is hypothesized to be endemic to South and Central America and to have first infected eucalypts via a host jump from native guava (Psidium guajava). Ten microsatellite markers were used to genotype 148 P. psidii samples from eucalypts and guava plus five additional myrtaceous hosts across a wide geographic range of south‐eastern Brazil and Uruguay. Principal coordinates analysis, a Bayesian clustering analysis and a minimum‐spanning network revealed two major genetic clusters among the sampled isolates, one associated with guava and another associated with eucalypts and three additional hosts. Multilocus genotypes infecting guava differed by multiple mutational steps at eight loci compared with those infecting eucalypts. Approximate Bayesian computation revealed that evolutionary scenarios involving a coalescence event between guava‐ and eucalypt‐associated pathogen populations within the past 1000 years are highly unlikely. None of the analyses supported the hypothesis that eucalypt‐infecting P. psidii in Brazil originated via host jump from guava following the introduction of eucalypts to Brazil approximately 185 years ago. The existence of host‐associated biotypes of P. psidii in Brazil indicates that this diversity must be considered when assessing the invasive threat posed by this pathogen to myrtaceous hosts worldwide.     

The incidence and distribution of viruses infecting lettuce, cultivated Brassica and associated natural vegetation in Spain

A virus survey was conducted during the spring and autumn of 2001 and 2002 to determine the presence, prevalence and distribution in Spain of the viruses that are most commonly found infecting lettuce and Brassica worldwide. Crop plants showing virus symptoms from the principal lettuce and Brassica-growing regions of Spain, and some samples of the annual and perennial flora nearby, were tested by enzyme-linked immunosorbent assays using specific commercial antibodies against the following viruses: Alfalfa mosaic virus (AMV), Broad bean wilt virus 1 (BBWV-1), Beet western yellows virus (BWYV), Cauliflower mosaic virus (CaMV), Cucumber mosaic virus (CMV), Lettuce mosaic virus (LMV), Pea seed-borne mosaic virus (PSbMV), Turnip mosaic virus (TuMV) and Tomato spotted wilt virus (TSWV). Samples were also tested with a Potyvirus genus antibody. Virus incidence was much lower in spring than in autumn, especially in 2001. In spring 2002, CMV and LMV were the most prevalent viruses in lettuce, while CaMV was the most important virus present in Brassica crops grown in Navarra, followed by CMV and BWYV. In the autumn, the spectrum of viruses was different; potyviruses were widespread in lettuce grown in Madrid, but TSWV and BWYV were predominant in the Murcia region. The prevalent Potyvirus detected in lettuce fields was LMV, but none of the samples collected were positive for PSbMV or TuMV. In Brassica crops, TSWV was the most abundant in autumn-sown crops, especially in the Navarra region. All of the viruses present in lettuce and Brassica were also frequently detected in their associated natural vegetation at the same time, suggesting that they probably play an important role as virus reservoirs. Sonchus spp. were particularly common and were frequently infected with CMV, LMV and BWYV. Another common species, Chenopodium album, was often infected with TSWV and BWYV. Multiple infections were common, especially in non-crop plants, and the most common combination was BWYV and TSWV. The role of weeds in the epidemiology of viruses that infect lettuce and Brassica crops in Spain is discussed.     

Deep sequencing reveals a novel closterovirus associated with wild rose leaf rosette disease

A bizarre virus‐like symptom of a leaf rosette formed by dense small leaves on branches of wild roses (Rosa multiflora Thunb.), designated as ‘wild rose leaf rosette disease’ (WRLRD), was observed in China. To investigate the presumed causal virus, a wild rose sample affected by WRLRD was subjected to deep sequencing of small interfering RNAs (siRNAs) for a complete survey of the infecting viruses and viroids. The assembly of siRNAs led to the reconstruction of the complete genomes of three known viruses, namely Apple stem grooving virus (ASGV), Blackberry chlorotic ringspot virus (BCRV) and Prunus necrotic ringspot virus (PNRSV), and of a novel virus provisionally named ‘rose leaf rosette‐associated virus’ (RLRaV). Phylogenetic analysis clearly placed RLRaV alongside members of the genus Closterovirus, family Closteroviridae. Genome organization of RLRaV RNA (17 653 nucleotides) showed 13 open reading frames (ORFs), except ORF1 and the quintuple gene block, most of which showed no significant similarities with known viral proteins, but, instead, had detectable identities to fungal or bacterial proteins. Additional novel molecular features indicated that RLRaV seems to be the most complex virus among the known genus members. To our knowledge, this is the first report of WRLRD and its associated closterovirus, as well as two ilarviruses and one capilovirus, infecting wild roses. Our findings present novel information about the closterovirus and the aetiology of this rose disease which should facilitate its control. More importantly, the novel features of RLRaV help to clarify the molecular and evolutionary features of the closterovirus.     

Horizontal transmission of begomoviruses between Bemisia tabaci biotypes

We have previously shown that the monopartite Tomato yellow leaf curl virus (TYLCV), a begomovirus (family Geminiviridae, genus Begomovirus) infecting tomato plants can be transmitted in a gender-dependent manner among its insect vector the whitefly Bemisia tabaci type B (Gennaduis) (Aleyrodidae: Hemiptera) during mating. Viruliferous females were able to transmit the virus to non-viruliferous males and vice versa, in the absence of any other virus source. The recipient insects were able to infect tomato plants. In this communication, we present evidence that two bipartite begomoviruses infecting cucurbits, Squash leaf curl virus (SLCV) and Watermelon chlorotic stunt virus (WmCSV) can be transmitted in a gender-dependent manner among whiteflies. In addition we show that TYLCV can be transmitted during mating among individuals from the same biotype (from B-males to B-females and vice versa; and from Q-males to Q-females and vice versa). However, viruliferous males of the B biotype are unable to transmit the virus to females of the Q biotype (and vice versa); similarly, viruliferous males of the Q biotype are unable to transmit the virus to females of the B biotype (and vice versa). These findings support the hypothesis that a pre-zygotic mating barrier between the Q and B biotypes is the cause for the absence of gene flow between the two biotypes, and that virus transmission can be used as a marker for inter-biotype mating. To be transmitted during mating, the virus needs to be present in the haemolymph of the donor insect. Abutilon mosaic virus (AbMV), a bipartite begomovirus that can be ingested but not transmitted by B. tabaci, is absent in the whitefly haemolymph, and cannot be transmitted during mating. Mating was a precondition for horizontal virus transfer from male to female, or female to male. Virus was not transmitted when viruliferous B. tabaci were caged with the non-vector non-viruliferous whitefly Trialeurodes vaporariorum (Westwood) (Aleyrodidae: Hemiptera) and vice versa.     

Occurrence and distribution of potyviruses infecting sorghum in Kaduna and Kano States,Nigeria

Field surveys were conducted during the 2012 wet season in five Local Government Areas (LGAs) each of Kaduna and Kano States, Nigeria, to determine the incidence of potyviruses infecting sorghum. Sorghum leaves (n = 450) showing mosaic, streak, dwarfism and asymptomatic ones were collected from 30 fields. Based on the fields, the virus disease incidence in Kaduna State was 19.3, 17.1, 53.7, 35.4 and 32.7% for Sabon Gari, Giwa, Lere, Kubau and Makarfi LGAs, respectively. In Kano State, the incidence was 13.3, 32.9, 28.9, 34.3 and 50.9% for Tsanyawa, Minjibir, Wudil, Sumaila and Garun Malam LGAs, respectively. Enzyme-linked immunosorbent assay (ELISA) was employed in the detection of the viruses. There was overall incidence of Sorghum mosaic virus 2.8%, Maize dwarf mosaic virus 2.0%, Sugarcane mosaic virus 3.3% and unidentified potyvirus 4.2% from the two states.