Bacilliform DNA-containing plant viruses in the tropics: commonalities within a genetically diverse group.
ABSTRACT: UNLABELLED:Plant viruses, possessing a bacilliform shape and containing double-stranded DNA, are emerging as important pathogens in a number of agricultural and horticultural crops in the tropics. They have been reported from a large number of countries in African and Asian continents, as well as from islands from the Pacific region. The viruses, belonging to two genera, Badnavirus and Tungrovirus, within the family Caulimoviridae, have genomes displaying a common plan, yet are highly variable, sometimes even between isolates of the same virus. In this article, we summarize the current knowledge with a view to revealing the common features embedded within the genetic diversity of this group of viruses. TAXONOMY:Virus; order Unassigned; family Caulimoviridae; genera Badnavirus and Tungrovirus; species Banana streak viruses, Bougainvillea spectabilis chlorotic vein banding virus, Cacao swollen shoot virus, Citrus yellow mosaic badnavirus, Dioscorea bacilliform viruses, Rice tungro bacilliform virus, Sugarcane bacilliform viruses and Taro bacilliform virus. MICROBIOLOGICAL PROPERTIES:Bacilliform in shape; length, 60-900?nm; width, 35-50?nm; circular double-stranded DNA of approximately 7.5?kbp with one or more single-stranded discontinuities. HOST RANGE:Each virus generally limited to its own host, including banana, bougainvillea, black pepper, cacao, citrus species, Dioscorea alata, rice, sugarcane and taro. DISEASE SYMPTOMS:Foliar streaking in banana and sugarcane, swelling of shoots in cacao, yellow mosaic in leaves and stems in citrus, brown spot in the tubers in yam and yellow-orange discoloration and stunting in rice. USEFUL WEBSITES:http://www.dpvweb.net.
Project description:Badnaviruses (Family: Caulimoviridae; Genus: Badnavirus) are non-enveloped bacilliform DNA viruses with a monopartite genome containing about 7.2 to 9.2 kb of dsDNA with three to seven open reading frames. They are transmitted by mealybugs and a few species by aphids in a semi-persistent manner. They are one of the most important plant virus groups and have emerged as serious pathogens affecting the cultivation of several horticultural crops in the tropics, especially banana, black pepper, cocoa, citrus, sugarcane, taro, and yam. Some badnaviruses are also known as endogenous viruses integrated into their host genomes and a few such endogenous viruses can be awakened, e.g., through abiotic stress, giving rise to infective episomal forms. The presence of endogenous badnaviruses poses a new challenge for the fool-proof diagnosis, taxonomy, and management of the diseases. The present review aims to highlight emerging disease problems, virus characteristics, transmission, and diagnosis of badnaviruses.
Project description:Sugarcane bacilliform viruses (SCBV), which belong to the genus Badnavirus, family Caulimoviridae, are an important DNA virus complex that infects sugarcane. To explore the genetic diversity of the sugarcane-infecting badnavirus complex in China, we tested 392 sugarcane leaf samples collected from Fujian, Yunnan, and Hainan provinces for the occurrence of SCBV by polymerase chain reaction (PCR) assays using published primers SCBV-F and SCBV-R that target the reverse transcriptase/ribonuclease H (RT/RNase H) regions of the viral genome. A total of 111 PCR-amplified fragments (726 bp) from 63 SCBV-positive samples were cloned and sequenced. A neighbor-joining phylogenetic tree was constructed based on the SCBV sequences from this study and 34 published sequences representing 18 different phylogroups or genotypes (SCBV-A to -R). All SCBV-tested isolates could be classified into 20 SCBV phylogenetic groups from SCBV-A to -T. Of nine SCBV phylogroups reported in this study, two novel phylogroups, SCBV-S and SCBV-T, that share 90.0-93.2% sequence identity and show 0.07-0.11 genetic distance with each other in the RT/ RNase H region, are proposed. SCBV-S had 57.6-92.2% sequence identity and 0.09-0.66 genetic distance, while SCBV-T had 58.4-90.0% sequence identity and 0.11-0.63 genetic distance compared with the published SCBV phylogroups. Additionally, two other Badnavirus species, Sugarcane bacilliform MO virus (SCBMOV) and Sugarcane bacilliform IM virus (SCBIMV), which originally clustered in phylogenetic groups SCBV-E and SCBV-F, respectively, are first reported in China. Our findings will help to understand the level of genetic heterogeneity present in the complex of Badnavirus species that infect sugarcane.
Project description:Yam (Dioscorea spp.) plants are potentially hosts to a diverse range of badnavirus species (genus Badnavirus, family Caulimoviridae), but their detection is complicated by the existence of integrated badnavirus sequences in some yam genomes. To date, only two badnavirus genomes have been characterised, namely, Dioscorea bacilliform AL virus (DBALV) and Dioscorea bacilliform SN virus (DBSNV). A further 10 tentative species in yam have been described based on their partial reverse transcriptase (RT)-ribonuclease H (RNaseH) sequences, generically referred to here as Dioscorea bacilliform viruses (DBVs). Further characterisation of DBV species is necessary to determine which represent episomal viruses and which are only present as integrated badnavirus sequences in some yam genomes. In this study, a sequence-independent multiply-primed rolling circle amplification (RCA) method was evaluated for selective amplification of episomal DBV genomes. This resulted in the identification and characterisation of nine complete genomic sequences (7.4-7.7 kbp) of existing and previously undescribed DBV phylogenetic groups from Dioscorea alata and Dioscorea rotundata accessions. These new yam badnavirus genomes expand our understanding of the diversity and genomic organisation of DBVs, and assist the development of improved diagnostic tools. Our findings also suggest that mixed badnavirus infections occur relatively often in West African yam germplasm.
Project description:Sugarcane Bacilliform Guadeloupe A Virus (SCBGAV, genus Badnavirus, family Caulimoviridae) is an emerging, deleterious pathogen of sugarcane which presents a substantial barrier to producing high sugarcane earnings. Sugarcane bacilliform viruses (SCBVs) are one of the main species that infect sugarcane. During the last 30 years, significant genetic changes in SCBV strains have been observed with a high risk of disease incidence associated with crop damage. SCBV infection may lead to significant losses in biomass production in susceptible sugarcane cultivars. The circular, double-stranded (ds) DNA genome of SCBGAV (7.4 Kb) is composed of three open reading frames (ORFs) on the positive strand that replicate by a reverse transcriptase. SCBGAV can infect sugarcane in a semipersistent manner via the insect vectors sugarcane mealybug species. In the current study, we used miRNA target prediction algorithms to identify and comprehensively analyze the genome-wide sugarcane (Saccharum officinarum L.)-encoded microRNA (miRNA) targets against the SCBGAV. Mature miRNA target sequences were retrieved from the miRBase (miRNA database) and were further analyzed for hybridization to the SCBGAV genome. Multiple computational approaches-including miRNA-target seed pairing, multiple target positions, minimum free energy, target site accessibility, maximum complementarity, pattern recognition and minimum folding energy for attachments-were considered by all algorithms. Among them, sof-miR396 was identified as the top effective candidate, capable of targeting the vital ORF3 of the SCBGAV genome. miRanda, RNA22 and RNAhybrid algorithms predicted hybridization of sof-miR396 at common locus position 3394. The predicted sugarcane miRNAs against viral mRNA targets possess antiviral activities, leading to translational inhibition by mRNA cleavage. Interaction network of sugarcane-encoded miRNAs with SCBGAV genes, created using Circos, allow analyze new targets. The finding of the present study acts as a first step towards the creation of SCBGAV-resistant sugarcane through the expression of the identified miRNAs.
Project description:Yams (Dioscorea spp.) host a diverse range of badnaviruses (genus Badnavirus, family Caulimoviridae). The first complete genome sequence of Dioscorea bacilliform RT virus 3 (DBRTV3), which belongs to the monophyletic species group K5, is described. This virus is most closely related to Dioscorea bacilliform SN virus (DBSNV, group K4) based on a comparison of genome sequences. Recombination analysis identified a unique recombination event in DBRTV3, with DBSNV likely to be the major parent and Dioscorea bacilliform AL virus (DBALV) the minor parent, providing the first evidence for recombination in yam badnaviruses. This has important implications for yam breeding programmes globally.
Project description:RNA silencing is an antiviral immunity that regulates gene expression through the production of small RNAs (sRNAs). In this study, deep sequencing of small RNAs was used to identify viruses infecting two taro plants. Blast searching identified five and nine contigs assembled from small RNAs of samples T1 and T2 matched onto the genome sequences of badnaviruses in the family Caulimoviridae. Complete genome sequences of two isolates of the badnavirus determined by sequence specific amplification comprised of 7,641 nucleotides and shared overall nucleotide similarities of 44.1%?55.8% with other badnaviruses. Six open reading frames (ORFs) were identified on the plus strand, showed amino acid similarities ranging from 59.8% (ORF3) to 10.2% (ORF6) to the corresponding proteins encoded by other badnaviruses. Phylogenetic analysis also supports that the virus is a new member in the genus Badnavirus. The virus is tentatively named as Taro bacilliform CH virus (TaBCHV), and it is the second badnavirus infecting taro plants, following Taro bacilliform virus (TaBV). In addition, analyzes of viral derived small RNAs (vsRNAs) from TaBCHV showed that almost equivalent number of vsRNAs were generated from both strands and the most abundant vsRNAs were 21 nt, with uracil bias at 5' terminal. Furthermore, TaBCHV vsRNAs were asymmetrically distributed on its entire circular genome at both orientations with the hotspots mainly generated in the ORF5 region.
Project description:Sugarcane-infecting badnaviruses (sugarcane bacilliform viruses, SCBVs) represent a genetically heterogeneous species complex, posing a serious threat to the yield and quality of sugarcane in all major producing regions. SCBVs are commonly transmitted across regions by the exchange of sugarcane germplasm. In this study, we develop two quick, sensitive, and reliable protocols for real-time quantitative PCR (qPCR) of Sugarcane bacilliform MO virus (SCBMOV) and Sugarcane bacilliform IM virus (SCBIMV) using two sets of TaqMan probes and primers targeting the reverse transcriptase/ribonuclease H (RT/RNase H) region. The two assays had a detection limit of 100 copies of plasmid DNA and were 100 times more sensitive than conventional PCR. High specificity of the two assays was observed with respect to SCBIMV and SCBMOV. A total of 176 sugarcane leaf tissue samples from Fujian and Yunnan provinces were collected and analyzed in parallel by conventional PCR, SCBIMV-qPCR, and SCBMOV-qPCR. The SCBIMV-qPCR and SCBMOV-qPCR assays indicated that 50% (88/176) and 47% (83/176) samples tested positive, respectively, whereas only 29% (51/176) tested positive with conventional PCR with the primer pairs SCBV-F and SCBV-R. We demonstrate for the first time that SCBIMV and SCBMOV occur in China and reveal coinfection of both Badnavirus species in 29% (51/176) of tested leaf samples. Our findings supply sensitive and reliable qPCR assays for the detection and quantitation of SCBV in sugarcane quarantine programs.
Project description:Common bean (Phaseolus vulgaris L.) is the primary source of protein and nutrients in the majority of households in sub-Saharan Africa. However, pests and viral diseases are key drivers in the reduction of bean production. To date, the majority of viruses reported in beans have been RNA viruses. In this study, we carried out a viral metagenomic analysis on virus symptomatic bean plants. Our virus detection pipeline identified three viral fragments of the double-stranded DNA virus Pelargonium vein banding virus (PVBV) (family, Caulimoviridae, genus Badnavirus). This is the first report of the dsDNA virus and specifically PVBV in legumes to our knowledge. In addition two previously reported +ssRNA viruses the bean common mosaic necrosis virus (BCMNVA) (Potyviridae) and aphid lethal paralysis virus (ALPV) (Dicistroviridae) were identified. Bayesian phylogenetic analysis of the Badnavirus (PVBV) using amino acid sequences of the RT/RNA-dependent DNA polymerase region showed the Kenyan sequence (SRF019_MK014483) was closely matched with two Badnavirus viruses: Dracaena mottle virus (DrMV) (YP_610965) and Lucky bamboo bacilliform virus (ABR01170). Phylogenetic analysis of BCMNVA was based on amino acid sequences of the Nib region. The BCMNVA phylogenetic tree resolved two clades identified as clade (I and II). Sequence from this study SRF35_MK014482, clustered within clade I with other Kenyan sequences. Conversely, Bayesian phylogenetic analysis of ALPV was based on nucleotide sequences of the hypothetical protein gene 1 and 2. Three main clades were resolved and identified as clades I-III. The Kenyan sequence from this study (SRF35_MK014481) clustered within clade II, and nested within a sub-clade; comprising of sequences from China and an earlier ALPV sequences from Kenya isolated from maize (MF458892). Our findings support the use of viral metagenomics to reveal the nascent viruses, their viral diversity and evolutionary history of these viruses. The detection of ALPV and PVBV indicate that these viruses have likely been underreported due to the unavailability of diagnostic tools.
Project description:Saccharum species such as sugarcane and energy cane are key players in the expanding bioeconomy for sugars, bioenergy, and production of high-value proteins. Genomic tools such as culm-regulated promoters would be of great value in terms of improving biomass characteristics through enhanced carbon metabolism for sugar accumulation and/or fiber content for biofuel feedstock. Unlike the situation in dicots, monocot promoters currently used are limited and mostly derived from highly expressed constitutive plant genes and viruses. In this study, a novel promoter region of Sugarcane bacilliform virus (SCBV; genus Badnavirus, family Caulimoviridae), SCBV21 was cloned and mapped by deletion analysis and functionally characterized transiently in monocot and dicot species and stably in sugarcane.In silico analysis of SCBV21 [1816 base pair (bp)] identified two putative promoter regions (PPR1 and PPR2) with transcription start sites (TSS1 and TSS2) and two TATA-boxes (TATAAAT and ATATAA), and several vascular-specific and regulatory elements. Deletion analysis revealed that the 710 bp region spanning PPR2 (with TSS2 and ATATAA) at the 3' end of SCBV21 retained the full promoter activity in both dicots and monocots, as shown by transient expression of the enhanced yellow fluorescent protein (EYFP) gene. In sugarcane young leaf segments, SCBV21 directed a 1.8- and 2.4-fold higher transient EYFP expression than the common maize ubiquitin 1 (Ubi1) and Cauliflower mosaic virus 35S promoters, respectively. In transgenic sugarcane, SCBV21 conferred a preferential expression of the ?-glucuronidase (GUS) gene in leaves and culms and specifically in the culm storage parenchyma surrounding the vascular bundle and in vascular phloem cells. Among the transgenic events and tissues characterized in this study, the SCBV21 promoter frequently produced higher GUS activity than the Ubi1 or 35S promoters in a manner that was not obviously correlated with the transgene copy number.The newly developed plant viral SCBV21 promoter is distinct from the few existing SCBV promoters in its sequence and expression pattern. The potential of SCBV21 as a tissue-regulated promoter with a strong activity in the culm vascular bundle and its storage parenchyma makes it useful in sugarcane engineering for improved carbon metabolism, increased bioenergy production, and enhanced stress tolerance.
Project description:Badnaviruses (family Caulimoviridae, genus Badnavirus) have emerged as serious pathogens especially affecting the cultivation of tropical crops. Badnavirus sequences can be integrated in host genomes, complicating the detection of episomal infections and the assessment of viral genetic diversity in samples containing a complex mixture of sequences. Yam (Dioscorea spp.) plants are hosts to a diverse range of badnavirus species, and recent findings have suggested that mixed infections occur frequently in West African yam germplasm. Historically, the determination of the diversity of badnaviruses present in yam breeding lines has been achieved by cloning and sequencing of polymerase chain reaction (PCR) products. In this study, the molecular diversity of partial reverse transcriptase (RT)-ribonuclease H (RNaseH) sequences from yam badnaviruses was analysed using PCR-dependent denaturing gradient gel electrophoresis (PCR-DGGE). This resulted in the identification of complex 'fingerprints' composed of multiple sequences of Dioscorea bacilliform viruses (DBVs). Many of these sequences show high nucleotide identities to endogenous DBV (eDBV) sequences deposited in GenBank, and fall into six monophyletic species groups. Our findings highlight PCR-DGGE as a powerful tool in badnavirus diversity studies enabling a rapid indication of sequence diversity as well as potential candidate integrated sequences revealed by their conserved nature across germplasm.