RESUMO
Sugarcane leaf fleck incited by Sugarcane bacilliform virus is emerging as a major disease and affecting exchange of sugarcane germplasm and cultivation worldwide. Roving surveys conducted in 162 fields belonging to 81 villages spread over 14 sugarcane growing districts of Andhra Pradesh during 2021-2022 revealed 8 to 44% incidence of the disease. Mean maximum fleck disease incidence was reported in Anakapalli district (33.00%) followed by Srikakulam district (22.66%), whereas least incidence was observed in Alluri Sitharamaraju district (9.33%). The early and sensitive detection of pathogens is vital and necessary to reduce the danger of introducing new diseases or pathogen strains into sugarcane growing regions. Both serological and molecular methods were used in proposed investigation to identify the virus at the protein and nucleic acid levels. DAS-ELISA results were positive for 50 suspected SCBV infected sugarcane leaf samples out of 81, with mean absorbance (A405) values ranging from 0.50 to 2.20. Further PCR assays were performed using SCBV-specific primers targeting RT/RNase H coding region which is frequently employed as a taxonomic marker for species delineation in Badnaviruses. Out of 81 symptomatic samples collected, 61 samples gave positive results, and no amplification was observed in healthy control and negative control. Results made it evident that PCR was more sensitive than DAS-ELISA. Low virus concentration or variation in virus strains may be the reason for the low detection rate in DAS-ELISA in the current study. Extensive roving surveys conducted for the incidence of leaf fleck disease for the first time in the state of Andhra Pradesh revealed severe occurrence of leaf fleck disease under field conditions.
Assuntos
Badnavirus , Saccharum , Badnavirus/genética , Plantas , Reação em Cadeia da PolimeraseRESUMO
Cocoa, Theobroma cacao, is an important tropical perennial crop grown widely in the humid tropics. The exchange of cocoa germplasm between germplasm collections and breeding centres is vital for varietal development. Intermediate quarantine facilities, such as the International Cocoa Quarantine Centre, Reading UK (ICQC-R) play a vital role in ensuring the transfer of germplasm whilst minimising the risk of spreading pests and diseases. Current screening procedures combine visual inspection and molecular techniques, which are effective in detecting Cocoa swollen shoot virus (CSSV), a badnavirus, which causes severe losses but are restricted to West Africa. However, the detection of latent or mild virus infections that produce no visual symptoms has been a challenge. Recently two badnavirus species of cocoa producing mild symptoms, cacao mild mosaic virus (CaMMV) and cacao yellow vein-banding virus (CYVBV), have been sequenced. Here, we report new assays for the detection of these two species, for the first time in non-symptomatic accessions. Evolutionary and bioinformatic analyses of the viruses suggest their most recent source was from Trinidad, though there is historic evidence that these viruses may have their origin in South America and then become widespread globally over the last century. We also report a novel colorimetric Loop-mediated isothermal amplification (LAMP) assay for the detection of CYVBV. This simple and accurate method could be employed in field virus testing.
Assuntos
Cacau/virologia , Vírus do Mosaico/classificação , Vírus do Mosaico/isolamento & purificação , África Ocidental , Badnavirus/classificação , Badnavirus/genética , Badnavirus/isolamento & purificação , Genoma Viral , Vírus do Mosaico/genética , Filogenia , Doenças das Plantas/virologia , Vírus de Plantas/classificação , Vírus de Plantas/genética , Vírus de Plantas/isolamento & purificação , Alinhamento de Sequência , Análise de Sequência de DNA , América do Sul , Trinidad e TobagoRESUMO
To analyze the DNA virome associated with cacao (Theobroma cacao L.) trees showing virus-like symptoms in Brazil (BR) and Puerto Rico (PR) during 2018-2019, total DNA was isolated from symptomatic leaves and subjected to high-throughput Illumina sequencing. The assembled complete badnaviral genome sequences were verified by PCR amplification, cloning, and DNA sequencing. Based on pairwise distances and phylogenetic analysis, three badnaviral genomes were identified, and these viruses were found to be isolates of the previously described cacao mild mosaic virus (CaMMV). The three genomes were 7,520, 7,524, and 7,514 bp in size for the isolates CaMMV-BR321, CaMMV-BR322, and CaMMV-PR3, respectively. Each genome contained four predicted open reading frames: ORFs 1-3 and ORFY. The CaMMV-PR3 isolate was identified as a probable recombinant, with a CaMMV-BR-like virus as the major parent.
Assuntos
Cacau/virologia , Genoma Viral/genética , Vírus do Mosaico/genética , Doenças das Plantas/virologia , Recombinação Genética/genética , Badnavirus/genética , Brasil , Sequenciamento de Nucleotídeos em Larga Escala , Fases de Leitura Aberta/genética , Filogenia , Porto Rico , Análise de Sequência de DNA/métodos , Sequenciamento Completo do Genoma/métodosRESUMO
Yam (Dioscorea spp.) is an important crop in tropical and subtropical regions. Many viruses have been recently identified in yam, hampering genetic conservation and safe international exchanges of yam germplasm. We report on the implementation of reliable and cost-effective PCR-based detection tools targeting eight different yam-infecting viruses. Viral indexing of the in vitro yam collection maintained by the Biological Resources Center for Tropical Plants (BRC-TP) in Guadeloupe (French West Indies) unveiled a high prevalence of potyviruses, badnaviruses, Dioscorea mosaic associated virus (DMaV) and yam asymptomatic virus 1 (YaV1) and a high level of coinfections. Infected yam accessions were subjected to a combination of thermotherapy and meristem culture. Sanitation levels were monitored using PCR-based and high-throughput sequencing-based diagnosis, confirming the efficacy and reliability of PCR-based detection tools. Sanitation rates were highly variable depending on viruses. Sixteen accessions were successfully sanitized, paving the way to safe yam germplasm exchanges and the implementation of clean seed production programs worldwide.
Assuntos
Dioscorea/virologia , Patologia Molecular/métodos , Doenças das Plantas/virologia , Vírus de Plantas/isolamento & purificação , Saneamento/métodos , Badnavirus/genética , Badnavirus/isolamento & purificação , Vírus de Plantas/genética , Reação em Cadeia da Polimerase/métodos , Potexvirus/genética , Potexvirus/isolamento & purificação , Reprodutibilidade dos Testes , Índias OcidentaisRESUMO
Thirty-five unique partial sugarcane bacilliform virus (SCBV) sequences extending over 529 bp were identified in sugarcane samples from Guadeloupe diagnosed by Immunocapture-PCR (IC-PCR) using specific badnavirus primers. Phylogenetic analysis of these sequences along with the two known genome sequences of Sugarcane bacilliform Mor virus (SCBMV) and Sugarcane bacilliform IM virus (SCBIMV) revealed high molecular variability in the SCBV genome. Seven phylogenetic groups, named A to G, were characterized: virus isolates from groups A-B, C and D are proposed to be members of three additional SCBV species. The two (7446 and 7444 bp) and one (7317 bp) complete sequences of SCBV isolates from groups A and D, respectively, likely represented the genome of two new species. Phylogenetic analysis of the complete genome and RT/RNase H sequences confirmed the polyphyletic structure of SCBV isolates and the absence of a clear separation between SCBV and Banana streak virus (BSV) isolates within badnavirus group 1. These results showed that reconsideration of taxonomy and classification of SCBV and BSV are necessary.