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      First natural crossover recombination between two distinct species of the family Closteroviridae leads to the emergence of a new disease

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          Abstract

          Lettuce chlorosis virus-SP (LCV-SP) (family Closteroviridae, genus Crinivirus), is a new strain of LCV which is able to infect green bean plants but not lettuce. In the present study, high-throughput and Sanger sequencing of RNA was used to obtain the LCV-SP full-length sequence. The LCV-SP genome comprises 8825 nt and 8672 nt long RNA1 and RNA2 respectively. RNA1 of LCV-SP contains four ORFs, the proteins encoded by the ORF1a and ORF1b are closely related to LCV RNA1 from California (FJ380118) whereas the 3´ end encodes proteins which share high amino acid sequence identity with RNA1 of Bean yellow disorder virus (BnYDV; EU191904). The genomic sequence of RNA2 consists of 8 ORFs, instead of 10 ORFs contained in LCV-California isolate. The distribution of vsiRNA (virus-derived small interfering RNA) along the LCV-SP genome suggested the presence of subgenomic RNAs corresponding with HSP70, P6.4 and P60. Results of the analysis using RDP4 and Simplot programs are the proof of the evidence that LCV-SP is the first recombinant of the family Closteroviridae by crossover recombination of intact ORFs, being the LCV RNA1 (FJ380118) and BnYDV RNA1 (EU191904) the origin of the new LCV strain. Genetic diversity values of virus isolates in the recombinant region obtained after sampling LCV-SP infected green bean between 2011 and 2017 might suggest that the recombinant virus event occurred in the area before this period. The presence of LCV-SP shows the role of recombination as a driving force of evolution within the genus Crinivirus, a globally distributed, emergent genus.

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          Full-length human immunodeficiency virus type 1 genomes from subtype C-infected seroconverters in India, with evidence of intersubtype recombination.

          The development of an effective human immunodeficiency virus type 1 (HIV-1) vaccine is likely to depend on knowledge of circulating variants of genes other than the commonly sequenced gag and env genes. In addition, full-genome data are particularly limited for HIV-1 subtype C, currently the most commonly transmitted subtype in India and worldwide. Likewise, little is known about sequence variation of HIV-1 in India, the country facing the largest burden of HIV worldwide. Therefore, the objective of this study was to clone and characterize the complete genome of HIV-1 from seroconverters infected with subtype C variants in India. Cocultured HIV-1 isolates were obtained from six seroincident individuals from Pune, India, and virtually full-length HIV-1 genomes were amplified, cloned, and sequenced from each. Sequence analysis revealed that five of the six genomes were of subtype C, while one was a mosaic of subtypes A and C, with multiple breakpoints in env, nef, and the 3' long terminal repeat as determined by both maximal chi2 analysis and phylogenetic bootstrapping. Sequences were compared for preservation of known cytotoxic T lymphocyte (CTL) epitopes. Compared with those of the HIV-1LAI sequence, 38% of well-defined CTL epitopes were identical. The proportion of nonconservative substitutions for Env, at 61%, was higher (P < 0.001) than those for Gag (24%), Pol (18%), and Nef (32%). Therefore, characterized CTL epitopes demonstrated substantial differences from subtype B laboratory strains, which were most pronounced in Env. Because these clones were obtained from Indian seroconverters, they are likely to facilitate vaccine-related efforts in India by providing potential antigens for vaccine candidates as well as for assays of vaccine responsiveness.
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            Statistical methods for detecting molecular adaptation.

            The past few years have seen the development of powerful statistical methods for detecting adaptive molecular evolution. These methods compare synonymous and nonsynonymous substitution rates in protein-coding genes, and regard a nonsynonymous rate elevated above the synonymous rate as evidence for darwinian selection. Numerous cases of molecular adaptation are being identified in various systems from viruses to humans. Although previous analyses averaging rates over sites and time have little power, recent methods designed to detect positive selection at individual sites and lineages have been successful. Here, we summarize recent statistical methods for detecting molecular adaptation, and discuss their limitations and possible improvements.
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              Nucleic acid-based approaches to investigate microbial-related cheese quality defects

              The microbial profile of cheese is a primary determinant of cheese quality. Microorganisms can contribute to aroma and taste defects, form biogenic amines, cause gas and secondary fermentation defects, and can contribute to cheese pinking and mineral deposition issues. These defects may be as a result of seasonality and the variability in the composition of the milk supplied, variations in cheese processing parameters, as well as the nature and number of the non-starter microorganisms which come from the milk or other environmental sources. Such defects can be responsible for production and product recall costs and thus represent a significant economic burden for the dairy industry worldwide. Traditional non-molecular approaches are often considered biased and have inherently slow turnaround times. Molecular techniques can provide early and rapid detection of defects that result from the presence of specific spoilage microbes and, ultimately, assist in enhancing cheese quality and reducing costs. Here we review the DNA-based methods that are available to detect/quantify spoilage bacteria, and relevant metabolic pathways in cheeses and, in the process, highlight how these strategies can be employed to improve cheese quality and reduce the associated economic burden on cheese processors.
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                Author and article information

                Contributors
                Role: ConceptualizationRole: Data curationRole: InvestigationRole: MethodologyRole: Writing – original draftRole: Writing – review & editing
                Role: InvestigationRole: Methodology
                Role: Methodology
                Role: Data curationRole: InvestigationRole: Methodology
                Role: ConceptualizationRole: Writing – original draftRole: Writing – review & editing
                Role: Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, CA USA )
                1932-6203
                13 September 2018
                2018
                : 13
                : 9
                : e0198228
                Affiliations
                [1 ] IFAPA Centro La Mojonera, IFAPA, La Mojonera, Almería, Spain
                [2 ] IFAPA Centro Churriana, IFAPA, Churriana, Málaga, Spain
                USDA Agricultural Research Service, UNITED STATES
                Author notes

                Competing Interests: The authors have declared that no competing interests exist.

                Author information
                http://orcid.org/0000-0002-8612-966X
                http://orcid.org/0000-0001-6638-1035
                http://orcid.org/0000-0001-9294-9237
                Article
                PONE-D-18-14377
                10.1371/journal.pone.0198228
                6136708
                30212464
                a6adfc77-49cd-44fd-b650-8c09639b12d7
                © 2018 Ruiz et al

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 13 May 2018
                : 1 September 2018
                Page count
                Figures: 5, Tables: 4, Pages: 18
                Funding
                Funded by: This work was supported by AVA201601.7, 80% cofinanced by European Regional Development Fund, within Operative Program FEDER of Andalusia Goverment 2014-2020.
                Award Recipient :
                This work was supported by AVA201601.7, 80% cofinanced by European Regional Development Fund, within Operative Program FEDER of Andalusia Goverment 2014-2020.
                Categories
                Research Article
                Biology and life sciences
                Molecular biology
                Molecular biology techniques
                Sequencing techniques
                RNA sequencing
                Research and analysis methods
                Molecular biology techniques
                Sequencing techniques
                RNA sequencing
                Biology and life sciences
                Molecular biology
                Molecular biology techniques
                Biomolecular isolation
                RNA isolation
                Research and analysis methods
                Molecular biology techniques
                Biomolecular isolation
                RNA isolation
                Biology and life sciences
                Genetics
                Gene expression
                Gene regulation
                Small interfering RNAs
                Biology and life sciences
                Biochemistry
                Nucleic acids
                RNA
                Non-coding RNA
                Small interfering RNAs
                Biology and Life Sciences
                Computational Biology
                Genome Analysis
                Gene Prediction
                Biology and Life Sciences
                Genetics
                Genomics
                Genome Analysis
                Gene Prediction
                Biology and life sciences
                Genetics
                DNA
                DNA recombination
                Biology and life sciences
                Biochemistry
                Nucleic acids
                DNA
                DNA recombination
                Biology and Life Sciences
                Molecular Biology
                Molecular Biology Techniques
                Sequencing Techniques
                Nucleotide Sequencing
                Research and Analysis Methods
                Molecular Biology Techniques
                Sequencing Techniques
                Nucleotide Sequencing
                Biology and Life Sciences
                Molecular Biology
                Molecular Biology Techniques
                Artificial Gene Amplification and Extension
                Polymerase Chain Reaction
                Reverse Transcriptase-Polymerase Chain Reaction
                Research and Analysis Methods
                Molecular Biology Techniques
                Artificial Gene Amplification and Extension
                Polymerase Chain Reaction
                Reverse Transcriptase-Polymerase Chain Reaction
                Biology and Life Sciences
                Computational Biology
                Genome Analysis
                Genomic Libraries
                Biology and Life Sciences
                Genetics
                Genomics
                Genome Analysis
                Genomic Libraries
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                All relevant data are within the paper and its Supporting Information files.

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