Showing papers by "Siobain Duffy published in 2011"

Single-Cell Genomics Reveals Organismal Interactions in Uncultivated Marine Protists

Abstract: Whole-genome shotgun sequence data from three individual cells isolated from seawater, followed by analysis of ribosomal DNA, indicated that the cells represented three divergent clades of picobiliphytes. In contrast with the recent description of this phylum, we found no evidence of plastid DNA nor of nuclear-encoded plastid-targeted proteins, which suggests that these picobiliphytes are heterotrophs. Genome data from one cell were dominated by sequences from a widespread single-stranded DNA virus. This virus was absent from the other two cells, both of which contained non-eukaryote DNA derived from marine Bacteroidetes and large DNA viruses. By using shotgun sequencing of uncultured marine picobiliphytes, we revealed the distinct interactions of individual cells.

Exploring the diversity of plant DNA viruses and their satellites using vector-enabled metagenomics on whiteflies

Abstract: Current knowledge of plant virus diversity is biased towards agents of visible and economically important diseases. Less is known about viruses that have not caused major diseases in crops, or viruses from native vegetation, which are a reservoir of biodiversity that can contribute to viral emergence. Discovery of these plant viruses is hindered by the traditional approach of sampling individual symptomatic plants. Since many damaging plant viruses are transmitted by insect vectors, we have developed "vector-enabled metagenomics" (VEM) to investigate the diversity of plant viruses. VEM involves sampling of insect vectors (in this case, whiteflies) from plants, followed by purification of viral particles and metagenomic sequencing. The VEM approach exploits the natural ability of highly mobile adult whiteflies to integrate viruses from many plants over time and space, and leverages the capability of metagenomics for discovering novel viruses. This study utilized VEM to describe the DNA viral community from whiteflies (Bemisia tabaci) collected from two important agricultural regions in Florida, USA. VEM successfully characterized the active and abundant viruses that produce disease symptoms in crops, as well as the less abundant viruses infecting adjacent native vegetation. PCR assays designed from the metagenomic sequences enabled the complete sequencing of four novel begomovirus genome components, as well as the first discovery of plant virus satellites in North America. One of the novel begomoviruses was subsequently identified in symptomatic Chenopodium ambrosiodes from the same field site, validating VEM as an effective method for proactive monitoring of plant viruses without a priori knowledge of the pathogens. This study demonstrates the power of VEM for describing the circulating viral community in a given region, which will enhance our understanding of plant viral diversity, and facilitate emerging plant virus surveillance and management of viral diseases.

Advances in plant virus evolution: translating evolutionary insights into better disease management.

Abstract: Recent studies in plant virus evolution are revealing that genetic structure and behavior of virus and viroid populations can explain important pathogenic properties of these agents, such as host resistance breakdown, disease severity, and host shifting, among others. Genetic variation is essential for the survival of organisms. The exploration of how these subcellular parasites generate and maintain a certain frequency of mutations at the intra- and inter-host levels is revealing novel molecular virus-plant interactions. They emphasize the role of host environment in the dynamic genetic composition of virus populations. Functional genomics has identified host factors that are transcriptionally altered after virus infections. The analyses of these data by means of systems biology approaches are uncovering critical plant genes specifically targeted by viruses during host adaptation. Also, a next-generation resequencing approach of a whole virus genome is opening new avenues to study virus recombination and the relationships between intra-host virus composition and pathogenesis. Altogether, the analyzed data indicate that systematic disruption of some specific parameters of evolving virus populations could lead to more efficient ways of disease prevention, eradication, or tolerable virus-plant coexistence.

Occurrence of grapevine leafroll-associated virus complex in Napa Valley.

Abstract: Grapevine leafroll disease (GLD) is caused by a complex of several virus species (grapevine leafroll-associated viruses, GLRaV) in the family Closteroviridae. Because of its increasing importance, it is critical to determine which species of GLRaV is predominant in each region where this disease is occurring. A structured sampling design, utilizing a combination of RT-PCR based testing and sequencing methods, was used to survey GLRaVs in Napa Valley (California, USA) vineyards (n = 36). Of the 216 samples tested for GLRaV-1, -2, -3, -4, -5, and -9, 62% (n = 134) were GLRaV positive. Of the positives, 81% (n = 109) were single infections with GLRaV-3, followed by GLRaV-2 (4%, n = 5), while the remaining samples (15%, n = 20) were mixed infections of GLRaV-3 with GLRaV-1, 2, 4, or 9. Additionally, 468 samples were tested for genetic variants of GLRaV-3, and of the 65% (n = 306) of samples positive for GLRaV-3, 22% were infected with multiple GLRaV-3 variants. Phylogenetic analysis utilizing sequence data from the single infection GLRaV-3 samples produced seven well-supported GLRaV-3 variants, of which three represented 71% of all GLRaV-3 positive samples in Napa Valley. Furthermore, two novel variants, which grouped with a divergent isolate from New Zealand (NZ-1), were identified, and these variants comprised 6% of all positive GLRaV-3 samples. Spatial analyses showed that GLRaV-3a, 3b, and 3c were not homogeneously distributed across Napa Valley. Overall, 86% of all blocks (n = 31) were positive for GLRaVs and 90% of positive blocks (n = 28) had two or more GLRaV-3 variants, suggesting complex disease dynamics that might include multiple insect-mediated introduction events.

Genetic Diversity in the 3′ Terminal 4.7-kb Region of Grapevine leafroll-associated virus 3

Abstract: Wang, J., Sharma, A. M., Duffy, S., and Almeida, R. P. P. 2011. Genetic diversity in the 3′ terminal 4.7-kb region of Grapevine leafroll-associated virus 3. Phytopathology 101:445-450. Grapevine leafroll-associated virus 3 (GLRaV-3; Ampelovirus, Closteroviridae), associated with grapevine leafroll disease, is an important pathogen found across all major grape-growing regions of the world. The genetic diversity of GLRaV-3 in Napa Valley, CA, was studied by sequencing 4.7 kb in the 3′ terminal region of 50 isolates obtained from Vitis vinifera ‘Merlot’. GLRaV-3 isolates were subdivided into four distinct phylogenetic clades. No evidence of positive selection was observed in the data set, although neutral selection (ratio of nonsynonymous to synonymous substitution rates = 1.1) was observed in one open reading frame (ORF 11, p4). Additionally, the four clades had variable degrees of overall nucleotide diversity. Moreover, no geographical structure among isolates was observed, and isolates belonging to different phylogenetic clades were found in distinct vineyards, with one exception. Considered with the evidence of purifying selection (i.e., against deleterious mutations), these data indicate that the population of GLRaV-3 in Napa Valley is not expanding and its effective population size is not increasing. Furthermore, research on the biological characterization of GLRaV-3 strains might provide valuable insights on the biology of this species that may have epidemiological relevance.

Genus-Specific Substitution Rate Variability Among Picornaviruses

Abstract: Picornaviruses have some of the highest nucleotide substitution rates among viruses, but there have been no comparisons of evolutionary rates within this broad family. We combined our own Bayesian coalescent analyses of VP1 regions from four picornaviruses with 22 published VP1 rates to produce the first within-family meta-analysis of viral evolutionary rates. Similarly, we compared our rate estimates for the RNA polymerase 3D(pol) gene from five viruses to four published 3D(pol) rates. Both a structural and a nonstructural gene show that enteroviruses are evolving, on average, a half order of magnitude faster than members of other genera within the Picornaviridae family.

Single-stranded genomic architecture constrains optimal codon usage.

Abstract: Viral codon usage is shaped by the conflicting forces of mutational pressure and selection to match host patterns for optimal expression. We examined whether genomic architecture (single- or double-stranded DNA) influences the degree to which bacteriophage codon usage differ from their primary bacterial hosts and each other. While both correlated equally with their hosts’ genomic nucleotide content, the coat genes of ssDNA phages were less well adapted than those of dsDNA phages to their hosts’ codon usage profiles due to their preference for codons ending in thymine. No specific biases were detected in dsDNA phage genomes. In all nine of ten cases of codon redundancy in which a specific codon was overrepresented, ssDNA phages favored the NNT codon. A cytosine to thymine biased mutational pressure working in conjunction with strong selection against non-synonymous mutations appears be shaping codon usage bias in ssDNA viral genomes.