Relative quantitation of virus population size in mixed genotype infections using sequencing chromatograms

Hall, Gerod S.; Little, Damon P.

doi:10.1016/j.jviromet.2007.05.029

Cited by 24 publications

(29 citation statements)

References 17 publications

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“…2). Analyzing sequence data in this manner has successfully been used to identify heterozygotes (10), to detect single nucleotide polymorphisms in pooled samples of DNA (3,34,36), and to distinguish viral variants (8). Figure 1C shows the change in proportion of each mutant virus compared with its own wild type after 7.5 days of incubation.…”

Section: Resultsmentioning

confidence: 99%

The Role of Evolutionary Intermediates in the Host Adaptation of Canine Parvovirus

Stucker

Pagán

Cifuente

et al. 2012

J Virol

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O ngoing viral evolution may generate important phenotypic variants, including strains that differ in host range, transmission mechanisms and efficiency, tissue tropism, antigenicity, and/or virulence. Although these novel biological functions often require multiple and concerted genomic changes, how such combinations of mutations can arise and be favored by natural selection is unclear. The complexity of the evolutionary pathways leading to the appearance of phenotypic variants is compounded when they alter the host range of viruses, as mutations may have different fitness consequences in different host species (24,27,32). Mutations may also be subject to complex selection pressures in the same host when, for example, receptor binding and antibody recognition sites overlap on the viral capsid, so that selection pressures differ between immunologically naïve and immune individuals (31). Understanding the processes by which viruses acquire new phenotypes in the face of such complex selective environments is critical for improving the prediction, prevention, and control strategies for emerging viral diseases.Feline panleukopenia virus (FPV) and closely related viruses that infect many hosts within the order Carnivora have undergone processes of cross-species transmission and adaptation during the last 3 decades, providing a powerful opportunity to examine the evolution and adaptation of a novel pandemic virus in the context of new host environments. Canine parvovirus type 2 (CPV-2) is a host-range variant of a virus closely related to FPV that gained the ability to infect dogs through the acquisition of capsid mutations that altered the interaction of virus capsids with the transferrin receptor type 1 (TfR) on the surface of canine cells (14,29). CPV-2 was the original virus strain in dogs that spread worldwide during 1978 and that was completely replaced during 1979 and 1980 by a new variant (CPV-2a). The CPV-2a strain is genetically and antigenically distinct from CPV-2 (17, 22) and is presumably better adapted to its canine host since it replaced CPV-2 in nature. It is now clear that the emergence of CPV involved a number of hostswitching events between cats, dogs, raccoons, and possibly other carnivores, with multiple transfers occurring among the different hosts (2). While the emergence of CPV-2a was previously attributed to host adaptation in dogs, recent studies have shown that this adaptive process likely involved transfer of CPV-2a to and from raccoons (2). Raccoon infection also involved a host-range change in the virus, as the raccoon viruses lost the canine host range and appeared to carry host-specific mutations that were likely adaptive for raccoons (2). The emergence of CPV-2a also involved a host-range expansion, as the original CPV-2 did not replicate in cats, while CPV-2a isolates replicated efficiently and caused disease in cats (28). This represented a novel hostadaptation event, as CPV-2a did not show significant reversion back to the original FPV sequences (28). CPV-2a isolates also showed al...

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Section: Resultsmentioning

confidence: 99%

The Role of Evolutionary Intermediates in the Host Adaptation of Canine Parvovirus

Stucker

Pagán

Cifuente

et al. 2012

J Virol

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“…RNA was isolated from systemic leaves upon the appearance of mosaic disease symptoms at 10 dpi. The relative amount of each virus was determined by quantitative sequencing (42). Each bar represents the mean of three independent plants ± SE.…”

Section: Discussionmentioning

confidence: 99%

“…Quantitative sequencing for the detection and quantification of the G-to-A TMV-V1087I point mutation was done as described in ref. 42. See SI Experimental Procedures for details.…”

Section: Methodsmentioning

confidence: 99%

“…At 10 dpi symptomatic systemic leaf tissue was harvested, and cDNA was generated from total RNA. Quantitative sequencing for the V1087I mutation was used to determine the relative concentrations of each virus in the systemic tissue (42). Results indicated that inoculation ratios of greater than five times TMV-V1087I to TMV were required for TMV-V1087I to be the predominant systemic virus (Fig.…”

Section: Stabilization Of Iaa26 Expressed From Its Native Phloem Prommentioning

confidence: 99%

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Tobacco mosaic virus-directed reprogramming of auxin/indole acetic acid protein transcriptional responses enhances virus phloem loading

Collum

Padmanabhan

Hsieh

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Vascular phloem loading has long been recognized as an essential step in the establishment of a systemic virus infection. In this study, an interaction between the replication protein of tobacco mosaic virus (TMV) and phloem-specific auxin/indole acetic acid (Aux/IAA) transcriptional regulators was found to modulate virus phloem loading in an age-dependent manner. Promoter expression studies show that in mature tissues TMV 126/183-kDa-interacting Aux/IAAs predominantly express and accumulate within the nuclei of phloem companion cells (CCs). Furthermore, CC Aux/IAA nuclear localization is disrupted upon infection with an interacting virus. In situ analysis of virus spread shows that the inability to disrupt Aux/IAA CC nuclear localization correlates with a reduced ability to load into the vascular tissue. Subsequent systemic movement assays also demonstrate that a virus capable of disrupting Aux/IAA localization is significantly more competitive at moving out of older plant tissues than a noninteracting virus. Similarly, CC expression and overaccumulation of a degradation-resistant Aux/IAA-interacting protein was found to inhibit TMV accumulation and phloem loading selectively in flowering plants. Transcriptional expression studies demonstrate a role for Aux/IAAinteracting proteins in the regulation of salicylic and jasmonic acid host defense responses as well as virus-specific movement factors, including pectin methylesterase, that are involved in regulating plasmodesmata size-exclusion limits and promoting virus cell-to-cell movement. Combined, these findings indicate that TMV directs the reprogramming of auxin-regulated gene expression within the vascular phloem of mature tissues as a means to enhance phloem loading and systemic spread.pathogen defense | plant hormone signaling | virus movement | plasmodesmata gating | age-related resistance

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“…Midguts were collected at 7, 14, 21, and 28 days following the second feeding. At 14 dpi, via polySNP analysis (Fitzpatrick et al, 2010; Hall and Little, 2007), mosquito midguts secondarily infected with 2K-V9M had a significantly higher proportion of the superinfecting genotype than did those superinfected by wildtype (Fig. 2) (Mann-Whitney, p value=0.0194).…”

Section: Resultsmentioning

confidence: 99%

A positively selected mutation in the WNV 2K peptide confers resistance to superinfection exclusion in vivo

Campbell¹,

Smith²,

Sánchez-Vargas³

et al. 2014

Virology

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Molecular epidemiologic studies of North American (NA) West Nile virus (WNV; Flaviviridae, Flavivirus) have documented the displacement of the introduced NY99 genotype with WN02. In addition, these studies have shown that particular substitutions are under positive selection. One occurs in the C-terminus of the NS4A coding sequence and results in a valine to methionine substitution at position nine of the 2K peptide. 2K-V9M confers the ability to overcome superinfection exclusion in vitro. We hypothesized that WNV strains bearing 2K-V9M have higher fitness than wildtype in Culex quinquefasciatus mosquitoes. Although infection rates and viral titers were not significantly different, virus dissemination rates were significantly higher with WNV 2K-V9M. As a super-infecting virus, WNV 2K-V9M was more successful than wildtype, however, in a mixed infection, 2K-V9M was not. These data support observations that 2K-V9M confers a context-specific selective advantage in mosquitoes and provides an in vivo mechanism for its positive selection.

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Relative quantitation of virus population size in mixed genotype infections using sequencing chromatograms

Cited by 24 publications

References 17 publications

The Role of Evolutionary Intermediates in the Host Adaptation of Canine Parvovirus

The Role of Evolutionary Intermediates in the Host Adaptation of Canine Parvovirus

Tobacco mosaic virus-directed reprogramming of auxin/indole acetic acid protein transcriptional responses enhances virus phloem loading

A positively selected mutation in the WNV 2K peptide confers resistance to superinfection exclusion in vivo

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