Title: Metatranscriptome Library Preparation Influences Analyses of Viral Community Activity During a Brown Tide Bloom
There is growing interest in the use of metatranscriptomics to study virus community dynamics. We used RNA samples collected from harmful brown tides caused by the eukaryotic alga Aureococcus anophagefferens within New York (United States) estuaries and in the process observed how preprocessing of libraries by either selection for polyadenylation or reduction in ribosomal RNA (rRNA) influenced virus community analyses. As expected, more reads mapped to the A. anophagefferens genome in polyadenylation-selected libraries compared to the rRNA-reduced libraries, with reads mapped in each sample correlating to one another regardless of preprocessing of libraries. Yet, this trend was not seen for reads mapping to the Aureococcus anophagefferens Virus (AaV), where significantly more reads (approximately two orders of magnitude) were mapped to the AaV genome in the rRNA-reduced libraries. In the rRNA-reduced libraries, there was a strong and significant correlation between reads mappings to AaV and A. anophagefferens . Overall, polyadenylation-selected libraries produced fewer viral contigs, fewer reads mapped to viral contigs, and different proportions across viral realms and families, compared to their rRNA-reduced pairs. This study provides evidence that libraries generated by rRNA reduction and not selected for polyadenylation are more appropriate for quantitative characterization of viral communities in aquatic ecosystems by metatranscriptomics. more »« less
Truchon, Alexander R.; Gann, Eric R.; Wilhelm, Steven W.
(, Microbiology Resource Announcements)
Dennehy, John J.
(Ed.)
ABSTRACT. Here, we report the genomic sequence of Aureococcus anophagefferens virus, assembled into one circular contig from both Nanopore and Illumina reads. The genome is 381,717 bp long with a GC content of 29.1%, which includes an additional 5-kb region between the previously predicted polar ends of the reference genome.
Chase, Emily E; Truchon, Alexander R; Schepens, William W; Wilhelm, Steven W
(, Microbiology Resource Announcements)
Hudson, André O
(Ed.)
ABSTRACT Here, we report the draft genome ofAureococcus anophagefferensstrain CCMP1851, which is susceptible to the virusKratosvirus quantuckense. CCMP1851 complements an available genome for a virus-resistant strain (CCMP1850) isolated from the same bloom. Future studies can now use this genome to examine genetic hints of virus resistance and susceptibility.
Jackson, Elliot W.; Wilhelm, Roland C.; Buckley, Daniel H.; Hewson, Ian
(, Journal of General Virology)
Echinoderms are a phylum of marine invertebrates that include model organisms, keystone species, and animals commercially harvested for seafood. Despite their scientific, ecological, and economic importance, there is little known about the diversity of RNA viruses that infect echinoderms compared to other invertebrates. We screened over 900 transcriptomes and viral metagenomes to characterize the RNA virome of 38 echinoderm species from all five classes (Crinoidea, Holothuroidea, Asteroidea, Ophiuroidea and Echinoidea). We identified 347 viral genome fragments that were classified to genera and families within nine viral orders - Picornavirales, Durnavirales, Martellivirales, Nodamuvirales, Reovirales, Amarillovirales, Ghabrivirales, Mononegavirales, and Hepelivirales . We compared the relative viral representation across three life stages (embryo, larvae, adult) and characterized the gene content of contigs which encoded complete or near-complete genomes. The proportion of viral reads in a given transcriptome was not found to significantly differ between life stages though the majority of viral contigs were discovered from transcriptomes of adult tissue. This study illuminates the biodiversity of RNA viruses from echinoderms, revealing the occurrence of viral groups in natural populations.
Ranaviruses (Iridoviridae), including Frog Virus 3 (FV3), are large dsDNA viruses that cause devastating infections globally in amphibians, fish, and reptiles, and contribute to catastrophic amphibian declines. FV3’s large genome (~105 kb) contains at least 98 putative open reading frames (ORFs) as annotated in its reference genome. Previous studies have classified these coding genes into temporal classes as immediate early, delayed early, and late viral transcripts based on their sequential expression during FV3 infection. To establish a high-throughput characterization of ranaviral gene expression at the genome scale, we performed a whole transcriptomic analysis (RNA-Seq) using total RNA samples containing both viral and cellular transcripts from FV3-infected Xenopus laevis adult tissues using two FV3 strains, a wild type (FV3-WT) and an ORF64R-deleted recombinant (FV3-∆64R). In samples from the infected intestine, liver, spleen, lung, and especially kidney, an FV3-targeted transcriptomic analysis mapped reads spanning the full-genome coverage at ~10× depth on both positive and negative strands. By contrast, reads were only mapped to partial genomic regions in samples from the infected thymus, skin, and muscle. Extensive analyses validated the expression of almost all of the 98 annotated ORFs and profiled their differential expression in a tissue-, virus-, and temporal class-dependent manner. Further studies identified several putative ORFs that encode hypothetical proteins containing viral mimicking conserved domains found in host interferon (IFN) regulatory factors (IRFs) and IFN receptors. This study provides the first comprehensive genome-wide viral transcriptome profiling during infection and across multiple amphibian host tissues that will serve as an instrumental reference. Our findings imply that Ranaviruses like FV3 have acquired previously unknown molecular mimics, interfering with host IFN signaling during evolution.
Abstract Aureococcus anophagefferens forms a model host-virus system with the “giant virus” Kratosvirus quantuckense. Studies to define its ribocell (uninfected) and virocell (virus-infected) forms are needed as these states co-occur during algal blooms. Previously, a link between light-derived energy, virus particle production, and virocell formation was noted. We explored how the time of day (morning, midday, or late day) of virus-host contact shaped virocell ontogeny. In parallel, we explored the dependence on light-derived energy in this mixotrophic plankter by inhibiting photosystem II, testing the role of heterotrophic energy in infection dynamics. Using flow cytometry and photochemical assessments, we examined the physiology of infected cells and controls, and estimated virus particle production. We observed differences between ribocell and virocell response to treatments, including reductions in virus particle production during reduced light duration) and PSII inhibition (i.e. “forced heterotrophy”). This work demonstrates the importance of light in shaping the fate of infected cells and provides insight into factors that constrain in situ blooms. Most significantly, we show that time of the solar day when a virus and host come into contact influences viral particle production, and therefore bloom dynamics; a factor that needs to be considered in bloom modeling work.
Gann, Eric R., Kang, Yoonja, Dyhrman, Sonya T., Gobler, Christopher J., and Wilhelm, Steven W. Metatranscriptome Library Preparation Influences Analyses of Viral Community Activity During a Brown Tide Bloom. Retrieved from https://par.nsf.gov/biblio/10293873. Frontiers in Microbiology 12. Web. doi:10.3389/fmicb.2021.664189.
Gann, Eric R., Kang, Yoonja, Dyhrman, Sonya T., Gobler, Christopher J., & Wilhelm, Steven W. Metatranscriptome Library Preparation Influences Analyses of Viral Community Activity During a Brown Tide Bloom. Frontiers in Microbiology, 12 (). Retrieved from https://par.nsf.gov/biblio/10293873. https://doi.org/10.3389/fmicb.2021.664189
Gann, Eric R., Kang, Yoonja, Dyhrman, Sonya T., Gobler, Christopher J., and Wilhelm, Steven W.
"Metatranscriptome Library Preparation Influences Analyses of Viral Community Activity During a Brown Tide Bloom". Frontiers in Microbiology 12 (). Country unknown/Code not available. https://doi.org/10.3389/fmicb.2021.664189.https://par.nsf.gov/biblio/10293873.
@article{osti_10293873,
place = {Country unknown/Code not available},
title = {Metatranscriptome Library Preparation Influences Analyses of Viral Community Activity During a Brown Tide Bloom},
url = {https://par.nsf.gov/biblio/10293873},
DOI = {10.3389/fmicb.2021.664189},
abstractNote = {There is growing interest in the use of metatranscriptomics to study virus community dynamics. We used RNA samples collected from harmful brown tides caused by the eukaryotic alga Aureococcus anophagefferens within New York (United States) estuaries and in the process observed how preprocessing of libraries by either selection for polyadenylation or reduction in ribosomal RNA (rRNA) influenced virus community analyses. As expected, more reads mapped to the A. anophagefferens genome in polyadenylation-selected libraries compared to the rRNA-reduced libraries, with reads mapped in each sample correlating to one another regardless of preprocessing of libraries. Yet, this trend was not seen for reads mapping to the Aureococcus anophagefferens Virus (AaV), where significantly more reads (approximately two orders of magnitude) were mapped to the AaV genome in the rRNA-reduced libraries. In the rRNA-reduced libraries, there was a strong and significant correlation between reads mappings to AaV and A. anophagefferens . Overall, polyadenylation-selected libraries produced fewer viral contigs, fewer reads mapped to viral contigs, and different proportions across viral realms and families, compared to their rRNA-reduced pairs. This study provides evidence that libraries generated by rRNA reduction and not selected for polyadenylation are more appropriate for quantitative characterization of viral communities in aquatic ecosystems by metatranscriptomics.},
journal = {Frontiers in Microbiology},
volume = {12},
author = {Gann, Eric R. and Kang, Yoonja and Dyhrman, Sonya T. and Gobler, Christopher J. and Wilhelm, Steven W.},
editor = {null}
}
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