scholarly journals Virus-Targeted Transcriptomic Analyses Implicate Ranaviral Interaction with Host Interferon Response in Frog Virus 3-Infected Frog Tissues

Viruses ◽  
2021 ◽  
Vol 13 (7) ◽  
pp. 1325
Author(s):  
Yun Tian ◽  
Francisco De Jesús Andino ◽  
Collins N. Khwatenge ◽  
Jiuyi Li ◽  
Jacques Robert ◽  
...  
Keyword(s):  
Transcriptome Profiling ◽  
Amphibian Declines ◽  
Transcriptomic Analysis ◽  
Open Reading Frames ◽  
Interferon Response ◽  
Dependent Manner ◽  
Frog Virus 3 ◽  
Frog Virus ◽  
Almost All ◽  
Genomic Regions

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.

scholarly journals Virus-Targeted Transcriptomic Analyses Implicate Ranaviral Interaction with Host Interferon Response in Frog Virus 3-infected Frog Tissues

2021 ◽  
Author(s):  
Yun Tian ◽  
Francisco De Jesús Andino ◽  
Jacques Robert ◽  
Yongming Sang
Keyword(s):  
Transcriptome Profiling ◽  
Amphibian Declines ◽  
Transcriptomic Analysis ◽  
Open Reading Frames ◽  
Interferon Response ◽  
Frog Virus 3 ◽  
Frog Virus ◽  
Genome Wide ◽  
Almost All ◽  
Genomic Regions

Frog Virus 3 (FV3) is a large dsDNA virus that cause global infections in amphibians, fish and reptiles, and contribute to amphibian declines. FV3's genome contains near 100 putative open reading frames (ORFs). 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 genome-wide characterize ranaviral gene expression, 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, a 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 annotated 98 ORFs and profiled their differential expression in a tissue-, virus-, and temporal class-dependent manners. 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 instrumental reference. It also presents evidence implying that ranaviruses like FV3 have acquired previously unknown molecular mimics interfering with host IFN signaling during evolution.

scholarly journals Targeted Transcriptomics of Frog Virus 3 in Infected Frog Tissues Reveal Non-Coding Regulatory Elements and microRNAs in the Ranaviral Genome and Their Potential Interaction with Host Immune Response

2021 ◽  
Vol 12 ◽  
Author(s):  
Yun Tian ◽  
Collins N. Khwatenge ◽  
Jiuyi Li ◽  
Francisco De Jesus Andino ◽  
Jacques Robert ◽  
...  
Keyword(s):  
Regulatory Elements ◽  
Potential Interaction ◽  
Amphibian Declines ◽  
Gene Expressions ◽  
Frog Virus 3 ◽  
Frog Virus ◽  
Eukaryotic Genes ◽  
A Genome ◽  
Cold Blood ◽  
Intergenic Regions

BackgroundFrog Virus 3 (FV3) is a large dsDNA virus belonging to Ranaviruses of family Iridoviridae. Ranaviruses infect cold-blood vertebrates including amphibians, fish and reptiles, and contribute to catastrophic amphibian declines. FV3 has a genome at ~105 kb that contains nearly 100 coding genes and 50 intergenic regions as annotated in its reference genome. Previous studies have mainly focused on coding genes and rarely addressed potential non-coding regulatory role of intergenic regions.ResultsUsing a whole transcriptomic analysis of total RNA samples containing both the viral and cellular transcripts from FV3-infected frog tissues, we detected virus-specific reads mapping in non-coding intergenic regions, in addition to reads from coding genes. Further analyses identified multiple cis-regulatory elements (CREs) in intergenic regions neighboring highly transcribed coding genes. These CREs include not only a virus TATA-Box present in FV3 core promoters as in eukaryotic genes, but also viral mimics of CREs interacting with several transcription factors including CEBPs, CREBs, IRFs, NF-κB, and STATs, which are critical for regulation of cellular immunity and cytokine responses. Our study suggests that intergenic regions immediately upstream of highly expressed FV3 genes have evolved to bind IRFs, NF-κB, and STATs more efficiently. Moreover, we found an enrichment of putative microRNA (miRNA) sequences in more than five intergenic regions of the FV3 genome. Our sequence analysis indicates that a fraction of these viral miRNAs is targeting the 3’-UTR regions of Xenopus genes involved in interferon (IFN)-dependent responses, including particularly those encoding IFN receptor subunits and IFN-regulatory factors (IRFs).ConclusionsUsing the FV3 model, this study provides a first genome-wide analysis of non-coding regulatory mechanisms adopted by ranaviruses to epigenetically regulate both viral and host gene expressions, which have co-evolved to interact especially with the host IFN response.

scholarly journals Amphibian (Xenopus laevis) Tadpoles and Adult Frogs Differ in Their Antiviral Responses to Intestinal Frog Virus 3 Infections

2021 ◽  
Vol 12 ◽  
Author(s):  
Kelsey A. Hauser ◽  
Julia C. Singer ◽  
Muhammad Riadul H. Hossainey ◽  
Tyler E. Moore ◽  
Emily S. Wendel ◽  
...  
Keyword(s):  
Xenopus Laevis ◽  
Myeloid Cells ◽  
Amphibian Declines ◽  
Type I ◽  
Frog Virus 3 ◽  
Frog Virus ◽  
Antiviral Responses ◽  
Major Route ◽  
Susceptibility And Resistance ◽  
Insight Into

The global amphibian declines are compounded by ranavirus infections such as Frog Virus 3 (FV3), and amphibian tadpoles more frequently succumb to these pathogens than adult animals. Amphibian gastrointestinal tracts represent a major route of ranavirus entry, and viral pathogenesis often leads to hemorrhaging and necrosis within this tissue. Alas, the differences between tadpole and adult amphibian immune responses to intestinal ranavirus infections remain poorly defined. As interferon (IFN) cytokine responses represent a cornerstone of vertebrate antiviral immunity, it is pertinent that the tadpoles and adults of the anuran Xenopus laevis frog mount disparate IFN responses to FV3 infections. Presently, we compared the tadpole and adult X. laevis responses to intestinal FV3 infections. Our results indicate that FV3-challenged tadpoles mount more robust intestinal type I and III IFN responses than adult frogs. These tadpole antiviral responses appear to be mediated by myeloid cells, which are recruited into tadpole intestines in response to FV3 infections. Conversely, myeloid cells bearing similar cytology already reside within the intestines of healthy (uninfected) adult frogs, possibly accounting for some of the anti-FV3 resistance of these animals. Further insight into the differences between tadpole and adult frog responses to ranaviral infections is critical to understanding the facets of susceptibility and resistance to these pathogens.

scholarly journals Frog Virus 3 Genomes Reveal Prevalent Recombination between Ranavirus Lineages and Their Origins in Canada

2019 ◽  
Vol 93 (20) ◽  
Author(s):  
Sibelle T. Vilaça ◽  
Joe-Felix Bienentreu ◽  
Craig R. Brunetti ◽  
David Lesbarrères ◽  
Dennis L. Murray ◽  
...  
Keyword(s):  
North America ◽  
Open Reading Frames ◽  
Anthropogenic Factors ◽  
Amphibian Species ◽  
Frog Virus 3 ◽  
Frog Virus ◽  
Wild Strains ◽  
Recent Origin ◽  
Midwife Toad ◽  
Recombination Breakpoints

ABSTRACT Ranaviruses are pathogens associated with the decline of amphibian populations across much of their distribution. In North America, frog virus 3 (FV3) is a widely distributed pathogen with wild populations of amphibians harboring different lineages and putative recombinants between FV3 and common midwife toad virus (CMTV). These recombinants have higher pathogenicity, and CMTV-derived genes associated with virulence are reported in wild strains in Canada. However, while FV3 is linked to amphibian die-offs in North America, CMTVs have been reported only in commercial frog farms in North America. We sequenced complete genomes of 18 FV3 isolates from three amphibian species to characterize genetic diversity of the lineages in Canada and infer possible recombinant regions. The 18 FV3 isolates displayed different signals of recombination, varying from none to interspersed recombination with previously isolated CMTV-like viruses. In general, most recombination breakpoints were located within open reading frames (ORFs), generating new ORFs and proteins that were a mixture between FV3 and CMTV. A combined spatial and temporal phylogeny suggests the presence of the FV3 lineage in Canada is relatively contemporary (<100 years), corroborating the hypothesis that both CMTV- and FV3-like viruses spread to North America when the international commercial amphibian trade started. Our results highlight the importance of pathogen surveillance and viral dynamics using full genomes to more clearly understand the mechanisms of disease origin and spread. IMPORTANCE Amphibian populations are declining worldwide, and these declines have been linked to a number of anthropogenic factors, including disease. Among the pathogens associated with amphibian mortality, ranaviruses have caused massive die-offs across continents. In North America, frog virus 3 (FV3) is a widespread ranavirus that can infect wild and captive amphibians. In this study, we sequenced full FV3 genomes isolated from frogs in Canada. We report widespread recombination between FV3 and common midwife toad virus (CMTV). Phylogenies indicate a recent origin for FV3 in Canada, possibly as a result of international amphibian trade.

scholarly journals Targeted Transcriptomics of Frog Virus 3 in Infected Frog Tissues Reveal Non-Coding Regulatory Elements and microRNAs in the Ranaviral Genome and Their Potential Interaction With Host Immune Response

2021 ◽  
Author(s):  
Yun (Simon) Tian ◽  
Collins Khwatenge ◽  
Jiuyi Li ◽  
Francisco De Jesus Andino ◽  
Jacques Robert ◽  
...  
Keyword(s):  
Regulatory Elements ◽  
Potential Interaction ◽  
Amphibian Declines ◽  
Gene Expressions ◽  
Frog Virus 3 ◽  
Frog Virus ◽  
Eukaryotic Genes ◽  
A Genome ◽  
Cold Blood ◽  
Intergenic Regions

Abstract Background: Frog Virus 3 (FV3) is a large dsDNA virus belonging to Ranaviruses of family Iridoviridae . Ranaviruses infect cold-blood vertebrates including amphibians, fish and reptiles, and contribute to catastrophic amphibian declines. FV3 has a genome at ~105 kb that contains nearly 100 coding genes and 50 intergenic regions as annotated in its reference genome. Previous studies have mainly focused on coding genes and rarely addressed potential non-coding regulatory role of intergenic regions. Results: Using a whole transcriptomic analysis of total RNA samples containing both the viral and cellular transcripts from FV3-infected frog tissues, we detected virus-specific reads mapping in non-coding intergenic regions, in addition to reads from coding genes. Further analyses identified multiple cis -regulatory elements ( CREs ) in intergenic regions neighboring highly transcribed coding genes. These CREs include not only a virus TATA-Box present in FV3 core promoters as in eukaryotic genes, but also viral mimics of CREs interacting with several transcription factors including CEBPs, CREBs, IRFs, NF-κB, and STATs, which are critical for regulation of cellular immunity and cytokine responses. Our study suggests that intergenic regions immediately upstream of highly expressed FV3 genes have evolved to bind IRFs, NF-κB, and STATs more efficiently. Moreover, we found an enrichment of putative microRNA (miRNA) sequences in more than five intergenic regions of the FV3 genome. Our sequence analysis indicates that a fraction of these viral miRNAs is targeting the 3’-UTR regions of Xenopus genes involved in interferon (IFN)-dependent responses, including particularly those encoding IFN receptor subunits and IFN-regulatory factors (IRFs). Conclusions: Using the FV3 model, this study provides a first genome-wide analysis of non-coding regulatory mechanisms adopted by ranaviruses to epigenetically regulate both viral and host gene expressions, which have co-evolved to interact especially in the host IFN response.

scholarly journals Discovery of granulocyte-lineage cells in the skin of the amphibian Xenopus laevis

FACETS ◽  
2020 ◽  
Vol 5 (1) ◽  
pp. 571-597
Author(s):  
Kelsey Hauser ◽  
Milan Popovic ◽  
Amulya Yaparla ◽  
Daphne V. Koubourli ◽  
Phillip Reeves ◽  
...  
Keyword(s):  
Xenopus Laevis ◽  
Batrachochytrium Dendrobatidis ◽  
Amphibian Declines ◽  
Immune Gene ◽  
Dependent Manner ◽  
Amphibian Skin ◽  
Positive Skin ◽  
Frog Virus ◽  
Polymorphonuclear Granulocyte ◽  
Surface Staining

The ranavirus Frog Virus 3 (FV3) and the chytrid fungus Batrachochytrium dendrobatidis ( Bd) are significant contributors to the global amphibian declines and both pathogens target the amphibian skin. We previously showed that tadpoles and adults of the anuran amphibian Xenopus laevis express notable levels of granulocyte chemokine genes ( cxcl8a and cxcl8b) within their skin and likely possess skin-resident granulocytes. Presently, we show that tadpole and adult X. laevis indeed possess granulocyte-lineage cells within their epidermises that are distinct from their skin mast cells, which are found predominantly in lower dermal layers. These esterase-positive cells responded to (r)CXCL8a and rCXCL8b in a concentration- and CXCR1/CXCR2-dependent manner, possessed polymorphonuclear granulocyte morphology, granulocyte marker surface staining, and exhibited distinct immune gene expression from conventional granulocytes. Our past work indicates that CXCL8b recruits immunosuppressive granulocytes, and here we demonstrated that enriching esterase-positive skin granulocytes with rCXCL8b (but not rCXCL8a) may increase tadpole susceptibility to FV3 and adult frog susceptibility to Bd. Furthermore, pharmacological depletion of skin-resident granulocytes increased tadpole susceptibility to FV3. This manuscript provides new insights into the composition and roles of immune cells within the amphibian skin, which is a critical barrier against pathogenic contributors to the amphibian declines.

scholarly journals Localization of Frog Virus 3 Conserved Viral Proteins 88R, 91R, and 94L

Viruses ◽  
2019 ◽  
Vol 11 (3) ◽  
pp. 276 ◽  
Author(s):  
Emily Penny ◽  
Craig Brunetti
Keyword(s):  
Protein Modification ◽  
Intracellular Localization ◽  
Genomic Analysis ◽  
Open Reading Frames ◽  
Comparative Genomic ◽  
Frog Virus 3 ◽  
Frog Virus ◽  
Viral Genes ◽  
Conserved Genes

The characterization of the function of conserved viral genes is central to developing a greater understanding of important aspects of viral replication or pathogenesis. A comparative genomic analysis of the iridoviral genomes identified 26 core genes conserved across the family Iridoviridae. Three of those conserved genes have no defined function; these include the homologs of frog virus 3 (FV3) open reading frames (ORFs) 88R, 91R, and 94L. Conserved viral genes that have been previously identified are known to participate in a number of viral activities including: transcriptional regulation, DNA replication/repair/modification/processing, protein modification, and viral structural proteins. To begin to characterize the conserved FV3 ORFs 88R, 91R, and 94L, we cloned the genes and determined their intracellular localization. We demonstrated that 88R localizes to the cytoplasm of the cell while 91R localizes to the nucleus and 94L localizes to the endoplasmic reticulum (ER).

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