scholarly journals Simulated vector transmission differentially influences dynamics of two viral variants of deformed wing virus in honey bees (Apis mellifera)

2021 ◽  
Vol 102 (11) ◽  
Author(s):  
Allyson M. Ray ◽  
Sheldon L. Davis ◽  
Jason L. Rasgon ◽  
Christina M. Grozinger
Keyword(s):  
Honey Bees ◽  
Disease Ecology ◽  
Nucleotide Diversity ◽  
Sequencing Analysis ◽  
Vector Transmission ◽  
Deformed Wing Virus ◽  
Functional Regions ◽  
Host Mortality ◽  
Amino Acid Mutations

Understanding how vectors alter the interactions between viruses and their hosts is a fundamental question in virology and disease ecology. In honey bees, transmission of deformed wing virus (DWV) by parasitic Varroa mites has been associated with elevated disease and host mortality, and Varroa transmission has been hypothesized to lead to increased viral titres or select for more virulent variants. Here, we mimicked Varroa transmission by serially passaging a mixed population of two DWV variants, A and B, by injection through in vitro reared honey bee pupae and tracking these viral populations through five passages. The DWV-A and DWV-B variant proportions shifted dynamically through passaging, with DWV-B outcompeting DWV-A after one passage, but levels of both variants becoming equivalent by Passage 5. Sequencing analysis revealed a dominant, recombinant DWV-B strain (DWV-A derived 5′ IRES region with the rest of the genome DWV-B), with low nucleotide diversity that decreased through passaging. DWV-A populations had higher nucleotide diversity compared to DWV-B, but this also decreased through passaging. Selection signatures were found across functional regions of the DWV-A and DWV-B genomes, including amino acid mutations in the putative capsid protein region. Simulated vector transmission differentially impacted two closely related viral variants which could influence viral interactions with the host, demonstrating surprising plasticity in vector-host-viral dynamics.

scholarly journals Attenuation of Bluetongue Virus (BTV) in an in ovo Model Is Related to the Changes of Viral Genetic Diversity of Cell-Culture Passaged BTV

Viruses ◽  
2019 ◽  
Vol 11 (5) ◽  
pp. 481 ◽  
Author(s):  
Fabian Z. X. Lean ◽  
Matthew J. Neave ◽  
John R. White ◽  
Jean Payne ◽  
Teresa Eastwood ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
Cell Culture ◽  
Bluetongue Virus ◽  
Genomic Analysis ◽  
Sequencing Analysis ◽  
Structural Protein ◽  
In Ovo ◽  
Novel Approach ◽  
Amino Acid Mutations

The embryonated chicken egg (ECE) is routinely used for the laboratory isolation and adaptation of Bluetongue virus (BTV) in vitro. However, its utility as an alternate animal model has not been fully explored. In this paper, we evaluated the pathogenesis of BTV in ovo using a pathogenic isolate of South African BTV serotype 3 (BTV-3) derived from the blood of an infected sheep. Endothelio- and neurotropism of BTV-3 were observed by immunohistochemistry of non-structural protein 1 (NS1), NS3, NS3/3a, and viral protein 7 (VP7) antigens. In comparing the pathogenicity of BTV from infectious sheep blood with cell-culture-passaged BTV, including virus propagated through a Culicoides-derived cell line (KC) or ECE, we found virus attenuation in ECE following cell-culture passage. Genomic analysis of the consensus sequences of segments (Seg)-2, -5, -6, -7, -8, -9, and -10 identified several nucleotide and amino-acid mutations among the cell-culture-propagated BTV-3. Deep sequencing analysis revealed changes in BTV-3 genetic diversity in various genome segments, notably a reduction of Seg-7 diversity following passage in cell culture. Using this novel approach to investigate BTV pathogenicity in ovo, our findings support the notion that pathogenic BTV becomes attenuated in cell culture and that this change is associated with virus quasispecies evolution.

Deformed wing virus: using reverse genetics to tackle unanswered questions about the most important viral pathogen of honey bees

2020 ◽  
Author(s):  
Luke Woodford ◽  
David J Evans
Keyword(s):  
Honey Bees ◽  
Reverse Genetics ◽  
Reverse Genetic ◽  
Strain Variation ◽  
Colony Losses ◽  
Viral Pathogen ◽  
Deformed Wing Virus ◽  
Host Tropism ◽  
Asymptomatic Infections

Abstract Deformed wing virus (DWV) is the most important viral pathogen of honey bees. It usually causes asymptomatic infections but, when vectored by the ectoparasitic mite Varroa destructor, it is responsible for the majority of overwintering colony losses globally. Although DWV was discovered four decades ago, research has been hampered by the absence of an in vitro cell culture system or the ability to culture pure stocks of the virus. The recent development of reverse genetic systems for DWV go some way to addressing these limitations. They will allow the investigation of specific questions about strain variation, host tropism and pathogenesis to be answered, and are already being exploited to study tissue tropism and replication in Varroa and non-Apis pollinators. Three areas neatly illustrate the advances possible with reverse genetic approaches; 1) strain variation and recombination, in which reverse genetics has highlighted similarities rather than differences between virus strains, 2) analysis of replication kinetics in both honey bees and Varroa, in studies which likely explain the near clonality of virus populations often reported and, 3) pathogen spillover to non-Apis pollinators, using genetically-tagged viruses to accurately monitor replication and infection.

scholarly journals A56 Visualization of recombination in deformed wing virus infecting bees

2019 ◽  
Vol 5 (Supplement_1) ◽  
Author(s):  
Diane Bigot ◽  
Andreas Gogol-Döring ◽  
Peter Koch ◽  
Robert J Paxton
Keyword(s):  
Honey Bees ◽  
Bumble Bees ◽  
Next Generation Sequencing Data ◽  
Sequencing Data ◽  
Wild Bees ◽  
Deformed Wing Virus ◽  
The Impact ◽  
Detection And Quantification

Abstract Honey bees suffer increasing colony mortality worldwide, partially caused by the spread of viral pathogens. Among these pathogens, deformed wing virus (DWV) is one of the major, widespread viruses of honey bees resulting in wing deformities and weakening colonies. DWV can be found in honey bees, bumble bees, and other wild bees as three major genotypes named DWV-A, -B (also named Varroa destructor virus 1), and -C. Various recombinants of DWV-A and -B have been previously found in honey bees, some of which have been suggested to have higher virulence over non-recombinant, parental virus. In most of these cases, recombinants were only shown as consensus sequences from previous assemblies and alignments and may not reflect the biological reality of all variants present within a host bee. It is therefore important to build a method of recombinant detection and quantification within mixed infections in single-host individuals, including both parental and various recombinant genomes, so as to evaluate the relevance of recombinants for viral genome evolution and the impact on hosts. Here, we propose to visualize and quantify these recombinants using next-generation sequencing data to better understand how these genomes evolve within bees. Our method will be performed directly from raw sequence reads from various datasets (including field and lab experiments as well as screening of public databases) in order to obtain an overview of DWV recombination in various in vivo and in vitro conditions. Recombination of viral genomes is a key point for virus evolution. The detection and quantification of recombination will facilitate analysis of the determinants of recombination and help in understanding the routes by which new viral variants emerge. The emergence of new (more virulent) recombinant viruses can result from acquisition of new capabilities, such as escape from host immunity or increased transmission rates. Recombination can also lead to adaptation to new environments and new hosts by a change in cell tropism, allowing cross-species transmission, which may be particularly relevant for bumble bees and wild bees infected by honey bee-derived DWV.

scholarly journals Pupal cannibalism by worker honey bees contributes to the spread of deformed wing virus

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Francisco Posada-Florez ◽  
Zachary S. Lamas ◽  
David J. Hawthorne ◽  
Yanping Chen ◽  
Jay D. Evans ◽  
...  
Keyword(s):  
Honey Bees ◽  
Control Strategies ◽  
Virus Transmission ◽  
Experimental Conditions ◽  
Colony Losses ◽  
Viral Pathogen ◽  
Deformed Wing Virus ◽  
Hygienic Behaviour ◽  
Pathogen Virulence ◽  
Comprehensive Knowledge

AbstractTransmission routes impact pathogen virulence and genetics, therefore comprehensive knowledge of these routes and their contribution to pathogen circulation is essential for understanding host–pathogen interactions and designing control strategies. Deformed wing virus (DWV), a principal viral pathogen of honey bees associated with increased honey bee mortality and colony losses, became highly virulent with the spread of its vector, the ectoparasitic mite Varroa destructor. Reproduction of Varroa mites occurs in capped brood cells and mite-infested pupae from these cells usually have high levels of DWV. The removal of mite-infested pupae by worker bees, Varroa Sensitive Hygiene (VSH), leads to cannibalization of pupae with high DWV loads, thereby offering an alternative route for virus transmission. We used genetically tagged DWV to investigate virus transmission to and between worker bees following pupal cannibalisation under experimental conditions. We demonstrated that cannibalization of DWV-infected pupae resulted in high levels of this virus in worker bees and that the acquired virus was then transmitted between bees via trophallaxis, allowing circulation of Varroa-vectored DWV variants without the mites. Despite the known benefits of hygienic behaviour, it is possible that higher levels of VSH activity may result in increased transmission of DWV via cannibalism and trophallaxis.

scholarly journals Genotypic and Phenotypic Analyses of Hepatitis C Virus from Patients Treated with JTK-853 in a Three-Day Monotherapy

2012 ◽  
Vol 57 (1) ◽  
pp. 436-444 ◽  
Author(s):  
Naoki Ogura ◽  
Yukiyo Toyonaga ◽  
Izuru Ando ◽  
Kunihiro Hirahara ◽  
Tsutomu Shibata ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Amino Acid ◽  
Viral Resistance ◽  
Hcv Rna ◽  
Amino Acid Substitutions ◽  
Sequencing Analysis ◽  
Resistance Mutations ◽  
In The Wild

ABSTRACTJTK-853, a palm site-binding NS5B nonnucleoside polymerase inhibitor, shows antiviral activityin vitroand in hepatitis C virus (HCV)-infected patients. Here, we report the results of genotypic and phenotypic analyses of resistant variants in 24 HCV genotype 1-infected patients who received JTK-853 (800, 1,200, or 1,600 mg twice daily or 1,200 mg three times daily) in a 3-day monotherapy. Viral resistance in NS5B was investigated using HCV RNA isolated from serum specimens from the patients. At the end of treatment (EOT) with JTK-853, the amino acid substitutions M414T (methionine [M] in position 414 at baseline was replaced with threonine [T] at EOT), C445R (cysteine [C] in position 445 at baseline was replaced with arginine [R] at EOT), Y448C/H (tyrosine [Y] in position 448 at baseline was replaced with cysteine [C] or histidine [H] at EOT), and L466F (leucine [L] in position 466 at baseline was replaced with phenylalanine [F] at EOT), which are known to be typical resistant variants of nonnucleoside polymerase inhibitors, were observed in a clonal sequencing analysis. These substitutions were also selected by a treatment with JTK-853in vitro, and the 50% effective concentration of JTK-853 in the M414T-, C445F-, Y448H-, and L466V-harboring replicons attenuated the susceptibility by 44-, 5-, 6-, and 21-fold, respectively, compared with that in the wild-type replicon (Con1). These findings suggest that amino acid substitutions of M414T, C445R, Y448C/H, and L466F are thought to be viral resistance mutations in HCV-infected patients receiving JTK-853 in a 3-day monotherapy.

scholarly journals Deformed wing virus variant shift from 2010 to 2016 in managed and feral UK honey bee colonies

2021 ◽  
Author(s):  
J. L. Kevill ◽  
K. C. Stainton ◽  
D. C. Schroeder ◽  
S. J. Martin
Keyword(s):  
Honey Bee ◽  
Honey Bees ◽  
Small Study ◽  
Deformed Wing Virus ◽  
Virus Variant ◽  
The Usa ◽  
The Uk ◽  
Bee Colonies ◽  
Existing Data

AbstractDeformed wing virus (DWV) has been linked to the global decline of honey bees. DWV exists as three master variants (DWV-A, DWV-B, and DWV-C), each with differing outcomes for the honey bee host. Research in the USA showed a shift from DWV-A to DWV-B between 2010 to 2016 in honey bee colonies. Likewise, in the UK, a small study in 2007 found only DWV-A, whereas in 2016, DWV-B was the most prevalent variant. This suggests a shift from DWV-A to DWV-B might have occurred in the UK between 2007 and 2016. To investigate this further, data from samples collected in 2009/10 (n = 46) were compared to existing data from 2016 (n = 42). These samples also allowed a comparison of DWV variants between Varroa-untreated (feral) and Varroa-treated (managed) colonies. The results revealed that, in the UK, DWV-A was far more prevalent in 2009/10 (87%) than in 2016 (43%). In contrast, DWV-B was less prevalent in 2009/10 (76%) than in 2016 (93%). Regardless if colonies had been treated for Varroa (managed) or not (feral), the same trend from DWV-A to DWV-B occurred. Overall, the results reveal a decrease in DWV-A and an increase in DWV-B in UK colonies.

scholarly journals Hypermethylation of APC2 Is a Predictive Epigenetic Biomarker for Chinese Colorectal Cancer

2018 ◽  
Vol 2018 ◽  
pp. 1-7 ◽  
Author(s):  
Yuan He ◽  
Li-Yue Sun ◽  
Jing Wang ◽  
Rui Gong ◽  
Qiong Shao ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Cox Regression ◽  
The Cancer Genome Atlas ◽  
Sequencing Analysis ◽  
Primary Tumors ◽  
Protein Levels ◽  
Normal Tissues ◽  
Ffpe Tissues ◽  
Epigenetic Biomarker

Objective. To investigate methylation of the adenomatosis polyposis coli homologue (APC2) promoter and its correlation with prognostic implications in Chinese colorectal cancer (CRC). Methods. The mRNA expression of APC2 in colorectal tissues was evaluated using the database of The Cancer Genome Atlas (TCGA). Methylation analysis of APC2 in tumor (n=66) and corresponding adjacent formalin-fixed and paraffin-embedded (FFPE) tissues (n=44) was performed by Sequenom EpiTYPER® and verified by cloning-based bisulfite sequencing analysis. Demethylation and retrieval of APC2 expression in cell lines HT29, HCT116, and SW480 were treated with 5-aza-2′-deoxycytidine (5-AZC). Results. Analysis of TCGA showed that APC2 mRNA was significantly downregulated in primary tumors when compared to normal tissues (p<0.05). APC2 methylation was upregulated (43.93% vs 7.31%, p<0.05) in tumors compared to adjacent FFPE tissues. In vitro experiments demonstrated that 5-AZC downregulated the methylation of APC2 and retrieved its expression of mRNA and protein levels (p<0.05). Multivariate Cox regression indicated that APC2_CPG_14 was an independent risk factor for overall survival (HR = 6.38, 95% CI: 1.59–25.64, p<0.05). Conclusion. This study indicates that APC2 is hypermethylated and may be a tumorigenesis biomarker for Chinese CRC patients.

scholarly journals Multi-host viruses in Argentine ants and honey bees: Increased viral disease in honey bees is associated with Argentine ants

2021 ◽  
Author(s):  
◽  
Jana Dobelmann
Keyword(s):  
Honey Bees ◽  
Rna Viruses ◽  
Population Control ◽  
Argentine Ant ◽  
Viral Dynamics ◽  
Invasive Ants ◽  
Argentine Ants ◽  
Deformed Wing Virus ◽  
Tomato Ringspot Virus ◽  
Bee Health

<p><b>Emerging infectious diseases threaten public health, livestock economies, and wildlife. Human-mediated species introductions can alter host and pathogen communities that shape the dynamics of infectious diseases. Several RNA viruses that have been linked to population declines in wild pollinators and losses of managed honey bees have been detected in multiple other species and are suspected to circulate within insect communities. Yet, we lack an understanding of how disease dynamics are affected by the introduction of novel species. These introduced species include invasive ants, which can disturb honey bees and become a pest in apiaries. The Argentine ant (Linepithema humile) is a globally successful invader that has been observed to attack bees and multiple bee-associated viruses have been detected in this ant species.</b></p> <p>Here, I studied interactions between Argentine ants and European honey bees (Apis mellifera) and how these interactions affect viral dynamics in beehives. I first tested a range of pollinators and associated insects for RNA viruses that are pathogenic to honey bees. Bee-associated viruses showed evidence for active viral replication in several pollinator species but also in species that cohabit in beehives such as ants, spiders, and cockroaches. Using phylogenetic analyses, I found that viral transmission within communities was shaped by geographic origin rather than being restricted by species barriers. Next, I used a longitudinal field study to test whether Argentine ant presence affected pathogen infections and survival in beehives. Argentine ants tested positive for three bee-associated viruses even before beehives were moved into ant-infested sites. Increased levels of deformed wing virus in beehives in autumn were associated with ant presence, although hive mortality was not affected by ants over the duration of this experiment. I used RNA sequencing on a subset of honey bee samples collected during autumn to study the RNA virome and identify transcriptomic responses associated with ant presence. Twelve RNA viruses were found in beehives, among those, three plant-associated viruses and an unclassified RNA virus that had not previously been observed in honey bees. Deformed wing virus showed the highest viral titres in most hives, but was only marginally affected by ant presence. Sacbrood virus and tomato ringspot virus levels were increased in hives with ants, however, both viruses are not known to infect Argentine ants and the plant-associated tomato ringspot virus seems unlikely to affect bee health.</p> <p>Lastly, I tested the feasibility of controlling Argentine ants in apiaries using a novel pest control strategy. RNA interference is a conserved cellular gene regulation mechanism that could be used to silence specific genes in ants. Using double-stranded RNA (dsRNA) to silence two immune-related genes in Argentine ants was expected to increase pathogen susceptibility, which could then lead to higher pathogen levels that reduce ant numbers. My results indicated that no consistent immune silencing could be achieved in the field. Immune gene expression changes were observed, but pathogen titres were not affected, and ant numbers stayed high. Argentine ant control using a conventional insecticide significantly increased bee survival, whereas many hives in the dsRNA and control group abandoned their hives due to ant attacks. Although population control was not successful using the two Argentine ant-specific dsRNAs, insights into ant immunity and ant-bee interactions could improve the development of novel control strategies.</p> <p>Bee-associated viruses have repeatedly been detected in ant species, yet, this is one of the first studies to investigate whether ants affect viral dynamics in honey bees. I showed that invasive Argentine ants are associated with increases in viral pathogens in honey bees. The mechanisms by which ants affect bee disease are unknown, although there is some evidence for ants transmitting viruses or causing stress responses in bees that affect immunity. The findings of this thesis highlight the risk of invasive ant species disrupting pollination services. New and environmentally-friendly methods to control invasive species are urgently needed to improve bee health and limit the spread of invasive ants, such as Argentine ants. The high prevalence of bee-associated viruses and viral diversity in ants suggests that pathogens that are suitable for population control might be present in ant populations, although risks of spillovers into other species need to be carefully considered.</p>

scholarly journals Trasposon Mediated Horizontal Gene Transfer (T-HGT) of Circulating Tumor DNA Reshapes MM Clonal Architecture

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 3065-3065
Author(s):  
Munevver Cinar ◽  
Steven Flygare ◽  
Marina Mosunjac ◽  
Ganji Nagaraju ◽  
Dongkyoo Park ◽  
...  
Keyword(s):  
Gene Transfer ◽  
Horizontal Gene Transfer ◽  
Drug Sensitivity ◽  
Genetic Material ◽  
Host Cells ◽  
Response To Treatment ◽  
Hierarchical Architecture ◽  
Tumor Evolution ◽  
Sequencing Analysis

Spatial genetic heterogeneity is a characteristic phenomenon that influences multiple myeloma's (MM) phenotype and drug sensitivity (Rasche L. et al and Bolli N et al.). Hence, the branch model of tumor evolution is not sufficient to explain the disorganized architecture observed in MM. In this study, we investigated whether MM ctDNA horizontal gene transfer (HGT) affect tumor genetic architecture and drug sensitivity, resembling what is seen in prokaryotes, and elucidated the mechanisms involved in the mobilization of genetic material from one cell to another. We identified that plasma from patients with MM transmits drug sensitivity or resistance to cells in culture. This transmission of drug sensitivity is mediated by ctDNA transfer of oncogenes to a host cell. Importantly, in vitro and in vivo demonstrated that ctDNA mainly targets cells resembling the cell of origin (tropism). Karyotype spreads and whole genome sequencing demonstrated that once patients ctDNA encounters host cells, it migrates into the nucleus where it ultimately integrates into the cell's genome. Integration to the genome was confirmed to be targeted to myeloma cells. Further sequencing analysis of multiple MM samples identified ctDNA tropism and integration is dependent on the 5' and 3' end presence of transposable elements (TE), particularly of the MIR and ALUsq family. These results were further validated by TE mediated delivery of GFP into MM cells in vitro and HSVTK in tumors of mouse xenografts. In conclusion, this data indicates for the first time that TE mediates MM ctDNA HGT into homologous tumor cells shaping the hierarchical architecture of tumor clones and affecting tumor response to treatment. Therapeutically, this unique quality of ctDNA can be exploited for targeted gene therapeutic approaches in MM and potentially other cancers. Disclosures Bernal-Mizrachi: Kodikas Therapeutic Solutions, Inc: Equity Ownership; TAKEDA: Research Funding; Winship Cancer Institute: Employment, Patents & Royalties.

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