Mammals Preferred: Reassortment of Batai and Bunyamwera orthobunyavirus Occurs in Mammalian but Not Insect Cells.
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ABSTRACT:
SUBMITTER: Heitmann A
PROVIDER: S-EPMC8473249 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Bunyamwera (BUNV), Batai (BATV) and Ngari (NRIV) are mosquito-borne viruses that are members of the genus Orthobunyavirus in the order Bunyavirales. These three viruses are enveloped with single-stranded, negative-sense RNA genomes consiting of three segments, denoted as Small (S), Medium (M) and Large (L). Ngari is thought to be the natural reassortant progeny of Bunyamwera and Batai viruses. The relationship between these 'parental' viruses and the 'progeny' poses an interesting question, especially given that there is overlap in their respective transmission ecologies, but differences in their infection host ranges and pathogenesis. We compared the in vivo kinetics of these three viruses in a common laboratory system and found no significant difference in growth kinetics. There was, however, a tendency of BATV to have smaller plaques than either BUNV or NRIV. Furthermore, we determined that all three viruses are stable in extracellular conditions and retain infectivity for a week in non-cellular media, which has public health and biosafety implications. The study of this understudied group of viruses addresses a need for basic characterization of viruses that have not yet reached epidemic transmission intensity, but that have the potential due to their infectivity to both human and animal hosts. These results lay the groundwork for future studies of these neglected viruses of potential public and One Health importance.
| S-EPMC8116939 | biostudies-literature
Project description:The bunyavirus nucleocapsid protein, N, is a multifunctional protein that encapsidates each of the three negative-sense genome segments to form ribonucleoprotein complexes that are the functional templates for viral transcription and replication. In addition, N protein molecules interact with themselves to form oligomers, with the viral L (RNA polymerase) protein, with the carboxy-terminal regions of either or both of the virion glycoproteins, and probably also with host cell proteins. Bunyamwera virus (BUNV), the prototype bunyavirus, encodes an N protein of 233 amino acids in length. To learn more about the roles of individual amino acids in the different interactions of N, we performed a wide-scale mutagenic analysis of the protein, and 110 single-point mutants were obtained. When the mutants were employed in a minireplicon assay to examine their effects on viral RNA synthesis, a wide range of activities compared to those of wild-type N protein were observed; changes at nine amino acid positions resulted in severely impaired RNA synthesis. Seventy-seven mutant clones were selected for use in the bunyavirus reverse genetics system, and 57 viable recombinant viruses were recovered. The recombinant viruses displayed a range of plaque sizes and titers in cell culture (from approximately 10(3) to 10(8) PFU/ml), and a number of viruses were shown to be temperature sensitive. Different assays were applied to determine why 20 mutant N proteins could not be recovered into infectious virus. Based on these results, a preliminary domain map of the BUNV N protein is proposed.
| S-EPMC2772805 | biostudies-literature
Project description:BackgroundA number of zoonotic mosquito-borne viruses have emerged in Europe in recent decades. Batai virus (BATV), a member of the genus Orthobunyavirus, is one example of a relatively newly emerged mosquito-borne virus, having been detected in mosquitoes and livestock. We conducted vector competency studies on three mosquito species at a low temperature to assess whether Aedes and Culex mosquito species are susceptible to infection with BATV.MethodsColonised lines of Aedes aegypti and Culex pipiens and a wild-caught species, Aedes detritus, were orally inoculated with BATV strain 53.2, originally isolated from mosquitoes trapped in Germany in 2009. Groups of blood-fed female mosquitoes were maintained at 20 °C for 7 or 14 days. Individual mosquitoes were screened for the presence of BATV in body, leg and saliva samples for evidence of infection, dissemination and transmission, respectively. BATV RNA was detected by reverse transcription-PCR, and positive results confirmed by virus isolation in Vero cells.ResultsAedes detritus was highly susceptible to BATV, with an infection prevalence of ≥ 80% at both measurement time points. Disseminated infections were recorded in 30.7-41.6% of Ae. detritus, and evidence of virus transmission with BATV in saliva samples (n = 1, days post-infection: 14) was observed. Relatively lower rates of infection for Ae. aegypti and Cx. pipiens were observed, with no evidence of virus dissemination or transmission at either time point.ConclusionsThis study shows that Ae. detritus may be a competent vector for BATV at 20 °C, whereas Ae. aegypti and Cx. pipiens were not competent. Critically, the extrinsic incubation period appears to be ≤ 7 days for Ae. detritus, which may increase the onward transmissibility potential of BATV in these populations.
| S-EPMC8567561 | biostudies-literature
Project description:Bunyamwera (BUNV), Batai (BATV), and Ngari (NRIV) are mosquito-borne viruses of the Bunyamwera serogroup in the Orthobunyavirus genus of the Bunyaviridae family. These three viruses have been found to cause disease in both livestock animals, avian species, and humans. Thus, these viruses pose a potential threat to human public health, animal health, and food security. This is especially the case in the developing nations, where BUNV and NRIV are found, mainly in Africa. BUNV and BATV are fairly well characterized, while NRIV is not well characterized owing to only sporadic detection in human and animal populations in Africa. Reassortment is common among bunyaviruses, but NRIV is believed to be the only natural reassortant of the Bunyamwera serogroup. It resulted from a combination of BUNV S and L segments and the BATV M segment. This indicates at least some level co-circulation of BUNV and BATV, which have no historically been reported to overlap in geographic distributions. But as these viruses are undercharacterized, there remains a gap in the understanding of how such reassortment could occur, and the consequences of such. Due to their combined wide range of hosts and vectors, geographic distributions, potential severity of associated diseases, and potential for transmissibility between vertebrate hosts, these viruses represent a significant gap in knowledge with important One Health implications. The goal of this review is to report available knowledge of and identify potential future directions for study of these viruses. As these are collectively understudied viruses, there is a relative paucity of data; however, we use available studies to discuss different perspectives in an effort to promote a better understanding of these three viruses and the public and One Health threat(s) they may pose.
| S-EPMC5906542 | biostudies-literature
Project description:The virion glycoproteins Gn and Gc of Bunyamwera virus (BUNV), the prototype of the Bunyaviridae family and also of the Orthobunyavirus genus, are encoded by the medium (M) RNA genome segment and are involved in both viral attachment and entry. After their synthesis Gn and Gc form a heterodimer in the endoplasmic reticulum (ER) and transit to the Golgi compartment for virus assembly. The N-terminal half of the Gc ectodomain was previously shown to be dispensable for virus replication in cell culture (X. Shi, J. Goli, G. Clark, K. Brauburger, and R. M. Elliott, J. Gen. Virol. 90:2483-2492, 2009.). In this study, the coding sequence for a fluorescent protein, either enhanced green fluorescent protein (eGFP) or mCherry fluorescent protein, was fused to the N terminus of truncated Gc, and two recombinant BUNVs (rBUNGc-eGFP and rBUNGc-mCherry) were rescued by reverse genetics. The recombinant viruses showed bright autofluorescence under UV light and were competent for replication in various mammalian cell lines. rBUNGc-mCherry was completely stable over 10 passages, whereas internal, in-frame deletions occurred in the chimeric Gc-eGFP protein of rBUNGc-eGFP, resulting in loss of fluorescence between passages 5 and 7. Autofluorescence of the recombinant viruses allowed visualization of different stages of the infection cycle, including virus attachment to the cell surface, budding of virus particles in Golgi membranes, and virus-induced morphological changes to the Golgi compartment at later stages of infection. The fluorescent protein-tagged viruses will be valuable reagents for live-cell imaging studies to investigate virus entry, budding, and morphogenesis in real time.
| S-EPMC2919021 | biostudies-literature
Project description:The M genome segment of Bunyamwera virus (BUNV)-the prototype of both the Bunyaviridae family and the Orthobunyavirus genus-encodes the glycoprotein precursor (GPC) that is proteolytically cleaved to yield two viral structural glycoproteins, Gn and Gc, and a nonstructural protein, NSm. The cleavage mechanism of orthobunyavirus GPCs and the host proteases involved have not been clarified. In this study, we investigated the processing of BUNV GPC and found that both NSm and Gc proteins were cleaved at their own internal signal peptides (SPs), in which NSm domain I functions as SP(NSm) and NSm domain V as SP(Gc) Moreover, the domain I was further processed by a host intramembrane-cleaving protease, signal peptide peptidase, and is required for cell fusion activities. Meanwhile, the NSm domain V (SP(Gc)) remains integral to NSm, rendering the NSm topology as a two-membrane-spanning integral membrane protein. We defined the cleavage sites and boundaries between the processed proteins as follows: Gn, from residue 17-312 or nearby residues; NSm, 332-477; and Gc, 478-1433. Our data clarified the mechanism of the precursor cleavage process, which is important for our understanding of viral glycoprotein biogenesis in the genus Orthobunyavirus and thus presents a useful target for intervention strategies.
| S-EPMC4978261 | biostudies-literature
Project description:Bunyamwera virus (BUNV) is the prototype of the genus Orthobunyavirus and the family Bunyaviridae. BUNV has a tripartite genome of negative-sense RNA composed of small (S), medium (M), and large (L) segments. Partially complementary untranslated regions (UTRs) flank the coding region of each segment. The terminal 11 nucleotides of these UTRs are conserved between the three segments and throughout the genus, while the internal regions are unique to each segment and largely nonconserved between different viruses. To investigate the functions of the UTR sequences, we constructed a series of BUNV S segment cDNA clones with deletions in the 3' and/or 5' UTR and then attempted to rescue these segments into recombinant viruses. We found that the genomic 5' UTR was much more sensitive to mutation than the 3' UTR and, in general, sequences proximal to the termini were more important than those flanking the coding region. Northern blot analyses of infected-cell RNA showed that the internal, nonconserved sequences of the S segment 3' UTR play a role in the regulation of transcription and replication and the balance between these two processes. In contrast, deletions in the 5' UTR caused attenuation of the recombinant virus but did not specifically affect levels of S segment RNAs or the encoded nucleocapsid protein. Thus, the internal regions of both UTRs are functional: most of the 5' UTR is essential to viral growth, and, while nonessential, the internal 3' UTR is important to the regulation of viral RNA synthesis.
| S-EPMC1235861 | biostudies-literature
Project description:The first line of defense against viral infection is the interferon (IFN) response, which culminates in the expression of hundreds of proteins with presumed antiviral activity, and must be overcome by a virus for successful replication. The nonstructural NSs protein is the primary IFN antagonist encoded by Bunyamwera virus (BUNV), the prototype of the Orthobunyavirus genus and the family Bunyaviridae. The NSs protein interferes with RNA polymerase II-mediated transcription, thereby inhibiting cellular mRNA production, including IFN mRNAs. A recombinant virus, rBUNdelNSs, that is unable to express the NSs protein does not inhibit cellular transcription and is a strong IFN inducer. We report here that cells stimulated into the antiviral state by IFN-? treatment were protected against wild-type BUNV and rBUNdelNSs infection but addition of IFN-? after infection had little effect on the replication cycle of either virus. By screening a panel of cell lines that overexpressed individual IFN-stimulated genes, we found that protein kinase R (PKR), MTAP44, and particularly viperin appreciably restricted BUNV replication. The enzymatic activities of PKR and viperin were required for their inhibitory activities. Taken together, our data show that the restriction of BUNV replication mediated by IFN is an accumulated effect of at least three IFN-stimulated genes that probably act on different stages of the viral replication cycle.
| S-EPMC3486307 | biostudies-literature
Project description:Bunyamwera orthobunyavirus and its serogroup can cause several diseases in humans, cattle, ruminants, and birds. The viral M-polyprotein helps the virus to enter the host body. Therefore, this protein might serve as a potential vaccine target against Bunyamwera orthobunyavirus. The present study applied the immunoinformatics technique to design an epitopic vaccine component that could protect against Bunyamwera infection. Phylogenetic analysis revealed the presence of conserved patterns of M-polyprotein within the viral serogroup. Three epitopes common for both B-cell and T-cell were identified, i.e., YQPTELTRS, YKAHDKEET, and ILGTGTPKF merged with a specific linker peptide to construct an active vaccine component. The low atomic contact energy value of docking complex between human TLR4 (TLR4/MD2 complex) and vaccine construct confirms the elevated protein-protein binding interaction. Molecular dynamic simulation and normal mode analysis illustrate the docking complex's stability, especially by the higher Eigenvalue. In silico cloning of the vaccine construct was applied to amplify the desired vaccine component. Structural allocation of both the vaccine and epitopes also show the efficacy of the developed vaccine. Hence, the computational research design outcomes support that the peptide-based vaccine construction is a crucial drive target to limit the infection of Bunyamwera orthobunyavirus to an extent.Supplementary informationThe online version contains supplementary material available at 10.1007/s10989-021-10322-9.
| S-EPMC8634745 | biostudies-literature
Project description:The NSs protein of Bunyamwera virus (Bunyaviridae) is an antiapoptotic interferon antagonist involved in silencing host protein expression by interfering with mRNA synthesis. Here, we show that the ability to inhibit both host transcription and the interferon response is linked to interaction of NSs with the MED8 component of Mediator, a protein complex necessary for mRNA production. The interacting domain on NSs was mapped to the C-terminal region, which contains amino acids conserved among orthobunyavirus NSs proteins. A recombinant virus in which the interacting domain in NSs was deleted had strongly reduced ability to inhibit host protein expression and was unable to inhibit the interferon response. This study provides further information on the mechanisms by which bunyavirus nonstructural proteins are involved in pathogenesis.
| S-EPMC1617248 | biostudies-literature