A shared transcription termination signal on negative and ambisense RNA genome segments of Rift Valley fever, sandfly fever Sicilian, and Toscana viruses.
(17/89)
The Phlebovirus genus (family Bunyaviridae) is composed of a diverse group of arboviruses that cause disease syndromes ranging from mild febrile illness to hemorrhagic fever with high fatality. Although antigenically similar, these viruses differ by approximately 25% at the genome level, and their ecologies, including geographic ranges, preferred vector species, and hosts, vary considerably. In contrast to other ambisense viruses, where RNA hairpin structures which serve as transcription termination signals are frequently found separating the opposite-sense open reading frames, no evidence of predicted high-energy hairpin structures was found at the ambisense junctions of phlebovirus S RNA segments. However, a conserved sequence motif was identified on both negative and ambisense genome segments that functions as a transcription termination signal for the N, NSs, and GPC mRNAs in three diverse phleboviruses, namely, Rift Valley fever, sandfly Sicilian, and Toscana viruses. The exact termination of nascent virus mRNA molecules was determined by 3' rapid amplification of cDNA ends. Surprisingly, analysis of the termini of mRNAs from both S and M segments of these three viruses revealed that transcription termination occurred immediately upstream of a conserved sequence motif with the general features 3'-C(1-3)GUCG/A-5'. In contrast, no corresponding sequence motif was found in the L segments, and analysis indicated a "runoff" transcript approach to L mRNA termination. The absolute requirement of the identified transcription termination motif was demonstrated by using a highly efficient Rift Valley fever virus reverse genetics system to generate live recombinant viruses with S segments lacking the termination signal motif for the NP or NSs mRNA and showing that these recombinant viruses generated mRNAs that failed to terminate correctly. (+info)
Ex vivo stability of the rodent-borne Hantaan virus in comparison to that of arthropod-borne members of the Bunyaviridae family.
(18/89)
The possible effect of virus adaptation to different transmission routes on virus stability in the environment is not well known. In this study we have compared the stabilities of three viruses within the Bunyaviridae family: the rodent-borne Hantavirus Hantaan virus (HTNV), the sand fly-borne Phlebovirus sandfly fever Sicilian virus (SFSV), and the tick-borne Nairovirus Crimean-Congo hemorrhagic fever virus (CCHFV). These viruses differ in their transmission routes: SFSV and CCHFV are vector borne, whereas HTNV is spread directly between its hosts, and to humans, via the environment. We studied whether these viruses differed regarding stability when kept outside of the host. Viral survival was analyzed at different time points upon exposure to different temperatures (4 degrees C, 20 degrees C, and 37 degrees C) and drying at 20 degrees C. We observed clearly different stabilities under wet conditions, particularly at 4 degrees C, where infectious SFSV, HTNV, and CCHFV were detectable after 528, 96, and 15 days, respectively. All three viruses were equally sensitive to drying, as shown by drying on aluminum discs. Furthermore, HTNV and SFSV partially survived for 2 min in 30% ethanol, whereas CCHFV did not. Electron microscopy images of HTNV, SSFSV, and CCHFV stored at 37 degrees C until infectivity was lost still showed the occurrence of virions, but with abnormal shapes and densities compared to those of the nonincubated samples. In conclusion, our study points out important differences in ex vivo stability among viruses within the Bunyaviridae family. (+info)
A novel, multipartite, negative-strand RNA virus is associated with the ringspot disease of European mountain ash (Sorbus aucuparia L.).
(19/89)
Four RNAs from a new plant-pathogenic virus, which we have tentatively named European mountain ash ringspot-associated virus (EMARAV), were identified and sequenced completely. All four viral RNAs could be detected in previous double-stranded RNA preparations. RNA 1 (7040 nt) encodes a protein with similarity to the RNA-dependent RNA polymerase of different members of the Bunyaviridae, a family containing five genera with viruses infecting invertebrates, vertebrates and plants. RNA 2 (2335 nt) encodes a 75 kDa protein containing a conserved motif of the glycoprotein precursor of the genus Phlebovirus. Immunological detection indicated the presence of proteins with the expected size of the precursor and one of its processing products. The amino acid sequence of protein p3 (35 kDa) encoded by RNA 3 shows similarities to a putative nucleocapsid protein of two still unclassified plant viruses. The fourth viral RNA encodes a 27 kDa protein that has no significant homology to any known protein. As is typical for members of the family Bunyaviridae, the 5' and 3' ends of all viral RNAs are complementary, which allows the RNA to form a panhandle structure. Comparison of these sequences demonstrates a conserved terminal part of 13 nt, similar to that of the bunyaviral genus Orthobunyavirus. Despite the high agreement of the EMARAV genome with several characteristics of the family Bunyaviridae, there are a few features that make it difficult to allocate the virus to this group. It is therefore more likely that this plant pathogen belongs to a novel virus genus. (+info)
TLR3 is essential for the induction of protective immunity against Punta Toro Virus infection by the double-stranded RNA (dsRNA), poly(I:C12U), but not Poly(I:C): differential recognition of synthetic dsRNA molecules.
(20/89)
In the wake of RNA virus infections, dsRNA intermediates are often generated. These viral pathogen-associated molecular patterns can be sensed by a growing number of host cell cytosolic proteins and TLR3, which contribute to the induction of antiviral defenses. Recent evidence indicates that melanoma differentiation-associated gene-5 is the prominent host component mediating IFN production after exposure to the dsRNA analog, poly(I:C). We have previously reported that Punta Toro virus (PTV) infection in mice is exquisitely sensitive to treatment with poly(I:C(12)U), a dsRNA analog that has a superior safety profile while maintaining the beneficial activity of the parental poly(I:C) in the induction of innate immune responses. The precise host factor(s) mediating protective immunity following its administration remain to be elucidated. To assess the role of TLR3 in this process, mice lacking the receptor were used to investigate the induction of protective immunity, type I IFNs, and IL-6 following treatment. Unlike wild-type mice, those lacking TLR3 were not protected against PTV infection following poly(I:C(12)U) therapy and failed to produce IFN-alpha, IFN-beta, and IL-6. In contrast, poly(I:C) treatment significantly protected TLR3(-/-) mice from lethal challenge despite some deficiencies in cytokine induction. There was no indication that the lack of protection was due to the fact that TLR3-deficient mice had a reduced capacity to fight infection because they were not found to be more susceptible to PTV. We conclude that TLR3 is essential to the induction of antiviral activity elicited by poly(I:C(12)U), which does not appear to be recognized by the cytosolic sensor of poly(I:C), melanoma differentiation-associated gene-5. (+info)
Phylogenetic relationships among sandfly fever group viruses (Phlebovirus: Bunyaviridae) based on the small genome segment.
(21/89)
The phleboviruses are more diverse in terms of arthropod vectors and antigenic relationships than most other genera of arthropod-borne viruses. In this study, 30 sandfly fever group viruses from the Naples, Sicilian, Punta Toro, Icoaraci and Frijoles serocomplexes were sequenced. Phylogenetic analyses were performed based on the sequence of the open reading frame for the nucleoprotein (N) and non-structural (NSs) protein genes of the small (S) segment. The five resultant genotypic lineages correlated with the serological grouping and were similar to analysis of M segment sequences. The sequence identity for N and NSs genes within the Sicilian, Naples, Punta Toro, Icoaraci and Frijoles serocomplexes was determined. The results indicated that genetic divergence for the S segment is lower than that for the M segment, suggesting that the S segment is more stable during evolution. (+info)
Prophylactic and therapeutic intervention of Punta Toro virus (Phlebovirus, Bunyaviridae) infection in hamsters with interferon alfacon-1.
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Assembly and polarized release of Punta Toro virus and effects of brefeldin A.
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Punta Toro virus (PTV), a member of the sandfly fever group of bunyaviruses, is assembled by budding at intracellular membranes of the Golgi complex. We have examined PTV glycoprotein transport, assembly, and release and the effects of brefeldin A (BFA) on these processes. Both the G1 and G2 proteins were transported out of the endoplasmic reticulum (ER) and retained in the Golgi complex in a stable structure, either during PTV infection or when expressed from a vaccinia virus recombinant. BFA treatment causes a rapid and dramatic change in the distribution of the G1 and G2 proteins, from a Golgi pattern to an ER pattern. The G1 and G2 proteins were found to be modified by medial but not trans Golgi network enzymes, in the presence or absence of BFA. We found that BFA blocks PTV release from cells but does not interfere with the intracellular assembly of infectious virions. Further, the BFA block of virus release is fully reversible, with high levels of virus release occurring upon removal of the inhibitor. It was also found that the release of PTV virions is polarized, occurring exclusively from the basolateral surfaces of the polarized Vero C1008 epithelial cell line. (+info)