Fig 4 - uploaded by Donald Netolitzky
Content may be subject to copyright.
Stem–loop structures in the 3h NTR. (a) Double stem–loop structures in SIN. (b) Double stem–loop structures in 3h NTR of WEE. Residues in the SIN-like 40 nt repeat are shaded. (c) Stem–loop structures in EEE.
Source publication
The complete nucleotide sequence of the 71V-1658 strain of western equine encephalitis virus (WEE) was determined (minus 25 nucleotides from the 5' end). A 5' RACE reaction was used to sequence the 5' terminus from WEE strain CBA87. The deduced WEE genome was 11508 nucleotides in length, excluding the 5' cap nucleotide and 3' poly(A) tail. The nucl...
Contexts in source publication
Context 1
... terminal repeats, as reported previously ( Ou et al., 1982), were found in WEE 71V-1658 (nt 11234-11273 and 11292-11331). However, the 3h NTR of WEE showed some surprising features that had not been described previously. The first 40 nt terminal repeat formed the backbone for the formation of a 57 nt double stem-loop structure (nt 11228-11284) (Fig. 4 b), consisting of an α and a β loop. The second 40 nt repeat of WEE formed a nearly identical 59 nt double stem-loop structure (nt 11285-11343), directly adjacent to the first structure. SIN, with three 40 nt repeats, forms three double stem-loops (Fig. 4 a), while EEE, which does not contain a SIN-like 40 nt repeat, contains the α and β ...
Context 2
... formed the backbone for the formation of a 57 nt double stem-loop structure (nt 11228-11284) (Fig. 4 b), consisting of an α and a β loop. The second 40 nt repeat of WEE formed a nearly identical 59 nt double stem-loop structure (nt 11285-11343), directly adjacent to the first structure. SIN, with three 40 nt repeats, forms three double stem-loops (Fig. 4 a), while EEE, which does not contain a SIN-like 40 nt repeat, contains the α and β loops (Fig. 4 ...
Context 3
... 4 b), consisting of an α and a β loop. The second 40 nt repeat of WEE formed a nearly identical 59 nt double stem-loop structure (nt 11285-11343), directly adjacent to the first structure. SIN, with three 40 nt repeats, forms three double stem-loops (Fig. 4 a), while EEE, which does not contain a SIN-like 40 nt repeat, contains the α and β loops (Fig. 4 ...
Context 4
... 3h NTRs of alphaviruses are characterized by wide- spread sequence divergence and yet contain small, strongly conserved motifs (reviewed in Strauss & Strauss, 1994 ;Pfeffer et al., 1998). Analysis of the 3h NTR indicated the presence of double stem-loop structures in SIN and WEE (Fig. 4 a, b). Interestingly, the 40 base repeat found in SIN and WEE is contained within the double stem-loop structure. SIN was found to contain three double stem-loop structures and WEE was found to contain two. In SIN, the spacing between the three double stem-loop structures was around 30 nucleotides, while in WEE, the distance was zero ...
Context 5
... Additional alphaviruses were assessed and it is interesting to note that double stem-loop structures were found in many of the WEE- and SIN-related viruses (SIN, Aura, Babanki, Ockelbo, Kyzylagach, Whataroa, WEE and HJ). The double stem-loop structures found in SIN and WEE consisted of the α loop [AUGUA(U\C)UU] and the β loop (GCAUAAU) (Fig. 4 b). Surprisingly, while EEE does not have the 40 base repeat element found in SIN and WEE, it contains the α and β loop structures (Fig. 4 c). The significance of these loop structures conserved between SIN, WEE and EEE has yet to be elucidated, although previous studies have suggested a role in virus replication and\or host specificity ...
Context 6
... and SIN-related viruses (SIN, Aura, Babanki, Ockelbo, Kyzylagach, Whataroa, WEE and HJ). The double stem-loop structures found in SIN and WEE consisted of the α loop [AUGUA(U\C)UU] and the β loop (GCAUAAU) (Fig. 4 b). Surprisingly, while EEE does not have the 40 base repeat element found in SIN and WEE, it contains the α and β loop structures (Fig. 4 c). The significance of these loop structures conserved between SIN, WEE and EEE has yet to be elucidated, although previous studies have suggested a role in virus replication and\or host specificity (Kuhn et al., 1990(Kuhn et al., , 1991. For example, a deletion of 26-318 nucleotides from the 3h end of SIN resulted in reduced virus ...
Citations
... 9 The positive sense single-stranded RNA genome of SINV allows a relatively large genome packaging capacity for proteins up to 5 kb. 10 WEEV is a hybrid with sequences derived from the Sindbis virus. 11 In VEEV, there are five structural proteins, which are considered as main targets for the acquired immune system. 12 ...
... The overall case-fatality rate of WEEV is 4%. 11 EEEV infection is responsible for very few cases per year in the US, with encephalitis cases showing high mortality (30%-70%) and neurological sequelae in survivors. 62,63 An epizootic disease is mostly produced by VEEV with 1% mortality. ...
... The Togaviridae family accommodates genus Alphavirus and 31 species, including several important human pathogens such as Eastern equine encephalitis virus (EEEV) [111], Western equine encephalitis virus (WEEV) [112], Venezuelan equine encephalitis virus (VEEV) [113], Sindbis virus (SINV) [114], Ross River virus (RRV) [115], Semliki Forest virus (SFV) [116], and Chikungunya virus [117]. Alphaviruses are arboviruses that are transmitted alternatively between insect vectors and vertebrate hosts [118,119]. ...
The cosmopolitan fungus Rhizoctonia solani has a wide host range and is the causal agent of numerous crop diseases, leading to significant economic losses. To date, no cultivars showing complete resistance to R. solani have been identified and it is imperative to develop a strategy to control the spread of the disease. Fungal viruses, or mycoviruses, are widespread in all major groups of fungi and next-generation sequencing (NGS) is currently the most efficient approach for their identification. An increasing number of novel mycoviruses are being reported, including double-stranded (ds) RNA, circular single-stranded (ss) DNA, negative sense (−)ssRNA, and positive sense (+)ssRNA viruses. The majority of mycovirus infections are cryptic with no obvious symptoms on the hosts; however, some mycoviruses may alter fungal host pathogenicity resulting in hypervirulence or hypovirulence and are therefore potential biological control agents that could be used to combat fungal diseases. R. solani harbors a range of dsRNA and ssRNA viruses, either belonging to established families, such as Endornaviridae, Tymoviridae, Partitiviridae, and Narnaviridae, or unclassified, and some of them have been associated with hypervirulence or hypovirulence. Here we discuss in depth the molecular features of known viruses infecting R. solani and their potential as biological control agents.
... The 5´ and 3´ ends encode four non-structural proteins (NSP1-4) and viral structural proteins, respectively. The heterodimer formed by glycoproteins E1 and E2 is strongly immunogenic, inducing the production of neutralising antibodies associated with protection (4,7,8,9,12,27,28,29,30,31). After viral attachment, mediated by E2, and pore formation and viral envelope-endosome membrane fusion mediated by E1, the nucleocapsid is released into the cytoplasm. ...
Alphaviral equine encephalomyelitis is a mosquito-borne infection that causes severe neurological disease and fatalities in horses and humans in the Americas. Consequently, the equine alphaviruses (Eastern, Western and Venezuelan) are of considerable concern worldwide and are notifiable to the World Organisation for Animal Health. In addition, these diseases are considered a potent potential biological weapon, emphasising the need to develop an effective vaccine. Alphaviral equine encephalomyelitis is caused by Eastern equine encephalomyelitis virus (EEEV), Western equine encephalomyelitis virus (WEEV) or Venezuelan equine encephalomyelitis virus (VEEV), which are related members of the Alphavirus genus in the Togaviridae family. Although related, the three viruses are genetically and antigenically distinct. The disease is characterised by fever, anorexia, depression and clinical signs of encephalomyelitis, and may be fatal in up to 90% of cases, for both humans and horses, particularly in the case of EEE. Surviving horses develop lifelong immunity but may have permanent neuropathology. The aim of this paper is to analyse the scientific information available on the evolution of EEE, WEE and VEE, and any potential vaccines.
... However, it was also demonstrated that, in vivo, FMV could infect Cs. melanura, the vector for EEEV and HJV . The genomes of two of the recombinants, WEEV and HJV, along with representative progenitors of the parental viruses, EEEV and SINV, have been described previously (Shirako et al., 1991;Weaver et al., 1993;Netolitzky et al., 2000;Allison and Stallknecht, 2009). However, the lack of the FMV genome has precluded a comprehensive comparative evolutionary analysis of the three recombinant viruses, although the genome of the variant Buggy Creek virus was recently sequenced (Forrester et al., 2012). ...
Western equine encephalitis virus (WEEV), Highlands J virus (HJV), and Fort Morgan virus (FMV) are the sole representatives of the WEE antigenic complex of the genus Alphavirus, family Togaviridae, that are endemic to North America. All three viruses have their ancestry in a recombination event involving eastern equine encephalitis virus (EEEV) and a Sindbis (SIN)-like virus that gave rise to a chimeric alphavirus that subsequently diversified into the present-day WEEV, HJV, and FMV. Here, we present a comparative analysis of the genetic, ecological, and evolutionary relationships among these recombinant-origin viruses, including the description of a nsP4 polymerase mutation in FMV that allows it to circumvent the host range barrier to Asian tiger mosquito cells, a vector species that is normally refractory to infection. Notably, we also provide evidence that the recombination event that gave rise to these three WEEV antigenic complex viruses may have occurred in North America.
... Infections of humans and horses can be fatal, and survivors often suffer permanent neurological sequelae (1,2). WEEV belongs to the genus Alphavirus in the family Togaviridae and has a positive-sense, single-stranded RNA genome approximately 11.5 kb in length, including two open reading frames (ORFs) flanked by 5=-and 3=-untranslated regions (UTRs) (3,4). One unusual feature of WEEV is that it is the descendant of an ancient recombination event between Sindbis virus (SINV)-like and eastern equine encephalitis virus (EEEV)-like ancestors (5,6). ...
Unlabelled:
Western equine encephalitis virus (WEEV) is an arbovirus from the genus Alphavirus, family Togaviridae, which circulates in North America between birds and mosquitoes, occasionally causing disease in humans and equids. In recent decades, human infection has decreased dramatically; the last documented human case in North America occurred in 1994, and the virus has not been detected in mosquito pools since 2008. Because limited information exists regarding the evolution of WEEV, we analyzed the genomic sequences of 33 low-passage-number strains with diverse geographic and temporal distributions and performed comprehensive phylogenetic analyses. Our results indicated that WEEV is a highly conserved alphavirus with only approximately 5% divergence in its most variable genes. We confirmed the presence of the previously determined group A and B lineages and further resolved group B into three sublineages. We also observed an increase in relative genetic diversity during the mid-20th century, which correlates with the emergence and cocirculation of several group B sublineages. The estimated WEEV population size dropped in the 1990s, with only the group B3 lineage being sampled in the past 20 years. Structural mapping showed that the majority of substitutions in the envelope glycoproteins occurred at the E2-E2 interface. We hypothesize that an event occurred in the mid-20th century that resulted in the increased genetic diversity of WEEV in North America, followed by genetic constriction due to either competitive displacement by the B3 sublineage or stochastic events resulting from a population decline.
Importance:
Western equine encephalitis virus (WEEV) has caused several epidemics that resulted in the deaths of thousands of humans and hundreds of thousands of equids during the past century. During recent decades, human infection decreased drastically and the virus has not been found in mosquito pools since 2008. Because limited information exists regarding the evolution of WEEV, we analyzed 33 complete genome sequences and conducted comprehensive phylogenetic analyses. We confirmed the presence of two major lineages, one of which diverged into three sublineages. Currently, only one of those sublineages is found circulating in nature. Understanding the evolution of WEEV over the past century provides a unique opportunity to observe an arbovirus that is in decline and to better understand what factors can cause said decline.
... Subunit vaccines consisting of recombinant forms of WEEV E2 or E1 have been reported to induce significant protection in animal models (6)(7)(8). Although E2 is the major neutralizing antigen, E1 is highly conserved among alphaviruses (9,10). Thus, E1 is an excellent vaccine candidate because it might offer broader ("pan-alphavirus") protection against fatal encephalitis. ...
... We purposefully chose this protein antigen for our LANAC formulation for several reasons. First, although E2 protein is the major neutralizing antigen, E1 is highly conserved among distinct alphaviral species (9,10), and antibody to E1 is protective (11). Second, the E1 of WEEV is a recent ancestor of the E1 from a Sindbis-like virus, which has been shown to induce cross-reactive and cross-protective antibodies (11,35). ...
Alphaviruses are mosquito-borne viruses that cause significant disease in animals and humans. Western and eastern equine encephalitis virus (WEEV and EEEV), two New World alphaviruses, can cause fatal encephalitis and EEEV is a select agent of concern in biodefense. However, we have no antiviral therapies against alphaviral disease and current vaccine strategies target only a single alphavirus species. In an effort to develop new tools for a broader response to outbreaks, we designed and tested a novel alphavirus vaccine comprised of cationic lipid nucleic acid complexes (CLNCs) and the ectodomain of WEEV E1 protein (E1ecto). Interestingly, we found that the CLNC component, alone, had therapeutic efficacy, as it increased survival of CD-1 mice following lethal WEEV infection. Immunization with the CLNC-WEEV E1ecto mixture (lipid-antigen-nucleic acid complexes; LANACs) using a prime/boost regimen provided 100% protection in mice challenged with WEEV subcutaneously, intranasally, or via mosquito. Mice immunized with LANAC mounted a strong humoral immune response, but did not produce neutralizing antibodies. Passive transfer of serum from LANAC E1-ecto immunized mice to non-immune CD-1 mice conferred protection to WEEV challenge, indicating that antibody is sufficient for protection. In addition, the LANAC E1-ecto immunization protocol significantly increased survival of mice following intranasal or subcutaneous challenge with EEEV. In summary, our LANAC formulation has therapeutic potential and is an effective vaccine strategy that offers protection against two distinct species of alphavirus irrespective of the route of infection. We discuss plausible mechanisms as well the potential utility of our LANAC formulation as a pan-alphavirus vaccine.
... e virus did in fact probably originate as a recombinant of EEEV and SINV. e capsid and nonstructural genes are derived from EEEV whilst the envelope protein genes are derived from a SIN-like virus [134][135][136][137]. ...
Alphaviruses are enveloped single-stranded positive sense RNA viruses of the family Togaviridae. The genus alphavirus contains nine viruses, which are of medical, theoretical, or economic importance, and which will be considered. Sindbis virus (SINV) and Semliki Forest (SFV), although of some medical importance, have largely been studied as models of viral pathogenicity. In mice, SINV and SFV infect neurons in the central nervous system and virulent strains induce lethal encephalitis, whereas avirulent strains of SFV induce demyelination. SFV infects the developing foetus and can be teratogenic. Venezuelan Equine Encephalitis virus, Eastern Equine Encephalitis virus, and Western Equine Encephalitis virus can induce encephalitis in horses and humans. They are prevalent in the Americas and are mosquito transmitted. Ross River virus, Chikungunya virus (CHIKV), and O’nyong-nyong virus (ONNV) are prevalent in Australasia, Africa and Asia, and Africa, respectively. ONNV virus is transmitted by Anopheles mosquitoes, while the other alphaviruses are transmitted by culicine mosquitoes. CHIKV has undergone adaptation to a new mosquito host which has increased its host range beyond Africa. Salmonid alphavirus is of economic importance in the farmed salmon and trout industry. It is postulated that future advances in research on alphavirus pathogenicity will come in the field of innate immunity.
... The latter authors postulate that bats may become infected, especially during epizootics in other hosts. The prototype virus of the genus Alphavirus is the Sindbis virus (SINV), which belongs together with the Highland J, Fort Morgan, Buggy Creek and Aura virus to the Western equine encephalomyelitis antigenic complex (Hubálek 2008;Lundström and Pfeffer 2010;Netolitzky et al. 2000). It was first recognized and isolated from Culex pipiens and Cx. ...
Bats are the only mammals with the capacity of powered flight. Nearly
1,000 species can be found all over the world except in the northern and southern
polar areas. They perform important ecosystem services such as control of insects,
reseeding of cut forests and pollination of plants, which provide food for humans
and animals. On the other side, they are also recognized to be natural reservoir hosts
of a large variety of zoonotic diseases with the ability to cross species barriers. To
date, more than 80 virus species of different groups and various parasites, which can
cause several diseases have been isolated or detected in bats. Especially their high
population density and gregarious roosting behaviour increase the likelihood of
intra- and inter-species transmission of infections. Another important factor, which
enables pathogens to spread long distances, is the migratory habit of some bat
species, resulting in a great dispersal capacity. The transmission of pathogens from
bats to humans or other animals occurs by direct contact with infected animals, their
blood and tissue or through vector species. One of the most important vector groups
are insects. With more than a million described species, they are the most diverse
group of animals. Especially haematophagous groups such as Cimicidae, Culicidae
or Phlebotominae are known as vectors for a variety of diseases. These include
bacteria, protozoan and metazoan parasites as well as viruses. We focused on
blood-feeding insects, because the presence of certain viruses in them as well as
in bats comprises a potential virus transmission from bats to humans through
mosquitoes or other blood-feeding insects. For this chapter, we could find 20 viruses from four different families and two parasitic pathogens detected in all three groups
of haematophagous insects.
... The homology with the 71V-1658 strain (GenBank Accession no. NC-003908) (Netolitzky et al. 2000) was 98.15% compared with the same E1 region (8,478~8,802bp). The virus stock was made with suckling mouse brains harvested 24 to 48 h after intracerebral inoculation and was prepared as a 10% W/V suspension. ...
Two western equine encephalomyelitis virus (WEEV) strains have been isolated in China. Our previous studies have verified that the mosquito Culex pipiens pallens Coquillett (Diptera: Culicidae) infected with WEEV was capable of transmitting this arbovirus, but it was not clear how the sequential multiplication and spread of virus occurred within the mosquito. In this study, we observed the distribution of WEEV antigen in orally-infected Cx. p. pallens by immunohistochemistry in order to better understand the initial infection, dissemination, and transmission of WEEV in the potential vector. Orally-infected WEEV dissemination varied within the different tissues of Cx. p. pallens, with virus antigen consistently observed in the salivary glands, foregut, midgut epithelial cells, Malpighian tubules, hindgut, and ovarian follicles of some individuals after various days of extrinsic incubation. We suggest that Cx. p. pallens, the potential vector of WEEV, has the ability to harbor the virus through the alimentary system, and the midgut epithelial cell may be the initial site of WEEV replication after ingestion of a viremic blood meal.
... Sequence analysis of WEEV strains-To identify potential virulence determinants at the molecular level, the coding regions of the full genomes of WEEV strains CO92-1356 (GenBank Accession Nos. FJ786260 and FJ786264), ON41-McMillan (FJ786263 and FJ786265), and TX71-TBT235 (FJ786261 and FJ786262) were sequenced and compared to a low virulence WEEV strain 71V-1658 [25,26]. Table 4 shows the deduced amino acid differences that are unique to each high-virulence WEEV strain (TBT235 and McMillan) when compared to both low virulence WEEV strains 71V-1658 and CO92. ...
We developed two types of chimeric Sindbis virus (SINV)/western equine encephalitis virus (WEEV) alphaviruses to investigate their potential use as live virus vaccines against WEE. The first-generation vaccine candidate, SIN/CO92, was derived from structural protein genes of WEEV strain CO92-1356, and two second-generation candidates were derived from WEEV strain McMillan. For both first- and second-generation vaccine candidates, the nonstructural protein genes were derived from SINV strain AR339. Second-generation vaccine candidates SIN/SIN/McM and SIN/EEE/McM included the envelope glycoprotein genes from WEEV strain McMillan; however, the amino-terminal half of the capsid, which encodes the RNA-binding domain, was derived from either SINV or eastern equine encephalitis virus (EEEV) strain FL93-939. All chimeric viruses replicated efficiently in mammalian and mosquito cell cultures and were highly attenuated in 6-week-old mice. Vaccinated mice developed little or no detectable disease and showed little or no evidence of challenge virus replication; however, all developed high titers of neutralizing antibodies. Upon intranasal challenge with high doses of virulent WEEV strains, mice vaccinated with >or=10(5)PFU of SIN/CO92 or >or=10(4)PFU of SIN/SIN/McM or SIN/EEE/McM were completely protected from disease. These findings support the potential use of these live-attenuated vaccine candidates as safe and effective vaccines against WEE.
