Genetic and antigenic characterization of complete genomes of Type 1 Porcine Reproductive and Respiratory Syndrome viruses (PRRSV) isolated in Denmark over a period of 10 years

Porcine Reproductive and Respiratory Syndrome (PRRS) caused by the PRRS virus (PRRSV) is considered one of the most devastating swine diseases worldwide. PRRS viruses are divided into two major genotypes, Type 1 and Type 2, with pronounced diversity between and within the genotypes. In Denmark more than 50% of the herds are infected with Type 1 and/or Type 2 PRRSV. The main objective of this study was to examine the genetic diversity and drift of Type 1 viruses in a population with limited introduction of new animals and semen. A total of 43 ORF5 and 42 ORF7 nucleotide sequences were obtained from viruses collected from 2003 to February 2013. Phylogenetic analysis of ORF5 nucleotide sequences showed that the Danish isolates formed two major clusters within the subtype 1. The nucleotide identity to the subtype 1 protogenotype Lelystad virus (LV) spanned 84.9–98.8% for ORF5 and 90.7–100% for ORF7. Among the Danish viruses the pairwise nucleotide identities in ORF5 and ORF7 were 81.2–100% and 88.9–100%, respectively. Sequencing of the complete genomes, including the 5′- and 3′-end nucleotides, of 8 Danish PRRSV Type 1 showed that the genome lengths differed from 14,876 to 15,098 nucleotides and the pairwise nucleotide identity among the Danish viruses was 86.5–97.3% and the identity to LV was 88.7–97.9%. The study strongly indicated that there have been at least two independent introductions of Type 1 PRRSV in Denmark and analysis of the full genomes revealed a significant drift in several regions of the virus.     

Molecular characterization and recombination analysis of porcine reproductive and respiratory syndrome virus emerged in southwestern China during 2012–2016

Porcine reproductive and respiratory syndrome virus (PRRSV) is an important swine pathogen causing tremendous economic losses to the swine industry. To investigate the prevalence of PRRSV of genotype 2 (North American type, NA-type) in southwestern China, the Nsp2 hypervariable region (Nsp2 HV) and ORF5 of 61 PRRS viruses collected during 2012–2016 were sequenced and analyzed. All the virus detected clustered into the JXA1-like (52/61), VR-2332-like (7/61), and NADC30-like (2/61) sub-genotypes. Five deletions in Nsp2 HV were detected in addition to the typical 30aa discontinuous deletion in HP-PRRSV, and two of these five were not reported previously. Strikingly, two PRRS virus (SCnj16 and SCcd16) isolated in 2016 contained the classic HP-PRRSV molecular marker in the Nsp2-coding region, but belonged to the NADC30-like sub-genotype on the ORF5 gene. Further recombination and phylogenetic analysis on the two complete genomic sequences revealed that they may have originated from recombination events between the NADC30 and Chinese HP-PRRSV strains. The present study suggests that the endemic PRRSVs in the region have continuously evolved and new vaccine strategies are necessary for more efficient control of the virus.     

Genetic diversity of the ORF5 gene of porcine reproductive and respiratory syndrome virus isolates in China from 2006 to 2008

Since April 2006, swine herds have experienced the outbreaks of a highly pathogenic porcine reproductive and respiratory syndrome (PRRS) in China. To explore the possible mechanism of the emergence of the highly pathogenic PRRS and more fully understand the extent of genetic diversity of PRRSV in China, we analyzed the ORF5 gene sequences of 159 representative PRRSV isolates in 16 provinces from 2006 to 2008. Sequence and phylogenetic analyses showed that all these 159 isolates belonged to the North American genotype and were further divided into six subgenotypes; 140 of 159 isolates were closely related to the highly pathogenic PRRSV with 98.5–100% nucleotide and 98.3–100% amino acid sequence identities and belonged to Subgenotype I; and 3, 8, 4, 3, 1 of 159 isolates were part of Subgenotypes II–VI, respectively. Amino acid analysis of the GP5 protein revealed that all the isolates in Subgenotypes I–III were found to be highly variable in the primary neutralizing epitope; most of the isolates in Subgenotypes I and IV had more glycosylation sites than those in Subgenotypes II, III, V and VI; and 1, 5, and 9 unique amino acid mutations were observed in Subgenotypes I, IV and VI, respectively. In conclusion, our study provides the evidence of coexistence of six different subgenotype isolates in pigs in China from 2006 to 2008, and emphasizes the importance of reinforcing PRRSV surveillance, especially after the emergence of highly pathogenic PRRS in China.     

The genetic diversity of European type PRRSV is similar to that of the North American type but is geographically skewed within Europe

Porcine reproductive and respiratory syndrome virus (PRRSV) is a recently emerged pathogen. Two PRRSV genotypes exist, North American and European, which are only 55-70% identical at the nucleotide level. Previous studies have shown high nucleotide diversity in the North American genotype and low nucleotide diversity in the European genotype. Here, we analyzed the ORF5 and ORF7 genes for a large number of new European type PRRSV isolates in conjunction with existing database sequences. This new analysis showed that contrary to previous assumptions, genetic diversity is at least as high in the European genotype as in the North American genotype. Furthermore, we showed that genetic diversity of European type PRRSV has a marked geographical pattern, with exceptionally high genetic diversity among Italian sequences. The geographical pattern of diversity in relation to the epidemiology of PRRSV in Europe is discussed. Discrepancies between ORF5- and ORF7-based genealogies were observed, and further analysis of the data set confirmed the presence of recombination. We were therefore able to report the first observation of recombination in wild-type isolates of European genotype PRRSV.     

Sequence analysis of NSP9 gene of 25 PRRSV strains from Guangdong province, subtropical southern China

In order to evaluate the trend in the prevalence of porcine reproductive and respiratory syndrome on farms, 25 porcine reproductive and respiratory syndrome virus (PRRSV) strains were collected from 2009 to 2012 from 11 districts in the Guangdong province. The complete gene sequences of NSP9 from the 25 PRRSV strains were amplified, sequenced, and then compared with the published sequences of other strains. The results showed that these sequences shared 97.6–99.4 % of the same nucleotides and 97.5–99.4 % of the same amino acid sequence identities with JXA1, 91.4–93.1 % of the same nucleotides and 95.4–92.6 % of the same amino acid sequence identities with VR2332, and 95.1–98.4 % of the same nucleotides and 95.3–97.0 % of the same amino acid sequences with the classical Chinese vaccine strain Ch-1a. Phylogenetic analyses showed that all of these 25 strains belonged to the North American genotype and were further divided into three sub-genotypes. This basic data allowed us to evaluate the prevalence of PRRS in the Guangdong province and the variation and evolution of the nsp9 gene in PRRSV.     

Genomic characterization of two Chinese isolates of <Emphasis Type="Italic">Porcine respiratory and reproductive syndrome virus</Emphasis>

Summary. The genomes of two isolates of Porcine respiratory and reproductive syndrome virus (PRRSV) from China, designated HB-1(sh)/2002 and HB-2(sh)/2002, were sequenced and analyzed. The size of the genomes of HB-1(sh)/2002 and HB-2(sh)/2002 were 15,411 and 15,373 nucleotides respectively, excluding the poly(A) tails. Comparative analysis with the genomic sequences of another Chinese isolate (BJ-4) and North American (VR2332) and European (Lelystad virus, LV) viruses revealed that HB-1(sh)/2002 shared 89.8% identity with BJ-4 and VR2332, but only 54.7% with LV; while HB-2(sh)/2002 shared 89.4% and 89.5% identity with BJ-4 and VR2332 respectively and 54.3% with LV, indicating that the two new Chinese isolates were related to the North American PRRSV genotype. Phylogenetic analysis based on the nucleotide sequence of the structural protein ORFs showed that the two new Chinese isolates belong to same genetic subgroup. HB-2(sh)/2002 additionally exhibited variations in the NSP2 nonstructural protein encoded by ORF1 and the structural protein GP3 encoded by ORF3 in comparison with other North American PRRSV isolates, namely a 12 amino acids deletion in Nsp2 and one amino acid deletion in GP3 were found in HB-2(sh)/2002. Therefore, HB-2(sh)/2002 was a novel strain with unique deletions.     

Molecular characterization of PL97-1, the first Korean isolate of the porcine reproductive and respiratory syndrome virus

We determined the complete nucleotide and predicted amino acid sequence of the genomic RNA of PL97-1, the first Korean strain of porcine reproductive and respiratory syndrome virus (PRRSV), which was isolated from the serum of an infected pig in 1997. We found that the 15411-nucleotide genome of PL97-1 consisted of a 189-nucleotide 5' noncoding region (NCR), a 15071-nucleotide protein-coding region, and a 151-nucleotide 3'NCR, followed by a poly (A) tail. The 5'-end of PL97-1 began with 1ATG ACG TAT AGG12. Comparison of the PL97-1 genome with the 11 fully sequenced PRRSV genomes currently available revealed sequence divergence ranging from 0.3% (the VR-2332-derived vaccine MLV RespPRRS/Repro strain) to 38% (the Dutch Lelystad strain). To better understand the genetic relationships between these different strains, phylogenetic analyses were performed on the full-length PRRSV genomes. Significantly, the phylogenetic tree based on the ORF1b or ORF7 genes most closely resembled the tree based on the full-length genomes. Thus, these single genes will be the most useful in revealing the genetic relationships between the different strains relative to their geographical distribution. Extensive phylogenetic analyses using the ORF7 sequences of 111 PRRSV isolates available revealed that PL97-1 is most closely related to the North American genotype VR-2332, a VR-2332-derived vaccine strain, and Chinese BJ-4. It is distantly related to the European genotype Lelystad. This study provides the largest full-length genome phylogenetic analysis of PRRSV that has been published to date, and supports an earlier genetic grouping of the many temporally and geographically diverse PRRSV strains currently isolated.     

Development of a heteroduplex mobility assay to identify field isolates of porcine reproductive and respiratory syndrome virus with nucleotide sequences closely related to those of modified live-attenuated vaccines

Porcine reproductive and respiratory syndrome has been devastating the swine industry since the late 1980s. The disease has been controlled, to some extent, through the use of modified live-attenuated (MLV) vaccines once available. However, such a practice periodically resulted in isolation or detection of vaccine-like viruses from pigs as determined by a partial genomic sequencing. In this study, we developed a heteroduplex mobility assay (HMA) for quickly identifying porcine reproductive and respiratory syndrome virus (PRRSV) isolates with significant nucleotide sequence identities (>/=98%) with the modified live-attenuated vaccines. The major envelope gene (ORF5) of 51 PRRSV field isolates recovered before and after the introduction of the vaccines was amplified, denatured, and reannealed with the HMA reference vaccine strains Ingelvac PRRS MLV and Ingelvac PRRS ATP, respectively. Nine of the 51 field isolates and the VR2332 parent virus of Ingelvac PRRS MLV, which were all highly related to Ingelvac PRRS MLV with 相似文献   

Sequence analysis of porcine reproductive and respiratory syndrome virus of the American type collected from Danish swine herds

Summary.  Vaccine-like viruses of American type of porcine reproductive and respiratory syndrome virus (PRRSV) were detected in serum samples by RT-PCR. The viruses were analysed by nucleotide sequencing of the genomic region encoding open reading frames 2 to 7. During the ongoing study of Danish isolates of PRRSV by means of nucleotide sequencing, RT-PCR reactions and subsequent nucleotide sequencing showed the presence of American type PRRSV in Danish breeding herds. Most likely, these atypical viruses originated from boars vaccinated with live vaccine of American type (MLV RespPRRS), which were taken to artificial insemination centres and there brought together with unvaccinated boars already at the centres. The nucleotide sequences of three Danish viruses of American type PRRSV were compared to those of known PRRSV isolates. The nucleotide sequence identities of the atypical Danish isolates were between 99.2–99.5% to the vaccine virus RespPRRS and 99.0–99.3% to VR2332 which are the parental virus to the vaccine virus. Phylogenetic analysis including field isolates of American type supports the conclusion that the introduction of American type PRRSV in Denmark was due to spread of vaccine virus. Received December 24, 1997 Accepted April 15, 1998     

Molecular epidemiological investigation of porcine reproductive and respiratory syndrome virus in Northwest China from 2007 to 2010

Porcine reproductive and respiratory syndrome (PRRS) is an economically important swine disease affecting swine worldwide. Northwest China has a sparse pig population and there is no comprehensive information currently available on PRRSV infection. In this study, we analyzed the epidemiological features and genetic diversity of PRRSV from this region. 322 field-isolated tissues or serum samples were collected from aborted pig fetuses or pigs with respiratory disease from 15 herds, twice over a period of 2 years. PRRSV infection was determined and virus strains were classified by the sequencing of GP5. We found that 35.9 % of the animals were PRRSV-positive, and the average prevalence in 2007-2008 and 2009-2010 was 46.5 and 29.3 %, respectively. To further investigate the genetic divergence of PRRSV samples collected from 2007 to 2010, 32 strains were isolated for GP5 sequencing and analysis, and phylogenetic trees were created based on GP5 amino acid sequences. All PRRSVs were of the North American genotype and belonged to the highly pathogenic HP-PRRSV subgenotype. Isolates from the Xinjiang province formed a tightly clustered branch and were closely related to an evolutionary intermediate subgroup isolate. Virus sequences from 2007 to 2008 were compared with those from 2009 to 2010 from the same herd. New mutations were found in isolates after 2009 and focused on nucleotides in the GP5 antibody decoy epitope. PRRSV strains in Northwest China from 2007 to 2010 were similar to those from other regions of China, with some regional characteristics. These results contribute to the knowledge of PRRSV epidemiology in China.     

Discriminating between serological responses to European-genotype live vaccine and European-genotype field strains of porcine reproductive and respiratory syndrome virus (PRRSV) by peptide ELISA

A peptide ELISA was developed based on an immunodominant and hypervariable epitope in the ORF4 envelope glycoprotein of porcine reproductive and respiratory syndrome virus (PRRSV). The peptide sequence was derived from the Porcilis live-attenuated PRRSV vaccine strain (genotype 1, European). Antibodies induced by the field PRRSVs currently circulating in Poland were not detected by the Porcilis ORF4 peptide ELISA. In contrast, Porcilis-vaccinated animals seroconverted in the ORF4 peptide ELISA at 21 days post-vaccination. Maximal titers were seen 30-92 days post-vaccination; most sera had endpoint titers between 1:1000 and 1:100,000. In a paired format, where sera were assayed in two separate ELISAs using ORF4 peptides derived from the genetically very closely related Porcilis and Lelystad PRRSV strains, it was possible to differentiate between antibodies induced by these two viruses. The Porcilis and Lelystad ORF4 peptide ELISAs had sensitivities of 89 and 100%, respectively. Thus, ORF4 peptide ELISA afforded specific detection of antibodies induced by an European-genotype live-attenuated vaccine PRRSV strain (Porcilis). The results suggest that specific ORF4 peptide ELISAs can be custom-made for European-genotype PRRSV strains, using general peptide design criteria described in this work. Thus, ORF4 ELISAs may be generally useful, to monitor safety and operational aspects of European-genotype live-attenuated PRRSV vaccine virus use in populations with circulating field European-genotype PRRSVs.     

Tracing the genetic history of porcine reproductive and respiratory syndrome viruses derived from the complete ORF 5-7 sequences: a Bayesian coalescent approach

To trace the genetic history of porcine reproductive and respiratory syndrome virus (PRRSV), we determined the complete sequences of ORFs 5 to 7 of four PRRSV isolates. These sequences were analyzed together with published sequences from 146 isolates from various parts of the world using a Bayesian coalescent approach as well as Bayesian inference and maximum-likelihood methods. All of the European-type (EU-type) viruses were classified into one of two groups or unclassified (4 isolates), while all North American–type (NA-type) viruses belonged to one of three major groups or were unclassified (5 isolates). Within each genotype, no apparent periodic and/or geographic influence on the evolution of PRRSVs was observed. The evolutionary rate of PRRSV isolates was estimated to be 1.55?×?10^?3 substitutions/site/year, and the time of the most recent common ancestor (TMRCA) was 491.2?years ago. Here, the TMRCA for the EU- and NA-type viruses was 58.7 and 62.6?years ago, respectively. A Bayesian skyline plot revealed that the viruses evolved at an almost constant population size until the late 1970s, when they experienced a population expansion that continued until the late 1980s. The population size then remained constant again until the early 2000s, when a rapid, sharp decline in the effective number of infections occurred.     

Phylogenetic analyses of the putative M (ORF 6) and N (ORF 7) genes of porcine reproductive and respiratory syndrome virus (PRRSV): implication for the existence of two genotypes of PRRSV in the U.S.A. and Europe

Summary The putative membrane (M) protein (ORF 6) and nucleocapsid (N) protein (ORF 7) genes of five U.S. isolates of porcine reproductive and respiratory syndrome virus (PRRSV) with differing virulence were cloned and sequenced. To determine the genetic variation and the phylogenetic relationship of PRRSV, the deduced amino acid sequences of the putative M and N proteins from these isolates were aligned, to the extent known, with other PRRSV isolates, and also other members of the proposed arterivirus group including lactate dehydrogenase-elevating virus (LDV) and equine arteritis virus (EAV). There was 96–100% amino acid sequence identity in the putative M and N genes among U.S. and Canadian PRRSV isolates with differing virulence. However, their amino acid sequences varied extensively from those of European PRRSV isolates, and displayed only 57–59% and 78–81% identity, respectively. The phylogenetic trees constructed on the basis of the putative M and N genes of the proposed arterivirus group were similar and indicated that both U.S. and European PRRSV isolates were related to LDV and were distantly related to EAV. The U.S. and European PRRSV isolates fell into two distinct groups, suggesting that U.S. and European PRRSV isolates represent two distinct genotypes.     

Heterogeneity in Nsp2 of European-like porcine reproductive and respiratory syndrome viruses isolated in the United States

Recently, isolates of porcine reproductive and respiratory syndrome virus (PRRSV) that possess nucleotide sequences similar to European isolates have been reported in United States herds. The origin, diversity and prevalence of European-like North American PRRSV isolates in the U.S. remain unknown. Nucleotide sequence analysis of the 12 kb ORF1 of a North American isolate, SDPRRS 01-08 (01-08), showed 93.7% identity with Lelystad virus (LV), the prototypic European isolate, but only 58% identity with VR-2332, the prototypic North American isolate. Comparisons between LV and 01-08 at the peptide sequence level of the predicted non-structural proteins (Nsp) showed that Nsp9 (98.9% amino acid identity) was the most conserved and the least conserved was Nsp2 at 90.6% identity. For the purpose of comparison, GP5, the principal envelope structural protein, showed a 93.5% identity between 01-08 and LV. The most dramatic differences between the Nsp2 proteins of LV and 01-08 were a single 17 amino acid deletion between residues 734 and 750, as well as several amino acid differences. The same deletion was identified in the Nsp2 in five of seven other EuroPRRSV isolates submitted to the South Dakota Animal Disease Research and Diagnostic Laboratory. The remaining two isolates contained small deletions, but in other regions of Nsp2. Peptide sequence diversity in the form of hypervariability and deletions in Nsp2 demonstrate that European-like PRRSV isolates in the USA represent a heterogeneous group. Furthermore, areas in Nsp2 with deletions and amino acid hypervariability localize to regions that are predicted to be immunologically important.     

Examination of the selective pressures on a live PRRS vaccine virus

Summary.  We determined the ORF5 and 7 sequences of 20 pathogenic revertants of a live PRRSV vaccine. The sequence analysis confirmed all 20 isolates to be of vaccine origin. Having established that clonal introduction of American (vaccine) PRRS virus had occurred in Denmark, we could perform analysis of the selective pressure this attenuated virus had experienced during reversion. An analysis of nucleotide mutations showed a similar rate of mutations in the two genes (ORF5 and 7). However, non-synonymous mutations in ORF7 were eliminated by purifying selection. In contrast, non-synonymous mutations in ORF5 were tolerated or even selected for. The cDNA sequencing of the 20 vaccine virus revertants identified two single nucleotide mutations located in ORF5 and in ORF6 that we suggest are involved or at least linked to the attenuation of the vaccine virus and to the subsequent reversion to virulence. Received April 10, 1999/Accepted July 13, 1999     

Comparison of the structural protein coding sequences of the VR-2332 and Lelystad virus strains of the PRRS virus

Summary The 3-portion of the genome of a U.S. isolate of the porcine reproductive and respiratory syndrome (PRRS) virus, ATCC VR-2332, was cloned and sequenced. The resultant 3358 nucleotides contain 6 open reading frames (ORFs) with homologies to ORFs 2 through 7 of the European strain of the PRRS virus and other members of the free-standing genus of arteriviruses. Both VR-2332 and the European isolate (called the Lelystad virus) have been identified as infectious agents responsible for the swine disease called PRRS. Comparative sequence analysis indicates that there are degrees of amino acid identity to the Lelystad virus open reading frames ranging from 55% in ORF 5 to 79% in ORF 6. Hydropathy profiles indicate that the ORFs of VR-2332 and Lelystad virus correspond structurally despite significant sequence differences. These results are consistent with the biological similarities but distinct serological properties of North American and European isolates of the virus.     

Genetic variation and geographic distribution of porcine reproductive and respiratory syndrome virus in Japan

Summary. Porcine reproductive and respiratory syndrome virus (PRRSV) has two genotypes, the North American-type (NA-type) and the European-type (EU-type), and each genotype is also genetically diverged. We sequenced the ORF5 gene of 30 PRRSVs isolated from 23 prefectures of Japan during 1992 and 1993 and during 2000 and 2001. All of the isolates were of the NA-type. Phylogenetic analysis of the overall NA-type viruses isolated from around the world identified five major genetic clusters. The 1992–1993 Japanese samples belonged to only two genetic clusters, while the 2000–2001 samples included more diverged ORF5 genomes. One genetic cluster, which included 63% (20/32) of Japanese isolates, one Taiwanese isolate and one Chinese isolate, was mainly found in the eastern part of Japan. Another genetic cluster, which was found in various areas around the world, was distributed in the western part of Japan.     

Genetic variation and pathogenicity of highly virulent porcine reproductive and respiratory syndrome virus emerging in China

A highly pathogenic swine disease designated as ‘porcine high fever disease (PHFD)’ appeared recently in China. Porcine reproductive and respiratory syndrome virus (PRRSV) was identified as an agent associated with PHFD, and two discontiguous sequence deletions were identified as a genetic marker in the Nsp2 region of the viral genome. To examine PHFD in Shandong province, a total of 10 PRRSV isolates were recovered from pig herds that had never been vaccinated for PRRS. Sequence analysis of open reading frame 5 (ORF5) showed that the level of identity among the 10 isolates ranged between 88.2 and 99.2%. For the non-structural protein 2 (Nsp2) gene, three isolates shared high sequence identity with VR-2332, the prototype virus of the North American genotype, while the remaining seven isolates exhibited two discontiguous sequence deletions that were identical to those of PHFD: a one-amino-acid (phenylalanine) deletion at position 482 and a 29-amino-acid deletion at positions 533–561 of Nsp2. Experimental infection of pigs with SD-JN, which was one of the seven isolates containing such deletions, resulted in severe clinical symptoms characterized by red discoloration on the body and hemorrhages in the lungs, kidneys, and inguinal lymph nodes, accompanied by higher mortality and longer duration of viremia. These symptoms were similar to those of PHFD observed in the field. Our results show that VR2332-like PRRSV coexists with PHFD-associated atypical PRRSV in pig herds in the Shandong area, and different PRRSV isolates differ greatly in their pathogenesis and virulence in pigs.     

Attenuation of porcine reproductive and respiratory syndrome virus strain MN184 using chimeric construction with vaccine sequence

Two genetically distinct infectious recombinant virus clones (pMLV, constructed from Ingelvac PRRS MLV and pMN184, constructed from virulent strain MN184) were developed to study attenuation of contemporary porcine reproductive and respiratory syndrome virus (PRRSV) strain MN184. Two reciprocal chimeric clones (pMLVORF1/MN184 and pMN184ORF1/MLV) were then constructed, such that the 5'UTR/ORF1 of one genotype was linked to ORF2-7/3'UTR from the other genotype. In vitro studies demonstrated that the rescued chimeric viruses possessed intermediate growth properties compared to recombinant rMLV and rMN184. Swine inoculation with rMN184 and rMLV verified that these viruses fully mimicked the respective parent virus. In addition, earlier and higher antibody responses were detected in animals infected with rMN184 in contrast to those infected with rMLV. Chimeric virus treatment groups showed similar antibody responses as seen with these parent viruses, but much less severe pathogenesis when compared to the rMN184 group. These data suggested that genetic aspects of Ingelvac PRRS MLV 5'UTR/ORF1 replicase region and/or the structural proteins/3'UTR can serve to attenuate virulent strain MN184. The data also indicated that designed PRRSV vaccines could be developed, keeping the known 5'UTR/replicase region of an early vaccine strain such as Ingelvac PRRS MLV intact, but replacing the structural protein/3'UTR domain with that of an emerging virulent virus.