Sequence of the Genome of Lactate Dehydrogenase-Elevating Virus: Heterogenicity between Strains P and C

AbstractThe complete nucleotide sequence of genomic RNA (14104 nt) of one strain of lactate dehydrogenase-elevating virus (LDV), LDV-P, is reported. It exhibits only about 80% nucleotide identity with the sequence reported for another LDV strain, LDV-C (Godeny et al., Virology 194, 585-596 (1993), and is 68 nucleotides shorter than the reported LDV-C sequence. The difference in length is largely due to the lack of a 59-nucleotide-long direct repeat in ORF 1a of the reported LDV-C sequence. Sequence analysis of a total of 1.4 kb of ORF 1a of LDV-C via reverse transcription/polymerase chain reaction (RT/PCR) technology failed to confirm the presence of this repeat in the LDV-C genome as well as of 24 deletions/insertions of single nucleotides that give rise to apparent transient reading frame differences between the LDV-P and LDV-C genomes and might have represented frameshift mutations. An additional 35 nucleotides in ORF 1a of the RT/PCR LDV-C products were the same as in the LDV-P rather than the reported LDV-C genome. The nucleotide sequences of the 5′ leader and the 3′ noncoding ends of the two genomes and the heptanucleotides involved in joining the 5′ leader to the bodies of the subgenomic mRNAs were highly conserved or identical. The predicted LDV-P proteins, however, differed from those predicted for the LDV-C proteins between 25% for the ORF 2 protein and 1% for the ORF 7 nuoleocapsid protein. All functional motifs of the ORF 1a and ORF 1b proteins were conserved. The ORF 1a protein possesses 11 potential transmembrane segments that flank the serine protease domain

Porcine Reproductive and Respiratory Syndrome Virus: Origin Hypothesis

Porcine reproductive and respiratory syndrome is a serious swine disease that appeared suddenly in the midwestern United States and central Europe approximately 14 years ago; the disease has now spread worldwide. In North America and Europe, the syndrome is caused by two genotypes of porcine reproductive and respiratory syndrome virus (PRRSV), an arterivirus whose genomes diverge by approximately 40%. My hypothesis, which explains the origin and evolution of the two distinct PRRSV genotypes, is that a mutant of a closely related arterivirus of mice (lactate dehydrogenase-elevating virus) infected wild boars in central Europe. These wild boars functioned as intermediate hosts and spread the virus to North Carolina in imported, infected European wild boars in 1912; the virus then evolved independently on the two continents in the prevalent wild hog populations for approximately 70 years until independently entering the domestic pig population

Effect of porcine circovirus type 2a or 2b on infection kinetics and pathogenicity of two genetically divergent strains of porcine reproductive and respiratory syndrome virus in the conventional pig model

To determine differences in infection kinetics of two temporally and genetically different type 2 porcine reproductive and respiratory syndrome virus (PRRSV) isolates in vivo with and without concurrent porcine circovirus (PCV) type 2a or 2b infection, 62 pigs were randomly assigned to one of seven groups: negative controls (n = 8); pigs coinfected with a 1992 PRRSV strain (VR-2385) and PCV2a (CoI-92-2a; n = 9), pigs coinfected with VR-2385 and PCV2b (CoI-92-2b; n = 9), pigs coinfected with a 2006 PRRSV strain (NC16845b) and PCV2a (CoI-06-2a; n = 9), pigs coinfected with NC16845b and PCV2b (CoI-06-2b; n = 9), pigs infected with VR-2385 (n = 9), and pigs infected with NC16845b (n = 9). Blood samples were collected before inoculation and at day post-inoculation (dpi) 3, 6, 9 and 12 and tested for the presence of PRRSV antibody and RNA, PCV2 antibody and DNA, complete blood counts, and interferon gamma (IFN-γ) levels. Regardless of concurrent PCV2 infection, VR-2385 initially replicated at higher levels and reached peak replication levels at dpi 6. Pigs infected with VR-2385 had significantly higher amounts of viral RNA in serum on both dpi 3 and dpi 6, compared to pigs infected with NC16845b. The peak of NC16845b virus replication occurred between dpi 9 and dpi 12 and was associated with a delayed anti-PRRSV antibody response in these pigs. PCV2 coinfection resulted in significantly more severe macroscopic and microscopic lung lesions and a stronger anti-PRRSV IgG response compared to pigs infected with PRRSV alone. This work further emphasizes in vivo replication differences among PRRSV strains and the importance of coinfecting pathogens

Phylogenetic characterization of genes encoding for glycoprotein 5 and membrane protein of PRRSV isolate HH08

A porcine reproductive and respiratory syndrome virus (PRRSV) was obtained from clinic samples. Genes 5 and 6 encoding for the viral glycoprotein 5 and a membrane protein of the PRRSV designated as HH08 were amplified by reverse transcription-PCR. These sequences were compared with reference sequences derived from different geographical locations. The results indicated that the virus belongs to the North American type rather than European. Comparative analyses of the genetic diversity between the PRRSV isolate HH08 and other Chinese as well as foreign reference strains of PRRSV were discussed based on the sequence comparison and the topology of phylogenetic trees constructed in this study

Attenuation of porcine reproductive and respiratory syndrome virus by molecular breeding of virus envelope genes from genetically divergent strains

Molecular breeding via DNA shuffling can direct the evolution of viruses with desired traits. By using a positive-strand RNA virus, porcine reproductive and respiratory syndrome virus (PRRSV), as a model, rapid attenuation of the virus was achieved in this study by DNA shuffling of the viral envelope genes from multiple strains. The GP5 envelope genes of 7 genetically divergent PRRSV strains and the GP5-M genes of 6 different PRRSV strains were molecularly bred by DNA shuffling and iteration of the process, and the shuffled genes were cloned into the backbone of a DNA-launched PRRSV infectious clone. Two representative chimeric viruses, DS722 with shuffled GP5 genes and DS5M3 with shuffled GP5-M genes, were rescued and shown to replicate at a lower level and to form smaller plaques in vitro than their parental virus. An in vivo pathogenicity study revealed that pigs infected with the two chimeric viruses had significant reductions in viral-RNA loads in sera and lungs and in gross and microscopic lung lesions, indicating attenuation of the chimeric viruses. Furthermore, pigs vaccinated with the chimeric virus DS722, but not pigs vaccinated with DS5M3, still acquired protection against PRRSV challenge at a level similar to that of the parental virus. Therefore, this study reveals a unique approach through DNA shuffling of viral envelope genes to attenuate a positive-strand RNA virus. The results have important implications for future vaccine development and will generate broad general interest in the scientific community in rapidly attenuating other important human and veterinary viruses

Dual Regulation of Host TRAIP Post-translation and Nuclear/Plasma Distribution by Porcine Reproductive and Respiratory Syndrome Virus Non-structural Protein 1α Promotes Viral Proliferation

In this study, we show that porcine reproductive and respiratory syndrome virus (PRRSV) non-structural protein 1α (nsp1α) facilitates PRRSV escape from innate immune by modulating nuclear to cytoplasmic translocation and distribution ratio of TRAIP to promote virus proliferation. Mechanistically, TRAIP interacts with PRRSV nsp1α via its K205 site, while NSP1α decreases the SUMOylation and K48 ubiquitination independent of the TRAIP interaction K205 site. Modulation of the dual modification of TRAIP by PRRSV nsp1α results in over-enrichment of TRAIP in the cytoplasm. Enrichment of nsp1α-induced cytoplasmic TRAIP in turn leads to excessive K48 ubiquitination and degradation of serine/threonine-protein kinase (TBK1), thereby antagonizing TBK1-IRF3-IFN signaling. This study proposes a novel mechanism by which PRRSV utilizes host proteins to regulate innate immunity. Findings from this study provides novel perspective to advance our understanding in the pathogenesis of PRRSV

The role of porcine reproductive and respiratory syndrome (PRRS) virus structural and non-structural proteins in virus pathogenesis

Porcine reproductive and respiratory syndrome (PRRS) is an economically devastating viral disease affecting the swine industry worldwide. The etiological agent, PRRS virus (PRRSV), possesses a RNA viral genome with nine open reading frames (ORFs). The ORF1a and ORF1b replicase-associated genes encode the polyproteins pp1a and pp1ab, respectively. The pp1a is processed in nine non-structural proteins (nsps): nsp1a, nsp1b, and nsp2 to nsp8. Proteolytic cleavage of pp1ab generates products nsp9 to nsp12. The proteolytic pp1a cleavage products process and cleave pp1a and pp1ab into nsp products. The nsp9 to nsp12 are involved in virus genome transcription and replication. The 30 end of the viral genome encodes four minor and three major structural proteins. The GP2a, GP3 and GP4 (encoded by ORF2a, 3 and 4), are glycosylated membrane associated minor structural proteins. The fourth minor structural protein, the E protein (encoded by ORF2b), is an unglycosylated membrane associated protein. The viral envelope contains two major structural proteins: a glycosylated major envelope protein GP5 (encoded by ORF5) and an unglycosylated membrane M protein (encoded by ORF6). The third major structural protein is the nucleocapsid N protein (encoded by ORF7). All PRRSV non-structural and structural proteins are essential for virus replication, and PRRSV infectivity is relatively intolerant to subtle changes within the structural proteins. PRRSV virulence is multigenic and resides in both the non-structural and structural viral proteins. This review discusses the molecular characteristics, biological and immunological functions of the PRRSV structural and nsps and their involvement in the virus pathogenesis

Genetic Diversity of the ORF5 Gene of Porcine Reproductive and Respiratory Syndrome Virus Isolates in Southwest China from 2007 to 2009

To gain insight into the molecular epidemiology and possible mechanisms of genetic variation of porcine reproductive and respiratory syndrome (PRRS) in Yunnan Province of China, the ORF5 gene of 32 PRRSV isolates from clinical samples collected from 2007 to 2009 were sequenced and analyzed. Nucleotide and amino acid analyses were carried out on 32 isolates and representative strains of the North American genotype, European genotype and two representative Chinese isolates. Results revealed that these isolates share 86.9–99.0% nucleotide and 87.5–98.0% amino acid identity with VR-2332 the prototypical North American PRRSV, 61.7–62.9% and 54.3–57.8% with Lelystad virus (LV) the representative strain of European genotype, 91.2–95.4% and 90.0–94.5% with CH-1a that was isolated in mainland China in 1996, 88.1–99.3% and 85.5–99.0% with JX-A1 the representative strain of High pathogenic PRRSV in China, and 86.2–99.8% and 85.5–100.0% between isolated strains of different years, respectively. Phylogenetic analysis revealed that all 32 PRRSV isolates belonged to the North American genotype and were further divided into two different subgenotypes. Subgenotype 1 comprised twenty two Yunnan isolates which divided into two branches. Subgenotype 2 comprised ten isolates which closely related to the RespPRRS vaccine and its parent strain VR-2332. The functional domains of GP5 such as the signal peptide, ectodomain, transmembrane regions and endodomain were identified and some motifs in GP5 with known functions, such as primary neutralizing epitope (PNE) and decoy epitope were also further analyzed. Our study shown the great genetic diversity of PRRSV in southwest China, rendering the guide for control and prevention of this disease