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Genomic evolution of porcine reproductive and respiratory syndrome virus (PRRSV) isolates revealed by deep sequencing.

ABSTRACT: Most studies on PRRSV evolution have been limited to a particular region of the viral genome. A thorough genome-wide understanding of the impact of different mechanisms on shaping PRRSV genetic diversity is still lacking. To this end, deep sequencing was used to obtain genomic sequences of a diverse set of 16 isolates from a region of Hong Kong with a complex PRRSV epidemiological record. Genome assemblies and phylogenetic typing indicated the co-circulation of strains of both genotypes (type 1 and type 2) with varying Nsp2 deletion patterns and distinct evolutionary lineages ("High Fever"-like and local endemic type). Recombination analyses revealed genomic breakpoints in structural and non-structural regions of genomes of both genotypes with evidence of many recombination events originating from common ancestors. Additionally, the high fold of coverage per nucleotide allowed the characterization of minor variants arising from the quasispecies of each strain. Overall, 0.56-2.83% of sites were found to be polymorphic with respect to cognate consensus genomes. The distribution of minor variants across each genome was not uniform indicating the influence of selective forces. Proportion of variants capable of causing an amino acid change in their respective codons ranged between 25-67% with many predicted to be non-deleterious. Low frequency deletion variants were also detected providing one possible mechanism for their sudden emergence as cited in previous reports.

SUBMITTER: Brar MS

PROVIDER: S-EPMC3974674 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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Genomic evolution of porcine reproductive and respiratory syndrome virus (PRRSV) isolates revealed by deep sequencing.

Brar Manreetpal Singh MS Shi Mang M Hui Raymond Kin-Hi RK Leung Frederick Chi-Ching FC

PloS one 20140403 4

Most studies on PRRSV evolution have been limited to a particular region of the viral genome. A thorough genome-wide understanding of the impact of different mechanisms on shaping PRRSV genetic diversity is still lacking. To this end, deep sequencing was used to obtain genomic sequences of a diverse set of 16 isolates from a region of Hong Kong with a complex PRRSV epidemiological record. Genome assemblies and phylogenetic typing indicated the co-circulation of strains of both genotypes (type 1 ...[more]

PMID: 24698958

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Project description:BackgroundPorcine reproductive and respiratory syndrome virus (PRRSV) is a causative agent of Porcine Reproductive and Respiratory Syndrome (PRRS). In this study, the complete nucleotide sequences of the selected two Thai PRRSV isolates, EU (01CB1) and US (01NP1) genotypes were determined since both isolates are the Thai prototypes.Results01CB1 and 01NP1 contain 14,943 and 15,412 nucleotides, respectively. The viruses compose 2 untranslated regions (5' UTR and 3' UTR) and 8 open reading frames (ORFs) designated as ORF1a, ORF1b and ORF2-7. Phylogenetic analysis of full length of the viruses also showed that the 01CB1 and 01NP1 were grouped into the EU and US genotype, respectively. In order to determine the genetic variation and genetic relatedness among PRRSV isolates, the complete nucleotide sequences of PRRSV isolated in Thailand, 01CB1 and 01NP1 were compared with those of 2 EU strains (Lelystad, and EuroPRRSV), 6 US strains (MLV, VR2332, PA8, 16244B, SP and HUN4). Our results showed that the 01CB1 genome shares approximately 99.2% (Lelystad) and 95.2% (EuroPRRSV) nucleotide identity with EU field strains. While, the 01NP1 genome has 99.9% nucleotide identity with a live vaccine strain (MLV) and 99.5% and 98.5% nucleotide identity with 2 other US isolates, VR2332 and 16244B, respectively. In addition, ORF5 nucleotide sequences of 9 PRRS viruses recovered in Thailand during 2002-2008 were also included in this study. Phylogenetic analysis of ORF5 showed high similarity among EU and US genotypes of the recent Thai PRRS viruses (2007-2008 viruses) with 01CB1 and 01NP1.ConclusionOverall, the results suggested that the Thai EU isolate (01CB1) may evolve from the EU prototype, Lelystad virus, whereas the Thai US isolate (01NP1) may originate and evolve from the vaccine virus or its derivatives. Interestingly, the US-MLV vaccine was not available in the Thai market in 2001. The Vaccine-like virus might have persisted in the imported pigs or semen and later spread in the Thai swine industry. This report is the first report of complete nucleotide sequences of the Thai PRRS viruses both EU and US genotypes.

Project description:BackgroundMechanisms of fetal death following maternal PRRSV2 infection remain uncharacterized, although hypoxia from umbilical cord lesions and/or placental detachment due to apoptosis are hypothesized. We performed two experiments examining hypoxia and apoptosis in PRRSV-infected and non-infected, third-trimester fetuses to elucidate possible associations with fetal death. Fetuses were selected based on four phenotypic infection groups: fetuses from non-challenged control gilts (CTRL); low viral load fetuses (LVL; Exp 1) or uninfected fetuses (UNINF; Exp 2) from inoculated gilts; viable high viral load fetuses (HVL-VIA); and HVL meconium-stained fetuses (HVL-MEC).ResultsIn experiment 1, paraffin embedded fetal tissues collected 21 days post maternal infection (DPI) were examined for DNA fragmentation associated with apoptosis. Positively stained foci were larger and more numerous (P < 0.05) in heart, liver, and thymus of HVL-VIA and HVL-MEC compared to CTRL and LVL fetuses. In experiment 2, group differences in gene expression within the hypoxia (HIF1a, IDO1, VEGFa, LDHA, NOS2, NOX1) and apoptosis (CASP3, CASP7, CASP8, CASP9, RIPK1, RIPK3) pathways were assessed by RT-qPCR in fetal tissues collected at 12 DPI. High viral load fetuses showed differential expression relative to the CTRL and UNINF (P < 0.05 for all). Brain tissue from HVL-VIA and HVL-MEC fetuses presented increased expression of CASP7, CASP8, RIPK3, HIF1a and IDO1. Fetal heart showed increased expression of CASP8, HIF1a, IDO and NOX1 and a decrease in NOS2 expression in infected groups. CASP7, CASP9, RIPK1 and RIPK3 were only increased in the heart of HVL-VIA while VEGFa was only increased for HVL-MEC fetuses. Thymus from HVL-MEC had decreased expression of CASP9 and there was increased IDO1 in all infected fetuses.ConclusionsThere is strong evidence of apoptosis occurring in the heart, liver and thymus of highly viral load fetuses at 21 DPI. Furthermore, there was clear upregulation of apoptotic genes in the heart of high viral load infected fetuses and less prominent upregulation in the brain of PRRSV-infected fetuses, whereas thymus appears to be spared at 12 DPI. There was no strong evidence of hypoxia at 12 DPI in brain and thymus but some indication of hypoxia occurring in fetal heart.

Dataset Information

Genomic evolution of porcine reproductive and respiratory syndrome virus (PRRSV) isolates revealed by deep sequencing.

Publications

Genomic evolution of porcine reproductive and respiratory syndrome virus (PRRSV) isolates revealed by deep sequencing.

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