Complete genomic characterization of two European-genotype porcine reproductive and respiratory syndrome virus isolates in Fujian province of China.

Porcine reproductive and respiratory syndrome (PRRS) is considered one of the most devastating swine diseases worldwide, resulting in immense economic losses. PRRS virus (PRRSV) is divided into two major genotypes, European (type 1) and the North American (type 2). Type 1 PRRSV have recently emerged in Fujian province (South China), and this might have a significant impact on the Chinese pig industry. From 2013 to 2014, two type 1 PRRSV strains, named FJEU13 and FJQEU14, were isolated from piglets and sows with respiratory problems and reproductive disorders in Fujian province. The full genome length of the two isolates was 14,869-15,062 nucleotides (nt), excluding the poly(A) tail. These isolates shared 86.0-89.9% sequence identity with the prototypic strains Lelystad virus (LV) and 82.8-92% with Chinese type 1 PRRSV strains, but only 59.9-60.1% with the North American reference strain VR-2332. However, they were 82.9% identical to each other. Nonstructural protein 2 (Nsp2) and ORF3-ORF5 were the most variable regions when compared to other type 1 PRRSV strains. Nsp2 and ORF3 contained multiple discontinuous deletions and a 204-bp deletion in NSP2 in isolate FJQEU14, which has never been described in other Chinese type 1 PRRSV strains. All of these results might be useful for understanding the epidemic status of type 1 PRRSV in China.

Multiplex PCR for the simultaneous detection of porcine reproductive and respiratory syndrome virus, classical swine fever virus, and porcine circovirus in pigs.

A multiplex polymerase chain reaction (PCR) was designed for the simultaneous detection of three viruses involved in reproductive and respiratory failure in pigs: porcine reproductive and respiratory syndrome virus (PRRSV), classical swine fever virus (CSFV) and porcine circovirus type 2 (PCV-2). Each target produced a specific amplicon with a size of 718 bp (PRRSV), 288 bp (CSFV), or 466 bp (PCV-2). The sensitivity of the multiplex PCR using purified plasmid constructs containing the specific viral target fragments was 2.0 × 10(4), 2.5 × 10(3), and 6.0 × 10(2) copies for PRRSV, CSFV, and PCV-2, respectively. Non-specific reactions were not observed when other viruses, bacteria, and PK-15/Marc-145 cells were used to assess the multiplex PCR. Among 82 clinical samples from Fujian province, co-infection by PRRSV and CSFV was 12.19%, co-infection by PRRSV and PCV-2 was 21.95%, CSFV and PCV-2 was 13.41%, and co-infection by the three viruses was 3.66%. In conclusion, the multiplex PCR should be useful for routine molecular diagnosis and epidemiology. The multiplex PCR was effective in detecting various combinations of one or more of these viruses in pig specimens.

Genetic diversity and evolutionary characterization of Chinese porcine reproductive and respiratory syndrome viruses based on NSP2 and ORF5.

To more fully understand the extent of genetic diversity of PRRSV in China, we analyzed the Nsp2 and ORF5 gene sequences of 35 representative PRRSV isolates from 2008 to 2012. Sequence analysis revealed that the Nsp2 and ORF5 genes have undergone genetic variation. Furthermore, the isolate FJLYDX04 contains five insertions at positions 599 to 603 and is the first isolate from China reported to have an insertion in Nsp2. Our results suggest that the highly pathogenic PRRSV has become the dominant strain in China and that Chinese PRRSV has undergone rapid evolution and can circumvent immune responses induced by currently used vaccines.

Protective Efficacy of an Inactive Vaccine Based on the LY02 Isolate against Acute Haemophilus parasuis Infection in Piglets.

Haemophilus parasuis can cause Glässer's disease characterized by fibrinous polyserositis, polyarthritis, and meningitis. The current prevention of Glässer's disease is mainly based on the inactive vaccines; however, the protective efficacy usually fails in heterogeneous or homologous challenges. Here, the predominant lineage of H. parasuis (LY02 strain) in Fujian province, China, characterized as serovar 5, was used to evaluate the protective immunity against acute H. parasuis infection in piglets after inactivation. Following challenging with H. parasuis, only mild lesions in the pigs immunized with the killed vaccine were observed, whereas the typical symptoms of Glässer's disease presented in the nonimmunized piglets. A strong IgG immune response was induced by the inactive vaccine. CD4(+) and CD8(+) T lymphocyte levels were increased, indicating the potent cellular immune responses were elicited. The significantly high levels of IL-2, IL-4, TGF-ß, and IFN-γ in sera from pigs immunized with this killed vaccine suggested that the mixed Th1 and Th2 immune responses were induced, associated with the high protection against H. parasuis infection compared to the nonimmunized animals. This study indicated that the inactivated LY02 strain of H. parasuis could serve as a potential vaccine candidate to prevent the prevalence of H. parasuis in Fujian province, China.