• Journal of Veterinary Science
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• v.22 no.6
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• pp.87.1-87.12
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• 2021

Background: African swine fever virus (ASFV), classical swine fever virus (CSFV), and porcine reproductive and respiratory syndrome virus (PRRSV) are still prevalent in many regions of China. Co-infections make it difficult to distinguish their clinical symptoms and pathological changes. Therefore, a rapid and specific method is needed for the differential detection of these pathogens. Objectives: The aim of this study was to develop a multiplex real-time quantitative reverse transcription polymerase chain reaction (multiplex qRT-PCR) for the simultaneous differential detection of ASFV, CSFV, and PRRSV. Methods: Three pairs of primers and TaqMan probes targeting the ASFV p72 gene, CSFV 5' untranslated region, and PRRSV ORF7 gene were designed. After optimizing the reaction conditions, including the annealing temperature, primer concentration, and probe concentration, multiplex qRT-PCR for simultaneous and differential detection of ASFV, CSFV, and PRRSV was developed. Subsequently, 1,143 clinical samples were detected to verify the practicality of the assay. Results: The multiplex qRT-PCR assay could specifically and simultaneously detect the ASFV, CSFV, and PRRSV with a detection limit of 1.78 × 10⁰ copies for the ASFV, CSFV, and PRRSV, but could not amplify the other major porcine viruses, such as pseudorabies virus, porcine circovirus type 1 (PCV1), PCV2, PCV3, foot-and-mouth disease virus, porcine parvovirus, atypical porcine pestivirus, and Senecavirus A. The assay had good repeatability with coefficients of variation of intra- and inter-assay of less than 1.2%. Finally, the assay was used to detect 1,143 clinical samples to evaluate its practicality in the field. The positive rates of ASFV, CSFV, and PRRSV were 25.63%, 9.36%, and 17.50%, respectively. The co-infection rates of ASFV+CSFV, ASFV+PRRSV, CSFV+PRRSV, and ASFV+CSFV+PRRSV were 2.45%, 2.36%, 1.57%, and 0.17%, respectively. Conclusions: The multiplex qRT-PCR developed in this study could provide a rapid, sensitive, specific diagnostic tool for the simultaneous and differential detection of ASFV, CSFV, and PRRSV.

• Korean Journal of Veterinary Research
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• v.48 no.3
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• pp.267-274
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• 2008

This study was to evaluate the efficacy of single dose Mycoplasma hyopneumoniae (M. hyo)-vaccination in the swine farms which had different serological patterns of porcine reproductive and respiratory syndrome virus (PRRSV) and porcine circovirus type 2 (PCV2). A minimum of 240 pigs from each farm was applied, allocating M. hyo vaccinated and control groups. The PRRSV and PCV2 infections were analyzed by serological method (commercial ELISA kit). After administrating pigs a single dose of M. hyo vaccine or control saline at 3 weeks of age, serum antibodies to M. hyo, PRRSV and PCV2 were monitored at 4, 10, 16 and 22 weeks of age. Mortality, weight changes, feed conversion ratio (FCR) and lung score were also evaluated. A single-dose vaccination of M. hyo bacterin was efficacious to reduce mycoplasmal lung lesions and induce good humoral immune response. However, FCR was improved only in one of the three farms where showed seronegative status to both PRRSV and PCV2 in the period from 4 to 16 weeks of age. These results might imply that M. hyo vaccine alone could not overcome the PRRSV and PCV2 infection-associated wasting in the field condition. Therefore, the control of PRRSV and PCV2 should be considered to obtain the better effects of M. hyo vaccination.

• Korean Journal of Veterinary Service
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• v.45 no.1
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• pp.63-69
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• 2022

Proliferative and necrotizing pneumonia (PNP) is a form of interstitial pneumonia that occurs in post-weaning pigs. In this study, we investigated the presence of swine influenza virus (SIV), porcine reproductive and respiratory syndrome virus (PRRSV), porcine circovirus type 2 (PCV2) and Aujeszky's disease virus (ADV) in PNP lesions in Jeju pigs. Based on the histopathologic criteria for PNP, a total of 50 cases were selected in Jeju pigs between 2008 and 2010. Coupled with histopathological examinations, the presence of ADV and SIV by polymerase chain reaction (PCR) and reverse transcription PCR (RT-PCR) and PRRSV and PCV2 by immunohistochemical (IHC) methods were investigated. Based on the PCR and RT-PCR methods, ADV and SIV nucleic acids were not detected in all cases. According to IHC, PRRSV was detected in 38 of the 50 cases examined (76%) and PCV2 in 25 cases (50%). PRRSV or PCV2 were detected in 19 (38%) or 6 (12%) cases, respectively. Both PRRSV and PCV2 were identified in other 19 cases (38%). Antigens of PRRSV and PCV2 were commonly observed in the cytoplasm of macrophages and clusters of necrotic cells in alveolar cavities. The results of the present study demonstrate that PRRSV is predominantly associated with PNP in Jeju pigs. Co-infection with PRRSV and PCV2 may enhance the severity of PNP lesions in affected pigs.

• Journal of Veterinary Clinics
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• v.28 no.4
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• pp.415-421
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• 2011

Currently, various diseases reside in Korean swine farms and affect production performance of the farms greatly. These damages from disease are further aggravated by the concurrent infection of other disease. In this study, y investigating the distribution of porcine reproductive and respiratory syndrome virus (PRRSV), porcine circovirus type 2 (PCV2), Salmonella spp. in farms, correlation between the damage and the prevalence of disease was analyzed. Ten selected Korean swine farms that uses PCV2 vaccine were tested for presence of antibody and antigen of PRRSV, PCV2, Salmonella spp. per ages of pigs, 4weeks, 7weeks, 11weeks and 17weeks, respectively. The results were analyzed by dividing the farms in to groups with MSY above 19, and that with MSY below 19. Then calculating the distribution of disease each ages of pigs. Farms with MSY below 19 showed high prevalence of disease by PRRSV, PCV2 and Salmonella spp.. In this group, the detection rate of PCV2 and Salmonella spp. was increased by the activation/viremia of PRRSV in the young ages of pigs. The results are proved that the correlation between disease prevalence and production performance in Korean swine farms were very significant. The prevalence of PRRSV is more important index which influence to the productivity in current prevalence of diseases.

• Korean Journal of Veterinary Research
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• v.38 no.2
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• pp.314-318
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• 1998

An establishment of effective control measures to PRRSV infection in swine industry depends on a sensitive and specific diagnosis to detect either viral antigen and/or antibodies to PRRSV. Several diagnostic methods are available to detect antibodies against PRRSV, including IPMA, IFA and ELISA tests have been successfully developed. Sensitivity of the indirect immunofluorescent assay in MA-104 cells using Korean field isolate PL96-1 was superior to that of VR-2332 and field isolate PL96-2. Sensitivity and specificity of the IFA test with PL96-1 were comparable to those of commercial ELISA test kit but ELISA test was more sensitive for the detection of declining antibodies to PRRSV in finishing pigs. In this study we concluded that IFA and ELISA test could be utilized to detect antibodies to PRRSV and the results generated from these two tests were comparable and there were no significant difference between these two tests.

• Korean Journal of Veterinary Research
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• v.53 no.4
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• pp.245-251
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• 2013

Porcine circovirus type 2 (PCV2) and porcine reproductive and respiratory syndrome virus (PRRSV) have been suspected to have immunosuppressive effects on pigs. To investigate the correlation between these virus infection and the lesions of lymph nodes including sub-mandibular and inguinal lymph node, 44 pigs (PCV2 single, n = 14; PRRSV single, n = 10; PCV2/PRRSV, n = 14; negative control, n = 6) were examined by histopathology and immunohistochemistry. Histopathologically, granulomatous lymphadenitis characterized by lymphoid depletion with histiocytic cells infiltration was observed in PCV-2 single and PCV-2/PRRSV group. Immunohistochemically, there were significant reduction of B and T lymphocytes in lymph nodes of these groups, while the number of macrophages was increased. In only PRRSV infected group, germinal center hypertrophy and lymphoid necrosis were observed. Immunohistochemically, the number of CD3+ T lymphocytes was slightly increased. Severe lymphocytic depletion in PCV-2 infection-related lymph nodes might be associated with producing immunocompromised state in pig. Comparing with PCV-2 infected group, PRRSV produced minor effects on the changes in immune cell population in the lymph nodes of pigs. PRRSV may increase susceptibility of the disease in pigs by disruption of the first defense lines in target organs, such as the alveolar macrophages in lungs.

• Korean Journal of Veterinary Service
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• v.37 no.1
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• pp.11-18
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• 2014

Porcine reproductive and respiratory syndrome (PRRS) causes a significant economic loss in the swine industry not only in Korea but also all over the world. Andong and Hapcheon region were selected for Area Regional Control (ARC) programme to reduce the shedding of PRRS virus (PRRSV) and decrease PRRS outbreaks. Before conducting the PRRS ARC, sera of pigs were tested for both antibody using ELISA and antigen using RT-PCR, then phylogenetic classifications was analysed. Pigs of 138/275 (50.2%) in Andong and 352/425 (82.8%) in Hapcheon were seropositive. Also, the RT-PCR results revealed that 27 heads (8.2%) in Andong, 112 heads (22.0%) in Hapcheon were positive for PRRSV antigen. PRRSVs were mainly detected between the ages of 40 to 60 days. PRRSV ORF5 regions were used to determine genetic clusters based on previous report. All PRRSV type I detected in both Andong and Hapcheon were classified as Cluster I. The PRRSV type II isolates in Andong were assorted to Cluster II, whereas the PRRSV type II isolates in Hapcheon were the viruses were unassembled into any cluster except one identified to Cluster III. Phylogenetic analysis indicated that new clusters of PRRSVs type II were prevalent in Hapcheon.

• Korean Journal of Veterinary Service
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• v.37 no.1
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• pp.1-9
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• 2014

Although it has been generally accepted that porcine reproductive and respiratory syndrome virus (PRRSV) induces weak and delayed protective immunity after infection, it is unclear that the same immunological features can be applicable to all PRRS viruses because huge genetic variation exists even among the same genotypes of PRRSV (Type 1 and 2). In the current study, two genetically distinct type 2 PRRSV strains (VR-2332 and JA142) which showed approximately 90% nucleotide homology based on ORF5 sequences were characterized by both in vitro and in vivo assessments to determine the immunological features of the viruses. For in vitro assessment, porcine alveolar macrophages (PAM) were infected with the viruses at $10^{-3}$ multiplicity of infection (MOI) and then supernatants and cells were collected separately at 36 hrs post infection to determine the relative expression levels of IL-$1{\alpha}$, IL-12, TNF-${\alpha}$ and INF-${\alpha}/{\beta}$ by quantitative RT-PCR. In addition, five PRRSV-free pigs were inoculated with either of JA142 or VR2332 for in vivo assessment. Serum samples were collected every week until 6 weeks post challenge. The serum samples were analyzed for the levels of viremia, PRRSV nucleocapsid-specific antibody and virus neutralizing antibody. Based on those assessments, the two viruses showed different patterns of cytokine expression in PAM and immune responses in pigs after infection. These results indicate that genetically distinct PRRSV strains have different immunological features, which might be criteria for virus classification and selection of candidate virus strains for vaccine development in the future.

• Asian-Australasian Journal of Animal Sciences
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• v.17 no.10
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• pp.1438-1445
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• 2004

The objective of this study was to determine the effects of dietary Zn level on performance, serum Zn concentrations, alkaline phosphatase activity (ALP), and immune response of pigs inoculated with Porcine Reproductive and Respiratory Syndrome virus (PRRSv) and Mycoplasma hyopneumoniae. A $2{\times}4$ factorial arrangement of treatments was used in a randomized design. Factors included; 1) PRRSv and M. hyopneumoniae inoculation (n=36 pigs) or sham inoculation (n=36 pigs) with media when pigs entered the grower facility (d 0) at 9 weeks of age and 2) 10, 50, 150 ppm supplemental Zn sulfate (${ZnSO}_4$) from weaning until the completion of the study, or 2,000 ppm supplemental ${ZnSO}_4$for two weeks in the nursery and then supplementation with 150 ppm ${ZnSO}_4$for the remainder of the trial. The basal diet contained 34 ppm Zn. Pigs were weighed on d 0, 10, 17, 24 and 31 and blood samples were collected on d 0, 7, 14, 21 and 28. Pigs inoculated with PRRSv were serologically positive at d 28 and control pigs remained negative to PRRSv. In contrast, the M hyopneumoniae inoculation was inconsistent with 33.3% and 52.8% of pigs serologically positive at d 28 in the control and infected groups, respectively. A febrile response was observed for approximately one week after inoculation with PRRSv. Feed intake (p<0.01) and gain (p<0.1) were less in PRRSv infected pigs than control pigs for the 31 d study. However, performance did not differ among pigs in the four levels of ${ZnSO}_4$. Assessments of immune responses failed to provide unequivocal influence of either PRRSv inoculation or ${ZnSO}_4$level. These data suggest that PRRSv and M. hyopneumoniae act to produce some performance deficits and the influence of Zn supplementation of nursery age pigs does not have clear effect in grower pigs affected with disease.

• Microbiology and Biotechnology Letters
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• v.43 no.1
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• pp.1-8
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• 2015

In this study, a DNA-launched reverse genetics system was developed from a type 2 porcine reproductive and respiratory syndrome virus (PRRSV) strain, KNU-12. The complete genome of 15,412 nucleotides was assembled as a single cDNA clone and placed under the eukaryotic CMV promoter. Upon transfection of BHK-tailless pCD163 cells with a full-length cDNA clone, viable and infectious type 2 progeny PRRSV were rescued. The reconstituted virus was found to maintain growth properties similar to those of the parental virus in porcine alveolar macrophage (PAM) cells. With the availability of this type 2 PRRSV infectious clone, we first explored the biological relevance of ORF5a in the PRRSV replication cycle. Therefore, we used a PRRSV reverse genetics system to generate an ORF5a knockout mutant clone by changing the ORF5a translation start codon and introducing a stop codon at the 7^th codon of ORF5a. The ORF5a knockout mutant was found to exhibit a lack of infectivity in both BHK-tailless pCD163 and PAM-pCD163 cells, suggesting that inactivation of ORF5a expression is lethal for infectious virus production. In order to restore the ORF5a gene-deleted PRRSV, complementing cell lines were established to stably express the ORF5a protein of PRRSV. ORF5a-expressing cells were capable of supporting the production of the replicationdefective virus, indicating complementation of the impaired ORF5a gene function of PRRSV in trans.