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Evaluation of immune responses to porcine reproductive and respiratory syndrome virus in pigs during early stage of infection under farm conditions.

Dwivedi V, Manickam C, Binjawadagi B, Linhares D, Murtaugh MP, Renukaradhya GJ - Virol. J. (2012)

Bottom Line: Irrespective of regular vaccination, control of PRRSV remains a challenge to swine farmers.Replicating PRRSV in both infected and contact pigs was found to be responsible for rapid modulation in NK cell-meditated cytotoxicity and alteration in the production of important immune cytokines.PRRSV-induced immunological changes observed simultaneously at both cellular and cytokine levels early post-infection appear to be responsible for the delay in generation of adaptive immunity.

View Article: PubMed Central - HTML - PubMed

Affiliation: Food Animal Health Research Program, Ohio Agricultural Research and Development Center, Department of Veterinary Preventive Medicine, The Ohio State University, Wooster, 44691, USA.

ABSTRACT

Background: Porcine reproductive and respiratory syndrome virus (PRRSV) causes chronic, economically devastating disease in pigs of all ages. Frequent mutations in the viral genome result in viruses with immune escape mutants. Irrespective of regular vaccination, control of PRRSV remains a challenge to swine farmers. In PRRSV-infected pigs, innate cytokine IFN-α is inhibited and the adaptive arm of the immunity is delayed. To elucidate both cellular and innate cytokine responses at very early stages of PRRSV infection, seven weeks old pigs maintained on a commercial pig farm were infected and analyzed.

Results: One pig in a pen containing 25 pigs was PRRSV infected and responses from this pig and one penmate were assessed two days later. All the infected and a few of the contact neighbor pigs were viremic. At day 2 post-infection, approximately 50% of viremic pigs had greater than 50% reduction in NK cell-mediated cytotoxicity, and nearly a 1-fold increase in IFN-α production was detected in blood of a few pigs. Enhanced secretion of IL-4 (in ~90%), IL-12 (in ~40%), and IL-10 (in ~20%) (but not IFN-γ) in PRRSV infected pigs was observed. In addition, reduced frequency of myeloid cells, CD4(-)CD8(+) T cells, and CD4(+)CD8(+) T cells and upregulated frequency of lymphocytes bearing natural T regulatory cell phenotype were detected in viremic pigs. Interestingly, all viremic contact pigs also had comparable immune cell modulations.

Conclusion: Replicating PRRSV in both infected and contact pigs was found to be responsible for rapid modulation in NK cell-meditated cytotoxicity and alteration in the production of important immune cytokines. PRRSV-induced immunological changes observed simultaneously at both cellular and cytokine levels early post-infection appear to be responsible for the delay in generation of adaptive immunity. As the study was performed in pigs maintained under commercial environmental conditions, this study has practical implications in design of protective vaccines.

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Reduced NK cell-mediated cytotoxicity in PRRSV infected pigs. Percent NK-specific cytotoxicity was measured in pig PBMC (effectors). (A) Day 0 pre-infection and (B) day 2 post-infection against K-562 target cells. Effectors and targets at indicated E:T ratios were co-cultured and the supernatant harvested after 24 h was analyzed spectrophotometrically (OD490 nm) for released LDH from the lysed cells using a LDH substrate. Each line corresponds to NK cell-specific lysis from one pig at the four different E:T ratios. Statistical analysis was performed using a paired t-test by comparing the percent specific NK cell lysis between day 0 and day 2 post-infection at respective E:T ratios: P = 0.0115 at 1:100; P = 0.0001 at 1:50; P = 0.0007 at 1:25; P = 0.0069 at 1:12.5.
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Figure 2: Reduced NK cell-mediated cytotoxicity in PRRSV infected pigs. Percent NK-specific cytotoxicity was measured in pig PBMC (effectors). (A) Day 0 pre-infection and (B) day 2 post-infection against K-562 target cells. Effectors and targets at indicated E:T ratios were co-cultured and the supernatant harvested after 24 h was analyzed spectrophotometrically (OD490 nm) for released LDH from the lysed cells using a LDH substrate. Each line corresponds to NK cell-specific lysis from one pig at the four different E:T ratios. Statistical analysis was performed using a paired t-test by comparing the percent specific NK cell lysis between day 0 and day 2 post-infection at respective E:T ratios: P = 0.0115 at 1:100; P = 0.0001 at 1:50; P = 0.0007 at 1:25; P = 0.0069 at 1:12.5.

Mentions: Prior to PRRSV infection, NK cell cytotoxicity was analyzed by a colorimetric assay using peripheral blood mononuclear cells (PBMC) from all 50 pigs and appreciable NK cell cytotoxicity was detected in 13 infected pigs and 12 contact control pigs (Figure 2A). Two days after infection, 10 of the 13 NK cell competent PRRSV-infected pigs had more than 50% reduction in NK cell cytotoxicity, whereas only three of the 12 contact pigs had a similar reduction in NK cell cytotoxicity (Figure 2B). The reduction in NK cytolytic activity in the 13 NK cell competent infected pigs was statistically significant at tested effector cell: target cell (E:T) ratios compared to day 0. Flow cytometric analysis detected an increased frequency of NK cells rich fraction at day 2 post-infection in both infected and contact pigs (Table 1).


Evaluation of immune responses to porcine reproductive and respiratory syndrome virus in pigs during early stage of infection under farm conditions.

Dwivedi V, Manickam C, Binjawadagi B, Linhares D, Murtaugh MP, Renukaradhya GJ - Virol. J. (2012)

Reduced NK cell-mediated cytotoxicity in PRRSV infected pigs. Percent NK-specific cytotoxicity was measured in pig PBMC (effectors). (A) Day 0 pre-infection and (B) day 2 post-infection against K-562 target cells. Effectors and targets at indicated E:T ratios were co-cultured and the supernatant harvested after 24 h was analyzed spectrophotometrically (OD490 nm) for released LDH from the lysed cells using a LDH substrate. Each line corresponds to NK cell-specific lysis from one pig at the four different E:T ratios. Statistical analysis was performed using a paired t-test by comparing the percent specific NK cell lysis between day 0 and day 2 post-infection at respective E:T ratios: P = 0.0115 at 1:100; P = 0.0001 at 1:50; P = 0.0007 at 1:25; P = 0.0069 at 1:12.5.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3298799&req=5

Figure 2: Reduced NK cell-mediated cytotoxicity in PRRSV infected pigs. Percent NK-specific cytotoxicity was measured in pig PBMC (effectors). (A) Day 0 pre-infection and (B) day 2 post-infection against K-562 target cells. Effectors and targets at indicated E:T ratios were co-cultured and the supernatant harvested after 24 h was analyzed spectrophotometrically (OD490 nm) for released LDH from the lysed cells using a LDH substrate. Each line corresponds to NK cell-specific lysis from one pig at the four different E:T ratios. Statistical analysis was performed using a paired t-test by comparing the percent specific NK cell lysis between day 0 and day 2 post-infection at respective E:T ratios: P = 0.0115 at 1:100; P = 0.0001 at 1:50; P = 0.0007 at 1:25; P = 0.0069 at 1:12.5.
Mentions: Prior to PRRSV infection, NK cell cytotoxicity was analyzed by a colorimetric assay using peripheral blood mononuclear cells (PBMC) from all 50 pigs and appreciable NK cell cytotoxicity was detected in 13 infected pigs and 12 contact control pigs (Figure 2A). Two days after infection, 10 of the 13 NK cell competent PRRSV-infected pigs had more than 50% reduction in NK cell cytotoxicity, whereas only three of the 12 contact pigs had a similar reduction in NK cell cytotoxicity (Figure 2B). The reduction in NK cytolytic activity in the 13 NK cell competent infected pigs was statistically significant at tested effector cell: target cell (E:T) ratios compared to day 0. Flow cytometric analysis detected an increased frequency of NK cells rich fraction at day 2 post-infection in both infected and contact pigs (Table 1).

Bottom Line: Irrespective of regular vaccination, control of PRRSV remains a challenge to swine farmers.Replicating PRRSV in both infected and contact pigs was found to be responsible for rapid modulation in NK cell-meditated cytotoxicity and alteration in the production of important immune cytokines.PRRSV-induced immunological changes observed simultaneously at both cellular and cytokine levels early post-infection appear to be responsible for the delay in generation of adaptive immunity.

View Article: PubMed Central - HTML - PubMed

Affiliation: Food Animal Health Research Program, Ohio Agricultural Research and Development Center, Department of Veterinary Preventive Medicine, The Ohio State University, Wooster, 44691, USA.

ABSTRACT

Background: Porcine reproductive and respiratory syndrome virus (PRRSV) causes chronic, economically devastating disease in pigs of all ages. Frequent mutations in the viral genome result in viruses with immune escape mutants. Irrespective of regular vaccination, control of PRRSV remains a challenge to swine farmers. In PRRSV-infected pigs, innate cytokine IFN-α is inhibited and the adaptive arm of the immunity is delayed. To elucidate both cellular and innate cytokine responses at very early stages of PRRSV infection, seven weeks old pigs maintained on a commercial pig farm were infected and analyzed.

Results: One pig in a pen containing 25 pigs was PRRSV infected and responses from this pig and one penmate were assessed two days later. All the infected and a few of the contact neighbor pigs were viremic. At day 2 post-infection, approximately 50% of viremic pigs had greater than 50% reduction in NK cell-mediated cytotoxicity, and nearly a 1-fold increase in IFN-α production was detected in blood of a few pigs. Enhanced secretion of IL-4 (in ~90%), IL-12 (in ~40%), and IL-10 (in ~20%) (but not IFN-γ) in PRRSV infected pigs was observed. In addition, reduced frequency of myeloid cells, CD4(-)CD8(+) T cells, and CD4(+)CD8(+) T cells and upregulated frequency of lymphocytes bearing natural T regulatory cell phenotype were detected in viremic pigs. Interestingly, all viremic contact pigs also had comparable immune cell modulations.

Conclusion: Replicating PRRSV in both infected and contact pigs was found to be responsible for rapid modulation in NK cell-meditated cytotoxicity and alteration in the production of important immune cytokines. PRRSV-induced immunological changes observed simultaneously at both cellular and cytokine levels early post-infection appear to be responsible for the delay in generation of adaptive immunity. As the study was performed in pigs maintained under commercial environmental conditions, this study has practical implications in design of protective vaccines.

Show MeSH
Related in: MedlinePlus