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Toll-like receptor responses to Peste des petits ruminants virus in goats and water buffalo.

Dhanasekaran S, Biswas M, Vignesh AR, Ramya R, Raj GD, Tirumurugaan KG, Raja A, Kataria RS, Parida S, Elankumaran S, Subbiah E - PLoS ONE (2014)

Bottom Line: Upon stimulation of PBMC with synthetic TLR3 and TLR7 agonists or PPRV, the levels of proinflammatory cytokines were found to be significantly higher while immunosuppressive interleukin (IL) 10 levels were lower in PPRV resistant Kanni and Salem Black breeds and water buffalo at transcriptional level, correlating with reduced viralloads in infected PBMC.Pre-treatment of Vero cells with human IFNα resulted in reduction of PPRV replication, confirming the role of IFNα in limiting PPRV replication.Treatment with IRS66, a TLR7 antagonist, resulted in the reduction of IFNα levels, with increased PPRV replication confirming the role of TLR7.

View Article: PubMed Central - PubMed

Affiliation: Department of Animal Biotechnology, Madras Veterinary College, Tamil Nadu Veterinary and Animal Sciences University, Chennai, Tamil Nadu, India.

ABSTRACT
Ovine rinderpest or goat plague is an economically important and contagious viral disease of sheep and goats, caused by the Peste des petits ruminants virus (PPRV). Differences in susceptibility to goat plague among different breeds and water buffalo exist. The host innate immune system discriminates between pathogen associated molecular patterns and self antigens through surveillance receptors known as Toll like receptors (TLR). We investigated the role of TLR and cytokines in differential susceptibility of goat breeds and water buffalo to PPRV. We examined the replication of PPRV in peripheral blood mononuclear cells (PBMC) of Indian domestic goats and water buffalo and demonstrated that the levels of TLR3 and TLR7 and downstream signalling molecules correlation with susceptibility vs resistance. Naturally susceptible goat breeds, Barbari and Tellichery, had dampened innate immune responses to PPRV and increased viral loads with lower basal expression levels of TLR 3/7. Upon stimulation of PBMC with synthetic TLR3 and TLR7 agonists or PPRV, the levels of proinflammatory cytokines were found to be significantly higher while immunosuppressive interleukin (IL) 10 levels were lower in PPRV resistant Kanni and Salem Black breeds and water buffalo at transcriptional level, correlating with reduced viralloads in infected PBMC. Water buffalo produced higher levels of interferon (IFN) α in comparison with goats at transcriptional and translational levels. Pre-treatment of Vero cells with human IFNα resulted in reduction of PPRV replication, confirming the role of IFNα in limiting PPRV replication. Treatment with IRS66, a TLR7 antagonist, resulted in the reduction of IFNα levels, with increased PPRV replication confirming the role of TLR7. Single nucleotide polymorphism analysis of TLR7 of these goat breeds did not show any marked nucleotide differences that might account for susceptibility vs resistance to PPRV. Analyzing other host genetic factors might provide further insights on susceptibility to PPRV and genetic polymorphisms in the host.

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Virus replication in PBMCs stimulated with TLR3 and TLR7 agonists.a) Reduction in TCID50 values of PPRV in goat PBMC on imiquimod treatment. Reduction in PPRV H gene expression levels in goat PBMC on treatment with b) poly I:C and c) imiquimod. Bars with the same superscript do not differ significantly. Significance is indicated when p<0.05. Values represent mean ± SD of 40-corrected CT in 5 individual animals per goat breed. Significantly higher PPRV viral loads observed in PBMC of Barbari and Tellicherry breeds as compared to Kanni and Salem Black. Significant reduction in PPRV levels on poly I:C and imiquimod treatment in all breeds.
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pone-0111609-g002: Virus replication in PBMCs stimulated with TLR3 and TLR7 agonists.a) Reduction in TCID50 values of PPRV in goat PBMC on imiquimod treatment. Reduction in PPRV H gene expression levels in goat PBMC on treatment with b) poly I:C and c) imiquimod. Bars with the same superscript do not differ significantly. Significance is indicated when p<0.05. Values represent mean ± SD of 40-corrected CT in 5 individual animals per goat breed. Significantly higher PPRV viral loads observed in PBMC of Barbari and Tellicherry breeds as compared to Kanni and Salem Black. Significant reduction in PPRV levels on poly I:C and imiquimod treatment in all breeds.

Mentions: We examined the relative basal expression levels of TLR3 and TLR7 mRNA in naive PBMC by quantitative real-time RT-PCR (qRT-PCR) in nine individual animals per breed (n = 9). The Kanni and Salem Black breeds revealed significantly higher basal TLR3 (p<0.001) and TLR7 (p<0.001) transcripts than Barbari goats while there were no significant difference (p>0.05) between Tellicherry and Barbari breeds (Figure 1). To understand their contribution to virus replication, PBMC from each of these four goat breeds (n = 5) were infected with 1×103.0 mean tissue culture infective dose (TCID50) of PPRV and the virus load analyzed at 24 h post infection (PI) by qRT-PCR, using primers specific to the PPRV-H gene and TCID50. PBMC from Barbari and Tellicherry goats supported significantly (p<0.01) higher PPRV replication than those from Kanni and Salem Black (Figure 2A) with the yields being similar in these two breeds. There was a significant reduction in virus yield by one log10 in Kanni/Salem Black PBMC (p<0.05) compared to Barbari/Tellicherry goats (Figure 2A).


Toll-like receptor responses to Peste des petits ruminants virus in goats and water buffalo.

Dhanasekaran S, Biswas M, Vignesh AR, Ramya R, Raj GD, Tirumurugaan KG, Raja A, Kataria RS, Parida S, Elankumaran S, Subbiah E - PLoS ONE (2014)

Virus replication in PBMCs stimulated with TLR3 and TLR7 agonists.a) Reduction in TCID50 values of PPRV in goat PBMC on imiquimod treatment. Reduction in PPRV H gene expression levels in goat PBMC on treatment with b) poly I:C and c) imiquimod. Bars with the same superscript do not differ significantly. Significance is indicated when p<0.05. Values represent mean ± SD of 40-corrected CT in 5 individual animals per goat breed. Significantly higher PPRV viral loads observed in PBMC of Barbari and Tellicherry breeds as compared to Kanni and Salem Black. Significant reduction in PPRV levels on poly I:C and imiquimod treatment in all breeds.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4219731&req=5

pone-0111609-g002: Virus replication in PBMCs stimulated with TLR3 and TLR7 agonists.a) Reduction in TCID50 values of PPRV in goat PBMC on imiquimod treatment. Reduction in PPRV H gene expression levels in goat PBMC on treatment with b) poly I:C and c) imiquimod. Bars with the same superscript do not differ significantly. Significance is indicated when p<0.05. Values represent mean ± SD of 40-corrected CT in 5 individual animals per goat breed. Significantly higher PPRV viral loads observed in PBMC of Barbari and Tellicherry breeds as compared to Kanni and Salem Black. Significant reduction in PPRV levels on poly I:C and imiquimod treatment in all breeds.
Mentions: We examined the relative basal expression levels of TLR3 and TLR7 mRNA in naive PBMC by quantitative real-time RT-PCR (qRT-PCR) in nine individual animals per breed (n = 9). The Kanni and Salem Black breeds revealed significantly higher basal TLR3 (p<0.001) and TLR7 (p<0.001) transcripts than Barbari goats while there were no significant difference (p>0.05) between Tellicherry and Barbari breeds (Figure 1). To understand their contribution to virus replication, PBMC from each of these four goat breeds (n = 5) were infected with 1×103.0 mean tissue culture infective dose (TCID50) of PPRV and the virus load analyzed at 24 h post infection (PI) by qRT-PCR, using primers specific to the PPRV-H gene and TCID50. PBMC from Barbari and Tellicherry goats supported significantly (p<0.01) higher PPRV replication than those from Kanni and Salem Black (Figure 2A) with the yields being similar in these two breeds. There was a significant reduction in virus yield by one log10 in Kanni/Salem Black PBMC (p<0.05) compared to Barbari/Tellicherry goats (Figure 2A).

Bottom Line: Upon stimulation of PBMC with synthetic TLR3 and TLR7 agonists or PPRV, the levels of proinflammatory cytokines were found to be significantly higher while immunosuppressive interleukin (IL) 10 levels were lower in PPRV resistant Kanni and Salem Black breeds and water buffalo at transcriptional level, correlating with reduced viralloads in infected PBMC.Pre-treatment of Vero cells with human IFNα resulted in reduction of PPRV replication, confirming the role of IFNα in limiting PPRV replication.Treatment with IRS66, a TLR7 antagonist, resulted in the reduction of IFNα levels, with increased PPRV replication confirming the role of TLR7.

View Article: PubMed Central - PubMed

Affiliation: Department of Animal Biotechnology, Madras Veterinary College, Tamil Nadu Veterinary and Animal Sciences University, Chennai, Tamil Nadu, India.

ABSTRACT
Ovine rinderpest or goat plague is an economically important and contagious viral disease of sheep and goats, caused by the Peste des petits ruminants virus (PPRV). Differences in susceptibility to goat plague among different breeds and water buffalo exist. The host innate immune system discriminates between pathogen associated molecular patterns and self antigens through surveillance receptors known as Toll like receptors (TLR). We investigated the role of TLR and cytokines in differential susceptibility of goat breeds and water buffalo to PPRV. We examined the replication of PPRV in peripheral blood mononuclear cells (PBMC) of Indian domestic goats and water buffalo and demonstrated that the levels of TLR3 and TLR7 and downstream signalling molecules correlation with susceptibility vs resistance. Naturally susceptible goat breeds, Barbari and Tellichery, had dampened innate immune responses to PPRV and increased viral loads with lower basal expression levels of TLR 3/7. Upon stimulation of PBMC with synthetic TLR3 and TLR7 agonists or PPRV, the levels of proinflammatory cytokines were found to be significantly higher while immunosuppressive interleukin (IL) 10 levels were lower in PPRV resistant Kanni and Salem Black breeds and water buffalo at transcriptional level, correlating with reduced viralloads in infected PBMC. Water buffalo produced higher levels of interferon (IFN) α in comparison with goats at transcriptional and translational levels. Pre-treatment of Vero cells with human IFNα resulted in reduction of PPRV replication, confirming the role of IFNα in limiting PPRV replication. Treatment with IRS66, a TLR7 antagonist, resulted in the reduction of IFNα levels, with increased PPRV replication confirming the role of TLR7. Single nucleotide polymorphism analysis of TLR7 of these goat breeds did not show any marked nucleotide differences that might account for susceptibility vs resistance to PPRV. Analyzing other host genetic factors might provide further insights on susceptibility to PPRV and genetic polymorphisms in the host.

Show MeSH
Related in: MedlinePlus