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In Vitro Coinfection and Replication of Classical Swine Fever Virus and Porcine Circovirus Type 2 in PK15 Cells.

Zhou N, Xing G, Zhou J, Jin Y, Liang C, Gu J, Hu B, Liao M, Wang Q, Zhou J - PLoS ONE (2015)

Bottom Line: However, CSFV reproduction decreased in a PCV2 dose-dependent manner.In addition, cellular apoptosis was not strengthened in PK15-CSFV cells infected with PCV2 in comparison with PCV2-infected PK15 cells.Moreover, using this coinfection model we further demonstrated PCV2-induced apoptosis might contribute to the impairment of CSFV HCLV strain replication in coinfected cells.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Animal Virology of Ministry of Agriculture, Zhejiang University, Hangzhou, PR China.

ABSTRACT
Increasing clinical lines of evidence have shown the coinfection/superinfection of porcine circovirus type 2 (PCV2) and classical swine fever virus (CSFV). Here, we investigated whether PCV2 and CSFV could infect the same cell productively by constructing an in vitro coinfection model. Our results indicated that PCV2-free PK15 cells but not ST cells were more sensitive to PCV2, and the PK15 cell line could stably harbor replicating CSFV (PK15-CSFV cells) with a high infection rate. Confocal and super-resolution microscopic analysis showed that PCV2 and CSFV colocalized in the same PK15-CSFV cell, and the CSFV E2 protein translocated from the cytoplasm to the nucleus in PK15-CSFV cells infected with PCV2. Moreover, PCV2-CSFV dual-positive cells increased gradually in PK15-CSFV cells in a PCV2 dose-dependent manner. In PK15-CSFV cells, PCV2 replicated well, and the production of PCV2 progeny was not influenced by CSFV infection. However, CSFV reproduction decreased in a PCV2 dose-dependent manner. In addition, cellular apoptosis was not strengthened in PK15-CSFV cells infected with PCV2 in comparison with PCV2-infected PK15 cells. Moreover, using this coinfection model we further demonstrated PCV2-induced apoptosis might contribute to the impairment of CSFV HCLV strain replication in coinfected cells. Taken together, our results demonstrate for the first time the coinfection/superinfection of PCV2 and CSFV within the same cell, providing an in vitro model to facilitate further investigation of the underlying mechanism of CSFV and PCV2 coinfection.

No MeSH data available.


Related in: MedlinePlus

PCV2 but not PCV2 genome or PCV2-encoded components affects CSFV replication in PK15-CSFV cells.Cells were inoculated with PCV2 at MOI of 1 or 10 and pretreated with inactivated PCV2, genomic DNA of PCV2 or PCV2-encoded components. (A and C) Percentage of TUNEL-positive cells. (B and D) Titration of CSFV progeny at 1 (B) and 10 (B) MOI of BPL-inactivated PCV2. Data are represented as means ± SD (n = 3 or more; ns, P > 0.05; *P < 0.05; **P < 0.01).
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pone.0139457.g008: PCV2 but not PCV2 genome or PCV2-encoded components affects CSFV replication in PK15-CSFV cells.Cells were inoculated with PCV2 at MOI of 1 or 10 and pretreated with inactivated PCV2, genomic DNA of PCV2 or PCV2-encoded components. (A and C) Percentage of TUNEL-positive cells. (B and D) Titration of CSFV progeny at 1 (B) and 10 (B) MOI of BPL-inactivated PCV2. Data are represented as means ± SD (n = 3 or more; ns, P > 0.05; *P < 0.05; **P < 0.01).

Mentions: To investigate whether PCV2-induced apoptosis involves viral components of PCV2, cells were treated with His-Rep, dCap, His-ORF3, His-ORF4 or genomic DNA of PCV2 and β-propiolactone (BPL)-inactivated PCV2, then the TUNEL assay and titration of CSFV were carried out. Data shown in Fig 8A and 8B revealed that none of the viral components of PCV2 tested caused apoptosis in PK15 or PK15-CSFV cells and did not affect CSFV replication. Similarly, PK15 and PK15-CSFV cells infected with the inactivated PCV2 could not further induce apoptosis and did not decrease the yield of CSFV in PK15-CSFV cells (MOI = 1 and 10) (Fig 8C and 8D). The results further confirmed that PCV2 replication but not viral components of PCV2 induced the apoptosis of PK15 and PK15-CSFV cells, and that the viral components of PCV2 did not interfere with the production of CSFV progeny.


In Vitro Coinfection and Replication of Classical Swine Fever Virus and Porcine Circovirus Type 2 in PK15 Cells.

Zhou N, Xing G, Zhou J, Jin Y, Liang C, Gu J, Hu B, Liao M, Wang Q, Zhou J - PLoS ONE (2015)

PCV2 but not PCV2 genome or PCV2-encoded components affects CSFV replication in PK15-CSFV cells.Cells were inoculated with PCV2 at MOI of 1 or 10 and pretreated with inactivated PCV2, genomic DNA of PCV2 or PCV2-encoded components. (A and C) Percentage of TUNEL-positive cells. (B and D) Titration of CSFV progeny at 1 (B) and 10 (B) MOI of BPL-inactivated PCV2. Data are represented as means ± SD (n = 3 or more; ns, P > 0.05; *P < 0.05; **P < 0.01).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4592061&req=5

pone.0139457.g008: PCV2 but not PCV2 genome or PCV2-encoded components affects CSFV replication in PK15-CSFV cells.Cells were inoculated with PCV2 at MOI of 1 or 10 and pretreated with inactivated PCV2, genomic DNA of PCV2 or PCV2-encoded components. (A and C) Percentage of TUNEL-positive cells. (B and D) Titration of CSFV progeny at 1 (B) and 10 (B) MOI of BPL-inactivated PCV2. Data are represented as means ± SD (n = 3 or more; ns, P > 0.05; *P < 0.05; **P < 0.01).
Mentions: To investigate whether PCV2-induced apoptosis involves viral components of PCV2, cells were treated with His-Rep, dCap, His-ORF3, His-ORF4 or genomic DNA of PCV2 and β-propiolactone (BPL)-inactivated PCV2, then the TUNEL assay and titration of CSFV were carried out. Data shown in Fig 8A and 8B revealed that none of the viral components of PCV2 tested caused apoptosis in PK15 or PK15-CSFV cells and did not affect CSFV replication. Similarly, PK15 and PK15-CSFV cells infected with the inactivated PCV2 could not further induce apoptosis and did not decrease the yield of CSFV in PK15-CSFV cells (MOI = 1 and 10) (Fig 8C and 8D). The results further confirmed that PCV2 replication but not viral components of PCV2 induced the apoptosis of PK15 and PK15-CSFV cells, and that the viral components of PCV2 did not interfere with the production of CSFV progeny.

Bottom Line: However, CSFV reproduction decreased in a PCV2 dose-dependent manner.In addition, cellular apoptosis was not strengthened in PK15-CSFV cells infected with PCV2 in comparison with PCV2-infected PK15 cells.Moreover, using this coinfection model we further demonstrated PCV2-induced apoptosis might contribute to the impairment of CSFV HCLV strain replication in coinfected cells.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Animal Virology of Ministry of Agriculture, Zhejiang University, Hangzhou, PR China.

ABSTRACT
Increasing clinical lines of evidence have shown the coinfection/superinfection of porcine circovirus type 2 (PCV2) and classical swine fever virus (CSFV). Here, we investigated whether PCV2 and CSFV could infect the same cell productively by constructing an in vitro coinfection model. Our results indicated that PCV2-free PK15 cells but not ST cells were more sensitive to PCV2, and the PK15 cell line could stably harbor replicating CSFV (PK15-CSFV cells) with a high infection rate. Confocal and super-resolution microscopic analysis showed that PCV2 and CSFV colocalized in the same PK15-CSFV cell, and the CSFV E2 protein translocated from the cytoplasm to the nucleus in PK15-CSFV cells infected with PCV2. Moreover, PCV2-CSFV dual-positive cells increased gradually in PK15-CSFV cells in a PCV2 dose-dependent manner. In PK15-CSFV cells, PCV2 replicated well, and the production of PCV2 progeny was not influenced by CSFV infection. However, CSFV reproduction decreased in a PCV2 dose-dependent manner. In addition, cellular apoptosis was not strengthened in PK15-CSFV cells infected with PCV2 in comparison with PCV2-infected PK15 cells. Moreover, using this coinfection model we further demonstrated PCV2-induced apoptosis might contribute to the impairment of CSFV HCLV strain replication in coinfected cells. Taken together, our results demonstrate for the first time the coinfection/superinfection of PCV2 and CSFV within the same cell, providing an in vitro model to facilitate further investigation of the underlying mechanism of CSFV and PCV2 coinfection.

No MeSH data available.


Related in: MedlinePlus