Ferreira Barbosa Jérémy A, Labrie Josée, Beaudry Francis, Gagnon Carl A, Jacques Mario
Centre de recherche en infectiologie porcine et avicole (CRIPA), Faculté de médecine vétérinaire, Université de Montréal, St-Hyacinthe, Québec, Canada.
Groupe de recherche sur les maladies infectieuses du porc (GREMIP), Faculté de médecine vétérinaire, Université de Montréal, St-Hyacinthe, Québec, Canada.
Virol J. 2015 Nov 14;12:188. doi: 10.1186/s12985-015-0404-3.
Porcine reproductive and respiratory syndrome virus (PRRSV) is one of the most important pathogens in the swine industry and causes important economic losses. No effective antiviral drugs against it are commercially available. We recently reported that the culture supernatant of Actinobacillus pleuropneumoniae, the porcine pleuropneumonia causative agent, has an antiviral activity in vitro against PRRSV in SJPL cells. Objectives of this study were (i) to identify the mechanism behind the antiviral activity displayed by A. pleuropneumoniae and (ii) to characterize the active molecules present in the bacterial culture supernatant.
Antibody microarray analysis was used in order to point out cellular pathways modulated by the A. pleuropneumoniae supernatant. Subsequent, flow cytometry analysis and cell cycle inhibitors were used to confirm antibody microarray data and to link them to the antiviral activity of the A. pleuropneumoniae supernatant. Finally, A. pleuropneumoniae supernatant characterization was partially achieved using mass spectrometry.
Using antibody microarray, we observed modulations in G2/M-phase cell cycle regulation pathway when SJPL cells were treated with A. pleuropneumoniae culture supernatant. These modulations were confirmed by a cell cycle arrest at the G2/M-phase when cells were treated with the A. pleuropneumoniae culture supernatant. Furthermore, two G2/M-phase cell cycle inhibitors demonstrated the ability to inhibit PRRSV infection, indicating a potential key role for PRRSV infection. Finally, mass spectrometry lead to identify two molecules (m/z 515.2 and m/z 663.6) present only in the culture supernatant.
We demonstrated for the first time that A. pleuropneumoniae is able to disrupt SJPL cell cycle resulting in inhibitory activity against PRRSV. Furthermore, two putative molecules were identified from the culture supernatant. This study highlighted the cell cycle importance for PRRSV and will allow the development of new prophylactic or therapeutic approaches against PRRSV.
猪繁殖与呼吸综合征病毒（PRRSV）是养猪业中最重要的病原体之一，会造成重大经济损失。目前尚无针对该病毒的有效商业抗病毒药物。我们最近报道，猪胸膜肺炎放线杆菌（猪胸膜肺炎的病原体）的培养上清液在体外对SJPL细胞中的PRRSV具有抗病毒活性。本研究的目的是：（i）确定猪胸膜肺炎放线杆菌显示抗病毒活性的机制；（ii）表征细菌培养上清液中存在的活性分子。
使用抗体微阵列分析来指出受猪胸膜肺炎放线杆菌上清液调节的细胞途径。随后，采用流式细胞术分析和细胞周期抑制剂来确认抗体微阵列数据，并将其与猪胸膜肺炎放线杆菌上清液的抗病毒活性联系起来。最后，通过质谱法部分实现了对猪胸膜肺炎放线杆菌上清液的表征。
使用抗体微阵列，我们观察到当用猪胸膜肺炎放线杆菌培养上清液处理SJPL细胞时，G2/M期细胞周期调控途径发生了变化。当用猪胸膜肺炎放线杆菌培养上清液处理细胞时，在G2/M期出现细胞周期停滞，从而证实了这些变化。此外，两种G2/M期细胞周期抑制剂显示出抑制PRRSV感染的能力，这表明其在PRRSV感染中可能起关键作用。最后，质谱分析鉴定出仅存在于培养上清液中的两种分子（质荷比分别为515.2和663.6）。
我们首次证明猪胸膜肺炎放线杆菌能够破坏SJPL细胞周期，从而产生对PRRSV的抑制活性。此外，从培养上清液中鉴定出两种假定的分子。本研究突出了细胞周期对PRRSV的重要性，并将有助于开发针对PRRSV的新的预防或治疗方法。
