Development of Immunoprecipitation-qPCR to Measure Infective Viruses in Environmental Water

2020 Fiscal Year Final Research Report

• Project/Area Number: 18K19676
• Research Category: Grant-in-Aid for Challenging Research (Exploratory)
• Allocation Type: Multi-year Fund
• Review Section: Medium-sized Section 58:Society medicine, nursing, and related fields
• Research Institution: Kyoto University
• Principal Investigator: Ihara Masaru 京都大学, 工学研究科, 特定助教 (70450202)
• Project Period (FY): 2018-06-29 – 2021-03-31
• Keywords: ウイルス / 下水 / 抗体 / 免疫沈降 / MS2

Outline of Final Research Achievements

We aimed to develop a method for selectively concentrating a virus from sewage using an antibody. The method was developed by using bacteriophage MS2. The cultured MS2 was immunoprecipitated by adding an anti-MS2 antibody. The amount of MS2 recovered was quantified by reverse transcription (RT)-qPCR. As a result, the amount of MS2 recovered increased depending on the amount of antibody input. Anti-MS2 antibody did not precipitate bacteriophage Qβ. Furthermore, it was confirmed by the culture method that culturable MS2 could be recovered by immunoprecipitation. Then we confirmed by reverse RT-qPCR that norovirus in sewage could also be concentrated by immunoprecipitation. From these results, it was demonstrated that the infectious virus could be selectively concentrated by immunoprecipitation.

Free Research Field

水中健康関連微生物、環境毒性学

Academic Significance and Societal Importance of the Research Achievements

下水でのウイルスモニタリングによる市中感染状況の把握は、市中での感染症流行抑制のための有効な方法として注目を集めているが、下水からのウイルス濃縮方法には改善の余地がある。本研究では、モデルウイルスを用いた実験によって、抗体を用いた免疫沈降によって下水中の感染性のあるウイルスを選択的に濃縮できる可能性を示すことができた。この技術は、ノロウイルスに限らず他のウイルスへも応用可能である。今後、大量の下水試料へ対応可能なスケールアップが実現できれば、下水中の様々なウイルスの選択的、高効率な濃縮の実現の可能性がある。