Engineered resistance to Nosema bombycis by in vitro expression of a single-chain antibody in Sf9-III cells

PLoS One. 2018 Feb 15;13(2):e0193065. doi: 10.1371/journal.pone.0193065. eCollection 2018.

Abstract

Nosema bombycis is a destructive, obligate intracellular parasite of the Bombyx mori. In this study, a single-chain variable fragment (scFv) dependent technology is developed for the purpose of inhibiting parasite proliferation in insect cells. The scFv-G4, which we prepared from a mouse G4 monoclonal antibody, can target the N. bombycis spore wall protein 12 (NbSWP12). Indirect immunofluorescence assays showed that NbSWP12 located mainly on the outside of the N. bombycis cytoskeleton, although some of it co-localized with β-tubulin in the meront-stage of parasites. When meront division began, NbSWP12 became concentrated at both ends of each meront. Western blotting showed that scFv-G4 could express in Sf9-III cells and recognized native NbSWP12. The transgenic Sf9-III cell line showed better resistance than the controls when challenged with N. bombycis, indicating that NbSWP12 is a promising target in this parasite and this scFv dependent strategy could be a solution for construction of N. bombycis-resistant Bombyx mori.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibodies, Monoclonal / genetics
  • Antibodies, Monoclonal / immunology
  • Blotting, Western
  • Bombyx / genetics
  • Bombyx / immunology*
  • Bombyx / microbiology
  • Cloning, Molecular
  • Disease Resistance / genetics
  • Disease Resistance / immunology*
  • Enzyme-Linked Immunosorbent Assay
  • Fluorescent Antibody Technique, Indirect
  • Nosema / immunology
  • Nosema / pathogenicity*
  • Organisms, Genetically Modified
  • Real-Time Polymerase Chain Reaction
  • Single-Chain Antibodies / genetics
  • Single-Chain Antibodies / immunology*

Substances

  • Antibodies, Monoclonal
  • Single-Chain Antibodies

Grants and funding

This work was supported by grants from the National Basic Research Program of China (No.2012CB114604), Fundamental Research Funds for the Central Universities (XDJK2015A010) to CFL. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.