Pilot project: "Establishment of the siRNA-mediated knock down in primary macrophages of different animal species"

Within the pilot project, a small interfering (si)RNA-mediated knock down (RNA interference/silencing) was established in primary avian and bovine macrophages to provide a suitable model for the molecular investigation of host-pathogen interactions in these animal species. Since the inducible nitric oxide synthase (iNOS) is an important factor in host defence against intracellular pathogens such as salmonella (Chakravortty, 2003), the method was incorporated using the example of iNOS synthesis. For this work, an in vitro infection model using primary avian blood monocyte-ripened macrophages and salmonella (Braukmann, 2015), already established in the "Infection Immunology Department Poultry" working group of the Friedrich Loeffler Institute, was available. The project used this already existing Salmonella (S.) infection model for primary avian macrophages to establish the iNOS siRNA knock out and control its efficiency. After successful establishment of RNA interference in primary avian cells, the methodology was also transferred to primary bovine blood monocyte-ripened macrophages and subsequently incorporated for infection with the S. serovars typhimurium and gallinarum to investigate the effects of iNOS silencing on host-pathogen interactions and the survival of salmonella during infection.

The pilot project will provide the zoonosis research community with an in vitro model of small interfering (si)RNA-mediated knock down in primary avian and bovine macrophages. With this, different host-adapted S. serovars (but also other bacterial species and genetically modified pathogens) can be investigated in the molecular interaction with the host.

Literature:

Braukmann, M., Methner, U., Berndt, A. (2015). "Immune Reaction and Survivability of Salmonella Typhimurium and Salmonella Infantis after Infection of Primary Avian Macrophages". PLoS ONE 10(3):e0122540
Chakravortty, D. and M. Hensel (2003). "Inducible nitric oxide synthase and control of intracellular bacterial pathogens." Microbes Infect 5(7): 621-627

Coordination: PD Dr. Angela Berndt (Friedrich-Loeffler-Institute, Institute for Molecular Pathogenesis)

Funding period: 12 months

Project start: 01.03.2015

Funding: BMBF