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Related Experiment Videos

Expression of Staphylococcus aureus clumping factor A in Lactococcus lactis subsp. cremoris using a new shuttle

Y A Que1, J A Haefliger, P Francioli

  • 1Division of Infectious Diseases, Department of Internal Medicine, Centre Hospitalier Universitaire Vaudois, 1011 Lausanne, Switzerland.

Infection and Immunity
|May 19, 2000
PubMed
Summary

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Researchers developed a novel system to express Staphylococcus aureus adhesins in Lactococcus lactis. This method allows for studying individual virulence factors, showing functional ClfA protein production in the new host.

Area of Science:

  • Microbiology
  • Molecular Biology
  • Bacterial Pathogenesis

Background:

  • Staphylococcus aureus possesses multiple adhesins crucial for infection.
  • Understanding the specific role of each adhesin is challenging due to redundancy.
  • A need exists for systems to study individual virulence factors outside their native context.

Purpose of the Study:

  • To develop a heterologous expression system for Staphylococcus aureus adhesins.
  • To evaluate the function of staphylococcal adhesins in a non-virulent host.
  • To create a tool for dissecting the contribution of individual adhesins to bacterial colonization.

Main Methods:

  • Construction of a novel Escherichia coli-Lactococcus lactis shuttle vector (pOri23) with a strong lactococcal promoter (P23).

Related Experiment Videos

  • Cloning and expression of the staphylococcal clumping factor A (clfA) gene in L. lactis.
  • Assessment of ClfA protein production and function using Western blots and plasma-induced clumping assays.
  • Quantification of bacterial clumping and fibrinogen binding.

    • Main Results:

    • The pOri23 vector enabled high-level constitutive expression of cloned genes in L. lactis, with no expression in E. coli.
    • Lactococci harboring pOri23-clfA successfully produced a functional 130-kDa ClfA protein on their cell surface.
    • ClfA-positive L. lactis exhibited significant clumping in plasma and comparable fibrinogen binding to S. aureus Newman.
    • The expression system successfully demonstrated the adhesive properties of ClfA in a heterologous host.

    Conclusions:

    • A robust Lactococcus lactis-based expression system for studying Staphylococcus aureus adhesins has been established.
    • This system allows for the functional characterization of individual staphylococcal virulence factors in a simplified genetic background.
    • The findings offer a complementary approach to existing mutagenesis and in vivo expression technologies for understanding bacterial pathogenesis.