A real-time impedance based method to assess Rhodococcus equi virulence.

Area of Science:

• Veterinary Microbiology
• Infectious Diseases
• Cellular Pathogenesis

Background:

• Rhodococcus equi is a significant cause of pneumonia in foals, acting as a facultative intracellular pathogen.
• Current methods for assessing R. equi virulence, such as bacterial enumeration post-macrophage lysis, are laborious and not high-throughput.
• A need exists for efficient, real-time methods to study R. equi-macrophage interactions and virulence.

Purpose of the Study:

• To develop and validate an impedance-based real-time method for characterizing Rhodococcus equi proliferation within macrophages.
• To compare the proliferation of virulent R. equi strains with attenuated strains (vapA gene or virulence plasmid deficient).
• To investigate the influence of bacterial factors and host cell responses on the impedance signal.

Main Methods:

• Utilized an impedance-based real-time biosensor system to monitor R. equi growth in macrophage cultures.
• Employed virulent and attenuated R. equi strains, including those lacking the vapA gene or virulence plasmid.
• Analyzed time-dependent cell response profiles (TCRP) and correlated them with bacterial virulence and host cell characteristics.

Main Results:

• The impedance-based method successfully characterized R. equi proliferation in real-time.
• The time-dependent cell response profile (TCRP) was influenced by macrophage cell size, roundness, and bacterial cytotoxicity.
• The amplitude and inflection point of the TCRP varied with the multiplicity of infection and the virulence of the R. equi strain, enabling differentiation between virulent and attenuated strains.

Conclusions:

• Impedance-based biosensing offers a high-throughput, real-time alternative for assessing R. equi virulence and intracellular growth.
• The method provides valuable insights into the interaction dynamics between R. equi and macrophages.
• This approach can aid in the rapid evaluation of R. equi virulence factors and the development of novel diagnostic or therapeutic strategies.