Quantification of intracellular bacteria

Background of the quantification of intracellular bacteria 

• Quantification of intracellular bacteria is fundamental in many areas of cellular and clinical microbiology to study acute and chronic infections.
• Rapid, accurate and low-cost methods represent valuable tools in determining bacterial ability to persist and proliferate within eukaryotic cells.
• Chlamydia species are obligate intracellular bacteria that require growth inside a mammalian host cell for propagation and survival.
   

Objectives of the model

• To develop a rapid and accurate real time PCR method for the detection of intracellular Chlamydia trachomatis.

   

Our approach at Vibiosphen 

• McCoy cell monolayers were infected by C. trachomatis strain LGV2 (3 infectious doses).
• Infected cells were harvested and lysed by heating or repeated freezing/thawing cycles to release intracellular bacteria.
• Intracellular bacteria were quantified by qPCR.
   

Outcomes of the quantification of intracellular bacteria 
 

• A dose-response relationship was detected for both lysis methods.
• Freezing/thawing cycles appeared to be the most efficient method to release intracellular bacteria.
• This approach can be applied for the quantification of bacterial internalization by host cells in a gentamycin protection assay (Neisseria, Salmonella,  Listeria…).
• This approach could also be optimized for in vivo studies.