Glenn Moonen and George Moutafis*

 

45 Poplar Rd,
Zoetis Australia Research & Manufacturing
Parkville, Victoria, Australia

 

Trueperella pyogenes (previously Arcanobacterium pyogenes) is an opportunistic pathogen that is considered to be the primary causative agent of metritis in cattle.  Approximately 90% of cattle become infected post parturition and most infections resolve naturally.  However, about 40% of these cases will advance to metritis and 20% of these, persist to endometritis.  The disease affects both milk production and reproductive capacity in the infected cows and current treatment is through the use of antibiotics and hormone therapies.  This organism possesses a number of virulence factors that contribute to its pathogenic potential, one of which is pyolysin, a cholesterol-dependent cytolysin.  This protein is a haemolysin, and is cytolytic for immune cells, including macrophages.  Expression of pyolysin is required for virulence of this bacterium.

This presentation will highlight how a wild-type T. pyogenes strain was developed into a GMP production organism by determining the factors that enhanced pyolysin production and finding methods to prevent the degradation of pyolysin by the host organism (thought to be a self-regulation mechanism).

T. pyogenes normally produces pyolysin at very low concentrations in complex microbiological media traditionally used to grow the organism, such as Brain Heart Infusion supplemented with foetal bovine serum.  It was identified that glucose present in the medium repressed expression of pyolysin and that it was only expressed following a glucose starvation event followed by the addition of an alternate carbon source for growth.  Pyolysin was then rapidly degraded by proteases produced by the organism presumed to limit the impact of the disease on the host.  The enzymes that degrade pyolysin are believed to be calcium-dependent proteases which are inhibited by the addition of EGTA to the fermentation medium.

An economical medium acceptable to government regulators that provided the essential vitamins, iron and fatty acids components was developed.  This improved medium in combination with optimised growth conditions provided a microaerophilic environment rich in carbon dioxide that enabled production of large quantities of pyolysin by T. pyogenes for the development of a vaccine antigen.

 

Biographic Details

Name: George Moutafis

Title: Technical Manager

Affiliation, Country: Zoetis Australia Research & Manufacturing, Australia

Phone: +61 3 83884383 Fax: +61 3 83884398  E-mail: george.mouafis@zoetis.com

Research interests: industrial microbiology, fermentation, veterinary microbiology

Venue

Room: 
Hawken N202