An artificial medium mimicking human sputum

Ref: 13611
This medium had been developed for laboratory use to culture respiratory bacteria in an environment that closely mimics human lung sputum.

Key advantages
  • Obtaining sufficient human sputum for culture is difficult and time consuming
  • ASMDM has all the basic components of sputum, plus those components needed to mimic sputum from cystic fibrosis patients.
  • The quantities and components can altered to suit any disease-specific lung sputum, e.g. COPD and bronchiectasis

Background

Respiratory bacteria such as P. aeruginosa, H. influenzae and S. aureus grow in human sputum or associated epithelial cells; therefore understanding their growth characteristics in sputum is an important step to understanding in vivo infection.

Currently laboratories grow respiratory bacteria on blood agar or enriched broth media such as Luria broth or Brain Heart Infusion Broth. These are general purpose media and are not specific to the level of nutrients and composition of any in vivo growth site. Bacteria express particular genes/proteins depending on the composition of their environment.

Bacterial gene expression can vary considerably in different types of lab media and this is often a problem for understanding which virulence genes and proteins are significant in bacterial pathogenesis in order to be targeted with therapeutic drugs.

An artificial sputum-containing medium have been made previously by Sriramulu et al (2005), while a totally synthetic sputum medium has also been made (Palmer et al 2007).

The invention

Growth of P. aeruginosa UCBPP-PA14, the Australian Epidemic Strain-1 isolate AES-1R

Growth of P. aeruginosa UCBPP-PA14, the Australian Epidemic Strain-1 isolate AES-1R and the non-epidemic isolate 34Bris in 24-well plates at 24 and 48 h post-inoculation, showing evidence of microcolony formation through the increasing density of the stained regions. The red color of the indicator oxidized by growing cells demarcates the boundaries of the expanding region of cell to cell attachment leading to microcolony formation.

Based on the knowledge on the chemical and nutritional composition of normal sputum, cystic fibrosis sputum and other artificial sputum media, we have refined and produced a media that closely mimics cystic fibrosis sputum.

This media has been validated with P. aeruginosa genomic expression studies and by comparison of the growth characteristics of P. aeruginosa in this medium with those known to occur in cystic fibrosis sputum (Fung et al 2010).

Applications

This media provides an ideal growth environment for respiratory bacteria from sputum and as such will help clinical and research scientists to look at the expression of specific genes/proteins in an “almost in-vivo” setting.

Furthermore, bacterial characteristics such as biofilm formation, tight microcolony formation, anaerobic growth and virulence factor expression can be monitored as they would be expected to occur in human sputum

Principal inventors

  • Dr Jim Manos
  • A/Prof Barbara Rose
  • A/Prof Cynthia Whitchurch
  • Dr Lynne Turnbull

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