The in-veso biofilm sensor

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The in-veso biofilm sensor
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Introduction

• Aim
• Characterise cell-free gene expression
• Use cell-free approach to detect quorum sensing
  within a bacterial biofilm

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Application
Detect bacterial biofilm on catheters
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Motivation

• Bacterial biofilms are impermeable to
  antimicrobial agents
• Biofilm-producing MRSA strains are associated
  with nosocomial infection this is a potential
  future application
• Cell-free approach eliminates need for bacterial
  chassis
• Attractive for detecting biofilms on medical
  devices

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Project Plan

• Proof of concept
• To model quorum sensing in biofilm using E.coli
  LuxI/LuxR system
• To control the concentration of the extracellular
  AHL signal
• To design a cell-free system that detects and
  reports the AHL signal
• To compare the cell-free detection system with
  E.coli detection system

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Specification

• The cell-free detection system should
• Have sensitivity to AHL 100ng/ml
• Output a signal visible to naked eye within 3
  hours
• Have a lifespan of 4 days
• Be applied through a gel spray or cream
• Work within temperatures between 20-30C
• Work in a pH range of 6-8

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Design

• In vitro transcription-translation
• Phospholipid vesicles
• S12/ S30 cell extract
• RNA polymerase
• Bacteriophage T7/ SP6 RNA
• Reporter
• Luciferase
• GFP
• Alpha-Hemolysin pores

Noireaux et. al. 2005
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Design
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Design
Biofilm bacteria
Cell-free system
LuxR
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Problems

• Noise leakage of pLux
• Cleaning the catheters after application
• Alpha-hemolysin might damage human cells

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Summary

• Non-microbial detection system
• Fast, simple and cheap
• Novel technique
• Lays a platform for future developments