Imperial/Infector Detector/Introduction

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    <li><a class="current" href="https://2007.igem.org/Imperial/Infector_Detector/Introduction" title=""><span>Introduction</span></a></li>
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    <li><a href="https://2007.igem.org/Imperial/Infector_Detector/Specification" title=""><span>Specifications</span></a></li>
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    <li><a href="https://2007.igem.org/Imperial/Infector_Detector/Design" title=""><span>Design</span></a></li>
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    <li><a href="https://2007.igem.org/Imperial/Infector_Detector/Modelling" title=""><span>Modelling</span></a></li>
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    <li><a href="https://2007.igem.org/Imperial/Infector_Detector/Implementation" title=""><span>Implementation</span></a></li>
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    <li><a href="https://2007.igem.org/Imperial/Infector_Detector/Testing" title=""><span>Testing</span></a></li>
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    <li><a href="https://2007.igem.org/Imperial/Infector_Detector/F2620 Comparison" title=""><span>F2620</span></a></li>
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    <li><a href="https://2007.igem.org/Imperial/Infector_Detector/Conclusion" title=""><span>Conclusion</span></a></li>
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__NOTOC__
= Infector Detector: Introduction =
= Infector Detector: Introduction =
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==Project Summary ==
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[[Image:IC07 cartoon.png|center|900px]] <br clear="all">
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In this project, we have decided to tackle the ongoing problem of catheter-associated urinary tract infections (CAUTI). To do this, we targetted biofilms formed by pathogens on the catheter, and designed a system which would be able to detect its presence. To do this, we identified the signaling molecule AHL, which is produced by these bacteria in response to biofilm formation. We picked the LuxR and lux promoter as components to detect the presence of AHL in the surroundings. We also implemented the cell free system, or the ''in vitro'' chassis to reduce the possible inherent sources of contamination of our device. Essentially, we have created an AHL detector which is able to differentiate between different concentrations of AHL and visibly report the presence of AHL above a specific threshold of 5nM.
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== Motivations ==
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Infector Detector tackles the ongoing problem of catheter-associated urinary tract infections. To do this, we looked at how infections develop - as biofilms - and designed a system which would be able to detect their presence. We have created a system which is capable of detecting one of the types of signalling molecules found in biofilms, AHL, and visibly report its presence by producing a fluorescent protein.
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=====Why urinary catheters?=====
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Urinary catheters are used commonly in all hospitals - their primary purpose is to drain the bladder of patients who are unable to do so themselves. Patients who are either confined to bed or undergoing surgery often need to be catheterized. However, the application of a urinary cathether can lead to urinary tract infection, the most common nosocomial infection in the hospital.
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So what are the risks of obtaining a catheter-associated urinary tract infection? Patients who receive one brief catheterization have a 1% chance of getting a urinary tract infection. With approximately 4-5 catheterizations per day, the chance of infection can reach a total of 5% daily. Mishandling of the catheter, or application by unspecialized staff can further increase the risk to around 20% [[Imperial/Infector_Detector/References#Ref1|(1.Kunin,1996)]].
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== Motivation ==
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Treatment of a urinary tract infection with antibiotics generally cost only US$1,000 [[Imperial/Infector_Detector/References#Ref2|(2.Maki,2001)]]. However, if further complications arise, the cost of treatment increases dramatically. Also, considering the high risk of obtaining such an infection and the large number of people who undergo catheterization every year (over 5 million in the US), the treatment costs total up to a phenominal number. Furthermore, further antibiotic treatment can result in the development of stronger resistant bacteria, making urinary tract infections harder to treat in the future.
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=== Urinary Catheter Infections ===
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'''Urinary catheters''' are the major source of '''hospital infections''' in the US: more than 40%, or '''1 million''' infections per year '''costing''' health services up to '''1 billion USD'''<sup>[[#References |1-4]]</sup>. It is a very uncomfortable and painful condition. Infection develops in up to '''25% of patients''' requiring a urinary catheter for more than seven days<sup>[[#References |5-6]]</sup>. They are the second most common cause of hospital bloodstream infection<sup>[[#References |8-10]]</sup>. Additionally, urinary catheters comprise the largest institutional reservoir of nosocomial '''antibiotic-resistant pathogens'''.<sup>[[#References |5-10]]</sup>
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E.coli alone is responsible for over a quarter of urinary catheter infections. Pseudomonas aeruginosa is another major agent. Such infections are caused by a bacterial biofilm that forms on the surface of the catheter, and then spreads up into the urethra.
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=====What causes urinary tract infections?=====
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{| style="v-align: top;"
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There are a large variety of bacteria that can be involved in urinary tract infection. Most common ones include ''Escherichia coli'', ''Enterococcus'' and ''Enterobacter'', all of which are gram-negative bacteria. These pathogens are generally opportunistic, and are not highly pathogenic. Occasionally, more pathogenic species, such as ''Staphylococcus aureus'' (which is responsible for MRSA), might also cause such infections.
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|-
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|[[image:IC07_catheter.gif|thumb|right|300px|Male and female urinary catheters. [http://www.oldcity.org.uk/cauda_equina/images/catheter.gif www.oldcity.org.uk]]]  || [[Image:IC07 infectedcatheter.jpg|thumb|An infected catheter<sup>[[#References |12]]</sup>.]] ||[[Image:IC07 infectedcatheter2.jpg|thumb|240px|Scanning electron micrograph of the surface of a catheter removed from patient with catheter-associated UTI<sup>[[#References |11]]</sup>.]]
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One important feature found in all these species of bacteria is their ability to form a thick bacteria biofilms. Biofilms consists of a complex of polysaccharides, excreted proteins and the bacteria glycocalyx. These mixture of molecules not only allow the bacteria to adhere together, but can also protect the bacteria from various foreign materials, most importantly antibiotics or anti-microbial agents. Some research groups have also shown that cleaning a cathether does not significantly reduce the risk of infection, possibly due to the resistance of bacteria to these foreign agents.
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There are several modes of entry for the bacteria into the urinary tract. The most common mode of infection is by the migration of bacteria on the tube into the urinary tract. Bacteria can contaminte the collection bag or the catheter tube by exogenous sources (such as the non-sterile gloves), or endogenous sources, such as the urine or other waste products of the patients. The process of the extension of the bacteria biofilm along the urinary tract cannot be determined by current medical procedures, since the infection has not begun, and the patient will not exhibit any symptoms.
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[[Image:IC07 biofilmspread.jpg|thumb|400px|right|Biofilm formation and spread: '''1''' Cells attach to a surface. '''2''' Cells begin producing polysaccharides. '''3''' A protective layer forms around the cells. '''4''' The biofilm grows. '''5''' Cells are released into the environment, continuing the cycle.]]
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=====How can we reduce the risk?=====
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=== Biofilms ===
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In our project, we have decided that the best way to prevent a patient from contracting catheter-associated urinary tract infection is to have a way to assay for the presence of this bacteria. Our device would detect for the presence of biofilm, which is an indicator of the bacteria. If the bacteria is shown to be present on the catheter, it can be changed immediately  to reduce the risk of the bacteria from migrating into the urinary tract. So how would we achieve this? Take a look at the specifications of our device!
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A biofilm forms when an organism attaches to a surface, and begins producing polysaccharides. This forms a protective matrix around the organism, to which other cells can attach and secrete even more polysaccharides. The biofilm grows until it begins to release cells and other debris into its environment. Released cells then attach to other parts of the surface, starting the cycle anew<sup>[[#References |13-15]]</sup>.
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== Achievements ==
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And like that, the biofilm spreads around and creeps along a surface. This process takes place between a few hours to a few days.
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Biofilms are very tough to eradicate - they are difficult to clean, and cells contained within are very hard to target with normal antibiotics or antiseptics.
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=== Cell signalling inside biofilms ===
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The behaviour of cells in a biofilm is very different from that of those in planktonic state. They talk to each other through molecular signalling mechanisms, such as quorum sensing and wall sensing. An important characteristic of biofilms, then, is that there are signal molecules floating around inside of it.
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[[Image:IC07 AHLmechanism.jpg|thumb|left|400px|The AHL quorum sensing mechanism. AHL enters the cell, where it binds to LuxR, forming a complex capable of activating the pLux promoter.]]
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One type of signalling molecule, AHL, is produced by Pseudomonas aeruginosa when forming biofilms<sup>[[#References |16]]</sup>. MRSA has a similar behaviour – the agr quorum sensing mechanism<sup>[[#References |17]]</sup>. Every biofilm has a similar mechanism, because this type of signalling controls the growth of the biofilm.
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Our system, Infector Detector, exploits exactly this mechanism in order to detect biofilms.
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== Achievements ==
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* We used synthetic biology to tackle the problem of urinary catheter infections. We achieved this using only parts available in the Registry, and a new in-vitro chassis.
 +
* We designed, built, and tested successfully a system capable of detecting AHL at concentrations as low as 5nM, and responding within 3 hours.
 +
* We tested the system in-vivo, using E.coli, and in-vitro, using titrated AHL. The next step is to test it on real biofilms.
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<center>  On to next stage : | [https://2007.igem.org/Imperial/Infector_Detector/Specification Specifications >>]
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</center>
== References ==
== References ==
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# Stamm WE. Catheter-associated urinary tract infections: Epidemiology, pathogenesis, and prevention. Am J Med 1991;91(Suppl 3B):65S-71S.
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# Burke JP, Riley DK. Nosocomial urinary tract infection. In: Mayhall CG, editor. Hospital epidemiology and infection control. Baltimore: Williams and Wilkins; 1996. p. 139-53.
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# Warren JW. Catheter-associated urinary tract infections. Infect Dis Clin North Am 1997;11:609-22.
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# Kunin CM. Care of the urinary catheter. In: Urinary tract infections: detection, prevention and management. Fifth ed. Baltimore: Williams and Wilkins; 1997. p. 227-99.
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# Kunin CM, McCormack RC. Prevention of catheter-induced urinary-tract infections by sterile closed drainage. N Engl J Med 1966;274:1155-61.
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# Garibaldi RA, et al. An evaluation of daily bacteriologic monitoring to identify preventable episodes of catheter associated UTI. Infect Control 1982;3:466-70.
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# Stark RP, Maki DG. Bacteriuria in the catheterized patient. N Engl J Med 1984;311:560-4.
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# Maki DG. Nosocomial bacteremia. An epidemiologic overview. Am J Med 1981;70:719-32.
 +
# Krieger JN, Kaiser DIL, Wenzel RP. Urinary tract etiology of bloodstream infections in hospitalized patients. J Infect Dis 1983;148:57-62.
 +
# Bryan CS, Reynolds KL. Hospital-acquired bacteremic urinary tract infection: epidemiology and outcome. J Urol 1984,132:494-8.
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# [http://www.medscape.com/viewarticle/416647_3 Bacterial Biofilms in Urology].Infect Urol 11(6):169-175, 1998.
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# Laube, N. Diamonds are a Urologist's Best Friend. [http://www.biofilmsonline.com/cgi-bin/biofilmsonline/00271.html BiofilmsOnline.com] 18-Nov-2004.
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#  Donlan RM. Biofilms: microbial life on surfaces. Emerg Infect Dis 2002 Sep; 8(9) 881-90. pmid:12194761. PubMed HubMed [biofilm4]
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# Donlan RM. Biofilm formation: a clinically relevant microbiological process. Clin Infect Dis 2001 Oct 15; 33(8) 1387-92.
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# Tenke P, Riedl CR, Jones GL, Williams GJ, Stickler D, and Nagy E. Bacterial biofilm formation on urologic devices and heparin coating as preventive strategy. Int J Antimicrob Agents 2004 Mar; 23 Suppl 1 S67-74. doi:10.1016/j.ijantimicag.2003.12.007
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# David J. Stickler, et al. Biofilms on Indwelling Urethral Catheters Produce Quorum-Sensing Signal Molecules In Situ and In Vitro. Applied and Environmental Microbiology, September 1998, p. 3486-3490, Vol. 64, No. 9
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# Manago K, et al. Biofilm formation by and accessory gene regulator typing of methicillin-resistant Staphylococcus aureus strains recovered from patients with nosocomial infections. Infect Control Hosp Epidemiol 2006 Feb; 27(2) 188-90. doi:10.1086/500620 pmid:16465637.

Latest revision as of 02:30, 27 October 2007




Infector Detector: Introduction

IC07 cartoon.png

Infector Detector tackles the ongoing problem of catheter-associated urinary tract infections. To do this, we looked at how infections develop - as biofilms - and designed a system which would be able to detect their presence. We have created a system which is capable of detecting one of the types of signalling molecules found in biofilms, AHL, and visibly report its presence by producing a fluorescent protein.

Motivation

Urinary Catheter Infections

Urinary catheters are the major source of hospital infections in the US: more than 40%, or 1 million infections per year costing health services up to 1 billion USD1-4. It is a very uncomfortable and painful condition. Infection develops in up to 25% of patients requiring a urinary catheter for more than seven days5-6. They are the second most common cause of hospital bloodstream infection8-10. Additionally, urinary catheters comprise the largest institutional reservoir of nosocomial antibiotic-resistant pathogens.5-10

E.coli alone is responsible for over a quarter of urinary catheter infections. Pseudomonas aeruginosa is another major agent. Such infections are caused by a bacterial biofilm that forms on the surface of the catheter, and then spreads up into the urethra.

Male and female urinary catheters. www.oldcity.org.uk
An infected catheter12.
Scanning electron micrograph of the surface of a catheter removed from patient with catheter-associated UTI11.


Biofilm formation and spread: 1 Cells attach to a surface. 2 Cells begin producing polysaccharides. 3 A protective layer forms around the cells. 4 The biofilm grows. 5 Cells are released into the environment, continuing the cycle.

Biofilms

A biofilm forms when an organism attaches to a surface, and begins producing polysaccharides. This forms a protective matrix around the organism, to which other cells can attach and secrete even more polysaccharides. The biofilm grows until it begins to release cells and other debris into its environment. Released cells then attach to other parts of the surface, starting the cycle anew13-15.

And like that, the biofilm spreads around and creeps along a surface. This process takes place between a few hours to a few days.

Biofilms are very tough to eradicate - they are difficult to clean, and cells contained within are very hard to target with normal antibiotics or antiseptics.


Cell signalling inside biofilms

The behaviour of cells in a biofilm is very different from that of those in planktonic state. They talk to each other through molecular signalling mechanisms, such as quorum sensing and wall sensing. An important characteristic of biofilms, then, is that there are signal molecules floating around inside of it.

The AHL quorum sensing mechanism. AHL enters the cell, where it binds to LuxR, forming a complex capable of activating the pLux promoter.

One type of signalling molecule, AHL, is produced by Pseudomonas aeruginosa when forming biofilms16. MRSA has a similar behaviour – the agr quorum sensing mechanism17. Every biofilm has a similar mechanism, because this type of signalling controls the growth of the biofilm.

Our system, Infector Detector, exploits exactly this mechanism in order to detect biofilms.


Achievements

  • We used synthetic biology to tackle the problem of urinary catheter infections. We achieved this using only parts available in the Registry, and a new in-vitro chassis.
  • We designed, built, and tested successfully a system capable of detecting AHL at concentrations as low as 5nM, and responding within 3 hours.
  • We tested the system in-vivo, using E.coli, and in-vitro, using titrated AHL. The next step is to test it on real biofilms.


On to next stage : | Specifications >>

References

  1. Stamm WE. Catheter-associated urinary tract infections: Epidemiology, pathogenesis, and prevention. Am J Med 1991;91(Suppl 3B):65S-71S.
  2. Burke JP, Riley DK. Nosocomial urinary tract infection. In: Mayhall CG, editor. Hospital epidemiology and infection control. Baltimore: Williams and Wilkins; 1996. p. 139-53.
  3. Warren JW. Catheter-associated urinary tract infections. Infect Dis Clin North Am 1997;11:609-22.
  4. Kunin CM. Care of the urinary catheter. In: Urinary tract infections: detection, prevention and management. Fifth ed. Baltimore: Williams and Wilkins; 1997. p. 227-99.
  5. Kunin CM, McCormack RC. Prevention of catheter-induced urinary-tract infections by sterile closed drainage. N Engl J Med 1966;274:1155-61.
  6. Garibaldi RA, et al. An evaluation of daily bacteriologic monitoring to identify preventable episodes of catheter associated UTI. Infect Control 1982;3:466-70.
  7. Stark RP, Maki DG. Bacteriuria in the catheterized patient. N Engl J Med 1984;311:560-4.
  8. Maki DG. Nosocomial bacteremia. An epidemiologic overview. Am J Med 1981;70:719-32.
  9. Krieger JN, Kaiser DIL, Wenzel RP. Urinary tract etiology of bloodstream infections in hospitalized patients. J Infect Dis 1983;148:57-62.
  10. Bryan CS, Reynolds KL. Hospital-acquired bacteremic urinary tract infection: epidemiology and outcome. J Urol 1984,132:494-8.
  11. Bacterial Biofilms in Urology.Infect Urol 11(6):169-175, 1998.
  12. Laube, N. Diamonds are a Urologist's Best Friend. BiofilmsOnline.com 18-Nov-2004.
  13. Donlan RM. Biofilms: microbial life on surfaces. Emerg Infect Dis 2002 Sep; 8(9) 881-90. pmid:12194761. PubMed HubMed [biofilm4]
  14. Donlan RM. Biofilm formation: a clinically relevant microbiological process. Clin Infect Dis 2001 Oct 15; 33(8) 1387-92.
  15. Tenke P, Riedl CR, Jones GL, Williams GJ, Stickler D, and Nagy E. Bacterial biofilm formation on urologic devices and heparin coating as preventive strategy. Int J Antimicrob Agents 2004 Mar; 23 Suppl 1 S67-74. doi:10.1016/j.ijantimicag.2003.12.007
  16. David J. Stickler, et al. Biofilms on Indwelling Urethral Catheters Produce Quorum-Sensing Signal Molecules In Situ and In Vitro. Applied and Environmental Microbiology, September 1998, p. 3486-3490, Vol. 64, No. 9
  17. Manago K, et al. Biofilm formation by and accessory gene regulator typing of methicillin-resistant Staphylococcus aureus strains recovered from patients with nosocomial infections. Infect Control Hosp Epidemiol 2006 Feb; 27(2) 188-90. doi:10.1086/500620 pmid:16465637.