|
|
| Line 1: |
Line 1: |
| | {{Edinburgh_OG}} | | {{Edinburgh_OG}} |
| | <html> | | <html> |
| − | <div class="photo-block" id="Description">
| |
| − | <div class="text left">
| |
| − | <div class="title">
| |
| − | Project Background
| |
| − | </div>
| |
| − | </div>
| |
| − | </div>
| |
| − | <div class="wiki-block">
| |
| − | <div id="article">
| |
| − |
| |
| − | </html>
| |
| | | | |
| − | ==Context== | + | <p style="color: red; font-weight: bold">>>>>> gd2md-html alert: ERRORs: 0; WARNINGs: 0; ALERTS: 2.</p> |
| − | <i>Campylobacter jejuni</i> is a Gram-negative, microaerophilic, corkscrew-shaped bacteria which has been implicated as being one of the most common causes of human gastroenteritis worldwide. WHO estimates that 1 in 10 people around the world fall ill with a <i>Campylobacter</i> infection annually.<ref name= "WHO">http://www.who.int/mediacentre/factsheets/fs255/en/</ref> Infection with this bacteria in the UK was recently estimated to cost health services £50 million (in 2008-2009 prices), with a cost of £85 per case.<ref>Tam CC, O’Brien SJ (2016) Economic Cost of Campylobacter, Norovirus and Rotavirus Disease in the United Kingdom. PLoS ONE11(2): e0138526. https://doi.org/10.1371/journal.pone.0138526</ref>
| + | <ul style="color: red; font-weight: bold"><li>See top comment block for details on ERRORs and WARNINGs. <li>In the converted Markdown or HTML, search for inline alerts that start with >>>>> gd2md-html alert: for specific instances that need correction.</ul> |
| | | | |
| − | Infection with <i>Campylobacter</i> causes common symptoms such as diarrhoea, abdominal pain, fever, headache, nausea and vomiting. <ref name= "WHO"/>
| + | <p style="color: red; font-weight: bold">Links to alert messages:</p><a href="#gdcalert1">alert1</a> |
| − | <i>C. jejuni</i> grows optimally between 37 and 42°C and is most commonly found on undercooked poultry, though it has been reported in other undercooked meat and meat products, raw milk, and in untreated drinking water <ref name= "WHO"/>. The high prevalence of <i>C. jejuni</i> makes it an interesting target for synthetic biology-based solutions.
| + | <a href="#gdcalert2">alert2</a> |
| | | | |
| − | ===Treatment and Prevention=== | + | <p style="color: red; font-weight: bold">>>>>> PLEASE check and correct alert issues and delete this message and the inline alerts.<hr></p> |
| | | | |
| − | Treatment for Campylobacteriosis (<i>Campylobacter</i>-caused diarrhoeal disease) is generally only given in the most severe cases or when patients are vulnerable, and usually consists of giving more fluids and glucose-electrolyte solutions to ensure dehydration does not occur. <ref name="eMedicineTreatment"> https://emedicine.medscape.com/article/213720-treatment </ref> Erythromycin therapy can also be given but little clinical benefit is seen if the treatment is begun after four days of symptom onset. <ref>https://emedicine.medscape.com/article/176400-overview</ref> However, the use of antibiotics can be recommended in the case of patients who are immunocompromised, pregnant, or with very severe symptoms.<ref name="eMedicineTreatment"/>
| |
| | | | |
| − | Prevention of this infection is very easy, with several strategies from the farm to the consumer recommended by WHO: <ref name ="WHO"/>
| + | <p> |
| − | *Enhanced biosecurity to avoid transmission of <i>Campylobacter</i> from the environment to the flock of birds – only in closed housing conditions
| + | <strong><span style="text-decoration:underline;">Project Background</span></strong> |
| − | *Good hygienic slaughtering practices carried out
| + | </p> |
| − | *Prevention methods in domestic kitchens are similar to those specified for other foodborne bacterial infections – ensure meat is cooked through and that milk has been pasteurized before consumption
| + | <p> |
| | + | One of the increasingly concerning unsustainable practice in human practices that needs to be addressed immediately is the growth of municipal pollution. Petrochemical plastics are undoubtedly among the top contributors to this due to their long degradation time. At the same time, it is challenging to detach plastic from our everyday life and the global economy as it provides many benefits thanks to their advantageous properties. The demand has increased and plastic production has doubled in the last 50 years, mainly because of its unrivalled properties, low cost and relatively easy production (MacArthur, 2016). Despite their advantages, the production and use of plastics is also responsible for the generation of greenhouse gas emissions and harmful impacts of substances used in their current life cycle (Higson, 2018). The current and projected production of plastics could lead to a scenario in where the ocean will contain more plastics than fish, according to a report from The Ellen MacArthur Foundation. |
| | + | </p> |
| | + | <p> |
| | + | A small estuary seahorse, Hippocampus kuda, drifts in the polluted waters near Sumbawa Besar, Sumbara Island, Indonesia. |
| | + | </p> |
| | + | <p> |
| | | | |
| − | ==Current Detection Methods==
| |
| | | | |
| − | ====Culture-based screening==== | + | <p id="gdcalert1" ><span style="color: red; font-weight: bold">>>>>> gd2md-html alert: inline image link here (to images/Untitled-document0.png). Store image on your image server and adjust path/filename if necessary. </span><br>(<a href="#">Back to top</a>)(<a href="#gdcalert2">Next alert</a>)<br><span style="color: red; font-weight: bold">>>>>> </span></p> |
| | | | |
| | | | |
| − | Traditional methods for detection of <i>Campylobacter</i> include culturing a swabbed sample on selective media and incubating for 48 hours at 42°C under microaerobic conditions.
| + | <img src="images/Untitled-document0.png" width="" alt="alt_text" title="image_tooltip"> |
| | | | |
| − | [[File:GlasgowCJejuniCulture.jpeg|thumb|center|500px|<b>Figure 1:</b> Campylobacter jejuni colonies isolated on blood-free, charcoal based selective medium (CSM)<ref>https://commons.wikimedia.org/wiki/File:Campylobacter_jejuni.jpg</ref>]]
| + | </p> |
| | + | <p> |
| | + | PHOTOGRAPH BY JUSTIN HOFMAN, SEALEGACY<a href="https://www.nationalgeographic.com/photography/proof/2017/09/seahorse-ocean-pollution/"> https://www.nationalgeographic.com/photography/proof/2017/09/seahorse-ocean-pollution/</a> |
| | + | </p> |
| | + | <p> |
| | + | |
| | + | </p> |
| | + | <p> |
| | + | With the growing concern for the environment due to the negative impacts of plastic degradability and climate change coupled with the over use of fossil fuels (regarding source exhaustion and price stability, for instance), increasing efforts have been made to establish more sustainable bio-based production platforms. Particular attention has been given to Polyhydroxyalkanoates (PHAs), a broad family of biodegradable, biologically produced polymers with a tremendous capacity for use as plastics. Many microbes synthesise PHAs when in an excess of carbon, but limited oxygen conditions, they synthesise them to act as an energy store (Shah, et al., 2008). |
| | + | </p> |
| | + | <p> |
| | | | |
| − | After isolating the bacteria in this way, identification through looking at the morphology and overall characteristics of the bacteria is carried out. Culture-based methods are relatively cheap to carry out and require less training than others techniques, but are incredibly time and labour intensive and therefore, in recent years, a move has been made towards use of rapid detection testing. <ref>http://www.rapidmicrobiology.com/test-method/campylobacter-detection-and-identification-methods/</ref>
| |
| | | | |
| − | ==== Rapid detection====
| + | <p id="gdcalert2" ><span style="color: red; font-weight: bold">>>>>> gd2md-html alert: inline image link here (to images/Untitled-document1.png). Store image on your image server and adjust path/filename if necessary. </span><br>(<a href="#">Back to top</a>)(<a href="#gdcalert3">Next alert</a>)<br><span style="color: red; font-weight: bold">>>>>> </span></p> |
| − | More recently, techniques such as enzyme immunoassay and lateral flow systems have been utilised to test for Campylobacter, as they require only between one and two hours until a result is given.
| + | |
| − | <ref>Giltner CL, Saeki S, Bobenchik AM, Humphries RM. Rapid Detection of Campylobacter Antigen by Enzyme Immunoassay Leads to Increased Positivity Rates. Journal of Clinical Microbiology. 2013;51(2):618-620. doi:10.1128/JCM.02565-12.</ref> However, use of these methods required much more highly trained employees to carry them out, and so therefore detection of Campylobacter in an industrial or agricultural setting would most likely have to be outsourced to a company specialising in these services. | + | |
| − | A particular study doing a comparison into three rapid detection systems showed a high number of false negative results, which can be considered an additional drawback when looking at detection of C. jejuni in order to reduce the incidence of disease outbreaks.
| + | |
| − | <ref>Granato PA, Chen L, Holiday I, et al. Comparison of Premier CAMPY Enzyme Immunoassay (EIA), ProSpecT Campylobacter EIA, and ImmunoCard STAT! CAMPY Tests with Culture for Laboratory Diagnosis of Campylobacter Enteric Infections . Journal of Clinical Microbiology. 2010;48(11):4022-4027. doi:10.1128/JCM.00486-10.</ref> | + | |
| | | | |
| − | ==Our device - <i>Campy</i>LOCATOR==
| |
| | | | |
| − | In an attempt to reduce incidences of food poisoning by <i>Campylobacter jejuni</i>, and to improve upon current methods of detection, we decided to create a biosensor that was able to detect the presence of <i>Campylobacter jejuni</i> quickly and accurately. To do this we aimed to create a two part biosensor that will detect two sensory inputs. The first, xylulose, is a rare sugar most commonly associated with the pentose-phosphate pathway, where formation of xylulose-5-phosphate is an important intermediate step. Interestingly, xylulose was found incorporated into the polysaccharide capsule of Campylobacter jejuni strain RM1221 <ref>Gilbert, M., Mandrell, R., Parker, C., Li, J., and Vinogradov, E. (2017). Structural Analysis of the Capsular Polysaccharide from Campylobacter jejuni RM1221</ref>. The presence of xylulose is not common in bacterial polysaccharide capsules, and additionally the glyosidic bonds which incorporate xylulose were found to be extremely acid-labile. A large focus of the project was aimed at xylulose. The other molecule we identified as a marker for Campylobacter was autoinducer-2 (AI-2). AI-2 is a secreted quorum sensing molecule. AI-2 is a significantly less specific biomarker, as many varied gram-positive and gram-negative bacterial species sense their population density and surrounding bacterial environment using this molecule <ref>Miller, M. and Bassler, B. (2001). Quorum Sensing in Bacteria. Annual Review of Microbiology, 55(1), pp.165-199.</ref> On the other hand, this ubiquity meant that AI-2 gene regulation was well characterised, with prior iGEM teams having worked on the natural E. coli AI-2 quorum sensing regulatory system.
| + | <img src="images/Untitled-document1.png" width="" alt="alt_text" title="image_tooltip"> |
| | | | |
| − | For the detection of xylulose two possible avenues were explored: utilising the [https://2017.igem.org/Team:Glasgow/mtlR mannitol regulatory system] from <i>Pseudomonas fluorescens</i>, which is known to detect xylulose but may have some off-target effects; and [https://2017.igem.org/Team:Glasgow/araC mutagenesis of the arabinose regulatory system] from <i>Escherichia coli</i>, with an aim to change its target sugar to xylulose. The detectors for both xylulose and [https://2017.igem.org/Team:Glasgow/QuorumSensing autoinducer-2] will form the two components of an [https://2017.igem.org/Team:Glasgow/ANDGate AND gate] that will ensure only a signal is seen when both xylulose and autoinducer-2 are present.
| + | </p> |
| | + | <p> |
| | + | Source: European Bioplastics 2017<a href="https://www.european-bioplastics.org/bioplastics/materials/"> https://www.european-bioplastics.org/bioplastics/materials/</a> |
| | + | </p> |
| | + | <p> |
| | + | Currently, over 150 different monomers of PHA have been identified, with a huge range of properties. Poly(3-hydroxybutyrate) (PHB), the simplest PHA, was the first identified, and remains the most studied PHA, and has properties similar to polystyrene and polypropylene –highly crystalline, and brittle. Despite it being able to be utilised in much of the existing infrastructure –in injection moulding and other processes -it possesses a very narrow processing range, limiting its use. One way of combatting such difficulties is to incorporate different monomers into the polymer chain, forming co-polymers and even ter-polymers. By introducing such monomers into the chain, the crystallinity of the resulting material is disrupted, producing a more flexible plastic, and is frequently easier to process (Babu, et al., 2013). |
| | + | </p> |
| | + | <p> |
| | + | One of the most studied PHB copolymer is poly-(3-hydroxybuterate-3-hydroxyvalerate) or PHBV due to its highly versatile applications. Higher amount of HV content in PHBV decreases the melting point – it was reported that the melting point of PHBV samples were about 100-150°C whereas the degradation temperatures were more than 220°C which will indefinitely increase PHBV's flexibility and ductility (Wang, et al., 2013). In addition, higher HV content also increases the degradation rate. It was shown that within 19 weeks of degradation in pH 7.4 phosphate buffer medium, the mass loss experienced by PHBV with 5 mol% and 12 mol% were approximately 6.2% and 9.2%, respectively (Liu, Pancholi, and Raghavan, 2010). |
| | + | </p> |
| | + | <p> |
| | + | … |
| | + | </p> |
| | + | <p> |
| | + | |
| | + | </p> |
| | + | <p> |
| | | | |
| − | Regarding the application of our biosensor we went through [https://2017.igem.org/Team:Glasgow/Applied_Design several design iterations] to create [https://2017.igem.org/Team:Glasgow/Hardware a device] that would house the biosensor to make the application simple and easy. In addition to this we also looked at [https://2017.igem.org/Team:Glasgow/XyluloseBiosynthesis producing xylulose in a cheap and efficient way] as the cost of xylulose held us back with regards to thorough testing of the biosensor elements. All of the above were done with thought of the implications with regards to [https://2017.igem.org/Team:Glasgow/HP/Gold_Integrated human practices] and how a genetically engineered biosensor would be received by [https://2017.igem.org/Team:Glasgow/Outreach the public.]
| + | 1. MacArthur, D. E., Waughray, D., & Stuchtey, M. R. ,2016. The New Plastics Economy, Rethinking the Future of Plastics. InWorld Economic Forum. |
| | + | </p> |
| | + | <p> |
| | + | |
| | + | </p> |
| | + | <p> |
| | | | |
| − | ==References==
| + | 2. Higson, A., 2018. Bio-based & Biodegradable Plastic in the UK, Market Perspective. In NNFCC The Bioeconomy Consultants. |
| − | <small><references/></small> | + | </p> |
| − | <html></div> | + | <p> |
| − | </div></html> | + | |
| | + | </p> |
| | + | <p> |
| | + | |
| | + | 3. Shah, A., Hasan, F., Hameed, A. & Ahmed, S., 2008. Biological degradation of plastics: A comprehensive review. <em>Biotechnology Advances</em>, 26(3), pp. 246-265. |
| | + | </p> |
| | + | <p> |
| | + | |
| | + | </p> |
| | + | <p> |
| | + | |
| | + | 4. Babu, R., O'Connor, K. & Seeram, R., 2013. Current progress on bio-based polymers and their future trends. <em>Progress in Biomaterials</em>, 2(1), p 8. |
| | + | </p> |
| | + | <p> |
| | + | |
| | + | </p> |
| | + | <p> |
| | + | |
| | + | 5. Wang, Y., Chen, R., Cai, J., Liu, Z., Zheng, Y., Wang, H., Li, Q. and He, N., 2013. Biosynthesis and thermal properties of PHBV produced from levulinic acid by Ralstonia eutropha. <em>PLoS One</em>, <em>8</em>(4), p.e60318. |
| | + | </p> |
| | + | <p> |
| | + | |
| | + | </p> |
| | + | <p> |
| | + | |
| | + | 6. Liu, H., Pancholi, M., Stubbs Iii, J. and Raghavan, D., 2010. Influence of hydroxyvalerate composition of polyhydroxy butyrate valerate (PHBV) copolymer on bone cell viability and in vitro degradation. <em>Journal of applied polymer science</em>, <em>116</em>(6), pp.3225-3231. |
| | + | </p> |
| | + | <p> |
| | + | |
| | + | </p> |
| | + | |
| | + | </html> |
