Team:WLC-Milwaukee/Background
Background
Bacteriophage Based Biosensors
While our use of bacteriophages or their receptor binding proteins (RBPs) as a biosensor is not a novel technique, our bacteriophage tail biosensor is essential because it allows for the detection of E. coli using a colorimetric assay. Some previous studies using bacteriophages as a biosensor have required the use of fluorescence and portable flow-cytometry for detection [1]. While techniques such as these are highly sensitive, the instruments required for such detection are rarely accessible to the average person. Derda et.al. developed a technique which requires that the bacteria be infected with the bacteriophage before detection can take place [2]. However, our concept does not require and indeed it is incapable of infection. Thus, if any E. coli is present in a sample, our bacteriophage sensor won’t affect the potential signal. In addition, our sensor will rely on a substrate and the use of horse radish peroxidase (HRP) to produce a detectable color change. This color change could either be observed with the naked eye at a testing site or quantified in a laboratory using photospectroscopy. Ultimately, there is a wide variety of bacteriophage based biosensor techniques that have been documented in the scientific literature [3]. However, very few of these diagnostic techniques have made any major impacts either in water testing or in other applicable fields such as medicine, agriculture, and food science [3]. Our project simply aims to provide a fast and easy to use alternative.
[1] http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0131466
[2] http://pubs.acs.org/doi/abs/10.1021/ac400961b
[3] https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3442824/
Water Contamination
Two billion people worldwide are utilizing contaminated drinking water [3]. Water can be easily contaminated, so the importance of testing drinking water is crucial. Contaminated water, namely that which is polluted with fecal matter, is a critical issue because it is a source of bacteria which can cause serious diseases. Water can be contaminated with untreated waste in a variety of ways. Storms can cause sewers to overflow, allowing untreated matter to run off into rivers. The Journal Sentinel reported that last year, “109.4 million gallons of untreated wastewater spilled into local waterways and Lake Michigan in overflows of combined sanitary and storm sewers during two heavy rain storms,” [4]. Heavy rainfall can also cause runoff in fields fertilized with manure. Pollution can also come from wild animals [2]. Because flooding and the instincts of wild animals cannot be controlled, this makes water contamination an especially important issue. Water contaminated with fecal matter contains a high level of bacteria. Since not every single type of bacteria can be monitored effectively, it is helpful to use an indicator. In the United States, testing for indicators is used for checking if drinking water is safe from a microbiological standpoint, ensuring if the water was treated correctly, ensuring that the water maintains its status of treatment as it is distributed, and ensuring that recreational water is safe [6]. The presence of E. coli is usually indicative of the presence of other pathogenic bacteria that are found in fecal matter [2]. These pathogenic bacteria can cause a number of diseases, some of which result in death. One common side effect of polluted water is diarrhea. According to the head of the World Health Organization’s health department, 500,000 deaths are caused by diarrhea from contaminated water every year [3]. More serious diseases include Cholera, Dysentery, Typhoid, Hepatitis A, Bacterial Gastroenteritis, and Polio [2, 3]. Some different diseases can be caused by contaminated water in tropical areas, such as Intestinal Worms, Schistosomiasis, and Trachoma [3]. There are vaccines available for some of these diseases, but being aware of the cleanliness of the water people are drinking is a better prevention measure.
[1] https://cfpub.epa.gov/si/si_public_record_report.cfm?dirEntryId=196784
[2] http://www.water-research.net/index.php/e-coli-in-water
[3] http://www.dw.com/en/new-who-report-raises-alarm-on-dirty-water/a-38408329
[6] https://www.ncbi.nlm.nih.gov/books/NBK215658/
Water Testing in Developing Countries
Water testing in developing countries is a problem. Most countries do some water testing, as it is required by law. However, they do not usually have the resources to test all drinking water sources. Rural areas in particular are often neglected by water testing. To be more efficient with money, countries spend most of their resources sampling and testing the water in the main pipelines that residents of the urban community use. Rural communities in developing countries do not always use pipelines, making it slightly more burdensome to have to test, so these tests can be neglected [1]. Additionally, each rural drinking water source serves a minimal amount of people, so using precious resources to benefit such a small group could seem wasteful from a budget perspective.
In the World Health Organization’s water testing guidelines, it mentions that for most water supplies, bacteriological testing should be done once initially and then only needs to be tested if the situation calls for it [2]. The situations that could require follow-up tests include, “change in environmental conditions, outbreak of waterborne disease, or increase in incidence of waterborne diseases” [2]. The issue here is that people may get sick with diseases associated with water-borne pathogens, but they may not test the water to know for sure that the water is not safe. Then, people will continue to drink unsafe water, putting them at risk of contracting a serious disease. This same problem can also happen with “improved water sources”. Improved water sources mean that they are a step up from where they were previously, but to imply that they are uncontaminated is a dangerous assumption [3].
The economic effect of water testing and providing clean water is also of high importance. A developing country’s resources can be put toward other important necessities, but then the people become less productive because they are incapacitated with illness [6]. This correlates with data suggesting that developing countries with clean water sources tend to have more economic growth than the poor countries that survive on contaminated water. In order to complete the long term goal of economic growth, one must start with clean water. Clean water will eliminate many types of sickness and keep the citizens of developing countries productive. In order to purify the water, it all starts with inexpensive testing to identify the contaminated water.
[1] https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4113879/
[2] http://www.who.int/water_sanitation_health/dwq/2edvol3d.pdf
[3] http://www.nationalacademies.org/hmd/~/media/0FF50A934DE64F8C9F1352CAF9BFD2E4.ashx
[5] http://pacinst.org/wp-content/uploads/2013/02/water_quality_facts_and_stats3.pdf
[6] http://www.who.int/mediacentre/factsheets/fs391/en/
Lambda Phage
Lambda phage is a virus that can infect E.coli by recognizing LamB, an integral protein that is embedded in the outer membrane of the E.coli. Lambda phage has siphoviridae morphology which consists of an icosahedral head and a noncontractile tail. The icosahedral head is typically the compartment where the virus stores their genetic material, and this is the case for Lambda phage. The non-contractile tail is used to detect and bind the specific target to inject its genetic material into the host, thereby infecting them [1]. The Lambda phage genome is about 48500 base pairs long. Lambda phage is one of the most studied bacteriophages that have been utilized as a model for biology to explore biological processes at the molecular level. As a model system, Lambda phages not only help us better understand the mechanisms behind biological processes, but also allows us to determine how this virus determines its own fate. Lambda phages are also known to be one of the temperate phages that can choose to be either in the lytic or lysogenic state. In their reproduction system, they utilize the lac operon to control which state they are in. In the lysogenic phase, the virus is in its prophage state, which stays inactivated. In lytic phase, the phages turn on the gene to activate the lac operon [3]. As mentioned above, Lambda phage infects E.coli and recognizes the membrane protein LamB with the noncontractile tail, also referred to as the phage tail. LamB is an integral outer membrane protein in E.coli. One of the major roles of the LamB protein is maltose uptake. In corresponding to the LamB protein in the outer membrane, there is also an IICman—IIDman complex of mannose transporter in the inner membrane. The two portions together make up for both the maltose and mannose transport, and the infection of Lambda phage. [4]
[1] https://www.sciencedirect.com/science/article/pii/B9780123739445000201
[2] http://www.sciencedirect.com/topics/neuroscience/lambda-phage
[3] https://www.ncbi.nlm.nih.gov/books/NBK21856/
[4] https://www.caister.com/jmmb/v/v3/v3n3/05.pdf
[5] http://onlinelibrary.wiley.com/doi/10.1111/j.13652958.2004.04242.x/abstract
