In many developing countries, people depend on reasonably priced and conveniently available street food. However, lack of action taken by governments to regulate street food vendors has led to the prevalence of severe street food-related illnesses. One of the primary microbial contaminants in street food is E. coli O157:H7, which acts by secreting the Shiga-like toxin (SLT). Currently, there is no detection method for SLT outside of a lab setting, thus putting the consumers of foods at risk. Our project aims to develop a device that would be used by street vendors and restaurant owners to verify the safety of their products. Through our device, we exploit the binding of Gb3 to subunit B of the Shiga toxin, and compare the migration pattern of the bound Gb3-subunit B complex to a non bound subunit B. A shift in the migration pattern on a PAGE gel will occur when Gb3 is bound, indicating the presence of the toxin in the food sample. If no shift occurs in the SLT migration pattern, this implies the absence of the toxin within the sample, and reflects the safety status of the food.
In many developing countries, people depend on reasonably priced and conveniently available street food. However, lack of action taken by governments to regulate street food vendors has led to the prevalence of severe street food-related illnesses. One of the primary microbial contaminants in street food is E. coli O157:H7, which acts by secreting the Shiga-like toxin (SLT). Currently, there is no detection method for SLT outside of a lab setting, thus putting the consumers of foods at risk. Our project aims to develop a device that would be used by street vendors and restaurant owners to verify the safety of their products. Through our device, we exploit the binding of Gb3 to subunit B of the Shiga toxin, and compare the migration pattern of the bound Gb3-subunit B complex to a non bound subunit B. A shift in the migration pattern on a PAGE gel will occur when Gb3 is bound, indicating the presence of the toxin in the food sample. If no shift occurs in the SLT migration pattern, this implies the absence of the toxin within the sample, and reflects the safety status of the food.
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Revision as of 12:16, 26 October 2017
Why is STEC a major concern?
Shiga toxin-producing
Escherichia coli (STEC) causes over 70,000 infections per year in the United States alone. A portion
of these individuals will experience kidney failure after 6 days, 50% of which will require renal replacement
therapy.[1] Conventional pathogen detection requires a laboratory setting using PCR, which requires multiple
annealing and extension steps that can take over 3 hours and involves expensive equipment.
Mode of Action
Symptoms of STEC infections
What are we doing?
Due to these difficulties in validating their prototype, and based on responses from food vendors in Pakistan and Indonesia,
we have produced a rapid, affordable, portable device that allows for the detection of STEC using loop-mediated
isothermal amplification (LAMP). This is a highly specific, efficient and rapid DNA amplification technique
that uses 4-6 primers that bind to 6-8 distinct regions of target DNA. The selectivity of our system was
tested using the rfbE gene, a non-toxic coding sequence required for O157-antigen synthesis. Reagents were
lyophilized into distinct sample wells in a PDMS chip, and sample DNA from broth inoculated with rfbE+ DH5alpha was introduced into
the system. The 65 ºC temperature was achieved using a Peltier Modular Cooling system with a 6 Volts, 1.5 Amperes external power supply adapter. The reaction was visualized under UV and blue light. It is envisioned that a smartphone will suffice for capturing the output of the reaction.
How are we doing it?
Why E.coLAMP?
iGEM NYU Abu Dhabi 2016 Project
Award: Silver medal
In many developing countries, people depend on reasonably priced and conveniently available street food. However, lack of action taken by governments to regulate street food vendors has led to the prevalence of severe street food-related illnesses. One of the primary microbial contaminants in street food is E. coli O157:H7, which acts by secreting the Shiga-like toxin (SLT). Currently, there is no detection method for SLT outside of a lab setting, thus putting the consumers of foods at risk. Our project aims to develop a device that would be used by street vendors and restaurant owners to verify the safety of their products. Through our device, we exploit the binding of Gb3 to subunit B of the Shiga toxin, and compare the migration pattern of the bound Gb3-subunit B complex to a non bound subunit B. A shift in the migration pattern on a PAGE gel will occur when Gb3 is bound, indicating the presence of the toxin in the food sample. If no shift occurs in the SLT migration pattern, this implies the absence of the toxin within the sample, and reflects the safety status of the food.
Our improvements
Shiga toxin is an exotoxin that consists of two subunits. Subunit B binds to Gb3 receptor expressed in the surface of target cells and permits the entry of subunit A, which inhibits protein synthesis.[2] 2016 Team NYU Abu Dhabi exploited the binding of Gb3 to subunit
B to detect for the presence of STEC. Their prototype compared the migration pattern of a bound Gb3-subunit
B complex to that of free subunit B using a PAGE gel. Their device was estimated to take 45 minutes and their
prototyping process ran into several issues that negatively impacted the specificity, affordability, and accessibility of the product.
[1] Borgatta, B.; Kmet-Lunaček, N.; Rello, J., E. coli O104:H4 outbreak and haemolytic–uraemic syndrome. Medicina Intensiva (English Edition) 2012, 36 (8), 576-583.
[2] Pacheco, A. R., Sperandio, V., Shiga toxin in enterohermorrhagic E. coli: regulation and novel anti-virulence strategies. Front. Cell. Infect. Microbiol. 2012, 2 (81), 1-12.
[3] New England BioLabs. Isothermal Amplification. Accessed October 19, 2017.
In many developing countries, people depend on reasonably priced and conveniently available street food. However, lack of action taken by governments to regulate street food vendors has led to the prevalence of severe street food-related illnesses. One of the primary microbial contaminants in street food is E. coli O157:H7, which acts by secreting the Shiga-like toxin (SLT). Currently, there is no detection method for SLT outside of a lab setting, thus putting the consumers of foods at risk. Our project aims to develop a device that would be used by street vendors and restaurant owners to verify the safety of their products. Through our device, we exploit the binding of Gb3 to subunit B of the Shiga toxin, and compare the migration pattern of the bound Gb3-subunit B complex to a non bound subunit B. A shift in the migration pattern on a PAGE gel will occur when Gb3 is bound, indicating the presence of the toxin in the food sample. If no shift occurs in the SLT migration pattern, this implies the absence of the toxin within the sample, and reflects the safety status of the food.
