Imagine detecting pathogens in your favorite foods right in the grocery store using your mobile phone.
It might not be long before you can do this, thanks to some recent groundbreaking research at Pittsburg State University (PSU), in southeast Kansas.
Detection with Nanotechnology
Tuhina Banerjee, PhD, Santimukul Santra, PhD, and James McAfee, PhD, faculty members in the PSU Department of Chemistry, along with six students, have successfully married magnetic resonance imaging (MRI) technology and fluorescence emission to detect E. coli O157:H7 in milk and lake water. The result is hybrid nanosensors that are able to screen quickly for target pathogens.
According to Dr. Banerjee, the PSU nanosensors are composed of special iron oxide particles blended with an optical dye, plus antibodies that specifically latch onto E. coli O157:H7 cells.
“When mixed into a solution with bacterial colonies, the nanosensors swarm around their target organism’s outer membrane and adhere to any such pathogens that are present, while ignoring nontargeted cells, even other strains of E. coli, or heat inactivated O157:H7,” she explains. “This aggregation is detectable using magnetic resonance. Similar to MRI technology used in human medicine, the bacteria detection procedure launches a magnetic field through the sample. But instead of measuring water molecules, as is the action of medical MRIs, the detector picks up iron-rich nanosensor clumps.”
In the presence of a small amount of bacteria (low colony forming units, CFUs), the magneto-fluorescent nanosensors (MFnS) cluster around the bacterial cell, which inhibits their interaction with the surrounding water protons, thus increasing magnetic relaxation (T2) values, Dr. Banerjee elaborates. “However, as bacterial concentration increases (high CFUs), the clustering decreases,” she says. “This causes the T2 signal to become saturated and the ability to quantify bacterial concentration is lessened.”
One huge benefit of the PSU nanosensor technology, Dr. Banerjee says, is its capability to scan both very small amounts of a pathogen, as well as very large amounts. “Magnetic resonance has the ability to detect bacteria in small quantities, but not in large ones,” she points out. “With fluorescence, the opposite is true.”
Dr. Banerjee explains that the dye glows when the nanosensors cling to bacteria. “When there are just few colonies, then detection is mainly mediated by monitoring T2 change (magnetic relaxation) and so even a minimum amount of bacterial contaminant can be detected,” she notes. “But if there are many cells, the sample will light up. The brighter the glow, the more contaminated the sample. We can detect as low as 1 CFU using our magnetic nanosensors.
“We believe that by tweaking the antibodies, we can adapt the technique to detect a wide range of pathogens, including bacteria and viruses, in foods, beverages, water, and even human blood samples,” she continues. “We plan to test the technology on lettuce and juice next.”
Another selling point is that the PSU nanosensors are currently able to detect bacterial contamination in less than an hour. “This is much quicker than current gold standard techniques, including real-time polymerase chain reaction, which can take up to 24 hours for data collection, sample amplification, and results,” Dr. Banerjee notes.
With further experimentation, the time required to get test results is expected to be reduced to a few minutes or even seconds, Dr. Banerjee predicts. “Magnetic relaxation is a powerful technique and we start to see a light change within a few minutes when we incubate MFnS with bacterial CFUs,” she relates. “Soon we expect to do multiplex samples for other contaminants besides E. coli in the same sample, so pathogen testing will be even faster then. And if one organism is more prominent than the other, we will be able to detect the less prominent one.”
The next big step, Dr. Banerjee says, is to convert the nanosensor technology into a chip device that will make it quick, easy, and inexpensive for consumers to determine outside of the lab if food or water is contaminated with pathogens. “That will take a bit more time and collaborations with engineers, but we believe the efforts will have long term benefits for the greater society,” she emphasizes. “Besides handy use in retail settings for grocery shoppers, potential global applications include food manufacturing and water analysis in developing countries.”
Liquid Droplet Test
Researchers at Massachusetts Institute of Technology (MIT) in Cambridge have developed a new test for E. coli based on a novel type of liquid droplet that can bind to bacterial proteins.
This interaction can be detected by either the naked eye or a smartphone, according to Timothy Swager, PhD, the John D. MacArthur Professor of Chemistry at MIT and the senior author of the study. “What we have here is something that can be faster and massively cheaper that traditional pathogen tests, with low entry costs,” he says.
In 2015, Dr. Swager’s lab developed a way to easily make complex droplets, including droplets called Janus emulsions. “These Janus droplets consist of two equally sized hemispheres, one made of a fluorocarbon and one made of a hydrocarbon,” Dr. Swager explains. “Fluorocarbon is denser than hydrocarbon, so when the droplets sit on a surface, the fluorocarbon half is always at the bottom.”
Two years later, Dr. Swager, his colleagues, and students decided to explore using these droplets as sensors because of their unique optical properties. “In their natural state, the Janus droplets are transparent when viewed from above, but they appear opaque if viewed from the side because of the way that light bends as it travels through the droplets,” Dr. Swager relates.
To turn the droplets into sensors, the researchers designed a surfactant molecule containing mannose sugar to self-assemble at the hydrocarbon–water interface, which makes up the top half of the droplet surface. “These molecules can bind to lectin proteins, which are found on the surface of some strains of E. coli,” Dr. Swager points out. “When E. coli is present, the droplets attach to the proteins and become clumped together. This knocks the particles off balance, so that light hitting them scatters in many directions, and the droplets become opaque when viewed from above.”
Dr. Swager says his team is using the native molecular recognition that these pathogens use. “They recognize each other with these weak carbohydrate-lectin binding schemes,” he notes. “We took advantage of the droplets’ multivalency to increase the binding affinity, and this is something that is very different than what other sensors are using.”
To demonstrate how these droplets could be used for sensing, the MIT researchers placed them into a petri dish atop a QR code that can be scanned with a smartphone. “When E. coli are present, the droplets clump together and the QR code can’t be read,” Dr. Swager relates.
The MIT team hopes to adapt its new technology into arrays of small wells, each containing droplets customized to detect a different pathogen and linked to a different QR code. “This could enable rapid, inexpensive detection of food contamination in most any venue using only a smartphone,” Dr. Swager emphasizes.
The MIT researchers are now working on optimizing the food sample preparation so they can be placed into the wells with the droplets. “We also plan to create droplets customized with more complex sugars that would bind to different bacterial proteins,” Dr. Swager says.
In their initial work, the team used a sugar that binds to a nonpathogenic type of E. coli, but they foresee adapting the sensor to other strains of E. coli and other pathogenic bacteria, Dr. Swager mentions. Another step would be to make really selective droplets to catch different bacteria, based on the sugar one puts on them.
“We are also working to improve the sensitivity of the sensor, which currently is similar to existing techniques but has the potential to be much greater,” Dr. Swager adds. “We expect to launch a company to commercialize the technology in the late summer of 2018.”