A French student research team won a silver medal during iGEM competition (synthetic biology). They designed tool for tuberculosis diagnosis, a disease that still kills 1,5 million people each year. The diagnosis test is cheaper and relies on simple technologies which can be used in developing countries.
The iGEM (international genetically engineered machine) competition is an international contest in which different teams of scientific students from all over the world present a genetic engineering project. Here, we follow French team iGEM Aix-Marseille. They came back from Boston on October 31 with a silver medal. But how did it all start? We need to go back a few months. The iGEM competition usually begins in February when the team registration opens. Students start to pair up with other students sharing the same interests. Brainstorming after brainstorming, they manage to form a multidisciplinary team on a special research subject. Every year, the “Giant Jamboree” meetup in Boston brings together these young researchers to share their project, their methods and their passions.
The 2019 Aix-Marseille team chose a special day to announce their theme. On the international day of tuberculosis, they stormed social media with their big announcement: their project was going to be on tuberculosis diagnosis. On April 24, they posted their official logo and slogan “We want TB free!”. They also gave more detail on what they were exactly going to accomplish: “an accessible, fast and reliable tuberculosis detection kit”.
What is tuberculosis and why work on it?
Tuberculosis is a bacterial infection caused by Mycobacterium tuberculosis, also named Koch bacillus, from the researcher who discovered it. It killed 1.5 million people in 2014 according to the WHO. A third of the world population is infected, though 90% of them do not know it. Why? Tuberculosis is a vicious disease that is mostly asymptomatic in a healthy person. But as soon as they get immunosuppressed, the bacterium wakes up from its dormant state, gets out of its hidden spot in the lungs. Immunosuppression can happen in the late stage of AIDS, with cancer or diabetes, people who had a transplant, or smokers. When it is active, a contaminated patient can infect 10 to 15 people a year. The symptoms can be intense: chronic cough with expectorations sometimes bloody, chest pains, constant fever, night sweats, weight loss... Sometimes, the bacterium can affect other organs than the lungs, like kidneys or the brain.
Don’t be alarmed! Tuberculosis can be cured. But the treatment is long: six months to two years of multiple antibiotics to avoid manifestation of drug resistant bacteria. The existing vaccine only protects one out of two adults. It’s better than nothing, and also prevents 90% of tuberculous meningitis in children. To get treatment, diagnosis is mandatory. For the latent disease, it is complex because it isn’t 100% accurate since the bacteria are hidden. Though you could suppose that, with the coughing and chest pain, diagnosis for symptomatic tuberculosis is easy, it actually is not perfect either. Thoracic radio imaging can be used to detect lesions but those are not specific to TB. Then, the microbiologic test, accurate and efficient, is also 3 days long and requires expensive lab equipment- which brings us back to the Aix-Marseille team. We need more efficient diagnosis so that patients can get access to early treatment and infect less healthy persons around them.
A long journey to get diagnosed
The usual microbiological diagnosis works thanks to DNA, the identification tag of any living thing on Earth. The patient spits in a cup. If he or she is infected, bacteria should be swarming in the spit. But to know that, scientists need to make sure the bacteria tag is here. They use a technique called Polymerase Chain Reaction (PCR). The spit is put in a test tube, along with elementary DNA bricks (nucleotides) and a protein, polymerase, that helps assemble. To get them in the right order, there also needs to be a
special DNA sequence called primer, here specific to the bacterial DNA. It will stick to the DNA sample if it is the right one and allow the polymerase to work. Based on the DNA sample, it will duplicate it until there are no more bricks. It helps scientists get a signal and identify the proper tag. If it matches Mycobacterium tuberculosis’ reference, then the patient will get treated. In Europe or the United States, PCR is easy. Every lab has a thermocycler, and every hospital can send samples to be diagnosed really fast. But in developing countries, access to those methods is limited: a PCR thermocycler costs at least 8.000€. Add to this the recurrent cost of DNA bricks and it makes it impossible to get diagnosed in a country with poor healthcare.
What does the test consist in?
The test our students from Aix-Marseille designed rests on a method similar to PCR, called Polymerase Spiral Reaction (PSR). Instead of having the DNA replicate in strands, it does so on a continuous line that spirals around. For that, you need a special set of primers, specific to the Mycobacterium tuberculosis DNA and that are made to spiral. Add the bricks, the polymerase protein, a 60°C bath and you’re all set. To be sure that when the test is negative it isn’t just because it didn’t work, you need a control tube. You can use a random gene you know and its primers. Getting the DNA amplified is good, but we also need to interpret the results, which is usually expensive. They made it work like a pregnancy test. Plunging a paper test in the PSR mix and it will reveal colored lines. The bacterial DNA is recognized by a special protein, a methylase, fused with a chromoprotein. When the complex is bound to the Mycobacterium tuberculosis DNA, the chromoprotein will color the paper in green. There also are two control lines, a green one to check if the methylase works and, as seen above, a red one for the control tube. To summarize: spit in a tube, warm it at 60°C for 1 hour, dip the paper test, wait 5 minutes and that’s it. Easy as pie. So, do you want TB Free?
To learn more about the project : https://2019.igem.org/Team:Aix-Marseille