WEIRD BUGS IN YOUR BRAIN?
Tapeworms and mould fungi could attack your brain and make you fatally ill. Often, doctors do not identify the attackers and are forced to "fire at random". However, a new, ground-breaking genetic method can now reveal the lethal organisms, before it is t
Feeling a bit off? It mightn’t be your attitude, it might just be the baby tapeworm lodged in your actual brain. Yes, it’s gross.
Doctors are unable to find out what is wrong with the 14-year-old girl. She is admitted to hospital in the summer of 2015 with severe fever and headache. After a few days, she becomes delirious, only speaks in brief sentences of no more than three words, and does not react, when spoken to. The symptoms indicate a type of meningitis, which could be caused by 100+ different pathogenic organisms, that all require different treatments. Scientists must find the cause to be able to give the girl the right drug, and so, they initiate a series of tests.
Each test is customized to reveal the presence of one single type of organism, and doctors test for one after the other, hoping to finally get a positive result. However, all test results are negative. The girl is infected with something that only rarely attacks the brain. There might be a test for the organism, but doctors have no idea what to look for. After 19 days, they give up and decide to send a sample of the girl’s cerebrospinal fluid to biochemist Joe DeRisi’s research laboratory at the University of California, San Francisco, USA, where a new test is in the process of revolutionizing medical science.
Test identifies microbes
According to DeRisi, it is “the mother of all tests”. Unlike the tests that doctors use, it is not meant to show the presence of one specific organism. Instead, it scans the sample to find evidence of everything that should not be there, and it does not distinguish between bacteria, fungi, animals, or other parasites.
The most important thing about the test is, however, that it does not only sound the alarm, when it spots an unauthorised organism in the sample. It immediately names the intruder, allowing the doctor to make a diagnosis and start the right treatment. It corresponds to a
burglar alarm, which immediately identifies the burglar and sends his name and address directly to the police.
As the girl’s symptoms gradually get worse, including cramps, stiffness, and trembling arms and legs, Joe DeRisi and his colleague, Michael Wilson, begin their analyses. The cerebrospinal fluid contains salts, sugar, hormones, fat, and a few proteins and white blood cells, but the two scientists are looking for something completely different. They are interested in evidence of genes that have escaped from cells in the young girl’s brain.
When brain cells die, they are broken down, and the residual products are drained from the brain with the cerebrospinal fluid. So, this is where to find evidence of the cells’ genetic material – not only of genes from the girl herself, but also from any contagious organism which might have invaded her brain. And that is the very "alien" genes that DeRisi and Wilson are looking for.
Scientists helped 300 patients
With DeRisi’s new test, doctors no longer need to try to guess what caused the infection and make a test for every guess. Valuable time could be wasted in this way, and like in the case of the 14-year-old girl, there is a risk that the many tests will never lead to a positive result. That is why in more than half of all cases of meningitis or cerebrospinal meningitis, doctors never manage to name the pathogenic organism.
Identifying the cause of the disease is key to the patients’ health, as whereas brain infections caused by bacteria can be combated with antibiotics, these drugs are ineffective in connection with virus, and fungicides are no good, when it comes to meningitis which is caused by a parasitic worm.
In 2013, DeRisi and Wilson first used the new test to identify an organism that had infected the brain of a patient who doctors were unable to diagnose in the normal way. Since then, the two scientists and several of their colleagues have helped more than 300 patients and their doctors establish difficult diagnoses.
With the assistance of physician Charles Chiu, who is also from the University of California, San Francisco, and others, they have systemized the analysis method, so it is more or less automatic. And since July 2017, American doctors have been able to order an analysis for their patients over the Internet at a price of some $3,000. The doctor only needs to take a sample from the patient, freeze it, and send it to San Francisco. And only 72 hours after the sample has arrived, the doctor receives the name of the organism.
Genome editor tidies up DNA
In the lab, DeRisi and his team are now analysing the sample from the 14-year-old girl. The small quantity of liquid contains millions of DNA sequences, but the scientists are only interested in a small handful of them – the ones coming from the pathogenic organism. So, the scientists aim to screen out the girl’s own DNA sequences, and that is done by means of the new CRISPR genome editing technology.
CRISPR is like a focused molecular editing tool that can be designed to cut up specific DNA sequences. DeRisi’s team has chosen to cut 266 different sequences, which are very common in people, but rarely observed in other organisms. All gene fragments with these typically human sequences are cut up and screened out during the very first steps of the process. So, a large quantity of genetic "noise" is removed from the sample, and the alien gene fragments stand out more clearly. According to DeRisi’s experiment, this process removes about one third of the unwanted DNA sequences from the specimen, making the test four times better at identifying the alien organism’s DNA.
In spite of the clear-out, the sample still contains almost eight million DNA sequences, of which more than 99.99 % belong to the girl herself. The scientists now need to read every single DNA sequence very accurately to find the small handful of sequences from the pathogenic organism. Previously, this would have required that the DNA sequences be read one at a time via a manual, time-consuming process. But a new method known as metagenomic nextgeneration sequencing (mNGS) makes it possible to sequence about 20 million DNA sequences at the same time and make the computer register every single sequence.
In short, the sequencing takes place by scientists peeling one strand off all DNA sequences, causing enzymes to start recreating the missing strands. Subsequently, a camera and a computer can monitor the recreation of every single strand. The equipment can determine which DNA bases are added and in which order. The result is that the computer registers the sequence of DNA bases of every single DNA strand.
DeRisi’s team now has a vast quantity of data – millions of sequences consisting of the letters A, T, C, and G. The next step is another round of clear-out to remove the remaining human genes. By means of sophisticated algorithms, a computer filters millions of sequences per hour. It compares the sequences to huge databases of human genes, allowing it to spot the girl’s DNA and filter it out. Finally, the scientists only have a small handful of sequences left. They are compared to known sequences from thousands of organisms, until DeRisi has a short, but complete list of all the life forms that have left evidence in the girl’s brain.
Mosquito carried rare disease
The list contains four organisms: a virus that only attacks tobacco plants, a blue-green alga, a soil bacterium, and the West Nile virus. The three first ones are harmless and very probably did not cause the infection in the girl’s brain. The fourth one, the West Nile virus, is very different. It is closely related to the Zika virus, dengue virus, and yellow fever virus, and it often attacks humans. Normally, the West Nile virus only causes flu-like symptoms, but in very rare cases, the virus is able to penetrate the brain’s protective barriers and attack brain cells.
DeRisi’s team immediately contact the girl’s doctors. They make an old-fashioned, targeted test, which is specifically designed to identify the presence of West Nile virus in the girl. The test is positive. DeRisi’s ground-breaking method has once again "hit the mark", and everything begins to make sense. Two days before the girl's first symptoms appeared, she was at a summer camp by a small lake in a national park outside Los Angeles. And when she came to the hospital, she had a large mosquito bite on her leg. None of the doctors paid any attention to it at the time, but the virus probably entered her body via an infected mosquito.
Today, there is no efficient treatment against the West Nile virus, but the girl’s disease has stabilized. She has gone back to school, and she is doing well – but she still has problems with speech and with her balance. Future drugs might ensure that she will get completely rid of the alien organism. However, DeRisi’s diagnosis already plays an important role for her, preventing that, due to ignorance, doctors would treat her with potentially hazardous drugs that have no effect on the disease.
Before the diagnosis, doctors had treated the girl with high doses of at least five different types of antibiotics and drugs against herpes and Epstein-Barr virus. All the drugs involved a long series of potential side effects, but none of them were efficient against West Nile virus. If DeRisi’s test had been as easily accessible to doctors as it is now, the diagnosis could have been made several weeks previously – and the
girl's chances of a quick recovery would have been improved.
Test also reveals cancer
So far, doctors are only allowed to use the test to analyse cerebrospinal fluid, so until further notice, it has only been used to diagnose brain infections. However, scientists throughout the world are struggling to show that the method can be used in connection with a long series of other diseases. Over the past five years, the annual number of scientific articles about the subject has been quintupled.
According to plan, doctors will also be able to have blood, tissue, urine, faeces, and slime samples from the respiratory organs analysed, meaning that in a matter of a few days, they will be able to find the cause of lethal cases of pneumonia or blood poisoning, which could both be caused by very different types of organisms.
DeRisi’s team has embarked on a new project involving eye infections, which are also difficult to diagnose in the traditional way. One of the patients who the scientists have examined had an eye infection for 16 years and had been checked by several doctors without an accurate diagnosis being established. The two scientists from San Francisco took a sample from the patient’s eyeball, and out of a total of 12,111,540 gene fragments, 10 turned out to come from the rubella virus, that normally causes measles.
However, the method is by no means limited to infectious diseases. It can also be used to reveal diseases caused by mutations in the patient’s own genes. In principle, scientists use the same method, but instead of comparing gene fragments in the sample to DNA sequences from alien organisms, they compare them to known, pathogenic mutations. In 2016, scientist David Dyment used the method to reveal the genetic cause of a number of otherwise inexplicable diseases in eight newborn children.
The most harmful genetic disease of all, cancer, is also under fire. The disease might develop as a result of inherited or new mutations, and previously, doctors have been forced to go through the patient’s genes one after the other to find the mutations. The new method scans thousands of genes at a time, and scientists are already well on their way to using DeRisi's method to make quick diagnoses of lung, prostate, and birthmark cancer.