By Paul Nicolaus
September 12, 2018 | New research is revealing that it may be possible to diagnose malaria in an unexpected way, and the hope is that it could lead to a non-invasive, breath-based field test with advantages over existing detection methods. At the heart of it all is a scientist, a grand vision, and the scent of sickness.
Many bacteria, fungi, and parasites produce smells just like flowers and fruits, according to Washington University in St. Louis microbiologist and infectious disease expert Audrey Odom John, and the flavor and fragrance industry already makes use of high-tech mass spectrometers to identify the compounds that contain the typical characteristics of foods, drinks, and cosmetics.
In other words, we know why an orange smells like an orange and a lemon like a lemon, she explained during a brief TEDx Talk delivered in 2015. Although mass spectrometers are too large and complex for use in typical care settings, devices can get progressively smaller once it is clear what you are looking for.
Her plan? Use this same type of flavor sniffing to figure out what certain diseases smell like. The technology has already arrived, after all. “All we need to do,” Odom John said, holding up a pocket-sized breathalyzer, “is tune the sensors to detect infections rather than alcohol.”
Since delivering this talk, she and her research team have taken steps to home in on malaria in particular—a global health concern that continues to impact hundreds of millions every year and a disease that appears to be a prime candidate for breath-based diagnosis.
Early Research Charts Path Forward
In recent years they have found, for example, that the malaria parasite releases volatile compounds in cultured red blood cells, Odom John told Diagnostics World, which led to the hypothesis that the same kind of volatile compounds produced in vitro would be found in human patients.
Their hunch does build upon the work of other researchers. There was a small study of adults who were intentionally infected with malaria parasites in order to study new anti-malarial drugs, for example. Although the researchers didn’t allow the participants to get sick, they did allow a small amount of parasites to enter the bloodstream, she explained, and discovered that some volatiles were different in those adults.
She and colleagues have also taken an interest in studies focused on mosquito behavior. In humans, mice, and birds, individuals infected with the malaria parasite are more attractive to mosquitos than the uninfected, which suggests that mosquitos can smell the difference between who has malaria and who doesn’t. If mosquitos can tell the difference, maybe a mass spectrometer could, too, they figured.
To explore further, Odom John and her research team carried out a field study in the African country of Malawi to identify compounds that are present in the breath of children with malaria. Breath volatiles from 17 children with and 18 children without uncomplicated Plasmodium falciparum malarial infection were collected. (For those who were infected, the average parasitemia level was 2.2% of erythrocytes.)
Participants blew into a chemically inert bag, which was then pumped over a small column that picks up compounds in the air. Those columns were shipped back to the United States, where a mass spectrometer performed the analysis.
The findings, published this year in The Journal of Infectious Diseases (DOI: 10.1093/infdis/jiy072), revealed that there were global changes to the compounds that are present in the breath of children with malaria compared to children who have a fever but do not have this disease. Although no single compound served as a classifier on its own, the use of six—methyl undecane, dimethyl decane, trimethyl hexane, nonanal, isoprene, and tridecane—led to the classification of malarial infection status with 83% accuracy (94% specificity and 71% sensitivity).
The study also revealed that there are higher levels of mosquito-attracting compounds in the breath of children with malaria, suggesting that parasite infection increases the likelihood of getting bit by a mosquito. In other words, the parasites may actually be intentionally hijacking mosquito behavior to increase transmission.
Overall, the results are promising, according to David Bell, a clinical and public health physician with a background in malariology, tropical health, and diagnostics product development who was not involved with the study. By using high-tech equipment in a centralized lab, the researchers were able to distinguish heavily-infected children from non-infected children with reasonable accuracy (although he pointed out that this was accomplished with a relatively costly collection system compared to current tests).
“This suggests an interesting direction for research, to determine whether sensitivity of detection of these compounds can be improved and ultimately simplified so that a non-invasive field test might be developed,” Bell, the director of global health technology for Intellectual Ventures and the Gates-backed Global Good, explained via email.
The results indicate we are still a long way from something practical in the field, he pointed out, but, “if a non-invasive test for malaria could be developed that had similar accuracy to current blood-based tests, and at a similar cost and simplicity, then this would be an important step forward in closing the gap on malaria diagnosis.”
The Move Away from Blood
Today, malaria diagnosis continues to revolve around blood. The demonstration of the parasitic stages in blood smears remains the gold standard, according to Satesh Bidaisee, professor of public health and preventive medicine at St. George's University in Grenada, West Indies.
Rapid diagnostic tests (RDTs) are another option. These use a finger prick blood sample and can be carried out in roughly 15 minutes by individuals with little training using test kits that don’t require electricity or specialized equipment.
However, a blood smear can be challenging to carry out in rural and resource-poor areas of the world where there may not be electricity or skilled technicians. And RDTs come with disadvantages, too. They are prone to false positives because the parasite-derived antigen can take weeks to leave the bloodstream after an infection is cured.
Compared to microscopy, RDTs lack sensitivity at low levels of parasitemia, Bidaisee noted, so those with early or mild infections may receive a negative diagnostic result. RDTs also lack the ability to quantify parasite density, which provides important information on the severity of infection and the response to treatment through changes in parasitic load.
The most widespread tests work by detecting the deadliest form of malaria, which is caused by Plasmodium falciparum, but “the parasite is exceptionally tricky,” Odom John explained, and in some areas where the tests have been widely used the parasite has actually stopped making the HRP2 protein that they are designed to detect.
The modeling suggests that parasites that do not make this protein and cannot be detected by the tests are spreading and will be widespread across Africa by 2030. “And if that’s the case,” she said, “we pretty urgently need to develop a new diagnostic test for malaria because we’re about to lose the best one we have.”
One of the biggest draws of a malaria breathalyzer would be the move away from blood samples and a move toward greater simplicity, but there are questions that remain. Because the Malawi patients were similar in respect to ethnicity, location, and diet, the findings need to be validated in different locations with different populations. At the moment, there are plans to follow up with a second cohort in Malawi and possibly another cohort in Kenya to build upon the initial results.
“We believe that these are compounds that are consistent across all parasites and across all people,” said Washington University pediatrician Indi Trehan, who has worked in rural Africa and southeast Asia for over a decade and led the breath collection aspect of the study, “but since we don’t know the exact biochemical pathway that creates this, we can’t say that for sure.” The research conducted so far has proven that the compounds are there, but “it doesn’t at all confirm that the compounds are there in a quantity that is easily testable by a small electronic device.”
Developing a diagnostic tool remains the ultimate goal, however. The research team includes Barani Raman, a Washington University bioengineer. “We’re optimistic that he can come up with a device that can find the compounds,” Trehan said, “and our job is to sort of get closer to finding which compounds it is we want him to find.”
As Trehan describes it, the hope is to come up with a reusable device roughly the size of a cell phone that would require little more than the occasional replacement of AA batteries (and not even that if a rechargeable solar battery could be incorporated). A person could breathe into it similar to an alcohol breathalyzer, and it could potentially fit over the nose, too, so that sufficient samples could be collected from comatose patients with cerebral malaria. Ideally, Trehan added, it would work for kids, pregnant women, and adults.
Cost would, of course, be a central issue. “It really depends on how many volatiles we have to detect and how stable the physical device is,” Odom John said. The goal would be to get down to less than 50 cents per test, so if a device can be used for years in a high humidity, high-temperature field environment, then the cost per use could be quite low. If the device can’t hold up for a lengthy duration, however, it wouldn’t be a reasonable investment.
So could breath be the right type of sample to use for the diagnosis of malaria? The answer isn’t clear just yet. At this stage, there is still a need to determine which volatiles are common to all infections and which ones are common to specific infections. “I think understanding the biological origin of these volatiles will tell us a lot about whether or not we can expect the kind of specificity that we’d like to have in a diagnostic test,” she added.
Paul Nicolaus is a freelance writer specializing in science, nature, and health. Learn more at www.nicolauswriting.com.