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CDC, Senegal Researchers Find 'Much Needed Alternative' to PCR in Meridian Malaria Assay

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NEW YORK (GenomeWeb) – Researchers at the US Centers for Disease Control and Prevention and the University Cheikh Anta Diop (UCAD) in Senegal have published an evaluation of the Illumigene Malaria test, describing it as a "much needed alternative" to standard PCR-based assays.

Published earlier this month in Scientific Reports, the study compared the performance of Meridian's loop mediated isothermal amplification (LAMP) test to a real-time PCR malaria reference test using laboratory and clinical samples. The researchers also assessed different sample prep methods for the test.

Close to half a million people die each year from malaria parasite infection, but eradication is ultimately an achievable goal, Naomi Lucchi, a researcher in the malaria branch of the CDC's center for global health division of parasitic diseases and a corresponding author on the study, said in an interview.

"If you are going to eradicate malaria ... you need sensitive tools to be able to detect that last parasite," Lucchi said.

The Illumigene Malaria test, she said, adds to a toolbox that includes lab-based PCR and field-deployable rapid diagnostic tests, or RDTs, which are typically lateral flow immunoassays.

Meridian's test can detect as little as one parasite per microliter of blood, while RDTs typically require a burden of 200 parasites per microliter or more to show a positive result.

She noted that community screening, which is a strategy that could be implemented in an eradication program, would likely yield many asymptomatic carriers of the parasite in malaria-endemic regions. "Microscopy and rapid diagnostic tests would not be able to pick those up," she said.

In addition, the test could find an application in follow-up tests after treatment or in vaccine trials, according to the study.

Meridian's test was CE marked earlier this year. The test is the tenth assay on the Illumigene platform, including others for Clostridium difficileGroup B StrepGroup A StrepMycoplasma pneumoniaeBordetella pertussis, HSV 1 and 2, chlamydia, and gonorrhea.

At the time of CE marking, Daouda Ndiaye, a parasitology researcher at UCAD who is also co-first author on the Scientific Reports study, noted the test could provide benefits in terms of diagnosing cases sooner, presumably at lower parasitic loads.

"Earlier diagnosis enables the correct treatment to be prescribed, which leads to better clinical outcomes for the person with malaria and keeps malaria treatments for the right people," Ndiaye said.

Lucchi said that Meridian had been meeting with the CDC for some time to discuss the malaria LAMP test in development, and the collaboration with Ndiaye's group in Senegal grew from CDC's connections.

"This had to be evaluated somewhere in the field. ... Being at the CDC, we have a lot of collaborators outside the US in endemic countries, and we proposed that they could try and work with Dr. Ndiaye," she said.

"I think the product is pretty promising," Lucchi said, noting that she had been working on LAMP assays for five or six years.

"The bottleneck was mostly the sample prep, that was always our biggest problem" she said. "When Meridian contacted us and they had this product, it really answered a lot of what we were looking for."

Besides simple sample prep — a lysis buffer-based method which takes 10 minutes from whole blood, as opposed to two hours using centrifuges and columns in standard methods — the LAMP assay can be done in any lab with access to electricity.

"It is much simpler [and] the whole workflow is very easy," Lucchi said, adding, "In the field, basically anybody can run it after a few hours training."

Furthermore, the test reagents are lyophilized, eliminating the need for cold chain transport or cold storage.

The sensitivity of the test is another pro, Lucchi said. Compared to microscopic examination looking for parasites in blood smears or to a qPCR test using two different sample prep methods, the test demonstrated specificity in the 88 percent to 99 percent range, and sensitivity of between 94 and 97 percent.

The study also showed the test was able to perform well on 209 clinical samples of whole blood in the field compared to dried blood spots that were used to confirm the results at the CDC using PCR. In this work, the researchers used venous blood, but the study argues that finger prick blood could also be effective.

The only possible drawbacks may be in the number of samples per run — which is currently limited to 10 — the requirement for electricity, and possibly cost in some regions. However, these paled in comparison to the advantages and could be overcome, in Lucchi's opinion. For example, to use the platform to run a large-scale testing program, customers could purchase two or three machines to increase throughput.  

Eradication is a tall order, she said, but efforts funded by the Bill and Melinda Gates Foundation and increased use of RDTs has been reducing the burden of malaria globally.

The CDC does not endorse or recommend specific tests, and Lucchi said the study is meant to encourage replication in other parts of the world, as well as World Health Organization and local health ministry evaluation. Assessing the test in a low-endemic region as opposed to highly endemic places like Senegal would be important, for example.

But there is still plenty of usefulness in RDTs, Lucchi said, and they have reduced the malaria burden globally via so-called "test and treat" programs. But the sensitivity is not as good, and people who are transmitting the parasite but carrying low levels can be missed.

For example, several recent studies, including one published in the Journal of Infectious Diseases last week, have uncovered strains of the malaria parasite that resist detection by RDTs. These strains have deleted the histidine-rich protein 2, and are now found in Congo, Ethiopia, and parts of South America, Lucchi said.

"You can imagine you are using these tests, and if the gene is not there you get a negative result," she said.

Yet, RDTs are "still the way forward" because of the ease of use, particularly if they can be made more sensitive and if the target can be changed to a marker that is not deleted in any strains, Lucchi said.

"Because we are not there yet, something like LAMP becomes an option because it is simple, and it is sensitive," she said.