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Quick Crispr-Based Test Detects Malaria in Blood

Malaria clinic

Malaria clinic in Mali (USAID.gov)

22 Sept. 2020. A blood test designed for low-resource field settings is shown to quickly detect and distinguish between different malaria parasites needed to accurately diagnose the disease. A team from the Wyss Institute, a biomedical engineering research center at Harvard University, describes the test in yesterday’s issue of Proceedings of the National Academy of Sciences.

Malaria, according to World Health Organization, affected 216 million people in 2016, which extracts heavy social and economic burdens in developing countries. In 2016, some 445,000 people died from malaria, of which 90 percent were in sub-Sahara Africa. Children under the age of 5 are particularly susceptible to the disease. The disease is caused by infections from the Plasmodium parasite transmitted by mosquitoes. In humans, the parasite multiplies in the liver, then infects red blood cells. Symptoms, including headache, fever, and vomiting, occur 10 to 15 days following transmission from a mosquito bite.

People with Plasmodium parasites but not showing symptoms are difficult to identify in many parts of the world, since sophisticated genomic tools are needed to reliably analyze blood samples, which are not available to many local health authorities. A team led by James Collins, part of the core biomedical engineering faculty at Wyss Institute, is seeking a simpler, yet fast and reliable technology that can be deployed in remote regions to detect the presence of Plasmodium parasites in the blood. The test also needs to distinguish between the main types of Plasmodium to accurately diagnose the disease and prescribe treatments.

The researchers adapted a diagnostic technique using the gene-editing process Crispr, short for clustered regularly interspaced short palindromic repeats. Crispr is a genome-editing process based on bacterial defense mechanisms that use RNA to identify and monitor precise locations in DNA. In this case, however, Crispr, is employed to edit RNA rather than DNA.

This extension of Crispr is called Sherlock, short for specific high-sensitivity enzymatic reporter unlocking, first developed at the Broad Institute, a joint genomics research center affiliated with Harvard and MIT. Sherlock uses Crispr editing enzymes that seek out specific genetic sequences in a specimen sample, and if detected in the sample, bind to and cut the RNA in nearby locations. In addition, Sherlock adds a reporter sequence to the RNA, a specific piece of synthetic RNA, which also gets cut by the editing enzyme, releasing a signal to identify the presence of the original target sequence. These reporter sequence signals can then be converted into a bioluminescent visual display that appears on an everyday material like paper and at room temperature.

The team further engineered the Sherlock process for use in low-resource field settings where refrigeration or even reliable electrical power are not available. Their process begins with a 10-minute blood sample-preparation stage with freeze-dried materials im ambient conditions, followed by analysis of the samples with Sherlock that provide either fluorescent or lateral flow strip, such as paper, readouts in about 60 minutes.

Tests in the lab with actual clinical blood samples accurately detected falciparum and vivax Plasmodium parasites with 100 percent true-positive sensitivity and 100 percent true-negative specificity. In addition, the tests were able to detect low volumes of parasites, meeting WHO’s recommended two parasites per microliter of blood detection target.

“This field-ready Sherlock diagnostic malaria assay surpasses the sensitivity and specificity requirements set by the WHO for a desired test that can be used to detect low parasite density in asymptomatic carriers of all major Plasmodium species,” says Collins in a Wyss Institute statement. “Its highly streamlined design could provide a viable solution to the present diagnostic bottleneck on the path to eliminate malaria, and more generally enable malaria surveillance in low-resource settings.”

A spin-off company, Sherlock Biosciences, licenses the Sherlock technology for fast diagnostics in the field, including for Covid-19, as reported by Science & Enterprise. However, the company is not developing an application for malaria, according to the competing interest statement in the journal article.

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