CRISPR-Based Assay for Rapid and Inexpensive Malaria Testing

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Researchers at the Harvard Wyss Institute have developed a rapid, inexpensive, and very sensitive malaria test that harnesses the power of CRISPR. The new technique does not require complicated sample preparation or processing, and it could allow for rapid on-site testing in low resource regions, aiding the campaign to eliminate malaria.

Determining where malaria is being contracted is key to eradicating and treating it. However, such regions are usually remote and typically lack medical resources, making testing challenging. Current tests for malaria have poor sensitivity and often require expensive and cumbersome lab equipment, making them unsuitable for use in the field.

“Unfortunately, available rapid diagnostic approaches cannot distinguish all four Plasmodium species from one other, which can be important to initiate the definitive course of treatment, and, most importantly, they are ineffective for detecting low numbers of Plasmodium parasites in asymptomatic individuals,” said Nira Pollock, one of the researchers that developed the new assay.

“These ‘asymptomatic carriers’ are silent reservoirs for ongoing transmission by malaria-spreading mosquitoes and extremely important for ongoing global efforts to eradicate malaria,” added Jeffrey Dvorin, another researcher involved in the study.

The new system allows clinicians to distinguish between different Plasmodium species and detect them at low levels. Impressively, the Specific High-sensitivity Enzymatic Reporter unLOCKing (SHERLOCK) assay can detect as little as two parasites in each microliter of blood.

The assay does not require the nucleic acid extraction step needed with other nucleic acid detection methods, simplifying sample preparation. The enzyme CRISPR-Cas12a is used to chew up specific single-stranded DNA reporter sequences, and the resulting cleavage products indicate whether a nucleic acid sequence from a Plasmodium species is present in the sample.

Sample preparation takes only ten minutes, and the Plasmodium can be detected within another 60 minutes using a hand-held fluorescence analysis device or a lateral flow strip, which displays a band in the presence of the assay end-product.

“Importantly, the assay is compatible with different sample types, such as whole blood, plasma, serum, and dried blood, and all components required for amplification, Cas12a activation and signal generation can be lyophilized in a single test tube to work together in a one-pot-reaction after they are reconstituted and mixed with patient sample,” said Rose Lee, another researcher involved in the project. “This avoids having to depend on a functional cold-chain and allows testing to be performed in low-resource settings with minimal expertise.”

Study in Proceedings of the National Academy of Sciences: Ultrasensitive CRISPR-based diagnostic for field-applicable detection of Plasmodium species in symptomatic and asymptomatic malaria

Via: Harvard Wyss Institute





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