Development and validation of a quantitative PCR for the detection of Guinea worm (Dracunculus medinensis)

Abstract

Dracunculus medinensis (Guinea worm) is a parasitic nematode that can cause the debilitating disease dracunculiasis (Guinea worm disease) in humans. The global Guinea Worm Eradication Program has led intervention and eradication efforts since the 1980s, and Guinea worm infections in people have decreased >99.99%. With the final goal of eradication drawing nearer, reports of animal infections from some remaining endemic countries pose unique challenges. Currently, confirmation of suspected Guinea worm infection relies on conventional molecular techniques such as polymerase chain reaction (PCR), which is not specific to Guinea worm and, therefore, requires sequencing of the PCR products to confirm the identity of suspect samples, a process that often takes a few weeks. To decrease the time required for species confirmation, we developed a quantitative PCR assay targeting the mitochondrial cytochrome b (cytb) gene of Guinea worm. Our assay has a limit of detection of 10 copies per reaction. The mean analytical parameters (± SE) were as follows: efficiency = 93.4 ± 7.7%, y-intercept = 40.93 ± 1.11, slope = -3.4896 ± 0.12, and the R2 = 0.999 ± 0.004. The assay did not amplify other nematodes found in Guinea worm-endemic regions and demonstrated 100% diagnostic sensitivity and specificity. Implementation of this quantitative PCR assay for Guinea worm identification could eliminate the need for DNA sequencing to confirm species. Thus, this approach can be implemented to provide more rapid confirmation of Guinea worm infections, leading to faster execution of Guinea worm interventions while increasing our understanding of infection patterns.

The current state of knowledge on dracunculiasis: a narrative review of a rare neglected disease

“…Conventional PCR and nematode DNA amplification have been used to some extent, but the turnaround time from the sample reception to species confirmation can be up to two weeks. Recently, a fast quantitative polymerase chain reaction (GW qPCR) test has shown high sensitivity and specificity in diagnosing suspected adult female Guinea worm infections in humans and in domestic or wild animals [29]. This approach could allow faster confirmation of Guinea worm infections, leading to more rapid interventions while improving our understanding of infection patterns.…”

Nanobody Technology and New Molecular Biology Methods to Advance Rapid Diagnostic Test for Neglected Tropical Diseases

“…At present, PCR technology is widely applied in diagnosis and other fields of biosciences. The technology is used for diagnosis of NTDs including Buruli ulcers [16], Dengue [17], Chikungunya [18], HAT [19], leishmaniasis [20], Dracunculiasis [21], deep mycoses [22], onchocerciasis [23], rabies [24], STH [25], and yaws [26] among others. PCR lends itself as a sensitive pathogen detection tool owing to its ability to amplify a single copy of genome to billions of copies.…”