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Tag: Onchocerciasis

Optimized strategy for real-time qPCR detection of Onchocerca volvulus DNA in pooled Simulium sp. blackfly vectors

Donna Huber on December 14, 2023

Fig 1. Workflow for the selection of the optimal qPCR assay.
Fig 1. Workflow for the selection of the optimal qPCR assay.

 

Background: Onchocerca volvulus is a filarial parasite that is a major cause of dermatitis and blindness in endemic regions primarily in sub-Saharan Africa. Widespread efforts to control the disease caused by O. volvulus infection (onchocerciasis) began in 1974 and in recent years, following successful elimination of transmission in much of the Americas, the focus of efforts in Africa has moved from control to the more challenging goal of elimination of transmission in all endemic countries. Mass drug administration (MDA) with ivermectin has reached more than 150 million people and elimination of transmission has been confirmed in four South American countries, with at least two African countries having now stopped MDA as they approach verification of elimination. It is essential that accurate data for active transmission are used to assist in making the critical decision to stop MDA, since missing low levels of transmission and infection can lead to continued spread or recrudescence of the disease.

Methodology/principal findings: Current World Health Organization guidelines for MDA stopping decisions and post-treatment surveillance include screening pools of the Simulium blackfly vector for the presence of O. volvulus larvae using a PCR-ELISA-based molecular technique. In this study, we address the potential of an updated, practical, standardized molecular diagnostic tool with increased sensitivity and species-specificity by comparing several candidate qPCR assays. When paired with heat-stable reagents, a qPCR assay with a mitochondrial DNA target (OvND5) was found to be more sensitive and species-specific than an O150 qPCR, which targets a non-protein coding repetitive DNA sequence. The OvND5 assay detected 19/20 pools of 100 blackfly heads spiked with a single L3, compared to 16/20 for the O150 qPCR assay.

Conclusions/significance: Given the improved sensitivity, species-specificity and resistance to PCR inhibitors, we identified OvND5 as the optimal target for field sample detection. All reagents for this assay can be shipped at room temperature with no loss of activity. The qPCR protocol we propose is also simpler, faster, and more cost-effective than the current end-point molecular assays.

 

Mary Doherty, Jessica R Grant, Nils Pilotte, Sasisekhar Bennuru, Kerstin Fischer, Peter U Fischer, Sara Lustigman, Thomas B Nutman, Kenneth Pfarr, Achim Hoerauf, Thomas R Unnasch, Hassan K Hassan, Samuel Wanji, Patrick J Lammie, Eric Ottesen, Charles Mackenzie, Steven A Williams. PLoS Negl Trop Dis. 2023 Dec 14;17(12):e0011815. doi: 10.1371/journal.pntd.0011815.

Onchocerciasis: Target product profiles of in vitro diagnostics to support onchocerciasis elimination mapping and mass drug administration stopping decisions

Donna Huber on August 3, 2022

In June 2021, the World Health Organization (WHO), recognizing the need for new diagnostics to support the control and elimination of onchocerciasis, published the target product profiles (TPPs) of new tests that would support the two most immediate needs: (a) mapping onchocerciasis in areas of low prevalence and (b) deciding when to stop mass drug administration programs. In both instances, the test should ideally detect an antigen specific for live, adult O. volvulus female worms. The preferred format is a field-deployable rapid test. For mapping, the test needs to be ≥ 60% sensitive and ≥ 99.8% specific, while to support stopping decisions, the test must be ≥ 89% sensitive and ≥ 99.8% specific. The requirement for extremely high specificity is dictated by the need to detect with sufficient statistical confidence the low seroprevalence threshold set by WHO. Surveys designed to detect a 1-2% prevalence of a given biomarker, as is the case here, cannot tolerate more than 0.2% of false-positives. Otherwise, the background noise would drown out the signal. It is recognized that reaching and demonstrating such a stringent specificity criterion will be challenging, but test developers can expect to be assisted by national governments and implementing partners for adequately powered field validation.

Marco A. Biamonte ,Paul T. Cantey,Yaya I. Coulibaly,Katherine M. Gass,Louise C. Hamill,Christopher Hanna,Patrick J. Lammie,Joseph Kamgno,Thomas B. Nutman,David W. Oguttu,Dieudonné P. Sankara,Wilma A. Stolk,Thomas R. Unnasch. PLoS Negl Trop Dis. 2022 Aug 3;16(8):e0010682. doi: 10.1371/journal.pntd.0010682.