Investigating genetic composition of Onchocerca worms pre‐ and post‐drug treatment: Improving the ability to test macrofilaricidal drug efficacy

Drugs for Neglected Diseases Initiative
PI: Poppy Lamberton

The World Health Organization (WHO) aims for elimination of onchocerciasis in Latin America, in selected African countries and in Yemen by 2020. Community Directed Treatment with Ivermectin (CDTI) is the current WHO recommended method of controlling onchocerciasis, yet this only kills the microfilariae and not the adult worms. Evidence for ivermectin resistance, with shorter embyrostasis duration in worms exposed to multiple treatments, has been reported. New drugs are being developed which need to be tested in endemic areas. However, none of the current diagnostic methods can distinguish between microfilariae derived from worms which have survived treatment, or those from reinfections.

We will optimise genetic fingerprinting methods to ascertain if microfilariae in patients post-treatment are due to reinfections or from adult worms which have survived drug treatment. We can successfully extract DNA from, and genotype, individual microfilariae, and obtain single nucleotide polymorphisms (SNPs), which enable inference of the parent adult worm genotypes. We will use double digest rad sequencing protocols to obtain SNPs. Using these methods Tristan Dennis infer if microfilariae post-treatment are siblings or half siblings of those collected pre-treatment (and therefore that the treatment was unsuccessful in removing a given proportion of the adult parent worms). These data will inform on drug efficacy by measuring the relative proportion of microfilariae that come from non-cleared parent infections.

In work package (WP) 1, Tristan Dennis will determine the minimal number of microfilariae required to accurately infer the adult breeding population at a range of baseline infection intensities. This will initially be performed on previously stored O. volvulus and O. ochengi samples to enable a non-invasive rapid start. The O. ochengi samples will further enable a controlled experiment to ascertain if the location and timing of skin snips affect our ability to infer the total adult worm population and how we can control for this. At the start of WP 2 the protocols optimised in WP1 will be trialled on microfilariae obtained from skin snips from patients in endemic areas harbouring different infection intensities. Bayesian algorithms will provide error measures and predictions on the proportions of treatment failures, when full pre-treatment genetic diversity is not fully quantified due to a limited number of skin snips. This will provide a proof of principle of methods for accurately determining O. volvulus siblings and half siblings in humans. These methods will then be used throughout WP2 to accurately quantify the proportion of adult worms surviving treatment in new macrofilaricide Phase III trials. These methods will also enable quantification of ivermectin resistance where necessary, by characterising the proportion of microfilariae repopulating the skin which are derived from treated worms versus those from ongoing transmission.