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Viability of Onchocerca volvulus in vitro
G Strote1, A Bokhof, J C Comley
1Bernhard Nocht Institute for Tropical Medicine, Department of Helminthology and Entomology, Hamburg, Germany.
Abstract:
The use of a selective schedule of tests to identify a viable population of isolated adult Onchocerca volvulus (Nematoda: Filarioidea) has been investigated in a large worm population. The study was initiated to develop methodology appropriate to test new candidate macrofilaricides for their in vitro activity against O. volvulus. After removal from the host the viability of isolated intact parasites was estimated by assessing the motility indices of male worms, and the colorimetric quantification of the reduction of the bioreducible tetrazolium reagent XTT and lactate output by female worms. Additionally the motility of whole females and the movement of inner organs of female worms were scored quantitatively. These response parameters were used to sort the adult worms into viability groups at the start of the in vitro culture. The adult worms were then observed for 6 days and viability was assessed regularly during the culture period. At the end of the culture period, the reduction of the water-insoluble tetrazolium reagent MTT was used to determine the formazan formed by the entire male and female worms. The response parameters used at the start of the culture proved to be highly predictive for detecting viable and non-viable adult worms. In the group of worms selected as 'viable' around 70% kept their motility and metabolic activity at a high level until the end of the culture compared to the initial level. In contrast, none of the female worms and only 13% of the male worms categorized as 'poorly viable' demonstrated a motility index or metabolic level at the end of the culture period that was comparable to that of the worms in the 'viable' groups. For female worms the lactate output correlated significantly with weight whereas no correlation was seen between MTT reduction and weight.
Insights
A selective testing schedule effectively identifies viable adult Onchocerca volvulus (O. volvulus) for drug screening. This method accurately predicts parasite survival and metabolic activity during in vitro culture, aiding macrofilaricide development.
Area of Science:
- Parasitology
- Drug Discovery
- Nematology
Background:
- Developing effective macrofilaricides against Onchocerca volvulus requires reliable methods to assess parasite viability.
- In vitro testing is crucial for screening new drug candidates against O. volvulus.
Purpose of the Study:
- To establish a selective testing schedule for identifying viable adult Onchocerca volvulus.
- To develop a methodology for assessing in vitro activity of candidate macrofilaricides against O. volvulus.
Main Methods:
- Assessing motility indices, XTT reduction, and lactate output for initial viability sorting.
- Quantitatively scoring female worm motility and internal organ movement.
- Monitoring worm viability over a 6-day in vitro culture period.
- Using MTT reduction to determine formazan formation at the culture's end.
Main Results:
- Initial viability parameters accurately predicted adult worm survival and metabolic activity.
- Approximately 70% of worms selected as 'viable' maintained high motility and metabolic activity.
- Poorly viable worms showed minimal motility and metabolic activity by the culture's end.
- Female worm lactate output correlated with weight, but MTT reduction did not.
Conclusions:
- The developed selective testing schedule is highly predictive of Onchocerca volvulus viability.
- This methodology supports the in vitro screening of macrofilaricides against O. volvulus.
- Accurate viability assessment is key for effective drug development against parasitic nematodes.