Development of a Caenorhabditis elegans model for the assessment of toxicity and its application in testing novel anthelmintics.

The nematode Caenorhabditis elegans is an alternative model used in biomedical research for the investigation of descriptive and mechanistic toxicity assessment of chemicals. There are considerable differences in published data, especially in terms of reproducibility and validation of toxicity endpoints, and the techniques used in the investigation of these endpoints. This thesis describes the evaluation of toxicological endpoints following the exposure of C. elegans to chemicals which include; zinc oxide nanoparticles (ZnONP), Diethylstilbestrol (DES) and derivatized target-specific anthelmintics. The results suggest that ZnONP prepared in anionic and cationic dispersants (AZNP and CZNP respectively) were the most toxic against the nematode resulting in the ‘bag of worms’ (BOW) phenotype which can be exploited as a marker for reproductive toxicity. Also, worms treated with ZnONP prepared in 0.1% FBS (FZNP), molecular grade water (WZNP) or E. coli OP50 supernatant (SZNP) presented three-fold embryo elongation showing fully differentiated tissues encapsulated within the eggshell and still within the hermaphrodite gravid adult. The phenotype has been named accelerated embryonic development (AED) and could be used as a developmental toxicity endpoint. The results suggest that the AED endpoint is the most sensitive while lethality endpoint appears to be the least sensitive despite its extensive use in the literature. Also, microRNA microarray expression appears to be the most sensitive molecular endpoint while behavioural endpoints such as speed should be interpreted with caution, especially when performed manually. Importantly, good C. elegans culture practice (GCeCP) is required for reproducible chemical toxicity assessment and different endpoints may be required for different types of toxicity assessment. Additionally, the thesis describes a second but related study which explores a potential for enhanced anthelmintic targeting. Novel fluorophore-based asparagine-containing oligopeptide substrate probes were used to target the helminth protease, legumain. These probes were selectively cleaved by legumain in C. elegans, Haemonchus contortus and Teladorsagia circumcincta. The protease-specific probes could potentially be exploited to achieve protease-mediated prodrug activation and drug delivery.