Egg hatching protocol and an in vitro scoring system in Parascaris univalens larvae after exposure to anthelmintic drugs

A scaris is a genus of parasitic worms (helminths) found in the small intestine of various mammalian hosts, including Ascaris lumbricoides in humans, Parascaris equorum and P univalens in horses, Ascaris suum in pigs, Toxocara cati in cats and Toxocara canis in dogs. To date, Parascaris spp. are the only Ascaris worms that have developed resistance to anthelmintic drugs. The mechanisms of resistance in Parascaris spp are incompletely understood, partly due to the absence of robust in-vitro models. Further complicating in-vitro studies, Parascaris spp lack a free-living larval stage as their larva only hatch within the host. The aim of this study was to develop in-vitro methods for hatching, scoring the viability of Parascaris L3 larvae and exposing them to the anthelmintic drugs ivermectin, pyrantel, thiabendazole, and the herbal extract carvacrol. This study shows that mechanical Ascaris egg breaking using a homogenizer resulted in a hatching rate of 98%. Our viability scoring system could distinguish an ivermectin resistant larvae from an ivermectin susceptible larvae derived from different farms. This indicates that this method may have utility for the screening of larvae ivermectin resistance on the level of farm populations. Interestingly, a highly paralytic effect observed after carvacrol exposure. Carvacrol shows direct paralytic effects on Parascaris larvae in a dose-dependent manner, as higher concentrations were lethal to all exposed larvae. This result presents a potential future opportunity for carvacrol used in the treatment of Ascaris infections. To conclude our results, we have successfully developed an in-vitro model as well as a scoring system for the viability of Parascaris L3 stage larvae, which can be used for assaying the effect on larvae after drug exposure