A dirofilariasis mouse model for heartworm preclinical research

Abstract

ABSTRACTUse of experimental cats and dogs in veterinary heartworm preclinical drug research is increasing. As a potential alternative primaryin vivoheartworm preventative drug screen, we assessed lymphopenic mice with ablation of the interleukin-2/7 common gamma chain (γc) as susceptible to the larval development phase ofD. immitis. Non-obese diabetic (NOD) Severe Combined ImmunoDeficient (SCID)γc-/-(NSG / NXG) mice consistently yielded viableD. immitislarvae at 2-4 weeks post-infection across multiple experiments, different batches of infectious larvae inoculates, different isolates ofD. immitisand at independent laboratories. Mice did not display any overt clinical signs associated with infection up to 4 weeks. Developing larvae were found in subcutaneous and muscle fascia tissues, the natural site of this stage of heartworm in dogs. Larvae retrieved from NSG / NXG mice were mid-L4 stage of development. Compared with 14-dayin vitropropagated larvae,in vivoderived L4 were significantly larger and contained expandedWolbachiaendobacteria titres, determined by QPCR and Fluorescentin situHybridisation (FISH). We established anex vivo6-day L4 paralytic screening system against nematodicidal agents (moxidectin, levamisole) which highlighted discrepancies in relative drug sensitivities in comparison within vitroreared L4D. immitis.We demonstrated effective depletion ofWolbachiaby 70-90% inD. immitisL4 following 2-7 day oralin vivoexposures of NSG / NXG infected mice with doxycycline or the rapid-acting investigational anti-Wolbachiadrug, AWZ1066S. We validated the NSG / NXG mouse model as a filaricide drug screen byin vivotreatments with single injections of moxidectin, which mediated 60-88% reduction in L4 larvae at 14-28 days. Future adoption of the mouse model as a first-line efficacy screen will benefit end-user laboratories conducting research and development of novel heartworm preventatives via increased access, rapid turnaround and reduced costs whilst simultaneously decreasing need for experimental cat or dog use.