Location: Animal Disease Research
Project Number: 2090-32000-039-023-T
Project Type: Trust Fund Cooperative Agreement
Start Date: Apr 1, 2018
End Date: Jul 1, 2021
1) Develop a continuous flow in vitro system for feeding of competent tick vectors of T. parva for isolation and quantification of pure sporozoites. 2) To abrogate the requirement of oxytetracycline co-treatment, determine if irradiation-attenuated, free T. parva sporozoites, or low doses of non-attenuated free sporozoites provide immune protection against tick challenge without causing significant morbidity. 3) Titrate a standardized dose for the improved, free-sporozoite vaccine and validate it in field immunization and challenge studies at livestock facilities in Malawi and Morocco.
The current infection and treatment method (ITM) vaccine for controlling T. parva uses live organisms within infected, macerated Rhipicephalus appendiculatus ticks. There are three major drawbacks to this approach. The first drawback is the inability to standardize sporozoite dose within vaccine stabilates. This occurs because there is no way to quantify sporozoites within tick salivary glands prior to maceration of ticks to produce the stabilate. Thus, each batch must be carefully titrated in large numbers of cattle to determine the dose of stabilate that, when given with oxytetracycline, will elicit an immune response without causing significant morbidity. The second major drawback is the inclusion of tick proteins in vaccine stabilates. Tick proteins are pharmacologically bioactive, have been shown to modify immune response development, and can enhance pathogen replication. The third major drawback is the necessary inclusion of oxytetracycline to prevent severe disease after inoculation with the current sporozoite vaccine. Oxytetracycline significantly increases the price per dose of ITM, and limits adoption of the vaccine by smallholder farmers. We propose to improve the well-established ITM vaccine by employing our recently developed continuous flow in vitro tick feeding system, which will greatly decrease the requirement of rabbits and cattle for ITM production, and enable isolation of pure T. parva sporozoites. The advantages of this new approach are: 1) Allows isolation of mature, infective sporozoites; 2) Allows accurate quantification of sporozoites for inter-batch standardization of ITM stabilates; 3) Eliminates the presence of immune-modulating tick protein contaminants in the stabilate; 4) Due to the availability of known quantities of free sporozoites, enables testing of methods for parasite attenuation (particularly irradiation) and dose manipulation to eliminate the need for antibiotic co-treatment in ITM. We hypothesize that inoculation with a known quantity of irradiation-attenuated, pure T. parva sporozoites, or a very low dose of non-attenuated, pure T. parva sporozoites, will establish sub-clinical infection and provide protection against T. parva-tick challenge without the need for concurrent oxytetracycline treatment. We will test this hypothesis in three specific aims: 1) Develop a continuous flow in vitro system for feeding of competent tick vectors of T. parva for isolation and quantification of pure sporozoites; 2) Determine if irradiation-attenuated, free T. parva sporozoites, or low doses of non-attenuated free sporozoites, provide immune protection against tick challenge without causing significant morbidity; 3) Titrate a standardized dose for the improved, free-sporozoite vaccine and validate it in field immunization and challenge studies at livestock facilities in Malawi and Morocco. Our goal is to decrease ITM per-dose costs by minimizing the use of animals for vaccine production, standardizing the infectious sporozoite dose to produce a more efficient and consistent vaccine stabilate, and eliminating the requirement for oxytetracycline.