Genetically modified babesia parasites expressing protective tick antigens and uses thereof
Inventors
Suarez, Carlos E. • Knowles, Jr., Donald P. • McElwain, Terry
Assignees
US Department of Agriculture USDA • Washington State University WSU
Publication Number
US-9707283-B2
Publication Date
2017-07-18
Expiration Date
2028-05-05
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Abstract
The present invention relates to methods for stable transfection of Babesia parasites, and for vaccines conferring immunity against parasitic arthropods.
Core Innovation
The invention provides methods for stable transfection of Babesia parasites to produce genetically modified strains expressing heterologous proteins, particularly protective tick antigens. This approach addresses the problem of parasitic arthropods, especially ticks, which are significant vectors of diseases affecting humans and livestock globally. Ticks transmit various pathogens causing diseases such as relapsing fever, Lyme disease, babesiosis, and others, leading to substantial medical and economic impacts, especially in the cattle industry.
Traditional methods to control ticks, including chemical acaricides and selective breeding of tick-resistant livestock, have limitations such as resistance development, environmental harm, and unpredictability. Existing recombinant vaccines require multiple inoculations and do not achieve full protection. The invention addresses the need for an effective vaccine that provides sustained immunity with a single inoculation, avoiding drawbacks associated with acaricides and breeding.
The invention uses stable transfection of Babesia parasites with expression vectors that include promoters operably linked to heterologous nucleic acids encoding protective tick antigens such as Bm86 or Bm95. These genetically modified Babesia parasites can be used as live attenuated vaccines that confer immunological resistance to ticks by expressing the protective antigens. The transfection methods include nucleofection and electroporation of infected erythrocytes or purified merozoites, allowing integration of foreign genes into the Babesia genome.
Claims Coverage
The patent claims include independent claims directed to genetically modified Babesia spp. producing heterologous proteins, methods for generating such stably transfected parasites, and compositions comprising the modified organisms. There are three independent claims covering the genetically modified organisms, the methods of preparation, and compositions containing them.
Genetically modified Babesia spp. producing heterologous proteins
A Babesia parasite that is stably transfected with a polynucleotide comprising a promoter operably linked to a heterologous nucleic acid encoding a heterologous protein. This stably transfected polynucleotide integrates into the Babesia genome, and the Babesia produces the heterologous protein.
Method for generating stably transfected Babesia spp. producing heterologous proteins
A method comprising: (i) preparing an expression vector with a promoter active in Babesia operably linked to heterologous DNA encoding a heterologous protein; (ii) transfecting this vector into Babesia merozoites or infected red blood cells, resulting in stable genome integration; and (iii) selecting for genetically modified Babesia that produce the heterologous protein.
Stably transfected Babesia spp. made by the described method
A Babesia parasite produced by the method of preparing an expression vector, transfecting Babesia merozoites or infected red blood cells, and selecting for stably transfected Babesia expressing the heterologous protein.
The claims cover stable genetic modification of Babesia parasites to express heterologous proteins, methods using expression vectors and transfection techniques such as nucleofection or electroporation, and compositions including these genetically modified parasites. The inventive features focus on stable genomic integration of heterologous expression cassettes encoding proteins such as tick antigens and their use in genetically modified live Babesia strains.
Stated Advantages
Provides a vaccine that delivers sustained immunity against ticks and Babesia with potentially a single inoculation, avoiding repeated vaccinations.
Circumvents problems associated with chemical acaricides such as resistance development, chemical residues, and environmental harm.
Allows development of live attenuated Babesia vaccines expressing protective tick antigens, combining protection against Babesia infection and tick infestation.
Enables stable transfection of Babesia for functional genetic studies and development of vaccine strains expressing heterologous antigens.
Documented Applications
Use of genetically modified Babesia parasites as live attenuated vaccines to confer immunological resistance against ticks and Babesia parasites in animals.
Functional characterization of Babesia genes by creating knock-out strains using stable transfection.
Identification and development of new subunit vaccine candidates for Babesia and related tick-borne diseases through transgenic Babesia expressing protective antigens.
Tracking and differentiating vaccinated animals from naturally infected animals via introduced selectable markers or antigenic tags expressed by the modified Babesia.
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