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United States Department of Agriculture

Agricultural Research Service

Research Project: IMMUNOLOGIC AND PHARMACOLOGICAL INTERVENTIONS OF VECTOR-BORNE BABESIOSIS

Location: Animal Diseases Research

2008 Annual Report


1a.Objectives (from AD-416)
The objectives of this research are: (1) the testing of the hypothesis that tick antigens capable of inducing anti-tick immunity can be delivered through a transfected parasite; (2) discovery of new tick antigen vaccine targets; (3) an understanding of the vector competence of certain U. S. ticks for transmission of equine babesiosis, and (4) the determination if certain anti-babesial drugs are capable of clearing horses of persistent babesial infection.


1b.Approach (from AD-416)
The above objectives will be approached through the use of transfection to create replication competent parasites containing tick antigens known to induce anti-tick immunity. In parallel with this approach new tick antigens will be discovered and characterized through genetic approaches including suppressive subtractive hybridization and Serial Analysis of Gene Expression. New antigens will first be tested by subunit immunization for their ability to induce anti-tick immunity. Tick vectors present in the U. S. and known to feed on horses will be tested for their ability to transmit B. equi and/or B. caballi and finally selected chemotherapeutics will be tested for their ability to clear persistent B. equi and/or B. caballi infections. Clearance will be defined as the lack of detectable anti-parasite antibody, the lack of PCR detectable genetic elements of B. equi and/or B. caballi and finally by testing for the ability of know competent ticks to acquire infection from treated horses. Formerly 5348-32000-020-00D (12/06).


3.Progress Report
This project is part of NP 103 Animal Health, component 7, Prevention and Control of Parasite Diseases, under 7B, Countermeasures to control and prevent parasitic diseases: Hemoparasitic Diseases. The causative protozoan parasites which cause bovine and equine babesiosis lead to disparate control issues for our primary partner (Animal Plant Health Inspection Service, APHIS) in controlling infection and disease. Research efforts are directed at providing risk analysis in terms of vector borne transmission and future solutions to control vector borne transmission of these parasites. Specifically, the following summarizes research progress directed at collaborating with APHIS in control of bovine and equine babesiosis. (1) completed development and technical transfer of accurate and validated serologic assays for Babesia bigemina (this is in addition to the assay for B. bovis, developed in the previous review cycle) ; (2) developed stable transfection of bovine babesial parasites and applied for a patent to cover this technology; (3) discovered a new distinct bovine gammadelta T-lymphocyte sub-population capable of participating in type 1 immune responses; (4) demonstrated relative vector-borne transmission efficiency of cattle and horses persistently infected with babesial parasites, and (5) completed and published the genomic sequence of B. bovis.


4.Accomplishments
1. Identify genes in Rhipicephalus (Boophilus) microplus that are expressed in the guts and ovaries in response to infection with Babesia bovis (cause of cattle fever).

Previously controlled, exotic disease transmitting ticks are developing resistance to chemical control. In order to develop anti-tick or transmission blocking vaccines it is necessary to identify tick genes that are expressed in response to infection. Working with Boophilus microplus scientists with the Animal Disease Research Unit and the Knipling-Bushland U. S. Livestock Insects Research Laboratory of ARS identified several genes that are expressed in the gut and ovary tissues in response to infection with Babesia bovis. These genes are candidates for anti-tick or transmission blocking vaccines. This accomplishment addresses NP #103, Animal Health, and Problem Statement #7B: Hemoparasitic Diseases.

2. Serologic diagnostic test for cattle persistently infected with Babesia bigemina.

Emergence of chemical resistant ticks has led to the need to detect cattle infected by babesial parasites transmitted by these ticks. Following through on our obligation to the Animal Plant Health Inspection Service (APHIS) scientists with the Animal Disease Research Unit of ARS and collaborators from Washington State University developed and transferred to the biomedical industry a cELISA for B. bigemina. This test and the previously developed test for B. bovis have been used in collaboration with APHIS to define the infectious status of certain cattle. The availability of these tests allows APHIS and diagnostic laboratories to accurately identified persistently infected populations of cattle and contributes to the ability to control transmission and economic losses due to disease outbreaks. This accomplishment addresses NP #103, Animal Health, and Problem Statement #7B: Hemoparasitic Diseases.

3. Stable transfection of Babesia bovis

Emergence of chemical resistant ticks has led to the need to develop novel vaccines intended to prevent transmission and/or control ticks through modulation of their ability to feed. Scientists with the Animal Disease Research Unit of ARS and collaborators from Washington State University successfully stably transfected (permanently) a foreign gene (green fluorescent protein) into the red blood cell stage of B. bovis. This now allows B. bovis to be stably transfected with other foreign genes such as those encoding for tick antigens which induce bovine anti-tick immunity. The impact of this work is novel vaccine design and delivery systems for control of vector-borne diseases. This accomplishment addresses NP #103, Animal Health, and Problem Statement #7B: Hemoparasitic Diseases.

4. Determination of relative transmission efficiency of cattle and horses persistently infected with babesial parasites which cause cattle fever and horse piroplasmosis

The emergence of chemically resistant ticks capable of transmitting the parasites of bovine and equine babesiosis has led to the need to determine the risk associated with persistently infected cattle and horses. Using a Rhipicephalus (Boophilus) microplus tick which is a natural transmission vector of babesial parasites, scientists from the Animal Disease Research Unit of ARS in collaboration with Washington State University showed that chronically infected cattle and horses are efficient reservoirs for parasite (disease) transmission. These data support the use of serologic diagnostic tests in the identification and movement restriction of infected animals. This work directly supports the infectious disease control mission of the Animal Plant Health Inspection Service. This accomplishment addresses NP #103, Animal Health, and Problem Statement #7B: Hemoparasitic Diseases.

5. Discovery of a novel T-lymphocyte involved in type 1 immune responses

A major impediment to successful vaccines against a number of important pathogens is the lack of knowledge concerning immune responses which contribute to protective immune responses. Gamma delta T lymphocytes are an important component of bovine innate immunity and have been shown to have regulatory, inflammatory and cytotoxic functions. Scientists with the Animal Disease Research Unit of ARS in collaboration with Washington State University discovered a new subset of gamma delta T lymphocytes which are capable of producing a robust IFN-gamma response, which is important in the immune control of bovine babesiosis (cattle fever). The discovery of this new T-lymphocytes subset provides a marker measurement for the induction of protective immune responses. This accomplishment addresses NP #103, Animal Health, and Problem Statement #7B: Hemoparasitic Diseases.

6. Completion of sequencing and annotation of the Babesia bovis genome

An important need in developing transfection based vaccines for parasitic diseases is a detailed understanding of their genomes including potential genomic sites for foreign gene insertion. Scientists with the Animal Disease Research Unit of ARS in collaboration with Washington State University completed sequencing and annotation of the B. bovis (cause of cattle fever) genome. This allows for the identification of novel parasite gene products for testing as vaccine targets. The publication of the complete genome allows the global scientific community to participate in discovery novel ways to develop vaccines which prevent or minimize economic losses from this important parasite of cattle. This accomplishment addresses NP #103, Animal Health, and Problem Statement #7B: Hemoparasitic Diseases.


5.Significant Activities that Support Special Target Populations
None


6.Technology Transfer

Number of New Patent Applications Filed1

Review Publications
Sunter, J.D., Patel, S.P., Skilton, R.A., Githaka, N., Knowles Jr, D.P., Scoles, G.A., Nene, V., De Villiers, E., Bishop, R. 2008. A novel SINE family occurs frequently in both genomic DNA and transcribed sequences in ixodid ticks of the arthropod sub-phylum Chelicerata. Gene. 415(1-2):13-22.

Suarez, C.E., Mcelwain, T.F. 2008. Transient transfection of purified Babesia bovis merozoites. Experimental Parasitology. 118(4):498-504.

Johnson, W.C., Bastos, R.G., Davis, W.C., Goff, W.L. 2008. Bovine WC1- gamma delta T-cells incubated with IL-15 express the natural cytotoxicity receptor CD335 (NKp46) and produce IFN-gamma in response to exogenous IL-12 and IL-18. Developmental and Comparative Immunology. 32:1002-1010.

Ueti, M.W., Palmer, G.H., Scoles, G.A., Kappmeyer, L.S., Knowles Jr, D.P. 2008. Persistently Infected Horses Are Reservoirs for Intrastadial Tick-Borne Transmission of the Apicomplexan Parasite Babesia equi. Infection and Immunity. 76(8):3525-3529.

Howell, J.M., Ueti, M.W., Palmer, G.H., Scoles, G.A., Knowles Jr, D.P. 2007. Persistently Infected Calves as Reservoirs for Acquisition and Transovarial Transmission of Babesia bovis by Rhipicephalus (Boophilus) microplus. Journal of Clinical Microbiology. 45(10):3155-3159

Brayton, K., Lau, A.O., Herndon, D.R., Hannick, L., Kappmeyer, L.S., Berens, S.J., Bidwell, S.L., Brown, W.C., Crabtree, J., Fadrosh, D., Feldblum, T., Forberger, H.A., Haas, B.J., Howell, J.M., Khouri, H., Koo, H., Mann, D.J., Norimine, J., Paulsen, I.T., Radune, D., Ren, Q., Smith, R.K., Suarez, C.E., White, O., Wortman, J.R., Knowles Jr, D.P., Mcelwain, T.F., Nene, V.M. 2007. Genome Sequence of Babesia bovis and Camparative Analysis of Apicomplexan Hemoprotozoa. PLoS Pathogens. 3(10):1401-1413.

Rachinsky, A., Guerrero, F.D., Scoles, G.A. 2007. Differential protein expression in ovaries of uninfected and Babesia-infected southern cattle ticks, Rhipicephalus (Boophilus) microplus. Insect Biochemistry and Molecular Biology. 37:1291-1308.

Rachinsky, A.S., Guerrero, F., Scoles, G.A. 2008. Proteomic profiling of Rhipicephalus (Boophilus) microplus midgut responses to infection with Babesia bovis. Veterinary Parasitology. 152(3-4):294-313.

Last Modified: 9/29/2014
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