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United States Department of Agriculture

Agricultural Research Service

Research Project: DETERMINANTS OF ANAPLASMA MARGINALE TRANSMISSION AT THE VECTOR/PATHOGEN INTERFACE
2011 Annual Report


1a.Objectives (from AD-416)
Our objective in this project is to investigate the factors influencing transmission of Anaplasma marginale by Dermacentor andersoni. We hypothesize that there are interactions between the vector and the pathogen that are determinants of transmission. Our first objective is to set up a field study to examine the relationship between tick vector competence and transmissibility of Anaplasma marginale strains at field sites selected for differences in vector abundance and pathogen strain composition. We will collect ticks annually and determine the susceptibility phenotype of the population at each site by determining the proportion of ticks that are susceptible to midgut infection with A. marginale; midgut susceptibility is a surrogate marker for vector competence. Using a longitudinal survey of a cohort of cattle at each site we will test the hypothesis that some strains of A. marginale are more highly transmissible than others. The first objective will also provide ticks and A. marginale isolates for study in the subsequent objectives. Our next 2 objectives target vector competence of the tick population. First we will attempt to establish if vector competence is a stable genetic characteristic of the tick populations at our field sites by testing the hypotheses that:.
1)the proportion of ticks that are susceptible to midgut infection with A. marginale within each population (i.e. the population susceptibility phenotype) is stable characteristic of the population from one year to the next, and.
2)that there is limited gene flow between populations of D. andersoni. Secondly, we will determine if tick innate immune responses regulate vector competence by testing the hypotheses that.
1)there are differences between tick populations in sequence or expression of tick defensins, and.
2)that these differences correlate with phenotypes which are associated with vector competence for A. marginale. Our final two objectives target A. marginale strain transmissibility. First, we will identify common genetic markers of highly transmissible A. marginale strains collected in our field study, and test the hypothesis that these strains share genetic determinants that are associated with, and are predictive of, more efficient transmission by ticks. We will then identify the outer membrane protein (OMP) structure of these highly transmissible A. marginale strains and test the hypotheses that.
1)highly transmissible A. marginale strains share conserved OMPs, and.
2)that immunization with conserved cross-linked OMPs will induce protection against challenge by heterologous A. marginale strains. By simultaneously approaching studies of the determinates of transmission of A. marginale from the prospective of tick vector competence and from the prospective of strain transmissibility we can begin to define the parameters that influence transmission, including parameters relating to the vector, the pathogen, and to their interaction.


1b.Approach (from AD-416)
Our objective in this project is to investigate the factors influencing transmission of Anaplasma marginale by Dermacentor andersoni. Our first approach is to set up a field study to examine the relationship between tick vector competence and transmissibility of Anaplasma marginale strains at field sites selected for differences in vector abundance and pathogen strain composition. We will collect ticks annually and determine the susceptibility phenotype of the population at each site by determining the proportion of ticks that are susceptible to midgut infection with A. marginale; midgut susceptibility is a surrogate marker for vector competence. Our next approach is to target vector competence of the tick population. We will establish if vector competence is a stable genetic characteristic of the tick populations at our field sites. And secondly, we will determine if tick innate immune responses regulate vector competence by testing if there are differences between tick populations in sequence or expression of tick defensins, and whether these differences correlate with phenotypes which are associated with vector competence for A. marginale. Our final approachs target A. marginale strain transmissibility. First, we will identify common genetic markers of highly transmissible A. marginale strains collected in our field study, and test the hypothesis that these strains share genetic determinants that are associated with, and are predictive of, more efficient transmission by ticks. We will then identify the outer membrane protein (OMP) structure of these highly transmissible A. marginale strains and test the hypotheses that.
1)highly transmissible A. marginale strains share conserved OMPs, and.
2)that immunization with conserved cross-linked OMPs will induce protection against challenge by heterologous A. marginale strains. Formerly 5348-32000-023-00D (12/06).


3.Progress Report
This was the final year of this project.

Anaplasma marginale, the causative agent of anaplasmosis, is a globally prevalent tick-borne pathogen of livestock. This bacterial pathogen causes a significant disease burden in cattle in the United States and is a barrier to trade. The tools currently available to control this disease are limited and rely on treatment of clinically affected animals and tick control. The overall goals of this project were to identify host and pathogen factors that lead to efficient tick transmission, and identify antigens that induce protective immunity with a goal of vaccine development.

This work resulted in 25 publications in peer-reviewed journals. The significant accomplishments from this work are:

1. Determinants of vector competence. In the U.S., the prevalence of A. marginale varies, despite the presence of a competent vector species. Understanding factors that dictate pathogen prevalence will allow for rational and targeted implementation of control strategies. Previously, Scoles et al. determined that populations of the ticks differ in their susceptibility to A. marginale. This work has been extended to demonstrate that this susceptibility is a stable genetic characteristic within a tick population. Additionally, tick population density is correlated with prevalence of A. marginale. Thus, both susceptibility of the tick vector to A. marginale as well as density of the tick population likely dictate prevalence of A. marginale in a particular region.

2. Pathogen determinants of efficient transmission. Understanding the mechanisms and molecules employed by the pathogen to successfully colonize the tick could lead to the development of control methods designed to block tick transmission. Toward this end, Ueti et al. determined that the tick midgut and salivary gland are two different barriers to tick transmission. The ability of the pathogen to replicate to high numbers in the salivary gland is a determinant of transmission efficiency of the pathogen. More specifically, sixteen outer membrane proteins that are specifically up-regulated in the tick as compared to the mammalian host have been identified, and thus are likely important during tick colonization and transmission.

3. Identification of antigens that induce protective immunity to A. marginale. A group of eleven surface-expressed A. marginale outer membrane proteins that induce protective immunity to challenge with A. marginale were identified. One of these vaccine candidates is Msp2, which is a highly antigenic outer membrane protein that allows for immune evasion and establishment of persistent infection in the mammalian host. Much work has been examining Msp2 as a vaccine candidate, however, Noh et al. determined that among immunized animals, the anti-Msp2 antibody response does not correlate with protective immunity. Thus, outer membrane proteins, other than Msp2, are deemed responsible for the effective vaccine-induced anti-A. marginale response. Identification of these proteins will guide future efforts in vaccine development.

New project 5348-32000-033-00D start 10-1-11.


4.Accomplishments
1. Development of tools required to understand the host-pathogen relationship. Understanding the mechanisms by which the pathogen, Anaplasma marginale, is able to colonize the tick may lead to novel methods to prevent tick transmission, and thus prevent disease. ARS researchers in concert with their collaborators at Washington State University determined that A. marginale carrying a green fluorescent protein (A. marginale-gfp) were able to complete an infection cycle, including establishment of persistent infection in the bovine host and colonization and subsequent transmission to a new host by the tick vector. These findings establish that A. marginale-gfp are a valid tool to study the interaction between the pathogen and host. Specifically, the ability to readily visualize the pathogen as it transits through the tick will provide a rapid means to measure the effects of potential transmission blocking agents.

2. Anaplasma marginale modulates the host immune system. The lack of a safe and effective vaccine for anaplasmosis, an economically significant disease caused by the tick-borne pathogen A. marginale is a hindrance to cattle production in many areas of the United States. ARS researchers in Pullman, WA, in concert with their collaborators at Washington State University determined that A. marginale-specific CD4+ T cells are deleted in the bovine during the transition from acute to persistent infection. The deletion of these immune cells, in part, explains the difficulty of induction of a protection immune response in the context of both infection and immunization. Formulation of a vaccine to stimulate A. marginale-specific CD4+T cells and overcome this pathogen-mediated immune modulation will likely greatly improve vaccine efficacy.


Review Publications
Han, S., Norimine, J., Brayton, K.A., Palmer, G.H., Scoles, G.A., Brown, W.C. 2011. Anaplasma marginale infection with persistent high-load bacteremia induces a dysfunctional memory CD4+ T lymphocyte response but sustained high IgG titers. Clinical and Vaccine Immunology. 17(12): 1881-1890. (doi:10.1128/CVI.00257-10).

Solomon, R.S., Schneider, D.A., Brayton, K.A., Ueti, M.W., Graca, T., Futse, J.E., Noh, S.M., Baszler, T.V., Palmer, G.H. 2011. Expression of Anaplasma marginale ankyrin repeat-containing proteins during infection of the mammalian host and tick vector. Infection and Immunity. 79(7):2847-2855.

Noh, S.M., Ueti, M.W., Palmer, G.H., Munderloh, U.G., Felsheim, R.F., Brayton, K.A. 2010. Stability and tick transmission phenotype of gfp-transformed Anaplasma marginale through a complete in vivo infection cycle. Applied and Environmental Microbiology. 77(1):330-334.

Galletti, M.F., Ueti, M.W., Knowles Jr, D.P., Brayton, K.A., Palmer, G.H. 2009. Independence of anaplasma marginale strains with high and low transmission efficiencies in the tick vector following simultaneous acquisition by feeding on a superinfected mammalian reservoir host. Infection and Immunity. 77(4):1459-64.

Last Modified: 10/21/2014
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