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Expression of Anaplasma marginale ankyrin repeat-containing proteins during infection of the mammalian host and tick vector.

ABSTRACT: Transmission of tick-borne pathogens requires transition between distinct host environments with infection and replication in host-specific cell types. Anaplasma marginale illustrates this transition: in the mammalian host, the bacterium infects and replicates in mature (nonnucleated) erythrocytes, while in the tick vector, replication occurs in nucleated epithelial cells. We hypothesized that proteins containing ankyrin motifs would be expressed by A. marginale only in tick cells and would traffic to the infected host cell nucleus. A. marginale encodes three proteins containing ankyrin motifs, an AnkA orthologue (the AM705 protein), AnkB (the AM926 protein), and AnkC (the AM638 protein). All three A. marginale Anks were confirmed to be expressed during intracellular infection: AnkA is expressed at significantly higher levels in erythrocytes, AnkB is expressed equally by both infected erythrocytes and tick cells, and AnkC is expressed exclusively in tick cells. There was no evidence of any of the Ank proteins trafficking to the nucleus. Thus, the hypothesis that ankyrin-containing motifs were predictive of cell type expression and nuclear localization was rejected. In contrast, AnkA orthologues in the closely related A. phagocytophilum and Ehrlichia chaffeensis have been shown to localize to the host cell nucleus. This difference, together with the lack of a nuclear localization signal in any of the AnkA orthologues, suggests that trafficking may be mediated by a separate transporter rather than by endogenous signals. Selection for divergence in Ank function among Anaplasma and Ehrlichia spp. is supported by both locus and allelic analyses of genes encoding orthologous proteins and their ankyrin motif compositions.

SUBMITTER: Ramabu SS

PROVIDER: S-EPMC3191954 | biostudies-literature | 2011 Jul

REPOSITORIES: biostudies-literature

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Expression of Anaplasma marginale ankyrin repeat-containing proteins during infection of the mammalian host and tick vector.

Ramabu Solomon S SS Schneider David A DA Brayton Kelly A KA Ueti Massaro W MW Graça Telmo T Futse James E JE Noh Susan M SM Baszler Timothy V TV Palmer Guy H GH

Infection and immunity 20110516 7

Transmission of tick-borne pathogens requires transition between distinct host environments with infection and replication in host-specific cell types. Anaplasma marginale illustrates this transition: in the mammalian host, the bacterium infects and replicates in mature (nonnucleated) erythrocytes, while in the tick vector, replication occurs in nucleated epithelial cells. We hypothesized that proteins containing ankyrin motifs would be expressed by A. marginale only in tick cells and would traf ...[more]

PMID: 21576345

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Project description:UnlabelledTick-borne transmission of bacterial pathogens in the order Rickettsiales is responsible for diverse infectious diseases, many of them severe, in humans and animals. Transmission dynamics differ among these pathogens and are reflected in the pathogen-vector interaction. Anaplasma marginale has been shown to establish and maintain infectivity within Dermacentor spp. for weeks to months while escaping the complex network of vacuolar peptidases that are responsible for digestion of the tick blood meal. How this prolonged maintenance of infectivity in a potentially hostile environment is achieved has been unknown. Using the natural vector Dermacentor andersoni, we demonstrated that A. marginale-infected tick vacuoles (AmVs) concurrently recruit markers of the early endosome (Rab5), recycling endosome (Rab4 and Rab11), and late endosome (Rab7), are maintained near neutral pH, do not fuse with lysosomes, exclude the protease cathepsin L, and engage the endoplasmic reticulum and Golgi apparatus for up to 21 days postinfection. Maintenance of this safe vacuolar niche requires active A. marginale protein synthesis; in its absence, the AmVs mature into acidic, protease-active phagolysosomes. Identification of this bacterially directed modeling of the tick midgut endosome provides a mechanistic basis for examination of the differences in transmission efficiency observed among A. marginale strains and among vector populations.ImportanceTicks transmit a variety of intracellular bacterial pathogens that cause significant diseases in humans and animals. For successful transmission, these bacterial pathogens must first gain entry into the tick midgut digestive cells, avoid digestion, and establish a replicative niche without harming the tick vector. Little is known about how this replicative niche is established and maintained. Using the ruminant pathogen A. marginale and its natural tick vector, D. andersoni, this study characterized the features of the A. marginale niche in the tick midgut and demonstrates that A. marginale protein synthesis is required for the maintenance of this niche. This work opens a new line of inquiry about the pathogen effectors and their targets within the tick that mediate tick-pathogen interactions and ultimately serve as the determinants of pathogen success.

Project description:BACKGROUND: The tick-borne pathogen Anaplasma marginale, which is endemic worldwide, is the type species of the genus Anaplasma (Rickettsiales: Anaplasmataceae). Rhipicephalus (Boophilus) microplus is the most important tick vector of A. marginale in tropical and subtropical regions of the world. Despite extensive characterization of the genetic diversity in A. marginale geographic strains using major surface protein sequences, little is known about the biogeography and evolution of A. marginale and other Anaplasma species. For A. marginale, MSP1a was shown to be involved in vector-pathogen and host-pathogen interactions and to have evolved under positive selection pressure. The MSP1a of A. marginale strains differs in molecular weight because of a variable number of tandem 23-31 amino acid repeats and has proven to be a stable marker of strain identity. While phylogenetic studies of MSP1a repeat sequences have shown evidence of A. marginale-tick co-evolution, these studies have not provided phylogeographic information on a global scale because of the high level of MSP1a genetic diversity among geographic strains. RESULTS: In this study we showed that the phylogeography of A. marginale MSP1a sequences is associated with world ecological regions (ecoregions) resulting in different evolutionary pressures and thence MSP1a sequences. The results demonstrated that the MSP1a first (R1) and last (RL) repeats and microsatellite sequences were associated with world ecoregion clusters with specific and different environmental envelopes. The evolution of R1 repeat sequences was found to be under positive selection. It is hypothesized that the driving environmental factors regulating tick populations could act on the selection of different A. marginale MSP1a sequence lineages, associated to each ecoregion. CONCLUSION: The results reported herein provided the first evidence that the evolution of A. marginale was linked to ecological traits affecting tick vector performance. These results suggested that some A. marginale strains have evolved under conditions that support pathogen biological transmission by R. microplus, under different ecological traits which affect performance of R. microplus populations. The evolution of other A. marginale strains may be linked to transmission by other tick species or to mechanical transmission in regions where R. microplus is currently eradicated. The information derived from this study is fundamental toward understanding the evolution of other vector-borne pathogens.

Project description:BackgroundThe intracellular bacteria Anaplasma phagocytophilum are emerging zoonotic pathogens affecting human and animal health, and a good model for the study of tick-host-pathogen interactions. This tick-borne pathogen is transmitted by Ixodes scapularis in the United States where it causes human granulocytic anaplasmosis. Tick midguts and salivary glands play a major role during tick feeding and development, and in pathogen acquisition, multiplication and transmission. Vertebrate host proteins are found in tick midguts after feeding and have been described in the salivary glands of fed and unfed ticks, suggesting a role for these proteins during tick feeding and development. Furthermore, recent results suggested the hypothesis that pathogen infection affects tick metabolic processes to modify host protein digestion and persistence in the tick with possible implications for tick physiology and pathogen life-cycle.MethodsTo address this hypothesis, herein we used I. scapularis female ticks fed on uninfected and A. phagocytophilum-infected sheep to characterize host protein content in midguts and salivary glands by proteomic analysis of tick tissues.ResultsThe results evidenced a clear difference in the host protein content between tick midguts and salivary glands in response to infection suggesting that A. phagocytophilum selectively manipulates the levels of vertebrate host proteins in ticks in a tissue-specific manner to facilitate pathogen infection, multiplication and transmission while preserving tick feeding and development. The mechanisms by which A. phagocytophilum manipulates the levels of vertebrate host proteins are not known, but the results obtained here suggested that it might include the modification of proteolytic pathways.ConclusionsThe results of this study provided evidence to support that A. phagocytophilum affect tick proteolytic pathways to selectively manipulate the levels of vertebrate host proteins in a tissue-specific manner to increase tick vector capacity. Investigating the biological relevance of host proteins in tick biology and pathogen infection and the mechanisms used by A. phagocytophilum to manipulate host protein content is essential to advance our knowledge of tick-host-pathogen molecular interactions. These results have implications for the identification of new targets for the development of vaccines for the control of tick-borne diseases.

Project description:Antigenic variation of major surface proteins is considered an immune-evasive maneuver used by pathogens as divergent as bacteria and protozoa. Likewise, major surface protein 2 (Msp2) of the tick-borne pathogen, Anaplasma marginale, is thought to be involved in antigenic variation to evade the mammalian host immune response. However, this dynamic process also works in the tick vector in the absence of immune selection pressure. We examined Msp2 variants expressed during infection of four tick and two mammalian cell-lines to determine if the presence of certain variants correlated with specific host cell types. Anaplasma marginale colonies differed in their development and appearance in each of the cell lines (P<0.001). Using Western blots probed with two Msp2-monospecific and one Msp2-monoclonal antibodies, we detected expression of variants with differences in molecular weight. Immunofluorescence-assay revealed that specific antibodies bound from 25 to 60% of colonies, depending on the host cell-line (P<0.001). Molecular analysis of cloned variant-encoding genes demonstrated expression of different predominant variants in tick (V1) and mammalian (V2) cell-lines. Analysis of the putative secondary structure of the variants revealed a change in structure when A. marginale was transferred from one cell-type to another, suggesting that the expression of particular Msp2 variants depended on the cell-type (tick or mammalian) in which A. marginale developed. Similarly, analysis of the putative secondary structure of over 200 Msp2 variants from ticks, blood samples, and other mammalian cells available in GenBank showed the predominance of a specific structure during infection of a host type (tick versus blood sample), demonstrating that selection of a possible structure also occurred in vivo. The selection of a specific structure in surface proteins may indicate that Msp2 fulfils an important role in infection and adaptation to diverse host systems. Supplemental Abstract in Spanish (File S1) is provided.

Dataset Information

Expression of Anaplasma marginale ankyrin repeat-containing proteins during infection of the mammalian host and tick vector.

Publications

Expression of Anaplasma marginale ankyrin repeat-containing proteins during infection of the mammalian host and tick vector.

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