Unknown,Transcriptomics,Genomics,Proteomics

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Comparison of Transcriptome Profiles of Human Ehrlichiosis Agents

ABSTRACT: This study is to determine and compare the transcriptomes in two eukaryotic hosts (mammalian host - DH82 canine cell line and tick vector - ISE6 Ixodes scapularis cell line) of eight Ehrlichia chaffeensis strains in three divergent genetic groups, including Arkansas, Heartland, Jax, Liberty, Osceola, St. Vincent, Wakulla, and West Paces, and one Ehrlichia sp. HF strain. RNA samples for the Arkansas (reference), Heartland, Jax, Liberty, Osceola, St. Vincent, Wakulla, and West Paces strains in both tick and canine cell lines, including two uninfected controls, will be isolated in triplicate. As such 108 libraries will be constructed and sequenced corresponding to (8 strains + 1 control) x 2 conditions (tick cells + dog cells) x 3 biological triplicates x 2 molecules sequenced (eukaryotic RNA + prokaryotic RNA). The transcriptomes are obtained with Illumina HiSeq reads obtained from paired end libraries. The bacterial reads will be mapped against the reference genome and compared through a differential expression analysis. The eukaryotic reads from the same cell types (i.e. tick or canine) will be assembled with Trinity. The reads will then be mapped back to this assembly for the differential expression analysis.

ORGANISM(S): Canis lupus familiaris

SUBMITTER: Sean Daugherty

PROVIDER: E-GEOD-56339 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Background Human monocytotropic ehrlichiosis is an emerging life-threatening zoonosis caused by obligately intracellular bacterium, Ehrlichia chaffeensis. E. chaffeensis is transmitted by the lone star tick, Amblyomma americanum, and replicates in mononuclear phagocytes in mammalian hosts. Differences in the E. chaffeensis transcriptome in mammalian and arthropod hosts are unknown. Thus, we determined host-specific E. chaffeensis gene expression in human monocyte (THP-1) and in Amblyomma and Ixodes tick cell lines (AAE2 and ISE6) using a whole genome microarray. Methodology/Principal Findings The majority (~80%) of E. chaffeensis genes were expressed during infection in human and tick cells. There were few differences observed in E. chaffeensis gene expression between the vector Amblyomma and non-vector Ixodes tick cells, but extensive host-specific and differential gene expression profiles were detected between human and tick cells, including higher transcriptional activity in tick cells and identification of gene subsets that were differentially expressed in the two hosts. Differentially and host-specifically expressed ehrlichial genes encoded major immunoreactive tandem repeat proteins (TRP), the outer membrane protein (OMP-1) family, and hypothetical proteins that were 30–80 amino acids in length. Consistent with previous observations, high expression of p28 and OMP-1B genes was detected in human and tick cells, respectively. Notably, E. chaffeensis genes encoding TRP32 and TRP47 were highly upregulated in the human monocytes and expressed as proteins; however, although TRP transcripts were expressed in tick cells, the proteins were not detected in whole cell lysates demonstrating that TRP expression was post transcriptionally regulated. Conclusions/Significance Ehrlichia gene expression is highly active in tick cells, and differential gene expression among a wide variety of host-pathogen associated genes occurs. Furthermore, we demonstrate that genes associated with host-pathogen interactions are differentially expressed and regulated by post transcriptional mechanisms. A microarray (4-plex) study using E. chaffeensis cultivated in each cell line (THP-1, AAE2 and ISE6), three biological replicates/cell line. For each cell line, RNA was also extracted from uninfected cells (negative controls) and was processed similar to the infected cells; these samples were used for background subtraction during data analysis.

Project description:Anaplasma phagocytophilum is an emerging zoonotic pathogen that causes human granulocytic anaplasmosis. These intracellular bacteria establish infection by affecting cell function in both the vertebrate host and the tick vector, Ixodes scapularis. Previous studies have characterized the tick transcriptome and proteome in response to A. phagocytophilum infection. However, in the post-genomic era, the integration of omics datasets through a systems biology approach allows network-based analyses to describe the complexity and functionality of biological systems such as host-pathogen interactions and the discovery of new targets for prevention and control of infectious diseases. This study reports for the first time a systems biology integration of metabolomics, transcriptomics and proteomics data to characterize essential metabolic pathways involved in the response of tick cells to A. phagocytophilum infection. The results showed that infection affected protein processing in endoplasmic reticulum and glucose metabolic pathways in tick cells. These results supported tick-Anaplasma co-evolution by providing new evidence of how tick cells limit pathogen infection, while the pathogen benefits from the tick cell response to establish infection. The results suggested that A. phagocytophilum induces protein misfolding to limit the tick cell response and facilitate infection, but requires protein degradation to prevent ER stress and cell apoptosis to survive in infected cells. Additionally, A. phagocytophilum may benefit from the tick cellâs ability to limit bacterial infection through PEPCK inhibition leading to decreased glucose metabolism, which also results in the inhibition of cell apoptosis that increases infection of tick cells. These results support the use of this experimental approach to systematically identify tick cell pathways and molecular mechanisms involved in tick-pathogen interactions Two samples with two replicates each were analyzed. Samples included Ixodes scapularis ISE6 cells uninfected (control) and infected with Anaplasma phagocytophilum human NY18 isolate.

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Comparison of Transcriptome Profiles of Human Ehrlichiosis Agents

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