Project description:A collection of 1145 clones from an EST project on female tick salivary gland genes was hybridized on glass slides to RNA extracted from several feeding stages of adult female tick salivary glands, including unfed and replete, and from adult male ticks, either unfed or fed in the presence or absence of female ticks. In the female ticks, the early fed (<50 mg) and partially fed (30-200 mg) groups were very similar. The fast feeding (350-500 mg) and replete ticks were similar to each other, but different from the partially fed. The unfed ticks were more similar to the fast feeding – replete groups than the early fed-partially fed groups. In the males, there were differences between the males fed in the presence or absence of females, but overall, these groups were very similar. The unfed ticks were significantly different from the fed ticks. Males showed clear differences with females in expression, as well. The unfed females had high levels of genes involved in protein synthesis, while genes possibly involved in survival on the host, such as anticoagulants, seemed to be most expressed in the early and partially fed states. By contrast, in the males, the protein synthesis genes were expressed more in all three groups, while the putative secreted genes for survival were expressed less. Keywords: time course, effect of feeding, sex, effect of presence of females
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.
Project description:A collection of 1145 clones from an EST project on female tick salivary gland genes was hybridized on glass slides to RNA extracted from several feeding stages of adult female tick salivary glands, including unfed and replete, and from adult male ticks, either unfed or fed in the presence or absence of female ticks. In the female ticks, the early fed (<50 mg) and partially fed (30-200 mg) groups were very similar. The fast feeding (350-500 mg) and replete ticks were similar to each other, but different from the partially fed. The unfed ticks were more similar to the fast feeding â replete groups than the early fed-partially fed groups. In the males, there were differences between the males fed in the presence or absence of females, but overall, these groups were very similar. The unfed ticks were significantly different from the fed ticks. Males showed clear differences with females in expression, as well. The unfed females had high levels of genes involved in protein synthesis, while genes possibly involved in survival on the host, such as anticoagulants, seemed to be most expressed in the early and partially fed states. By contrast, in the males, the protein synthesis genes were expressed more in all three groups, while the putative secreted genes for survival were expressed less. Keywords: time course, effect of feeding, sex, effect of presence of females All samples were compared to the partially fed females. Females consisted of five groups: unfed, early fed, partially fed, fast feeding and replete. Four or five biological replicates were done of each, with the dyes used in both possible ways. In the males, three groups were used: unfed, feeding in the presence of females, and feeding in the absence of females. Two biological replicates were done of the feeding males, and one of extracts was hybridized twice for the males fed in the presence of females. Unfed males used one RNA sample, extracted from a large pool of ticks.
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:Ticks are blood feeding arthropod ectoparasites that transmit pathogens, which cause diseases in humans and animals worldwide. In the past ten decades, the continuous human exploitation of environmental resources and the increase in human outdoor activities has promoted contact with arthropod vectors normally present in the wild, resulting in increased transmission of vector-borne pathogens. In addition, vector populations are expanding in response to climate change and human interventions that impact reservoir host movement and human exposure to infected vectors. Among these emerging vector-borne pathogens, Anaplasma phagocytophilum (Rickettsiales: Anaplasmataceae) has become an important tick-borne pathogen in the United States, Europe and Asia, with increasing numbers of infected people and animals every year. Diseases caused by A. phagocytophilum include human granulocytic anaplasmosis (HGA), equine and canine granulocytic anaplasmosis and tick-borne fever (TBF) in ruminants. The natural infection cycle of A. phagocytophilum is dependent upon the presence of infected vertebrate reservoir hosts and Ixodid tick vectors. In the United States and Europe the main vector species are Ixodes scapularis, Ixodes pacificus, and Ixodes ricinus, while a wide range of mammals, lizards, and birds serve as reservoir hosts for various A. phagocytophilum genotypes. A. phagocytophilum initially infects tick midgut cells and then subsequently develops in salivary glands for transmission to susceptible hosts during tick feeding where the pathogen infects granulocytic cells, primarily neutrophils. Anaplasma phagocytophilum develops within membrane-bound inclusions in the host cell cytoplasm. This pathogen has evolved with its tick and vertebrate hosts through dynamic processes involving genetic traits of the pathogen and hosts that collectively mediate pathogen infection, development, persistence, and survival. However, the mechanisms used by A. phagocytophilum for molecular mechanisms involved in tick-pathogen interactions have not been fully characterized. The objective of this study is to characterize the dynamics of the microRNA response in the tick vector Ixodes scapularis in response to A. phagocytophilum infection. To address this objective, the composition of tick microRNAs was characterize using RNA sequencing in I. scapularis tick cells in response to A. phagocytophilum infection. The discovery of these mechanisms provides evidence that a control strategy could be developed targeted at both vertebrate and tick hosts for more complete control of A. phagocytophilum and its associated diseases.
Project description:Polymerase chain reaction analysis of 204 Amblyomma americanum and 28 A. maculatum ticks collected in August 1999 near the homes of patients with southern tick-associated rash illness and in control areas in Choctaw County, Alabama, showed Borrelia lonestari flagellin gene sequence from two adult A. americanum. The presence of B. lonestari in A. americanum ticks from Alabama suggests that this suspected pathogen may be widespread in the southeastern United States.
Project description:Pioneering studies (PXD014844) have identified many interesting molecules in tick saliva by LC-MS/MS proteomics, but the protein databases used to assign mass spectra were based on short Illumina reads of the Amblyomma americanum transcriptome and may not have captured the diversity and complexity of longer transcripts. Here we apply long-read Pacific Bioscience technologies to complement the previously reported short-read Illumina transcriptome-based proteome in an effort to increase spectrum assignments. Our dataset reveals a small increase in assignable spectra to supplement the previously released short-read transcriptome-based proteome.