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Bioinformatic prediction of G protein-coupled receptor encoding sequences from the transcriptome of the foreleg, including the Haller's organ, of the cattle tick, Rhipicephalus australis.

ABSTRACT: The cattle tick of Australia, Rhipicephalus australis, is a vector for microbial parasites that cause serious bovine diseases. The Haller's organ, located in the tick's forelegs, is crucial for host detection and mating. To facilitate the development of new technologies for better control of this agricultural pest, we aimed to sequence and annotate the transcriptome of the R. australis forelegs and associated tissues, including the Haller's organ. As G protein-coupled receptors (GPCRs) are an important family of eukaryotic proteins studied as pharmaceutical targets in humans, we prioritized the identification and classification of the GPCRs expressed in the foreleg tissues. The two forelegs from adult R. australis were excised, RNA extracted, and pyrosequenced with 454 technology. Reads were assembled into unigenes and annotated by sequence similarity. Python scripts were written to find open reading frames (ORFs) from each unigene. These ORFs were analyzed by different GPCR prediction approaches based on sequence alignments, support vector machines, hidden Markov models, and principal component analysis. GPCRs consistently predicted by multiple methods were further studied by phylogenetic analysis and 3D homology modeling. From 4,782 assembled unigenes, 40,907 possible ORFs were predicted. Using Blastp, Pfam, GPCRpred, TMHMM, and PCA-GPCR, a basic set of 46 GPCR candidates were compiled and a phylogenetic tree was constructed. With further screening of tertiary structures predicted by RaptorX, 6 likely GPCRs emerged and the strongest candidate was classified by PCA-GPCR to be a GABAB receptor.

SUBMITTER: Munoz S

PROVIDER: S-EPMC5322884 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Bioinformatic prediction of G protein-coupled receptor encoding sequences from the transcriptome of the foreleg, including the Haller's organ, of the cattle tick, Rhipicephalus australis.

Munoz Sergio S Guerrero Felix D FD Kellogg Anastasia A Heekin Andrew M AM Leung Ming-Ying MY

PloS one 20170223 2

The cattle tick of Australia, Rhipicephalus australis, is a vector for microbial parasites that cause serious bovine diseases. The Haller's organ, located in the tick's forelegs, is crucial for host detection and mating. To facilitate the development of new technologies for better control of this agricultural pest, we aimed to sequence and annotate the transcriptome of the R. australis forelegs and associated tissues, including the Haller's organ. As G protein-coupled receptors (GPCRs) are an im ...[more]

PMID: 28231302

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Project description:BackgroundDisease emergence and production loss caused by cattle tick infestations have focused attention on genetic selection strategies to breed beef cattle with increased tick resistance. However, the mechanisms behind host responses to tick infestation have not been fully characterised. Hence, this study examined gene expression profiles of peripheral blood leukocytes from tick-naive Brangus steers (Bos taurus x Bos indicus) at 0, 3, and 12 weeks following artificial tick challenge experiments with Rhipicephalus australis larvae. The aim of the study was to investigate the effect of tick infestation on host leukocyte response to explore genes associated with the expression of high and low host resistance to ticks.ResultsAnimals with high (HR, n = 5) and low (LR, n = 5) host resistance were identified after repeated tick challenge. A total of 3644 unique differentially expressed genes (FDR < 0.05) were identified in the comparison of tick-exposed (both HR and LR) and tick-naive steers for the 3-week and 12-week infestation period. Enrichment analyses showed genes were involved in leukocyte chemotaxis, coagulation, and inflammatory response. The IL-17 signalling, and cytokine-cytokine interactions pathways appeared to be relevant in protection and immunopathology to tick challenge. Comparison of HR and LR phenotypes at timepoints of weeks 0, 3, and 12 showed there were 69, 8, and 4 differentially expressed genes, respectively. Most of these genes were related to immune, tissue remodelling, and angiogenesis functions, suggesting this is relevant in the development of resistance or susceptibility to tick challenge.ConclusionsThis study showed the effect of tick infestation on Brangus cattle with variable phenotypes of host resistance to R. australis ticks. Steers responded to infestation by expressing leukocyte genes related to chemotaxis, cytokine secretion, and inflammatory response. The altered expression of genes from the bovine MHC complex in highly resistant animals at pre- and post- infestation stages also supports the relevance of this genomic region for disease resilience. Overall, this study offers a resource of leukocyte gene expression data on matched tick-naive and tick-infested steers relevant for the improvement of tick resistance in composite cattle.

Project description:Ticks are the vector of many human and animal diseases; and host detection is critical to this process. Ticks have a unique sensory structure located exclusively on the 1st pairs of legs; the fore-tarsal Haller's organ, not found in any other animals, presumed to function like the insect antennae in chemosensation but morphologically very different. The mechanism of tick chemoreception is unknown. Utilizing next-generation sequencing and comparative transcriptomics between the 1st and 4th legs (the latter without the Haller's organ), we characterized 1st leg specific and putative Haller's organ specific transcripts from adult American dog ticks, Dermacentor variabilis. The analysis suggested that the Haller's organ is involved in olfaction, not gustation. No known odorant binding proteins like those found in insects, chemosensory lipocalins or typical insect olfactory mechanisms were identified; with the transcriptomic data only supporting a possible olfactory G-protein coupled receptor (GPCR) signal cascade unique to the Haller's organ. Each component of the olfactory GPCR signal cascade was identified and characterized. The expression of GPCR, G?o and ?-arrestin transcripts identified exclusively in the 1st leg transcriptome, and putatively Haller's organ specific, were examined in unfed and blood-fed adult female and male D. variabilis. Blood feeding to repletion in adult females down-regulated the expression of all three chemosensory transcripts in females but not in males; consistent with differences in post-feeding tick behavior between sexes and an expected reduced chemosensory function in females as they leave the host. Data are presented for the first time of the potential hormonal regulation of tick chemosensation; behavioral assays confirmed the role of the Haller's organ in N,N-diethyl-meta-toluamide (DEET) repellency but showed no role for the Haller's organ in host attachment. Further research is needed to understand the potential role of the GPCR cascade in olfaction.

Project description:The protein tyrosine phosphatase receptor type-C (PTPRC) gene encodes the common leukocyte antigen (CD45) receptor. CD45 affects cell adhesion, migration, cytokine signalling, cell development, and activation state. Four families of the gene have been identified in cattle: a taurine group (Family 1), two indicine groups (Families 2 and 4) and an African "taurindicine" group (Family 3). Host resistance in cattle to infestation with ticks is moderately heritable and primarily manifests as prevention of attachment and feeding by larvae. This study was conducted to describe the effects of PTPRC genotype on immune-response phenotypes in cattle that display a variable immune responsiveness to ticks. Thirty tick-naïve Santa-Gertrudis cattle (a stabilized composite of 5/8 taurine and 3/8 indicine) were artificially infested with ticks weekly for 13 weeks and ranked according to their tick counts. Blood samples were taken from control and tick-challenged cattle immediately before, then at 21 d after infestation and each subsequent week for 9 weeks. Assays included erythrocyte profiles, white blood cell counts, the percentage of cellular subsets comprising the peripheral blood mononuclear cell (PBMC) population, and the ability of PBMC to recognize and proliferate in response to stimulation with tick antigens in vitro. The cattle were PTPRC genotyped using a RFLP assay that differentiated Family 1 and 3 together (220 bp), from Family 2 (462 bp), and from Family 4 (486 bp). The PTPRC allele frequencies were Family 1/3 = 0.34; Family 2 = 0.47; Family 4 = 0.19. There was no significant association between PTPRC genotype and tick count. Each copy of the Family 1/3 allele significantly decreased total leucocyte count (WCC) and CD8+ cells. Increasing dosage of Family 2 alleles significantly increased red blood cell count (RCC), haematocrit (PCV), and haemoglobin (Hb) concentration in blood. Increasing dosage of the Family 4 allele was associated with increased WCC, reduced RCC, reduced PCV and reduced Hb. Homozygote Family 1/3 animals had consistently lower IgG1 in response to tick Ag than homozygote Family 2 animals. The PTPRC genotype influences the bovine immune response to ticks but was not associated with the observed variation in resistance to tick infestation in this study.

Project description:BACKGROUND: Ticks are regarded as the most relevant vectors of disease-causing pathogens in domestic and wild animals. The cattle tick, Rhipicephalus (Boophilus) microplus, hinders livestock production in tropical and subtropical parts of the world where it is endemic. Tick microbiomes remain largely unexplored. The objective of this study was to explore the R. microplus microbiome by applying the bacterial 16S tag-encoded FLX-titanium amplicon pyrosequencing (bTEFAP) technique to characterize its bacterial diversity. Pyrosequencing was performed on adult males and females, eggs, and gut and ovary tissues from adult females derived from samples of R. microplus collected during outbreaks in southern Texas. RESULTS: Raw data from bTEFAP were screened and trimmed based upon quality scores and binned into individual sample collections. Bacteria identified to the species level include Staphylococcus aureus, Staphylococcus chromogenes, Streptococcus dysgalactiae, Staphylococcus sciuri, Serratia marcescens, Corynebacterium glutamicum, and Finegoldia magna. One hundred twenty-one bacterial genera were detected in all the life stages and tissues sampled. The total number of genera identified by tick sample comprised: 53 in adult males, 61 in adult females, 11 in gut tissue, 7 in ovarian tissue, and 54 in the eggs. Notable genera detected in the cattle tick include Wolbachia, Coxiella, and Borrelia. The molecular approach applied in this study allowed us to assess the relative abundance of the microbiota associated with R. microplus. CONCLUSIONS: This report represents the first survey of the bacteriome in the cattle tick using non-culture based molecular approaches. Comparisons of our results with previous bacterial surveys provide an indication of geographic variation in the assemblages of bacteria associated with R. microplus. Additional reports on the identification of new bacterial species maintained in nature by R. microplus that may be pathogenic to its vertebrate hosts are expected as our understanding of its microbiota expands. Increased awareness of the role R. microplus can play in the transmission of pathogenic bacteria will enhance our ability to mitigate its economic impact on animal agriculture globally. This recognition should be included as part of analyses to assess the risk for re-invasion of areas like the United States of America where R. microplus was eradicated.

Project description:BACKGROUND: The southern cattle tick, Rhipicephalus (Boophilus) microplus, is an economically important parasite of cattle and can transmit several pathogenic microorganisms to its cattle host during the feeding process. Understanding the biology and genomics of R. microplus is critical to developing novel methods for controlling these ticks. RESULTS: We present a global comparative genomic analysis of a gene index of R. microplus comprised of 13,643 unique transcripts assembled from 42,512 expressed sequence tags (ESTs), a significant fraction of the complement of R. microplus genes. The source material for these ESTs consisted of polyA RNA from various tissues, lifestages, and strains of R. microplus, including larvae exposed to heat, cold, host odor, and acaricide. Functional annotation using RPS-Blast analysis identified conserved protein domains in the conceptually translated gene index and assigned GO terms to those database transcripts which had informative BlastX hits. Blast Score Ratio and SimiTri analysis compared the conceptual transcriptome of the R. microplus database to other eukaryotic proteomes and EST databases, including those from 3 ticks. The most abundant protein domains in BmiGI were also analyzed by SimiTri methodology. CONCLUSION: These results indicate that a large fraction of BmiGI entries have no homologs in other sequenced genomes. Analysis with the PartiGene annotation pipeline showed 64% of the members of BmiGI could not be assigned GO annotation, thus minimal information is available about a significant fraction of the tick genome. This highlights the important insights in tick biology which are likely to result from a tick genome sequencing project. Global comparative analysis identified some tick genes with unexpected phylogenetic relationships which detailed analysis attributed to gene losses in some members of the animal kingdom. Some tick genes were identified which had close orthologues to mammalian genes. Members of this group would likely be poor choices as targets for development of novel tick control technology.

Dataset Information

Bioinformatic prediction of G protein-coupled receptor encoding sequences from the transcriptome of the foreleg, including the Haller's organ, of the cattle tick, Rhipicephalus australis.

Publications

Bioinformatic prediction of G protein-coupled receptor encoding sequences from the transcriptome of the foreleg, including the Haller's organ, of the cattle tick, Rhipicephalus australis.

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