Gene_locus Report for: 9acar-g3mm97

Rickettsia rickettsii is a tick-borne obligate intracellular bacterium that causes Rocky Mountain Spotted Fever (RMSF). In Brazil, two species of ticks in the genus Amblyomma, A. sculptum and A. aureolatum, are incriminated as vectors of this bacterium. Importantly, these two species present remarkable differences in susceptibility to R. rickettsii infection, where A. aureolatum is more susceptible than A. sculptum. In the current study, A. aureolatum and A. sculptum ticks were fed on suitable hosts previously inoculated with R. rickettsii, mimicking a natural infection. As control, ticks were fed on non-infected animals. Both midgut and salivary glands of all positively infected ticks were colonized by R. rickettsii. We did not observe ticks with infection restricted to midgut, suggesting that important factors for controlling rickettsial colonization were produced in this organ. In order to identify such factors, the total RNA extracted from the midgut (MG) was submitted to next generation RNA sequencing (RNA-seq). The majority of the coding sequences (CDSs) of A. sculptum differentially expressed by infection were upregulated, whereas most of modulated CDSs of A. aureolatum were downregulated. The functional categories that comprise upregulated CDSs of A. sculptum, for instance, metabolism, signal transduction, protein modification, extracellular matrix, and immunity also include CDSs of A. aureolatum that were downregulated by infection. This is the first study that reports the effects of an experimental infection with the highly virulent R. rickettsii on the gene expression of two natural tick vectors. The distinct transcriptional profiles of MG of A. sculptum and A. aureolatum upon infection stimulus strongly suggest that molecular factors in this organ are responsible for delineating the susceptibility to R. rickettsii. Functional studies to determine the role played by proteins encoded by differentially expressed CDSs in the acquisition of R. rickettsii are warranted and may be considered as targets for the development of strategies to control the tick-borne pathogens as well as to control the tick vectors.

Tick secretory proteins modulate haemostasis, inflammation and immune responses of the host and are attractive recombinant anti-tick vaccine candidates. Yet, many of the proteins have not been characterised due to the limited sequence availability for ticks and other arthropods for homology-based annotation. To address this limitation, we sequenced the salivary glands of the economically important adult male and female Rhipicephalus appendiculatus ticks during feeding. The quality-filtered Illumina sequencing reads were de novo assembled to generate a R. appendiculatus sialotranscriptome of 21,410 transcripts. A non-redundant set of 12,761 R. appendiculatus proteins was predicted from the transcripts, including 2134 putative secretory and 8237 putative housekeeping proteins. Secretory proteins accounted for most of the expression in the salivary gland transcriptome (63%). Of the secretory protein class, the Glycine-rich superfamily contributed 66% and the Lipocalin family 12% of the transcriptome expression. Differential expression analysis identified 1758 female and 2346 male up-regulated transcripts, suggesting varying blood-feeding mechanisms employed between female and male ticks. The sialotranscriptome assembled in this work, greatly improves on the sequence information available for R. appendiculatus and is a valuable resource for potential future vaccine candidate selection.

Hard ticks feed for several days or weeks on their hosts. Blood feeding is assisted by tick saliva, which is injected in the host skin regularly, alternating with blood ingestion. Tick saliva contains hundreds or thousands of different peptides and other bioactive compounds that assist feeding by inhibiting their hosts' blood clotting, platelet aggregation, vasoconstriction, as well as pain and itching. Immunomodulatory and antimicrobial peptides are also found in tick saliva. Molecular characterization of tick salivary compounds, or its sialome (from the Greek sialos = saliva), helps identification of possible antigens that might confer anti-tick immunity, as well as identifying novel pharmacologically active compounds. Amblyomma americanum is a major nuisance tick in Eastern and Southern US, being a vector of Theileria and Ehrlichia bacteria to animals and humans. Presently we report an RNAseq study concerning the salivary glands of adult female A. americanum ticks, which involved sequencing of four libraries collected at different times of feeding. A total of 5,792 coding sequences were deduced from the transcriptome assembly, 3,139 of which were publicly deposited, expanding from the previously available 146 salivary sequences found in GenBank. A remarkable time-dependent transcript expression was found, mostly related to secretory products, supporting the idea that ticks may have several "sialomes" that are expressed at different times during feeding. The molecular nature of this sialome switching remains unknown. The hyperlinked spreadsheet containing the deduced coding sequences can be found at http://exon.niaid.nih.gov/transcriptome/Amb_americanum/Ambame-web.xlsx.

Rickettsia rickettsii is a tick-borne obligate intracellular bacterium that causes Rocky Mountain Spotted Fever (RMSF). In Brazil, two species of ticks in the genus Amblyomma, A. sculptum and A. aureolatum, are incriminated as vectors of this bacterium. Importantly, these two species present remarkable differences in susceptibility to R. rickettsii infection, where A. aureolatum is more susceptible than A. sculptum. In the current study, A. aureolatum and A. sculptum ticks were fed on suitable hosts previously inoculated with R. rickettsii, mimicking a natural infection. As control, ticks were fed on non-infected animals. Both midgut and salivary glands of all positively infected ticks were colonized by R. rickettsii. We did not observe ticks with infection restricted to midgut, suggesting that important factors for controlling rickettsial colonization were produced in this organ. In order to identify such factors, the total RNA extracted from the midgut (MG) was submitted to next generation RNA sequencing (RNA-seq). The majority of the coding sequences (CDSs) of A. sculptum differentially expressed by infection were upregulated, whereas most of modulated CDSs of A. aureolatum were downregulated. The functional categories that comprise upregulated CDSs of A. sculptum, for instance, metabolism, signal transduction, protein modification, extracellular matrix, and immunity also include CDSs of A. aureolatum that were downregulated by infection. This is the first study that reports the effects of an experimental infection with the highly virulent R. rickettsii on the gene expression of two natural tick vectors. The distinct transcriptional profiles of MG of A. sculptum and A. aureolatum upon infection stimulus strongly suggest that molecular factors in this organ are responsible for delineating the susceptibility to R. rickettsii. Functional studies to determine the role played by proteins encoded by differentially expressed CDSs in the acquisition of R. rickettsii are warranted and may be considered as targets for the development of strategies to control the tick-borne pathogens as well as to control the tick vectors.

BACKGROUND: Ticks secrete a diverse mixture of secretory proteins into the host to evade its immune response and facilitate blood-feeding, making secretory proteins attractive targets for the production of recombinant anti-tick vaccines. The largely neglected tick species, Rhipicephalus zambeziensis, is an efficient vector of Theileria parva in southern Africa but its available sequence information is limited. Next generation sequencing has advanced sequence availability for ticks in recent years and has assisted the characterisation of secretory proteins. This study focused on the de novo assembly and annotation of the salivary gland transcriptome of R. zambeziensis and the temporal expression of secretory protein transcripts in female and male ticks, before the onset of feeding and during early and late feeding. RESULTS: The sialotranscriptome of R. zambeziensis yielded 23,631 transcripts from which 13,584 non-redundant proteins were predicted. Eighty-six percent of these contained a predicted start and stop codon and were estimated to be putatively full-length proteins. A fifth (2569) of the predicted proteins were annotated as putative secretory proteins and explained 52% of the expression in the transcriptome. Expression analyses revealed that 2832 transcripts were differentially expressed among feeding time points and 1209 between the tick sexes. The expression analyses further indicated that 57% of the annotated secretory protein transcripts were differentially expressed. Dynamic expression profiles of secretory protein transcripts were observed during feeding of female ticks. Whereby a number of transcripts were upregulated during early feeding, presumably for feeding site establishment and then during late feeding, 52% of these were downregulated, indicating that transcripts were required at specific feeding stages. This suggested that secretory proteins are under stringent transcriptional regulation that fine-tunes their expression in salivary glands during feeding. No open reading frames were predicted for 7947 transcripts. This class represented 17% of the differentially expressed transcripts, suggesting a potential transcriptional regulatory function of long non-coding RNA in tick blood-feeding. CONCLUSIONS: The assembled sialotranscriptome greatly expands the sequence availability of R. zambeziensis, assists in our understanding of the transcription of secretory proteins during blood-feeding and will be a valuable resource for future vaccine candidate selection.

Tick secretory proteins modulate haemostasis, inflammation and immune responses of the host and are attractive recombinant anti-tick vaccine candidates. Yet, many of the proteins have not been characterised due to the limited sequence availability for ticks and other arthropods for homology-based annotation. To address this limitation, we sequenced the salivary glands of the economically important adult male and female Rhipicephalus appendiculatus ticks during feeding. The quality-filtered Illumina sequencing reads were de novo assembled to generate a R. appendiculatus sialotranscriptome of 21,410 transcripts. A non-redundant set of 12,761 R. appendiculatus proteins was predicted from the transcripts, including 2134 putative secretory and 8237 putative housekeeping proteins. Secretory proteins accounted for most of the expression in the salivary gland transcriptome (63%). Of the secretory protein class, the Glycine-rich superfamily contributed 66% and the Lipocalin family 12% of the transcriptome expression. Differential expression analysis identified 1758 female and 2346 male up-regulated transcripts, suggesting varying blood-feeding mechanisms employed between female and male ticks. The sialotranscriptome assembled in this work, greatly improves on the sequence information available for R. appendiculatus and is a valuable resource for potential future vaccine candidate selection.

Hard ticks feed for several days or weeks on their hosts. Blood feeding is assisted by tick saliva, which is injected in the host skin regularly, alternating with blood ingestion. Tick saliva contains hundreds or thousands of different peptides and other bioactive compounds that assist feeding by inhibiting their hosts' blood clotting, platelet aggregation, vasoconstriction, as well as pain and itching. Immunomodulatory and antimicrobial peptides are also found in tick saliva. Molecular characterization of tick salivary compounds, or its sialome (from the Greek sialos = saliva), helps identification of possible antigens that might confer anti-tick immunity, as well as identifying novel pharmacologically active compounds. Amblyomma americanum is a major nuisance tick in Eastern and Southern US, being a vector of Theileria and Ehrlichia bacteria to animals and humans. Presently we report an RNAseq study concerning the salivary glands of adult female A. americanum ticks, which involved sequencing of four libraries collected at different times of feeding. A total of 5,792 coding sequences were deduced from the transcriptome assembly, 3,139 of which were publicly deposited, expanding from the previously available 146 salivary sequences found in GenBank. A remarkable time-dependent transcript expression was found, mostly related to secretory products, supporting the idea that ticks may have several "sialomes" that are expressed at different times during feeding. The molecular nature of this sialome switching remains unknown. The hyperlinked spreadsheet containing the deduced coding sequences can be found at http://exon.niaid.nih.gov/transcriptome/Amb_americanum/Ambame-web.xlsx.