Exploring the midgut proteome of partially fed female cattle tick (Rhipicephalus (Boophilus) microplus) (original) (raw)
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Transmembrane proteins – Mining the cattle tick transcriptome
Ticks and Tick-borne Diseases, 2015
Managing the spread and load of pathogen-transmitting ticks is an important task worldwide. The cattle tick, Rhipicephalus microplus, not only impacts the economy through losses in dairy and meat production, but also raises concerns for human health in regards to the potential of certain transmitted pathogens becoming zoonotic. However, novel strategies to control R. microplus are hindered by lack of understanding tick biology and the discovery of suitable vaccine or acaricide targets. The importance of transmembrane proteins as vaccine targets are well known, as is the case in tick vaccines with Bm86 as antigen. In this study, we describe the localization and functional annotation of 878 putative transmembrane proteins. Thirty proteins could be confirmed in the R. microplus gut using LC-MS/MS analysis and their roles in tick biology are discussed. To the best of our knowledge, 19 targets have not been reported before in any proteomics study in various tick species and the possibility of using the identified proteins as targets for tick control are discussed. Although tissue expression of identified putative proteins through expansive proteomics is necessary, this study demonstrates the possibility of using bioinformatics for the identification of targets for further evaluation in tick control strategies. Protein identity Antigen name Species Source Protein type(s) Experimental host(s) Vaccine efficacy Reference 5'-nucleotidase 5'-nucleotidase (4F8) Rhipicephalus microplus Malpighian tubules Recombinant Cattle NSE Hope et al., 2010 Akirin rAKR Various Whole mosquito Recombinant Red deer 25-33% reduction in tick infestation Carreón et al. 2012 Angiotensin-converting enzyme Bm91 Rhipicephalus microplus Salivary glands Recombinant Cattle 6 and 8% reproductive efficiency and egg viability Lambertz et al., 2012 Angiotensin-converting enzyme Bm91 Rhipicephalus microplus Salivary glands Native Cattle ~37% reduction in eggs Riding et al., 1994 Aquaporin Aquaporin Rhipicephalus microplus Midgut Recombinant Cattle 75% and 68% Guerrero et al. 2014 Aspartic proteinase BYC Rhipicephalus microplus Eggs Native Cattle 14%and 36% da Silva Vaz et al., 1998; Seixas et al., 2012 Aspartic proteinase BYC Rhipicephalus microplus Eggs Recombinant Cattle 25% Leal et al., 2006; Seixas et al., 2012 Cement protein 64TRP Rhipicephalus appendiculatus Salivary glands Recombinant Guinea pigs ~62% mortality Trimnell et al., 2002 Cement protein 64TRP Rhipicephalus sangiuneus Salivary glands Recombinant Guinea pigs ~47% mortality Trimnell et al., 2005 Cement protein TrP64 Rhipicephalus appendiculatus N/i Recombinant Rabbits 43.9% Saimo et al. 2011 Component of vitellin GP80 Rhipicephalus microplus Various Native Sheep 68% Tellam et al., 2002 Component of vitellogenin, vitellin VIT87 Rhipicephalus microplus Eggs Native Sheep 68% Tellam et al., 2002 Elongation factor 1 alpha Ef1a Rhipicephalus microplus Various Recombinant Cattle 31% Almazán et al., 2012 Protein identity Antigen name Species Source Protein type(s) Experimental host(s) Vaccine efficacy Reference Extracellular matrix protein, Glycine-rich protein RH50 Rhipicephalus haemaphysaloides Salivary glands Recombinant Rabbits 30.5% mortality Zhou et al., 2006 Ferritin 2 RaFER2 Rhipicephalus annulatus Midgut Recombinant Cattle 72% Hajdusek et al., 2010 Ferritin 2 RmFER2 Rhipicephalus microplus Midgut Recombinant Cattle 64% Hajdusek et al., 2010 Glutathione S-transferase GST-Hl Rhipicephalus microplus Various Recombinant Cattle 57% Parizi et al., 2011 Glutathione S-transferase, Vitellin-degrading enzyme and Aspartic proteinase GST-Hl, VTDCE and BYC Rhipicephalus microplus Various Recombinant Cattle 35.3 to 61.6% protection against infestation Parizi et al., 2012 Mating factor, voraxinα Voraxinα Rhipicephalus appendiculatus Testis Recombinant Rabbits ~50% reduction in eggs Yamada et al., 2009 Metalloprotease BrRm-MP4 Rhipicephalus microplus Larvae Recombinant Cattle 60% Ali et al. 2015 Mucin BMA7 Rhipicephalus microplus Whole ticks, membrane fractions Native Cattle ~21% reduction in eggs McKenna et al., 1998 Ribosomal protein P0 pP0-KHL Rhipicephalus sanguineus Various Synthetic peptide Rabbits 90.25% Rodríguez-Mallon et al., 2012 Serpin-3 (RAS), Serpin-4 (RAS-4) and 36kDa immuno-dominant protein (RIM36) RAS-3, RAS-4 and RIM36 Rhipicephalus appendiculatus Salivary glands Recombinant Cattle ~27% mortality Imamura et al., 2008 Strain variant of Bm86 Bm95 Rhipicephalus microplus Midgut Recombinant Cattle 58-89% Garcia-Garcia et al., 2000 Strain variant of Bm86 Bm95 Rhipicephalus haemaphysaloides Midgut Recombinant Cattle 78.9-84.6% Sugumar et al., 2011 Protein identity Antigen name Species Source Protein type(s) Experimental host(s) Vaccine efficacy Reference Trypsin inhibitor BmLTI Rhipicephalus microplus Various Recombinant Cattle 32% Andreotti et al.,, 2012 Trypsin inhibitor BmTI Rhipicephalus microplus Salivary glands Native Cattle 72.8% Andreotti et al., 2002 Trypsin inhibitor BmTI-A Rhipicephalus microplus Salivary glands Synthetic Cattle ~18.4% Andreotti et al., 2007 Ubiquitin UBE Rhipicephalus annulatus Various Recombinant Cattle 15% and 22% depending construct preparation Almazán et al., 2010, Almazán et al., 2012 Ubiquitin UBE Rhipicephalus microplus Various Recombinant Cattle 55% Almazán et al., 2010 Unknown ARS antigen 1 Rhipicephalus microplus Midgut Recombinant Cattle 73-76%
Insect Biochemistry and Molecular Biology, 2007
We used gel electrophoresis and mass spectrometry to investigate differences in protein expression in ovarian tissues from Babesia bovis-infected and uninfected southern cattle tick, Rhipicephalus (Boophilus) microplus. Soluble and membrane proteins were extracted from ovaries of adult female ticks, and analyzed by isoelectric focusing (IEF) and one-dimensional or two-dimensional (2-D) gel electrophoresis. Protein patterns were analyzed for differences in expression between infected and uninfected ticks. 2-D separation of proteins revealed a number of proteins that appeared to be up-or down-regulated in response to infection with Babesia, in particular membrane/membrane-associated proteins and proteins in a low molecular mass range between 6 and 36 kDa. A selection of differentially expressed proteins was subjected to analysis by capillary-HPLC-electrospray tandem mass spectrometry (HPLC-ESI-MS/MS). Among the ovarian proteins that were up-regulated in infected ticks were calreticulin, two myosin subunits, an endoplasmic reticulum protein, a peptidyl-prolyl cis-trans isomerase (PPIase), a cytochrome c oxidase subunit, a glutamine synthetase, and a family of Kunitz-type serine protease inhibitors. Among the down-regulated ovarian proteins were another PPIase, a hemoglobin subunit, and a lysozyme. This study is part of an ongoing effort to establish a proteome database that can be utilized to investigate specific proteins involved in successful pathogen transmission.
Parasites & Vectors, 2015
Background: The argasid tick Ornithodoros erraticus is the vector of African swine fever virus and of several Borrelia species that cause human relapsing fever in the Iberian Peninsula. The tick midgut is part of the ectoparasite-host interface and expresses proteins that are vital for the survival of the tick. Midgut proteins are therefore potential targets for drug and/or vaccine design aimed at the development of new strategies for tick control. Thus, the aim of this work was the characterization of the proteome of the O. erraticus midgut before and after a blood meal trying to elucidate the induced changes upon blood feeding. Methods: Midgut tissues from unfed and engorged O. erraticus females were dissected and proteins were fractionated by centrifugation and SDS-PAGE, and the corresponding gel pieces analysed by LC-MS/MS. The identified proteins were classified according to their Protein Class and Molecular Function and the differences between fed and unfed specimens were analysed. Results: Overall 555 tick proteins were identified: 414 in the midgut of the unfed specimens and 376 in the fed specimens, of which 235 were present in both groups. The proteins with catalytic, binding and structural functions were the most numerous and abundant, consistent with their role in the intracellular processing of the blood meal. The analysis of some groups of proteins putatively involved directly in blood meal digestion, including protein digestion (peptidase activity), iron metabolism, enzymes involved in oxidative stress and detoxification and membrane traffic and transport proteins, detected some differences between the fed and unfed ticks Conclusions: This work reports for the first time the collection and analysis of the midgut proteome of an argasid tick species and provides molecular information about the argasid machinery involved in blood digestion. This information represents a starting point for the identification and selection of new targets for the development of alternative control strategies.
PLoS ONE, 2014
The cattle tick Rhipicephalus (Boophilus) microplus is one of the most harmful parasites affecting bovines. Similarly to other hematophagous ectoparasites, R. microplus saliva contains a collection of bioactive compounds that inhibit host defenses against tick feeding activity. Thus, the study of tick salivary components offers opportunities for the development of immunological based tick control methods and medicinal applications. So far, only a few proteins have been identified in cattle tick saliva. The aim of this work was to identify proteins present in R. microplus female tick saliva at different feeding stages. Proteomic analysis of R. microplus saliva allowed identifying peptides corresponding to 187 and 68 tick and bovine proteins, respectively. Our data confirm that (i) R. microplus saliva is complex, and (ii) that there are remarkable differences in saliva composition between partially engorged and fully engorged female ticks. R. microplus saliva is rich mainly in (i) hemelipoproteins and other transporter proteins, (ii) secreted cross-tick species conserved proteins, (iii) lipocalins, (iv) peptidase inhibitors, (v) antimicrobial peptides, (vii) glycine-rich proteins, (viii) housekeeping proteins and (ix) host proteins. This investigation represents the first proteomic study about R. microplus saliva, and reports the most comprehensive Ixodidae tick saliva proteome published to date. Our results improve the understanding of tick salivary modulators of host defense to tick feeding, and provide novel information on the tick-host relationship.
Parasites & Vectors, 2014
Background: Ticks represent a significant health risk to animals and humans due to the variety of pathogens they can transmit during feeding. The traditional use of chemicals to control ticks has serious drawbacks, including the selection of acaricide-resistant ticks and environmental contamination with chemical residues. Vaccination with the tick midgut antigen BM86 was shown to be a good alternative for cattle tick control. However, results vary considerably between tick species and geographic location. Therefore, new antigens are required for the development of vaccines controlling both tick infestations and pathogen infection/transmission. Tick proteins involved in tick-pathogen interactions may provide good candidate protective antigens for these vaccines, but appropriate screening procedures are needed to select the best candidates. Methods: In this study, we selected proteins involved in tick-Anaplasma (Subolesin and SILK) and tick-Babesia (TROSPA) interactions and used in vitro capillary feeding to characterize their potential as antigens for the control of cattle tick infestations and infection with Anaplasma marginale and Babesia bigemina. Purified rabbit polyclonal antibodies were generated against recombinant SUB, SILK and TROSPA and added to uninfected or infected bovine blood to capillary-feed female Rhipicephalus (Boophilus) microplus ticks. Tick weight, oviposition and pathogen DNA levels were determined in treated and control ticks. Results: The specificity of purified rabbit polyclonal antibodies against tick recombinant proteins was confirmed by Western blot and against native proteins in tick cell lines and tick tissues using immunofluorescence. Capillary-fed ticks ingested antibodies added to the blood meal and the effect of these antibodies on tick weight and oviposition was shown. However, no effect was observed on pathogen DNA levels. Conclusions: These results highlighted the advantages and some of the disadvantages of in vitro tick capillary feeding for the characterization of candidate tick protective antigens. While an effect on tick weight and oviposition was observed, the effect on pathogen levels was not evident probably due to high tick-to-tick variations among other factors. Nevertheless, these results together with previous results of RNA interference functional studies suggest that these proteins are good candidate vaccine antigens for the control of R. microplus infestations and infection with A. marginale and B. bigemina.
Veterinary Parasitology, 2008
Differences in protein expression in midgut tissue of uninfected and Babesia bovis-infected southern cattle ticks, Rhipicephalus (Boophilus) microplus, were investigated in an effort to establish a proteome database containing proteins involved in successful pathogen transmission. The electrophoretic separation of midgut membrane proteins was greatly improved by using liquid-phase isoelectric focusing combined with one-dimensional or two-dimensional (2-D) gel electrophoresis. A selection of differentially expressed proteins were subjected to analysis by capillary-HPLC-electrospray tandem mass spectrometry (HPLC-ESI-MS/MS). Among the identified Babesia-affected tick midgut proteins were six proteins that are implicated in signaling processes, including three Ca 2+-binding proteins, a guanine nucleotide-binding protein, a protein with signal peptide activity and a translocon-associated receptor protein. Up-regulation of five metabolic enzymes indicated parasite-induced changes in electron and proton transport, protein processing and retinoic acid metabolism. Among the down-regulated proteins were a molecular chaperone, a cytoskeletal protein and a multifunctional protein of the prohibitin family. Identification of these proteins may provide new insights into the molecular interactions between B. bovis and its tick vector, and could lead to identification of anti-tick and transmission-blocking vaccine candidates.
Parasites & vectors, 2017
The argasid tick Ornithodoros moubata is the main African vector of the human relapsing fever agent Borrelia duttoni and the African swine fever virus. Together with saliva, the tick midgut forms part of the host-tick-pathogen interface, and numerous midgut proteins play key functions in the blood digestion-related process and the infection and transmission of pathogens. This work explores the composition of the midgut proteome of unfed and fed O. moubata females with the aim to complete the biological information already obtained from the midgut transcriptome and provide a more robust and comprehensive perspective of this biological system. Midgut tissues taken from females before feeding and 48 h after feeding were subjected to LC/MS-MS analysis. After functional characterization and classification of the proteins identified, the differences in the proteome between unfed and fed females were analysed and discussed. Additionally, a detailed analysis of particular groups of proteins...
Parasites & Vectors
Background There have been ongoing efforts to identify anti-tick vaccine targets to protect cattle from infestation with cattle fever ticks Rhipicephalus (Boophilus) microplus. Two commercial vaccines based on the tick gut protein Bm86 have had variable effectiveness, which has led to poor acceptance, and numerous studies have attempted to identify vaccine antigens that will provide more consistently effective protection. Transcriptomic analysis of R. microplus led to identification of three aquaporin genes annotated to code for transmembrane proteins involved in the transport of water across cell membranes. Previous work showed that vaccination with full-length recombinant aquaporin 1 (RmAQP1) reduced tick burdens on cattle. Targeted silencing of aquaporin 2 (RmAQP2) expression suggested it might also be a good anti-tick vaccination target. Methods Three synthetic peptides from the predicted extracellular domains of RmAQP2 were used to vaccinate cattle. Peptides were conjugated to ...
Gene expression profiling of adult female tissues in feeding Rhipicephalus microplus cattle ticks
International Journal for Parasitology, 2013
The southern cattle tick, Rhipicephalus microplus, is an economically important pest, especially for resource-poor countries, both as a highly adaptive invasive species and prominent vector of disease. The increasing prevalence of resistance to chemical acaricides and variable efficacy of current tick vaccine candidates highlight the need for more effective control methods. In the absence of a fully annotated genome, the wealth of available expressed sequence tag (EST) sequence data for this species presents a unique opportunity to study the genes that are expressed in tissues involved in blood meal acquisition, digestion and reproduction during feeding. Utilizing a custom oligonucleotide microarray designed from available singletons (BmiGI Version 2.1) and EST sequences of R. microplus, the expression profiles in feeding adult female midgut, salivary glands and ovarian tissues were compared. From 13,456 assembled transcripts, 588 genes expressed in all three tissues were identified from fed adult females 20 days post infestation. The greatest complement of genes relate to translation and protein turnover. Additionally, a number of unique transcripts were identified for each tissue that relate well to their respective physiological/biological function/role(s). These transcripts include secreted anti-hemostatics and defense proteins from the salivary glands for acquisition of a blood meal, proteases as well as enzymes and transporters for digestion and nutrient acquisition from ingested blood in the midgut, and finally proteins and associated factors involved in DNA replication and cell-cycle control for oogenesis in the ovaries. Comparative analyses of adult female tissues during feeding enabled the identification of a catalogue of transcripts that may be essential for successful feeding and reproduction in the cattle tick, R. microplus. Future studies will increase our understanding of basic tick biology, allowing the identification of shared proteins/pathways among different tissues that may offer novel targets for the development of new tick control strategies.