The argasid tick Ornithodoros moubata is the main vector of the African swine fever and the human relapsing fever in Africa. As part of the host-parasite-pathogen interface, the tick midgut expresses key proteins for tick survival and tick-borne pathogen transmission. Accordingly, midgut proteins are potential targets for the development of new drugs and vaccines aimed at tick control, and obtaining proteomic and transcriptomic data from the O. moubata midgut would facilitate the identification of such target candidates. With this aim, we have assembled and characterized the midgut transcriptome of O. moubata females before and 48h after a blood meal, and identified the genes that are differentially expressed in the midgut after feeding. Overall, 23,863 transcripts were obtained, and of them, 9,164 were identified and annotated. The most represented molecular functions were catalytic and binding activities, and the most represented biological processes were metabolic, cellular and single-organism processes. KEGG analysis of the annotated sequences assigned up to 3,053 of them to 130 active pathways, among which, the top 30 pathways were mostly metabolic routes. Differential expression analysis between unfed and fed ticks detected 8,026 Differentially Expressed Genes (DEGs), 4,093 up-regulated and 3,933 down-regulated, respectively. The biological significance of these DEGs was further investigated using the KEEG, Pfam and GO databases. The functional groups of the genes/proteins predicted to be involved in the processes of blood digestion, nutrient transport and metabolism, and in responses related to defence and oxidative stress are discussed in more detail. This work reports the first midgut transcriptome analysis of an argasid tick species, and provides a wealth of novel molecular information about the argasid machinery involved in blood feeding and digestion. This information represents a starting point for the development of alternative strategies for tick control.

The filarial parasite Loa loa, the causative agent of loiasis, is endemic in Central and Western Africa infecting 3–13 million people. L. loa has been associated with fatal encephalopathic reactions in high Loa-infected individuals receiving ivermectin during mass drug administration programs for the control of onchocerciasis and lymphatic filariasis. In endemic areas, the only diagnostic method routinely used is the microscopic examination of mid-day blood samples by thick blood film. Improved methods for detection of L. loa are needed in endemic regions with limited resources, where delayed diagnosis results in high mortality. We have investigated the use of a loop-mediated isothermal amplification (LAMP) assay to facilitate rapid, inexpensive, molecular diagnosis of loiasis. Primers for LAMP were designed from a species-specific repetitive DNA sequence from L. loa retrieved from GenBank. Genomic DNA of a L. loa adult worm was used to optimize the LAMP conditions using a thermocycler or a conventional heating block. Amplification of DNA in the LAMP mixture was visually inspected for turbidity as well as addition of fluorescent dye. LAMP specificity was evaluated using DNA from other parasites; sensitivity was evaluated using DNA from L. loa 10-fold serially diluted. Simulated human blood samples spiked with DNA from L. loa were also tested for sensitivity. Upon addition of fluorescent dye, all positive reactions turned green while the negative controls remained orange under ambient light. After electrophoresis on agarose gels, a ladder of multiple bands of different sizes could be observed in positive samples. The detection limit of the assay was found to be as little as 0.5 ag of L. loa genomic DNA when using a heating block. We have designed, for the first time, a highly sensitive LAMP assay for the detection of L. loa which is potentially adaptable for field diagnosis and disease surveillance in loiasis-endemic areas.

BackgroundThe 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.MethodsMidgut 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.ResultsOverall 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 ticksConclusionsThis 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.Electronic supplementary materialThe online version of this article (doi:10.1186/s13071-015-1148-z) contains supplementary material, which is available to authorized users.

Ticks are parasites of great medical and veterinary importance since they are vectors of numerous pathogens that affect humans, livestock and pets. Among the argasids, several species of the genus Ornithodoros transmit serious diseases such as tick-borne human relapsing fever (TBRF) and African Swine Fever (ASF). In particular, Ornithodoros erraticus is the main vector of these two diseases in the Mediterranean while O. moubata is the main vector in Africa. The presence of these Ornithodoros ticks in domestic and peridomestic environments may greatly hinder the eradication of TBRF and ASF from endemic areas. In addition, there is a constant threat of reintroduction and spreading of ASF into countries from where it has been eradicated (Spain and Portugal) or where it was never present (the Caucasus, Russia and Eastern Europe). In these countries, the presence of Ornithodoros vectors could have a tremendous impact on ASF transmission and long-term maintenance. Therefore, elimination of these ticks from at least synanthropic environments would contribute heavily to the prevention and control of the diseases they transmit. Tick control is a difficult task and although several methods for such control have been used, none of them has been fully effective against all ticks and the problems they cause. Nevertheless, immunological control using anti-tick vaccines offers an attractive alternative to the traditional use of acaricides. The aim of the present paper is to offer a brief overview of the current status in control measure development for Ornithodoros soft ticks, paying special attention to the development of vaccines against O. erraticus and O. moubata. Thus, our contribution includes an analysis of the chief attributes that the ideal antigens for an anti-tick vaccine should have, an exhaustive compilation and analysis of the scant anti-soft tick vaccine trials carried out to date using both concealed and salivary antigens and, finally, a brief description of the new reverse vaccinology approaches currently used to identify new and more effective protective tick antigens.

Ticks are hematophagous vectors of great medical and veterinary importance because they transmit numerous pathogenic microorganisms to humans and animals. The argasid Ornithodoros erraticus is the main vector of tick-borne human relapsing fever and African swine fever in the Mediterranean Basin. Tick enterocytes express bioactive molecules that perform key functions in blood digestion, feeding, toxic waste processing and pathogen transmission. To explore new strategies for tick control, in this work we have obtained and compared the midgut transcriptomes of O. erraticus female ticks before and after a blood meal and identified the genes whose expression is differentially regulated after feeding. The transcript sequences were annotated, functionally and structurally characterised and their expression levels compared between both physiological conditions (unfed females and fed females at 2 days post-engorgement). Up to 29,025 transcripts were assembled, and 9290 of them corresponded to differentially expressed genes (DEGs) after feeding. Of these, 4656 genes were upregulated and nearly the same number of genes was downregulated in fed females compared to unfed females. BLASTN and BLASTX analyses of the 29,025 transcripts allowed the annotation of 9072 transcripts/proteins. Among them, the most numerous were those with catalytic and binding activities and those involved in diverse metabolic pathways and cellular processes. The analyses of functional groups of upregulated DEGs potentially related to the digestion of proteins, carbohydrates and lipids, and the genes involved in the defence response and response to oxidative stress, confirm that these processes are narrowly regulated in ticks, highlighting their complexity and importance in tick biology. The expression patterns of six genes throughout the blood digestion period revealed significant differences between these patterns, strongly suggesting that the transcriptome composition is highly dynamic and subjected to important variation along the trophogonic cycle. This may guide future studies aimed at improving the understanding of the molecular physiology of tick digestion and digestion-related processes. The current work provides a more robust and comprehensive understanding of the argasid tick digestive system.

Title Function-guided selection of midgut antigens from Ornithodoros erraticus ticks and an evaluation of their protective efficacy in rabbits.

BackgroundNew candidate protective antigens for tick vaccine development may be identified by selecting and testing antigen candidates that play key biological functions. After blood-feeding, tick midgut overexpresses proteins that play essential functions in tick survival and disease transmission. Herein, Ornithodoros erraticus midgut transcriptomic and proteomic data were examined in order to select functionally significant antigens upregulated after feeding to be tested as vaccine candidate antigens.MethodsTranscripts annotated as chitinases, tetraspanins, ribosomal protein P0 and secreted proteins/peptides were mined from the recently published O. erraticus midgut transcriptome and filtered in a second selection step using criteria based on upregulation after feeding, predicted antigenicity and expression in the midgut proteome. Five theoretical candidate antigens were selected, obtained as recombinant proteins and used to immunise rabbits: one chitinase (CHI), two tetraspanins (TSPs), the ribosomal protein P0 (RPP0) and one secreted protein PK-4 (PK4).ResultsRabbit vaccination with individual recombinant candidates induced strong humoral responses that mainly reduced nymph moulting and female reproduction, providing 30.2% (CHI), 56% (TSPs), 57.5% (RPP0) and 57.8% (PK4) protection to O. erraticus infestations and 19.6% (CHI), 11.1% (TSPs), 0% (RPP0) and 8.1% (PK4) cross-protection to infestations by the African tick Ornithodoros moubata. The joint vaccine efficacy of the candidates was assessed in a second vaccine trial reaching 66.3% protection to O. erraticus and 25.6% cross-protection to O. moubata.ConclusionsThese results (i) indicate that argasid chitinases and RPP0 are promising protective antigens, as has already been demonstrated for ixodid chitinases and RPP0, and could be included in vaccines targeting multiple tick species; (ii) reveal novel protective antigens tetraspanins and secreted protein PK-4, never tested before as protective antigens in ticks; and (iii) demonstrate that multi-antigenic vaccines increased vaccine efficacy compared with individual antigens. Lastly, our data emphasize the value of the tick midgut as a source of protective candidate antigens in argasids for tick control.

BackgroundThe 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.MethodsMidgut 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 that are involved in the processes of nutrient digestion and responses to the oxidative stress was carried out.Results1491 non-redundant tick proteins were identified: 1132 of them in the midgut of unfed ticks, 1138 in the midgut of fed ticks, and up to 779 shared by both physiological conditions. Overall, the comparative analysis of the midgut proteomes of O. moubata females before and after feeding did not reveal great differences in the number or class of proteins expressed, enzymatic composition or functional classification.ConclusionsThe hemoglobinolytic system in ixodids and argasids is very similar in spite of the fact that they display very different feeding and reproductive strategies. Although the main source of nutrients in ticks are proteins, lipids and carbohydrates also constitute significant nutritional sources and play an important part in the process of blood digestion. The genes and proteins involved in intracellular transport mechanisms, defensive responses, detoxifying responses and stress responses seem to be closely regulated, highlighting the complexity and importance of these processes in tick biology, which in turn assigns them a great interest as targets for therapeutic and immunological interventions.Electronic supplementary materialThe online version of this article (doi:10.1186/s13071-017-2300-8) contains supplementary material, which is available to authorized users.