Tick Physiology Research Articles

Anthropogenic electromagnetic radiation alters the transcription levels of the genes encoding the SIFamide and myoinhibitory peptide and their receptors in Ixodes ricinus synganglion

The research of the influences of man-made electromagnetic fields on tick physiology has been very sparse and long neglected since the pioneer studies published in 1996 and 2000. Once multiple behavioral tests confirmed an attraction and possible perception of electromagnetic fields in ticks, a new interest in this topic erupted in recent years. In this study, qRT-PCR is utilized to determine the changes in the mRNA transcript levels of neuropeptides SIFamide and myoinhibitory peptide (mip and sifa) and their representative receptors (mip-r1 and sifa-r1) in the synganglia of the tick Ixodes ricinus irradiated by 900 MHz radiofrequency electromagnetic field. It was determined that 40 V/m intensity has a significant suppressory effect on the transcript levels of all genes after at least 60 minutes of constant exposure in both sexes. Commonly occurring intensity of radiation in urban areas (2 V/m) produced an elevation in mRNA levels after various timespans in every gene. A significant decrease of transcript abundances was detected in females after one hour of exposure to 2 V/m. Results of this study widen the knowledge of EMF-induced alterations in the neurophysiology of I. ricinus, the most commonly distributed hard tick in Europe.

A multi-omics approach for understanding blood digestion dynamics in Ixodes scapularis and identification of anti-tick vaccine targets

Ixodes scapularis, the black-legged tick, is a major arthropod vector that transmits the causative agents of Lyme disease and several other pathogens of human significance. The tick midgut is the main tissue involved in blood acquisition and digestion and the first organ to have contact with pathogens ingested through the blood meal. Gene expression in the midgut before, during, and after a blood meal may vary in response to the physiological changes due to blood feeding. A systems biology approach based on RNA and protein sequencing was used to gain insight into the changes in tick midgut transcripts and proteins during blood ingestion (unfed and partially fed) and digestion (1-, 2-, 7-, and 14 days post detachment from the host) by the Ixodes scapularis female ticks. A total of 2,726 differentially expressed transcripts, and 449 proteins were identified across the time points. Genes involved in detoxification of xenobiotics, proteases, protease inhibitors, metabolism, and immunity were differentially expressed in response to blood feeding. Similarly, proteins corresponding to the same groups were also differentially expressed. Nine genes from major gene categories were chosen as potential vaccine candidates, and, using RNA interference, the effect of these gene knockdowns on tick biology was investigated. Knockdown of these genes had variable negative impacts on tick physiology, such as the inability to engorge fully and to produce eggs and increased mortality. These and additional gene targets provide opportunities to explore novel tick control strategies.

Microbiome of two adult tick species and their laboratory-reared offspring shows intra- and inter-species differences

Tick-borne pathogens are a significant threat to human and animal health. Exposing the microbial composition of ticks elucidates their potential role in transmitting pathogens and causing disease as well as uncovering their potential interaction with the hosting tick. Our study focused on characterizing the tick microbiome of adult females and their lab-reared larval offspring of two prevalent tick species found on dogs in Nigeria [Rhipicephalus sanguineus s.l. tropical lineage (R. linnaei) and Haemaphysalis leachi]. We investigated the relative phyla abundance, the alpha and beta diversities of microbial communities comparing tick species, and different development stages (adults versus larvae). To the best of our knowledge, this is the first analysis on H. leachi microbiome described from West Africa. Our findings revealed a diverse microbiome with significant differences across species and their developmental stages, highlighting the dominance of the Proteobacteria phylum, followed by Firmicutes and Actinobacteriota. In contrast to H. leachi, for R. linnaei we observed significant differences in the alpha and beta diversities of the microbiome of larvae and adult females. Predominant bacterial genera were identified in R. linnaei, particularly Arsenophonus and Coxiella, which showed increased abundance in adult ticks. In H. leachi, other predominant genera were detected, including Sphingomonas, Comamonas, and Williamsia. Our results contribute to the understanding of microbiome dynamics within ticks and offers insights of tick physiology for addressing public health concerns and developing effective strategies for pathogen control.

Revealing tick diversity: Chemical profiling and dynamics in scanning microscopy and molecular phylogenetics.

This study presents a comprehensive investigation into the evolutionary trajectories of Rhipicephalus ticks (Ixodidae) through the interpretation of molecular phylogenetics, elucidating their chromatographic spectrum. The use of advanced chromatographic tools in this study explored the dynamics chemical profiling, providing valuable insights into the evolutionary history and ecological adaptations. Prevalence of Rhipicephalus ticks was 4.5% in sheep and 3.9% in goats. The ITS2 sequence of the Rhipicephalus sanguineus (OK642408) and Rhipicephalus microplus (OK642409) form a distinct clade with sequences from other countries. The 16S rRNA sequences of R. sanguineus (OK560870) clustered with sequences form three lineages, tropical, temperate, and south-eastern. The Cox I gene-identified Rhipicephalus turanicus (OK623472) and R. microplus (OK623463) form separate clades with sequences. The HPLC chromatogram of tick samples reveals a diverse array of identified hydrocarbons, explained the complex chemical composition of their exoskeletons. This analytical approach provides valuable insights into the specific hydrocarbon profiles, allowing for potential applications in species differentiation, ecological studies, and a deeper understanding of the functional roles played by hydrocarbon compounds in tick physiology. The findings revealed the potential of applying molecular phylogenetics tools with chromatography not only to enhance our understanding of tick evolution but also to inform strategies for disease control and management in regions where Rhipicephalus ticks (Ixodidae) are endemic. RESEARCH HIGHLIGHTS: Chemical mapping utilizing advanced chromatographic techniques. Scanning microscopic insights high-resolution scanning tool to observe structural and morphological features of ticks at a molecular level. Molecular phylogeny data elucidate the evolutionary relationships among tick species.

Evaluation of the biological function of ribosomal protein S18 from cattle tick Rhipicephalus microplus

Rhipicephalus (Boophilus) microplus, also known as the cattle tick, causes severe parasitism and transmits different pathogens to vertebrate hosts, leading to massive economic losses. In the present study, we performed a functional characterization of a ribosomal protein from R. microplus to investigate its importance in blood feeding, egg production and viability. Ribosomal protein S18 (RPS18) is part of the 40S subunit, associated with 18S rRNA, and has been previously pointed to have a secondary role in different organisms. Rhipicephalus microplus RPS18 (RmRPS18) gene expression levels were modulated in female salivary glands during blood feeding. Moreover, mRNA levels in this tissue were 10 times higher than those in the midgut of fully engorged female ticks. Additionally, recombinant RmRPS18 was recognized by IgG antibodies from sera of cattle naturally or experimentally infested with ticks. RNAi-mediated knockdown of the RmRPS18 gene was performed in fully engorged females, leading to a significant (29 %) decrease in egg production. Additionally, egg hatching was completely impaired, suggesting that no viable eggs were produced by the RmRPS18-silenced group. Furthermore, antimicrobial assays revealed inhibitory activities against gram-negative Escherichia coli and gram-positive Staphylococcus aureus bacteria, affecting bacterial growth. Data presented here show the important role of RmRPS18 in tick physiology and suggest that RmRPS18 can be a potential target for the development of novel strategies for tick control.

Organ-specific bacterial microbiota in the engorged female Haemaphysalis longicornis ticks

ABSTRACT Bacterial microbiota are involved in tick physiology and varies with various factors. The variation of bacterial microbiota among tick organs is less investigated. In this study, the bacterial microbiota of four organs (ovaries, salivary glands, midgut, and Malpighian tubules) from the engorged female Haemaphysalis longicornis were surveyed using 16S rRNA high-throughput sequencing. Results indicated that the bacterial richness and diversity varied among tick organs, and ovaries had the lowest level of bacterial richness and diversity. The ovaries and salivary glands had relatively stable bacterial compositions when considering bacterial abundance. Proteobacteria was the dominant bacterial phylum in the four organs, followed by Firmicutes, Bacteroidota and Actinobacteriota. The ovaries carried more Proteobacteria, while the Firmicutes abundance was higher in salivary glands. The environmental bacteria including Staphylococcus, Pseudomonas, Brevundimonas, Acinetobacter, Enterobacter, and Stenotrophomonas had high abundance in the midgut and salivary glands, suggesting that ticks can acquire environmental bacteria through blood meals or host skin. The predominant proportion of Coxiella in ovaries ensures its vertical transmission, and Coxiella in Malpighian tubules can provide nutrients to the host. In conclusion, organ-specific bacterial microbiota was observed in the engorged female H. longicornis, and the functions of these bacteria in organs remain to be investigated.

From the fat body to the hemolymph: Profiling tick immune and storage proteins through transcriptomics and proteomics

Ticks are blood-feeding arachnids that are known to transmit various pathogenic microorganisms to their hosts. During blood feeding, ticks activate their metabolism and immune system to efficiently utilise nutrients from the host's blood and complete the feeding process. In contrast to insects, in which the fat body is known to be a central organ that controls essential metabolic processes and immune defense mechanisms, the function of the fat body in tick physiology is still relatively unexplored. To fill this gap, we sought to uncover the repertoire of genes expressed in the fat body associated with trachea (FB/Tr) by analyzing the transcriptome of individual, partially fed (previtellogenic) Ixodes ricinus females. The resulting catalog of individual mRNA sequences reveals a broad repertoire of transcripts encoding proteins involved in nutrient storage and distribution, as well as components of the tick immune system. To gain a detailed insight into the secretory products of FB/Tr specifically involved in inter-tissue transport and humoral immunity, the transcriptomic data were complemented with the proteome of soluble proteins in the hemolymph of partially fed female ticks. Among these proteins, the hemolipoglyco-carrier proteins were predominant. When comparing immune peptides and proteins from the fat body with those produced by hemocytes, we found that the fat body serves as a unique producer of certain immune components. Finally, time-resolved transcriptional regulation of selected immune transcripts from the FB/Tr was examined in response to experimental challenges with model microbes and analyzed by RT-qPCR. Overall, our data show that the fat body of ticks, similar to insects, is an important metabolic tissue that also plays a remarkable role in immune defense against invading microbes. These findings improve our understanding of tick biology and its impact on the transmission of tick-borne pathogens.

A longitudinal transcriptomic analysis of Rhipicephalus microplus midgut upon feeding

Rhipicephalus microplus, a highly host-specific tick that primarily feeds on cattle, posing a significant threat to livestock production. The investigation of tick physiology is crucial for identifying potential targets in tick control. Of particular interest adult female ticks undergo a significant expansion of the midgut during feeding, leading to an over 100-fold increase in body weight. Beyond the functions of storing and digesting blood meals, the tick midgut plays a crucial role in acquiring and transmitting pathogens. However, our understanding of tick midgut physiology remains limited. In this study we conducted a comprehensive longitudinal transcriptome analysis of the midgut from adult female R. microplus ticks collected at various feeding stages, providing an overview of the transcriptional modulation in this organ as feeding progress. By employing a de novo assembly approach followed by coding-sequences (CDS) extraction, 60,599 potential CDS were identified. In preparation for functional annotation and differential expression analysis, transcripts that showed an average transcript per million (TPM) ≥ 3 in at least one of the biological conditions were extracted. This selection process resulted in a total of 10,994 CDS, which were categorized into 24 functional classes. Notably, our differential expression analysis revealed three main transcriptional profiles. In the first one, representing the slow-feeding stage, the most abundant functional classes were the “protein synthesis” and “secreted” groups, reflecting the highly active state of the tick midgut. The second profile partially accounts for the rapid-feeding stage, in which a high number of differentially expressed transcripts was observed. Lastly, the third transcriptional profile represents post-detached ticks. Notably the highest number of modulated transcripts was observed up to 48 h post-detachment (hpd), however no major differences was observed up to 168 hpd. Overall, the data presented here offers a temporal insight into tick midgut physiology, contributing to the identification of potential targets for the development of anti-tick control strategies.

A microRNA profile of the saliva in the argasid ticks Ornithodoros erraticus and Ornithodoros moubata and prediction of specific target genes

Ornithodoros erraticus and Ornithodoros moubata ticks are the main vectors of the agents of human relapsing fever (TBRF) and African swine fever (ASF) in the Mediterranean Basin and Africa, respectively. Tick ​​saliva is crucial for complete tick feeding and pathogen transmission, as it contains numerous molecules such as proteins, lipids, and non-coding RNAs (ncRNA) including microRNAs (miRNA). MiRNAs are small ncRNAs capable of regulating the expression of their target messenger RNA (mRNA) leading to degradation or inhibition of its translation into protein. Research on miRNAs from ixodid ticks has revealed that miRNAs are involved in the regulation of different physiological processes of ticks, as well as in the modulation of host gene expression, immune response to tick bite and pathogen transmission. Regarding argasid ticks, there is not information about their miRNAs or their potential involvement in tick physiology and/or in the regulation of the tick-host-pathogen interactions. The aim of this work was to profile the miRNAs expressed in the saliva of O. erraticus and O. moubata, and the in silico prediction and functional analysis of their target genes in the swine host. As a whole, up to 72 conserved miRNAs families were identified in both species: 35 of them were shared and 23 and 14 families were unique to O. erraticus and O. moubata, respectively. The most abundant miRNAs families were mir-1, mir-10 and let-7 in O. erraticus and let-7, mir-252, mir-10 in O. moubata. Four miRNAs sequences of each species were validated by RT-qPCR confirming their presence in the saliva. Target gene prediction in the host (Sus scrofa) and functional analysis showed that the selected miRNAs are mainly involved in processes related to signal transduction, regulation of mRNA transcription and gene expression, synapse regulation, immune response, angiogenesis and vascular development. These results suggest that miRNAs could play an important role at the tick-host interface, providing new insights into this complex relationship that may contribute to a more precise selection of tick molecules for the development of therapeutic and immune strategies to control tick infestations and tick-borne pathogens.

Impact of endosymbionts on tick physiology and fitness.

Ticks transmit pathogens and harbour non-pathogenic, vertically transmitted intracellular bacteria termed endosymbionts. Almost all ticks studied to date contain 1 or more of Coxiella, Francisella, Rickettsia or Candidatus Midichloria mitochondrii endosymbionts, indicative of their importance to tick physiology. Genomic and experimental data suggest that endosymbionts promote tick development and reproductive success. Here, we review the limited information currently available on the potential roles endosymbionts play in enhancing tick metabolism and fitness. Future studies that expand on these findings are needed to better understand endosymbionts’ contributions to tick biology. This knowledge could potentially be applied to design novel strategies that target endosymbiont function to control the spread of ticks and pathogens they vector.

A longitudinal transcriptomic analysis from unfed to post-engorgement midguts of adult female Ixodes scapularis

The hematophagy behavior has evolved independently several times within the Arthropoda phylum. Interestingly, the process of acquiring a blood meal in ticks is considerably distinct from that observed in other blood-feeding arthropods. Instead of taking seconds to minutes to complete a blood meal, an adult female Ixodes scapularis tick can remain attached to its host for numerous days. During this extended feeding period, the tick undergoes drastic morphological changes. It is well established that the tick midgut plays a pivotal role not only in blood meal digestion but also in pathogen acquisition and transmission. However, our understanding of the underlying molecular mechanisms involved in these events remains limited. To expedite tick research, we conducted a comprehensive longitudinal RNA-sequencing of the tick midgut before, during, and after feeding. By collecting ticks in different feeding stages (unfed, slow feeding, rapid feeding, and early post-detached), we obtained a comprehensive overview of the transcripts present in each stage and the dynamic transcriptional changes that occur between them. This provides valuable insights into tick physiology. Additionally, through unsupervised clustering, we identified transcripts with similar patterns and stage-specific sequences. These findings serve as a foundation for selecting targets in the development of anti-tick control strategies and facilitate a better understanding of how blood feeding and pathogen infection impact tick physiology.

Universal Tick Vaccines: Candidates and Remaining Challenges.

Recent advancements in molecular biology, particularly regarding massively parallel sequencing technologies, have enabled scientists to gain more insight into the physiology of ticks. While there has been progress in identifying tick proteins and the pathways they are involved in, the specificities of tick-host interaction at the molecular level are not yet fully understood. Indeed, the development of effective commercial tick vaccines has been slower than expected. While omics studies have pointed to some potential vaccine immunogens, selecting suitable antigens for a multi-antigenic vaccine is very complex due to the participation of redundant molecules in biological pathways. The expansion of ticks and their pathogens into new territories and exposure to new hosts makes it necessary to evaluate vaccine efficacy in unusual and non-domestic host species. This situation makes ticks and tick-borne diseases an increasing threat to animal and human health globally, demanding an urgent availability of vaccines against multiple tick species and their pathogens. This review discusses the challenges and advancements in the search for universal tick vaccines, including promising new antigen candidates, and indicates future directions in this crucial research field.

Comparison of salivary gland and midgut microbiome in the soft ticks Ornithodoros erraticus and Ornithodoros moubata.

Ornithodoros erraticus and Ornithodoros moubata are the main vectors of African swine fever virus (ASFV) and the human relapsing fever spirochetes Borrelia hispanica and Borrelia crocidurae in the Mediterranean region and Borrelia duttoni in continental Africa. Manipulation of the tick microbiome has been shown to reduce vector fitness and competence in tick vectors, suggesting that the identification of key microbial players associated with tick tissues can inform interventions such as anti-microbiota vaccines to block pathogen development in the midgut and/or salivary glands. In this study, we analyzed and compared the microbiome of the salivary glands and midgut of O. erraticus and O. moubata. For the taxonomic and functional characterization of the tissue-specific microbiome, we used 16S rRNA amplicon sequencing and prediction of metabolic profiles using PICRUSt2. Co-occurrence networks were built to characterize the community assembly and identify keystone taxa in each tick species. Our results revealed differences in the composition, diversity, and assembly of the bacterial microbiome of salivary glands and midgut within each tick species, but differences were more noticeable in O. moubata. Differences were also found in the microbiome of each tissue, salivary gland and midgut, between species. However, the 'Core Association Networks (CAN)' analysis revealed conserved patterns of interacting taxa in tissues within and between tick species. Different keystone taxa were identified in O. erraticus and O. moubata tissues, but Muribaculaceae and Alistipes were found as keystone taxa in the salivary glands of both tick species which justifies their use as anti-microbiota vaccine candidates to alter the microbiome and reduce tick fitness and/or block pathogen transmission. The high similarity of predicted metabolic pathways profiles between tissues of the two tick species suggests that taxonomic variability of the microbiome is not associated with significant changes in microbial functional profiles. We conclude that the taxonomic structure of the microbiome in O. erraticus and O. moubata is tissue-specific, suggesting niche partitioning of bacterial communities associated to these soft ticks. However, shared keystone taxa and conserved patterns of interacting taxa between tissues and tick species suggest the presence of key microbial players that could be used as anti-microbiota vaccine candidates to affect tick physiology and/or pathogen colonization.

Multi-omics analysis shed light on the mechanisms of tick reproductive inhibition by antibiotics

Ticks are vector pests and their biology and control are of worldwide concern. Microbiota play an important role in tick physiology, and antibiotic treatments are mostly used to explore the interactions between ticks and symbiotic microorganisms. In addition to altering the host microbial community, antibiotics also exhibit toxic effects on the host. In the tick Haemaphysalis longicornis, the engorged female ticks showed reproductive disruption after microinjection of tetracycline, as evidenced by prolonged oviposition time, reduced reproductive efficiency and hatchability, and abnormal oocyte development.

Microbiota Community Structure and Interaction Networks within Dermacentor silvarum, Ixodes persulcatus, and Haemaphysalis concinna.

Ticks carry and transmit a variety of pathogens, which are very harmful to humans and animals. To characterize the microbial interactions in ticks, we analysed the microbiota of the hard ticks, Dermacentor silvarum, Ixodes persulcatus, and Haemaphysalis concinna, using 16S rRNA, showing that microbial interactions are underappreciated in terms of shaping arthropod microbiomes. The results show that the bacterial richness and microbiota structures of these three tick species had significant differences. Interestingly, the bacterial richness (Chao1 index) of all ticks decreased significantly after they became engorged. All the operational taxonomic units (OTUs) were assigned to 26 phyla, 67 classes, 159 orders, 279 families, and 627 genera. Microbial interactions in D. silvarum demonstrated more connections than in I. persulcatus and H. concinna. Bacteria with a high abundance were not important families in microbial interactions. Positive interactions of Bacteroidaceae and F_Solibacteraceae Subgroup 3 with other bacterial families were detected in all nine groups of ticks. This study provides an overview of the microbiota structure and interactions of three tick species and improves our understanding of the role of the microbiota in tick physiology and vector capacity, thus being conducive to providing basic data for the prevention of ticks and tick-borne diseases.

Transcriptome of the synganglion in the tick Ixodes ricinus and evolution of the cys-loop ligand-gated ion channel family in ticks

BackgroundTicks represent a major health issue for humans and domesticated animals. Exploring the expression landscape of the tick’s central nervous system (CNS), known as the synganglion, would be an important step in understanding tick physiology and in managing tick-borne diseases, but studies on that topic are still relatively scarce. Neuron-specific genes like the cys-loop ligand-gated ion channels (cys-loop LGICs, or cysLGICs) are important pharmacological targets of acaricides. To date their sequence have not been well catalogued for ticks, and their phylogeny has not been fully studied.ResultsWe carried out the sequencing of transcriptomes of the I. ricinus synganglion, for adult ticks in different conditions (unfed males, unfed females, and partially-fed females). The de novo assembly of these transcriptomes allowed us to obtain a large collection of cys-loop LGICs sequences. A reference meta-transcriptome based on synganglion and whole body transcriptomes was then produced, showing high completeness and allowing differential expression analyses between synganglion and whole body. Many of the genes upregulated in the synganglion were associated with neurotransmission and/or localized in neurons or the synaptic membrane. As the first step of a functional study of cysLGICs, we cloned the predicted sequence of the resistance to dieldrin (RDL) subunit homolog, and functionally reconstituted the first GABA-gated receptor of Ixodes ricinus. A phylogenetic study was performed for the nicotinic acetylcholine receptors (nAChRs) and other cys-loop LGICs respectively, revealing tick-specific expansions of some types of receptors (especially for Histamine-like subunits and GluCls).ConclusionsWe established a large catalogue of genes preferentially expressed in the tick CNS, including the cysLGICs. We discovered tick-specific gene family expansion of some types of cysLGIC receptors, and a case of intragenic duplication, suggesting a complex pattern of gene expression among different copies or different alternative transcripts of tick neuro-receptors.

Serpins in Tick Physiology and Tick-Host Interaction.

Tick saliva has been extensively studied in the context of tick-host interactions because it is involved in host homeostasis modulation and microbial pathogen transmission to the host. Accumulated knowledge about the tick saliva composition at the molecular level has revealed that serine protease inhibitors play a key role in the tick-host interaction. Serpins are one highly expressed group of protease inhibitors in tick salivary glands, their expression can be induced during tick blood-feeding, and they have many biological functions at the tick-host interface. Indeed, tick serpins have an important role in inhibiting host hemostatic processes and in the modulation of the innate and adaptive immune responses of their vertebrate hosts. Tick serpins have also been studied as potential candidates for therapeutic use and vaccine development. In this review, we critically summarize the current state of knowledge about the biological role of tick serpins in shaping tick-host interactions with emphasis on the mechanisms by which they modulate host immunity. Their potential use in drug and vaccine development is also discussed.

The Symbiotic Continuum Within Ticks: Opportunities for Disease Control.

Among blood-sucking arthropods, ticks are recognized as being of prime global importance because of their role as vectors of pathogens affecting human and animal health. Ticks carry a variety of pathogenic, commensal, and symbiotic microorganisms. For the latter, studies are available concerning the detection of endosymbionts, but their role in the physiology and ecology of ticks remains largely unexplored. This review paper focuses on tick endosymbionts of the genera Coxiella, Rickettsia, Francisella, Midichloria, and Wolbachia, and their impact on ticks and tick-pathogen interactions that drive disease risk. Tick endosymbionts can affect tick physiology by influencing nutritional adaptation, fitness, and immunity. Further, symbionts may influence disease ecology, as they interact with tick-borne pathogens and can facilitate or compete with pathogen development within the vector tissues. Rickettsial symbionts are frequently found in ticks of the genera of Ixodes, Amblyomma, and Dermacentor with relatively lower occurrence in Rhipicephalus, Haemaphysalis, and Hyalomma ticks, while Coxiella-like endosymbionts (CLEs) were reported infecting almost all tick species tested. Francisella-like endosymbionts (FLEs) have been identified in tick genera such as Dermacentor, Amblyomma, Ornithodoros, Ixodes, and Hyalomma, whereas Wolbachia sp. has been detected in Ixodes, Amblyomma, Hyalomma, and Rhipicephalus tick genera. Notably, CLEs and FLEs are obligate endosymbionts essential for tick survival and development through the life cycle. American dog ticks showed greater motility when infected with Rickettsia, indirectly influencing infection risk, providing evidence of a relationship between tick endosymbionts and tick-vectored pathogens. The widespread occurrence of endosymbionts across the tick phylogeny and evidence of their functional roles in ticks and interference with tick-borne pathogens suggests a significant contribution to tick evolution and/or vector competence. We currently understand relatively little on how these endosymbionts influence tick parasitism, vector capacity, pathogen transmission and colonization, and ultimately on how they influence tick-borne disease dynamics. Filling this knowledge gap represents a major challenge for future research.

First molecular and functional characterisation of ferritin 2 proteins from Ornithodoros argasid ticks

Ferritins are iron-binding proteins that play critical functions in iron metabolism. Tick ferritins are essential in blood feeding, reproduction, iron transport, and protection of ticks from the iron-mediated oxidative stress during blood feeding and digestion. In ixodids, ferritin 2 (Fer2) is responsible for iron transport into peripheral tissues, it is critically involved in tick reproduction and has been identified as a good candidate antigen to be included in anti-tick vaccines. In argasids, information on the molecular and functional characteristics of ferritins is almost nonexistent. Given the potential of ixodid Fer2 as a vaccine target, the aim of the current study was to characterise the Fer2 orthologues in Ornithodoros erraticus (OEFer2) and O. moubata (OMFer2), including functional analyses by RNAi gene knockdown and the assessment of the protective efficacy of recombinant Fer2 protein in an animal vaccination trials. Characterisation and analysis of the OMFer2 and OEFer2 amino acid sequences showed high similarity to each other, and high similarity to the Fer2 sequences of ixodid species as well, confirming that Fer2 is highly conserved between both tick families and suggesting a similar function in the physiology of both argasid and ixodid ticks. Fer2 gene knockdown in O. moubata reduced egg hatchability rate and the subsequent number of emerging nymphs-1 up to 71%. Conversely, Fer2 gene knockdown in O. erraticus did not affect the treated ticks even though the Fer2 mRNA expression level was reduced by 90%. The recombinant form of O. moubata Fer2 (tOMFer2) was highly immunogenic and induced strong humoral responses when administered to rabbits formulated with Montanide adjuvant. The protective effect of the anti-tOMFer2 response was limited. While in O. erraticus, we did not observe any protective effect, in O. moubata it induced a significant reduction in oviposition without affecting the other parameters analysed. Accordingly, Fer2 seems to be involved in O. moubata embryogenesis. This study provides the first data on the molecular and functional characterisation of Fer2 in soft tick species and paves the way for further studies aimed at unveiling the functional aspects of Fer2 in soft ticks and confirming its potential as a vaccine candidate antigen.

Neuropeptides in Rhipicephalus microplus and other hard ticks

The synganglion is the central nervous system of ticks and, as such, controls tick physiology. It does so through the production and release of signaling molecules, many of which are neuropeptides. These peptides can function as neurotransmitters, neuromodulators and/or neurohormones, although in most cases their functions remain to be established. We identified and performed in silico characterization of neuropeptides present in different life stages and organs of Rhipicephalus microplus, generating transcriptomes from ovary, salivary glands, fat body, midgut and embryo. Annotation of synganglion transcripts led to the identification of 32 functional categories of proteins, of which the most abundant were: secreted, energetic metabolism and oxidant metabolism/detoxification. Neuropeptide precursors are among the sequences over-represented in R. microplus synganglion, with at least 5-fold higher transcription compared with other stages/organs. A total of 52 neuropeptide precursors were identified: ACP, achatin, allatostatins A, CC and CCC, allatotropin, bursicon A/B, calcitonin A and B, CCAP, CCHamide, CCRFamide, CCH/ITP, corazonin, DH31, DH44, eclosion hormone, EFLamide, EFLGGPamide, elevenin, ETH, FMRFamide myosuppressin-like, glycoprotein A2/B5, gonadulin, IGF, inotocin, insulin-like peptides, iPTH, leucokinin, myoinhibitory peptide, NPF 1 and 2, orcokinin, proctolin, pyrokinin/periviscerokinin, relaxin, RYamide, SIFamide, sNPF, sulfakinin, tachykinin and trissin. Several of these neuropeptides have not been previously reported in ticks, as the presence of ETH that was first clearly identified in Parasitiformes, which include ticks and mites. Prediction of the mature neuropeptides from precursor sequences was performed using available information about these peptides from other species, conserved domains and motifs. Almost all neuropeptides identified are also present in other tick species. Characterizing the role of neuropeptides and their respective receptors in tick physiology can aid the evaluation of their potential as drug targets.