Search

Background Lyme disease is the most prevalent vector-borne disease in the United States, yet its host factors are poorly understood and diagnostic tests are limited. We evaluated patients in a large health system to uncover the role of cholesterol in the susceptibility, severity, and machine learning-based diagnosis of Lyme disease. Methods A longitudinal health system cohort comprised 1,019,175 individuals with electronic health record data and 50,329 with linked genetic data. Associations of blood cholesterol level, a cholesterol genetic score comprising common genetic variants, and burden of rare loss-of-function (LoF) variants in cholesterol metabolism genes with Lyme disease were investigated. A portable machine learning model was constructed and tested to predict Lyme disease using routine lipid and clinical measurements. Results There were 3,832 cases of Lyme disease. Increasing cholesterol was associated with greater risk of Lyme disease and hypercholesterolemia was more prevalent in Lyme disease cases than controls. Cholesterol genetic scores and rare LoF variants in CD36 and LDLR were associated with elevated Lyme disease risk. Serological profiling of cases revealed parallel trajectories of rising cholesterol and immunoglobulin levels over the disease course, including marked increases in individuals with LoF variants and high cholesterol genetic scores. The machine learning model predicted Lyme disease solely using routine lipid panel, blood count, and metabolic measurements. Conclusions These results demonstrate the value of large-scale genetic and clinical data to reveal host factors underlying infectious disease biology, risk, and prognosis, and the potential for their clinical translation to machine learning diagnostics that do not need specialized assays.

The immune deficiency (IMD) pathway directs host defense in arthropods upon bacterial infection. In Pancrustacea, peptidoglycan recognition proteins sense microbial moieties and initiate nuclear factor-κB-driven immune responses. Proteins that elicit the IMD pathway in non-insect arthropods remain elusive. Here, we show that an Ixodes scapularis homolog of croquemort (Crq), a CD36-like protein, promotes activation of the tick IMD pathway. Crq exhibits plasma membrane localization and binds the lipid agonist 1-palmitoyl-2-oleoyl- sn -glycero-3-phosphoglycerol. Crq regulates the IMD and jun N-terminal kinase signaling cascades and limits the acquisition of the Lyme disease spirochete B. burgdorferi . Additionally, nymphs silenced for crq display impaired feeding and delayed molting to adulthood due to a deficiency in ecdysteroid synthesis. Collectively, we establish a distinct mechanism for arthropod immunity outside of insects and crustaceans.

Ancestral signaling pathways serve critical roles in metazoan development, physiology, and immunity. We report an evolutionary interspecies communication pathway involving a central Ixodes scapularis tick receptor termed Dome1, which acquired a mammalian cytokine receptor motif exhibiting high affinity for interferon-gamma (IFN-γ). Host-derived IFN-γ facilitates Dome1-mediated activation of the Ixodes JAK–STAT pathway. This accelerates tick blood meal acquisition and development while upregulating antimicrobial components. The Dome1–JAK–STAT pathway, which exists in most Ixodid tick genomes, regulates the regeneration and proliferation of gut cells—including stem cells—and dictates metamorphosis through the Hedgehog and Notch–Delta networks, ultimately affecting Ixodes vectorial competence. We highlight the evolutionary dependence of I. scapularis on mammalian hosts through cross-species signaling mechanisms that dually influence arthropod immunity and development.

Ticks exist across diverse environments and transmit numerous pathogens. Due to their long and unique life cycles, these arthropods likely evolved robust epigenetic mechanisms that provide sustainable responses and buffers against extreme environmental conditions. Herein, we highlight how the study of the epigenetic basis of tick biology and vectorial capacity will enrich our knowledge of tick-borne infections.

Metabolism influences biochemical networks, and arthropod vectors are endowed with an immune system that affects microbial acquisition, persistence, and transmission to humans and other animals. Here, we aim to persuade the scientific community to expand their interests in immunometabolism beyond mammalian hosts and towards arthropod vectors. Immunometabolism investigates the interplay of metabolism and immunology. We provide a conceptual framework for investigators from diverse disciplines and indicate that relationships between microbes, mammalian hosts and their hematophagous arthropods may result in cost-effective (mutualism) or energetically expensive (parasitism) interactions. We argue that disparate resource allocations between species may partially explain why some microbes act as pathogens when infecting humans and behave as mutualistic or commensal organisms when colonizing arthropod vectors.

Borrelia burgdorferi is the causative agent of Lyme disease and is transmitted to vertebrate hosts by Ixodes spp. ticks. The spirochaete relies heavily on its arthropod host for basic metabolic functions and has developed complex interactions with ticks to successfully colonize, persist and, at the optimal time, exit the tick. For example, proteins shield spirochaetes from immune factors in the bloodmeal and facilitate the transition between vertebrate and arthropod environments. On infection, B. burgdorferi induces selected tick proteins that modulate the vector gut microbiota towards an environment that favours colonization by the spirochaete. Additionally, the recent sequencing of the Ixodes scapularis genome and characterization of tick immune defence pathways, such as the JAK–STAT, immune deficiency and cross-species interferon-γ pathways, have advanced our understanding of factors that are important for B. burgdorferi persistence in the tick. In this Review, we summarize interactions between B. burgdorferi and I. scapularis during infection, as well as interactions with tick gut and salivary gland proteins important for establishing infection and transmission to the vertebrate host., Borrelia burgdorferi has a complex life cycle with several different hosts, causing Lyme disease when it infects humans. In this Review, Fikrig and colleagues discuss how B. burgdorferi infects and interacts with its tick vector to ensure onward transmission.

Lyme disease, caused by some Borrelia burgdorferi sensu lato, is the most common tick-borne illness in the Northern Hemisphere and the number of cases, and geographic spread, continue to grow. Previously identified B. burgdorferi proteins, lipid immunogens, and live mutants lead the design of canonical vaccines aimed at disrupting infection in the host. Discovery of the mechanism of action of the first vaccine catalyzed the development of new strategies to control Lyme disease that bypassed direct vaccination of the human host. Thus, novel prevention concepts center on proteins produced by B. burgdorferi during tick transit and on tick proteins that mediate feeding and pathogen transmission. A burgeoning area of research is tick immunity as it can unlock mechanistic pathways that could be targeted for disruption. Studies that shed light on the mammalian immune pathways engaged during tick-transmitted B. burgdorferi infection would further development of vaccination strategies against Lyme disease., A vaccine to protect against Lyme disease is needed and feasible. Different types of vaccines can target the bacteria and the tick vector pathogen-transport apparatus for human, animal, and ecological applications.

Extracellular vesicles are thought to facilitate pathogen transmission from arthropods to humans and other animals. Here, we reveal that pathogen spreading from arthropods to the mammalian host is multifaceted. Extracellular vesicles from Ixodes scapularis enable tick feeding and promote infection of the mildly virulent rickettsial agent Anaplasma phagocytophilum through the SNARE proteins Vamp33 and Synaptobrevin 2 and dendritic epidermal T cells. However, extracellular vesicles from the tick Dermacentor andersoni mitigate microbial spreading caused by the lethal pathogen Francisella tularensis. Collectively, we establish that tick extracellular vesicles foster distinct outcomes of bacterial infection and assist in vector feeding by acting on skin immunity. Thus, the biology of arthropods should be taken into consideration when developing strategies to control vector-borne diseases., Extracellular vesicles have been implicated in the transmission of pathogens from the arthropod to the human host. Here the authors show that tick-derived extracellular vesicles play a role in feeding and modulate the outcome of bacterial infection.

Ixodes scapularis and Ixodes pacificus, are the predominant vectors of multiple human pathogens including Borrelia burgdorferi, one of the causative agents of Lyme disease, in North America. Differences in the habitats and host preferences of these closely-related tick species presents an opportunity to examine key aspects of the tick microbiome. While, advances in sequencing technologies, have accelerated a descriptive understanding of the tick microbiome, molecular and mechanistic insights into the tick microbiome is only beginning to emerge. Progress is stymied by technical difficulties to manipulate the microbiome and by biological variables related to the life cycle of Ixodid ticks. This review will highlight these challenges and examine avenues to understand the significance of the tick microbiome in tick biology.

Tick-borne illnesses pose a serious concern to human and veterinary health and their prevalence is on the rise. The interactions between ticks and the pathogens they carry are largely undefined. However, the genus Anaplasma, a group of tick-borne bacteria, has been instrumental in uncovering novel paradigms in tick biology. The emergence of sophisticated technologies and the convergence of entomology with microbiology, immunology, metabolism and systems biology has brought tick–Anaplasma interactions to the forefront of vector biology with broader implications for the infectious disease community. Here, we discuss the use of Anaplasma as an instrument for the elucidation of novel principles in arthropod-microbe interactions. We offer an outlook of the primary areas of study, outstanding questions and future research directions.

Vector-borne illnesses comprise a significant portion of human maladies, representing 17% of global infections. Transmission of vector-borne pathogens to mammals primarily occurs by hematophagous arthropods. It is speculated that blood may provide a unique environment that aids in the replication and pathogenesis of these microbes. Lipids and their derivatives are one component enriched in blood and are essential for microbial survival. For instance, the malarial parasitePlasmodium falciparumand the Lyme disease spirocheteBorrelia burgdorferi, among others, have been shown to scavenge and manipulate host lipids for structural support, metabolism, replication, immune evasion, and disease severity. In thisReview, we will explore the importance of lipid hijacking for the growth and persistence of these microbes in both mammalian hosts and arthropod vectors.

Ticks transmit a diverse array of microbes to vertebrate hosts, including human pathogens, which has led to a human-centric focus in this vector system. Far less is known about pathogens of ticks themselves. Here, we discover that a toxin in blacklegged ticks (Ixodes scapularis) horizontally acquired from bacteria-called domesticated amidase effector 2 (dae2)-has evolved to kill mammalian skin microbes with remarkable efficiency. Secreted into the saliva and gut of ticks, Dae2 limits skin-associated staphylococci in ticks while feeding. In contrast, Dae2 has no intrinsic ability to kill Borrelia burgdorferi, the tick-borne Lyme disease bacterial pathogen. These findings suggest ticks resist their own pathogens while tolerating symbionts. Thus, just as tick symbionts can be pathogenic to humans, mammalian commensals can be harmful to ticks. Our study underscores how virulence is context-dependent and bolsters the idea that "pathogen" is a status and not an identity.

SummaryHard ticks are blood-feeding arthropods that carry and transmit microbes to their vertebrate hosts1. Tick-borne disease cases have been on the rise over the last several decades, drawing much-needed attention to the molecular interplay between transmitted pathogens and their human hosts. However, far less is known about how ticks control their own microbes, which is critical for understanding how zoonotic transmission cycles persist. We previously found that ticks horizontally acquired an antimicrobial toxin gene from bacteria known as domesticated amidase effector 2 (dae2)2. Here we show that this effector from the tick vector Ixodes scapularis (Dae2Is) has structurally and biochemically diverged from ancestral bacterial representatives, expanding its antibacterial targeting range to include host skin microbes. Disruption of dae2Is increases the burden of skin-associated staphylococci within I. scapularis and adversely affects tick fitness, suggesting resistance of host microbes may be important for the parasitic blood-feeding lifestyle. In contrast, Dae2Is has no intrinsic ability to kill Borrelia burgdorferi, the tick-borne bacterium of Lyme disease. Our observations suggest that ticks have evolved to tolerate their own symbionts while resisting host skin commensals, which we discover are natural opportunistic pathogens of ticks. This work moves our understanding of vector biology beyond a human-centric view: just as tick commensals are pathogenic to humans, so too do our commensals pose a threat to ticks. These observations illuminate how a complex and mirrored set of interkingdom interactions between blood-feeding vectors, their hosts, and associated microbes can ultimately lead to disease.

The E3 ubiquitin ligase X-linked inhibitor of apoptosis (XIAP) acts as a molecular rheostat for the immune deficiency (IMD) pathway of the tick Ixodes scapularis. How XIAP activates the IMD pathway in response to microbial infection remains ill defined. Here, we identified the XIAP enzymatic substrate p47 as a positive regulator of the I. scapularis IMD network. XIAP polyubiquitylates p47 in a lysine 63-dependent manner and interacts with the p47 ubiquitin-like (UBX) module. p47 also binds to Kenny (IKKγ/NEMO), the regulatory subunit of the inhibitor of nuclear factor (NF)- κB kinase complex. Replacement of the amino acid lysine to arginine within the p47 linker region completely abrogated molecular interactions with Kenny. Furthermore, mitigation of p47 transcription levels through RNA interference in I. scapularis limited Kenny accumulation, reduced phosphorylation of IKKβ (IRD5), and impaired cleavage of the NF-κB molecule Relish. Accordingly, disruption of p47 expression increased microbial colonization by the Lyme disease spirochete Borrelia burgdorferi and the rickettsial agent Anaplasma phagocytophilum. Collectively, we highlight the importance of ticks for the elucidation of paradigms in arthropod immunology. Manipulating immune signaling cascades within I. scapularis may lead to innovative approaches to reducing the burden of tick-borne diseases.

The insect immune deficiency (IMD) pathway resembles the tumour necrosis factor receptor network in mammals and senses diaminopimelic-type peptidoglycans present in Gram-negative bacteria. Whether unidentified chemical moieties activate the IMD signalling cascade remains unknown. Here, we show that infection-derived lipids 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol (POPG) and 1-palmitoyl-2-oleoyl diacylglycerol (PODAG) stimulate the IMD pathway of ticks. The tick IMD network protects against colonization by three distinct bacteria, that is the Lyme disease spirochete Borrelia burgdorferi and the rickettsial agents Anaplasma phagocytophilum and A. marginale. Cell signalling ensues in the absence of transmembrane peptidoglycan recognition proteins and the adaptor molecules Fas-associated protein with a death domain (FADD) and IMD. Conversely, biochemical interactions occur between x-linked inhibitor of apoptosis protein (XIAP), an E3 ubiquitin ligase, and the E2 conjugating enzyme Bendless. We propose the existence of two functionally distinct IMD networks, one in insects and another in ticks., This work was supported by the National Institutes of Health (R01 AI093653 and R01AI116523 to J.H.F.P.) and the University of Maryland, Baltimore School of Medicine. E.E.M. was a trainee under the Institutional Training Grant T32AI007540 from the National Institute of Allergy and Infectious Diseases.

Vector-borne diseases cause over 700,000 deaths annually and represent 17% of all infectious illnesses worldwide. This public health menace highlights the importance of understanding how arthropod vectors, microbes and their mammalian hosts interact. Currently, an emphasis of the scientific enterprise is at the vector–host interface where human pathogens are acquired and transmitted. At this spatial junction, arthropod effector molecules are secreted, enabling microbial pathogenesis and disease. Extracellular vesicles manipulate signaling networks by carrying proteins, lipids, carbohydrates and regulatory nucleic acids. Therefore, they are well positioned to aid in cell-to-cell communication and mediate molecular interactions. This Review briefly discusses exosome and microvesicle biogenesis, their cargo, and the role that nanovesicles play during pathogen spread, host colonization and disease pathogenesis. We then focus on the role of extracellular vesicles in dictating microbial pathogenesis and host immunity during transmission of vector-borne pathogens.

Tick saliva contains a number of effector molecules that inhibit host immunity and facilitate pathogen transmission. How tick proteins regulate immune signaling, however, is incompletely understood. Here, we describe that loop 2 of sialostatin L2, an anti-inflammatory tick protein, binds to annexin A2 and impairs the formation of the NLRC4 inflammasome during infection with the rickettsial agent Anaplasma phagocytophilum . Macrophages deficient in annexin A2 secreted significantly smaller amounts of interleukin-1β (IL-1β) and IL-18 and had a defect in NLRC4 inflammasome oligomerization and caspase-1 activation. Accordingly, Annexin a2 -deficient mice were more susceptible to A. phagocytophilum infection and showed splenomegaly, thrombocytopenia, and monocytopenia. Providing translational support to our findings, better binding of annexin A2 to sialostatin L2 in sera from 21 out of 23 infected patients than in sera from control individuals was also demonstrated. Overall, we establish a unique mode of inflammasome evasion by a pathogen, centered on a blood-feeding arthropod.

The saliva of ixodid ticks contains a mixture of bioactive molecules that target a wide spectrum of host defense mechanisms to allow ticks to feed on the vertebrate host for several days. Tick salivary proteins cluster in multigenic protein families, and individual family members display redundancy and pluripotency in their action to ameliorate or evade host immune responses. It is now clear that members of different protein families can target the same cellular or molecular pathway of the host physiological response to tick feeding. Here we present and discuss our hypothesis that the redundancy and pluripotency has evolved in tick salivary immunomodulators in order to evade immune recognition by the host, while retaining the immunomodulatory potential of their saliva.

Recent scientific breakthroughs have significantly expanded our understanding of arthropod vector immunity. Insights in the laboratory have demonstrated how the immune system provides resistance to infection, and in what manner innate defenses protect against a microbial assault. Less understood, however, is the effect of biotic and abiotic factors on microbial-vector interactions and the impact of the immune system on arthropod populations in nature. Furthermore, the influence of genetic plasticity on the immune response against vector-borne pathogens remains mostly elusive. Herein, we discuss evolutionary forces that shape arthropod vector immunity. We focus on resistance, pathogenicity and tolerance to infection. We posit that novel scientific paradigms should emerge when molecular immunologists and evolutionary ecologists work together.

Next generation sequencing and proteomics have helped to comprehensively characterize gene expression in tick salivary glands at both the transcriptome and the proteome level. Functional data are, however, lacking. Given that tick salivary secretions are critical to the success of the tick transmission lifecycle and, as a consequence, for host colonization by the pathogens they spread, we thoroughly review here the literature on the known interactions between tick saliva (or tick salivary gland extracts) and the innate and adaptive vertebrate immune system. The information is intended to serve as a reference for functional characterization of the numerous genes and proteins expressed in tick salivary glands with an ultimate goal to develop novel vector and pathogen control strategies.We overview all the known interactions of tick saliva with the vertebrate immune system. The provided information is important, given the recent developments in high-throughput transcriptomic and proteomic analysis of gene expression in tick salivary glands, since it may serve as a guideline for the functional characterization of the numerous newly-discovered genes expressed in tick salivary glands.

It is estimated that approximately one billion people are at risk of infection with obligate intracellular bacteria, but little is known about the underlying mechanisms that govern their life cycles. The difficulty in studying Chlamydia spp., Coxiella spp., Rickettsia spp., Anaplasma spp., Ehrlichia spp. and Orientia spp. is, in part, due to their genetic intractability. Recently, genetic tools have been developed; however, optimizing the genomic manipulation of obligate intracellular bacteria remains challenging. In this Review, we describe the progress in, as well as the constraints that hinder, the systematic development of a genetic toolbox for obligate intracellular bacteria. We highlight how the use of genetically manipulated pathogens has facilitated a better understanding of microbial pathogenesis and immunity, and how the engineering of obligate intracellular bacteria could enable the discovery of novel signalling circuits in host-pathogen interactions.

Ticks transmit a variety of human pathogens, including Borrelia burgdorferi, the etiological agent of Lyme disease. Multiple pathogens that are transmitted simultaneously, termed “coinfections”, are of increasing importance and can affect disease outcome in a host. Arthropod immunity is central to pathogen acquisition and transmission by the tick. Pattern recognition receptors (PRRs) recognize pathogen-associated molecular patterns (PAMPs) and induce humoral responses through the Toll and Immune Deficiency (IMD) pathways. Comparative analyses between insects and ticks reveal that while the Toll pathway is conserved, the IMD network exhibits a high degree of variability. This indicates that major differences in humoral immunity exist between insects and ticks. While many variables can affect immunity, one of the major forces that shape immune outcomes is the microbiota. In light of this, we discuss how the presence of commensal bacteria, symbionts and/or coinfections can lead to altered immune responses in the tick that impact pathogen persistence and subsequent transmission. By investigating non-insect arthropod immunity, we will not only better comprehend tick biology, but also unravel the intricate effects that pathogen coinfections have on vector competence and tick-borne disease transmission.

Host immunity can be divided into two branches: innate and adaptive. The Toll-like and Nod-like receptor pathways are the two best characterized innate signaling cascades. Parasitic organisms have evolved a wide range of methods to invade and persist on and/or within hosts. The main crux of this life strategy hinges on the ability to evade or interrupt the host immune response. In the context of microbial pathogenesis, secreted effector molecules are an immunomodulatory staple. Similarly, ectoparasitic organisms, such as a tick, use salivary molecules to modulate immune processes that favor blood feeding and survival. In this chapter, we will discuss the commonalities between the immunomodulatory properties of tick saliva and microbial-derived effector molecules in the context of Nod-like receptor signaling. Given the extensive body of literature surrounding microbial effectors, we offer the opinion that this can be used to guide discovery and molecular characterization of tick salivary molecules.

Ubiquitination is a posttranslational modification that regulates protein degradation and signaling in eukaryotes. Although it is acknowledged that pathogens exploit ubiquitination to infect mammalian cells, it remains unknown how microbes interact with the ubiquitination machinery in medically relevant arthropods. Here, we show that the ubiquitination machinery is present in the tick Ixodes scapularis and demonstrate that the E3 ubiquitin ligase named x-linked inhibitor of apoptosis protein (XIAP) restricts bacterial colonization of this arthropod vector. We provide evidence that xiap silencing significantly increases tick colonization by the bacterium Anaplasma phagocytophilum, the causative agent of human granulocytic anaplasmosis. We also demonstrate that (i) XIAP polyubiquitination is dependent on the really interesting new gene (RING) catalytic domain, (ii) XIAP polyubiquitination occurs via lysine (K)-63 but not K-48 residues, and (iii) XIAP-dependent K-63 polyubiquitination requires zinc for catalysis. Taken together, our data define a role for ubiquitination during bacterial colonization of disease vectors.

Ubiquitination (ubiquitylation) is a common protein modification that regulates a multitude of processes within the cell. This modification is typically accomplished through the covalent binding of ubiquitin to a lysine residue onto a target protein and is catalysed by the presence of three enzymes: an activating enzyme (E1), ubiquitin-conjugating enzyme (E2) and ubiquitin-protein ligase (E3). In recent years, ubiquitination has risen as a major signalling regulator of immunity and microbial pathogenesis in the mammalian system. Still, little is known about how ubiquitin relates specifically to vector immunology. Here, we provide a brief overview of ubiquitin biochemistry and describe how ubiquitination regulates immune responses in arthropods of medical relevance. We also discuss scientific gaps in the literature and suggest that, similar to mammals, ubiquitin is a major regulator of immunity in medically important arthropods.

Having emerged during the early part of the Cretaceous period, ticks are an ancient group of hematophagous ectoparasites with significant veterinary and public health importance worldwide. The success of their life strategy can be attributed, in part, to saliva. As we enter into a scientific era where the collection of massive data sets and structures for biological application is possible, we suggest that understanding the molecular mechanisms that govern the life cycle of ticks is within grasp. With this in mind, we discuss what is currently known regarding the manipulation of Toll-like (TLR) and Nod-like (NLR) receptor signaling pathways by tick salivary proteins, and how these molecules impact pathogen transmission.

Rickettsial agents are sensed by pattern recognition receptors but lack pathogen-associated molecular patterns commonly observed in facultative intracellular bacteria. Due to these molecular features, the order Rickettsiales can be used to uncover broader principles of bacterial immunity. Here, we used the bacterium Anaplasma phagocytophilum, the agent of human granulocytic anaplasmosis, to reveal a novel microbial surveillance system. Mechanistically, we discovered that upon A. phagocytophilum infection, cytosolic phospholipase A2 cleaves arachidonic acid from phospholipids, which is converted to the eicosanoid prostaglandin E2 (PGE2) via cyclooxygenase 2 (COX2) and the membrane associated prostaglandin E synthase-1 (mPGES-1). PGE2-EP3 receptor signaling leads to activation of the NLRC4 inflammasome and secretion of interleukin (IL)-1β and IL-18. Importantly, the receptor-interacting serine/threonine-protein kinase 2 (RIPK2) was identified as a major regulator of the immune response against A. phagocytophilum. Accordingly, mice lacking COX2 were more susceptible to A. phagocytophilum, had a defect in IL-18 secretion and exhibited splenomegaly and damage to the splenic architecture. Remarkably, Salmonella-induced NLRC4 inflammasome activation was not affected by either chemical inhibition or genetic ablation of genes associated with PGE2 biosynthesis and signaling. This divergence in immune circuitry was due to reduced levels of the PGE2-EP3 receptor during Salmonella infection when compared to A. phagocytophilum. Collectively, we reveal the existence of a functionally distinct NLRC4 inflammasome illustrated by the rickettsial agent A. phagocytophilum., Author Summary Elimination of bacteria is orchestrated by the immune system. Intracellular bacteria are generally recognized by cytosolic molecules named Nod-like receptors (NLRs). One such protein scaffold that senses needle-like structures and globular proteins, namely, the bacterial type III secretion (T3SS) and flagellin, is the NLRC4 inflammasome. The NLRC4 inflammasome induces caspase-1 autoproteolysis and secretion of the pro-inflammatory cytokines interleukin (IL)-1β and IL-18. Here, we show that the obligate intracellular rickettsial pathogen Anaplasma phagocytophilum, which does not have a T3SS or flagellin-coding genes, induces a distinct NLRC4 inflammasome circuitry through the eicosanoid prostaglandin E2 and the EP3 receptor. Conceptually, these findings establish the existence of a distinct microbial surveillance system where the NLRC4 inflammasome senses an obligate intracellular pathogen of public health relevance. Therefore, we propose that rickettsial agents can be used to uncover broader principles of immune surveillance given their unique life style to survive inside the mammalian host and lack of pathogen associated molecular patterns commonly present in most facultative intracellular bacteria.

Ticks transmit more pathogens to humans and animals than any other arthropod. We describe the 2.1 Gbp nuclear genome of the tick, Ixodes scapularis (Say), which vectors pathogens that cause Lyme disease, human granulocytic anaplasmosis, babesiosis and other diseases. The large genome reflects accumulation of repetitive DNA, new lineages of retro-transposons, and gene architecture patterns resembling ancient metazoans rather than pancrustaceans. Annotation of scaffolds representing ∼57% of the genome, reveals 20,486 protein-coding genes and expansions of gene families associated with tick–host interactions. We report insights from genome analyses into parasitic processes unique to ticks, including host ‘questing’, prolonged feeding, cuticle synthesis, blood meal concentration, novel methods of haemoglobin digestion, haem detoxification, vitellogenesis and prolonged off-host survival. We identify proteins associated with the agent of human granulocytic anaplasmosis, an emerging disease, and the encephalitis-causing Langat virus, and a population structure correlated to life-history traits and transmission of the Lyme disease agent., This project has been funded in part with federal funds from the National Institute of Allergy and Infectious Diseases, National Institutes of Health, Department of Health and Human Services (NIAID, NIH, DHHS) under contract numbers N01-AI30071, HHSN272200900007C, HHSN266200400001C and 5R01GM77117-5. Its contents are solely the responsibility of the authors and do not represent the official views of the NIH. Additional grants and contracts supporting work described in this manuscript were from the NIH-NIAID (HHSN266200400039C and HHSN272200900039C) to F.H.C., and a subcontract under HHSN272200900039C to C.A.H. and J.M.M., the Australian Research Council Discovery Project (DP120100240) to S.C.B. and R.S., the Ministerio de Ciencia e Innovación of Spain (BFU2007–6292; BFU2010–15484) to J.R., BIO2009–07990 and BIO2012–37926 to J.V. NIH-1R01AI090062 to Y.P., L.S., and J.K., NIH 1R21AI096268 and NSF IOS-0949194 to R.M.R., the Xunta de Galicia of Spain (10PXIB918057PR) to J.M.C.T. and M.T., BFU2011–23896 and EU FP7 ANTIGONE (278976) to J.F., the USDA-NRI/CREES (2008-35302-18820) and Texas AgriLife Research Vector Biology grant to P.V.P. and European Research Council Starting Independent Researcher Grant (205202) to R.B., J.M.R was supported by the intramural program of the NIAID, R.M.W. by a Marie Curie International Outgoing Fellowship PIOF-GA-2011–303312, E.M.Z. by Swiss National Science Foundation awards 31003A-125350 and 31003A-143936, J.M.G. by an NIH-NCATS award TL1 TR000162 and NSF Graduate Research Fellowship (DGE 1333468), V.C. by a Boehringer Ingelheim Ph.D. Fellowship, F.G.V. by a Fundação para a Ciência e a Tecnologia, Portugal fellowship (SFRH/BD/22360/2005), C.J.P.G. and F.H. by The Lundbeck Foundation (Denmark), and J.J.G. by NIH awards HHSN272200900040C, R01AI017828 and R01AI043006. Support from the Broad Genomics Platform is gratefully acknowledged.

The Gram-negative obligate intracellular bacterium Anaplasma phagocytophilum is the causative agent of human granulocytic anaplasmosis (HGA), an emerging tick-borne infectious disease occurring worldwide. HGA is generally self-limiting; however, the underlying mechanisms, particularly the innate immune pathways that mediate the immune clearance of A. phagocytophilum, are less understood. We herein report an unexpected role for Receptor interacting protein-2 (Rip2), the adaptor protein for the Nod-like receptors (NLRs), Nod1/Nod2, in the host immune response against A. phagocytophilum infection. Although A. phagocytophilum genome is reported to lack the genes encoding the known ligands of Nod1 and Nod2, its infection upregulated the transcription of Rip2 in human primary neutrophils. Our results revealed that Rip2-deficient mice had significantly higher bacterial load than wild-type controls throughout the infection period. In addition, the Rip2-deficient mice took strikingly longer duration to clear A. phagocytophilum infection. Detailed analysis identified that interferon gamma (IFNγ) and interleukin (IL)-18 but not IL-12, macrophage inflammatory protein-2, and KC response were diminished in A. phagocytophilum-challenged Rip2-deficient mice. Together, these results revealed that Rip2 plays important roles in the immune control of A. phagocytophilum and may contribute to our understanding of the host response to Rickettsiales.

Anaplasma phagocytophilum is a tick-borne rickettsial pathogen that provokes an acute inflammatory response during mammalian infection. The illness caused by A. phagocytophilum , human granulocytic anaplasmosis, occurs irrespective of pathogen load and results instead from host-derived immunopathology. Thus, characterizing A. phagocytophilum genes that affect the inflammatory process is critical for understanding disease etiology. By using an A. phagocytophilum Himar1 transposon mutant library, we showed that a single transposon insertion into the A. phagocytophilum dihydrolipoamide dehydrogenase 1 gene ( lpda1 [ APH _ 0065 ]) affects inflammation during infection. A. phagocytophilum lacking lpda1 revealed enlargement of the spleen, increased splenic extramedullary hematopoiesis, and altered clinicopathological abnormalities during mammalian colonization. Furthermore, LPDA1-derived immunopathology was independent of neutrophil infection and correlated with enhanced reactive oxygen species from NADPH oxidase and nuclear factor (NF)-κB signaling in macrophages. Taken together, these findings suggest the presence of different signaling pathways in neutrophils and macrophages during A. phagocytophilum invasion and highlight the importance of LPDA1 as an immunopathological molecule.

Ticks are mostly obligatory blood feeding ectoparasites that have an impact on human and animal health. In addition to direct damage due to feeding, some tick species serve as the vectors for the causative agents of several diseases, such as the spirochetes of the genus Borrelia causing Lyme disease, the virus of tick-borne encephalitis, various Rickettsial pathogens or even protozoan parasites like Babesia spp. Hard ticks are unique among bloodfeeders because of their prolonged feeding period that may last up to two weeks. During such a long period of blood uptake, the host develops a wide range of mechanisms to prevent blood loss. The arthropod ectoparasite, in turn, secretes saliva in the sites of bite that assists blood feeding. Indeed, tick saliva represents a rich source of proteins with potent pharmacologic action that target different mechanisms of coagulation, platelet aggregation and vasoconstriction. Tick adaptation to their vertebrate hosts led to the inclusion of a powerful protein armamentarium in their salivary secretion that has been investigated by high-throughput methods. The resulting knowledge can be exploited for the isolation of novel antihemostatic agents. Here we review the tick salivary antihemostatics and their characterized functions at the molecular and cellular levels.

Fucosylated structures participate in a wide range of pathological processes in eukaryotes and prokaryotes. The impact of fucose on microbial pathogenesis, however, has been less appreciated in arthropods of medical relevance. Thus, we used the tick-borne bacterium Anaplasma phagocytophilum- the agent of human granulocytic anaplasmosis to understand these processes. Here we show that A. phagocytophilum uses alpha1,3-fucose to colonize ticks. We demonstrate that A. phagocytophilum modulates the expression of alpha1,3-fucosyltransferases and gene silencing significantly reduces colonization of tick cells. Acquisition but not transmission of A. phagocytophilum was affected when alpha1,3-fucosyltransferases were silenced during tick feeding. Our results uncover a novel mechanism of pathogen colonization in arthropods. Decoding mechanisms of pathogen invasion in ticks might expedite the development of new strategies to interfere with the life cycle of A. phagocytophilum.

Nod-like receptors have emerged as an important family of sensors in host defense. These receptors are expressed in macrophages, dendritic cells and monocytes and play an important role in microbial immunity. Some Nod-like receptors form the inflammasome, a protein complex that activates caspase-1 in response to several stimuli. Caspase-1 activation leads to processing and secretion of pro-inflammatory cytokines such as interleukin (IL)-1β and IL-18. Here, we discuss recent advances in the inflammasome field with an emphasis on host defense. We also compare differential requirements for inflammasome activation in dendritic cells, macrophages and monocytes.

Tick saliva facilitates tick feeding and infection of the host. Gene expression analysis of tick salivary glands and other tissues involved in host-pathogen interactions has revealed a wide range of bioactive tick proteins. Transcriptomic analysis has been a milestone in the field and has recently been enhanced by next-generation sequencing (NGS). Furthermore, the application of quantitative proteomics to ticks with unknown genomes has provided deeper insights into the molecular mechanisms underlying tick hematophagy, pathogen transmission, and tick-host-pathogen interactions. We review current knowledge on the transcriptomics and proteomics of tick tissues from a systems-biology perspective and discuss future challenges in the field.

Ixodes ricinus is the main tick vector of the microbes that cause Lyme disease and tick-borne encephalitis in Europe. Pathogens transmitted by ticks have to overcome innate immunity barriers present in tick tissues, including midgut, salivary glands epithelia and the hemocoel. Molecularly, invertebrate immunity is initiated when pathogen recognition molecules trigger serum or cellular signalling cascades leading to the production of antimicrobials, pathogen opsonization and phagocytosis. We presently aimed at identifying hemocyte transcripts from semi-engorged female I. ricinus ticks by mass sequencing a hemocyte cDNA library and annotating immune-related transcripts based on their hemocyte abundance as well as their ubiquitous distribution.De novo assembly of 926,596 pyrosequence reads plus 49,328,982 Illumina reads (148 nt length) from a hemocyte library, together with over 189 million Illumina reads from salivary gland and midgut libraries, generated 15,716 extracted coding sequences (CDS); these are displayed in an annotated hyperlinked spreadsheet format. Read mapping allowed the identification and annotation of tissue-enriched transcripts. A total of 327 transcripts were found significantly over expressed in the hemocyte libraries, including those coding for scavenger receptors, antimicrobial peptides, pathogen recognition proteins, proteases and protease inhibitors. Vitellogenin and lipid metabolism transcription enrichment suggests fat body components. We additionally annotated ubiquitously distributed transcripts associated with immune function, including immune-associated signal transduction proteins and transcription factors, including the STAT transcription factor.This is the first systems biology approach to describe the genes expressed in the haemocytes of this neglected disease vector. A total of 2,860 coding sequences were deposited to GenBank, increasing to 27,547 the number so far deposited by our previous transcriptome studies that serves as a discovery platform for studies with I. ricinus biochemistry and physiology.

Background Ixodes ricinus is the main tick vector of the microbes that cause Lyme disease and tick-borne encephalitis in Europe. Pathogens transmitted by ticks have to overcome innate immunity barriers present in tick tissues, including midgut, salivary glands epithelia and the hemocoel. Molecularly, invertebrate immunity is initiated when pathogen recognition molecules trigger serum or cellular signalling cascades leading to the production of antimicrobials, pathogen opsonization and phagocytosis. We presently aimed at identifying hemocyte transcripts from semi-engorged female I. ricinus ticks by mass sequencing a hemocyte cDNA library and annotating immune-related transcripts based on their hemocyte abundance as well as their ubiquitous distribution. Methodology/principal findings De novo assembly of 926,596 pyrosequence reads plus 49,328,982 Illumina reads (148 nt length) from a hemocyte library, together with over 189 million Illumina reads from salivary gland and midgut libraries, generated 15,716 extracted coding sequences (CDS); these are displayed in an annotated hyperlinked spreadsheet format. Read mapping allowed the identification and annotation of tissue-enriched transcripts. A total of 327 transcripts were found significantly over expressed in the hemocyte libraries, including those coding for scavenger receptors, antimicrobial peptides, pathogen recognition proteins, proteases and protease inhibitors. Vitellogenin and lipid metabolism transcription enrichment suggests fat body components. We additionally annotated ubiquitously distributed transcripts associated with immune function, including immune-associated signal transduction proteins and transcription factors, including the STAT transcription factor. Conclusions/significance This is the first systems biology approach to describe the genes expressed in the haemocytes of this neglected disease vector. A total of 2,860 coding sequences were deposited to GenBank, increasing to 27,547 the number so far deposited by our previous transcriptome studies that serves as a discovery platform for studies with I. ricinus biochemistry and physiology., Author Summary Ixodes ricinus transmits Lyme disease and tick-borne encephalitis in Europe. Tick-borne pathogens ingested with a blood meal persist in the tick gut and, during the next feeding stage, disseminate through the haemolymph to pass into a vertebrate host via the salivary glands. Invertebrate immunity is initiated when tick pathogen recognition molecules trigger serum or cellular signalling cascades. We aimed to identify haemocyte transcripts from semi-engorged female I. ricinus ticks by deep sequencing a haemocyte cDNA library and by annotating immune-related transcripts based on their abundance in haemocytes and their sequence divergence when compared to their arthropod homo(ortho)logues. 15,716 extracted coding sequences (CDS) were generated in total, and 327 transcripts were significantly overexpressed in haemocytes. These included those coding for scavenger receptors, antimicrobial peptides, pathogen recognition proteins, proteases, and protease inhibitors. Transcriptional enrichment of vitellogenin and lipid metabolism effectors suggests that body fat components mediate tick innate immunity. Annotation of ubiquitously distributed transcripts associated with immune function and their phylogenetic analysis provide insights into the evolution of major protein families involved in tick innate immunity.

Chapter 110, Second Edition.-- et al., Anaplasma phagocytophilum is a rickettsial pathogen transmitted by ixodid ticks. A. phagocytophilum colonises myeloid and non-myeloid cells and causes human granulocytic anaplasmosis – an important disease in the United States, Europe and Asia. Research has illustrated how A. phagocytophilum successfully invades and proliferates inside host cells, causing a systemic disease. Here, we address A. phagocytophilum biology and the underlying mechanisms involved in bacterial pathogenesis and disease development. Clinical features, diagnostics and treatment are also discussed, as well as the contribution of the tick vector for microbial transmission.