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A divergent Anaplasma phagocytophilum variant in an Ixodes tick from a migratory bird; Mediterranean basin.

ABSTRACT: Anaplasma phagocytophilum (AP) has vast geographical and host ranges and causes disease in humans and domesticated animals. We investigated the role of northward migratory birds in the dispersal of tick-borne AP in the African-Western Palearctic. Ticks were collected from northward migratory birds trapped during spring migration of 2010 at two localities in the central Mediterranean Sea. AP DNA was detected by PCR (gltA and 16S rRNA) and variant determination was performed using ankA sequences. In total, 358 ticks were collected. One of 19 ticks determined as Ixodes was confirmed positive for AP DNA. The tick was collected from a woodchat shrike (Lanius senator senator) trapped in Greece, and molecularly determined to belong to the I. ricinus complex and sharing highest (95%) 16S RNA sequence identity to I. gibbosus. The ankA AP sequence exhibited highest similarity to sequences from rodents and shrews (82%) and ruminants (80%). Phylogenetic analyses placed it convincingly outside other clades, suggesting that it represents a novel AP variant. The divergent Ixodes species harboring a novel AP variant could either indicate an enzootic cycle involving co-evolution with birds, or dissemination from other regions by avian migration. None of the 331 Hyalomma marginatum sensu lato ticks, all immature stages, were positive for AP DNA, lending no evidence for the involvement of Hyalomma ticks transported by birds in the ecology of AP.

SUBMITTER: Hoffman T

PROVIDER: S-EPMC7144310 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Ticks are blood feeding arthropod ectoparasites that transmit pathogens, which cause diseases in humans and animals worldwide. In the past ten decades, the continuous human exploitation of environmental resources and the increase in human outdoor activities has promoted contact with arthropod vectors normally present in the wild, resulting in increased transmission of vector-borne pathogens. In addition, vector populations are expanding in response to climate change and human interventions that impact reservoir host movement and human exposure to infected vectors. Among these emerging vector-borne pathogens, Anaplasma phagocytophilum (Rickettsiales: Anaplasmataceae) has become an important tick-borne pathogen in the United States, Europe and Asia, with increasing numbers of infected people and animals every year. Diseases caused by A. phagocytophilum include human granulocytic anaplasmosis (HGA), equine and canine granulocytic anaplasmosis and tick-borne fever (TBF) in ruminants. The natural infection cycle of A. phagocytophilum is dependent upon the presence of infected vertebrate reservoir hosts and Ixodid tick vectors. In the United States and Europe the main vector species are Ixodes scapularis, Ixodes pacificus, and Ixodes ricinus, while a wide range of mammals, lizards, and birds serve as reservoir hosts for various A. phagocytophilum genotypes. A. phagocytophilum initially infects tick midgut cells and then subsequently develops in salivary glands for transmission to susceptible hosts during tick feeding where the pathogen infects granulocytic cells, primarily neutrophils. Anaplasma phagocytophilum develops within membrane-bound inclusions in the host cell cytoplasm. This pathogen has evolved with its tick and vertebrate hosts through dynamic processes involving genetic traits of the pathogen and hosts that collectively mediate pathogen infection, development, persistence, and survival. However, the mechanisms used by A. phagocytophilum for molecular mechanisms involved in tick-pathogen interactions have not been fully characterized. The objective of this study is to characterize the dynamics of the microRNA response in the tick vector Ixodes scapularis in response to A. phagocytophilum infection. To address this objective, the composition of tick microRNAs was characterize using RNA sequencing in I. scapularis tick cells in response to A. phagocytophilum infection. The discovery of these mechanisms provides evidence that a control strategy could be developed targeted at both vertebrate and tick hosts for more complete control of A. phagocytophilum and its associated diseases.

Project description:The obligate intracellular pathogen, Anaplasma phagocytophilum, is the causative agent of human, equine, and canine granulocytic anaplasmosis and tick-borne fever (TBF) in ruminants. A. phagocytophilum has become an emerging tick-borne pathogen in the United States, Europe, Africa, and Asia, with increasing numbers of infected people and animals every year. It has been recognized that intracellular pathogens manipulate host cell metabolic pathways to increase infection and transmission in both vertebrate and invertebrate hosts. However, our current knowledge on how A. phagocytophilum affect these processes in the tick vector, Ixodes scapularis is limited. In this study, a genome-wide search for components of major carbohydrate metabolic pathways was performed in I. scapularis ticks for which the genome was recently published. The enzymes involved in the seven major carbohydrate metabolic pathways glycolysis, gluconeogenesis, pentose phosphate, tricarboxylic acid cycle (TCA), glyceroneogenesis, and mitochondrial oxidative phosphorylation and ?-oxidation were identified. Then, the available transcriptomics and proteomics data was used to characterize the mRNA and protein levels of I. scapularis major carbohydrate metabolic pathway components in response to A. phagocytophilum infection of tick tissues and cultured cells. The results showed that major carbohydrate metabolic pathways are conserved in ticks. A. phagocytophilum infection inhibits gluconeogenesis and mitochondrial metabolism, but increases the expression of glycolytic genes. A model was proposed to explain how A. phagocytophilum could simultaneously control tick cell glucose metabolism and cytoskeleton organization, which may be achieved in part by up-regulating and stabilizing hypoxia inducible factor 1 alpha in a hypoxia-independent manner. The present work provides a more comprehensive view of the major carbohydrate metabolic pathways involved in the response to A. phagocytophilum infection in ticks, and provides the basis for further studies to develop novel strategies for the control of granulocytic anaplasmosis.

Project description:The causative agent of tick-borne fever and human granulocytic anaplasmosis, Anaplasma phagocytophilum, is transmitted by Ixodes ricinus, and is currently considered an emerging disease throughout Europe. In this study, we established a model of A. phagocytophilum sheep infection and I. ricinus transmission using the European Norway variant 2 ovine strain (NV2Os) propagated in both IDE8 and ISE6 tick cells. Two sheep were inoculated with IDE8 tick cells infected with NV2Os. Both sheep developed A. phagocytophilum infection as determined by qPCR and PCR, the presence of fever 4 days post inoculation (dpi), the observation of morulae in granulocytes at 6 dpi, and the detection of A. phagocytophilum antibodies at 14 dpi. A. phagocytophilum was detected by PCR in skin, lung, small intestine, liver, spleen, uterus, bone marrow, and mesenteric lymph node from necropsies performed at 14 and 15 dpi. One sheep was infested during the acute phase of infection with I. ricinus nymphs from a pathogen-free colony. After molting, A. phagocytophilum transstadial transmission in ticks was validated with qPCR positive bacterial detection in 80% of salivary glands and 90% of midguts from female adults. Infected sheep blood collected at 14 dpi was demonstrated to be able to infect ISE6 tick cells, thus enabling the infection of two additional naive sheep, which then went on to develop similar clinical signs to the sheep infected previously. One of the sheep remained persistently infected until 115 dpi when it was euthanized, and transmitted bacteria to 70 and 2.7% of nymphs engorged as larvae during the acute and persistent infection stages, respectively. We then demonstrated that these infected nymphs were able to transmit the bacteria to one of two other naive infested sheep. As expected, when I. ricinus females were engorged during the acute phase of infection, no A. phagocytophilum transovarial transmission was detected. The development of this new experimental model will facilitate future research on this tick-borne bacterium of increasing importance, and enable the evaluation of any new tick/transmission control strategies.

Project description:Anaplasma phagocytophilum are transmitted by Ixodes spp. ticks and have become one of the most common and relevant tick-borne pathogens due to their impact on human and animal health. Recent results have increased our understanding of the molecular interactions between Ixodes scapularis and A. phagocytophilum through the demonstration of tissue-specific molecular pathways that ensure pathogen infection, development and transmission by ticks. However, little is known about the Ixodes ricinus genes and proteins involved in the response to A. phagocytophilum infection. The tick species I. scapularis and I. ricinus are evolutionarily closely related and therefore similar responses are expected in A. phagocytophilum-infected cells. However, differences may exist between I. scapularis ISE6 and I. ricinus IRE/CTVM20 tick cells associated with tissue-specific signatures of these cell lines. To address this hypothesis, the transcriptional response to A. phagocytophilum infection was characterized by RNA sequencing and compared between I. scapularis ISE6 and I. ricinus IRE/CTVM20 tick cell lines. The transcriptional response to infection of I. scapularis ISE6 cells resembled that of tick hemocytes while the response in I. ricinus IRE/CTVM20 cells was more closely related to that reported previously in infected tick midguts. The inhibition of cell apoptosis by A. phagocytophilum appears to be a key adaptation mechanism to facilitate infection of both vertebrate and tick cells and was used to investigate further the tissue-specific response of tick cell lines to pathogen infection. The results supported a role for the intrinsic pathway in the inhibition of cell apoptosis by A. phagocytophilum infection of I. scapularis ISE6 cells. In contrast, the results in I. ricinus IRE/CTVM20 cells were similar to those obtained in tick midguts and suggested a role for the JAK/STAT pathway in the inhibition of apoptosis in tick cells infected with A. phagocytophilum. Nevertheless, tick cell lines were derived from embryonated eggs and may contain various cell populations with different morphology and behavior that could affect transcriptional response to infection. These results suggested tissue-specific signatures in I. scapularis ISE6 and I. ricinus IRE/CTVM20 tick cell line response to A. phagocytophilum infection that support their use as models for the study of tick-pathogen interactions.

Project description:The obligate intracellular pathogen, Anaplasma phagocytophilum, is the causative agent of life-threatening diseases in humans and animals. A. phagocytophilum is an emerging tick-borne pathogen in the United States, Europe, Africa and Asia, with increasing numbers of infected people and animals every year. It is increasingly recognized that intracellular pathogens modify host cell metabolic pathways to increase infection and transmission in both vertebrate and invertebrate hosts. Recent reports have shown that amino acids are central to the host-pathogen metabolic interaction. In this study, a genome-wide search for components of amino acid metabolic pathways was performed in Ixodes scapularis, the main tick vector of A. phagocytophilum in the United States, for which the genome was recently published. The enzymes involved in the synthesis and degradation pathways of the twenty amino acids were identified. Then, the available transcriptomics and proteomics data was used to characterize the mRNA and protein levels of I. scapularis amino acid metabolic pathway components in response to A. phagocytophilum infection of tick tissues and ISE6 tick cells. Our analysis was focused on the interplay between carbohydrate and amino acid metabolism during A. phagocytophilum infection in ISE6 cells. The results showed that tick cells increase the synthesis of phosphoenolpyruvate (PEP) from tyrosine to control A. phagocytophilum infection. Metabolic pathway analysis suggested that this is achieved by (i) increasing the transcript and protein levels of mitochondrial phosphoenolpyruvate carboxykinase (PEPCK-M), (ii) shunting tyrosine into the tricarboxylic acid (TCA) cycle to increase fumarate and oxaloacetate which will be converted into PEP by PEPCK-M, and (iii) blocking all the pathways that use PEP downstream gluconeogenesis (i.e., de novo serine synthesis pathway (SSP), glyceroneogenesis and gluconeogenesis). While sequestering host PEP may be critical for this bacterium because it cannot actively carry out glycolysis to produce PEP, excess of this metabolite may be toxic for A. phagocytophilum. The present work provides a more comprehensive view of the major amino acid metabolic pathways involved in the response to pathogen infection in ticks, and provides the basis for further studies to develop novel strategies for the control of granulocytic anaplasmosis.

Project description:Anaplasma phagocytophilum is an emerging zoonotic pathogen transmitted by Ixodes scapularis that causes human granulocytic anaplasmosis. Here, a high throughput quantitative proteomics approach was used to characterize A. phagocytophilum proteome during rickettsial multiplication and identify proteins involved in infection of the tick vector, I. scapularis. The first step in this research was focused on tick cells infected with A. phagocytophilum and sampled at two time points containing 10-15% and 65-71% infected cells, respectively to identify key bacterial proteins over-represented in high percentage infected cells. The second step was focused on adult female tick guts and salivary glands infected with A. phagocytophilum to compare in vitro results with those occurring during bacterial infection in vivo. The results showed differences in the proteome of A. phagocytophilum in infected ticks with higher impact on protein synthesis and processing than on bacterial replication in tick salivary glands. These results correlated well with the developmental cycle of A. phagocytophilum, in which cells convert from an intracellular reticulated, replicative form to the nondividing infectious dense-core form. The analysis of A. phagocytophilum differentially represented proteins identified stress response (GroEL, HSP70) and surface (MSP4) proteins that were over-represented in high percentage infected tick cells and salivary glands when compared to low percentage infected cells and guts, respectively. The results demonstrated that MSP4, GroEL and HSP70 interact and bind to tick cells, thus playing a role in rickettsia-tick interactions. The most important finding of these studies is the increase in the level of certain bacterial stress response and surface proteins in A. phagocytophilum-infected tick cells and salivary glands with functional implication in tick-pathogen interactions. These results gave a new dimension to the role of these stress response and surface proteins during A. phagocytophilum infection in ticks. Characterization of Anaplasma proteome contributes information on host-pathogen interactions and provides targets for development of novel control strategies for pathogen infection and transmission.

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A divergent Anaplasma phagocytophilum variant in an Ixodes tick from a migratory bird; Mediterranean basin.

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