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New England Journal of Medicine 01/2013; 368(3):291-3. · 53.30 Impact Factor
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ABSTRACT: Ixodes scapularis transmits the agent of human granulocytic anaplasmosis, among other pathogens. The mechanisms used by the tick to control Anaplasma phagocytophilum are not known. We demonstrate that the I. scapularis Janus kinase (JAK)-signaling transducer activator of transcription (STAT) pathway plays a critical role in A. phagocytophilum infection of ticks. The A. phagocytophilum burden increases in salivary glands and hemolymph when the JAK-STAT pathway is suppressed by RNA interference. The JAK-STAT pathway exerts its anti-Anaplasma activity presumably through STAT-regulated effectors. A salivary gland gene family encoding 5.3-kDa antimicrobial peptides is highly induced upon A. phagocytophilum infection of tick salivary glands. Gene expression and electrophoretic mobility shift assays showed that the 5.3-kDa antimicrobial peptide-encoding genes are regulated by tick STAT. Silencing of these genes increased A. phagocytophilum infection of tick salivary glands and transmission to mammalian host. These data suggest that the JAK-STAT signaling pathway plays a key role in controlling A. phagocytophilum infection in ticks by regulating the expression of antimicrobial peptides.
The Journal of Infectious Diseases 08/2012; 206(8):1233-41. · 6.41 Impact Factor
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ABSTRACT: The maintenance of Borrelia burgdorferi in its complex tick-mammalian enzootic life cycle is dependent on the organism's adaptation to its diverse niches. To this end, the RpoN-RpoS regulatory pathway in B. burgdorferi plays a central role in microbial survival and Lyme disease pathogenesis by up- or down-regulating the expression of a number of virulence-associated outer membrane lipoproteins in response to key environmental stimuli. Whereas a number of studies have reported on the expression of RpoS and its target genes, a more comprehensive understanding of when activation of the RpoN-RpoS pathway occurs, and when induction of the pathway is most relevant to specific stage(s) in the life cycle of B. burgdorferi, has been lacking.
Herein, we examined the expression of rpoS and key lipoprotein genes regulated by RpoS, including ospC, ospA, and dbpA, throughout the entire tick-mammal infectious cycle of B. burgdorferi. Our data revealed that transcription of rpoS, ospC, and dbpA is highly induced in nymphal ticks when taking a blood meal. The RpoN-RpoS pathway remains active during the mammalian infection phase, as indicated by the sustained transcription of rpoS and dbpA in B. burgdorferi within mouse tissues following borrelial dissemination. However, dbpA transcription levels in fed larvae and intermolt larvae suggested that an additional layer of control likely is involved in the expression of the dbpBA operon. Our results also provide further evidence for the downregulation of ospA expression during mammalian infection, and the repression of ospC at later phases of mammalian infection by B. burgdorferi.
Our study demonstrates that the RpoN-RpoS regulatory pathway is initially activated during the tick transmission of B. burgdorferi to its mammalian host, and is sustained during mammalian infection.
BMC Microbiology 03/2012; 12:44. · 3.04 Impact Factor
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ABSTRACT: Ixodes ticks harbour several human pathogens belonging to the order Rickettsiales, including Anaplasma phagocytophilum, the agent of human anaplasmosis. When ticks feed on A. phagocytophilum-infected mice, the pathogen enters the ticks' gut. The bacteria then migrate from the gut to infect the salivary glands of the ticks and are transmitted to the next host via the saliva. The molecular mechanisms that enable the migration of A. phagocytophilum from the gut to the salivary glands are poorly understood. Here we show that a secreted tick protein, P11, is important in this process. We show that P11 enables A. phagocytophilum to infect tick haemocytes, which are required for the migration of A. phagocytophilum from the gut to the salivary glands. Silencing of p11 impaired the A. phagocytophilum infection of tick haemocytes in vivo and consequently decreased pathogen infection of the salivary glands. In vitro experiments showed that P11 could bind to A. phagocytophilum and thus facilitate its infection of tick cells. This report provides new insights into A. phagocytophilum infection of ticks and reveals new avenues to interrupt the life cycle of Anaplasma and related Rickettsial pathogens.
EMBO Reports 09/2011; 12(11):1196-203. · 7.36 Impact Factor
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Tim J Schuijt,
Jeroen Coumou, Sukanya Narasimhan,
Jianfeng Dai,
Kathleen Deponte,
Diana Wouters,
Mieke Brouwer,
Anneke Oei,
Joris J T H Roelofs,
Alje P van Dam,
Tom van der Poll,
Cornelis Van't Veer,
Joppe W Hovius,
Erol Fikrig
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ABSTRACT: The Lyme disease agent Borrelia burgdorferi is primarily transmitted to vertebrates by Ixodes ticks. The classical and alternative complement pathways are important in Borrelia eradication by the vertebrate host. We recently identified a tick salivary protein, designated P8, which reduced complement-mediated killing of Borrelia. We now discover that P8 interferes with the human lectin complement cascade, resulting in impaired neutrophil phagocytosis and chemotaxis and diminished Borrelia lysis. Therefore, P8 was renamed the tick salivary lectin pathway inhibitor (TSLPI). TSLPI-silenced ticks, or ticks exposed to TSLPI-immune mice, were hampered in Borrelia transmission. Moreover, Borrelia acquisition and persistence in tick midguts was impaired in ticks feeding on TSLPI-immunized, B. burgdorferi-infected mice. Together, our findings suggest an essential role for the lectin complement cascade in Borrelia eradication and demonstrate how a vector-borne pathogen co-opts a vector protein to facilitate early mammalian infection and vector colonization.
Cell host & microbe 08/2011; 10(2):136-46. · 13.02 Impact Factor
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ABSTRACT: Borrelia burgdorferi, the causative agent of Lyme disease, is transmitted to humans by bite of Ixodes scapularis ticks. The mechanisms by which the bacterium is transmitted from vector to host are poorly understood. In this study, we show that the F(ab)(2) fragments of BBE31, a B.burgdorferi outer-surface lipoprotein, interfere with the migration of the spirochete from tick gut into the hemolymph during tick feeding. The decreased hemolymph infection results in lower salivary glands infection, and consequently attenuates mouse infection by tick-transmitted B. burgdorferi. Using a yeast surface display approach, a tick gut protein named TRE31 was identified to interact with BBE31. Silencing tre31 also decreased the B. burgdorferi burden in the tick hemolymph. Delineating the specific spirochete and arthropod ligands required for B. burgdorferi movement in the tick may lead to new strategies to interrupt the life cycle of the Lyme disease agent.
PLoS Pathogens 06/2011; 7(6):e1002079. · 9.13 Impact Factor
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ABSTRACT: Anaplasma phagocytophilum causes human granulocytic anaplasmosis, one of the most common tick-borne diseases in North America. This unusual obligate intracellular pathogen selectively persists within polymorphonuclear leucocytes. In this study, using the yeast surrogate model we identified an A. phagocytophilum virulence protein, AptA (A. phagocytophilum toxin A), that activates mammalian Erk1/2 mitogen-activated protein kinase. This activation is important for A. phagocytophilum survival within human neutrophils. AptA interacts with the intermediate filament protein vimentin, which is essential for A. phagocytophilum-induced Erk1/2 activation and infection. A. phagocytophilum infection reorganizes vimentin around the bacterial inclusion, thereby contributing to intracellular survival. These observations reveal a major role for the bacterial protein, AptA, and the host protein, vimentin, in the activation of Erk1/2 during A. phagocytophilum infection.
Cellular Microbiology 01/2011; 13(1):47-61. · 5.46 Impact Factor
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Tim J Schuijt, Sukanya Narasimhan,
Sirlei Daffre,
Kathleen DePonte,
Joppe W R Hovius,
Cornelis Van't Veer,
Tom van der Poll,
Kamran Bakhtiari,
Joost C M Meijers,
Eric T Boder,
Alje P van Dam,
Erol Fikrig
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ABSTRACT: Repeated exposure of rabbits and other animals to ticks results in acquired resistance or immunity to subsequent tick bites and is partially elicited by antibodies directed against tick antigens. In this study we demonstrate the utility of a yeast surface display approach to identify tick salivary antigens that react with tick-immune serum. We constructed an Ixodes scapularis nymphal salivary gland yeast surface display library and screened the library with nymph-immune rabbit sera and identified five salivary antigens. Four of these proteins, designated P8, P19, P23 and P32, had a predicted signal sequence. We generated recombinant (r) P8, P19 and P23 in a Drosophila expression system for functional and immunization studies. rP8 showed anti-complement activity and rP23 demonstrated anti-coagulant activity. Ixodes scapularis feeding was significantly impaired when nymphs were fed on rabbits immunized with a cocktail of rP8, rP19 and rP23, a hall mark of tick-immunity. These studies also suggest that these antigens may serve as potential vaccine candidates to thwart tick feeding.
PLoS ONE 01/2011; 6(1):e15926. · 4.09 Impact Factor
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ABSTRACT: 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.
Cellular Microbiology 03/2010; 12(9):1222-34. · 5.46 Impact Factor
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ABSTRACT: Ticks are distributed worldwide and affect human and animal health by transmitting diverse infectious agents. Effective vaccines against most tick-borne pathogens are not currently available. In this study, we characterized a tick histamine release factor (tHRF) from Ixodes scapularis and addressed the vaccine potential of this antigen in the context of tick engorgement and B. burgdorferi transmission. Results from western blotting and quantitative Reverse Transcription-PCR showed that tHRF is secreted in tick saliva, and upregulated in Borrelia burgdorferi-infected ticks. Further, the expression of tHRF was coincident with the rapid feeding phase of the tick, suggesting a role for tHRF in tick engorgement and concomitantly, for efficient B. burgdorferi transmission. Silencing tHRF by RNA interference (RNAi) significantly impaired tick feeding and decreased B. burgdorferi burden in mice. Interfering with tHRF by actively immunizing mice with recombinant tHRF, or passively transferring tHRF antiserum, also markedly reduced the efficiency of tick feeding and B. burgdorferi burden in mice. Recombinant tHRF was able to bind to host basophils and stimulate histamine release. Therefore, we speculate that tHRF might function in vivo to modulate vascular permeability and increase blood flow to the tick bite-site, facilitating tick engorgement. These findings suggest that blocking tHRF might offer a viable strategy to complement ongoing efforts to develop vaccines to block tick feeding and transmission of tick-borne pathogens.
PLoS Pathogens 01/2010; 6(11):e1001205. · 9.13 Impact Factor
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ABSTRACT: Traditionally, vaccines directly target a pathogen or microbial toxin. Lyme disease, caused by Borrelia burgdorferi, is a tick-borne illness for which a human vaccine is not currently available. B. burgdorferi binds a tick salivary protein, Salp15, during transmission from the vector, and this interaction facilitates infection of mice. We now show that Salp15 antiserum significantly protected mice from B. burgdorferi infection. Salp15 antiserum also markedly enhanced the protective capacity of antibodies against B. burgdorferi antigens, such as OspA or OspC. Mice actively immunized with Salp15 were also significantly protected from tick-borne Borrelia. In vitro assays showed that Salp15 antiserum increased the clearance of Salp15-coated B. burgdorferi by phagocytes, suggesting a mechanism of action. Vaccination with a vector molecule that a microbe requires for infection of the mammalian host suggests a new strategy for the prevention of Lyme disease, and this paradigm may be applicable to numerous arthropod-borne pathogens of medical importance.
Cell host & microbe 11/2009; 6(5):482-92. · 13.02 Impact Factor
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ABSTRACT: Borrelia burgdorferi, the agent of Lyme disease, is recognized by Toll-like receptor (TLR) 1 and 2 heterodimers. Microarray analysis of in vivo B. burgdorferi gene expression in murine skin showed that several genes were altered in TLR1/2-deficient animals compared with wild-type mice. For example, expression of bbe21 (a gene involved in B. burgdorferi lp25 plasmid maintenance) and bb0665 (a gene encoding a glycosyl transferase) were higher in TLR1/2-deficient mice than in control animals. In contrast, messenger RNA levels for bb0731 (a spoJ-like gene) and bba74 (a gene encoding a periplasmic protein) were lower in TLR1/2-deficient mice than in wild-type animals. The expression profiles of some of these genes were altered similarly in B. burgdorferi-infected ticks fed on control or TLR1/2-deficient mice. Quantitative reverse-transcription polymerase chain reaction analysis supported the microarray analysis and suggested that spirochete gene expression is altered by the milieu created by specific host TLRs, both in the murine host and in the arthropod vector.
The Journal of Infectious Diseases 10/2009; 200(8):1331-40. · 6.41 Impact Factor
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ABSTRACT: The saliva of hematophagous arthropods contains potent anti-inflammatory and antihemostatic activities that promote acquisition of the blood meal and enhance infection with pathogens. We have shown that polymorphonuclear leukocytes (PMN) treated with the saliva of the tick Ixodes scapularis have reduced expression of beta(2) integrins, impaired PMN adherence, and reduced killing of Borrelia burgdorferi, the causative agent of Lyme disease. Here we describe two Ixodes proteins that are induced upon tick feeding and expressed predominantly in the salivary glands. Using saliva harvested from ticks with reduced levels of ISL 929 and ISL 1373 through targeted RNA interference knockdown, as well as purified recombinant proteins, we show the effects of these proteins on downregulation of PMN integrins and inhibition of the production of O(2)(-) by PMN in vitro. Mice immunized with ISL 929/1373 had increased numbers of PMN at the site of tick attachment and a lower spirochete burden in the skin and joints 21 days after infection compared to control-immunized animals. Our results suggest that ISL 929 and ISL 1373 contribute to the inhibition of PMN functions shown previously with tick saliva and support important roles for these inhibitory proteins in the modulation of PMN function in vivo.
Infection and immunity 04/2009; 77(6):2320-9. · 4.21 Impact Factor
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Sukanya Narasimhan,
Bindu Sukumaran,
Ulas Bozdogan,
Venetta Thomas,
Xianping Liang,
Kathleen DePonte,
Nancy Marcantonio,
Raymond A Koski,
John F Anderson,
Fred Kantor,
Erol Fikrig
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ABSTRACT: The tick Ixodes scapularis is an efficient vector for microbes, including the Lyme disease agent Borrelia burgdorferi. Ticks engorging on vertebrates induce recruitment of inflammatory cells to the bite site. For efficient transmission to the vector, pathogens have to traffic through this complex feeding site while avoiding the deleterious effects of immune cells. We show that a tick protein, Salp25D, plays a critical role-in the mammalian host-for acquisition of Borrelia burgdorferi by the vector. Silencing salp25D in tick salivary glands impaired spirochete acquisition by ticks engorging on B. burgdorferi-infected mice. Immunizing mice against Salp25D also decreased Borrelia acquisition by I. scapularis. Salp25D detoxified reactive oxygen species at the vector-pathogen-host interface, thereby providing a survival advantage to B. burgdorferi at the tick feeding site in mice. These data demonstrate that pathogens can exploit arthropod molecules to defuse mammalian responses in order to successfully enter the vector.
Cell host & microbe 08/2007; 2(1):7-18. · 13.02 Impact Factor
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Sukanya Narasimhan,
Kathleen Deponte,
Nancy Marcantonio,
Xianping Liang,
Thomas E Royce,
Kenneth F Nelson,
Carmen J Booth,
Benjamin Koski,
John F Anderson,
Fred Kantor,
Erol Fikrig
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ABSTRACT: In North America, the black-legged tick, Ixodes scapularis, an obligate haematophagus arthropod, is a vector of several human pathogens including Borrelia burgdorferi, the Lyme disease agent. In this report, we show that the tick salivary gland transcriptome and proteome is dynamic and changes during the process of engorgement. We demonstrate, using a guinea pig model of I. scapularis feeding and B. burgdorferi transmission, that immunity directed against salivary proteins expressed in the first 24 h of tick attachment - and not later - is sufficient to evoke all the hallmarks of acquired tick-immunity, to thwart tick feeding and also to impair Borrelia transmission. Defining this subset of proteins will promote a mechanistic understanding of novel I. scapularis proteins critical for the initiation of tick feeding and for Borrelia transmission.
PLoS ONE 02/2007; 2(5):e451. · 4.09 Impact Factor
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ABSTRACT: We previously examined the physiological role of the anticoagulant salivary protein 14 (salp14) in adult Ixodes scapularis and showed that Salp14 played a role in tick feeding and engorgement. We now analyze whether the disruption of the salp14 family expression by RNA interference affects tick weight in naïve nymph I. scapularis. Salp14 expression after dsRNA injection was significantly reduced, as shown by mRNA and protein analysis. However, nymph engorgement weight was not altered in salp9pac (salp14 paralog) dsRNA-injected ticks. We also determined Borrelia burgdorferi and Anaplasma phagocytophilum acquisition in I. scapularis nymphs that had reduced Salp14 expression. B. burgdorferi and A. phagocytophilum acquisition was not affected 72 hours after feeding. Our results suggest that different mechanisms govern nymph and adult feeding in I. scapularis.
The American journal of tropical medicine and hygiene 11/2006; 75(4):677-82. · 2.59 Impact Factor
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ABSTRACT: Anaplasma phagocytophilum is the agent of human anaplasmosis, the second most common tick-borne illness in the United States. This pathogen, which is closely related to obligate intracellular organisms in the genera Rickettsia, Ehrlichia, and Anaplasma, persists in ticks and mammalian hosts; however, the mechanisms for survival in the arthropod are not known. We now show that A. phagocytophilum induces expression of the Ixodes scapularis salp16 gene in the arthropod salivary glands during vector engorgement. RNA interference-mediated silencing of salp16 gene expression interfered with the survival of A. phagocytophilum that entered ticks fed on A. phagocytophilum-infected mice. A. phagocytophilum migrated normally from A. phagocytophilum-infected mice to the gut of engorging salp16-deficient ticks, but up to 90% of the bacteria that entered the ticks were not able to successfully infect I. scapularis salivary glands. These data demonstrate the specific requirement of a pathogen for a tick salivary protein to persist within the arthropod and provide a paradigm for understanding how Rickettsia-like pathogens are maintained within vectors.
Journal of Experimental Medicine 07/2006; 203(6):1507-17. · 13.85 Impact Factor
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ABSTRACT: We outline in this review how Borrelia burgdorferi, the causative agent of Lyme disease, moves from the tick to the vertebrate host, and what molecules are potentially involved in this challenging commute. The survival strategies utilized by the spirochete during transmission and the initial stages of infection are discussed.
Microbes and Infection 05/2006; 8(5):1390-9. · 3.10 Impact Factor
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Nandhini Ramamoorthi, Sukanya Narasimhan,
Utpal Pal,
Fukai Bao,
Xiaofeng F Yang,
Durland Fish,
Juan Anguita,
Michael V Norgard,
Fred S Kantor,
John F Anderson,
Raymond A Koski,
Erol Fikrig
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ABSTRACT: The Lyme disease agent, Borrelia burgdorferi, is maintained in a tick-mouse cycle. Here we show that B. burgdorferi usurps a tick salivary protein, Salp15 (ref. 3), to facilitate the infection of mice. The level of salp15 expression was selectively enhanced by the presence of B. burgdorferi in Ixodes scapularis, first indicating that spirochaetes might use Salp15 during transmission. Salp15 was then shown to adhere to the spirochaete, both in vitro and in vivo, and specifically interacted with B. burgdorferi outer surface protein C. The binding of Salp15 protected B. burgdorferi from antibody-mediated killing in vitro and provided spirochaetes with a marked advantage when they were inoculated into naive mice or animals previously infected with B. burgdorferi. Moreover, RNA interference-mediated repression of salp15 in I. scapularis drastically reduced the capacity of tick-borne spirochaetes to infect mice. These results show the capacity of a pathogen to use a secreted arthropod protein to help it colonize the mammalian host.
Nature 08/2005; 436(7050):573-7. · 36.28 Impact Factor
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ABSTRACT: Ixodes scapularis ticks transmit many pathogens, including Borrelia burgdorferi, Anaplasma phagocytophilum, and Babesia microti. Vaccines directed against arthropod proteins injected into the host during tick engorgement could prevent numerous infectious diseases. Salp14, a salivary anticoagulant, poses a key target for such intervention. Salp14 is the prototypic member of a family of potential I. scapularis anticoagulants, expressed and secreted in tick saliva during tick feeding. RNA interference was used to assess the role of Salp14 in tick feeding. Salp14 and its paralogs were silenced, as demonstrated by the reduction of mRNA and protein specific for these antigens. Tick salivary glands lacking Salp14 had reduced anticoagulant activity, as revealed by a 60-80% reduction of anti-factor Xa activity. Silencing the expression of salp14 and its paralogs also reduced the ability of I. scapularis to feed, as demonstrated by a 50-70% decline in the engorgement weights. Because ticks have several anticoagulants, it is likely that the expression of multiple anticoagulants in I. scapularis saliva would have to be ablated simultaneously to abolish tick feeding. These studies demonstrate that RNA interference can silence I. scapularis genes and disrupt their physiologic function in vivo, and they identify vaccine candidates that can alter vector engorgement.
Proceedings of the National Academy of Sciences 03/2004; 101(5):1141-6. · 9.68 Impact Factor
