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1.
Clin Microbiol Infect ; 26(4): 513.e1-513.e6, 2020 Apr.
Article in English | MEDLINE | ID: mdl-31404672

ABSTRACT

OBJECTIVES: Borrelia miyamotoi is a relapsing fever Borrelia, transmitted by hard (Ixodes) ticks, which are also the main vector for Borrelia burgdorferi. A widely used test for serodiagnosis of Lyme borreliosis is an enzyme immunoassay (EIA) based on the C6 peptide of the B. burgdorferi sl VlsE protein. We set out to study C6 reactivity upon infection with B. miyamotoi in a large well-characterized set of B. miyamotoi disease (BMD) patient sera and in experimental murine infection. METHODS: We performed in silico analyses, comparing the C6-peptide to immunodominant B. miyamotoi variable large proteins (Vlps). Next, we determined C6 reactivity in sera from mice infected with B. miyamotoi and in a unique longitudinal set of 191 sera from 46 BMD patients. RESULTS: In silico analyses revealed similarity of the C6 peptide to domains within B. miyamotoi Vlps. Cross-reactivity against the C6 peptide was confirmed in 21 out of 24 mice experimentally infected with B. miyamotoi. Moreover, 35 out of 46 BMD patients had a C6 EIA Lyme index higher than 1.1 (positive). Interestingly, 27 out of 37 patients with a C6 EIA Lyme index higher than 0.9 (equivocal) were negative when tested for specific B. burgdorferi sl antibodies using a commercially available immunoblot. CONCLUSIONS: We show that infection with B. miyamotoi leads to cross-reactive antibodies to the C6 peptide. Since BMD and Lyme borreliosis are found in the same geographical locations, caution should be used when relying solely on C6 reactivity testing. We propose that a positive C6 EIA with negative immunoblot, especially in patients with fever several weeks after a tick bite, warrants further testing for B. miyamotoi.


Subject(s)
Antibodies, Bacterial/blood , Bacterial Proteins/immunology , Borrelia/immunology , Cross Reactions , Lyme Disease/immunology , Relapsing Fever/immunology , Animals , Computer Simulation , Female , Humans , Immunoblotting , Ixodes/microbiology , Longitudinal Studies , Lyme Disease/diagnosis , Mice , Mice, Inbred C3H , Peptides/immunology , Reagent Kits, Diagnostic , Relapsing Fever/diagnosis , Serologic Tests
2.
Ticks Tick Borne Dis ; 4(1-2): 160-3, 2013 Feb.
Article in English | MEDLINE | ID: mdl-23141105

ABSTRACT

Prior exposure of vertebrate hosts to tick salivary proteins can induce specific immunity to tick infestation, as well as affording protection against tick-transmitted Borrelia burgdorferi infection in the mammalian host. Vaccination using an adenovirus expression system to deliver 4 tick salivary proteins (Ad-Salps) derived from Ixodes scapularis, Salp15, Salp25A, Salp25D, and Isac, was explored. Results indicate that vaccination with tick salivary proteins in an adenoviral vector can be used to modulate a Th1 response in the host and partially control spirochete load in immunized mice after infected tick challenge.


Subject(s)
Adenoviridae/genetics , Ixodes/metabolism , Lyme Disease/transmission , Salivary Proteins and Peptides/immunology , Tick Infestations/prevention & control , Vaccines/immunology , Animals , Arthropod Proteins/genetics , Arthropod Proteins/immunology , Heart/microbiology , Ixodes/microbiology , Mice , Salivary Proteins and Peptides/metabolism , Urinary Bladder/microbiology
3.
Ticks Tick Borne Dis ; 2(1): 20-6, 2011 Mar.
Article in English | MEDLINE | ID: mdl-21771533

ABSTRACT

B. burgdorferi, B. afzelii, and B. bavariensis show resistance to mouse and human complement. B. garinii and B. valaisiana are sensitive to mouse and human complement. We evaluated whether the absence of C3 in mice influenced infectivity and pathogenicity of different Borrelia species. C3 knockout mice (C3-/-) and syngeneic C57Bl/6 wild-type (WT) mice were challenged with 5 different Borrelia species. After 2 weeks, quantitative PCR (qPCR), culture, histopathology, and immunofluorescence were performed on heart, joint, brain, bladder, and skin. Spirochaetes were detected by qPCR after infection with B. burgdorferi, B. afzelii, or B. bavariensis strains. In joints of C3-/-, but not WT mice challenged with B. burgdorferi, spirochaetes were detected by qPCR. No other significant differences between C3-/- and WT mice were seen. Histopathology demonstrated concordance between borrelia load and inflammation score. Only after B. burgdorferi and B. afzelii infection, spirochaetes were detected by immunofluorescence microscopy. B. burgdorferi was cultured from heart, joint, bladder, and skin from all mice within 2 weeks. B. afzelii and B. bavariensis grew only from heart tissue from both C3-/- and WT mice after 2-6 weeks. The infectivity and pathogenicity of complement-resistant Borrelia strains is unchanged in complement-deficient mice. Complement-susceptible strains do not become infectious in the absence of C3.


Subject(s)
Borrelia burgdorferi Group/pathogenicity , Complement C3/deficiency , Lyme Disease/immunology , Animals , Ankle Joint/pathology , Arthritis/etiology , Arthritis/pathology , Culture Techniques , Disease Susceptibility , Lyme Disease/complications , Lyme Disease/microbiology , Lyme Disease/pathology , Mice , Mice, Inbred C57BL , Mice, Knockout , Real-Time Polymerase Chain Reaction
4.
Trends Parasitol ; 27(1): 40-7, 2011 Jan.
Article in English | MEDLINE | ID: mdl-20594913

ABSTRACT

Lyme disease, or Lyme borreliosis, the most prevalent arthropod-borne disease in the Western world, is caused by spirochetes belonging to the Borrelia burgdorferi sensu lato group and is predominantly transmitted through Ixodes ticks. There is currently no vaccine available to prevent Lyme borreliosis in humans. Borrelia outer membrane proteins are reviewed which have been investigated as vaccine candidates. In addition, several tick proteins are discussed, on which anti-tick vaccines have been based, or are interesting future candidates, to prevent transmission of the spirochete from the tick vector to the mammalian host. Finally, novel vaccination strategies to prevent Lyme borreliosis are proposed, based on multiple Borrelia antigens, tick antigens or a combination of both Borrelia as well as tick antigens.


Subject(s)
Bacterial Vaccines/immunology , Borrelia burgdorferi/immunology , Lyme Disease/prevention & control , Animals , Arachnid Vectors/immunology , Arachnid Vectors/microbiology , Humans , Ixodes/immunology , Ixodes/microbiology , Lyme Disease/immunology , Lyme Disease/transmission
5.
J Infect Dis ; 198(8): 1189-97, 2008 Oct 15.
Article in English | MEDLINE | ID: mdl-18752445

ABSTRACT

BACKGROUND: Ixodes ticks are the main vectors for Borrelia burgdorferi sensu lato. In the United States, B. burgdorferi is the sole causative agent of Lyme borreliosis and is transmitted by Ixodes scapularis. In Europe, 3 Borrelia species-B. burgdorferi, B. garinii, and B. afzelii-are prevalent, which are transmitted by Ixodes ricinus. The I. scapularis salivary protein Salp15 has been shown to bind to B. burgdorferi outer surface protein (Osp) C, protecting the spirochete from antibody-mediated killing. METHODS AND RESULTS: We recently identified a Salp15 homologue in I. ricinus, Salp15 Iric-1. Here, we have demonstrated, by solid-phase overlays, enzyme-linked immunosorbent assay, and surface plasmon resonance, that Salp15 Iric-1 binds to B. burgdorferi OspC. Importantly, this binding protected the spirochete from antibody-mediated killing in vitro and in vivo; immune mice rechallenged with B. burgdorferi preincubated with Salp15 Iric-1 displayed significantly higher Borrelia numbers and more severe carditis, compared with control mice. Furthermore, Salp15 Iric-1 was capable of binding to OspC from B. garinii and B. afzelii, but these Borrelia species were not protected from antibody-mediated killing. CONCLUSIONS: Salp15 Iric-1 interacts with all European Borrelia species but differentially protects B. burgdorferi from antibody-mediated killing, putatively giving this Borrelia species a survival advantage in nature.


Subject(s)
Antigens, Bacterial/metabolism , Bacterial Outer Membrane Proteins/metabolism , Borrelia burgdorferi/drug effects , Borrelia burgdorferi/metabolism , Ixodes/microbiology , Saliva/metabolism , Salivary Proteins and Peptides/pharmacology , Animals , Antibodies, Bacterial/blood , Antibodies, Bacterial/immunology , Borrelia burgdorferi/immunology , Borrelia burgdorferi/pathogenicity , Enzyme-Linked Immunosorbent Assay , Female , Mice , Mice, Inbred C3H , Rabbits , Salivary Proteins and Peptides/metabolism , Surface Plasmon Resonance
6.
Vector Borne Zoonotic Dis ; 7(3): 296-303, 2007.
Article in English | MEDLINE | ID: mdl-17896872

ABSTRACT

The 15-kDa Ixodes scapularis salivary gland protein Salp15 protects Borrelia burgdorferi sensu stricto from antibody-mediated killing and facilitates infection of the mammalian host. In addition, Salp 15 has been shown to inhibit T-cell activation. We determined whether Ixodes ricinus, the major vector for Lyme borreliosis in Western Europe, also express salp15-related genes. We show that engorged I. ricinus express salp15 and we have identified three Salp15 homologues within these ticks by reverse transcriptase-polymerase chain reaction (RT-PCR). One of the predicted proteins showed 80% similarity to I. scapularis Salp15, evenly distributed over the entire amino acid sequence, whereas the two other predicted proteins showed approximately 60% similarity, mainly confined to the signal sequence and C-terminus. Comparison of the DNA and protein sequences with those deposited in several databases indicates that these proteins are part of a Salp15 family of which members are conserved among different Ixodes species, all capable of transmitting B. burgdorferi sensu lato. This suggests that these Salp15 homologues could also play a role in the transmission of diverse Borrelia species and in inhibition of T-cell activation.


Subject(s)
Ixodes/genetics , Salivary Proteins and Peptides/genetics , Amino Acid Sequence , Animals , Base Sequence , Female , Molecular Sequence Data , Phylogeny , Salivary Glands/metabolism , Salivary Proteins and Peptides/chemistry , Sequence Alignment , Sequence Homology, Nucleic Acid
7.
Insect Mol Biol ; 16(4): 469-79, 2007 Aug.
Article in English | MEDLINE | ID: mdl-17651236

ABSTRACT

Ixodes ticks are vectors of several pathogens including Borrelia burgdorferi. Tick saliva contains numerous molecules that facilitate blood feeding without host immune recognition and rejection. We have expressed, purified, and characterized Ixodes scapularis salivary protein 20 (Salp20), a potential inhibitor of the alternative complement pathway that shares homology with the Isac protein family. When analysed by SDS-PAGE and size exclusion chromatography, Salp20 was approximately 48 kDa, more than double its predicted mass, primarily due N- and O-linked glycosylations. Recombinant Salp20 inhibited the alternative complement pathway by dissociating the C3 convertase, and partially protected a serum sensitive species of Borrelia from lysis by normal human serum. We propose that Salp20 facilitates tick feeding and possibly protects tick-borne pathogens from complement components.


Subject(s)
Complement Activation/drug effects , Ixodes/genetics , Ixodes/metabolism , Salivary Proteins and Peptides/genetics , Salivary Proteins and Peptides/pharmacology , Amino Acid Sequence , Animals , Borrelia , Molecular Sequence Data , Salivary Proteins and Peptides/chemistry , Salivary Proteins and Peptides/metabolism
8.
Res Vet Sci ; 79(2): 99-103, 2005 Oct.
Article in English | MEDLINE | ID: mdl-15924926

ABSTRACT

Blood samples were collected from 98 horses suspected of having borreliosis or granulocytic ehrlichiosis in Connecticut and New York State, USA during 1985, 1995, and 1996. Serum antibodies to Borrelia burgdorferi were detected by an enzyme-linked immunosorbent assay (ELISA), based on whole-cell and recombinant antigens, in 82 (84%) horses. Of the 181 sera tested, 59% were positive, using whole-cell antigens, compared to 48% with protein (p)37 and 35% with VlsE antigens. An ELISA containing either of these fusion proteins can be used as an adjunct to general screening by an ELISA or immunoblotting in animals not vaccinated for this disease.


Subject(s)
Antibodies, Bacterial/blood , Antigens, Bacterial/immunology , Borrelia burgdorferi/immunology , Horse Diseases/diagnosis , Lyme Disease/veterinary , Animals , Enzyme-Linked Immunosorbent Assay/veterinary , Horse Diseases/epidemiology , Horses , Recombinant Proteins/immunology , Reproducibility of Results , United States/epidemiology
9.
Insect Mol Biol ; 11(6): 641-50, 2002 Dec.
Article in English | MEDLINE | ID: mdl-12421422

ABSTRACT

Using biochemical and molecular approaches, we have identified a 9.8 kDa protein in the saliva of Ixodes scapularis that inhibits the intrinsic pathway of coagulation. The 9.8 kDa anticoagulant protein was purified by reverse-phase HPLC and its N-terminal amino acid sequence determined. The N-terminal sequence showed homology with Salp14, an immuno-dominant antigen present in the saliva of engorging I. scapularis nymphs. Recombinant Salp14 expressed in Escherichia coli prolonged the activated partial thromboplastin time (APTT) of human plasma in a dose-dependent manner and was a specific inhibitor of factor Xa. A cDNA encoding a 9.3 kDa protein, Salp9Pac, was subsequently isolated from an I. scapularis salivary gland cDNA library. Salp9Pac showed 93% identity to the N-terminal sequence of the anticoagulant purified by HPLC. These data indicate that the anticoagulant protein purified by HPLC, Salp9Pac and Salp14 are members of a family of novel coagulation protease inhibitors present in tick saliva. While recombinant Salp9Pac did not show biological activity in the assays tested currently, it is likely to be mechanistically different from its paralogues. This raises the possibility that ticks may enhance their adaptive ability to cope with a wide spectrum of proteases, by transcribing such structurally related anticoagulant proteins with different functions.


Subject(s)
Anticoagulants/isolation & purification , Insect Proteins/metabolism , Ixodes/physiology , Saliva/metabolism , Amino Acid Sequence , Animals , Anticoagulants/chemistry , Cloning, Molecular , DNA Primers , Escherichia coli/genetics , Gene Library , Insect Proteins/chemistry , Insect Proteins/genetics , Molecular Sequence Data , Recombinant Proteins/biosynthesis , Recombinant Proteins/chemistry , Sequence Alignment , Sequence Homology, Amino Acid
10.
J Immunol ; 167(9): 5273-7, 2001 Nov 01.
Article in English | MEDLINE | ID: mdl-11673542

ABSTRACT

West Nile (WN) virus is a mosquito-borne flavivirus that emerged in the United States in 1999 and can cause fatal encephalitis. Envelope (E) protein cDNA from a WN virus isolate recovered from Culex pipiens in Connecticut was expressed in Escherichia coli. The recombinant E protein was purified and used as Ag in immunoblot assays and immunization experiments. Patients with WN virus infection had Abs that recognized the recombinant E protein. C3H/HeN mice immunized with E protein developed E protein Abs and were protected from infection with WN virus. Passive administration of E protein antisera was also sufficient to afford immunity. E protein is a candidate vaccine to prevent WN virus infection.


Subject(s)
Vaccines, Synthetic/immunology , Viral Envelope Proteins/immunology , Viral Vaccines/immunology , West Nile virus/immunology , Animals , Antibodies, Viral/immunology , Immunization , Mice , Mice, Inbred C3H , Recombinant Proteins/immunology
11.
J Infect Dis ; 184(8): 1056-64, 2001 Oct 15.
Article in English | MEDLINE | ID: mdl-11574922

ABSTRACT

Rabbits or guinea pigs infested with Ixodes scapularis acquire resistance to tick bites, a phenomenon, known as tick immunity, that is partially mediated by antibody. To determine the salivary gland antigens that elicit antibodies in the host, an I. scapularis salivary gland cDNA expression library was probed with serum from tick-immune rabbits. Sera from sensitized rabbits strongly recognized 47 of 100,000 library clones in an antibody-screening assay. These 47 clones encoded 14 different I. scapularis genes, including a glutathione peroxidase homologue. Expression of these 14 genes in engorged tick salivary glands was confirmed by reverse-transcription polymerase chain reaction. The I. scapularis glutathione peroxidase homologue, named salp25D, was expressed in both unfed and fed nymphal salivary glands. Recombinant Salp25D was able to catalyze the reduction of hydrogen peroxide in the presence of reduced glutathione and glutathione reductase. These results categorize the prominent salivary gland proteins in I. scapularis and demonstrate the presence of a potent antioxidant in tick saliva.


Subject(s)
Antigens/analysis , Antioxidants/analysis , Guinea Pigs/immunology , Insect Proteins/immunology , Ixodes/immunology , Salivary Glands/immunology , Amino Acid Sequence , Animals , Cattle/immunology , Cloning, Molecular , Consensus Sequence , Enzyme-Linked Immunosorbent Assay , Female , Gene Library , Glutathione Peroxidase/chemistry , Glutathione Peroxidase/genetics , Humans , Immunoglobulin G/blood , Insect Proteins/genetics , Ixodes/classification , Ixodes/genetics , Mice , Mice, Inbred C3H , Molecular Sequence Data , Nematoda/immunology , RNA, Messenger/genetics , Rabbits/immunology , Salivary Glands/chemistry , Sequence Homology, Amino Acid , Species Specificity , Time Factors
12.
J Immunol ; 167(6): 3383-90, 2001 Sep 15.
Article in English | MEDLINE | ID: mdl-11544329

ABSTRACT

Spirochete adaptation in vivo is associated with preferential Borrelia burgdorferi gene expression. In this paper, we show that the administration of B. burgdorferi-immune sera to IFN-gammaR-deficient mice that have been infected with B. burgdorferi N40 for 4 days causes spirochete clearance. In contrast, immune sera-mediated clearance of B. burgdorferi N40 is not apparent in immunocompetent mice, suggesting a role for IFN-gamma-mediated responses in B. burgdorferi N40 host adaptation. B. burgdorferi-immune sera also induces clearance of B. burgdorferi N40 that have been passaged in vitro 75 times (B. burgdorferi N40-75), a derivative of B. burgdorferi N40 that does not rapidly adapt in vivo in immunocompetent mice. B. burgdorferi N40-75 produce lower levels of IFN-gamma and IL-12 in mice than does B. burgdorferi N40, and the administration of these cytokines to B. burgdorferi N40-75-infected mice results in an increased spirochetal burden, further indicating that IFN-gamma-mediated events promote B. burgdorferi survival. Differential immunoscreening and RT-PCR demonstrate that IFN-gamma-mediated signals facilitate spirochete recombination at the variable major protein like sequence locus, a site for early antigenic variation in vivo, and that recombination rates by B. burgdorferi N40 are lower in IFN-gammaR-deficient mice than in control animals. These results suggest that the murine immune response can promote the in vivo adaptation of B. burgdorferi.


Subject(s)
Antigens, Bacterial/genetics , Antigens, Surface/genetics , Bacterial Proteins , Borrelia burgdorferi/physiology , DNA, Bacterial/genetics , Lipoproteins/genetics , Lyme Disease/microbiology , Recombination, Genetic , Adaptation, Physiological/genetics , Animals , Antibodies, Bacterial/immunology , Antigens, Bacterial/immunology , Antigens, Surface/immunology , Base Sequence , Borrelia burgdorferi/genetics , Borrelia burgdorferi/isolation & purification , CD4-Positive T-Lymphocytes/immunology , Gene Expression Regulation , Immune Sera , Immunocompetence , Inflammation , Interferon-gamma/biosynthesis , Interferon-gamma/genetics , Interferon-gamma/pharmacology , Interleukin-12/biosynthesis , Interleukin-12/genetics , Interleukin-12/pharmacology , Lipoproteins/immunology , Lyme Disease/pathology , Mice , Mice, Inbred C3H , Mice, Knockout , Molecular Sequence Data , Receptors, Interferon/deficiency , Receptors, Interferon/genetics , Sequence Alignment , Sequence Homology, Nucleic Acid , Interferon gamma Receptor
13.
Eur J Clin Microbiol Infect Dis ; 20(7): 482-5, 2001 Jul.
Article in English | MEDLINE | ID: mdl-11561804

ABSTRACT

Recombinant protein 44, expressed and purified as a maltose-binding protein fusion peptide of the human granulocytic ehrlichiosis (HGE) agent (Ehrlichia phagocytophila genogroup), was used as antigen in enzyme-linked immunosorbent assays (ELISAs) to detect total antibodies, immunoglobulin (Ig) M antibodies, and IgG antibodies. Of the 67 human sera obtained from 64 HGE patients 3-5 weeks after the onset of illness and confirmed as having total immunoglobulins to whole-cell antigen by indirect fluorescent antibody analyses, 63 were positive in a polyvalent ELISA. Fifty-six and 61 sera had IgM or IgG antibodies, respectively. Fifty sera had both IgM and IgG antibodies. In specificity tests of 110 sera, one serum sample from a patient who had Lyme borreliosis reacted to the protein 44 antigen in the analysis for IgM antibody (specificity, 99%). There were no false-positive results in an ELISA for IgG antibodies. With their high sensitivity and specificity, class-specific ELISAs can be used in conjunction with indirect fluorescent antibody analyses or immunoblotting methods to help diagnose human granulocytic ehrlichiosis.


Subject(s)
Antibodies, Bacterial/blood , Ehrlichia/isolation & purification , Ehrlichiosis/diagnosis , Enzyme-Linked Immunosorbent Assay/methods , Bacterial Proteins/blood , Bacterial Proteins/immunology , Blotting, Western , Ehrlichiosis/blood , Female , Humans , Immunoglobulin G/blood , Immunoglobulin G/immunology , Immunoglobulin M/blood , Immunoglobulin M/immunology , Male , Probability , Prospective Studies , Recombinant Fusion Proteins/blood , Recombinant Fusion Proteins/immunology , Sensitivity and Specificity , Serologic Tests/methods
14.
Am J Vet Res ; 62(9): 1365-9, 2001 Sep.
Article in English | MEDLINE | ID: mdl-11560261

ABSTRACT

OBJECTIVE: To test serum samples of dogs and horses by use of class-specific recombinant-based ELISA for establishing a diagnosis of granulocytic ehrlichiosis attributable to infection with organisms from the Ehrlichia phagocytophila genogroup. SAMPLE POPULATION: Serum samples from 43 client-owned dogs and 131 horses (81 with signs of acute illness and 50 without signs of disease). PROCEDURE: Serum samples were analyzed, using ELISA with a recombinant 44-kd protein antigen for IgM and IgG antibodies to the human granulocytic ehrlichiosis (HGE) agent (NCH-1 strain). Western blot analyses, using infected human promyelocytic leukemia cells, were conducted on 38 serum samples of horses and 11 serum samples of dogs to verify reactivity to the 44-kd peptide. RESULTS: IgM or IgG antibodies to the HGE agent were detected in 5 to 28% of dog serum samples and 5 to 37% of horse serum samples. Thirty-five of 38 (92%) horse serum samples had corresponding results on both tests (2 positive results for 26 samples and 2 negative results for 9 samples), using an ELISA for IgG antibodies or immunoblotting for total immunoglobulins. All 11 serum samples of dogs had positive results for both methods. CONCLUSION AND CLINICAL RELEVANCE: These ELISA with recombinant 44-kd antigen are suitable for detecting IgM or IgG antibodies to the HGE agent in serum samples of dogs and horses. Positive results for serum samples of horses from Connecticut, New York, Virginia, and Georgia indicate that the HGE agent is widely distributed in tick-infested areas of the eastern United States.


Subject(s)
Dog Diseases/immunology , Ehrlichia/immunology , Ehrlichiosis/veterinary , Enzyme-Linked Immunosorbent Assay/veterinary , Horse Diseases/immunology , Animals , Antibodies, Bacterial/blood , Blotting, Western/veterinary , Connecticut , Dog Diseases/blood , Dog Diseases/microbiology , Dogs , Ehrlichia/growth & development , Ehrlichiosis/blood , Ehrlichiosis/immunology , Ehrlichiosis/microbiology , Enzyme-Linked Immunosorbent Assay/methods , Fluorescent Antibody Technique, Indirect/veterinary , Georgia , Horse Diseases/blood , Horse Diseases/microbiology , Horses , Humans , Immunoglobulin G/blood , Immunoglobulin M/blood , New York , Recombinant Proteins , Virginia
15.
Infect Immun ; 69(9): 5577-88, 2001 Sep.
Article in English | MEDLINE | ID: mdl-11500432

ABSTRACT

The agent of human granulocytic ehrlichiosis (HGE) is an obligate intracellular bacterium with a tropism for neutrophils; however, the mechanisms of bacterial dissemination are not yet understood. Interleukin-8 (IL-8) is a chemokine that induces neutrophil migration to sites of infection for host defense against pathogens. We now show that HGE bacteria, and the HGE-44 protein, induce IL-8 secretion in a promyelocytic (HL-60) cell line that has been differentiated along the neutrophil lineage with retinoic acid and in neutrophils. Infected HL-60 cells also demonstrate upregulation of CXCR2, an IL-8 receptor, but not CXCR1. Human neutrophils migrate towards Ehrlichia sp.-infected cells in a chemotaxis chamber assay, and this movement can be blocked with antibodies to IL-8. Finally, immunocompetent and severe combined immunodeficient mice administered CXCR2 antisera, and CXCR2(-/-) mice that lack the human IL-8 receptor homologue, are much less susceptible to granulocytic ehrlichiosis than are control animals. These results demonstrate that HGE bacteria induce IL-8 production by host cells and, paradoxically, appear to exploit this chemokine to enhance infection.


Subject(s)
Chemotaxis, Leukocyte , Ehrlichia/immunology , Ehrlichiosis/immunology , Interleukin-8/pharmacology , Neutrophils/immunology , Animals , Antigens, Bacterial/immunology , Ehrlichiosis/microbiology , Granulocytes , HL-60 Cells , Humans , Mice , Mice, Inbred BALB C , Mice, Inbred C3H , Mice, SCID , Neutrophils/microbiology , Receptors, Interleukin-8B/immunology , Receptors, Interleukin-8B/metabolism , Recombination, Genetic , Spleen/cytology , Spleen/immunology
17.
J Immunol ; 166(12): 7398-403, 2001 Jun 15.
Article in English | MEDLINE | ID: mdl-11390491

ABSTRACT

Borrelia burgdorferi outer surface protein (Osp) A is preferentially expressed by spirochetes in the Ixodes scapularis gut and facilitates pathogen-vector adherence in vitro. Here we examined B. burgdorferi-tick interactions in vivo by using Abs directed against OspA from each of the three major B. burgdorferi sensu lato genospecies: B. burgdorferi sensu stricto, Borrelia afzelii, and Borrelia garinii. Abs directed against B. burgdorferi sensu stricto (isolate N40) destroy the spirochete and can protect mice from infection. In contrast, antisera raised against OspA from B. afzelii (isolate ACA-1) and B. garinii (isolate ZQ-1) bind to B. burgdorferi N40 but are not borreliacidal against the N40 isolate. Our present studies assess whether these selected OspA Abs interfere with B. burgdorferi-tick attachment in a murine model of Lyme disease with I. scapularis. We examined engorged ticks that had fed on B. burgdorferi N40-infected scid mice previously treated with OspA (N40, ACA-1, ZQ-1, or mAb C3.78) or control Abs. OspA-N40 antisera or mAb C3.78 destroyed B. burgdorferi N40 within the engorged ticks. In contrast, treatment of mice with OspA-ACA-1 and OspA-ZQ-1 antisera did not kill B. burgdorferi N40 within the ticks but did effectively interfere with B. burgdorferi-I. scapularis adherence, thereby preventing efficient colonization of the vector. These studies show that nonborreliacidal OspA Abs can inhibit B. burgdorferi attachment to the tick gut, highlighting the importance of OspA in spirochete-arthropod interactions in vivo.


Subject(s)
Anti-Bacterial Agents/administration & dosage , Antibodies, Bacterial/administration & dosage , Antigens, Surface/immunology , Bacterial Outer Membrane Proteins/immunology , Borrelia burgdorferi Group/immunology , Ixodes/immunology , Ixodes/microbiology , Lipoproteins , Lyme Disease Vaccines/immunology , Animals , Anti-Bacterial Agents/chemistry , Anti-Bacterial Agents/metabolism , Antibodies, Bacterial/chemistry , Antibodies, Bacterial/metabolism , Antibodies, Monoclonal/administration & dosage , Antibodies, Monoclonal/chemistry , Antibodies, Monoclonal/metabolism , Antigens, Surface/genetics , Antigens, Surface/metabolism , Bacterial Outer Membrane Proteins/genetics , Bacterial Outer Membrane Proteins/metabolism , Bacterial Vaccines , Binding Sites, Antibody/genetics , Borrelia burgdorferi Group/genetics , Digestive System/immunology , Digestive System/metabolism , Digestive System/microbiology , Epitopes/genetics , Epitopes/metabolism , Female , Immune Sera/administration & dosage , Immune Sera/chemistry , Immune Sera/metabolism , Injections, Intraperitoneal , Injections, Subcutaneous , Ixodes/anatomy & histology , Ixodes/metabolism , Lyme Disease/immunology , Lyme Disease/prevention & control , Lyme Disease Vaccines/genetics , Lyme Disease Vaccines/metabolism , Mice , Mice, SCID , Mutation , Protein Structure, Tertiary/genetics , Recombinant Proteins/immunology , Recombinant Proteins/metabolism
18.
Infect Immun ; 69(5): 3359-71, 2001 May.
Article in English | MEDLINE | ID: mdl-11292759

ABSTRACT

Lyme disease and human granulocytic ehrlichiosis (HGE) are tick-borne illnesses caused by Borrelia burgdorferi and the agent of HGE, respectively. We investigated the influence of dual infection with B. burgdorferi and the HGE agent on the course of murine Lyme arthritis and granulocytic ehrlichiosis. Coinfection resulted in increased levels of both pathogens and more severe Lyme arthritis compared with those in mice infected with B. burgdorferi alone. The increase in bacterial burden during dual infection was associated with enhanced acquisition of both organisms by larval ticks that were allowed to engorge upon infected mice. Coinfection also resulted in diminished interleukin-12 (IL-12), gamma interferon (IFN-gamma), and tumor necrosis factor alpha levels and elevated IL-6 levels in murine sera. During dual infection, IFN-gamma receptor expression on macrophages was also reduced, implying a decrease in phagocyte activation. These results suggest that coinfection of mice with B. burgdorferi and the HGE agent modulates host immune responses, resulting in increased bacterial burden, Lyme arthritis, and pathogen transmission to the vector.


Subject(s)
Ehrlichiosis/immunology , Lyme Disease/immunology , Neutrophils/microbiology , Animals , Antibodies, Bacterial/blood , CD40 Ligand/analysis , Cytokines/blood , DNA, Bacterial/analysis , Ehrlichiosis/complications , Ehrlichiosis/microbiology , Humans , Lyme Disease/complications , Lyme Disease/microbiology , Mice , Mice, Inbred C3H , Mice, SCID , Ticks/microbiology
19.
Infect Immun ; 69(5): 3507-9, 2001 May.
Article in English | MEDLINE | ID: mdl-11292785

ABSTRACT

Clonal Borrelia burgdorferi N40 (cN40) passaged 75 times in vitro (N40-75) infects mice but does not cause disease. N40-75 passaged 45 times further in vitro (N40-120) was no longer infectious and lacked genes encoded on linear plasmids 38 and 28-1, among other differences. These data suggest that B. burgdorferi cN40, N40-75, and N40-120 have distinct phenotypes that can be used to dissect the genetic elements responsible for pathogenicity and infectivity.


Subject(s)
Borrelia burgdorferi Group/genetics , Borrelia burgdorferi Group/pathogenicity , Animals , Disease Models, Animal , Lyme Disease/etiology , Mice , Mice, Inbred C3H , Plasmids
20.
Am J Vet Res ; 62(1): 29-32, 2001 Jan.
Article in English | MEDLINE | ID: mdl-11197555

ABSTRACT

OBJECTIVE: To develop and evaluate a polyvalent ELISA incorporating a highly specific recombinant antigen (p44) for diagnosis of granulocytic ehrlichiosis in dogs and horses. ANIMALS: 32 dogs and 43 horses. PROCEDURE: Results of the ELISA were compared with results of indirect fluorescent antibody (IFA) staining and western immunoblotting incorporating whole-cell antigen. RESULTS: For the canine and equine samples, percentages of samples with positive IFA staining, western immunoblotting, and ELISA results were similar. For 29 (91 %) canine samples and 30 (70%) equine samples, results of IFA staining, western immunoblotting, and the ELISA were in complete agreement. Results of the ELISA for 3 canine serum samples known to contain antibodies to Ehrlichia canis and 12 equine serum samples known to contain antibodies to E risticii were negative. CONCLUSIONS AND CLINICAL RELEVANCE: Results of the present study suggest that a polyvalent ELISA incorporating a recombinant p44 antigen is suitable for detecting antibodies to E equi in dogs and horses.


Subject(s)
Dog Diseases/diagnosis , Ehrlichiosis/veterinary , Horse Diseases/diagnosis , Animals , Blotting, Western , Dog Diseases/blood , Dog Diseases/microbiology , Dogs , Ehrlichiosis/blood , Ehrlichiosis/diagnosis , Enzyme-Linked Immunosorbent Assay/methods , Fluorescent Antibody Technique, Indirect , Horse Diseases/blood , Horse Diseases/microbiology , Horses , Recombinant Proteins
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