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Jere W McBride

Publications and source records attributed to Jere W McBride.

At least 19 recordsLinked to original sources
PubMed2006

Enzyme-linked immunosorbent assay with conserved immunoreactive glycoproteins gp36 and gp19 has enhanced sensitivity and provides species-specific immunodiagnosis of Ehrlichia canis infection.

Ehrlichia canis is the primary etiologic agent of canine monocytic ehrlichiosis, a globally distributed and potentially fatal disease of dogs. We previously reported on the identification of two conserved major immunoreactive antigens, gp36 and gp19, which are the first proteins to elicit an E. canis-specific antibody response, and gp200 and p28, which elicit strong antibody responses later in the acute phase of the infection. In this report, the sensitivities and specificities of five recombinant E. canis proteins for the immunodiagnosis of E. canis infection by an enzyme-linked immunosorbent assay (ELISA) were evaluated. Recombinant polypeptides gp36, gp19, and gp200 (N and C termini) exhibited 100% sensitivity and specificity for immunodiagnosis by the recombinant glycoprotein ELISA compared with the results obtained by an indirect fluorescent-antibody assay (IFA) for the detection of antibodies in dogs that were naturally infected with E. canis. Moreover, the enhanced sensitivities of gp36 and gp19 for immunodiagnosis by the recombinant glycoprotein ELISA compared to those obtained by IFA were demonstrated with dogs experimentally infected with E. canis, in which antibodies were detected as much as 2 weeks earlier, on day 14 postinoculation. gp36 and gp19 were not cross-reactive with antibodies in sera from E. chaffeensis-infected dogs and thus provided species-specific serologic discrimination between E. canis and E. chaffeensis infections. This is the first demonstration of the improved detection capability of the recombinant protein technology compared to the capability of the "gold standard" IFA and may eliminate the remaining obstacles associated with the immunodiagnosis of E. canis infections, including species-specific identification and the lack of sensitivity associated with low antibody titers early in the acute phase of the infection.

Animals↗
PubMed2006

Identification of a glycosylated Ehrlichia canis 19-kilodalton major immunoreactive protein with a species-specific serine-rich glycopeptide epitope.

Ehrlichia canis has a small subset of major immunoreactive proteins that includes a 19-kDa protein that elicits an early Ehrlichia-specific antibody response in infected dogs. We report herein the identification and molecular characterization of this highly conserved 19-kDa major immunoreactive glycoprotein (gp19) ortholog of the Ehrlichia chaffeensis variable-length PCR target (VLPT) protein. E. canis gp19 has substantial carboxyl-terminal amino acid homology (59%) with E. chaffeensis VLPT and the same chromosomal location; however, the E. chaffeensis VLPT gene (594 bp) has tandem repeats that are not present in the E. canis gp19 gene (414 bp). Consistent with other ehrlichial glycoproteins, the gp19 protein exhibited a larger-than-predicted mass (approximately 3 kDa), O-linked glycosylation sites were predicted in an amino-terminal serine/threonine/glutamate (STE)-rich patch (26 amino acids), carbohydrate was detected on the recombinant gp19 protein, and the neutral sugars glucose and galactose were detected on the recombinant amino-terminal polypeptide. E. canis gp19 composition consists of five predominant amino acids, cysteine, glutamate, tyrosine, serine, and threonine, concentrated in the STE-rich patch and a carboxyl-terminal domain predominated by cysteine and tyrosine (55%). The amino-terminal STE-rich patch contained a major species-specific antibody epitope strongly recognized by serum from an E. canis-infected dog. The recombinant glycopeptide epitope was substantially more reactive with antibody than the synthetic (nonglycosylated) peptide, and periodate treatment of the recombinant glycopeptide epitope reduced its immunoreactivity, demonstrating the importance of a carbohydrate immunodeterminant(s). The gp19 protein was present on reticulate and dense-cored cells, and it was found extracellularly in the fibrillar matrix and associated with the morula membrane, the host cell cytoplasm, and the nucleus.

Amino Acid Sequence↗
PubMed2006

Restriction and expansion of Ehrlichia strain diversity.

Ehrlichia are tick-borne gram negative, obligately intracellular bacteria. The 16S rRNA gene DNA sequences are highly conserved among strains of each Ehrlichia species. The 28-kDa/Map-1 outer membrane protein genes are highly diversified among strains of Ehrlichia chaffeensis and E. ruminantium, but are highly conserved among E. canis isolates. The diversity of the immunodominant proteins of E. chaffeensis and E. ruminantium in contrast with the conservation of the immunodominant proteins of E. canis suggests that E. chaffeensis and E. ruminantium face more host immune pressure than E. canis or that E. chaffeensis and E. ruminantium evolved earlier than E. canis and have diverged.

Bacterial Outer Membrane Proteins↗
PubMed2006

A preliminary investigation of Ehrlichia species in ticks, humans, dogs, and capybaras from Brazil.

A molecular epidemiologic investigation in two Brazilian states (Rondônia and São Paulo) was undertaken to determine if Ehrlichia species responsible for human and animal ehrlichioses in North America could be found in Brazilian vectors, potential natural mammalian reservoirs and febrile human patients with a tick bite history. Samples, including 376 ticks comprising 9 Amblyomma species, 29 capybara (Hydrochaeris hydrochaeris) spleens, 5 canine blood, and 75 human blood samples from febrile patients with history of tick bites were tested by a real-time PCR assay targeting a fragment of the Ehrlichia dsb gene. Ehrlichia DNA was not detected in any tick, capybara or human samples. In contrast, 4 out of 5 dogs contained Ehrlichia canis DNA in their blood, which were sequenced, representing the first report of E. canis infecting dogs in the Amazon region of Brazil. Further studies are needed to evaluate the presence of other agents of human and animal ehrlichioses in Brazil.

Animals↗
PubMed2006

Differentially expressed and secreted major immunoreactive protein orthologs of Ehrlichia canis and E. chaffeensis elicit early antibody responses to epitopes on glycosylated tandem repeats.

Ehrlichia canis major immunoreactive proteins of 36 and 19 kDa elicit the earliest detectable antibody responses during the acute phase of canine monocytic ehrlichiosis. Genes encoding the major immunoreactive 36-kDa protein of E. canis and the corresponding ortholog of E. chaffeensis (47 kDa) were identified and the proteins characterized. The molecular masses of the strongly immunoreactive recombinant proteins were larger than predicted (26.7 and 32.9 kDa, respectively) but were consistent with those of the corresponding native proteins (36 and 47 kDa). Similar to other reported ehrlichial immunoreactive glycoproteins, carbohydrate was detected on the recombinant expressed proteins, indicating that they were glycoproteins. Both glycoproteins (gp36 and gp47) have carboxy-terminal serine/threonine-rich tandem repeat regions containing repeats that vary in number (4 to 16 repeats) and amino acid sequence among different isolates of each species. E. canis gp36 was recognized by early acute-phase antibodies (day 14), and species-specific antibody epitopes were mapped to C-terminal nonhomologous repeat units of gp36 and gp47. Periodate treatment of recombinant gp36 reduced the antibody reactivity, and nonglycosylated synthetic peptide repeat units from E. canis gp36 and E. chaffeensis gp47 were substantially less immunoreactive than corresponding recombinant peptides, demonstrating that glycans are important epitope determinants that are structurally conserved on the recombinant proteins expressed in Escherichia coli. E. canis gp36 and E. chaffeensis gp47 were differentially expressed only on the surface of dense-cored ehrlichiae and detected in the Ehrlichia-free supernatants, indicating that these proteins are released extracellularly during infection.

Amino Acid Sequence↗
PubMed2006

Early diagnosis of rickettsioses by electrochemiluminscence.

The diagnosis of acute rickettsioses during the acute phase of the disease is challenging. We present preliminary evidence that antigen-capture using streptavidin-coated magnetic beads with biotinylated anti-rickettsia typhi rabbit polyclonal antibodies followed by electrochemiluminescent detection with ruthenylated antibodies of the same specificity could be used for the diagnosis of rickettsial diseases in the acute phase.

Animals↗
PubMed2005

Detection of medically important Ehrlichia by quantitative multicolor TaqMan real-time polymerase chain reaction of the dsb gene.

Ehrlichia species are the etiological agents of emerging and life-threatening tick-borne human zoonoses, in addition to causing serious and fatal infections in companion animals and livestock. We developed the first tricolor TaqMan real-time polymerase chain reaction assay capable of simultaneously detecting and discriminating medically important ehrlichiae in a single reaction. Analytical sensitivity of 50 copies per reaction was attained with templates from Ehrlichia chaffeensis, Ehrlichia ewingii, and Ehrlichia canis by amplifying the genus-specific disulfide bond formation protein gene (dsb). Ehrlichia genus-specific dsb primers amplified DNA from all known Ehrlichia species but not from other rickettsial organisms including Anaplasma platys, Anaplasma phagocytophilum, Rickettsia conorii, or Rickettsia typhi. High species specificity was attained as each species-specific TaqMan probe (E. chaffeensis, E. ewingii, and E. canis) identified homologous templates but did not cross-hybridize with heterologous Ehrlichia templates at concentrations as high as 10(8) copies. Identification of E. chaffeensis, E. ewingii, and E. canis from natural and experimental infections, previously confirmed by polymerase chain reaction and serological or microscopic evidence, demonstrated the comparable specificity and sensitivity of the dsb real-time assay. This assay provides a powerful tool for prospective medical diagnosis for human and canine ehrlichioses and for ecologic and epidemiological studies involving arthropod and mammalian hosts.

Animals↗
PubMed2005

An immunoreactive 38-kilodalton protein of Ehrlichia canis shares structural homology and iron-binding capacity with the ferric ion-binding protein family.

Ehrlichiae are tick-transmitted, gram-negative, obligately intracellular bacteria that live and replicate in cytoplasmic vacuoles, but little is known about iron acquisition mechanisms necessary for their survival. In this study, a genus-conserved immunoreactive ferric ion-binding protein (Fbp) of Ehrlichia canis was identified and its iron-binding capability was investigated. E. canis Fbp was homologous to a family of periplasmic Fbp's involved in iron acquisition and transport in gram-negative bacteria. E. canis Fbp had a molecular mass (38 kDa) consistent with those of Fbp's in other bacteria and exhibited substantial immunoreactivity in its native conformation. The predicted three-dimensional structure of E. canis Fbp demonstrated conservation of important Fbp family structural motifs: two domains linked with a polypeptide "hinge" region. Under iron-binding conditions, the recombinant Fbp exhibited an intense red color and an absorbance spectrum indicative of iron binding, and it bound Fe(III) but not Fe(II). Fbp was observed primarily in the cytoplasm of the reticulate forms of E. canis and Ehrlichia chaffeensis but was notably found on extracellular morula fibers in morulae containing dense-cored organisms. Although expression of Fbp is regulated through an operon of three functionally linked genes in other gram-negative bacteria, the absence of an intact fbp operon in Ehrlichia spp. suggests that genes involved in ehrlichial iron acquisition have been subject to reductive evolution.

Animals↗
PubMed2005

Essential role for humoral immunity during Ehrlichia infection in immunocompetent mice.

Although cellular immunity is essential for host defense during intracellular bacterial infections, humoral immunity can also play a significant role in host defense during infection by some intracellular bacteria, including the ehrlichiae. Antibodies can protect susceptible SCID mice from fatal Ehrlichia chaffeensis infection, an observation that has been hypothesized to involve the opsonization of bacteria released from host cells. To determine whether humoral immunity plays an essential role during ehrlichia infection in immunocompetent mice, we utilized a murine model of fatal monocytotropic ehrlichiosis caused by Ixodes ovatus ehrlichia. Mice lacking either B cells or FcgammaRI were unable to resolve a low-dose (sublethal) I. ovatus ehrlichia infection, which suggested that humoral immunity is essential for resistance. Polyclonal sera generated in I. ovatus ehrlichia-infected mice recognized a conserved ehrlichia outer membrane protein and, when administered to infected mice, caused a significant decrease in bacterial infection. Mice experimentally depleted of complement, or deficient for complement receptors 1 and 2, were also susceptible to sublethal I. ovatus ehrlichia infection, as were mice that lacked the phox91 subunit of NADPH oxidase. The data are consistent with a mechanism whereby bacteria released from infected cells are lysed directly by complement or undergo antibody-mediated FcgammaR-dependent phagocytosis and subsequent exposure to reactive oxygen intermediates. The findings suggest mechanisms whereby antibodies contribute to immunity against intracellular bacteria in immunocompetent mice.

Animals↗
PubMed2005

Analysis of ehrlichial p28 gene expression in a murine model of persistent infection.

Historically, ehrlichioses were tick-borne diseases of veterinary medical importance and are now important emerging infectious diseases in humans. p28s are encoded by multigene families with ORFs tandemly arranged with intergenic spaces of variable lengths. We reported initial sequencing of the Ehrlichia muris p28 locus. A model of persistent infection was described and provided tools for study of persistent ehrlichial infection. We completed the sequence of the E. muris p28 locus and examined mRNA expression.

Animals↗
PubMed2004

Overproduction of TNF-alpha by CD8+ type 1 cells and down-regulation of IFN-gamma production by CD4+ Th1 cells contribute to toxic shock-like syndrome in an animal model of fatal monocytotropic ehrlichiosis.

Human monocytotropic ehrlichiosis (HME) is an emerging, life-threatening, infectious disease caused by Ehrlichia chaffeensis, an obligate intracellular bacterium that lacks cell wall LPS. We have previously developed an animal model of severe HME using a strain of Ehrlichia isolated from Ixodes ovatus ticks (IOE). To understand the basis of susceptibility to severe monocytotropic ehrlichiosis, we compared low and high doses of the highly virulent IOE strain and the less virulent Ehrlichia muris strain that are closely related to E. chaffeensis in C57BL/6 mice. Lethal infections caused by high or low doses of IOE were accompanied by extensive liver damage, extremely elevated levels of TNF-alpha in the serum, high frequency of Ehrlichia-specific, TNF-alpha-producing CD8(+) T cells in the spleen, decreased Ehrlicha-specific CD4(+) T cell proliferation, low IL-12 levels in the spleen, and a 40-fold decrease in the number of IFN-gamma-producing CD4(+) Th1 cells. All groups contained negligible numbers of IL-4-producing cells in the spleen. Transfer of Ehrlichia-specific polyclonal Abs and IFN-gamma-producing Ehrlichia-specific CD4(+) and CD8(+) type 1 cells protected naive mice against lethal IOE challenge. Interestingly, infection with high dose E. muris provided protection against rechallenge with a lethal dose of IOE. Cross-protection was associated with substantial expansion of IFN-gamma-producing CD4(+) and CD8(+) cells, but not TNF-alpha-producing CD8(+) T cells, a high titer of IgG2a, and a low serum level of TNF-alpha. In conclusion, uncontrolled TNF-alpha production by CD8(+) T cells together with a weak CD4(+) Th1 cell response are associated with immunopathology and failure to clear IOE in the fatal model of HME.

Adoptive Transfer↗
PubMed2004

Histologic, serologic, and molecular analysis of persistent ehrlichiosis in a murine model.

Human monocytotropic ehrlichiosis caused by Ehrlichia chaffeensis was reported in 1987. An animal model to study acute fatal ehrlichiosis in mice that has been developed closely resembles the fatal form of human monocytotropic ehrlichiosis. However, animal models for persistent infection in the genus Ehrlichia in immunocompetent mice have not been characterized. We report the histopathological progression of Ehrlichia muris infection in immunocompetent mice (AKR and C57BL/6 strains) correlated with their antibody response determined by indirect immunofluorescence and Western immunoblotting, and the distribution and quantity of the ehrlichial load by immunohistochemistry, polymerase chain reaction (PCR), and real-time PCR in lungs, liver, and spleen. Mild to moderate correlation was observed between histopathological grading in these organs and relative ehrlichial loads. The highest ehrlichial loads were present between days 4 and 14 after infection. E. muris was detected in tissues examined up to 150 days after infection by real-time PCR. Analysis of the serological response revealed several immunodominant antigens, including 200-, 180-, 100-, 73/75-, 45-, and 28-kd proteins. In conclusion, we have provided for the first time a complete histopathological, serological, immunohistochemical, and quantitative analysis of an animal model for the study of persistent ehrlichial infection.

Animals↗
PubMed2004

Rickettsia species infecting Amblyomma cooperi ticks from an area in the state of São Paulo, Brazil, where Brazilian spotted fever is endemic.

Owing to the potential role of the tick Amblyomma cooperi in the enzootic cycle of Rickettsia rickettsii, the etiologic agent of Brazilian spotted fever (BSF), this study evaluated infection by Rickettsia species in A. cooperi ticks collected from an area in Brazil where BSF is endemic. Among a total of 40 A. cooperi adult ticks collected in an area of BSF endemicity in the state of São Paulo, PCR analysis detected DNA of Rickettsia bellii in 16 ticks (40%), and 3 other ticks (7.5%) were positive for a previously unidentified spotted-fever-group (SFG) rickettsia. Cultivation in Vero cell cultures by the shell vial technique with individual A. cooperi ticks resulted in two isolates of R. bellii and one isolate genotypically characterized as an SFG rickettsia. The two R. bellii isolates were established in Vero cell cultures in the laboratory and were confirmed to be R. bellii by molecular analysis of the gltA and 17-kDa protein-encoding genes and by electron microscopic analysis. The SFG rickettsial isolate could not be stably passaged in cell culture in the laboratory, but molecular analysis of early passages suggested that it was closely related to Rickettsia parkeri, Rickettsia africae, and Rickettsia sibirica. These results do not support the role of A. cooperi in the ecology of R. rickettsii in the area studied, but they add two more species of rickettsiae to the poorly developed list of species occurring in ticks in South America.

Animals↗
PubMed2004

Molecular evidence for a spotted fever group Rickettsia species in the tick Amblyomma longirostre in Brazil.

Two Amblyomma longirostre adult male ticks were collected from a Brazilian porcupine Coendou prehensilis L. in the state of Rondonia, Western Amazon, Brazil. The two ticks were pooled for DNA extraction and tested for the presence of rickettsial DNA by amplifying portions of the gltA, 17-kDa, ompA, and ompB rickettsial genes by polymerase chain reaction (PCR). Portions of the four genes were amplified from the sample and subsequently sequenced. These results indicated the presence of a Rickettsia strain infecting A. longirostre, which was designated as strain Aranha. Compared with homologous ompA rickettsial sequences, "Rickettsia amblyommii" ompA seemed to be the closest relative to Aranha (similarity values: 99.0-99.3%). Phylogenetic analyses of more conserved genes including 17-kDa and gtlA partial sequences indicated that this Rickettsia sp. is a spotted fever group rickettsia. The partial ompB sequence of strain Aranha was distinct from all homologous sequences available in GenBank. Although our ompA analysis suggested a very close molecular phylogenetic relationship of Aranha with "R. amblyommii," we cannot at this time determine if Aranha is a new strain of "R. amblyommii" or a new Rickettsia species in South America.

Animals↗
PubMed2004

Detection of Rickettsia africae in patients and ticks along the coastal region of Cameroon.

Rickettsia africae was identified in seven (6%) of 118 patients with acute fevers of unknown etiology proven not to be malaria or typhoid fever from clinics along the coastal region of Cameroon by polymerase chain reaction (PCR) amplification and sequencing of the citrate synthase (gltA) and outer membrane protein A (ompA) genes of Rickettsia. The majority (71%) of the patients were female. Clinical manifestations included fever (100%), headache (71%), myalgia (71%), arthralgia (43%), pulmonary involvement (29%), and diffuse rash (14%). Moreover, R. africae was detected by PCR amplification and sequence analysis of the gltA and ompA genes in 62 (75%) of 83 adult Amblyomma variegatum ticks collected from cattle in the same region. These results confirm the presence of a previously unrecognized infectious disease in the indigenous Cameroonian population, as well as extend the established range of R. africae.

Adult↗
PubMed2004

Ehrlichia chaffeensis: a prevalent, life-threatening, emerging pathogen.

Ehrlichia chaffeensis are small, obligately intracellular, endosomal bacteria with tropism for macrophages. Persistent infection in reservoir white-tailed deer is transmitted by lone star ticks. Flu-like illness can progress to severe multisystem disease with toxic shock-like syndrome, meningitis, or ARDS. The case-fatality rate is 2.7%. Leukopenia and thrombocytopenia are diagnostically useful. Granulomas are associated with control of the infection. Ehrlichial proteins and glycoproteins have been sequenced and expressed for diagnostic serology and vaccine development. Mouse models (mild disease and persistent infection with E. muris and fatal monocytotropic ehrlichiosis with a Japanese tick isolate) revealed that CD4 and CD8 T type 1 lymphocyte responses, IFN-gamma, TNF-alpha, and antibodies play roles in protective immunity, while a weak CD4 T-helper response, overproduction of TNF-alpha, and very high IL-10 are associated with toxic shock-like mortality. Protection against fatal ehrlichiosis was achieved by prior infection with low virulence E. muris. Acute clinical diagnosis is difficult except by PCR. Response to doxycycline is dramatic.

Animals↗
PubMed2003

L-selectin and E-selectin expressed on monocytes mediating Ehrlichia chaffeensis attachment onto host cells.

Ehrlichia chaffeensis, the agent of human monocytic ehrlichiosis, is an obligatory intracellular bacterium that exhibits monocytic host cell tropism. Ehrlichiae must enter the host cell, and then establish infection. The tropism of E. chaffeensis for monocytes suggests that the cell contains some specific surface components that mediate E. chaffeensis attachment and entry into host cells. In this study, host cell surface components that play a role in ehrlichial attachment were identified using a human monocyte/macrophage cell line, THP-1. E. chaffeensis attachment to THP-1 cells was partially blocked in the presence of antibodies to E-selectin and L-selectin, but not by antibodies to P-selectin, integrin alpham, integrin alphax, or normal mouse IgG as determined by real time polymerase chain reaction. Conversely, in HeLa cells that do not exhibit surface expression of E-selectin and L-selectin, antibodies to these cell surface proteins did not inhibit E. chaffeensis attachment. These findings indicate that E-selectin and L-selectin are cell surface proteins that might act as co-receptors and contribute to E. chaffeensis attachment and entry into THP-1.

Bacterial Adhesion↗