Search PubMed⌕ Search

Biomedical subjects

A G Barbour

Publications and source records attributed to A G Barbour.

At least 37 records · Page 2Linked to original sources
PubMed1999

Esterases in serum-containing growth media counteract chloramphenicol acetyltransferase activity in vitro.

The spirochete Borrelia burgdorferi was unexpectedly found to be as susceptible to diacetyl chloramphenicol, the product of the enzyme chloramphenicol acetyltransferase, as it was to chloramphenicol itself. The susceptibilities of Escherichia coli and Bacillus subtilis, as well as that of B. burgdorferi, to diacetyl chloramphenicol were then assayed in different media. All three species were susceptible to diacetyl chloramphenicol when growth media were supplemented with rabbit serum or, to a lesser extent, human serum. Susceptibility of E. coli and B. subtilis to diacetyl chloramphenicol was not observed in the absence of serum, when horse serum was used, or when the rabbit or human serum was heated first. In the presence of 10% rabbit serum, a strain of E. coli bearing the chloramphenicol acetyltransferase (cat) gene had a fourfold-lower resistance to chloramphenicol than in the absence of serum. A plate bioassay for chloramphenicol activity showed the conversion by rabbit, mouse, and human sera but not bacterial cell extracts or heated serum of diacetyl chloramphenicol to an inhibitory compound. Deacetylation of acetyl chloramphenicol by serum components was demonstrated by using fluorescent substrates and thin-layer chromatography. These studies indicate that esterases of serum can convert diacetyl chloramphenicol back to an active antibiotic, and thus, in vitro findings may not accurately reflect the level of chloramphenicol resistance by cat-bearing bacteria in vivo.

Animals↗
PubMed1999

In vitro activities of the everninomicin SCH 27899 and other newer antimicrobial agents against Borrelia burgdorferi.

The in vitro activity of the everninomicin antibiotic SCH 27899 against 17 isolates of Borrelia spp. was investigated. MICs ranged from 0.06 to 0.5 microg/ml. Time-kill studies with the B31 strain of B. burgdorferi demonstrated >/=3-log10-unit killing after 72 h with concentrations representing four times the MIC. The in vitro activity of four other newer antimicrobial agents, meropenem, cefepime, quinupristin-dalfopristin, and linezolid, was also tested against the B31 strain. Meropenem was the most potent of the latter agents, with an MIC of 0.125 microg/ml.

Aminoglycosides↗
PubMed1999

Access of antibody or trypsin to an integral outer membrane protein (P66) of Borrelia burgdorferi is hindered by Osp lipoproteins.

The outer membrane of Borrelia burgdorferi, the Lyme disease agent, contains lipoproteins anchored by their lipid moieties and integral proteins with membrane-spanning regions. We used the techniques of in situ proteolysis, immunofluorescence, in vitro growth inhibition, and cross-linking with formaldehyde to characterize topological relationships between P66, an integral membrane protein, and selected Osp lipoproteins of B. burgdorferi. Protease treatment of intact spirochetes cleaved P66 and Osp proteins but not the periplasmic flagellin or the BmpA protein of the cytoplasmic membrane. P66 of cells lacking OspA, OspB, and OspC was more susceptible to trypsin cleavage than was P66 of cells with these Osp proteins. A monoclonal antibody against the surface loop of P66 bound, agglutinated, and inhibited the growth of viable spirochetes lacking OspA, OspB, OspC, and OspD but not of the cells that expressed OspA, OspC, and/or OspD. When cells were fixed, the antibody bound to cells that express OspD and OspC but still not to cells with OspA. The close association of OspA and P66 was confirmed by the crosslinking of the two proteins by formaldehyde. These results show that Osp proteins, particularly OspA, limit the access of antibody or trypsin to the surface loop region of P66. The proximity and possible contact between P66 and OspA (or other Osp proteins) may hinder the effectiveness of antibodies to what otherwise would be an appropriate vaccine target.

Antibodies, Bacterial↗
PubMed1999

Comparative analysis and immunological characterization of the Borrelia Bdr protein family.

Multiple circular and linear plasmids of Lyme disease and relapsing fever Borrelia spirochetes carry genes for members of the Bdr (Borrelia direct repeat) protein family. To define their common and divergent attributes, we first comprehensively compared the known homologs. Bdr proteins with predicted sizes ranging from 10.7 to 30. 6 kDa formed five homology groups, based on variable numbers of short direct repeats in a central domain and diverse N- and C-terminal domains. In a further characterization, Western blots were probed with rabbit antisera raised against either of two purified recombinant Bdr proteins from Borrelia burgdorferi B31. The results showed that antibodies cross-react and several Bdr paralogs 19.5 to 30.5 kDa in size are expressed by cultured strain B31 in a temperature-independent manner. In situ proteolysis, immunofluorescence, and growth inhibition assays indicated that Bdr proteins are not surface exposed. Distinct patterns of cross-reacting proteins of 17.5 to 33 kDa were also detected in other B. burgdorferi, Borrelia garinii, and Borrelia afzelii strains as well as in relapsing fever spirochetes Borrelia hermsii and Borrelia turicatae. Last, we examined whether these proteins are antibody targets during Lyme disease. Analysis of 47 Lyme disease patient sera by immunoblotting and enzyme-linked immunosorbent assays showed that 24 (51%) and 20 (43%), respectively, had detectable antibodies to one or more of the Bdr proteins. Together, these data indicate that Bdr proteins constitute a family of cross-reactive Borrelia proteins which are expressed in the course of Lyme disease and in vitro.

Amino Acid Sequence↗
PubMed1999

Characterization of VspB of Borrelia turicatae, a major outer membrane protein expressed in blood and tissues of mice.

Serotypes A and B of the relapsing fever spirochete Borrelia turicatae produce different disease manifestations in infected mice. Whereas serotype B causes more severe arthritis and reaches higher densities in the blood of mice than serotype A, serotype A invades the central nervous system earlier than serotype B during infection. These differences between serotypes A and B in mice are associated with the expression of different surface proteins, VspA and VspB, respectively, in the culture medium. To determine whether these proteins, in particular, VspB, are also expressed in vivo, scid mice infected with B. turicatae were studied. The expression of VspB by spirochetes in the blood was demonstrated in Coomassie blue-stained polyacrylamide gels and Western blots with a specific monoclonal antibody. Indirect immunofluorescence and immunoperoxidase studies confirmed the expression of VspB in the blood and also demonstrated VspB expression in the joints and heart. The gene for VspB was next identified and cloned by using partial amino acid sequencing, reverse transcriptase PCR, and a specific monoclonal antibody. The vspB gene encodes a protein of 216 amino acids that is 68% identical to VspA of B. turicatae and 44 to 56% identical to representative Vsp and OspC lipoproteins of other Borrelia spp. The processed VspB protein was distinguished from 26 other Vsp and OspC proteins by a high predicted isoelectric point at 9.39. The promoter region for vspB was similar to the promoter region for the vsp33 gene of Borrelia hermsii and for the ospC gene of Borrelia burgdorferi, two genes known to be environmentally regulated. These studies established that the virulence-associated VspB protein is expressed by spirochetes in the mouse and that VspB is a novel member of the Vsp-OspC family of proteins.

Amino Acid Sequence↗
PubMed1998

Neuroborreliosis during relapsing fever: review of the clinical manifestations, pathology, and treatment of infections in humans and experimental animals.

The spirochetal disease relapsing fever is caused by different Borrelia species. Relapsing fever is well recognized as an infection of the blood, but little is known about its predilection for the nervous system and the eyes. To investigate neurological and ocular involvement during relapsing fever, we reviewed the clinical manifestations, pathology, and treatment of relapsing fever of humans and experimental animals. The results indicate that Borrelia turicatae and Borrelia duttonii, the agents of tick-borne relapsing fever in southwestern North America and sub-Saharan Africa, respectively, cause neurological involvement as often as Borrelia burgdorferi in Lyme disease. Evidence of this is the frequent occurrence of lymphocytic meningitis and peripheral facial palsy in human disease; the identification of spirochetes in the brain and other nervous tissues of humans, animals, and arthropod vectors; and the persistence of brain infection after treatment with antibiotics that do not readily penetrate the blood-brain barrier.

Animals↗
PubMed1998

Fall and rise of Lyme disease and other Ixodes tick-borne infections in North America and Europe.

Lyme disease is a spirochaetal infection with acute and chronic manifestations. Lyme disease and other infections transmitted by Ixodes species ticks are increasing in temperate and Holarctic regions of the Northern hemisphere. These zoonotic infections are most commonly acquired in suburban residential areas and outdoor recreation areas close to cities. Different enzootic cycles, which include a variety of large and small mammals as well as migratory birds, maintain and distribute in nature the Borrelia species that cause Lyme disease. The rise in cases of Lyme disease and the other Ixodes tick-borne infections is, in part, the consequence of reforestation and the increase in deer populations in developed countries.

Europe↗
PubMed1998

Population structure of the relapsing fever spirochete Borrelia hermsii as indicated by polymorphism of two multigene families that encode immunogenic outer surface lipoproteins.

The tick-borne relapsing fever spirochete Borrelia hermsii evades the mammalian immune system by periodically switching expression among members of two multigene families that encode immunogenic, antigenically distinct outer surface proteins. The type strain, B. hermsii HS1, has at least 40 complete genes and pseudogenes that participate in this multiphasic antigenic variation. Originally termed vmp (for variable major protein) genes, they have been reclassified as vsp (for variable small protein) and vlp (for variable large protein) genes, based on size and amino acid sequence similarities. To date, antigenic variation in B. hermsii has been studied only in the type strain, HS1. Nucleotide sequence comparisons of 23 B. hermsii HS1 genes revealed five distinct groups, the vsp gene family and four subfamilies of vlp genes. We used PCR with family- and subfamily-specific primers, followed by restriction fragment length polymorphism analysis, to compare the vsp and vlp repertoires of HS1 and seven other B. hermsii isolates from Washington, Idaho, and California. This analysis, together with pulsed-field gel electrophoresis genome profiles, revealed that the eight isolates formed three distinct groups, which likely represent clonal lineages. Members of the three groups coexisted in the same geographic area, but they could also be isolated across large geographical distances. This population structure may result from immune selection by the host, as has been proposed for other pathogens with polymorphic antigens.

Borrelia↗
PubMed1998

A surface-exposed region of a novel outer membrane protein (P66) of Borrelia spp. is variable in size and sequence.

A model of the 66-kDa outer membrane protein (P66) of Lyme disease Borrelia spp. predicts a surface-exposed loop near the C terminus. This region contains an antigen commonly recognized by sera from Lyme disease patients. In the present study, this region of P66 and homologous proteins of other Borrelia spp. were further investigated by using monoclonal antibodies, epitope mapping of P66 of Borrelia burgdorferi, and DNA sequencing. A monoclonal antibody specific for B. burgdorferi bound to the portion of P66 that was accessible to proteolysis in situ. The linear epitope for the antibody was mapped within a variable segment of the surface-exposed region. To further study this protein, the complete gene of Borrelia hermsii for a protein homologous to P66 was cloned. The deduced protein was 589 amino acids in length and 58% identical to P66 of B. burgdorferi. The B. hermsii P66 protein was predicted to have a surface-exposed region in the same location as that of B. burgdorferi's P66 protein. With primers designed on the basis of conserved sequences and PCR, we identified and cloned the same regions of P66 proteins of Borrelia turicatae, Borrelia parkeri, Borrelia coriaceae, and Borrelia anserina. The deduced protein sequences from all species demonstrated two conserved hydrophobic regions flanking a surface-exposed loop. The loop sequences were highly variable between different Borrelia spp. in both sequence and size, varying between 35 and 45 amino acids. Although the actual function of P66 of Borrelia spp. is unknown, the results suggest that its surface-exposed region is subject to selective pressure.

Amino Acid Sequence↗
PubMed1997

Antigenic variation in Lyme disease borreliae by promiscuous recombination of VMP-like sequence cassettes.

We have identified and characterized an elaborate genetic system in the Lyme disease spirochete Borrelia burgdorferi that promotes extensive antigenic variation of a surface-exposed lipoprotein, VlsE. A 28 kb linear plasmid of B. burgdorferi B31 (lp28-1) was found to contain a vmp-like sequence (vls) locus that closely resembles the variable major protein (vmp) system for antigenic variation of relapsing fever organisms. Portions of several of the 15 nonexpressed (silent) vls cassette sequences located upstream of vlsE recombined into the central vlsE cassette region during infection of C3H/HeN mice, resulting in antigenic variation of the expressed lipoprotein. This combinatorial variation could potentially produce millions of antigenic variants in the mammalian host.

Amino Acid Sequence↗
PubMed1997

An OspA-based DNA vaccine protects mice against infection with Borrelia burgdorferi.

Immunization with recombinant OspA protein of Borrelia burgdorferi protects against experimental Lyme disease. In the present study, mice were injected intramuscularly with plasmid DNA (VR2210) encoding strain B31 OspA. In this vector, the ospA-coding sequence was under transcriptional control of the cytomegalovirus immediate early promoter. For negative and positive controls, mice were immunized with either the plasmid vector without an osp-coding sequence or recombinant OspA protein, respectively. Mice immunized with VR2210 DNA produced OspA-specific antibodies that bound to B. burgdorferi in a whole cell ELISA and inhibited the growth of a homologous strain of B. burgdorferi. Immunization with VR2210 protected mice against challenge with 2 infectious strains of B. burgdorferi, Sh-2-82 and N40. These results indicate that vaccination with plasmid DNA expressing OspA is an efficacious method for providing a protective response against B. burgdorferi infection.

Amino Acid Sequence↗
PubMed1997

Arthritis severity and spirochete burden are determined by serotype in the Borrelia turicatae-mouse model of Lyme disease.

Immunodeficient mice infected with Borrelia turicatae, a relapsing fever agent, have a disorder that resembles disseminated Lyme disease. Two serotypes, A and B, differed in their arthritogenicity in both CB-17 SCID and C3H SCID mice. In CB-17 SCID mice infected with serotype A or B, arthritis was assessed by measurement of tibiotarsal diameter, functional ability on a beam walk test, and microscopic assessment of joint inflammation. Serotype B-infected mice had greater joint swelling, functional disability, and leukocytic infiltration in the joints than serotype A-infected mice. Joint swelling and disability peaked at 2 weeks of infection and then decreased, while leukocyte infiltration in the joints persisted. To investigate the basis for the differences in arthritogenicity of serotypes A and B, spirochete burdens in infected mice were measured by quantitative PCR of spirochete DNA in joints, direct immunofluorescence of spirochetes in joints, and counts of spirochetes in the blood. At 2 weeks of infection there were seven times more spirochetes in the joints of serotype B-infected mice than in those of serotype A-infected mice, measured by both quantitative PCR and direct enumeration. Although serotypes A and B had the same infectivity and growth rate in vivo, serotype B spirochetes were eightfold more abundant in the blood than serotype A spirochetes and produced greater fatality in newborn mice. These findings indicate that differences in disease severity in mice infected with serotype A or B are attributable to differences in the spirochete burden in the joints and blood.

Animals↗
PubMed1997

Immunologic and genetic analyses of VmpA of a neurotropic strain of Borrelia turicatae.

In mice infected with serotype A but not serotype B of the relapsing fever spirochete Borrelia turicatae, early invasion of the brain occurs. Serotypes A and B are further distinguished by the abundant surface protein they produce: VmpA and VmpB, respectively. Western blotting with monoclonal antibodies, one-dimensional peptide mapping, and partial amino acid sequencing demonstrated regions of the VmpA protein that differed from VmpB. Oligonucleotide primers based on the partial amino acid sequences of unique regions were used to amplify a portion of the VmpA gene (vmpA) by PCR, and the product was used as a probe in Southern blot and Northern blot analyses. These experiments showed that (i) expression of the vmpA sequence was determined at the level of transcription and (ii) the vmpA sequence was in two locations in serotype A and one location in serotype B. The vmpA gene at the expression-linked locus of serotype A was cloned and sequenced. An open reading frame would encode a polypeptide of 214 amino acids. The polypeptide expressed by Escherichia coli was bound by VmA-specific but not VmpB-specific antibody. Primer extension analysis identified a consensus sigma70-type promoter for vmpA at the expression locus. Phylogenetic analysis revealed that VmpA is homologous to small Vmp (Vsp) proteins of B. hermsii and to OspC proteins of B. burgdorferi. These findings indicate that a function of the Vsp-OspC family of proteins of Borrelia spp. may be differential localization in organs, including the brain, during infection.

Amino Acid Sequence↗