Biomedical subjects

Tom G Schwan

Publications and source records attributed to Tom G Schwan.

At least 19 recordsLinked to original sources

PubMed2006

Dual role of MyD88 in rapid clearance of relapsing fever Borrelia spp.

Relapsing fever Borrelia spp. undergo antigenic variation, achieve high levels in blood, and require rapid production of immunoglobulin M (IgM) for clearance. MyD88-deficient mice display defective clearance of many pathogens; however, the IgM response to persistent infection is essentially normal. Therefore, MyD88(-/-) mice provided a unique opportunity to study the effect of nonantibody, innate host defenses to relapsing fever Borrelia. Infected MyD88(-/-) mice harbored extremely high levels of B. hermsii in the blood compared to wild-type littermates. In the comparison of MyD88(-/-) mice and B- and T-cell-deficient scid mice, two features stood out: (i) bacterial numbers in blood were at least 10-fold greater in MyD88(-/-) mice than scid mice, even though the production of IgM still occurred in MyD88(-/-) mice; and (ii) many of the MyD88(-/-) mice were able to exert partial clearance, although with delayed kinetics relative to wild-type mice, a feature not seen in scid mice. Further analysis revealed a delay in the IgM response to lipoproteins expressed by the original inoculum; however, by 6 days of infection antibodies were produced in MyD88(-/-) mice that could clear spirochetemia in scid mice. While these results indicated that the production of IgM was delayed in MyD88(-/-) mice, they also point to a second, antibody-independent role for MyD88 signaling in host defense to relapsing fever Borrelia. This second defect was apparent only when antibody levels were limiting.

Animals↗

PubMed2006

Relapsing fever spirochaetes produce a serine protease that provides resistance to oxidative stress and killing by neutrophils.

The spirochaetes that cause tick-borne relapsing fever and Lyme disease are closely related human pathogens, yet they differ significantly in their ecology and pathogenicity. Genome sequencing of two species of relapsing fever spirochaetes, Borrelia hermsii and Borrelia turicatae, identified a chromosomal open reading frame, designated bhpA, not present in the Lyme disease spirochaete Borrelia burgdorferi. The predicted amino acid sequence of bhpA was homologous with the HtrA serine proteases, which are involved with stress responses and virulence in other bacteria. B. hermsii produced an active serine protease that was recognized by BhpA antibodies and the recombinant BhpA protein-degraded beta-casein. bhpA was transcribed in vitro at all growth temperatures and transcription levels were slightly elevated at higher temperatures. These results correlated with the synthesis of BhpA during B. hermsii infection in mice. With the exception of Borrelia recurrentis, the bhpA gene, protein and enzymatic activity were found in all relapsing fever spirochaetes, but not in Lyme disease or related spirochaetes. Heterologous expression of bhpA in B. burgdorferi increased the spirochaete's resistance to both oxidative stress and killing by human neutrophils. Therefore, we propose that bhpA encodes a unique and functional serine protease in relapsing fever spirochaetes. This periplasmic enzyme may prevent the accumulation of proteins damaged by the innate immune response and contribute to the ability of the relapsing fever spirochaetes to achieve high cell densities in blood.

Animals↗

PubMed2006

Antigenic variation by Borrelia hermsii occurs through recombination between extragenic repetitive elements on linear plasmids.

The relapsing fever agent Borrelia hermsii undergoes multiphasic antigenic variation through gene conversion of a unique expression site on a linear plasmid by an archived variable antigen gene. To further characterize this mechanism we assessed the repertoire and organization of archived variable antigen genes by sequencing approximately 85% of plasmids bearing these genes. Most archived genes shared with the expressed gene a <or= 62 nucleotide (nt) region, the upstream homology sequence (UHS), that surrounded the start codon. The 59 archived variable antigen genes were arrayed in clusters with 13 repetitive, 214 nt long downstream homology sequence (DHS) elements distributed among them. A fourteenth DHS element was downstream of the expression locus. Informative nucleotide polymorphisms in UHS regions and DHS elements were applied to the analysis of the expression site of relapse serotypes from 60 infected mice in a prospective study. For most recombinations, the upstream crossover occurred in the UHS's second half, and the downstream crossover was in the DHS's second half. Usually the closest archival DHS element was used, but occasionally a more distant DHS was employed. The downstream extragenic crossover site in B. hermsii contrasts with the upstream [corrected] extragenic crossover site for antigenic variation in African trypanosomes.

Antigenic Variation↗

PubMed2005

Defining plasmids required by Borrelia burgdorferi for colonization of tick vector Ixodes scapularis (Acari: Ixodidae).

Maintenance in nature of Borrelia burgdorferi, the pathogenic bacterium that causes Lyme disease, requires transmission through an infectious cycle that includes a tick vector and a mammalian host. The genetic requirements for persistence in these disparate environments have not been well defined. B. burgdorferi has a complex genome composed of a chromosome and >20 plasmids. Previous work has demonstrated that B. burgdorferi requires two plasmids, lp25 and lp28-1, in the mammalian host. To investigate the requirement for these same two plasmids during tick infection, we experimentally infected larval ticks with B. burgdorferi lacking either lp25 or lp28-1 and then assessed the spirochete load in ticks at different points of the infection. Whereas plasmid lp28-1 was dispensable in ticks, plasmid lp25 was essential for tick infection. Furthermore, we investigated the requirement in ticks for the lp25 gene bbe22, which encodes a nicotinamidase that is necessary and sufficient for mammalian infection by B. burgdorferi clones lacking lp25. This gene was also sufficient in ticks to restore survival of spirochetes lacking lp25. This is the first study to investigate the requirement for specific plasmids by B. burgdorferi within the tick vector, and it begins to establish the genomic components required for persistence of this pathogen throughout its natural infectious cycle.

Animals↗

PubMed2005

Variable tick protein in two genomic groups of the relapsing fever spirochete Borrelia hermsii in western North America.

Borrelia hermsii is the primary cause of tick-borne relapsing fever in North America. When its tick vector, Ornithodoros hermsi, acquires these spirochetes from the blood of an infected mammal, the bacteria switch their outer surface from one of many bloodstream variable major proteins (Vmps) to a unique protein, Vtp (Vsp33). Vtp may be critical for successful tick transmission of B. hermsii; however, the gene encoding this protein has been described previously in only one isolate. Here we identified and sequenced the vtp gene in 31 isolates of B. hermsii collected over 40 years from localities throughout much of its known geographic distribution. Seven major Vtp types were found. Little or no sequence variation existed within types, but between them significant variation was observed, similar to the pattern of diversity described for the outer surface protein C (OspC) gene in Lyme disease spirochetes. The pattern of sequence relatedness among the Vtp types was incongruent in two branches compared to two genomic groups identified among the isolates by multilocus sequence typing of the 16S rRNA, flaB, gyrB, and glpQ genes. Therefore, both horizontal transfer and recombination within and between the two genomic groups were responsible for some of the variation observed in the vtp gene. O. hermsi ticks were capable of transmitting spirochetes in the newly identified genomic group. Therefore, given the longevity of the tick vector and persistent infection of spirochetes in ticks, these arthropods rather than mammals may be the likely host where the exchange of spirochetal DNA occurs.

Amino Acid Sequence↗

PubMed2005

Relapsing fever spirochetes contain chromosomal genes with unique direct tandemly repeated sequences.

Genome sequencing of the relapsing fever spirochetes Borrelia hermsii and Borrelia turicatae identified three open reading frames (ORFs) on the chromosomes that contained internal, tandemly repeated amino acid sequences that were absent in the Lyme disease spirochete Borrelia burgdorferi. The predicted amino acid sequences of these genes (BH0209, BH0512, and BH0553) have hydrophobic N termini, indicating that these proteins may be secreted. B. hermsii transcribed the three ORFs in vitro, and the BH0512- and BH0553-encoded proteins (PBH-512 and PBH-553) were produced in vitro and in experimentally infected mice. PBH-512 and PBH-553 were on the spirochete's outer surface, and antiserum to these proteins reduced the adherence of B. hermsii to red blood cells. PCR analyses of 28 isolates of B. hermsii and 8 isolates of B. turicatae demonstrated polymorphism in each gene correlated with the number of repeats. Serum samples from relapsing fever patients reacted with recombinant PBH-512 and PBH-553, suggesting that these proteins are produced during human infection. These polymorphic proteins may be involved in the pathogenicity of these relapsing fever spirochetes and provide a mechanism for antigenic heterogeneity within their populations.

Amino Acid Sequence↗

PubMed2005

Phylogenetic analysis of the spirochetes Borrelia parkeri and Borrelia turicatae and the potential for tick-borne relapsing fever in Florida.

Isolates of Borrelia turicatae, Borrelia parkeri, and the Florida canine borrelia (FCB) were examined to further phylogenetically characterize the identities of these spirochetes in the United States. DNA sequences of four chromosomal loci (the 16S rRNA gene, flaB, gyrB, and glpQ) were determined for eight isolates of B. turicatae and six isolates of B. parkeri, which grouped the spirochetes into two distinct but closely related taxa (>98% sequence identity) separate from Borrelia hermsii. The FCB was clearly separated with the group identified as B. turicatae, confirming this bacterium as a relapsing fever spirochete. Therefore, the potential for tick-borne relapsing fever in humans and other animals exists in Florida and future efforts are needed to determine the enzootic hosts and distribution of this spirochete in the southeastern United States. Analysis of plasmids demonstrated both linear and circular forms in B. turicatae but only linear plasmids in B. parkeri, which should be of interest to investigators concerned with plasmid diversity and evolution within this group of spirochetes.

Bacterial Proteins↗

PubMed2004

Outer-surface protein C of the Lyme disease spirochete: a protein induced in ticks for infection of mammals.

Environmentally responsive synthesis of surface proteins represents a hallmark of the infectious cycle of the Lyme disease agent, Borrelia burgdorferi. Here we created and analyzed a B. burgdorferi mutant lacking outer-surface protein C (OspC), an abundant Osp that spirochetes normally synthesize in the tick vector during the blood meal and down-regulate after transmission to the mammal. We demonstrate that B. burgdorferi strictly requires OspC to infect mice but not to localize or migrate appropriately in the tick. The induction of a spirochetal virulence factor preceding the time and host in which it is required demonstrates a developmental sequence for transmission of this arthropod-borne pathogen.

Animals↗

PubMed2004

Isolation and characterization of Borrelia hermsii associated with two foci of tick-borne relapsing fever in California.

Relapsing fever, caused by the spirochete Borrelia hermsii and transmitted by the soft tick Ornithodoros hermsi, is endemic in many rural mountainous areas of California. Between 1996 and 1998, 12 cases of relapsing fever associated with two exposure sites in northern California were investigated. Follow-up at exposure sites included collection of soft ticks and serum specimens from sylvatic rodents. Attempts to cultivate spirochetes were made through inoculation of patient blood into mice and by feeding Ornithodoros ticks on mice. Three isolates of B. hermsii were recovered from two blood specimens and one pool of ticks. The protein and plasmid profiles of the three isolates were comparable to those of previous B. hermsii isolates from the western United States. Western immunoblotting of patient sera demonstrated an expanding immunologic response to antigens within four distinct molecular weight regions by 3 to 4 weeks postonset. Antibody to B. hermsii was detected in sera from 4 of 11 yellow-pine chipmunks (Tamias amoenus); no other rodent species collected were seropositive.

Adult↗

PubMed2004

Glycerophosphodiester phosphodiesterase gene (glpQ) of Borrelia lonestari identified as a target for differentiating Borrelia species associated with hard ticks (Acari:Ixodidae).

A glpQ ortholog was identified in DNA from Borrelia lonestari-positive Amblyomma americanum, providing further evidence that B. lonestari is more closely related to the relapsing fever group spirochetes than to borreliae that cause Lyme disease. This finding provides a basis for developing diagnostic assays to differentiate species of borrelia transmitted by hard ticks.

Amino Acid Sequence↗

PubMed2003

Bacterial pathogens modulate an apoptosis differentiation program in human neutrophils.

Human polymorphonuclear leukocytes (PMNs or neutrophils) are essential to the innate immune response against bacterial pathogens. Recent evidence suggests that PMN apoptosis facilitates resolution of inflammation during bacterial infection. Although progress has been made toward understanding apoptosis in neutrophils, very little is known about transcriptional regulation of this process during bacterial infection. To gain insight into the molecular processes that facilitate resolution of infection, we measured global changes in PMN gene expression during phagocytosis of a diverse group of bacterial pathogens. Genes encoding key effectors of apoptosis were up-regulated, and receptors critical to innate immune function were down-regulated during apoptosis induced by phagocytosis of Burkholderia cepacia, Borrelia hermsii, Listeria monocytogenes, Staphylococcus aureus, and Streptococcus pyogenes. Importantly, we identified genes that comprise a common apoptosis differentiation program in human PMNs after phagocytosis of pathogenic bacteria. Unexpectedly, phagocytosis of Str. pyogenes induced changes in neutrophil gene expression not observed with other pathogens tested, including down-regulation of 21 genes involved in responses to IFN. Compared with other bacteria, PMN apoptosis was significantly accelerated by Str. pyogenes and was followed by necrosis. Thus, we hypothesize that there are two fundamental outcomes for the interaction of bacterial pathogens with neutrophils: (i) phagocytosis of bacteria induces an apoptosis differentiation program in human PMNs that contributes to resolution of bacterial infection, or (ii) phagocytosis of microorganisms such as Str. pyogenes alters the apoptosis differentiation program in neutrophils, resulting in pathogen survival and disease.

Apoptosis↗

PubMed2003

Spirochete-platelet attachment and thrombocytopenia in murine relapsing fever borreliosis.

Thrombocytopenia is common in persons infected with relapsing fever Borreliae. We previously showed that the relapsing fever spirochete Borrelia hermsii binds to and activates human platelets in vitro and that, after platelet activation, high-level spirochete-platelet attachment is mediated by integrin alpha IIb beta 3, a receptor that requires platelet activation for full function. Here we established that B hermsii infection of the mouse results in severe thrombocytopenia and a functional defect in hemostasis caused by accelerated platelet loss. Disseminated intravascular coagulation, immune thrombocytopenic purpura, or splenic sequestration did not play a discernible role in this model. Instead, spirochete-platelet complexes were detected in the blood of infected mice, suggesting that platelet attachment by bacteria might result in platelet clearance. Consistent with this, splenomegaly and thrombocytopenia temporally correlated with spirochetemia, and the severity of thrombocytopenia directly correlated with the degree of spirochetemia. Activation of platelets and integrin alpha IIb beta 3 were apparently not required for bacterium-platelet binding or platelet clearance because the bacterium-bound platelets in the circulation were not activated, and platelet binding and thrombocytopenia during infection of beta 3-deficient and wild-type mice were indistinguishable. These findings suggest that thrombocytopenia of relapsing fever is the result of platelet clearance after beta 3-independent bacterial attachment to circulating platelets.

Animals↗

PubMed2003

Regulation of expression of the Borrelia burgdorferi beta(3)-chain integrin ligand, P66, in ticks and in culture.

Borrelia burgdorferi is maintained in an infection cycle between mammalian and arthropod hosts. Appropriate gene expression by B. burgdorferi at different stages of this cycle is probably essential for transmission and establishment of infection. The B. burgdorferi beta(3) integrin ligand P66 is expressed by the bacteria in mammals, laboratory culture, and engorged but not unfed ticks. No in vitro culture conditions in which P66 expression reflected that in the unfed tick were found, suggesting that there are aspects of B. burgdorferi-tick interaction that remain unexplored.

Animals↗

PubMed2003

Glycerol-3-phosphate acquisition in spirochetes: distribution and biological activity of glycerophosphodiester phosphodiesterase (GlpQ) among Borrelia species.

Relapsing-fever spirochetes achieve high cell densities (>10(8)/ml) in their host's blood, while Lyme disease spirochetes do not (<10(5)/ml). This striking contrast in pathogenicity of these two groups of bacteria suggests a fundamental difference in their ability to either exploit or survive in blood. Borrelia hermsii, a tick-borne relapsing-fever spirochete, contains orthologs to glpQ and glpT, genes that encode glycerophosphodiester phosphodiesterase (GlpQ) and glycerol-3-phosphate transporter (GlpT), respectively. In other bacteria, GlpQ hydrolyzes deacylated phospholipids to glycerol-3-phosphate (G3P) while GlpT transports G3P into the cytoplasm. Enzyme assays on 17 isolates of borreliae demonstrated GlpQ activity in relapsing-fever spirochetes but not in Lyme disease spirochetes. Southern blots demonstrated glpQ and glpT in all relapsing-fever spirochetes but not in the Lyme disease group. A Lyme disease spirochete, Borrelia burgdorferi, that was transformed with a shuttle vector containing glpTQ from B. hermsii produced active enzyme, which demonstrated the association of glpQ with the hydrolysis of phospholipids. Sequence analysis of B. hermsii identified glpF, glpK, and glpA, which encode the glycerol facilitator, glycerol kinase, and glycerol-3-phosphate dehydrogenase, respectively, all of which are present in B. burgdorferi. All spirochetes examined had gpsA, which encodes the enzyme that reduces dihydroxyacetone phosphate (DHAP) to G3P. Consequently, three pathways for the acquisition of G3P exist among borreliae: (i) hydrolysis of deacylated phospholipids, (ii) reduction of DHAP, and (iii) uptake and phosphorylation of glycerol. The unique ability of relapsing-fever spirochetes to hydrolyze phospholipids may contribute to their higher cell densities in blood than those of Lyme disease spirochetes.

Animals↗

PubMed2003

Experimental infection of Ixodes scapularis larvae (Acari: Ixodidae) by immersion in low passage cultures of Borrelia burgdorferi.

We describe a procedure for the introduction of Borrelia burgdorferi, the spirochetal agent of Lyme disease, into larvae of the tick vector Ixodes scapularis. Internalized spirochetes were observed in larvae examined after 15 or 45 min immersion at 32 degrees C in liquid culture suspensions of low passage B. burgdorferi strain B31. Larval ticks immersed in low passage strain B31 were able to feed to repletion on white-footed mice. Midguts of larvae contained many spirochetes 1 wk postengorgement, while larvae incubated with high passage strain B31 were free of detectable spirochetes at the same interval. Larvae incubated with low passage strain B31 were competent to transmit the pathogen to mice, as shown by serology, reisolation of B. burgdorferi from mice, and xenodiagnosis. Ticks maintained the infection transstadially to the nymphal stage and transmitted the infection to naive mice, replicating an essential aspect of natural infection. This method requires no special equipment and allows artificial infection of large numbers of ticks at the larval stage. It will facilitate studies of the contribution of specific B. burgdorferi genetic loci to tick colonization.

Animals↗

PubMed2003

Tick-borne relapsing fever caused by Borrelia hermsii, Montana.

Five persons contracted tick-borne relapsing fever after staying in a cabin in western Montana. Borrelia hermsii was isolated from the blood of two patients, and Ornithodoros hermsi ticks were collected from the cabin, the first demonstration of this bacterium and tick in Montana. Relapsing fever should be considered when patients who reside or have vacationed in western Montana exhibit a recurring febrile illness.

Adolescent↗

PubMed2002

Population dynamics of a heterogeneous Borrelia burgdorferi B31 strain in an experimental mouse-tick infectious cycle.

We have recently shown that low-passage, infectious Borrelia burgdorferi strain B31 MI is a heterogeneous mixture of clones varying in colony morphology, growth rate, protein profiles, plasmid content and infectivity. In this study, we asked whether there is a selection for certain clonal populations during the infectious cycle when uncloned B31 MI is used as the starting strain. B31 MI derivatives were reisolated from various tissues of two mice after completion of a mouse-tick-mouse infectious cycle and their protein and plasmid profiles were analyzed. Both mice developed ostensibly clonal infections despite the fact that the infectious cycle was started with a heterogeneous strain. Moreover, the mice became infected with two different clones varying in protein profile and growth phenotype. Comparison of the mouse reisolates to uncloned B31 MI and clonal variants derived from B31 MI before mouse-tick-mouse passage suggests that they were derived from clonal populations present in the uncloned B31 MI. Our results indicate the presence of at least two distinct populations within B31 MI that are competent to complete an experimental mouse-tick infectious cycle. The study provides insight into infectivity profiles and infection dynamics of different clonal populations present in a low-passage, infectious B. burgdorferi strain.

Animals↗

PubMed2002

Role of Yersinia murine toxin in survival of Yersinia pestis in the midgut of the flea vector.

Transmission by flea bite is a relatively recent adaptation that distinguishes Yersinia pestis, the plague bacillus, from closely related enteric bacteria. Here, a plasmid-encoded phospholipase D (PLD), previously characterized as Yersinia murine toxin (Ymt), was shown to be required for survival of Y. pestis in the midgut of its principal vector, the rat flea Xenopsylla cheopis. Intracellular PLD activity appeared to protect Y. pestis from a cytotoxic digestion product of blood plasma in the flea gut. By enabling colonization of the flea midgut, acquisition of this PLD may have precipitated the transition of Y. pestis to obligate arthropod-borne transmission.

Animals↗