Endogenous Linear Plasmids lp28-4 and lp25 are Required for Infectivity and Restriction Protection in the Lyme Disease Spirochete Borrelia mayonii
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ABSTRACT:
ORGANISM(S): Borreliella mayonii
PROVIDER: GSE220655 | GEO | 2023/02/13
REPOSITORIES: GEO
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Project description:Borrelia mayonii is a newly recognized causative agent of Lyme disease in the Upper Midwestern United States, with distinct clinical presentations compared to classical Lyme disease caused by other Lyme Borrelia species. However, little is known about the B. mayonii genetic determinants required for establishing infection or perpetuating disease in mammals. Extrachromosomal plasmids in Borrelia species often encode proteins necessary for infection and pathogenesis, and spontaneous loss of these plasmids can lead to the identification of virulence determinant genes. Here, we describe infection of Lyme disease-susceptible C3H mice with B. mayonii, and show bacterial dissemination and persistence in peripheral tissues. Loss of endogenous plasmids, including lp28-4, lp25, and lp36 correlated with reduced infectivity in mice. The apparent requirement for lp28-4 during murine infection suggests the presence of a novel virulence determinant, as this plasmid does not encode homologs of any known virulence determinant. We also describe transformation and stable maintenance of a self-replicating shuttle vector in B. mayonii, and show that loss of either lp25 or lp28-4 correlated with increased transformation competency. Finally, we demonstrate that linear plasmids lp25 and lp28-4 each encode functional restriction modification systems with distinct but partially overlapping target modification sequences, which likely accounts for the observed decrease in transformation efficiency when those plasmids are present. Taken together, this study describes a role for endogenous plasmids in mammalian infection and restriction protection in the Lyme disease spirochete Borrelia mayonii.
| S-EPMC10016076 | biostudies-literature
Project description:Single Molecule Real Time (SMRT) sequencing was utilized to map the genome-wide DNA methylation pattern, and to identify consensus modified motifs in B. burgdorferi as well as isogenic methyltransferase mutants. Four conserved m6A modification motifs were identified, and were fully attributable to the presence of specific methyltransferases. Next, RNA-seq was compared between strains to determine the effects of altering global DNA methylation patterns on gene expression. Whole-genome transcriptome changes were observed in conjunction with the loss of methyltransferase enzymes, indicating that DNA methylation by restriction modification systems has effects on gene expression in B. burgdorferi.
2018-06-05 | GSE115308 | GEO
Project description:Borrelia hermsii, an agent of tick-borne relapsing fever, was found to contain multiple circular plasmids approximately 30 kb in size. Sequencing of a DNA library constructed from circular plasmid fragments enabled assembly of a composite DNA sequence that is homologous to the cp32 plasmid family of the Lyme disease spirochete, B. burgdorferi. Analysis of another relapsing fever bacterium, B. parkeri, indicated that it contains linear homologs of the B. hermsii and B. burgdorferi cp32 plasmids. The B. hermsii cp32 plasmids encode homologs of the B. burgdorferi Mlp and Bdr antigenic proteins and BlyA/BlyB putative hemolysins, but homologs of B. burgdorferi erp genes were absent. Immunoblot analyses demonstrated that relapsing fever patients produced antibodies to Mlp proteins, indicating that those proteins are synthesized by the spirochetes during human infection. Conservation of cp32-encoded genes in different Borrelia species suggests that their protein products serve functions essential to both relapsing fever and Lyme disease spirochetes. Relapsing fever borreliae replicate to high levels in the blood of infected animals, permitting direct detection and possible functional studies of Mlp, Bdr, BlyA/BlyB, and other cp32-encoded proteins in vivo.
| S-EPMC101665 | biostudies-literature
Project description:To determine whether the Lyme disease spirochete Borrelia lusitaniae is associated with lizards, we compared the prevalence and genospecies of spirochetes present in rodent- and lizard-associated ticks at a site where this spirochete frequently infects questing ticks. Whereas questing nymphal Ixodes ricinus ticks were infected mainly by Borrelia afzelii, one-half of the infected adult ticks harbored B. lusitaniae at our study site. Lyme disease spirochetes were more prevalent in sand lizards (Lacerta agilis) and common wall lizards (Podarcis muralis) than in small rodents. Although subadult ticks feeding on rodents acquired mainly B. afzelii, subadult ticks feeding on lizards became infected by B. lusitaniae. Genetic analysis confirmed that the spirochetes isolated from ticks feeding on lizards are members of the B. lusitaniae genospecies and resemble type strain PotiB2. At our central European study site, lizards, which were previously considered zooprophylactic for the agent of Lyme disease, appear to perpetuate B. lusitaniae.
| S-EPMC1489336 | biostudies-literature
Project description:Defining the metabolic capabilities and regulatory mechanisms controlling gene expression is a valuable step in understanding the pathogenic properties of infectious agents such as Borrelia burgdorferi. The present studies demonstrated that B. burgdorferi encodes functional Pfs and LuxS enzymes for the breakdown of toxic products of methylation reactions. Consistent with those observations, B. burgdorferi was shown to synthesize the end product 4,5-dihydroxy-2,3-pentanedione (DPD) during laboratory cultivation. DPD undergoes spontaneous rearrangements to produce a class of pheromones collectively named autoinducer 2 (AI-2). Addition of in vitro-synthesized DPD to cultured B. burgdorferi resulted in differential expression of a distinct subset of proteins, including the outer surface lipoprotein VlsE. Although many bacteria can utilize the other LuxS product, homocysteine, for regeneration of methionine, B. burgdorferi was found to lack such ability. It is hypothesized that B. burgdorferi produces LuxS for the express purpose of synthesizing DPD and utilizes a form of that molecule as an AI-2 pheromone to control gene expression.
| S-EPMC1082833 | biostudies-literature
Project description:The genome of the Lyme disease pathogen Borrelia burgdorferi contains about a dozen linear DNA molecules that carry covalently closed hairpin telomeres as a specialized mechanism for dealing with the end-replication problem. The hairpin telomeres are generated from replicative intermediates through a two-step transesterification promoted by the telomere resolvase ResT. Although the genome of B. burgdorferi has been sequenced, the sequence of most telomeres has remained unknown because of difficulties in recovering and completely sequencing the covalently closed hairpin ends. In this study we report a new approach for the direct sequencing Borrelia telomeres and report the sequence, characterization, and in vitro reaction properties of 19 unique telomeres. Surprisingly, a variation of greater than 160-fold in the initial reaction rates of in vitro ResT-mediated telomere resolution was observed between the most active and least active telomeres. Moreover, three of the hairpin telomeres were completely inactive in vitro, but their in vivo functionality was demonstrated. Our results provide important new information on the structure and function of the B. burgdorferi telomeres and suggest the possibility that factors besides the telomere resolvase ResT may influence the reaction in vivo and rescue those telomeres that are not functional in vitro with ResT alone.
| S-EPMC2652300 | biostudies-literature
Project description:Lyme disease is the most common vector-borne disease in the United States. The causative agent is the spirochete Borrelia burgdorferi. The copy number and organization of the genes encoding the rRNAs of this organism were determined. There is a single gene for 16S rRNA and two copies each of the 23S rRNA and 5S rRNA genes. All of the genes are located within a chromosomal fragment of approximately 9.5 to 10.0 kb. The 23S and 5S rRNA genes are tandemly duplicated in the order 23S-5S-23S-5S and are apparently not linked to the 16S rRNA gene, which is situated over 2 kb upstream from the 23S-5S duplication. The individual copies of the 23S-5S duplication are separated by a 182-bp spacer. Within each 23S-5S unit, an identical 22-bp spacer separates the 23S and 5S rRNA sequences from each other. The genome organization of the 23S-5S gene cluster in a number of different B. burgdorferi isolates obtained at a number of different geographical locations, as well as in several other species of Borrelia, was investigated. All isolates of B. burgdorferi tested displayed the tandem duplication, whereas the closely related species B. hermsii, B. anserina, and B. turicatae all contained a single copy of each of the genes. In addition, different geographical isolates of B. burgdorferi can be differentiated on the basis of a restriction fragment length polymorphism associated with the 23S-5S gene cluster. This polymorphism can be a useful tool for the determination of genetic relatedness between different isolates of B. burgdorferi.
| S-EPMC206066 | biostudies-other
Project description:Six bacterial genomes, Geobacter metallireducens GS-15, Chromohalobacter salexigens, Vibrio breoganii 1C-10, Bacillus cereus ATCC 10987, Campylobacter jejuni subsp. jejuni 81-176 and Campylobacter jejuni NCTC 11168, all of which had previously been sequenced using other platforms were re-sequenced using single-molecule, real-time (SMRT) sequencing specifically to analyze their methylomes. In every case a number of new N6-methyladenine (m6A) and N4-methylcytosine (m4C) methylation patterns were discovered and the DNA methyltransferases (MTases) responsible for those methylation patterns were assigned. In 15 cases it was possible to match MTase genes with MTase recognition sequences without further sub-cloning. Two Type I restriction systems required sub-cloning to differentiate their recognition sequences, while four MTases genes that were not expressed in the native organism were sub-cloned to test for viability and recognition sequences. No attempt was made to detect 5-methylcytosine (m5C) recognition motifs from the SMRT sequencing data because this modification produces weaker signals using current methods. However, all predicted m6A and m4C MTases were detected unambiguously. This study shows that the addition of SMRT sequencing to traditional sequencing approaches gives a wealth of useful functional information about a genome showing not only which MTase genes are active, but also revealing their recognition sequences. Examination of the methylomes of six different strains of bacteria using kinetic data from single-molecule, real-time (SMRT) sequencing on the PacBio RS.
2012-10-04 | E-GEOD-40133 | biostudies-arrayexpress
Project description:Endogenous Linear Plasmids lp28-4 and lp25 are Required for Infectivity and Restriction Protection in the Lyme Disease Spirochete Borrelia mayonii
| PRJNA910814 | ENA
Project description:The pathogenic spirochete Borrelia burgdorferi senses and responds to changes in the environment, including changes in nutrient availability, throughout its enzootic cycle in Ixodes ticks and vertebrate hosts. This study examined the role of DnaK suppressor protein (DksA) in the transcriptional response of B. burgdorferi to starvation. Wild-type and dksA mutant B. burgdorferi strains were subjected to starvation by shifting cultures grown in rich complete medium, Barbour-Stoenner-Kelly II (BSK II) medium, to a defined mammalian tissue culture medium, RPMI 1640, for 6?h under microaerobic conditions (5% CO2, 3% O2). Microarray analyses of wild-type B. burgdorferi revealed that genes encoding flagellar components, ribosomal proteins, and DNA replication machinery were downregulated in response to starvation. DksA mediated transcriptomic responses to starvation in B. burgdorferi, as the dksA-deficient strain differentially expressed only 47 genes in response to starvation compared to the 500 genes differentially expressed in wild-type strains. Consistent with a role for DksA in the starvation response of B. burgdorferi, fewer CFU of dksA mutants were observed after prolonged starvation in RPMI 1640 medium than CFU of wild-type B. burgdorferi spirochetes. Transcriptomic analyses revealed a partial overlap between the DksA regulon and the regulon of RelBbu, the guanosine tetraphosphate and guanosine pentaphosphate [(p)ppGpp] synthetase that controls the stringent response; the DksA regulon also included many plasmid-borne genes. Additionally, the dksA mutant exhibited constitutively elevated (p)ppGpp levels compared to those of the wild-type strain, implying a regulatory relationship between DksA and (p)ppGpp. Together, these data indicate that DksA, along with (p)ppGpp, directs the stringent response to effect B. burgdorferi adaptation to its environment.IMPORTANCE The Lyme disease bacterium Borrelia burgdorferi survives diverse environmental challenges as it cycles between its tick vectors and various vertebrate hosts. B. burgdorferi must withstand prolonged periods of starvation while it resides in unfed Ixodes ticks. In this study, the regulatory protein DksA is shown to play a pivotal role controlling the transcriptional responses of B. burgdorferi to starvation. The results suggest that DksA gene regulatory activity impacts B. burgdorferi metabolism, virulence gene expression, and the ability of this bacterium to complete its natural life cycle.
| S-EPMC6351744 | biostudies-literature