title: Culturing selects for specific genotypes of Borrelia burgdorferi in an enzootic cycle in Colorado.
abstract: In Colorado, Borrelia burgdorferi sensu stricto, the etiologic agent of Lyme disease, is maintained in an enzootic cycle between Ixodes spinipalpis ticks and Neotoma mexicana rats (27). The frequencies of flagellin (fla), 66-kDa protein (p66), and outer surface protein A (ospA) alleles were examined in 71 B. burgdorferi isolates from samples from Colorado. Approximately two-thirds of these samples were isolates from I. spinipalpis ticks that had been cultured in BSK-H medium prior to DNA extraction. The remaining samples were from total DNA extracted directly from infected I. spinipalpis ticks. A portion of each gene was amplified by PCR and screened for genetic variability by single-strand conformation polymorphism (SSCP) analysis. We identified three alleles in the fla gene, seven in the p66 gene, and seven in the ospA gene. Sequencing verified that the amplified products originated from B. burgdorferi template DNA and indicated 100% sensitivity and specificity of the SSCP analysis. The frequencies of the p66 and ospA alleles were significantly different between cultured and uncultured spirochetes. The number of three-locus genotypes and the genetic diversity of alleles at all loci were consistently lower in cultured spirochetes, suggesting that culturing of B. burgdorferi in BSK-H medium may select for specific genotypes.
authors.affiliation: Department of Microbiology, Colorado State University, Ft. Collins 80523, USA.
authors.forename: D E
authors.initials: DE
authors.lastname: Norris
authors.forename: B J
authors.initials: BJ
authors.lastname: Johnson
authors.forename: J
authors.initials: J
authors.lastname: Piesman
authors.forename: G O
authors.initials: GO
authors.lastname: Maupin
authors.forename: J L
authors.initials: JL
authors.lastname: Clark
authors.forename: W C
authors.initials: WC
authors.lastname: Black
mesh_terms: D000483:Alleles; D000818:Animals; D000954:Antigens, Surface / Q000235:genetics*; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics*; D001428:Bacterial Vaccines; D015748:Borrelia burgdorferi Group / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification*; D003120:Colorado / Q000453:epidemiology; D004269:DNA, Bacterial / Q000235:genetics; D005408:Flagellin / Q000235:genetics*; D005798:Genes, Bacterial; D014644:Genetic Variation; D018884:Ixodes / Q000382:microbiology*; D008074:Lipoproteins; D008193:Lyme Disease / Q000453:epidemiology / Q000235:genetics*; D017720:Molecular Epidemiology; D010802:Phylogeny; D016133:Polymerase Chain Reaction; D018807:Polymorphism, Single-Stranded Conformational; D012680:Sensitivity and Specificity; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA
chemical_list: D000954:Antigens, Surface; D001425:Bacterial Outer Membrane Proteins; D001428:Bacterial Vaccines; D004269:DNA, Bacterial; D008074:Lipoproteins; C072045:OspA protein; D005408:Flagellin