Genetic linkages in B. burgdorferi | CDC EID

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Volume 16, Number 7–July 2010
Dispatch
Geographic Differences in Genetic Locus Linkages for Borrelia burgdorferi
Bridgit Travinsky, Jonas Bunikis, and Alan G. Barbour
Author affiliation: University of California, Irvine, California, USA

Suggested citation for this article

Abstract
Borrelia burdorferi genotype in the northeastern United States is associated with Lyme borreliosis severity. Analysis of DNA sequences of the outer surface protein C gene and rrs-rrlA intergenic spacer from extracts of Ixodes spp. ticks in 3 US regions showed linkage disequilibrium between the 2 loci within a region but not consistently between regions.
Most bacterial pathogens comprise a variety of strains in various proportions. For Borrelia burgdorferi, an agent of Lyme borreliosis, strains differ in their reservoir host preferences (1), propensities to disseminate in humans (2,3), and prevalences in ticks by geographic area (4,5). Strain identification of B. burgdorferi now is predominantly based on DNA sequences of either of 2 genetic loci: 1) the plasmid-borne, highly polymorphic ospC gene, which encodes outer surface protein C (6,7), or 2) the intergenic spacer (IGS) between the rrs and rrlA rDNA, here called IGS1. Other loci for genotyping are the plasmid-borne ospA gene (7) and the rrfA-rrlB rDNA intergenic spacer, here called IGS2 (8). The apparent clonality of B. burgdorferi was justification for inferring strain identity from a single locus (9,10), but the extent of genomewide genetic exchange in this species may have been underestimated (6).

Given reports of an association between disease severity and B. burgdorferi genotype (2,3), prediction of a strain’s virulence potential from its genotype has clinical, diagnostic, and epidemiologic relevance. But is a single locus sufficient for this assessment?

The Study
To investigate this issue, we determined sequences of ospC and IGS1 loci, and in selected cases the ospA and IGS2 loci, in 1,522 DNA extracts from B. burgdorferi–infected Ixodes scapularis nymphs collected from the northeastern, mid-Atlantic, and north-central United States during the summers of 2004, 2005, 2006, and 2007, as described (4,11). We also included results from 214 infected I. pacificus nymphs collected in Mendocino County, California (5); 20 infected I. pacificus adults from Contra Costa County, California (J. Bunikis and A.G. Barbour, unpub. data); and 10 B. burgdorferi genomes (strains B31, ZS7, 156a, 64b, 72a, 118a, WI91-23, 94a, 29805, and CA-11.2a), for which sequences are publicly available (www.ncbi.nlm.nih.gov). Multilocus sequence typing (MLST), based on 8 chromosomal housekeeping genes, had been carried out for several strains represented in the extracts (Table) (4,12). The corresponding MLST types of the 10 genome sequences were assigned by reference to a B. burgdorferi MLST database (http://borrelia.mlst.net) (12). For this study, we also determined the MLST type of strain CA8.

The methods for 1) DNA extraction from ticks (11), 2) PCR amplification of ospC, ospA, and IGS1 (7), 3) amplification of IGS2 (8), and 4) amplification of 8 chromosomal loci for MLST (12) have been described. Sequences for both strands were determined from either PCR products or cloned fragments with custom primers (7). We followed the basic nomenclature of Wang et al. (13) until, after exhausting the alphabet, we assigned both a letter and, arbitrarily, the number 3 (e.g., C3) when a new nucleotide sequence differed by >8% from known ospC alleles. We distinguished ospC variants with <1% sequence difference by adding a lowercase letter, e.g., Da and Db. Except for ospC D3 and Oa, novel polymorphisms were confirmed in at least 1 other sample. To simplify IGS1 nomenclature, we numbered types sequentially, beginning with the original 9 types (7); ospA alleles (7) and IGS2 loci were likewise sequentially numbered. The Appendix Table provides accession numbers for all sequences, as well as original and revised names for IGS1 sequences.

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Suggested Citation for this Article
Travinsky T, Bunikis J, Barbour AG. Geographic differences in genetic locus linkages for Borrelia burgdorferi. Emerg Infect Dis [serial on the Internet]. 2010 Jul [date cited]. http://www.cdc.gov/EID/content/16/7/1147.htm

DOI: 10.3201/eid1607.091452