Phylogenetic Analysis of a Virulent Borrelia Species Isolated from Patients with Relapsing Fever

Abstract

Multilocus sequence analysis (MLSA) was used to clarify the taxonomic status of a virulent Borrelia organism previously isolated from patients with relapsing fever and from ticks in Spain that is designated the Spanish relapsing fever (SRF) Borrelia. This species has been used extensively in experimental infection models because of its continued virulence. Seven genes were amplified to analyze the phylogenetic relationships among several Spanish isolates of SRF Borrelia and other relapsing fever Borrelia species. The genes targeted in this study included rrs and flaB, which have commonly been used in phylogenetic studies; the rrf-rrl intergenic spacer (IGS), which is highly discriminatory; and four additional genes, p66, groEL, glpQ, and recC, which are located on the chromosome and which have therefore evolved in a clonal way. The species included in this study were Borrelia duttonii, B. recurrentis, B. crocidurae, and B. hispanica as Old World Borrelia species and B. turicatae and B. hermsii as New World Borrelia species. The results obtained by MLSA of the SRF Borrelia on the basis of 1% of the genomic sequence data analyzed confirmed that the SRF Borrelia isolates are B. hispanica. However, the prototype isolates of B. hispanica used in this study have an uncertain history and display unique phenotypic characteristics that are not shared with the SRF Borrelia. Therefore, we propose to use strain SP1, isolated from a relapsing fever patient in 1994 in southern Spain, as the type strain for B. hispanica.The members of the Borrelia genus comprise two major groups: those causing Lyme disease and those causing relapsing fever (RF). Relapsing fever is distributed all over the world; and its agents are traditionally classified according to their geographic origins, vector, and infectivity in various animal species. In Spain, relapsing fever has been reported sporadically throughout the last century and has been associated with Borrelia hispanica (2,11,17,19,27,28). From 1994 to 1996, spirochetes were isolated from the blood of two patients with RF symptoms and from Ornithodoros erraticus, and we have designated these spirochetes Spanish relapsing fever (SRF) Borrelia. This SRF organism is refractory to in vitro cultivation, and this represents a unique phenotypic characteristic not shared by B. hispanica (1). SRF Borrelia infects C3H/ HeN, BALB/c, C57/B51, and Swiss outbred mice (1), and it is highly infectious. A murine model of SRF was developed (13) and has been used extensively in experimental studies on pathogenesis and the host response (5-7, 14, 16, 20). A preliminary genetic analysis comparing the rrs and flaB loci from B. hispanica to those from the Spanish isolates suggested that SRF could represent a new species (1).To date, most of the ecological and epidemiological studies on relapsing fever Borrelia have used a single locus. Therefore, characterization of relapsing fever spirochetes has relied upon the amplification and sequencing of certain genes, such as rrs (24), flaB (12), and more recently, the n...

Complete Genome Sequence of Borrelia crocidurae

“…The B. crocidurae Achema strain (11), grown in BarbourStoenner-Kelly H medium (Sigma, Saint-Quentin-Fallavier, France) supplemented with 6% rabbit serum (Eurobio, Courtaboeuf, France) at 33°C, was sequenced using pyrosequencing technology on a Roche 454 GS FLX sequencer (Roche, BoulogneBillancourt, France). An average 24-fold genomic coverage was obtained over 22,610 paired-end reads, which were assembled using the Newbler 2.3 assembler (Roche), generating 41 scaffolds.…”

Phylogenetic Analysis of the Spirochete Borrelia microti, a Potential Agent of Relapsing Fever in Iran

“…Concatenated coding genes can be used to draw phylogenetic inference, while IGS, being noncoding, provides a more discriminatory typing tool. Our findings revealed a high degree of homology between B. microti and isolates of B. duttonii and B. recurrentis, while typing of IGS proved to be more discriminatory than typing of other loci but could show overlap between subgroups of B. duttonii and B. recurrentis (7,25,28). The greatest nucleotide homology between B. microti and B. duttonii over the loci analyzed ( Fig.…”

“…suggests that it may be maintained by enzootic cycles that involve rodents and other small mammals (12,13) and that thus resemble the life cycle of B. crocidurae (3,5,29). Interestingly, the isolate of B. duttonii, which showed the greatest homology with our B. microti isolate, was derived from O. erraticus ticks in West Africa (strain CR2A), initially being classified as B. crocidurae rather than B. duttonii (28). It is tempting to speculate whether such as yet undisclosed links hold the key to the evolutionary origins of these isolates.…”

Rapid Identification of Borrelia by High Resolution Melting Analysis of the groEL Gene

“…was differentiated as a distinct entity. The usefulness of the groEL gene in differentiating relapsing fever group borreliae and establishing new species was shown by TOLEDO et al (2010).…”