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, 3 (12), e4002

Fine-scale Phylogeographic Structure of Borrelia Lusitaniae Revealed by Multilocus Sequence Typing

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Fine-scale Phylogeographic Structure of Borrelia Lusitaniae Revealed by Multilocus Sequence Typing

Liliana R Vitorino et al. PLoS One.

Abstract

Borrelia lusitaniae is an Old World species of the Lyme borreliosis (LB) group of tick-borne spirochetes and prevails mainly in countries around the Mediterranean Basin. Lizards of the family Lacertidae have been identified as reservoir hosts of B. lusitaniae. These reptiles are highly structured geographically, indicating limited migration. In order to examine whether host geographic structure shapes the evolution and epidemiology of B. lusitaniae, we analyzed the phylogeographic population structure of this tick-borne bacterium using a recently developed multilocus sequence typing (MLST) scheme based on chromosomal housekeeping genes. A total of 2,099 questing nymphal and adult Ixodes ricinus ticks were collected in two climatically different regions of Portugal, being approximately 130 km apart. All ticks were screened for spirochetes by direct PCR. Attempts to isolate strains yielded 16 cultures of B. lusitaniae in total. Uncontaminated cultures as well as infected ticks were included in this study. The results using MLST show that the regional B. lusitaniae populations constitute genetically distinct populations. In contrast, no clear phylogeographic signals were detected in sequences of the commonly used molecular markers ospA and ospC. The pronounced population structure of B. lusitaniae over a short geographic distance as captured by MLST of the housekeeping genes suggests that the migration rates of B. lusitaniae are rather low, most likely because the distribution of mediterranean lizard populations is highly parapatric. The study underlines the importance of vertebrate hosts in the geographic spread of tick-borne microparasites.

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1
Figure 1. Bayesian phylogenetic tree of B. lusitaniae using concatenated sequences of clpA, clpX, nifS, pepX, pyrG, recG, rplB.
The tree was rooted with B. burgdorferi strain B31. Posterior probabilities values are indicated to provide branch support. The scale bar represents 1% sequence divergence. B. lusitaniae samples derived from Grândola are highlighted.
Figure 2
Figure 2. Bayesian phylogenetic tree of B. lusitaniae strains based on the IGS.
The tree was rooted with B. burgdorferi strain B31. Posterior probabilities values are indicated to provide branch support. The scale bar represents 5% sequence divergence. B. lusitaniae samples derived from Grândola are highlighted.
Figure 3
Figure 3. Bayesian phylogenetic tree of B. lusitaniae strains based on ospA.
The tree was rooted with B. burgdorferi strain B31. Posterior probabilities values are indicated to provide branch support. The scale bar represents 1% sequence divergence. B. lusitaniae samples derived from Grândola are highlighted.
Figure 4
Figure 4. Bayesian phylogenetic tree of B. lusitaniae strains based on ospC.
The tree was rooted with B. burgdorferi strain B31. Posterior probabilities values are indicated to provide branch support. The scale bar represents 5% sequence divergence. B. lusitaniae samples derived from Grândola are highlighted.
Figure 5
Figure 5. Distribution of pairwise genetic distance for B. lusitaniae housekeeping genes (A) and ospC (B).
Figure 6
Figure 6. Variable sites for each housekeeping gene shown for ST64, ST69, STLus2, STLus3 and STLus4.
The numbers above the sequences refer to the position in the concatenated alignment. Regions corresponding to the individual genes are separated by a line and gene names are given on the top. The likely recombination event between ST64 and ST69 in clpX is indicated by an arrow.
Figure 7
Figure 7. Network analysis.
An analysis of B. lusitaniae MLST data (concatenated housekeeping gene sequences) using SplitDecomposition provided a network at the split separating the strains from Mafra which coincides with a recombination event in clpX. The two populations from Mafra and Grândola are well separated.

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