Multilocus sequence typing of Borrelia burgdorferi suggests existence of lineages with differential pathogenic properties in humans

Abstract

The clinical manifestations of Lyme disease, caused by Borrelia burgdorferi, vary considerably in different patients, possibly due to infection by strains with varying pathogenicity. Both rRNA intergenic spacer and ospC typing methods have proven to be useful tools for categorizing B. burgdorferi strains that vary in their tendency to disseminate in humans. Neither method, however, is suitable for inferring intraspecific relationships among strains that are important for understanding the evolution of pathogenicity and the geographic spread of disease. In this study, multilocus sequence typing (MLST) was employed to investigate the population structure of B. burgdorferi recovered from human Lyme disease patients. A total of 146 clinical isolates from patients in New York and Wisconsin were divided into 53 sequence types (STs). A goeBURST analysis, that also included previously published STs from the northeastern and upper Midwestern US and adjoining areas of Canada, identified 11 major and 3 minor clonal complexes, as well as 14 singletons. The data revealed that patients from New York and Wisconsin were infected with two distinct, but genetically and phylogenetically closely related, populations of B. burgdorferi. Importantly, the data suggest the existence of B. burgdorferi lineages with differential capabilities for dissemination in humans. Interestingly, the data also indicate that MLST is better able to predict the outcome of localized or disseminated infection than is ospC typing.

Figure 1. A population snapshot of B. burgdorferi in the northeastern and midwestern United States and Canada.

The snapshot comprises of 88(420 human and tick samples) and was created by goeBURST v1.2 using data from this study and the previously published data sets downloaded from http://borrelia.mlst.net/ , –. Circle size and color correspond to MLST sample size and the source, respectively. Gray, strains found in ticks; white, strains found in humans. Colored lines connecting STs indicate descending order of certainty; black lines are inferred without tiebreak rules, blue lines are inferred using tiebreak rule 1 (number of SLV), and green lines are inferred using tiebreak rule 2 (number of DLV). STs connected by a black line are single locus variants and STs connected by gray line are double locus variants. The inferred founders of clonal complexes are numbered in bold. STs found in patients with localized infection are outlined in green, those found in patients with disseminated infection are outlined in red, those found in both patients with localized and patients with disseminated infection are outlined in orange and those found in patients with undetermined clinical status are outlined in black. STs that are not outlined were only found in ticks. The ospC major groups are shown for all 53 STs which were found in humans.

Figure 2. Unrooted ML tree of B. burgdorferi based on concatenated sequences of eight MLST housekeeping genes.

The tree was created using data from this study and the previously published data sets downloaded from http://borrelia.mlst.net/ , –. A total of 420 B. burgdorferi samples (88 STs) found in humans and ticks from the northeastern United States and Canada were used. The aLRT statistical values and nonparametric bootstrap values for highly supported nodes in both maximum parsimony (with >70% support) and maximum likelihood (with aLRT >0.9 support) are indicated above and below the branches, respectively. STs newly identified in this study are in bold. The grouping of STs into major clonal complexes (CCs) is indicated by right brackets. The STs found only in humans are shown in blue, those found only in ticks are shown in red and those found in both humans and ticks are shown in green. The type of infection is indicated next to the ST using solid square (ST found in patients with localized infection), solid triangle (ST found in patients with disseminated infection) and solid diamond (ST found in both patients with localized and patients with disseminated infection). Geographical origin of STs found in humans and identified in this study is indicated in brackets next to the STs (NY – New York; WI – Wisconsin).

Figure 3. Venn diagrams depicting the geographical distribution of B. burgdorferi in Lyme disease patients.

A) Geographical distribution of B. burgdorferi STs found in Lyme disease patients from New York and Wisconsin. B) Geographical distribution of B. burgdorferi ospC major groups identified in skin samples of Lyme disease patients from New York and Wisconsin.