Plasmid based protein composition of <i>B</i>. <i>mayonii</i> in comparison to 4 other Bbsl genospecies and 1 RF <i>Borrelia</i> species.

<p>The mean amino acid identity is indicated for proteins present on each of the 14 <i>B</i>. <i>mayonii</i> plasmids. Comparisons are to protein databases constructed for <i>B</i>. <i>burgdorferi</i> (B.b.s.s.; green), <i>B</i>. <i>garinii</i> (B.g.; blue), <i>B</i>. <i>afzelii</i> (B. a.; red), <i>B</i>. <i>bissettii</i> (B.b.; yellow), and <i>B</i>. <i>miyamotoi</i> (B.m.; grey).</p

Comparison of <i>B</i>. <i>burgdorferi</i> B31 and <i>B</i>. <i>mayonii</i> lp54.

<p>Heat map of nucleotide identity between lp54 plasmids of <i>B</i>. <i>burgdorferi</i> B31 (top) and <i>B</i>. <i>mayonii</i> MN14-1420 (bottom). Nucleotide identity along the entire lp54 plasmids is displayed as colored blocks (Yellow = highest, Blue = lowest). Orientation of arrows indicates gene orientation on the forward or reverse DNA strand. The variable PF54 gene array is at the 3’ end of the plasmid. Specific gene loci discussed in text are indicated.</p

Whole genome sequencing and annotation statistics for <i>B</i>. <i>mayonii</i> MN14-1420 and MN14-1539.

<p>Whole genome sequencing and annotation statistics for <i>B</i>. <i>mayonii</i> MN14-1420 and MN14-1539.</p

Phylogenetic analysis of <i>B</i>. <i>mayonii</i> based on a ~6.5 kbp region of cp26.

<p>The ~6.5 kbp region of cp26 encompasses open reading frames from BB_B14 through BB_B18 of <i>B</i>. <i>burgdorferi</i> B31. DNA sequences from 8 other Bbsl genospecies (22 strains) were retrieved from GenBank. Bootstrap values greater than 50% are shown. The scale bar corresponds to 0.02 substitutions per nucleotide position.</p

Nucleotide differences detected between <i>B. mayonii</i> strains MN14-1420 and MN14-1539^*.

<p>Nucleotide differences detected between <i>B. mayonii</i> strains MN14-1420 and MN14-1539^{<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0168994#t003fn001" target="_blank">*</a>}.</p

<i>vls</i> containing linear plasmid lp28-10 from <i>B</i>. <i>mayonii</i> MN14-1420 and MN14-1539.

<p>A) Heat map displaying nucleotide identity (blue—lowest; yellow—highest) between the active <i>vlsE</i> nucleotide sequence (magenta arrow) and silent <i>vls</i> cassettes within the two <i>B</i>. <i>mayonii</i> strains. Position and copy number of the silent cassette array is indicated by the blue to yellow ribbons. Red bars indicate silent <i>vls</i> loci present in MN14-1420 and missing in MN14-1539. Genes flanking the <i>vls</i> locus are indicated by black arrows. Orientation of arrows indicates gene orientation on the forward or reverse DNA strand. B) Multi-alignment of the 26 amino acid C6 peptide within VlsE from <i>B</i>. <i>mayonii</i>, <i>B</i>. <i>burgdorferi</i> B31 and <i>B</i>. <i>garinii</i> IP90. Identical (*), conserved (:) and differentially charged (.) amino acid residues are indicated.</p

Putative <i>B</i>. <i>burgdorferi</i> plasmid-borne proteins absent from the <i>B</i>. <i>mayonii</i> genome.

<p>Putative <i>B</i>. <i>burgdorferi</i> plasmid-borne proteins absent from the <i>B</i>. <i>mayonii</i> genome.</p

Whole Genome Sequence and Comparative Genomics of the Novel Lyme Borreliosis Causing Pathogen, <i>Borrelia mayonii</i>

<div><p><i>Borrelia mayonii</i>, a <i>Borrelia burgdorferi sensu lato</i> (Bbsl) genospecies, was recently identified as a cause of Lyme borreliosis (LB) among patients from the upper midwestern United States. By microscopy and PCR, spirochete/genome loads in infected patients were estimated at 10⁵ to 10⁶ per milliliter of blood. Here, we present the full chromosome and plasmid sequences of two <i>B</i>. <i>mayonii</i> isolates, MN14-1420 and MN14-1539, cultured from blood of two of these patients. Whole genome sequencing and assembly was conducted using PacBio long read sequencing (Pacific Biosciences RSII instrument) followed by hierarchical genome-assembly process (HGAP). The <i>B</i>. <i>mayonii</i> genome is ~1.31 Mbp in size (26.9% average GC content) and is comprised of a linear chromosome, 8 linear and 7 circular plasmids. Consistent with its taxonomic designation as a new Bbsl genospecies, the <i>B</i>. <i>mayonii</i> linear chromosome shares only 93.83% average nucleotide identity with other genospecies. Both <i>B</i>. <i>mayonii</i> genomes contain plasmids similar to <i>B</i>. <i>burgdorferi sensu stricto</i> lp54, lp36, lp28-3, lp28-4, lp25, lp17, lp5, 5 cp32s, cp26, and cp9. The <i>vls</i> locus present on lp28-10 of <i>B</i>. <i>mayonii</i> MN14-1420 is remarkably long, being comprised of 24 silent <i>vls</i> cassettes. Genetic differences between the two <i>B</i>. <i>mayonii</i> genomes are limited and include 15 single nucleotide variations as well as 7 fewer silent <i>vls</i> cassettes and a lack of the lp5 plasmid in MN14-1539. Notably, 68 homologs to proteins present in <i>B</i>. <i>burgdorferi sensu stricto</i> appear to be lacking from the <i>B</i>. <i>mayonii</i> genomes. These include the complement inhibitor, CspZ (BB_H06), the fibronectin binding protein, BB_K32, as well as multiple lipoproteins and proteins of unknown function. This study shows the utility of long read sequencing for full genome assembly of Bbsl genomes, identifies putative genome regions of <i>B</i>. <i>mayonii</i> that may be linked to clinical manifestation or tissue tropism, and provides a valuable resource for pathogenicity, diagnostic and vaccine studies.</p></div

<i>B</i>. <i>mayonii</i> proteins with greatest sequence diversity as compared to 4 other Bbsl genospecies and 1 RF Borrelia species.

<p>Bar graph depicting <i>B</i>. <i>mayonii</i> proteins greater than 100 amino acids with less than 60% amino acid identity to other Bbsl proteins. Amino acid identity is shown with respect to homologs present in <i>B</i>. <i>burgdorferi</i> (B.b.s.s.; green), <i>B</i>. <i>garinii</i> (B.g.; blue), <i>B</i>. <i>afzelii</i> (B. a.; red), <i>B</i>. <i>bissettii</i> (B.b.; yellow) and <i>B</i>. <i>miyamotoi</i> (B.m.; grey).</p

Temperature curves.

<p>Mice (n = 14 mice for each of the four <i>F. tularensis</i> infection groups) infected in round 1 and round 2 with <i>F. tularensis</i> groups (A1a, A1b, A2 and type B) are shown. Mice were infected with <i>F. tularensis</i> and temperature was monitored every 1–2 hours until mice expired.</p