Genome-wide effect of MRS-deficient hypermutability on the mutagenesis of mononucleotide SSRs.

<p>The bars show the fold increase in the mutations per year that occurred in mononucleotide G:C SSRs and A:T SSRs, and in other types of mutation after <i>P. aeruginosa</i> PACS2 became mutator<b>.</b></p

Involvement of mononucleotide SSRs in genes involved in <i>P. aeruginosa</i> adaptation during CF chronic infection.

<p>The bar graphs show (i) the percentages of the 60 genes mutated during CF lung chronic infection as reported by Smith <i>et al. </i><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0080514#pone.0080514-Smith1" target="_blank">[1]</a> that harbor mononucleotide G:C SSRs (A) and A:T SSRs (B), relative to the length of the SSR (black bars); (ii) the percentages of those genes carrying mononucleotide SSRs that were mutated during the process of chronic infection, relative to the length of the SSR (gray bars).</p

Mononucleotide SSRs in the <i>P. aeruginosa</i> PAO1 genome.

<p>The plots show the counts for mononucleotide SSRs (red circles) in the whole <i>P. aeruginosa</i> PAO1 genome and in random sequences generated by various predictive models (black symbols). b and m1: homogeneous models (Bernoulli and first-order Markov); b-b, b-bp, m1-m1, m1-m1p, m1-c and m1-c1: heterogeneous models (see Methods). Counts are shown of mononucleotide G:C and A:T SSRs in the coding and non-coding regions of the genome.</p

Evidence for parallel evolution. Genes identified as being independently mutated in at least a half of the coexisting CFA I–IV and CFD I–VI sub-lineages.

a<p>The categories used for functional classification were as described in the The <i>Pseudomonas aeruginosa</i> Community Annotation Project (<a href="http://www.pseudomonas.com" target="_blank">http://www.pseudomonas.com</a>).</p><p>Evidence for parallel evolution. Genes identified as being independently mutated in at least a half of the coexisting CFA I–IV and CFD I–VI sub-lineages.</p

Minimum spanning trees (MSTs) of genomes among CFA and CFD lineages.

<p>MSTs for CFA (A) and CFD (B) were constructed based on the total number of genes altered by nonsynonymous SNPs and indel mutations in the respective genomes. Links between nodes represents the minimum distance in terms of mutated genes. Numbers above each link indicate the total amount of mutated genes between the two connected nodes (<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004651#pgen.1004651.s008" target="_blank">Table S6</a>). For tree construction, ancestors CFA_2004/01 and CFD_1991/01 were considered as origins.</p

Average total catabolic function of isolates from CFA and CFD lineages.

<p>Total catabolic function was calculated relative to CFA_2004/01 and CFD_1991/01 as a weighted average across all substrates for each CFA (A) and CFD (B) isolate. Total catabolic function was defined as 1 for the reference levels (CFA_2004/01 and CFD_1991/01). Lower values indicate decay. Isolates CFA_2010/01, CFA_2010/11, and CFA_2010/31 were excluded from the analysis because significant dispersion was observed in the duplicates.</p

Mutational spectra and top mutated homopolymeric G∶C SSRs in CFA_2010 and CFD_2011 contemporary isolates.

<p>(A) Percentage of 1–4 bp insertions/deletions located in G∶C and A∶T homopolymeric sequences. (B) The heat map represents individual indels mutations in homopolymeric G∶C SSRs of ≥6 bp, which were mutated in at least half of the coexisting isolates in both CFA and CFD lineages. The color-code indicates the type of mutation. <i>Right</i>: Percentage of MRS-deficient isolates harboring a indel mutation in each analyzed G∶C SSR. (C) Mutations in CFA_2010 and CFD_2011 isolates were analyzed based on the percentage of transitions, transversions, and insertions/deletions.</p

Pathoadaptive genes convergently mutated in CFA and CFD sub-lineages.

<p>The analysis was performed based only on non-synonymous mutated genes that were altered independently in at least half of the 10 evolving sub-lineages CFA I–IV and CFD I–VI.</p

Isolate sampling points and patient life spans.

<p><i>P. aeruginosa</i> isolates were collected from two CF patients: CFA and CFD. Hollow symbols: single bacterial isolates. Solid circles: cross-sectional populations of 90 bacterial isolates. *: estimated start of chronic infection. Gray bar: patient life span.</p

Evolutionary relationships among isolates from CFA and CFD lineages.

<p>Maximum-parsimony phylogenetic trees of CFA (A) and CFD (B) were constructed based on the accumulation of new SNPs relative to ancestors CFA_2004/01 and CFD_1991/01. Alleles of <i>P. aeruginosa</i> reference strain PAO1 were used to root the trees. Lengths of branches are proportional to the number of accumulated SNPs. Branches are designated by capital letters. MRCA: most recent common ancestor.</p