In vivo competition and horizontal gene transfer among distinct Staphylococcus aureus lineages as major drivers for adaptational changes during long-term persistence in humans

Abstract

Background: The airways of the majority of adolescent cystic fibrosis (CF) patients are persistently colonized or infected by Staphylococcus aureus. Using whole genome sequencing, we studied the evolutionary traits within a S. aureus population in the airways of a CF patient hypothesizing that horizontal gene transfer (HGT) and inter-bacterial interaction play a major role in adaptation during long-term persistence.

Results: Whole genome sequencing of 21 S. aureus isolates spanning 13 years resulted in seven lineages defined by the spa types t012, t021, t331, t338, t364, t056, and t2351. Of these, the successfully persisting lineages t012 and t021 were closely related suggesting the evolution of t021 from t012, which was further corroborated by a nearly identical, syntenic set of mobile genetic elements. During transformation from t012 to t021, an increase of genomic changes including HGT from other S. aureus lineages was detected.

Conclusions: In summary, our in vivo data enabled us to conceptualize an evolutionary model showing the impact of HGT and inter-bacterial interaction on bacterial long-term adaptation to the human host during CF.

Keywords: Adaptation; Cystic fibrosis; Genome sequencing; Horizontal gene transfer; Staphylococcus aureus.

Fig. 1

K-mer based unrooted UPGMA tree of all 21 S. aureus isolates used in this study. The tree was generated using CLC Genomics Workbench (k-mer size 15; distance calculation method was “Mahalanobis”) using the de novo assembled contigs or complete genome sequences of the respective isolates. The four isolates marked with an asterisk were isolated from another sampling source compared to the unmarked isolates from the same spa type obtained in the same year without exhibiting any differences on the genomic level (Additional File 5). Moreover, the MLST sequence types are given for each sample in parenthesis

Fig. 2

Comparison of mobile genomic elements of the isolates t012#00 and t021#02. The colored boxes on black horizontal lines (i. e. the chromosomes) represent the different mobile elements. Identical elements share the same color, are named accordingly, and are linked with a dashed line between the two isolates. Green bars flanking the mobile elements illustrate identical flanking genomic sequences in both isolates. Small insertions (In) and deletions (Del) in the pairwise comparison of the mobile elements are marked in the t021#02 isolate. Insertions within the shorter copy of SaPIn1 and the shortest copy of phage 112CMu50 are displayed in Fig. 3a and c, respectively. The deletions within SaPIn2 are shown in detail in Fig. 3b. The insertions within the longer copy of phage 112CMu50 are displayed in Fig. 3d

Fig. 3

Schemes of the three structural differences on the genomic level between the isolates t012#00 and t021#02 within the mobile elements SaPIn1, SaPIn2, and SA bacteriophage 112C Mu 50 B. Genes on the forward strand are depicted as right-sided arrows; genes located on the reverse strand are depicted as left-sided arrows. Genes showing synteny between both genomes are highlighted with the same color and are linked with a dashed line. Genes that are present in just one of both genomes are colored in red. The locus tags of the genes and the genomic position in relation to the early (for t012#00) and late (for t021#02) genome sequence, respectively, are given and listed in Additional File 7. a Genomic differences within the S. aureus pathogenicity island SaPIn1. In both isolates, the islands are flanked by a gene encoding putative permease (upstream site) and by a gene encoding heat shock protein 60 family chaperone GroEL (downstream site). b Deletion of three genes encoding tandem lipoprotein within pathogenicity islandSaPIn2 in isolate t021#02. c Genomic differences within the shorter copy of phage SA bacteriophage 112C Mu 50 B, which is inserted in the identical genomic region. d Genomic differences within the longer copy of phage SA bacteriophage 112C Mu 50 B, which is inserted in the identical genomic region

Fig. 4

Genomic organization of the novel 14 kb transposon and its insertion site in isolate t021#04. Genes on the forward strand are depicted as right-sided arrows; genes located on the reverse strand are depicted as left-sided arrows. The transposon inserted into the gene encoding the carboxylesterase type B, whose parts border the insertion element both up- and downstream. The inserted genes are highlighted in turquoise; the inverted repeats (IR) are marked with red and the direct repeats (DR) are highlighted with light blue. For each gene, the locus tags are given (see also Additional File 6 for further details)

Fig. 5

Model of the in vivo evolution and interactions of the CF S. aureus lineages from 1995 to 2008. The isolates within the airways are shown as colored circles; every lineage has its own color. Hypothetical isolates, which were not isolated but probably took place in the direct line of evolution, are depicted as dashed circles. Black arrows connect sequential isolates of the same lineage; dashed lines connect hypothetical isolates. Colored (+) and (−) indicate the acquisition or loss of genetic material. If genetic material was likely to be transferred from one lineage to another, dotted colored arrows show the assumed direction and the transferred material is listed. In case the origin was ambiguous, the transferred material is shown in brackets. In addition, the 25.7 kb DNA transposon (yellow line), the 30 kb plasmid (green circle), and the 14.2 kb DNA transposon (brown line) are shown. If applicable, boxes display in a pairwise comparison of consecutive isolates the number of differing synonymous (S), non-synonymous (N), or intergenic (I) SNPs, the number of insertions (In) and deletions (Del), and the d_N/d_S value. N/A, not applicable