Helicobacter pylori genome evolution during human infection

Abstract

High genetic diversity is a hallmark of the gastric pathogen Helicobacter pylori. We used 454 sequencing technology to perform whole-genome comparisons for five sets of H. pylori strains that had been sequentially cultured from four chronically infected Colombians (isolation intervals=3-16 y) and one human volunteer experimentally infected with H. pylori as part of a vaccine trial. The four sets of genomes from Colombian H. pylori differed by 27-232 isolated SNPs and 16-441 imported clusters of polymorphisms resulting from recombination. Imports (mean length=394 bp) were distributed nonrandomly over the chromosome and frequently occurred in groups, suggesting that H. pylori first takes up long DNA fragments, which subsequently become partially integrated in multiple shorter pieces. Imports were present at significantly increased frequency in members of the hop family of outer membrane gene paralogues, some of which are involved in bacterial adhesion, suggesting diversifying selection. No evidence of recombination and few other differences were identified in the strain pair from an infected volunteer, indicating that the H. pylori genome is stable in the absence of mixed infection. Among these few differences was an OFF/ON switch in the phase-variable adhesin gene hopZ, suggesting strong in vivo selection for this putative adhesin during early colonization.

Fig. 1.

Sequential accumulation of isolated SNPs and clusters of nucleotide polymorphisms in two triplets of sequential isolates, NQ392/1707/4060 and NQ367/1671/4191, cultured after 0, 3, and 16 y of chronic infection. Black numbers indicate the numbers of SNPs or CNPs in pair-wise genome comparisons. Because there was only a partial overlap of differences for the two pair-wise comparisons, a common ancestor of both follow-up isolates was inferred (black dot) based on the overlapping differences, and gray numbers indicate differences between the sequenced isolates and the inferred intermediate clone. Overlapping and nonoverlapping events of the second isolates do not always add up to the total number, because there are cases where the presence or absence of the event in the third isolate could not be determined.

Fig. 2.

Analysis of the aligned genomes of sequential H. pylori isolate pairs. Each line represents 50 kb of sequence. Nonaligned regions are shown in white. Regions colored in black are clonal, regions in green are imported, and regions in red are not imported but in a cluster of imports. The intensity of each color represents statistical uncertainty. Polymorphism between the paired genomes is shown by gray triangles.

Fig. 3.

Proportion of genes affected by imports for different functional classes of encoded proteins. Import frequency for a category is calculated as the percentage of genes affected by imports. Color-coding is shown below the graphs. Asterisks indicate significantly increased frequency of imports (Fisher exact test; *P < 0.05; **P < 0.01). The composition of the group of outer membrane proteins (yellow bars) and the hop subfamily (hatched yellow bars) is based on ref. . SI Appendix, Fig. S6 has an extended graph including additional outer membrane protein subfamilies.