Phylogeographic origin of Helicobacter pylori is a determinant of gastric cancer risk

Abstract

Background and aims: Helicobacter pylori colonises the stomach in half of all humans, and is the principal cause of gastric cancer, the second leading cause of cancer death worldwide. While gastric cancer rates correlate with H pylori prevalence in some areas, there are regions where infection is nearly universal, but rates of gastric cancer are low. In the case of Colombia, there is a 25-fold increase in gastric cancer rate in the Andean mountain (high risk) region compared to the coastal (low risk) region, despite similarly high (∼90%) prevalence of H pylori in the two locations. Our aim was to investigate the ancestral origin of H pylori strains isolated from subjects in these high- and low-risk regions and to determine whether this is a predictive determinant of precancerous lesions.

Methods: Multi-locus sequence typing was used to investigate phylogeographic origins of infecting H pylori strains isolated from subjects in the Pacific coast and Andes Mountains in the state of Nariño, Colombia. We analysed 64 subjects infected with cagA+ vacA s1m1 strains. Gastric biopsy slides from each individual were scored for histological lesions and evaluated for DNA damage by immunohistochemistry.

Results: We show that strains from the high-risk region were all of European phylogeographic origin, whereas those from the low risk region were of either European (34%) or African origin (66%). European strain origin was strongly predictive of increased premalignant histological lesions and epithelial DNA damage, even in the low-risk region; African strain origin was associated with reduced severity of these parameters.

Conclusion: The phylogeographic origin of H pylori strains provides an explanation for geographic differences in cancer risk deriving from this infection.

Figure 1

MLST analysis of 64 cagA⁺ vacA s1m1 Helicobacter pylori strains from Colombia. (A) Neighbor-joining tree; branches are drawn to scale to represent evolutionary distance. ◯ with green lines, isolates from LR region (low risk of gastric cancer); ● with blue lines, isolates from HR region (high risk of gastric cancer). The brackets denote the classification of the strains into two groups. (B) Distribution of strains into phylogenetic groups defined in (A), according to risk region of origin.

Figure 2

MLST analysis of 64 cagA⁺ vacA s1m1 Colombian isolates from the HR and LR regions, along with 380 reference strains that were previously classified into distinct ancestral haplogroups [34]. (A) Neighbor-joining tree built using bootstrapping with 10,000 trials. Branches are drawn to scale to represent evolutionary distance. ◯, isolates from LR region; ● isolates from HR region. The colours of the branches represent the classification of strains into distinct ancestral haplogroups [34]. (B) Distribution of strains into phylogeographic groups defined in (A), according to risk region of origin.

Figure 3

Histopathologic analysis of biopsies from subjects infected with Colombian H. pylori strains. Histologic scoring on a 1 to 6 scale (defined in Methods; each symbol represents the score for an individual subject. (A) Histopathology scores by risk region of origin. (B) Scores grouped by phylogenetic category of the strains. Horizontal bars represent the mean; p values were obtained by a linear multivariate model analysis.

Figure 4

Histopathologic analysis of biopsies from subjects infected with Colombian H. pylori strains. Histologic scoring on a 1 to 6 scale (defined in Methods; each symbol represents the score for an individual subject. (A) Scores grouped according to phylogeny within a given risk region of origin. Horizontal bars represent the mean; pairwise comparisons were made using Dunnett's test with Sidak's adjustment. (B) Representative photomicrographs of hematoxylin and eosin staining of gastric tissue as follows: (left panel) LR subject infected with hpAfrica1 strain, showing non-atrophic gastritis; (center panel) LR subject infected with hpEurope strain, demonstrating intestinal metaplasia with changes indefinite for dysplasia; and (right panel) HR subject infected with hpEurope strain with intestinal metaplasia. Magnification in B was 200×.

Figure 5

Detection of oxidative DNA damage in gastric biopsies by quantification of 8-OHdG-positive epithelial cell nuclei, expressed as % positively-stained nuclei. (A) 8-OHdG levels, by risk region of origin. (B) Cases grouped according to strain phylogeny. Horizontal bars represent the mean; statistics were performed as in figure 3.

Figure 6

Detection of oxidative DNA damage in gastric biopsies by quantification of 8-OHdG-positive epithelial cell nuclei, expressed as % positively-stained nuclei. (A) Cases grouped according to strain phylogeny within a given risk region of origin. Horizontal bars represent the mean; statistics were performed as in figure 4. (B) Representative photomicrographs of 8-OHdG staining on gastric tissues as follows: (left panel) LR subject infected with hpAfrica1 strain; (center panel) LR subject infected with hpEurope strain; and (right panel) HR subject infected with hpEurope strain. Nuclei showing DNA damage are stained brown. Magnification in B was 400×.