Helicobacter colonization of the biliary tree: commensal, pathogen, or spurious finding?

The authors hypothesized that Helicobacter species may be present in the bile and gallbladder wall of patients with chronic cholecystitis who live in a region with a high prevalence of gallbladder cancer. They attempted to identify such species by obtaining both bile and resected gallbladder tissue from 46 patients who underwent cholecystectomy. Tissue specimens were stained with hematoxylin and eosin as well as other stains used specifically for the identification of Helicobacter species, and culture was attempted using specialized media on samples from tissue and bile. Unfortunately, the authors were unable to culture any Helicobacter species, and the yield from histopathology was also poor with silver stains identifying curved bacteria suggestive, but not diagnostic, of Helicobacter species in only two cases. Molecular techniques were more successful. DNA was extracted from both tissue and bile and amplified by polymerase chain reaction (PCR) using a specific primer. The amplicons they identified were then compared with known Helicobacter proteins using a Southern blot approach. PCR amplification was relatively successful with 9 of 23 gallbladder samples and 13 of the 23 bile samples coming up positive for Helicobacter species using two specific primers. These specimens were also positive by Southern blot hybridization. The cloning and sequencing of the 16S ribosomal RNA amplicons in eight cases verified true Helicobacter origin with a phylogenetic analysis showing greater than 99.3% similarity. Five of the amplicons clustered with H. bilis, two with Flexispira rappini, and one with H. pullorum. The authors concluded that despite their being unable to identify organisms directly, the stringent PCR technique with amplicon sequencing confirmed that Helicobacter species could be identified within the bile and gallbladder tissue of patients with chronic cholecystitis in a region with high incidence of gallbladder cancer. They indicated that further studies are needed to ascertain whether similar species have a causative role in the development of gallbladder cancer.