Identification of a genetic marker of Helicobacter pylori strains involved in gastric extranodal marginal zone B cell lymphoma of the MALT-type

Abstract

Background and aims: Gastric extranodal marginal zone B cell lymphoma of the mucosa associated lymphoid tissue (MALT)-type (MZBL) is a rare complication of Helicobacter pylori infection. Currently, no bacterial factor has been associated with the development of this disease. Our aim was to identify genes associated with lymphoma development. Methods: We used subtractive hybridisation as a tool for comparative genomics between H pylori strains isolated from a patient with gastric MZBL and from a patient with gastritis only. Results: When gastric MZBL strains were compared with gastritis strains, two open reading frames (ORFs) were significantly associated with gastric MZBL: JHP950 (74.4% v 48.7%, respectively; p = 0.023) and JHP1462 (25.6% v 2.6%, respectively; p = 0.004). The prevalence of JHP950 was 48.8% (p = 0.024) in duodenal ulcer strains and 39.3% (p = 0.006) in gastric adenocarcinoma strains, which makes this ORF a specific marker for gastric MZBL strains. In contrast, the prevalence of JHP1462 was 16% (p = 0.545) and 35.7% (p = 0.429) in duodenal ulcer and adenocarcinoma strains, respectively. These ORFs were present in reference strain J99 but not in reference strain 26695. JHP950 is located in the plasticity zone whereas the other, JHP1462, is located outside. Both encode for H pylori putative proteins with unknown functions. Conclusion: Despite its low prevalence, the ORF JHP1462 can be considered a candidate marker for H pylori strains involved in severe gastroduodenal diseases. In contrast, the ORF JHP950 has a high prevalence, and is the first candidate marker for strains giving rise to an increased risk of gastric MZBL strains. Further confirmation in other studies is needed.

Identification of Markers for Helicobacter pylori Strains Isolated from Children with Peptic Ulcer Disease by Suppressive Subtractive Hybridization

“…Only one sequence showed a region with no homology to database sequences, whereas in other similar studies original sequences were identified in H. pylori (3,39). There are two possible explanations for this result: either no original sequence exists in the strain tested in the present study or such sequences exist and were missed, indicating that a more exhaustive screening of the E. coli colonies should have been performed.…”

“…In terms of functional categories, the most abundant class of strain-specific genes is unique to H. pylori with unknown function, as was reported in other studies (39,50). The homologies of the other classes of strain-specific genes imply functions that may aid the bacterium under certain circumstances or in certain hosts, such as OMP and LPS genes, and also restriction modification systems that help promote the genetic diversity and thus contribute to the adaptation of the bacterium.…”

“…The restriction enzyme used was AluI (New England Biolabs, Ipswich, MA) according to the method of Akopyants et al (3). The treatment of both tester and driver DNAs with the AluI restriction enzyme generated fragments with sizes between 1,250 and 178 bp, as previously reported (3,39). The analysis of the subtraction efficacy was performed by real-time PCR with the SYBR-Green format in a LightCycler thermocycler (Roche Diagnostics GmbH, Mannheim, Germany), using the H. pylori 16S rRNA gene as the target gene.…”

“…In addition, a French collection of 43 H. pylori strains obtained from gastric MALT lymphoma patients (62.8% male; mean age, 48.2 Ϯ 13.4 years) described previously (39) was also included.…”

“…All of the sequences with a PCR-positive result for the tester-strain and PCR negative for the driver-strain were analyzed by dot blot DNA hybridization according to conditions previously reported (39,44) in the collection of H. pylori strains, in order to look for possible associations. The J99 and 26695 strains were also included as controls for hybridization.…”