Helicobacter pylori infection and DNMT3a polymorphism are associated with the presence of premature coronary artery disease and subclinical atherosclerosis. Data from the GEA Mexican Study.

BACKGROUND: The association between H. pylori infection and coronary artery disease (CAD) is well-known. Alterations in DNA methylation in CAD have been reported, which can be induced by H. pylori through the DNA demethylases (DNMTs). The objective was to analyze the association and interaction of H. pylori infection and DMNT3a gene polymorphisms with premature CAD (pCAD) and subclinical atherosclerosis (SA). METHODS: The study included 561 patients with pCAD, 318 subjects with SA, and 599 healthy controls. Antibodies against H. pylori and DNMT3a rs13420827, rs752208, and rs1550117 polymorphisms were determined. RESULTS: The pCAD group presented the highest seroprevalence of H. pylori infection (87.7%) compared to the SA (74.5%, p = 1 × 10-6) and the control group (63.1%, p = 7 × 10-23). A significant association was observed between H. pylori infection and pCAD (OR = 2.729, p = 1.0 × 10-6). The rs13420827 polymorphism was associated with a high risk of H. pylori infection in the whole population (padditive = 0.009, pdominant = 0.018, and pcodominant2 = 0.013) and in individuals with SA (padditive = 0.003, pdominant = 0.020, precessive = 0.013, and pcodominant2 = 0.005). The coexistence of H. pylori infection and the rs13420827GG genotype increases the risk of pCAD (pinteraction = 1.1 × 10-5). CONCLUSIONS: According to the model adjusted for more confounding variables, H. pylori infection was associated with almost three times the risk of developing pCAD. The rs13420827G allele was associated with an increased risk of H. pylori infection in the whole population and in individuals with SA. Individuals in whom H. pylori infection and the rs13420827GG genotype coexist are at increased risk of pCAD.

Infection with Helicobacter bilis but not Helicobacter hepaticus was Associated with Extrahepatic Cholangiocarcinoma.

BACKGROUND AND AIMS: The biliary tract cancer or cholangiocarcinoma (CCA) represents the sixth leading cause of gastrointestinal tumors in the Western world, and mortality varies across the world, with regions such as Chile, Thailand, Japan, and northeastern India presenting the highest rates. CCA may develop in the bile duct, gallbladder, or ampulla of Vater; and risk factors include obesity, parity, genetic background, geographical and environmental factors. Inflammation induced by bacterial infections might play a role in the pathogenesis of CCA. In this work, we investigated whether there is an association between extrahepatic cholangiocarcinoma (ECCA) and infection with S. typhi, H. hepaticus, or H. bilis in a Mexican population. METHODS: A total of 194 patients were included and divided into 91 patients with benign biliary pathology (controls) and 103 with ECCA (cases). Tumor samples were taken during endoscopic retrograde cholangiopancreatography by biliary brushing, followed by DNA extraction and PCR testing for infections. RESULTS: We found that 44/103 cases were positive for H. bilis, compared with 19/91 controls (p = 0.002; OR 2.83, 95% CI 1.49-5.32), and when analyzed by sub-site, H. bilis infection was significantly more associated with cancer in the common bile duct (p = 0.0005; OR 3.56, 95% CI 1.77-7.17). In contrast, H. hepaticus infection was not different between cases (17/103) and controls (13/91) (p = 0.82; OR 1.19, 95% CI 0.54-2.60). None of the samples were positive for S. typhi infection. CONCLUSION: In conclusion, infection with H. bilis but neither H. Hepaticus nor S. typhi was significantly associated with ECCA, particularly with tumors located in the common bile duct.

In vivo expression of Helicobacter pylori virulence genes in patients with gastritis, ulcer, and gastric cancer.

The best-studied Helicobacter pylori virulence factor associated with development of peptic ulcer disease or gastric cancer (GC) rather than asymptomatic nonatrophic gastritis (NAG) is the cag pathogenicity island (cagPAI), which encodes a type IV secretion system (T4SS) that injects the CagA oncoprotein into host epithelial cells. Here we used real-time reverse transcription-PCR (RT-PCR) to measure the in vivo expression of genes on the cagPAI and of other virulence genes in patients with NAG, duodenal ulcer (DU), or GC. In vivo expression of H. pylori virulence genes was greater overall in gastric biopsy specimens of patients with GC than in those of patients with NAG or DU. However, since in vitro expression of cagA was not greater in H. pylori strains from patients with GC than in those from patients with NAG or DU, increased expression in GC in vivo is likely a result of environmental conditions in the gastric mucosa, though it may in turn cause more severe pathology. Increased expression of virulence genes in GC may represent a stress response to elevated pH or other environmental conditions in the stomach of patients with GC, which may be less hospitable to H. pylori colonization than the acidic environment in patients with NAG or DU.

Infections With Enterohepatic Non-H. pylori Helicobacter Species in X-Linked Agammaglobulinemia: Clinical Cases and Review of the Literature.

The genus Helicobacter is classified into two main groups according to its habitat: gastric and enterohepatic. Patients with X-linked agammaglobulinemia (XLA) appear to be associated with invasive infection with enterohepatic non-Helicobacter pylori species (NHPH), mainly H. cinaedi and H. bilis. Such infections are difficult to control and have a high potential for recurrence. The spectrum of illnesses caused by these species includes recurrent fever, bacteremia, arthritis, osteomyelitis, cellulitis, abdominal abscesses, and pyoderma gangrenosum-like ulcer. The presence of these Helicobacters is particularly difficult to diagnose and eradicate, as they are very fastidious bacteria and present resistance to several types of antibiotics. We report two clinical cases of XLA patients infected with H. bilis. These infections were chronic in these patients and could not be eradicated in one of them. We also review the cases of enterohepatic non-Helicobacter pylori species (NHPH) in patients with this inborn error of immunity.

Toxigenic Helicobacter pylori infection precedes gastric hypochlorhydria in cancer relatives, and H. pylori virulence evolves in these families.

PURPOSE: Helicobacter pylori infection by virulent strains is associated with gastric adenocarcinoma. We aimed to determine whether infection with virulent H. pylori preceded precancerous gastric hypochlorhydria and atrophy in gastric cancer relatives and quantify the extent of virulence factor evolution. EXPERIMENTAL DESIGN: H. pylori strains from 51 Scottish gastric cancer relatives were characterized by genetic fingerprinting and typing the vacuolating cytotoxin gene (vacA), the cytotoxin-associated gene (cagA), and housekeeping genes. We phenotyped strains by coculture with gastric epithelial cells and assessing vacuolation (microscopy), CagA tyrosine phosphorylation (immunoblot), and interleukin-8 secretion (ELISA). RESULTS: Toxigenic (vacA type s1/m1) H. pylori was associated with precancerous gastric hypochlorhydria (P<0.01). Adult family members with this type of H. pylori had the same strain as currently noncohabiting adult family members in 68% cases, implying acquisition during childhood from each other or a common source. We analyzed different isolates of the same strain within families and showed that H. pylori commonly microevolved to change virulence: this occurred in 22% individuals and a striking 44% cases where the strain was shared within families. Microevolution in vacA occurred by extragenomic recombination and in cagA by this or duplication/deletion. Microevolution led to phenotypic changes in virulence. Passage of microevolved strains could be tracked within families. CONCLUSIONS: Toxigenic H. pylori infection precedes and so likely causes gastric hypochlorhydria, suggesting that virulent H. pylori increases cancer risk by causing this condition. Microevolution of virulence genes is common within families of gastric cancer patients and changes H. pylori virulence.

Effect of the flavonoid quercetin on inflammation and lipid peroxidation induced by Helicobacter pylori in gastric mucosa of guinea pig.

BACKGROUND: Helicobacter pylori infection induces an inflammatory response in the gastric mucosa. Activation of polymorphonuclear leukocytes can produce oxidative damage to gastric tissue through intermediary radicals of oxygen and nitrogen. Vegetable extracts containing polyphenols of the flavonoid family have antibacterial activity, and the flavonoid quercetin possesses anti-H. pylori activity in vitro. The aim of this study was to analyze the effect of oral administration of pure quercetin on inflammation and lipid peroxidation induced by H. pylori in the gastric mucosa of the guinea pig. METHODS: Sixty days after oral infection with H. pylori guinea pigs received 200 mg/kg of quercetin daily by mouth for 15 days. The infiltration index of inflammatory cells and bacterial density in both the pyloric antrum and corpus were histologically determined by myeloperoxidase histochemistry, hematoxylin-eosin, and modified Giemsa stains. The lipid hydroperoxide content was assessed by the orange xylenol spectrophotometric method. RESULTS: Quercetin significantly reduced the infiltration index of mononuclear cell and bacterial colonization in the pyloric antrum and corpus. In the antrum of infected quercetin-treated animals, a significant diminution of neutrophil leukocyte infiltration was observed compared with the infected nonquercetin-treated animals. In the antrum, the lipid hydroperoxide concentration was significantly decreased in infected animals treated with quercetin, whereas in the corpus no significant differences were observed. CONCLUSIONS: Our results indicate that in vivo oral quercetin administration decreases H. pylori infection in the gastric mucosa and reduces both the inflammatory response and lipid peroxidation.

On the Search to Elucidate the Role of Microbiota in the Genesis of Cancer: The Cases of Gastrointestinal and Cervical Cancer.

The microbiota that inhabits the human body plays an important role in health and disease, by their fundamental role in food digestion, training of the immune system or protection against pathogen colonization. However, when the equilibrium with its host is altered, some diseases like cancer might be promoted. In this review we describe the information collected in recent studies between the microbiota and its association with cancer. We conducted the review of the relation of microbiome and cancer etiology focusing on the gastrointestinal and cervical cancer. The MEDLINE database was used for the search. The gastrointestinal tract harbours a diverse and site specific microbiota, and several studies have demonstrated that perturbation of these microbial communities might be associated with different types of cancer. In particular, alteration of the colorectal, gastric and oesophageal microbiota have been reported associated with cancer development. Likewise, cervical microbiome studies suggest that some members of the cervical microbiota are possible modifiers of the cytokine profile of the cervical microenvironment during the development of cervical lesions and cervical cancer. Larger prospective studies are needed to examine whether microbiome dysbiosis could cause cancer, and to evaluate the utility of microbiome profiles as biomarkers for prevention and early diagnosis. This is an important area of research if we consider that microbiota may be a modifiable factor by the use of pre- and probiotics, in order to prevent cancer evolution or even to potentiate cancer treatment.

Whole Genome Sequence and Phylogenetic Analysis Show Helicobacter pylori Strains from Latin America Have Followed a Unique Evolution Pathway.

Helicobacter pylori (HP) genetics may determine its clinical outcomes. Despite high prevalence of HP infection in Latin America (LA), there have been no phylogenetic studies in the region. We aimed to understand the structure of HP populations in LA mestizo individuals, where gastric cancer incidence remains high. The genome of 107 HP strains from Mexico, Nicaragua and Colombia were analyzed with 59 publicly available worldwide genomes. To study bacterial relationship on whole genome level we propose a virtual hybridization technique using thousands of high-entropy 13 bp DNA probes to generate fingerprints. Phylogenetic virtual genome fingerprint (VGF) was compared with Multi Locus Sequence Analysis (MLST) and with phylogenetic analyses of cagPAI virulence island sequences. With MLST some Nicaraguan and Mexican strains clustered close to Africa isolates, whereas European isolates were spread without clustering and intermingled with LA isolates. VGF analysis resulted in increased resolution of populations, separating European from LA strains. Furthermore, clusters with exclusively Colombian, Mexican, or Nicaraguan strains were observed, where the Colombian cluster separated from Europe, Asia, and Africa, while Nicaraguan and Mexican clades grouped close to Africa. In addition, a mixed large LA cluster including Mexican, Colombian, Nicaraguan, Peruvian, and Salvadorian strains was observed; all LA clusters separated from the Amerind clade. With cagPAI sequence analyses LA clades clearly separated from Europe, Asia and Amerind, and Colombian strains formed a single cluster. A NeighborNet analyses suggested frequent and recent recombination events particularly among LA strains. Results suggests that in the new world, H. pylori has evolved to fit mestizo LA populations, already 500 years after the Spanish colonization. This co-adaption may account for regional variability in gastric cancer risk.

Stomach microbiota composition varies between patients with non-atrophic gastritis and patients with intestinal type of gastric cancer.

We aimed to characterize microbiota of the gastric mucosa as it progress to intestinal type of cancer. Study included five patients each of non-atrophic gastritis (NAG), intestinal metaplasia (IM) and intestinal-type gastric cancer (GC). Gastric tissue was obtained and DNA extracted for microbiota analyses using the microarray G3 PhyloChip. Bacterial diversity ranged from 8 to 57, and steadily decreased from NAG to IM to GC (p = 0.004). A significant microbiota difference was observed between NAG and GC based on Unifrac-presence/absence and weighted-Unifrac-abundance metrics of 283 taxa (p < 0.05). HC-AN analyses based on presence/absence of 238 taxa revealed that GC and NAG grouped apart, whereas IM overlapped with both. An ordinated analyses based on weighted-Unifrac distance given abundance of 44 taxa showing significance across categories revealed significant microbiota separation between NAG and GC. This study is the first to show a gradual shift in gastric microbiota profile from NAG to IM to GC.

Genome Sequence of a Helicobacter pylori Strain Isolated from a Mexican Patient with Intestinal Gastric Cancer.

Helicobacter pylori strains are the major risk factor for gastric cancer. Strains vary in their content of disease-associated genes, so genome-wide analysis of cancer-isolated strains will help elucidate their pathogenesis and genetic diversity. We present the draft genome sequence of H. pylori isolated from a Mexican patient with intestinal gastric cancer.

Genetic analysis of Helicobacter pylori strain populations colonizing the stomach at different times postinfection.

Genetic diversity of the human gastric pathogen Helicobacter pylori in an individual host has been observed; whether this diversity represents diversification of a founding strain or a mixed infection with distinct strain populations is not clear. To examine this issue, we analyzed multiple single-colony isolates from two to four separate stomach biopsies of eight adult and four pediatric patients from a high-incidence Mexican population. Eleven of the 12 patients contained isolates with identical random amplified polymorphic DNA, amplified fragment length polymorphism, and vacA allele molecular footprints, whereas a single adult patient had two distinct profiles. Comparative genomic hybridization using whole-genome microarrays (array CGH) revealed variation in 24 to 67 genes in isolates from patients with similar molecular footprints. The one patient with distinct profiles contained two strain populations differing at 113 gene loci, including the cag pathogenicity island virulence genes. The two strain populations in this single host had different spatial distributions in the stomach and exhibited very limited genetic exchange. The total genetic divergence and pairwise genetic divergence between isolates from adults and isolates from children were not statistically different. We also analyzed isolates obtained 15 and 90 days after experimental infection of humans and found no evidence of genetic divergence, indicating that transmission to a new host does not induce rapid genetic changes in the bacterial population in the human stomach. Our data suggest that humans are infected with a population of closely related strains that vary at a small number of gene loci, that this population of strains may already be present when an infection is acquired, and that even during superinfection genetic exchange among distinct strains is rare.

Intensity of inflammation, density of colonization and interleukin-8 response in the gastric mucosa of children infected with Helicobacter pylori.

BACKGROUND: Few reports exist on inflammation and interleukin (IL)-8 response in H. pylori-infected children. The aim of this study was to determine the intensity of inflammation, density of colonization and magnitude of IL-8 response in children with and without H. pylori infection. MATERIALS AND METHODS: We studied 45 children with dyspeptic symptoms, 21 infected with H. pylori and 24 without infection. Antrum and corpus gastric biopsies were obtained and studied for H. pylori infection with an immunofluorescence technique and for IL-8 with an immunohistochemical assay. Biopsy specimens were stained with hematoxilin and eosin and gastritis was graded according to the Sydney system. The magnitudes of the IL-8 response and H. pylori colonization were estimated microscopically with image analyzer software. RESULTS: In H. pylori-infected children, mild mono-nuclear cell infiltration was found in 50%, and no neutrophils in 40% of cases. In the antrum but not in the corpus, the intensity of colonization correlated with neutrophil and mononuclear cell infiltration. The IL-8 response was significantly higher in the antrum (p <.05) and corpus (p <.02) of infected children, and was localized mainly in the surface and crypts of the epithelium. No correlation was found between the magnitude of the IL-8 response and the infiltration of either neutrophil or mononuclear cells. CONCLUSIONS: In H. pylori-infected children, poor mononuclear and neutrophil infiltration was observed. Infection was associated with a higher IL-8 response by gastric epithelial cells. The density of colonization but not the IL-8 response correlated with neutrophil cell infiltration.

Evolution of the Helicobacter pylori vacuolating cytotoxin in a human stomach.

We describe two subclones of Helicobacter pylori, isolated contemporaneously from a human stomach, which differ markedly in the vacuolating cytotoxin gene, vacA, but whose near identity in sequences outside this locus implies a very recent common origin. The differences are consistent with homologous recombination with DNA from another strain and result in a changed vacA midregion and, importantly, in changed toxicity.