Bile enhances cell surface hydrophobicity and biofilm formation of bifidobacteria.

Twenty-four human bifidobacterial strains were analysed for cell surface hydrophobicity (CSH) using a salt aggregation test (SAT) and a Congo red binding (CRB) assay. Three strains were selected for a systematic study on the CSH and biofilm formation: Bifidobacterium breve 46, Bifidobacterium animalis ssp. lactis 8:8 and a reference strain B. animalis ssp. lactis JCM 10602. CRB of the B. breve 46 and B. animalis ssp. lactis JCM 10602 was significantly enhanced (P < 0.05) when grown in deMan-Rogosa-Sharpe cysteine (MRSC) broth supplemented with taurocholic acid (TA) or native porcine bile (PB). An enhanced CSH of the strains grown with PB and gastric mucin correlated with an increased mucin binding and an enhanced biofilm formation in prebiotic oligosaccharide-supplemented cultures. The three strains showed late bile-induced biofilm (72 h) under an anaerobic growth condition, and both B. animalis ssp. lactis strains showed a late bile-induced biofilm formation under aerobic conditions shown by crystal violet staining. These two strains were thus considered to be oxygen tolerant and more robust. Furthermore, enhanced biofilm formation of these robust bifidobacterial strains in the presence of prebiotics may allow for strong colonisation in the gastrointestinal tract when administered to in vivo models as a "synbiotic supplement".

Enterococcus faecalis infection causes inflammation, intracellular oxphos-independent ROS production, and DNA damage in human gastric cancer cells.

BACKGROUND: Achlorhydria caused by e.g. atrophic gastritis allows for bacterial overgrowth, which induces chronic inflammation and damage to the mucosal cells of infected individuals driving gastric malignancies and cancer. Enterococcus faecalis (E. faecalis) can colonize achlohydric stomachs and we therefore wanted to study the impact of E. faecalis infection on inflammatory response, reactive oxygen species (ROS) formation, mitochondrial respiration, and mitochondrial genetic stability in gastric mucosal cells. METHODS: To separate the changes induced by bacteria from those of the inflammatory cells we established an in vitro E. faecalis infection model system using the gastric carcinoma cell line MKN74. Total ROS and superoxide was measured by fluorescence microscopy. Cellular oxygen consumption was characterized non-invasively using XF24 microplate based respirometry. Gene expression was examined by microarray, and response pathways were identified by Gene Set Analysis (GSA). Selected gene transcripts were verified by quantitative real-time polymerase chain reaction (qRT-PCR). Mitochondrial mutations were determined by sequencing. RESULTS: Infection of MKN74 cells with E. faecalis induced intracellular ROS production through a pathway independent of oxidative phosphorylation (oxphos). Furthermore, E. faecalis infection induced mitochondrial DNA instability. Following infection, genes coding for inflammatory response proteins were transcriptionally up-regulated while DNA damage repair and cell cycle control genes were down-regulated. Cell growth slowed down when infected with viable E. faecalis and responded in a dose dependent manner to E. faecalis lysate. CONCLUSIONS: Infection by E. faecalis induced an oxphos-independent intracellular ROS response and damaged the mitochondrial genome in gastric cell culture. Finally the bacteria induced an NF-κB inflammatory response as well as impaired DNA damage response and cell cycle control gene expression. TRANSCRIPT PROFILING: Array Express accession number E-MEXP-3496.

Effect of heparin, fucoidan and other polysaccharides on adhesion of enterohepatic helicobacter species to murine macrophages.

Helicobacter species have been isolated and cultured from both the gastric and enterohepatic niches of the gastrointestinal tract and are associated with a wide spectrum of diseases. Some members of the enterohepatic Helicobacter species (EHS), which include Helicobacter bilis, Helicobacter hepaticus and Helicobacter pullorum, are associated with chronic inflammatory and proliferative bowel inflammation, hepatitis and in experimental murine studies with hepatic cancer. The present study aimed to explore if polysulphated polysaccharides can prevent adhesion of EHS to the murine macrophage cell line J774A.1. A competitive binding assay showed that heparin and heparan sulphate at a concentration of 1.25 mg/ml reduced binding of H. hepaticus and H. pullorum to the host cells, but not H. bilis. Of the tested Helicobacter spp, the highest inhibition by heparin was demonstrated for H. pullorum (P < 0.01), the most hydrophilic strain. Partially or completely de-sulphated heparin derivatives lost the ability to inhibit adherence of EHS, indicating the importance of sulphated groups of heparin. The most efficient inhibitor of EHS binding to macrophages was fucoidan, which reduced bacterial adhesion of the three enterohepatic Helicobacter species to a greater extent than heparin, 60-90% inhibition vs 30-70% inhibition by heparin. Identification of receptors that EHS ligands bind to is important for understanding the development of infection and may provide a rational target to prevent infection and therapy.

Helicobacter species and common gut bacterial DNA in gallbladder with cholecystitis.

AIM: To analyze the association between Helicobacter spp. and some common gut bacteria in patients with cholecystitis. METHODS: A nested-polymerase chain reaction (PCR), specific to 16S rRNA of Helicobacter spp. was performed on paraffin-embedded gallbladder samples of 100 cholecystitis and 102 control cases. The samples were also analyzed for some common gut bacteria by PCR. Positive samples were sequenced for species identification. RESULTS: Helicobacter DNA was found in seven out of 100 cases of acute and chronic cholecystitis. Sequence analysis displayed Helicobacter pullorum (H. pullorum) in six cases and Helicobacter pylori in one; H. pullorum was only found in cases with metaplasia. Control samples were negative for Helicobacter spp. and some common gut bacteria. There was a significant difference (P = 0.007) between cholecystitis and control samples for Helicobacter DNA. CONCLUSION: A possible relationship was detected between Helicobacter DNA and cholecystitis. Further serological and immunohistochemical studies are needed to support these data.

Detection of Helicobacter rodentium-like DNA in the liver tissue of patients with chronic liver diseases by polymerase chain reaction-denaturing gradient gel electrophoresis and DNA sequence analysis.

Many Helicobacter spp. were isolated from the stomach, intestinal tract, and liver of different animals and humans. The association between Helicobacter spp. and hepatobiliary diseases, including hepatocellular carcinoma, was thoroughly examined, indicating a potential role of the bacteria in the progression toward cancer. In our work, we screened 97 liver biopsies from patients with chronic liver diseases for the presence of Helicobacter spp. DNA. With the use of genus-specific polymerase chain reaction (PCR)-denaturing gradient gel electrophoresis (DGGE) and subsequent sequencing, we found that the majority of Helicobacter spp. DNA detected was similar to Helicobacter rodentium DNA (71%). The DNA of other detected Helicobacter spp. was similar to Helicobacter pylori DNA. This is the first indication of H. rodentium-like DNA presence in human liver tissue. We also conclude that PCR-DGGE is a useful screening method for assigning species designation and heterogeneity.

Strong lymphoid nuclear expression of SOX11 transcription factor defines lymphoblastic neoplasms, mantle cell lymphoma and Burkitt's lymphoma.

BACKGROUND: We surveyed lymphomas to determine the range of expression of the mantle cell lymphoma-associated SOX11 transcription factor and its relation to cyclin D1. DESIGN AND METHODS: On hundred and seventy-two specimens were immunostained for the SOX11 N and C termini. Cyclin D1 was detected by immunohistochemistry and quantitative reverse transcriptase polymerase chain reaction; in situ hybridization for t(11;14) was applied when needed. RESULTS: Nuclear SOX11 was strongly expressed in most B and T-lymphoblastic leukemia/lymphomas and half of childhood Burkitt's lymphomas, but only weakly expressed in some hairy cell leukemias. Chronic lymphocytic leukemia/lymphoma, marginal zone, follicular and diffuse large B-cell lymphomas were negative for SOX11, as were all cases of intermediate Burkitt's lymphomas/diffuse large B-cell lymphoma, myeloma, Hodgkin's lymphomas and mature T-cell and NK/T-cell lymphomas. CONCLUSIONS: In addition to mantle cell lymphoma, SOX11 is strongly expressed only in lymphoblastic malignancies and Burkitt's lymphomas. Its expression is independent of cyclin D1 (except for weak expression in hairy cell leukemias) and unlikely to be due to translocations in lymphoid neoplasia.

Cross-reactivity between immune responses to Helicobacter bilis and Helicobacter pylori in a population in Thailand at high risk of developing cholangiocarcinoma.

Helicobacter bilis DNA has been detected in human tissue and is a candidate for etiologic investigations on the causes of hepatic and biliary tract diseases, but reliable serologic tests need to be developed in order to pursue such investigations. The scope of this study was to assess the specificity of two assays for H. bilis immune response allowing for H. pylori, and their cross-reactivity in a population in Thailand at high risk for cholangiocarcinoma. Plasma samples from 92 Thai volunteers were independently tested in two laboratories (Massachusetts Institute of Technology [MIT] and Lund). MIT performed three analyses of H. pylori and H. bilis based either on (i) outer membrane protein (OMP) with no preabsorption or on antigens derived from whole-cell sonicate before (ii) or after (iii) preabsorption with H. pylori sonicate protein. Lund used cell surface proteins from H. pylori and H. bilis as antigens. Testing for H. bilis was preabsorbed with a whole-cell lysate of H. pylori. More than 80% of the samples were positive for H. pylori in both laboratories. As tested by MIT, 58.7% (95% confidence interval, 47.9 to 68.9%) were positive for H. bilis by OMP and 44.5% (34.1 to 55.3%) were positive for H. bilis sonicate protein, but only 15.2% (8.6 to 24.2%) remained positive after preabsorption with H. pylori sonicate protein. Lund found 34.5% of the samples positive for H. bilis (22.0 to 41.0%), which was statistically compatible with all three MIT results. Serologic responses to OMPs of the two bacteria coincided in 66 and 45% of the samples in the MIT and Lund assays, respectively. We found high cross-reactivity between the immune responses to H. pylori and H. bilis antigens. More-specific H. bilis antigens need to be isolated to develop serologic tests suitable for epidemiological studies.

Determination of lamivudine-resistant variants of hepatitis B virus by denaturing gradient gel electrophoresis: a novel approach to monitoring drug resistance.

BACKGROUND: A DGGE-based assay for hepatitis B virus (HBV) drug-resistance monitoring was designed and checked for feasibility. It detects mutations within the YMDD motif related to lamivudine resistance. MATERIAL/METHODS: The YMDD motif of HBV polymerase was amplified by the set of primers designed in this study. DGGE analysis of the amplicons was performed on 9% polyacrylamide gels containing a 20-40% gradient of urea plus formamide and electrophoresis was performed. DNA sequencing was performed using a standard protocol. RESULTS: Based on the DGGE pattern of previously sequenced HBV variants, a reference ladder consisting of bands was constructed within and near the YMDD motif of HBV with excellent resolution. The genotypes of all the fragments included in the ladder were confirmed by sequencing after DGGE analysis. The flexibility of DGGE was demonstrated by the ability to add more bands to the migration ladder when new variants were discovered during the analysis of patient specimens. Clinical samples from HBV-infected patients were also used to demonstrate the performance of this approach. CONCLUSIONS: This preliminary feasibility study of HBV drug-resistance monitoring by means of DGGE shows the potential advantage of this approach for low-cost screening for viral drug resistance in clinical settings. The presented example can be extended to detect other mutations related to drug resistance in the HBV genome as well as other viruses.

The effect of different strains of Helicobacter pylori on platelet aggregation.

BACKGROUND AND AIMS: Helicobacter pylori is the major causative agent in peptic ulcer disease and is strongly implicated in the development of gastric cancer. It has also been linked, less strongly, to cardiovascular disease. The mechanisms by which certain strains of H pylori induce platelet aggregation through interactions with platelet glycoprotein Ib have been previously described. METHODS: In the present study, 21 different strains of H pylori, varying in their vacuolating toxin gene, cytotoxic-associated gene A status and other pathogenicity factors, were tested for their ability to induce platelet aggregation. RESULTS: Ten of the 21 strains induced platelet aggregation, a response that appeared to be independent of their vacuolating toxin gene and cytotoxic-associated gene A status. CONCLUSIONS: Platelet aggregation has been suggested to be one of the possible mechanisms involved in the effects on the cardiovascular system induced by H pylori. Our results suggest that any putative role H pylori plays in cardiovascular disease may be strain dependent. Further work to identify the H pylori factors involved in induction of platelet aggregation may allow for identification of 'higher risk' strains for cardiovascular disease.

Does Helicobacter pylori infection per se cause gastric cancer or duodenal ulcer? Inadequate evidence in Mongolian gerbils and inbred mice.

A role for Helicobacter pylori infection in the development of gastric cancer in humans is well established; however, evidence for its carcinogenicity in animals remains inadequate. Mongolian gerbils and mice are commonly used to investigate the carcinogenicity of H. pylori, yet it is unclear whether H. pylori infection per se causes gastric cancer or duodenal ulcers in these animal models. Gastric adenocarcinoma in the gerbils was reported over 10 years ago, but this species has proved an unreliable model for studying H. pylori infection-associated gastric cancer. Helicobacter pylori infection alone appears insufficient to induce gastric cancer in these animals; additional carcinogenic insult is required. The development of invasive adenocarcinoma in inbred mice is rare regardless of the mouse or bacterial strain, and many long-term studies have failed to induce gastric cancer in these animals. Helicobacter pylori infection is also an established causative factor for duodenal ulcer in humans. However, few studies have attempted to develop animal models of H. pylori infection-induced duodenal ulcer. We therefore conclude that both Mongolian gerbils and inbred mice may be inadequate models for studying H. pylori infection-associated gastric cancer and that there is no animal model of H. pylori infection-induced duodenal ulcer.

Combined serum IgG response to Helicobacter pylori VacA and CagA predicts gastric cancer.

Helicobacter pylori is a major factor for the development of gastric cancer. The aim of this study was to define serum antibody patterns associated with H. pylori infection in patients with gastric cancer using a Western blot technique. Serum samples collected from 115 patients with gastric cancer and 110 age- and gender-matched patients without gastrointestinal diseases were tested for IgG antibodies to H. pylori antigens (outer membrane proteins and whole cell preparations). No significant differences were found between patients with and without gastric cancer using outer membrane proteins (82% and 73%, P>0.05) or whole cell antigens (84% and 76%, P>0.05), respectively. The significant differences between patients with and without gastric cancer were associated with bands of 94 kDa (54% and 20%, P<0.001) and 30 kDa (65% and 44%, P<0.01). A combination of antibodies to 85 kDa (VacA) and 120 kDa (CagA) was significantly (P<0.01) more frequent in gastric cancer patients than in patients without gastric cancer. The detection of antibodies to 94- and 30-kDa bands, in association with the determination of serum antibodies to CagA+/VacA+, may have a prospective value in assessment of the risk of developing of gastric cancer.

Gastric inflammatory markers and interleukins in patients with functional dyspepsia treated with astaxanthin.

The chronic active inflammation caused by Helicobacter pylori is dominated by neutrophils, macrophages, lymphocytes and plasma cells. Several interleukins are involved in the inflammatory process. The aim of this study was to investigate the effect of astaxanthin on gastric inflammation in patients with functional dyspepsia. Forty-four consecutive patients were included, and biopsies were examined for IL-4, IL-6, IL-8, IL-10, interferon-gamma, CD4, CD8, CD14, CD19, CD25 and CD30. Patients were randomized: 21 patients were treated with 40 mg of astaxanthin daily, and 23 patients were treated with a placebo. There was a significant decrease in gastric inflammation in H. pylori-positive patients from both groups. There were no significant changes in the density of H. pylori or in any of the interleukins during or after treatment. There was a significant up-regulation of CD4 and down-regulation of CD8 in patients with H. pylori treated with astaxanthin. Astaxanthin had an effect on the inflammation and on the density of H. pylori in mice in a study where the diet could be standardized without antioxidants (Bennedsen et al., 1999). These dietary conditions are impossible in studies involving humans, and may be due to the minor effect when the host have access to antioxidants in their diet.

Swedish moist snuff accelerates gastric cancer development in Helicobacter pylori-infected wild-type and gastrin transgenic mice.

The Swedish variant of moist oral smokeless tobacco (snus) is popular in Sweden and Norway, banned from sale within the European Union and is currently being introduced in USA. The aim of the present study was to determine if snus is carcinogenic to the stomach, particularly in Helicobacter pylori (H.P.)-infected hosts at increased risk for gastric cancer development. Snus (Generaltrade mark; Swedish Match, Sweden) was mixed with powdered standard mouse chow at a concentration of 5-9% (wt/wt) and given to wild-type (WT, FVB) and gastrin transgenic (INS-GAS, FVB) mice for 6 months with or without H.P. (strain 67:21, CagA+, VacA+) infection. At necropsy, pathological evaluation of stomachs from uninfected snus-treated WT mice showed mild morphological changes, whereas 50% snus-treated INS-GAS mice developed carcinoma in situ (CIS), compared with 25% not exposed to snus. When snus was given to H.P.-infected mice, 9 of 17 WT mice developed CIS with intramucosal invasion, and the remaining 8 of 17 WT mice developed high-grade dysplasia (score >1.5) that was associated with increased gastritis, epithelial defects, oxyntic atrophy, hyperplasia and intestinal metaplasia. Twelve of 12 H.P.-infected INS-GAS mice developed CIS with intramucosal invasion and submucosal herniation. We suggest that snus is a potential gastric carcinogen in mice. The development of CIS was associated with increased rates of the epithelial cell proliferation and apoptosis, common features of gastric carcinogenesis.

Role of Helicobacter pylori in conjunctival mucosa-associated lymphoid tissue lymphoma.

OBJECTIVE: Conjunctiva-associated lymphoid tissue is the conjunctival equivalent to mucosa-associated lymphoid tissue (MALT). Mucosa-associated lymphoid tissue lymphoma has been shown to be associated with Helicobacter pylori. In this study, the prevalence and possible role of H. pylori infection in conjunctival MALT lymphoma were evaluated. DESIGN: Retrospective noncomparative case series. PARTICIPANTS: Thirteen cases of conjunctival MALT lymphoma were investigated. Five samples of conjunctival lymphoid hyperplasia and 20 biopsies of normal conjunctiva served as controls. METHODS: The specimens were investigated for the presence of H. pylori with immunohistochemistry (IHC) and nested polymerase chain reaction (PCR) techniques. For each case of conjunctival MALT lymphoma, information regarding gender, age at presentation, conjunctival localization, and information of generalized MALT lymphoma were collected. MAIN OUTCOME MEASURES: Detection of H. pylori and patient characteristics. RESULTS: The 13 conjunctival MALT lymphomas originated from 8 women and 5 men with an average age of 62 years (range, 25-87). Only 1 patient had evidence of systemic MALT lymphoma. H. pylori could not be identified in any of the conjunctival MALT lymphomas, in conjunctival lymphoid hyperplasia, or in normal conjunctival biopsies using IHC and PCR techniques. CONCLUSIONS: An association between H. pylori and localized conjunctival MALT lymphoma could not be verified. Antigens other than H. pylori may take part in the development of conjunctival MALT lymphoma.

SabA is the H. pylori hemagglutinin and is polymorphic in binding to sialylated glycans.

Adherence of Helicobacter pylori to inflamed gastric mucosa is dependent on the sialic acid-binding adhesin (SabA) and cognate sialylated/fucosylated glycans on the host cell surface. By in situ hybridization, H. pylori bacteria were observed in close association with erythrocytes in capillaries and post-capillary venules of the lamina propria of gastric mucosa in both infected humans and Rhesus monkeys. In vivo adherence of H. pylori to erythrocytes may require molecular mechanisms similar to the sialic acid-dependent in vitro agglutination of erythrocytes (i.e., sialic acid-dependent hemagglutination). In this context, the SabA adhesin was identified as the sialic acid-dependent hemagglutinin based on sialidase-sensitive hemagglutination, binding assays with sialylated glycoconjugates, and analysis of a series of isogenic sabA deletion mutants. The topographic presentation of binding sites for SabA on the erythrocyte membrane was mapped to gangliosides with extended core chains. However, receptor mapping revealed that the NeuAcalpha2-3Gal-disaccharide constitutes the minimal sialylated binding epitope required for SabA binding. Furthermore, clinical isolates demonstrated polymorphism in sialyl binding and complementation analysis of sabA mutants demonstrated that polymorphism in sialyl binding is an inherent property of the SabA protein itself. Gastric inflammation is associated with periodic changes in the composition of mucosal sialylation patterns. We suggest that dynamic adaptation in sialyl-binding properties during persistent infection specializes H. pylori both for individual variation in mucosal glycosylation and tropism for local areas of inflamed and/or dysplastic tissue.

Lactic acid bacteria as probiotics.

A number of Lactobacillus species, Bifidobacterium sp, Saccharomyces boulardii, and some other microbes have been proposed as and are used as probiotic strains, i.e. live microorganisms as food supplement in order to benefit health. The health claims range from rather vague as regulation of bowel activity and increasing of well-being to more specific, such as exerting antagonistic effect on the gastroenteric pathogens Clostridium difficile, Campylobacter jejuni, Helicobacter pylori and rotavirus, neutralising food mutagens produced in colon, shifting the immune response towards a Th2 response, and thereby alleviating allergic reactions, and lowering serum cholesterol (Tannock, 2002). Unfortunately, most publications are case reports, uncontrolled studies in humans, or reports of animal or in vitro studies. Whether or not the probiotic strains employed shall be of human origin is a matter of debate but this is not a matter of concern, as long as the strains can be shown to survive the transport in the human gastrointestinal (GI) tract and to colonise the human large intestine. This includes survival in the stressful environment of the stomach - acidic pH and bile - with induction of new genes encoding a number of stress proteins. Since the availability of antioxidants decreases rostrally in the GI tract production of antioxidants by colonic bacteria provides a beneficial effect in scavenging free radicals. LAB strains commonly produce antimicrobial substance(s) with activity against the homologous strain, but LAB strains also often produce microbicidal substances with effect against gastric and intestinal pathogens and other microbes, or compete for cell surface and mucin binding sites. This could be the mechanism behind reports that some probiotic strains inhibit or decrease translocation of bacteria from the gut to the liver. A protective effect against cancer development can be ascribed to binding of mutagens by intestinal bacteria, reduction of the enzymes beta-glucuronidase and beta-glucosidase, and deconjugation of bile acids, or merely by enhancing the immune system of the host. The latter has attracted considerable interest, and LAB have been tested in several clinical trials in allergic diseases. Characteristics ascribed to a probiotic strain are in general strain specific, and individual strains have to be tested for each property. Survival of strains during production, packing and storage of a viable cell mass has to be tested and declared.

Gastric inflammation, metaplasia, and tumor development in gastrin-deficient mice.

BACKGROUND & AIMS: Gastrin deficiency and proton pump inhibitor treatment cause achlorhydria, which predisposes to disease. To elucidate the underlying molecular biology, we examined the changes in gastric gene expression in both types of achlorhydria. We also explored the associated changes in the gastric microflora and the long-term consequences of gastrin-deficient achlorhydria. METHODS: Expression profiles were generated from gastric RNA from wild-type mice, gastrin knockout (KO) mice, gastrin KO mice after 1 week of gastrin infusion, and wild-type mice treated for 1 month with a proton pump inhibitor. The results were confirmed using real-time polymerase chain reaction and immunohistochemistry. Selective media were used to characterize the gastric microflora. RESULTS: The number of gastric bacteria was increased in both gastrin KO and PPI-treated mice. The expression profiles revealed activation of immune defense genes, interferon-regulated response genes, and intestinal metaplasia of the gastric mucosa. In young gastrin-deficient mice, gastrin infusions reversed the changes. Over time, the changes accumulated, became irreversible, and progressed into metaplasia and polyp development. Finally, the study showed that gastrin regulated the expression of genes encoding extracellular matrix proteins. CONCLUSIONS: Independently of gastrin, achlorhydria is associated with gastric bacterial overgrowth and intestinal gene expression patterns and is associated with predisposition to disease. Gastrin is therefore essential for prevention of gastric disease, mainly through control of acid secretion but to a lesser extent also through control of gastric gene expression. The gastrin-deficient mouse serves as a useful new model for gastric metaplasia and neoplasia.

Chronic Helicobacter pylori infection in a population in southern Sweden analysed by histopathology, immunoblot and ELISA serology.

BACKGROUND: Many individuals are infected with the bacterium Helicobacter pylori. Some develop ulcers or mucosal atrophy. AIMS: To correlate the histological characteristics of the H. pylori -induced gastritis to the immunoblot pattern of the H. pylori infection and to compare the presence of H. pylori bacteria in tissue specimens with ELISA serology and immunoblot analysis. METHODS: One hundred and sixty-six consecutive patients were referred to gastroscopy. Forty patients were excluded for various reasons and 126 were included in the study. RESULTS: Twenty-three patients had ulcerations and 25 erosions. Ninety-two (73%) had a chronic gastritis and in 90 (71%) it involved both the antrum and corpus. Ninety-one (72%), of whom 96% had a chronic gastritis, had visible bacteria in the tissue specimens, used as the 'gold standard' for the detection of infection. In patients with chronic gastritis 65 (70%) had positive H. pylori ELISA serology, 27 (30%) had negative H. pylori ELISA, while 76 (83%) had a positive immunoblot pattern. The ELISA positive patients had more advanced chronic gastritis but a lower frequency of metaplasia and atrophy. Acute inflammatory activity in the chronic gastritis had a high immunoreactivity to 120 kDa (CagA) protein and was significantly correlated to antibody reactivity to proteins in the 53-65 kDa range (heat shock proteins) and to a 43 kDa subunit. Metaplasia and atrophy in antrum was associated with a 62 kDa protein band. CONCLUSION: Almost all H. pylori-infected patients had a pangastritis, visible in both antrum and corpus. Acute inflammatory activity in the chronic gastritis and the presence of metaplasia and atrophy in antrum were associated with a specific immunoblot pattern, indicating infection with more virulent strains. Immunoblot analysis had a better sensitivity than ELISA H. pylori serology.

Helicobacter species ribosomal DNA in the pancreas, stomach and duodenum of pancreatic cancer patients.

AIM: To determine whether gastric and enteric Helicobacter species are associated with pancreatic cancer. METHODS: Patients with exocrine pancreatic cancer (n = 40), neuroendocrine cancer (n = 14), multiple endocrine neoplasia type 1 (n = 8), and chronic pancreatitis (n = 5) were studied. Other benign pancreatic diseases (n = 10) and specimens of normal pancreas (n = 7) were included as controls. Pancreatic tissue specimens were analyzed by Helicobacter-specific PCR-assay and products were characterized by denaturing gradient electrophoresis and DNA-sequencing. From a subset of the pancreatic cancer patients, gastric and/or duodenal tissue as well as gallbladder and ductus choledochus tissue were analyzed. Gallbladder and choledochus samples were included as controls. Stomach and duodenum samples were investigated to analyze whether a gastric helicobacter might disseminate to the pancreas in pancreatic cancer patients. Pancreatic specimens were analyzed by Bacteroides-specific PCR for detecting the translocation of indigenous gut microbes to the diseased pancreas. RESULTS: Helicobacter DNA was detected in pancreas (tumor and/or surrounding tissue) of 75% of patients with exocrine cancer, 57% of patients with neuroendocrine cancer, 38% of patients with multiple endocrine neoplasia, and 60% of patients with chronic pancreatitis. All samples from other benign pancreatic diseases and normal pancreas were negative. Thirty-three percent of the patients were helicobacter-positive in gastroduodenal specimens. Surprisingly, H. bilis was identified in 60% of the positive gastroduodenal samples. All gallbladder and ductus choledochus specimens were negative for helicobacter. Bacteroides PCR-assay was negative for all pancreatic samples. CONCLUSION: Helicobacter DNA commonly detected in pancreatic cancer suggests a possible role of the emerging pathogens in the development of chronic pancreatitis and pancreatic cancer.