Comparison of diagnostic methods for detection of Helicobacter pylori

Of two hundred Ethiopian patients with dyspepsia, multiple biopsies were taken from the antrum of the stomach. Helicobacter pylori was cultured from 85% of duodenal ulcer and in 75% of chronic antral gastritis patients. The overall Helicobacter pylori positivity was 70%. The sensitivity, specificity, positive and negative predictive values of the tests as compared to culture were as follows, respectively: direct urease test 100%/87%/95%/100%, direct gram stain 60%/98%/99%/51%, histological gram stain 66%/97%/98%/56%, Giemsa stain 100%/97%/99%/100% and Gimenez stain 100%/87%/95%/100%. It is concluded that gram staining of direct tissue smear or histology is an insensitive method in the diagnosis of Helicobacter pylori. All the other tests, are shown to be valid. Urease test is an excellent test for provision of presumptive diagnosis of Helicobacter pylori while awaiting confirmation either by culture of histology.     

Comparison of five PCR methods for detection of Helicobacter pylori DNA in gastric tissues

Five different PCR methods for the detection of Helicobacter pylori were evaluated. The results of this study indicate that of the five PCR methods examined, the ureC (glmM) gene PCR is the most sensitive and specific for the detection of H. pylori in gastric biopsy specimens.     

Prospective screening of dyspeptic patients by Helicobacter pylori serology

A 24-year-old Chinese woman presented with cough, chest pain, weight loss, low grade fever and bronchial breath sounds on auscultation. The diagnosis of chronic eosinophilic pneumonia was made on characteristic systemic and pulmonary clinical manifestations, blood eosinophilia and the striking chest radiographic appearance. This rare, idiopathic but benign condition responds well to corticosteroid treatment and the long term prognosis is excellent. The typical chest radiographic pattern of 'photographic negative of pulmonary oedema' in this condition is emphasised.     

Detection methods of Helicobacter pylori: accuracy and costs

A variety of methods exist for determining gastric colonization with Helicobacter pylori, which has been implicated in the development of peptic ulcer disease. The goal of this study was to evaluate four of the current methods available in a clinical surgical practice setting through a prospective evaluation of 40 consecutive patients undergoing upper diagnostic endoscopy. All patients underwent six antral gastric biopsies for use with the following detection methods: histologic demonstration of organisms (hematoxylin and eosin stain), direct detection of urease activity (Remel Selective Rapid Urea, Lenexa, KS), and culture of H. pylori. All patients also had measurement of serum immunoglobulin G for H. pylori by the enzyme-linked immunosorbent assay method (Corning Clinical Laboratories, St. Louis, MO). The infection status was established by a concordance of test results. The results show that H. pylori can be assessed equally well with histology, a rapid urease test, and serology, with all three tests having good sensitivity (92-100%) and specificity (85-96%). The culturing of the organism had poor sensitivity (42%). The benefits of the urease test are a much more rapid response time and a much lower cost as compared to histologic and serologic testing. In conclusion, the rapid urease test is the method of choice to detect H. pylori in those patients undergoing endoscopy in whom the identification of H. pylori will change their management.     

Accuracy of IgG serology and other tests in confirming Helicobacter pylori eradication

BACKGROUND: In this study we assessed the accuracy of IgG serology and other tests in confirming Helicobacter pylori eradication. METHODS: The outcome of anti-H. pylori therapy was established by at least two of the following tests: rapid urease test (RUT), culture, 14C urea breath test (non-capsule or capsule UBT), and IgG serology (Orion Diagnostica Pyloriset New EIA-G). RESULTS: Successful H. pylori eradication was confirmed in 698 of 794 patients (88%). The percentage decrease in IgG antibody titre was related to the patients' pre-treatment IgG titre and time interval after treatment. A decrease in IgG titres of 40% or more confirmed H. pylori eradication with 100% specificity, whereas the sensitivity was 82%, 90%, 98%, and 98% 3, 4, 5, and 6 months after therapy, respectively. The 40% cut-off confirmed eradication 3 to 6 months after therapy in 328 of 339 patients (97%) with pre-treatment IgG titres of >700, in 36 of 45 patients (80%) with pre-treatment titres of 300-700, and in 5 of 12 patients (42%) with pretreatment titres of <300. The sensitivity and specificity of the other tests 2 months after treatment were as follows: RUT, 84% and 100%; culture, 88% and 100%; non-capsule UBT, 100% and 89%; and capsule UBT, 100% and 97%. CONCLUSION: A decrease in IgG antibody titre of 40% or more 3 to 6 months after therapy and the capsule 14C UBT at the 2-month follow-up were both highly accurate in confirming H. pylori eradication.     

Comparison of two rapid urease tests for detection of Helicobacter pylori infection

The selection of NIH 3T3 cells expressing a hydroxytamoxifen-inducible c-Raf-1-estrogen receptor fusion protein (c-Raf-1-BxB-ER) in the absence or presence of the inducer results in dramatic differences in the expression levels of the fusion protein. Hydroxytamoxifen-mediated constitutive activation of the Raf signal favors the selection of cells expressing low levels of c-Raf-1-BxB-ER. Cells selected in the absence of hydroxytamoxifen express up to 20 times higher levels of the inducible Raf kinase. Activation of the oncogenic Raf kinase in cells expressing low levels leads to a weak activation of the Raf/Mek/Erk cascade and the induction of S phase in confluent cells. The activation of cells expressing high levels of the kinase leads to a strong persistent signal and inhibits DNA synthesis and mitosis in proliferating cells. The inhibition of DNA synthesis and cell division is presumably due to the elevated expression of the cyclin-dependent kinase inhibitor p21cip1, similar to cells exposed to ionizing radiation. Despite the inhibition of DNA synthesis and mitosis, the constitutive activity of the Raf signaling pathway is still able to initiate cell growth. Activation of the high-intensity Raf signal in arrested serum-starved cells induces cell growth up to a size corresponding to that of M-phase cells in the absence of DNA synthesis. High-intensity Raf signals in proliferating cells consistently lead to an accumulation of cells with the size of M-phase cells and the DNA content of G1 cells or G2-M-phase cells. Therefore, the activation of Raf kinase is sufficient to drive cell growth, even in the presence of high levels of the cyclin-dependent kinase inhibitor p21cip1.     

Helicobacter pylori detection by polymerase chain reaction in gastric juice and its correlation with the histology (Giemsa)]

HP infection is involved in the pathogenesis of several gastroduodenal diseases, as type B chronic gastritis, duodenal and gastric ulcer, MALT lymphoma and gastric cancer. The recent availability of molecular techniques, specifically the PCR, allow us to detect very low amounts of the bacterium. The aim of the study is to evaluate the presence of HP in gastric juice by PCR technique and to correlate this findings with histology (Giemsa) of gastric mucosa. Gastric juice PCR positive findings were found in 10/31 (32.3%) HP positive patients at histology. We concluded that HP in gastric juice is possible to detect by molecular techniques. In our study 32.3% of the patients showed the presence of HP in gastric juice.     

Positive result by serology indicates active Helicobacter pylori infection in patients with atrophic gastritis

Patients with atrophic corpus gastritis and elevated Helicobacter pylori antibody titers but 13C-urea breath test (13C-UBT) and histology results negative for H. pylori were randomized into eradication therapy or follow-up only. Antibody levels decreased significantly in six out of seven patients in the eradication group, while in the follow-up group, the titers declined in only one out of eight patients. In patients with atrophic corpus gastritis, positive serology results may indicate an ongoing infection in spite of negative 13C-UBT and histology results.     

Acid, protons and Helicobacter pylori

The anti-ulcer drugs that act as covalent inhibitors of the gastric acid pump are targeted to the gastric H+/K+ ATPase by virtue of accumulation in acid and conversion to the active sulfenamide. This results in extremely effective inhibition of acid secretion. Appropriate dosage is able to optimize acid control therapy for reflux and peptic ulcer disease as compared to H2 receptor antagonists. However, clinical data on recurrence show that Helicobacter pylori eradication should accompany treatment of the lesion. These drugs have been found to synergize with many antibiotics for eradication. The survival of aerobes depends on their ability to maintain a driving force for protons across their inner membrane, the sum of a pH and potential difference gradient, the protonmotive force (pmf). The transmembrane flux of protons across the F1F0 ATPase, driven by the pmf, is coupled to the synthesis of ATP. The internal pH of H. pylori was measured using the fluorescent dye probe, BCECF, and the membrane potential defined by the uptake of the carbocyanine dye, DiSC3 [5] at different pHs to mimic the gastric environment. The protonmotive force at pH 7.0 was composed of a delta pH of 1.4 (-84mV) and a delta potential difference of -131mV, to give a pmf of -215 mV. The effect of variations in external pH on survival of the bacteria in the absence of urea correlated with the effect of external pH on the ability of the bacteria to maintain a pmf. The effect of the addition of 5 mM urea on the pmf was measured at different medium pH values. Urea restored the pmf at pH 3.0 or 3.5, but abolished the pmf at pH 7.0 or higher, due the production of the alkalinizing cation, NH3. Hence H. pylori is an acid-tolerant neutrophile due to urease activity, but urease activity also limits its survival to an acidic environment. These data help explain the occupation of the stomach by the organism and its distribution between fundus and antrum. This distribution and its alteration by proton pump inhibitors also explains the synergism of proton pump inhibition and antibiotics such as amoxicillin and clarithromycin in H. pylori eradication.     

Viral serology and detection

Viral detection is an important part of clinical hepatology. For many years practical clinical tests have been serological but recently newer molecular techniques have become available for virus detection, although these have yet to become routine and some, such as PCR of viral nucleic acid in blood or tissue are not yet consistently reliable. Serology remains the mainstay at present for routine diagnosis. Hepatitis A testing in clinical practice is entirely serological, the IgM response representing acute infection and the IgG response immunity, although more sophisticated molecular techniques have been applied experimentally. A second agent of epidemic enteral hepatitis, the hepatitis E virus, has recently been cloned and sequenced and serological tests for this virus are available, although experience in their use is necessarily limited and a commercial IgM assay has yet to be produced. Serological tests for the hepatitis B virus are well developed. The IgM anticore response differentiates acute infection from chronic, the latter being characterized by the persistence of hepatitis B surface antigen for over six months. Chronic carriers are at risk of liver damage and this risk is best assessed by the amount of viral DNA circulating, which can be determined using a hybridization assay. More sensitive techniques such as the branched chain DNA assay or PCR can detect lower levels of viral DNA but their clinical relevance remains to be established. The hepatitis D virus is defective and relies on hepatitis B to replicate. Serology for antibody and antigen is well established although PCR for circulating viral genome may come to supplant hepatic viral antigen as a test for hepatitis D replication. For hepatitis C serology is feasible only for antibodies, not antigens; although early tests were prone both to false positives and false negatives, current versions are more reliable. PCR has been much used for detection of hepatitis C RNA in blood and tissues and a bDNA assay is now commercially available. Cytomegalovirus detection is confounded by the problem of distinguishing asymptomatic viral replication from disease. Serology is helpful, especially in primary infections, but viral culture is a widely used method. PCR (especially quantitative modifications) or the pp65 antigenaemia assay are experimental approaches which may prove specific enough for general use.     

Helicobacter pylori: the mouth, stomach, and gut axis

The aim of this study was to identify the natural reservoir and route of transmission of Helicobacter pylori infection. Two hundred eight (208) dyspeptic patients (114 males, 94 females; peak age of cohort, 50-59.9) were recruited. Specimens were collected from saliva, supra- and subgingival dental plaque, tongue scrapings, and oropharyngeal swabs. At subsequent endoscopy, gastric antral biopsy was performed for the rapid urease test (RUT), microbiological culture, and, in some patients, histology. Gastric juice samples were aspirated, and in 50 patients duodenal aspirate was collected. Polymerase chain reaction (PCR) with primers targeted to the 16S rRNA sequence of H. pylori was also employed for each of the specimens. In those patients where H. pylori was detected from multiple sites (dental plaque, gastric juice, gastric biopsy, and duodenal aspirate), restriction endonuclease digestion with Hae III was performed to determine if they were epidemiologically linked. The results indicated that 15/208 patients (7%) tested positively for H. pylori by PCR in dental plaque; only 2 samples were positive by culture. In none of the other oral sites sampled was H. pylori detected by any test used in the study. Gastric juice and gastric biopsy specimens from 36/ 208 patients (17%) and 114/208 patients (55%), respectively, were positive by PCR. Duodenal aspirate from 6/50 patients (12%) also tested positively by PCR. All specimens tested by restriction endonuclease digestion with Hae III (15/15 patients) were positive in both antral biopsy and gastric juice specimens, as well as 5 specimens from the duodenal aspirate. Four of the dental plaque strains had restriction patterns similar to those of the stomach and duodenal sites, providing evidence that these sites were infected with the same strain of H. pylori. In conclusion, the results suggest that H. pylori selects the gastric mucosa as its preferred site. The detection in dental plaque could indicate that the oral cavity may act as a reservoir or sanctuary for the organism. Whether H. pylori is a resident or transient oral microorganism is still unclear, although it is more likely to be transient in nature.     

Value of serology as a noninvasive method for evaluating the efficacy of treatment of Helicobacter pylori infection

The systemic humoral response to Helicobacter pylori was studied in 86 infected adult patients before antimicrobial therapy and at intervals following therapy. Endoscopy with collection of biopsy specimens was performed immediately before treatment; a 13C-labeled urea breath test was performed, and blood specimens were collected before treatment and at 1, 3, 6, 9, and 12 months after treatment. Serum samples from three patient groups (eradication success [n = 50], eradication failure [n = 16], and no treatment [n = 20]) were assayed for IgA and IgG antibodies to H. pylori by enzyme-linked immunosorbent assay. Levels of antibody to H. pylori before treatment were similar in all three groups. As expected, the no treatment and eradication failure groups had no significant changes in antibody levels during the study period. In contrast, for the eradication success group, the specific IgA and IgG antibody levels decreased progressively and significantly. We conclude that serology is a potentially useful way to monitor the success of treatment of H. pylori infection without using invasive or more expensive methods.     

Duodenal ulcer, Helicobacter pylori, and gastric secretion

Patients with chronic dyspepsia were categorised by macroscopic appearance at oesophagogastroduodenoscopy as having duodenal ulceration (DU), other diagnosed lesions such as reflux oesophagitis, carcinoma of stomach, etc, or no organic lesion (non-ulcer dyspepsia, NUD). Material was collected to identify gastric infection with Helicobacter pylori (H pylori) by CP urease test, culture, and histological examination and to make the microscopic diagnosis of active chronic gastritis. Each patient in the DU and NUD categories was then invited to volunteer for a gastric secretion study in which maximal gastric secretion in response to histamine was measured. Sixty two gastric secretion tests were performed (31 DU, 31 NUD). The presence of H pylori was associated with active chronic gastritis (100%). DU patients secreted more acid than the NUD patients. H pylori positivity was associated with decreased maximal gastric secretion in both groups. There was a positive correlation between smoking and maximal acid output shown only in H pylori negative but not in H pylori positive patients. These findings were clear cut when all corrections of maximal gastric secretion were made for pyloric loss, duodenogastric reflux, and stature. This study failed to show any aetiological link between H pylori and DU by increased maximal gastric secretion.     

Helicobacter pylori, gastric pathology, and histopathological diagnosis

Structural analogues of ZAPA, Z-3-[(aminoiminomethyl)thio]prop-2-enoic acid, an isothiouronium analogue of GABA, are potent GABAA agonists as seen in the isolated guinea-pig ileum and in the facilitation of [3H]diazepam binding to rat brain membranes. Compounds with guanidino or amidine groups replacing the amino functionality of GABA were also found to be active. The highest activity was displayed by the isothiouronium salts in which the conformational flexibility of the molecule is restricted by a Z-substituted carbon-carbon double bond. A series of bis-isothiouronium compounds was prepared from aliphatic alpha, omega-bis-thioureas as mixtures of E and Z adducts. Maximum GABAA agonist activity for this series was found with a C6-C8 carbon chain, and the results were consistent with an interaction at the GABAA receptor with only one end of the molecule, rather than the more potent effect expected of a molecule bridging two active sites. GABAA antagonist/partial agonist activity was observed on the guinea pig isolated ileum for a number of different analogue types, with the most potent being bis-isothiouronium derivatives. None of the substituted derivatives of ZAPA was as active as ZAPA itself, and maximum GABAA activity was found in the n-pentyl and n-hexyl analogues.     

Evaluation of eight commercial kits for Helicobacter pylori IgG antibody detection

Eight commercial kits and an in-house ELISA for detection of IgG antibodies against Helicobacter pylori were evaluated for their use in diagnosis of H. pylori infection and in epidemiological research: Helico-GTM (Porton-Cambridge), G. A. P. test (Bio-Rad), H. pylori antibodies ELISA (Biometra), Anti-H. pylori IgG EIA (Roche), 2nd generation H. pylori EIA (Roche), Anti-H. pylori MTP-assay (Roche), Pylori stat test kit (Whittaker), Pyloriset latex agglutination kit (Orion), and the in-house ELISA based on heat-stable antigens. Fifty-four patients with dyspepsia (31 H. pylori positive by culture or microscopy) and 68 asymptomatic persons were tested. Sensitivities for the eight kits were 71%, 77%, 90%, 84%, 87%, 94%, 90%, 87%, and 87%, specificities were 74%, 65%, 74%, 74%, 83%, 83%, 70%, 65%, and 65%, respectively. For epidemiological use the estimated seroprevalence varied within approximately 15% in all age groups. Sensitivities and specificities obtained in different studies reveal as great differences in the results with the same kit as between results obtained with different kits in the same study. Kits with the highest sensitivities tend to be the same in all studies. It is therefore more important to test a kit in the population to which it is to be applied than to choose a specific kit.