The capacity for intracellular growth is an important survival strategy for a large group of common pathogens. Helicobacter pylori, the etiological agent for gastritis and duodenal ulcer, has been shown by both in vivo and in vitro studies to have the capacity to invade epithelial cells. In vitro models are used to study the effect of antibiotics on microoganisms. Most investigations are performed in broth culture or on agar plates, but kinetic models for bacteria in broth have been described. We present a new, kinetic model adapted for intracellular pathogens. A glass chamber, with a metal rack fitting Falcon cell culture inserts, was connected to a pump by rubber tubes. Different tube diameters and pump speeds were evaluated, and the assay was designed to mimic the half-lives of the antibiotics in vivo, i.e., 11.5 h for azithromycin, 5 h for clarithromycin, and 1 h for amoxicillin. Monolayers of HEp-2 cells were grown in the inserts for 2 days, after which H. pylori (clinical strain 88-23), was added to the system. Internalization was allowed for 12 h, and extracellular H. pylori cells were eradicated with gentamicin. The inserts were moved to the glass chamber, containing medium with 12.5 mg of either amoxicillin or azithromycin per liter or 2.4 mg of clarithromycin per liter. This represents 12.5, 50, and 80 times the extracellular minimum bactericidal concentration value, respectively. Samples were taken at 0, 2, 4, 6, 8, and 24 h. The HEp-2 cells were lysed, and intracellular bacteria were counted by plating. Inserts with infected cells grown in drug-free medium were included as controls for each time interval. A 3-log10 reduction of H. pylori was achieved in the experiments with azithromycin, and a 4-log10 reduction was achieved in the clarithromycin experiments, while no intracellular effect was seen when amoxicillin was used. The antibiotic concentrations at the sampling intervals were 12.5, 3.1, 0.8, 0.2, 0.05, and 0 mg/liter for amoxicillin; 12.5, 11.5, 10, 9, 8, and 3 mg/liter for azithromycin; and 2.4, 1.8, 1.4, 1, 0.8, and 0 mg/liter for clarithromycin. This new model for pharmacokinetic studies provides a useful tool, with applications for a broad range of microorganisms.
In this study, we evaluated a rapid whole-blood test, BM-test Helicobacter pylori, for detection of H. pylori infection in 144 and 48 patients with other gastrointestinal symptoms and with gastric cancer, respectively. The sensitivity, specificity, positive predictive value, negative predictive value, and diagnostic accuracy of the test correlated well with the standards used for the calculation, i.e., serology by enzyme-linked immunosorbent assay or culture and histology.As the market is flooded with new diagnostic kits for the rapid serological detection of Helicobacter pylori in the offices of general practitioners, there is an urgent need for prospective evaluation studies for different patient groups in various countries. In the present study, we included 192 Swedish endoscopy patients, both patients with gastric cancer (GC) and patients with other gastrointestinal (GI) symptoms. We compared results from culture, histology, and an ordinary enzyme-linked immunosorbent assay (ELISA) with those from a rapid wholeblood test, BM-test Helicobacter pylori (Boehringer Mannheim) for the detection of H. pylori. Culture, histology, and serology by ELISA were used to confirm existing H. pylori infection. The BM-test Helicobacter pylori was evaluated for both groups of patients, separately and together. We also correlated antibody titers, obtained by ELISA, to the visual scorings of the whole-blood diagnostic test. The aims of the present study were to evaluate the overall diagnostic accuracy, sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) for the rapid whole-blood BM-test Helicobacter pylori for patients with GI symptoms and to compare these to the results obtained for patients with GC.Patients. Forty-eight patients with GC and 144 patients with other GI symptoms were tested. The patients were included at endoscopy, and their ages ranged from 26 to 92 years for the GC group (mean age, 76 years) and from 34 to 90 years for the GI group (mean age, 74 years). Biopsies as well as serum and whole-blood samples were collected for analysis. Sera were not available from five patients (two GC and three GI group patients).Diagnosis of H. pylori infection. At endoscopy, biopsies for culture and histology were taken from the antrum and corpus areas of the stomach. Two biopsies from each location were homogenized together and cultured for isolation of bacteria according to standard procedures. Mucosal samples for histology were stained with hematoxylin and eosin stain, Alcian Blue-periodic acid-Schiff stain, and Giemsa stain. Immunohistochemical staining was performed with anti-H. pylori antibody (Dakopatts, Glostrup, Denmark). Sections were scored according to degree of inflammation and presence of H. pylori.
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