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Introduction Helicobacter pylori-infected individuals may present low-density infection, undetectable by conventional tests such as histology, rapid urease test, or urea breath test. Droplet digital polymerase chain reaction (ddPCR) is more sensitive than other polymerase chain reaction methods. We aimed to evaluate the ability of ddPCR to detect H. pylori infection in patients diagnosed as negative by conventional tests. Methods Dyspeptic patients (n = 236) were tested for H. pylori by histology, urea breath test, and rapid urease test. Patients were classified as having 3 positive (n = 25, control group), 2 positive (n = 12), one positive (n = 41), or zero positive (n = 158) diagnostic tests. DNA was extracted from gastric biopsies. Triplicate ddPCR testing for each of the 16S rDNA, ureA, and vacA(s) genes was performed using a QX200 ddPCR system (Bio-Rad). A gene was considered positive when detected by at least 2 of 3 repeated ddPCRs. H. pylori positivity was defined as having 2 or more positive genes. Results All the biopsies of the control patients were positive for all 3 16S rDNA, ureA, and vacA(s) genes. H. pylori infection was detected in 57 (36%), 22 (54%), and 9 (75%) patients with zero, 1, and 2 positive diagnostic tests, respectively. The density of infection was 5, 121, 599, and 3,133 copies of H. pylori genome equivalents for patients with zero, 1, and 2 of 3 positive test results and for the control group, respectively. Discussion ddPCR detected low-density "occult" H. pylori infection in a significant proportion (36%) of patients diagnosed as negative by conventional methods. The number of conventional positive tests was related to the density of infection.

The purpose of this study was to explore the circulating-microRNA profile fromOne hundred and twenty-three patients were enrolled and assigned to the discovery or the validation sets. In the discovery phase, circulating-microRNAs were measured by dye-based quantitative polymerase chain reaction and a selection of circulating-microRNAs was validated by probe-based quantitative polymerase chain reaction. A quality control protocol was used.One hundred and sixty-seven circulating-microRNAs were detected. Precursor lesions of gastric cancer and gastric cancer patients showed the downregulation of eight and five circulating-microRNAs, respectively. We further validated the deregulation of miR-196a-5p in precursor lesions of gastric cancer and the deregulation of miR-134-5p, miR-144-3p and miR-451a in gastric cancer. However, circulating-microRNAs exhibited moderate diagnostic performance due to the overlap of circulating-microRNA expression between non-cancer and cancer patients. miR-144-3p/miR-451a expression levels were correlated. Interestingly, these microRNAs are in 17q11.2, a site of rearrangements associated with gastric cancer.Circulating-microRNAs are deregulated in precancerous and gastric cancer patients but efforts are needed to improve their diagnostic accuracy.

33 Background: The development of gastric cancer (GC) is a stepwise progression from non-active gastritis (NAG), chronic active gastritis (CAG) and precursor lesions of GC (PLGC: atrophy, metaplasia). Early detection of GC improves survival rates. Recent evidence has proposed circulating microRNAs (c-miRNAs) as biomarkers. The aim was to explore the c-miRNA profile of a cohort of non-GC and GC patients in order to identify early biomarkers of GC. Methods: Thirty-eight patients were enrolled: ten with NAG, nine with H. pylori infected with CAG, eight with PLGC, and ten with GC. GC patients were at TNM stages I and II. As quality controls, synthetic RNA was added during the isolation, cDNA synthesis, and PCR steps. MiRNA profiling was performed using Exiqon Human Panel I. Exploratory significance was set at P