Background-The development of macrolide resistance in Helicobacter pylori is considered an essential reason for failure of antibiotic eradication therapies. The predominant mechanism of resistance to macrolides, particularly clarithromycin, is based on three defined mutations within 23S rRNA, resulting in decreased binding of the antibiotic to the bacterial ribosome. Aim-To develop an rRNA based whole cell hybridisation method to detect Helicobacter species in situ within gastric tissue, simultaneously with its clarithromycin resistance genotype. Methods-A set of fluorescent labelled oligonucleotide probes was developed, binding either to H pylori 16S rRNA or 23S rRNA sequences containing specific point mutations responsible for clarithromycin resistance. After hybridisation and stringent washing procedures, labelling of intact single bacteria was monitored by fluorescence microscopy. The new approach was compared with PCR based assays, histology, and microbiological culture. Results-In comparison with the phenotypic resistance measurement by E test, the genotypic clarithromycin resistance correlated perfectly (100%) for 35 H pylori isolates analysed. In a set of gastric biopsy specimens (27) H pylori infection was confirmed by histology (17/27) and correctly detected by whole cell hybridisation. Five clarithromycin resistant strains were identified in gastric tissue specimens directly. Furthermore, non-cultivable coccoid forms of H pylori were easily detectable by whole cell hybridisation. Conclusions-Whole cell hybridisation of rRNA holds great promise for cultivation independent, reliable, and rapid (three hours) genotypic determination of clarithromycin resistance in H pylori. Compared with PCR techniques it is independent of nucleic acid preparations, not prone to inhibition, and allows semiquantitative visualisation of the bacteria within intact tissue samples. (Gut 2000;46:608-614)

Background: Patients who have had a colorectal adenoma are likely to develop a metachronous adenoma and therefore need to be kept under surveillance. It is essential to avoid unnecessary examinations by tailoring the frequency of follow up examinations to individual risk. Methods: A total of 3134 patients undergoing endoscopic removal of colorectal adenomas were prospectively recorded on the Erlangen Registry of Colorectal Polyps between 1978 and 1996. A multivariate analysis of 1159 patients on long term follow up was performed to identify risk factors determining surveillance intervals for patients with metachronous adenomas of advanced pathologythat is, adenomas >10 mm or with high grade dysplasia or invasive carcinoma. Results: Univariate analysis revealed that sex, parental history of colorectal carcinoma, and characteristics of the initial findings-that is, size, multiplicity, and amount of villous structure-were significant predictors of metachronous adenomas of advanced pathology. On the basis of multivariate analysis, two risk groups were identified: (1) patients with no parental history of colorectal carcinoma with only small (<10 mm) tubular adenomas at the initial clearing examination have a very low risk, and we estimated that 10% will develop advanced metachronous adenomas after 10 years; (2) the high risk group contained all other patients, 10% of whom will show metachronous adenomas of advanced pathology at follow up after only three years. Conclusions: The risk of developing metachronous adenomas with advanced pathology can be stratified for various patient and adenoma characteristics. Surveillance intervals can be scheduled for low risk (10 years) and high risk (three years) patients. Risk related follow up thus helps to avoid unnecessary examinations.

The thymus is believed to play a central role in the pathogenesis of Myasthenia gravis (MG). According to a previous hypothesis, MG is initiated within the thymus by immunogenic presentation of locally produced nicotinic acetylcholine receptor (AChR) to potentially autoimmune T cells. Data of 10 consecutive MG patients demonstrate two critical features of MG thymuses that support the concept of intrathymic activation of autoreactive, AChR-specific lymphocytes. Morphologically, the thymuses showed lympho-follicular hyperplasia in nine cases and benign thymoma in one case. The paramount feature revealed by immunohistological double marker analyses was the intimate association of myoid cells (antigen producing) with interdigitating reticulum cells (potentially antigen presenting cells), both of which were surrounded by T3+ lymphocytes in thymus medulla. All 10 thymuses contained T lymphocytes reactive with AChR. This was in contrast to the peripheral immune compartment (blood) where in only 3 of 10 patients, significant T cell responses to AChR were observed. AChR-specific T cell lines could be established from 8 of 10 thymuses, all members of the helper/inducer subset as indicated by the expression of markers T3 and T4.

Thirty-nine resection specimens of gastric carcinomas have been investigated for the presence of EBV RNA sequences using a highly sensitive non-radioactive in situ hybridization technique. Transcribed EBER sequences were found in seven (18%), including four cases of undifferentiated carcinoma with prominent lymphoid infiltration and three gastric adenocarcinomas. In the positive tumours all, or nearly all, tumour nuclei were distinctly labelled. No positive signals could be detected in the non-dysplastic epithelial cells or the reactive inflammatory infiltrate. Clonality analysis using specific probes to the variable tandem repeat region of the EBV yielded single episomal bands in all four cases tested, two of which were undifferentiated carcinomas and two adenocarcinomas. By means of immunohistology, no expression of the EBV-related proteins LMP or EBNA-2 was present in tumour cells of positive cases in in situ or blotting attempts. Our results suggest that infection of gastric carcinomas by the EB virus occurs early in tumourigenesis but, in contrast to nasopharyngeal carcinomas, does not result in the expression of EBV-specific proteins.

Selective expansion of CD4+, CD45RO+ memory-type T cells could reflect an antigen-specific and/or chemokine-mediated effect in HAG. Locally produced CC chemokines such as RANTES have been detected in HAG.