R. Roosendaal

Long-term sequelae of Helicobacter pylori gastritis.

Chronic Helicobacter pylori gastritis has been put forward as a risk factor for development of gastric mucosal atrophy and gastric cancer. The purpose of our study was to investigate the long-term effects of H pylori gastritis on the gastric mucosa. We prospectively studied 49 subjects negative for H pylori and 58 positive subjects for a mean follow-up of 11.5 years (range 10-13 years). Serum samples were obtained at the initial and follow-up visits for determination of H pylori IgG antibodies. Gastroscopies with biopsy sampling were done in all patients at both visits. Biopsy specimens were used for assessment of H pylori infection and histology. Development of atrophic gastritis and intestinal metaplasia occurred in 2 (4%) uninfected and 16 (28%) infected subjects. Regression of atrophy was noted in 4 (7%) infected subjects. Development of atrophic gastritis and intestinal metaplasia was significantly associated with H pylori infection (p = 0.0014; odds ratio 9.0, 95% CI 1.9-41.3). The proportion of atrophic gastritis in the study population showed an annual increase of 1.15% (0.5-1.8%). We conclude that H pylori infection is a significant risk factor for development of atrophic gastritis and intestinal metaplasia. Our findings support strongly the causative role of this infection in gastric carcinogenesis.

Presence of Helicobacter pylori in the oral cavity, oesophagus, stomach and faeces of patients with gastritis

The presence ofHelicobacter pylori in the oral cavity (6 sites), oesophagus, stomach and bowel of 20 dyspeptic patients was investigated. Samples were cultured on three selective media and analyzed by 16S rDNA polymerase chain reaction (PCR) and southern hybridization.Helicobacter pylori DNA was detected by PCR from oral-cavity samples of three (20 %) and from faeces samples of only one (7 %) of the patients whose stomach biopsies were positive forHelicobacter pylori. When culture was used, the microorganisms rate of recovery from the oral cavity and faeces was 13 % and 7%, respectively. One patient had aHelicobacter pylori-like organism in samples collected from the tongue and palate. Both strains were urease, catalase and oxidase positive and grew microaerophilically but were negative on PCR analysis. This demonstrates the possibility of false identification ofHelicobacter pylori by use of routine enzyme reactions. Interestingly, specimens collected from the cheeks of three patients were positive forHelicobacter pylori by PCR analysis. This is the first instance of detection of this micro-organism in the cheek.

Comparison of different primer sets for detection of Chlamydia trachomatis by the polymerase chain reaction.

The sensitivity and specificity of the polymerase chain reaction (PCR) method was studied in vitro with HeLa cells infected with Chlamydia trachomatis serovar L2. Three different primer sets were studied; they were derived from the endogenous plasmid, the nonvariable part of the MOMP gene and the 16S ribosomal RNA (rRNA) gene. The plasmid primers were the most sensitive in the PCR method and detected at least 0.1 infectious unit of C. trachomatis in the presence of a superfluous amount of human DNA. Application of this plasmid PCR to 13 C. trachomatis culture-positive cervical smears containing < 10- > 200 inclusion-forming units showed that it was the most sensitive of the three methods and detected C. trachomatis in all samples. This correlates with the observation that the plasmid PCR method could detect C. trachomatis in cervical smears of four symptomatic patients for up to 3 weeks after the start of treatment with doxycycline. In contrast, the MOMP gene- and rRNA gene-directed PCR, as well as culture and direct immunofluorescence, gave negative results within 1 week. Therefore, we conclude that the plasmid primers are the best candidates for use in the PCR method in C. trachomatis screening programmes and clinical follow-up studies.

Detection of Chlamydia pneumoniae using a general Chlamydia polymerase chain reaction with species differentiation after hybridisation

Abstract A general polymerase chain reaction (PCR) for the detection of Chlamydia trachomatis, C. psittaci and C. pneumoniae was developed. By computer assisted sequence analysis two sets of sense primers CM1 and CM2 and one set of antisense primers CM3, each consisting of a mixture of two primers, were selected from conserved regions on the gene coding for the major outer membrane protein (MOMP-gene). Internal species specific oligoprobes were selected for the differentiation of the three species. The general PCR using primer combinations CM1/CM3 and CM2/CM3 generated a DNA fragment of approximately 360 and 320 bp respectively with 15 serovars of C. trachomatis , 6 strains of C. pneumoniae and 2 strains of C. psittaci . All amplified fragments hybridised with the general Chlamydia probe. The species specific probes detected only the Chlamydia strains of the corresponding species. Although both combinations gave rise to an identical specificity in the PCR, the CM2/CM3-PCR was 10–100 times more sensitive. The sensitivity of the CM2/CM3-PCR was one inclusion forming unit (IFU) or ten copies of isolated MOMP-DNA. Application of this general Chlamydia PCR on a throat sample weakly positive in culture for C. pneumoniae (one IFU) scored also positive after hybridisation with the general probe and the C. pneumoniae probe. The results indicate that the general Chlamydia PCR can be used for the sensitive detection of all Chlamydia species and could prove useful in the detection of Chlamydia pneumoniae in clinical specimens.

Chlamydia pneumoniae and Mycoplasma pneumoniae in Children with Acute Respiratory Infection in General Practices in The Netherlands

In this retrospective study Chlamydia pneumoniae and Mycoplasma pneumoniae infections were detected by polymerase chain reaction (PCR) in samples (n = 457) from children presenting with acute respiratory infection to general practitioners during 1992-97. Samples were collected in autumn and winter, and from 1994 onwards in spring and summer also. Overall, C. pneumoniae and M. pneumoniae were detected in throat or nasal samples by PCR in 3.1% and 2.4% of the cases, respectively. The proportion of both C. pneumoniae and M. pneumoniae infections varied between 0% and 6.9% over the years studied, whereas seasonal proportions varied from 1.8 to 9.1% and 1.2 to 4.5%, respectively. For both microorganisms the lowest proportion was detected during winter and the highest in summer. C. pneumoniae could already be detected by PCR in patients under 4 y of age, an observation not made in sero-epidemiological studies. In conclusion, both C. pneumoniae and M. pneumoniae infections play a minor role in children presenting with acute respiratory infection.In this retrospective study Chlamydia pneumoniae and Mycoplasma pneumoniae infections were detected by polymerase chain reaction (PCR) in samples (n=457) from children presenting with acute respiratory infection to general practitioners during 1992-97. Samples were collected in autumn and winter, and from 1994 onwards in spring and summer also. Overall, C. pneumoniae and M. pneumoniae were detected in throat or nasal samples by PCR in 3.1% and 2.4% of the cases, respectively. The proportion of both C. pneumoniae and M. pneumoniae infections varied between 0% and 6.9% over the years studied, whereas seasonal proportions varied from 1.8 to 9.1% and 1.2 to 4.5%, respectively. For both microorganisms the lowest proportion was detected during winter and the highest in summer. C. pneumoniae could already be detected by PCR in patients under 4 y of age, an observation not made in sero-epidemiological studies. In conclusion, both C. pneumoniae and M. pneumoniae infections play a minor role in children presenting with acute respiratory infection.