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Laboratory diagnosis of Helicobacter pylori
OBERMAJEROVÁ, Jana
Helicobacter pylori is a Gram-negative, microaerophilic bacterium requiring for its growth and development a microaerophilic environment and carbon dioxide. The most important biochemical properties include the production of urease, catalase, alkaline phosphatase, and phospholipase as well as other enzymes. Helicobacter pylori colonises the gastric mucosa in the area of the antrum and may be transferred into the body of the stomach (Corpus gastricum) in the event of massive infection. It contributes to the development of gastric mucosal inflammation, especially the antrum mucosa, which later, in the majority of cases, passes to the chronic stage. It also contributes considerably to the development of both duodenal and gastric peptic ulcers. The connection between carcinoma and gastric MALT lymphoma has been proved. Diagnosis of Helicobacter pylori infection can be made by invasive and non-invasive methods. In order to proceed to the invasive method, a biopsy sample of the gastric mucosa must be taken. This sample is later submitted for histological and microbiological examination. The microbiological methods include microscopic examination, rapid urease test and H. pylori cultivation test. In principal, non-invasive methods (breath test, stool antigen test) are used for the verification of eradical infection treatment. The topic of the thesis dealt with the comparison of individual microbiological examination methods ? microscopy, cultivation and rapid urease test which are used at the Bacteriology Institute of Hospital of České Budějovice, a.s. in order to diagnose Helicobacter pylori. Biopsy samples of the gastric mucosa sent to the Bacteriology Institute in 2011 were used as the main material for the given study. The samples were most frequently taken at the Gastroenterology Department and Children´s Department. Each biopsy sample was divided into three parts for individual methods. A microscopic preparation was elaborated from a part of the sample. The stain device - MIRASTAINER? system was used for the Gram staining. During observations in the optic microscope (immersion lens system with 1000×) fine S-shaped Gram-negative rods were visible in the event of a positive finding. Other parts of the sample were used for the urease hydrolyses test (Christensen´s) in the liquid media. The test was assessed the following day, in the case of a positive finding the media went red. The remaining part of the sample was used for the cultivation examination. The material was inoculated into the HPFA diagnostic cultivation media provided by the Dulab Company and, at the same moment, on non-selective blood agar (in order to exclude contaminating microflora). The cultivation was executed in a microaerophilic atmosphere (obtained in the LAS system) from 5 to 7 days at 37 °C. In the event of a positive cultivation finding (often represented in case of H. pylori by typical, finer, slightly grey and partially transparent colonies) the sensitivity to antibiotics was tested. In 2011, the Bacteriology Department at the Hospital of České Budějovice, a.s. examined 599 biopsy samples of the gastric mucosa. The examination proved the presence of Helicobacter pylori infection in the bodies of 161 patients. The results of the study reveal that the sensitivity of the cultivation examination is comparable to microscopy. The lowest sensitivity was proved in the case of the urease test. The infection was the most commonly diagnosed in the age group between 40 and 59 years (47.5 % of men and 52.2 % of women). The sensitivity to selected antibiotics was determined in the case of 137 phyla. 42 % of phyla were resistant to metronidazole and 18 % of phyla to azithromycin.
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