Abstract
AimRapid detection of H.pylori strains by PCR-Sequencing.Methods16S rDNA amplification by PCR from template genomic DNA, confirmation of amplicon size by agarose gel electrophoresis, sequencing of amplicons by automated sequencer, analysis of sequences by NCBI –BLAST software.ResultsThe PCR –Sequencing and analysis of the sequence data by BLAST resulted in detection of the strain to be of H.pylori strain#26695.ConclusionThe pathogenicity of H.pylori depends on the strain of the bacteria, PCR-Sequencing and analysis of the sequence data by BLAST can be a very quick and useful diagnostic method of the pathogen.
Highlights
There is an increasing demand worldwide for the application of intelligent, fast and inexpensive measurement systems in clinical diagnosis
We present in this study a molecular approach for H.pylori detection in which the primary identification of the causative agent was accomplished by using broad range primers to amplify part of the 16S rDNA gene followed by sequencing of the amplicon and database search
Genomic DNA of two strains of H.pylori, Strain ATCC # 700824D and 70039D were purchased from American type culture collection (ATCC), Manassas, VA, USA. 16S rDNA was amplified with the universal bacterial primers [5]; Forward: 5’ AGA GTT TGA TCC TGG CTC AG3’
Summary
There is an increasing demand worldwide for the application of intelligent, fast and inexpensive measurement systems in clinical diagnosis. In the field of Clinical Microbiology, current techniques generally require 24-48 hours to identify and characterize a pathogenic microorganism following a series of biochemical tests. New molecular biological and serological test have been introduced recently, they still have not replaced cultural methods and microscopy. Increased capital costs, need of highly skilled personnel and contamination, reduces the efficiency of these methods in the diagnosis of diseases like H.pylori infection and Tuberculosis [1,2]. The infection by H.pylori produces a superficial chronic gastritis, which can develop to Peptic Ulcer Disease, Gastric and Duodenal ulcer. In recent years it was found that H.pylori infection may play a role in the early stage of the sequence:atrophic gastritis, intestinal metaplasia, dysplasia, and gastric cancer. The relation between H.pylori infection and cancer is so close that in 1994, in the United States, the international Agency for Research in Cancer as well as the World Health Organization considered these bacteria as the most important causative agent of cancer clarifying it as a class 1 carcinogen
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