Abstract
Because of its association with severe gastric pathologies, including gastric cancer, Helicobacter pylori has been subject of research for more than 30 years. Its capacity to adapt and survive in the human stomach can be attributed to its genetic flexibility. Its natural competence and its capacity to turn genes on and off allows H. pylori to adapt rapidly to the changing conditions of its host. Because of its genetic variability, it is difficult to establish the uniqueness of each strain obtained from a human host. The methods considered to-date to deliver the best result for differentiation of strains are Rapid Amplification of Polymorphic DNA (RAPD), Multilocus Sequence Typing (MLST) and Whole Genome Sequencing (WGS) analysis. While RAPD analysis is cost-effective, it requires a stable genome for its reliability. MLST and WGS are optimal for strain identification, however, they require analysis of data at the bioinformatics level. Using the StainFree method, which modifies tryptophan residues on proteins using 2, 2, 2, - trichloroethanol (TCE), we observed a strain specific pattern of tryptophan in 1D acrylamide gels. In order to establish the effectiveness of tryptophan fingerprinting for strain identification, we compared the graphic analysis of tryptophan-labelled bands in the gel images with MLST results. Based on this, we find that tryptophan banding patterns can be used as an alternative method for the differentiation of H. pylori strains. Furthermore, investigating the origin for these differences, we found that H. pylori strains alters the number and/or position of tryptophan present in several proteins at the genetic code level, with most exchanges taking place in membrane- and cation-binding proteins, which could be part of a novel response of H. pylori to host adaptation.
Highlights
Helicobacter pylori is a Gram (−) bacterium colonizing the human stomach
Tryptophan Fingerprinting detects variation within H. pylori strains undetected in Rapid Amplification of Polymorphic DNA (RAPD)
RAPD results for strains from patient 49 show both strains as being identical, which is confirmed with the presence of identical tryptophan pattern (Tryptophan Fingerprinting (TryF) for the same strains) (Fig. 1A)
Summary
Helicobacter pylori is a Gram (−) bacterium colonizing the human stomach. It has been estimated that around 50% of the human population are colonized by H. pylori, most of them during childhood[1,2]. One of the preferred methods for comparison of strains and their identity is the RAPD (Rapid Amplification of Polymorphic DNA) This technique uses a short DNA primer containing a sequence that is able to anneal to several regions of the DNA amplifying fragments variable in size and creating a pattern of fragments after their separation in a gel. The results are used to confirm the uniqueness of the isolates[6] This method, highly accurate, implies higher costs than the detection of DNA fragments and their patterns as in RAPD. We validated the use of tryptophan patterns from full protein lysates of H. pylori as a method for identification of strain uniqueness and investigated if different usage changes of tryptophan in H. pylori proteins by each strain are encoded in the bacterial genome and which implications this might have
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