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- Dicksved, Johan, et al.
(författare)
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Molecular characterization of the stomach microbiota in patients with gastric cancer and in controls
- 2009
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Ingår i: Journal of Medical Microbiology. - : Microbiology Society. - 0022-2615 .- 1473-5644. ; 58:4, s. 509-516
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Tidskriftsartikel (refereegranskat)abstract
- Persistent infection of the gastric mucosa by Helicobacter pylori can initiate an inflammatory cascade that progresses into atrophic gastritis, a condition associated with reduced capacity for secretion of gastric acid and an increased risk of developing gastric cancer. The role of H. pylori as an initiator of inflammation is evident but the mechanism for development into gastric cancer has not yet been proven. A reduced capacity for gastric acid secretion allows survival and proliferation of other microbes that normally are killed by the acidic environment. It has been postulated that some of these species may be involved in the development of gastric cancer; however, their identities are poorly defined. In this study, the gastric microbiota from ten patients with gastric cancer was characterized and compared with that from five dyspeptic controls using the molecular profiling approach terminal restriction fragment length polymorphism (T-RFLP), in combination with 16S rRNA gene cloning and sequencing. T-RFLP analysis revealed a complex bacterial community in the cancer patients that was not significantly different from that in the controls. Sequencing of 140 clones revealed 102 phylotypes, with representatives from five bacterial phyla (Firmicutes, Bacteroidetes, Proteobacteria, Actinobacteria and Fusobacteria). The data revealed a relatively low abundance of H. pylori and showed that the gastric cancer microbiota was instead dominated by different species of the genera Streptococcus, Lactobacillus, Veillonella and Prevotella. The respective role of these species in development of gastric cancer remains to be determined.
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| 2. |
- Hugerth, Luisa W., et al.
(författare)
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DegePrime, a Program for Degenerate Primer Design for Broad-Taxonomic-Range PCR in Microbial Ecology Studies
- 2014
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Ingår i: Applied and Environmental Microbiology. - 0099-2240 .- 1098-5336. ; 80:16, s. 5116-5123
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Tidskriftsartikel (refereegranskat)abstract
- The taxonomic composition of a microbial community can be deduced by analyzing its rRNA gene content by, e. g., high-throughput DNA sequencing or DNA chips. Such methods typically are based on PCR amplification of rRNA gene sequences using broad-taxonomic-range PCR primers. In these analyses, the use of optimal primers is crucial for achieving an unbiased representation of community composition. Here, we present the computer program DegePrime that, for each position of a multiple sequence alignment, finds a degenerate oligomer of as high coverage as possible and outputs its coverage among taxonomic divisions. We show that our novel heuristic, which we call weighted randomized combination, performs better than previously described algorithms for solving the maximum coverage degenerate primer design problem. We previously used DegePrime to design a broad-taxonomic-range primer pair that targets the bacterial V3-V4 region (341F-805R) (D. P. Herlemann, M. Labrenz, K. Jurgens, S. Bertilsson, J. J. Waniek, and A. F. Andersson, ISME J. 5:1571-1579, 2011, http://dx.doi.org/10.1038/ismej.2011.41), and here we use the program to significantly increase the coverage of a primer pair (515F-806R) widely used for Illumina-based surveys of bacterial and archaeal diversity. By comparison with shotgun metagenomics, we show that the primers give an accurate representation of microbial diversity in natural samples.
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| 3. |
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| 4. |
- Lindberg, Mathilda
(författare)
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The human gastric microbiota in health and disease
- 2010
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The human body comprises of complex microbial ecosystems that have co-evolved with its host and play important roles in the maintenance of health and in the etiology and outcome of various disease states. The microbiota that resides in the stomach has not been completely identified, and focus has previously been mainly on the gastric pathogen Helicobacter pylori. This bacterium colonizes a substantial part of the human population, almost 40% of the population in many western countries, often in a life-long persistence. H. pylori infection as a cause of gastric disease has been extensively studied and some, but not all, risk factors have been identified. This gives rise to the question of involvement of other factors such as presence of other bacteria and their possible protecting or aggravating effects. In the normal acidic stomach a sparse cultivable microbiota has been found, but in studies using the 16S rRNA gene to determine the bacterial microbiota an increased diversity has been seen. To what extent this represents resident or transient populations of ingested microbes is not known. In this thesis both molecular and cultivation based approaches were used to study the bacterial composition in the human gut. One specific aim was to further develop the 454 pyrosequencing as a tool for assessing changes in the human microbiota. Using the 454 pyrosequencing approach PCR-products can be sequenced without prior cloning and sample-specific sequence tags enable sequencing of hundreds of samples in one single run. In order to function well in samples with both high and low bacterial/host cell ratios, primers were selected not only to maximize the number of hits to bacterial 16S rRNA genes but also minimize matches to sequences in the human genome. Using this sequence tag approach the gastric microbiota in healthy individuals compared to individuals with corpus predominant atrophic gastritis and individuals treated with proton pump inhibitors was analyzed and a shift in the microbiota between stomachs with atrophy compared to controls. A second aim was to determine changes in the gastric microbiota in gastric cancer patients compared to healthy controls using a different molecular approach, terminal-restriction fragment length polymorphism (T-RFLP) in combination with cloning and sequencing. The composition between the dyspeptic controls and stomachs with cancer was similar with a dominance of streptococci and Firmicutes. There are bacterial groups that are known to be able to survive in an acidic environment and one of these groups is the Lactobacillus sp. group. Some of these lactobacilli may be able to colonize the human stomach and also co-exist with H. pylori. A cultivation based approach was chosen since the aim was to investigate the possible colonization of this specific group in the human stomach. In molecular based approaches, when using extracted DNA in the analyses, there can be no distinction between DNA coming from dead or alive bacterial cells. Our results show that Lactobacillus spp. are as common in the human stomach as H. pylori. We also found Lactobacillus strains that were present in the same stomach when sampling four years apart. This indicates that Lactobacillus spp. are constantly present in the stomach and could even be long-term colonizers. In conclusion, the gastric microbiota is highly diverse but still very similar between individuals. However, shift in the gastric microbiota was found in individuals with corpus predominant atrophic gastritis.
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