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Humans host complex microbial communities believed to contribute to health maintenance and, when in imbalance, to the development of diseases. Determining the microbial composition in patients and healthy controls may thus provide novel therapeutic targets. For this purpose, high-throughput, cost-effective methods for microbiota characterization are needed. We have employed 454-pyrosequencing of a hyper-variable region of the 16S rRNA gene in combination with sample-specific barcode sequences which enables parallel in-depth analysis of hundreds of samples with limited sample processing. In silico modeling demonstrated that the method correctly describes microbial communities down to phylotypes below the genus level. Here we applied the technique to analyze microbial communities in throat, stomach and fecal samples. Our results demonstrate the applicability of barcoded pyrosequencing as a high-throughput method for comparative microbial ecology.

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.

Organizations state that their responsibilities are growing and that one of the most common issues is how to integrate CSR internally with the business, where the main problem is how CSR should be evaluated. The connection between CSR integration and improvement has been unclear regarding what improvement means and how it is achieved. Interviews have been conducted with large businesses who have experience in CSR evaluation and sustainability reporting, in order to investigate; In the context of large business organizations, how can CSR be evaluated in terms of sustainability reporting, in order to understand the internal connection between integration and improvement? Research findings emphasize that CSR has transformed into the wider concept sustainability. Sustainability reporting has become institutionalized in large businesses. However, how to conduct sustainability reporting is not fully institutionalized since businesses can select what to include. Internal techniques are considered more important for the evaluation than the report itself. The analytical model is confirmed to an extent because the connection is concluded as a constant process. Particularly, because improvement is not always aimed for, since it can hinder other essential techniques. Finally, improvement is diffuse with different meanings in different industries, which should be encountered carefully.