Helicobacter pylori causes one of the most common, chronic bacterial infections and is a primary cause of severe gastric disorders. To unravel the bacterial factors necessary for the process of gastric colonization and pathogenesis, signature tagged mutagenesis (STM) was adapted to H. pylori. The Mongolian gerbil (Meriones unguiculatus) was used as model system to screen a set of 960 STM mutants. This resulted in 47 H. pylori genes, assigned to 9 different functional categories, representing a set of biological functions absolutely essential for gastric colonization, as verified and quantified for many mutants by competition experiments. Identification of previously known colonization factors, such as the urease and motility functions validated this method, but also novel and several hypothetical genes were found. Interestingly, a secreted collagenase, encoded by hp0169, could be identified and functionally verified as a new essential virulence factor for H. pylori stomach colonization. Furthermore, comB4, encoding a putative ATPase being part of a DNA transformation-associated type IV transport system of H. pylori was found to be absolutely essential for colonization, but natural transformation competence was apparently not the essential function. Thus, this first systematic STM application identified a set of previously unknown H. pylori colonization factors and may help to potentiate the development of novel therapies against gastric Helicobacter infections.

Summary: This study investigated the diversity of cyanobacterial mat communities of three meltwater ponds – Fresh, Orange and Salt Ponds, south of Bratina Island, McMurdo Ice Shelf, Antarctica. A combined morphological and genetic approach using clone libraries was used to investigate the influence of salinity on cyanobacterial diversity within these ecosystems without prior cultivation or isolation of cyanobacteria. We were able to identify 22 phylotypes belonging to Phormidium sp., Oscillatoria sp. and Lyngbya sp. In addition, we identified Antarctic Nostoc sp., Nodularia sp. and Anabaena sp. from the clone libraries. Fresh (17 phylotypes) and Orange (nine phylotypes) Ponds showed a similar diversity in contrast to that of the hypersaline Salt Pond (five phylotypes), where the diversity within cyanobacterial mats was reduced. Using the comparison of identified phylotypes with existing Antarctic sequence data, it was possible to gain further insight into the different levels of distribution of phylotypes identified in the investigated cyanobacterial mat communities of McMurdo Ice Shelf.

Bone is a dynamically remodeled tissue that requires gravity-mediated mechanical stimulation for maintenance of mineral content and structure. Homeostasis in bone occurs through a balance in the activities and signaling of osteoclasts, osteoblasts, and osteocytes, as well as proliferation and differentiation of their stem cell progenitors. Microgravity and unloading are known to cause osteoclast-mediated bone resorption; however, we hypothesize that osteocytic osteolysis, and cell cycle arrest during osteogenesis may also contribute to bone loss in space. To test this possibility, we exposed 16-week-old female C57BL/6J mice (n = 8) to microgravity for 15-days on the STS-131 space shuttle mission. Analysis of the pelvis by µCT shows decreases in bone volume fraction (BV/TV) of 6.29%, and bone thickness of 11.91%. TRAP-positive osteoclast-covered trabecular bone surfaces also increased in microgravity by 170% (p = 0.004), indicating osteoclastic bone degeneration. High-resolution X-ray nanoCT studies revealed signs of lacunar osteolysis, including increases in cross-sectional area (+17%, p = 0.022), perimeter (+14%, p = 0.008), and canalicular diameter (+6%, p = 0.037). Expression of matrix metalloproteinases (MMP) 1, 3, and 10 in bone, as measured by RT-qPCR, was also up-regulated in microgravity (+12.94, +2.98 and +16.85 fold respectively, p<0.01), with MMP10 localized to osteocytes, and consistent with induction of osteocytic osteolysis. Furthermore, expression of CDKN1a/p21 in bone increased 3.31 fold (p<0.01), and was localized to osteoblasts, possibly inhibiting the cell cycle during tissue regeneration as well as conferring apoptosis resistance to these cells. Finally the apoptosis inducer Trp53 was down-regulated by −1.54 fold (p<0.01), possibly associated with the quiescent survival-promoting function of CDKN1a/p21. In conclusion, our findings identify the pelvic and femoral region of the mouse skeleton as an active site of rapid bone loss in microgravity, and indicate that this loss is not limited to osteoclastic degradation. Therefore, this study offers new evidence for microgravity-induced osteocytic osteolysis, and CDKN1a/p21-mediated osteogenic cell cycle arrest.