A whole-genome shotgun optical map of Yersinia pestis strain KIM

Yersinia pestis is the causative agent of the bubonic, septicemic, and pneumonic plagues (also known as black death) and has been responsible for recurrent devastating pandemics throughout history. To further understand this virulent bacterium and to accelerate an ongoing sequencing project, two whole-genome restriction maps (XhoI and PvuII) of Y. pestis strain KIM were constructed using shotgun optical mapping. This approach constructs ordered restriction maps from randomly sheared individual DNA molecules directly extracted from cells. The two maps served different purposes; the XhoI map facilitated sequence assembly by providing a scaffold for high-resolution alignment, while the PvuII map verified genome sequence assembly. Our results show that such maps facilitated the closure of sequence gaps and, most importantly, provided a purely independent means for sequence validation. Given the recent advancements to the optical mapping system, increased resolution and throughput are enabling such maps to guide sequence assembly at a very early stage of a microbial sequencing project.     

Short-Term Summer Inundation as a Measure to Counteract Acidification in Rich Fens

In regions with intensive agriculture, water level fluctuation in wetlands has generally become constricted within narrow limits. Water authorities are, however, considering the re-establishment of fluctuating water levels as a management tool in biodiverse, base-rich fens (‘rich fens’). This includes temporary inundation with surface water from ditches, which may play an important role in counteracting acidification in order to conserve and restore biodiversity. Inundation may result in an increased acid neutralizing capacity (ANC) for two reasons: infiltration of base-rich inundation water into peat soils, and microbial alkalinity generation under anaerobic conditions. The main objectives of this study were to test whether short-term (2 weeks) summer inundation is more effective than short-term winter inundation to restore the ANC in the upper 10 cm of non-floating peat soils, and to explain potential differences. Large-scale field experiments were conducted for five years in base-rich fens and Sphagnum-dominated poor fens. Winter inundation did not result in increased porewater ANC, because infiltration was inhibited in the waterlogged peat and evapotranspiration rates were relatively low. Also, low temperatures limit microbial alkalinity generation. In summer, however, when temperature and evapotranspiration rates are higher, inundation resulted in increased porewater Ca and HCO₃^- concentrations, but only in areas with characteristic rich fen bryophytes. This increase was not only due to stronger infiltration into the soil, but also to higher microbial alkalinity generation under anaerobic conditions. In contrast, porewater ANC did not increase in Sphagnum-plots as a result of the ability of Sphagnum spp. to acidify their environment. In both rich and poor fens, flooding-induced P-mobilization remained sufficiently low to safeguard P-limited vegetation. NO₃^- and NH₄⁺ dynamics showed no considerable changes either. In conclusion, short-term summer inundation with base-rich and nutrient-poor surface water is considered beneficial in the management of non-floating rich fens, and much more effective than winter inundation.     

Apoptosis in developing anthers and the role of ABA in this process during androgenesis in Hordeum vulgare L.

Intra-nucleosomal cleavage of DNA into fragments of about 200 bp was demonstrated to occur in developing anthers, in which microspores had developed into the mid-late to late uni-nucleate stage in situ, i.e. at the verge of mitosis. The same was observed, but to a much larger extent, if these anthers were pre- treated by a hyper-osmotic shock. Pretreatment of anthers before the actual culture of microspores was required for optimal androgenesis of microspores. The use of the TUNEL reaction, which specifically labels 3 ends of DNA breaks, after intra-nucleosomal cleavage of DNA, revealed that DNA fragmentation mainly occurred in the loculus wall cells, tapetum cells and filament cells. TUNEL staining was absent or infrequently observed in the microspores of developing anthers in situ. Electron microscopy studies showed condensed chromatin in nuclei of loculus wall cells in the developing anthers. These observations at the chromatin and DNA level are known characteristics of programmed cell death, also known as apoptosis. Features of apoptosis were infrequently found in microspores from freshly isolated mature anthers. However, most tapetum cells had disappeared in these anthers and the remaining cell structures showed loss of cellular content. The viability of microspores in pre-treated anthers was comparable to those in freshly isolated anthers and almost four times higher than in anthers from control experiments. This observation was correlated with three to four times less microspores showing TUNEL staining and a two times higher level of ABA in the anther plus medium samples than in controls. Addition of ABA to the controls enhanced the viability and lowered the occurrence of apoptosis linked characteristics in the microspores. These data suggest that pre-treatment is effective in stimulating androgenesis because it leads to an increase in ABA levels which protects microspores from dying by apoptosis.     

B-cell maturation in chimaeric mice deficient for the heat stable antigen (HSA/mouse CD24)

The murine differentiation marker heat stable antigen (HSA) is a GPI-anchored surface glycoprotein showing strong expression on immature B- and T-lymphocytes and gradually reduced expression during maturation. Although HSA has been suggested to be involved in adhesion and/or signalling, its function has not been clearly demonstrated so far. In order to elucidate the function of HSA, we analysed chimaeric mice that were generated by targeted disruption of both HSA alleles in ES cells. These mice contain normal numbers of peripheral B-cells and normal serum IgM and IgG titres of ES cell-derived allotype, demonstrating that HSA expression on B-cells is not an absolute requirement for their maturation. However, a reduction in immature B-cells in the bone marrow and an altered degree of bone marrow and blood chimaerism suggest that HSA expression influences the maturation of B-cells.