No abstract

Worldwide, methicillin-resistant Staphylococcus aureus (MRSA) pose an increased risk for healthcare- and community-associated infections. Since the first report of MRSA in England in 1961, several distinct clones or strains have emerged. Changes within the MRSA population of whole countries, small regions or of single hospitals have been observed with some clones replacing others. In this study, the clonal replacement of MRSA isolates in a South-eastern German tertiary care hospital between 2000 and 2010 is described based on microarray analyses of 778 isolates and at least 50 MRSA per year. Within these eleven years, four common epidemic strains, CC22-MRSA-IV, CC45-MRSA-IV, CC5/ST228-MRSA-I (including a variant with a truncated SCCmec element) and CC5-MRSA-II were identified. The PVL-negative CC22-MRSA-IV strain (Barnim Epidemic Strain, UK-EMRSA-15) was detected for the first time in 2001 and its abundance increased since then to 58.6% in 2010. CC5-MRSA-II increased from 2% (2000) to about 30% (2003), and since then it fluctuates between 23 and 37% of isolates. CC5/ST228-MRSA-I decreased from about the half of tested isolates (2000) to 2.3% (2010). A similar trend was observed for CC45-MRSA-IV, which decreased drastically down to 3.4% in 2010 after reaching a maximum of 62.0% in 2002. Seventeen other PVL-negative MRSA strains were identified sporadically with no significant trend being observed. Seven PVL-positive MRSA strains were found, but they remained rare during the study period accounting together for 2.7% of isolates.

The HtrA family of serine proteases is found in most bacteria, and plays an essential role in the virulence of the gastric pathogen Helicobacter pylori. Secreted H. pylori HtrA (HtrAHp) cleaves various junctional proteins such as E-cadherin disrupting the epithelial barrier, which is crucial for bacterial transmigration across the polarized epithelium. Recent studies indicated the presence of two characteristic HtrAHp forms of 55 and 52 kDa (termed p55 and p52, respectively), in worldwide strains. In addition, p55 and p52 were produced by recombinant HtrAHp, indicating auto-cleavage. However, the cleavage sites and their functional importance are yet unclear. Here, we determined the amino-terminal ends of p55 and p52 by Edman sequencing. Two proteolytic cleavage sites were identified (H46/D47 and K50/D51). Remarkably, the cleavage site sequences are conserved in HtrAHp from worldwide isolates, but not in other Gram-negative pathogens, suggesting a highly specific assignment in H. pylori. We analyzed the role of the amino-terminal cleavage sites on activity, secretion and function of HtrAHp. Three-dimensional modeling suggested a trimeric structure and a role of amino-terminal processing in oligomerization and regulation of proteolytic activity of HtrAHp. Furthermore, point and deletion mutants of these processing sites were generated in the recently reported Campylobacter jejuni ΔhtrA/htrAHp genetic complementation system and the minimal sequence requirements for processing were determined. Polarized Caco-2 epithelial cells were infected with these strains and analyzed by immunofluorescence microscopy. The results indicated that HtrAHp processing strongly affected the ability of the protease to disrupt the E-cadherin-based cell-to-cell junctions. Casein zymography confirmed that the amino-terminal region is required for maintaining the proteolytic activity of HtrAHp. Furthermore, we demonstrated that this cleavage influences the secretion of HtrAHp in the extracellular space as an important prerequisite for its virulence activity. Taken together, our data demonstrate that amino-terminal cleavage of HtrAHp is conserved in this pathogen and affects oligomerization and thus, secretion and regulatory activities, suggesting an important role in the pathogenesis of H. pylori.

Over the last decade, the cultivation of mindfulness has become a common part of the curriculum in classrooms around the world. A recent survey indicates that nearly 50% of teachers are sharing mindfulness with children. To date, researchers have predominately used outcome-based trial designs to understand the practice's efficacy for improving wellness in children. Less research has been directed towards understanding how children perceive mindfulness experiences. This gap inspired the research question-What are students' perspectives of learning mindfulness practices at school? Thematic analysis was employed to understand and interpret 38 elementary school students' mindfulness journals. Findings suggest that mindfulness enhances student wellbeing and helps children develop a greater awareness of their body, mind and emotions. Implications of these findings are discussed. Future research is required to determine how mindfulness practices enhance and sustain student wellbeing and learning.