Effective assembly of fimbriae in Escherichia coli depends on the translocation assembly module nanomachine

Outer membrane proteins are essential for Gram-negative bacteria to rapidly adapt to changes in their environment. Intricate remodelling of the outer membrane proteome is critical for bacterial pathogens to survive environmental changes, such as entry into host tissues1,​2,​3. Fimbriae (also known as pili) are appendages that extend up to 2 μm beyond the cell surface to function in adhesion for bacterial pathogens, and are critical for virulence. The best-studied examples of fimbriae are the type 1 and P fimbriae of uropathogenic Escherichia coli, the major causative agent of urinary tract infections in humans. Fimbriae share a common mode of biogenesis, orchestrated by a molecular assembly platform called ‘the usher’ located in the outer membrane. Although the mechanism of pilus biogenesis is well characterized, how the usher itself is assembled at the outer membrane is unclear. Here, we report that a rapid response in usher assembly is crucially dependent on the translocation assembly module. We assayed the assembly reaction for a range of ushers and provide mechanistic insight into the β-barrel assembly pathway that enables the rapid deployment of bacterial fimbriae

Automated external defibrillation training on the left or the right side – a randomized simulation study

Mathilde Stærk,1 Henrik Bødtker,1 Kasper G Lauridsen,1–3 Bo Løfgren,1,3,4 1Research Center for Emergency Medicine, Aarhus University Hospital, Aarhus, 2Clinical Research Unit, 3Department of Internal Medicine, Randers Regional Hospital, Randers, 4Institute of Clinical Medicine, Aarhus University, Aarhus, Denmark Background: Correct placement of the left automated external defibrillator (AED) electrode is rarely achieved. AED electrode placement is predominantly illustrated and trained with the rescuer sitting on the right side of the patient. Placement of the AED electrodes from the left side of the patient may result in a better overview of and access to the left lateral side of the thorax. This study aimed to investigate if training in automated external defibrillation on the left side compared to the right side of a manikin improves left AED electrode placement.Methods: Laypeople attending basic life support training were randomized to learn automated external defibrillation from the left or right side of a manikin. After course completion, participants used an AED and placed AED electrodes in a simulated cardiac arrest scenario.Results: In total, 40 laypersons were randomized to AED training on the left (n=19 [missing data =1], 63% female, mean age: 47.3 years) and right (n=20, 75% female, mean age: 48.7 years) sides of a manikin. There was no difference in left AED electrode placement when trained on the left or right side: the mean (SD) distances to the recommended left AED electrode position were 5.9 (2.1) cm vs 6.9 (2.2) cm (p=0.15) and to the recommended right AED electrode position were 2.6 (1.5) cm vs 1.8 (0.8) cm (p=0.06), respectively.Conclusion: Training in automated external defibrillation on the left side of a manikin does not improve left AED electrode placement compared to training on the right side. Keywords: automated external defibrillator, pads, basic life support, trainin

Differences in implementation strategies of the European Resuscitation Council Guidelines 2015 in Danish hospitals – a nationwide study

Mathilde Stærk,1–3 Kasper G Lauridsen,1–3 Troels Mygind-Klausen,1–3 Bo Løfgren2–5 1Clinical Research Unit, Randers Regional Hospital, Randers, Denmark; 2Research Center for Emergency Medicine, Aarhus University Hospital, Aarhus, Denmark; 3Department of Internal Medicine, Randers Regional Hospital, Randers, Denmark; 4Department of Clinical Medicine, Aarhus University, Aarhus, Denmark; 5Department of Cardiology, Aarhus University Hospital, Aarhus, Denmark Introduction: Guideline implementation is essential to improve survival following cardiac arrest. This study aimed to investigate awareness, expected time frame, and strategy for implementation of the European Resuscitation Council (ERC) Guidelines 2015 in Danish hospitals.Methods: All public, somatic hospitals with a cardiac arrest team in Denmark were included. A questionnaire was sent to hospital resuscitation committees one week after guideline publication. The questionnaire included questions on awareness of ERC Guidelines 2015 and time frame and strategy for implementation.Results: In total, 41 hospitals replied (response rate: 87%) between October 22 and December 22, 2015. Overall, 37% hospital resuscitation committees (n=15) were unaware of the guideline content. Most hospitals (80%, n=33) expected completion of guideline implementation within 6 months and 93% hospitals (n=38) expected the staff to act according to the ERC Guidelines 2015 within 6 months. In contrast, 78% hospitals (n=32) expected it would take between 6 months to 3 years for all staff to have completed a resuscitation course based on ERC Guidelines 2015. Overall, 29% hospitals (n=12) planned to have a strategy for implementation later than a month after guideline publication and 10% (n=4) hospitals did not plan to make a strategy.Conclusion: There are major differences in guideline implementation strategies among Danish hospitals. Many hospital resuscitation committees were unaware of guideline content. Most hospitals expected hospital staff to follow ERC Guidelines 2015 within six months after the publication even though they did not offer information or skill training to all staff members within that time frame. Keywords: resuscitation, guidelines, implementatio

A Method for Quantification of Epithelium Colonization Capacity by Pathogenic Bacteria

Most bacterial infections initiate at the mucosal epithelium lining the gastrointestinal, respiratory, and urogenital tracts. At these sites, bacterial pathogens must adhere and increase in numbers to effectively breach the outer barrier and invade the host. If the bacterium succeeds in reaching the bloodstream, effective dissemination again requires that bacteria in the blood, reestablish contact to distant endothelium sites and form secondary site foci. The infectious potential of bacteria is therefore closely linked to their ability to adhere to, colonize, and invade epithelial and endothelial surfaces. Measurement of bacterial adhesion to epithelial cells is therefore standard procedure in studies of bacterial virulence. Traditionally, such measurements have been conducted with microtiter plate cell cultures to which bacteria are added, followed by washing procedures and final quantification of retained bacteria by agar plating. This approach is fast and straightforward, but yields only a rough estimate of the adhesive properties of the bacteria upon contact, and little information on the ability of the bacterium to colonize these surfaces under relevant physiological conditions. Here, we present a method in which epithelia/endothelia are simulated by flow chamber-grown human cell layers, and infection is induced by seeding of pathogenic bacteria on these surfaces under conditions that simulate the physiological microenvironment. Quantification of bacterial adhesion and colonization of the cell layers is then performed by in situ time-lapse fluorescence microscopy and automatic detection of bacterial surface coverage. The method is demonstrated in three different infection models, simulating Staphylococcus aureus endothelial infection and Escherichia coli intestinal- and uroepithelial infection. The approach yields valuable information on the fitness of the bacterium to successfully adhere to and colonize epithelial surfaces and can be used to evaluate the influence of specific virulence genes, growth conditions, and antimicrobial treatment on this process