Assessment of the hypothetical protein BB0616 in the murine infection of Borrelia burgdorferi.

bb0616 of Borrelia burgdorferi, the Lyme disease pathogen, encodes a hypothetical protein of unknown function. In this study, we showed that BB0616 was not surface-exposed or associated with the membrane through localization analyses using proteinase K digestion and cell partitioning assays. The expression of bb0616 was influenced by a reduced pH but not by growth phases, elevated temperatures, or carbon sources during in vitro cultivation. A transcriptional start site for bb0616 was identified by using 5' rapid amplification of cDNA ends, which led to the identification of a functional promoter in the 5' regulatory region upstream of bb0616. By analyzing a bb0616-deficient mutant and its isogenic complemented counterparts, we found that the infectivity potential of the mutant was significantly attenuated. The inactivation of bb0616 displayed no effect on borrelial growth in the medium or resistance to oxidative stress, but the mutant was significantly more susceptible to osmotic stress. In addition, the production of global virulence regulators such as BosR and RpoS as well as virulence-associated outer surface lipoproteins OspC and DbpA was reduced in the mutant. These phenotypes were fully restored when gene mutation was complemented with a wild-type copy of bb0616. Based on these findings, we concluded that the hypothetical protein BB0616 is required for the optimal infectivity of B. burgdorferi, potentially by impacting B. burgdorferi virulence gene expression as well as survival of the spirochete under stressful conditions.

Role of the Hypothetical Protein BB0563 during Borrelia burgdorferi Infection in Animals.

The alternative sigma factor RpoS in Borrelia burgdorferi, the etiological agent of Lyme disease, has long been postulated to regulate virulence-associated genes other than ospC and dbpA. Here, we demonstrate that bb0563, a gene encoding a hypothetical protein, is regulated by RpoS and contributes to the optimal infectivity of B. burgdorferi. When B. burgdorferi was exposed to environmental stimuli, bb0563 showed similar expression patterns as rpoS, ospC, and dbpA. Expression of bb0563 was significantly downregulated when rpoS was inactivated and was restored in the complemented strain. By using rapid amplification of cDNA ends (RACE) and luciferase reporter assays, a functional promoter was identified in the regulatory region upstream of bb0563. Gene expression from this promoter was drastically decreased in the rpoS mutant. We next investigated the role of bb0563 during animal infection. By using quantitative reverse transcription-PCR (RT-PCR), we found that bb0563 was highly expressed in mouse tissues during infection. We further created a bb0563-deficient mutant in a bioluminescent B. burgdorferi strain and examined infection dynamics using in vivo imaging. Relative to the parental and complemented strains, the mutant showed a delayed infection pattern and bacterial load was reduced. Another bb0563 deletion mutant was also created in the strain 297 background, and quantitative PCR (qPCR) analysis revealed a significantly lower spirochetal burden in tissue samples collected from animals infected with the mutant. In addition, localization studies indicate that BB0563 is not exposed on the cell surface but is associated with outer membrane. Taken together, these results suggest that bb0563 is required for optimal infectivity of B. burgdorferi during experimental infection.

Trauma-Informed Pedagogy to Foster Resilience in Graduate Nursing Education.

ABSTRACT: The experiences of graduate nursing students during the COVID-19 pandemic necessitate a trauma-informed approach to education. Three hundred graduate nursing students responded to a discussion assignment in a doctoral-level health care policy course. Thematic analysis identified common themes of fear, anxiety, frustration, and exhaustion ( n = 93). Conflict and strain were identified in relation to all major roles (provider, student, and family member), ultimately creating physical and mental barriers to fulfilling each of the roles. Curricular standards must maintain rigor while incorporating flexibility into design standards to assist students when faced with trauma or crisis.

BB0761, a MepM homolog, contributes to Borrelia burgdorferi cell division and mammalian infectivity.

M23 family endopeptidases play important roles in cell division and separation in a wide variety of bacteria. Recent studies have suggested that these proteins also contribute to bacterial virulence. However, the biological function of M23 peptidases in pathogenic spirochetes remains unexplored. Here, we describe Borrelia burgdorferi, the bacterial pathogen causing Lyme disease, requires a putative M23 family homolog, BB0761, for spirochete morphology and cell division. Indeed, the inactivation of bb0761 led to an aberrant filamentous phenotype as well as the impairment of B. burgdorferi growth in vitro. These phenotypes were complemented not only with B. burgdorferi bb0761, but also with the mepM gene from E. coli. Moreover, the bb0761 mutant showed a complete loss of infectivity in a murine model of Lyme borreliosis. Resistance of the mutant to osmotic and oxidative stresses was markedly reduced. Our combined results indicate that BB0761 contributes to B. burgdorferi cell division and virulence.

Effect of intraoperative personalized goal-directed hemodynamic management on acute myocardial injury in high-risk patients having major abdominal surgery: a post-hoc secondary analysis of a randomized clinical trial.

Acute myocardial injury is common after noncardiac surgery and associated with mortality. Impaired intraoperative cardiovascular dynamics are a risk factor for acute myocardial injury. Optimizing intraoperative cardiovascular dynamics may thus reduce acute myocardial injury. We aimed to investigate the effect of intraoperative personalized goal-directed hemodynamic management on the incidence of acute myocardial injury. We hypothesized that personalized goal-directed hemodynamic management reduces the incidence of acute myocardial injury compared to routine hemodynamic management in high-risk patients having major abdominal surgery. We performed a post-hoc secondary analysis of a randomized clinical trial including 180 high-risk major abdominal surgery patients that were randomized to personalized goal-directed hemodynamic management or routine hemodynamic management. We compared the incidences of acute myocardial injury-defined according to the Fourth Universal Definition of Myocardial Infarction (2018)-between patients randomized to personalized goal-directed hemodynamic management or routine hemodynamic management by calculating the relative and absolute risk reduction together with 95% Wald confidence intervals and P values. Acute myocardial injury occurred in 4 of 90 patients (4%) in the personalized goal-directed hemodynamic management group and in 12 of 90 patients (13%) in the routine hemodynamic management group (relative risk: 0.33, 95% confidence interval: 0.11 to 0.99, P = 0.036; absolute risk reduction: - 9%, 95% confidence interval: - 17% to - 0.68%, P = 0.034). In this post-hoc secondary analysis, intraoperative personalized goal-directed hemodynamic management reduced the incidence of acute myocardial injury compared to routine hemodynamic management in high-risk patients having major abdominal surgery. This needs to be confirmed in larger prospective trials.

Rehabilitation Following Fracture in Dystrophinopathy, A Case Series.

BACKGROUND: Boys with dystrophinopathies (DMD) are at increased risk of low bone mineral density and fracture. Femoral fracture is the most common extremity fracture and is accompanied by significant risk of functional loss. Care considerations for DMD have stressed that aggressive management may be needed to maintain ambulation and that surgical fixation allows early mobilization. OBJECTIVES: Describe 5 cases of femoral fracture in ambulatory boys with DMD and the course of care undertaken to optimize function. PATIENTS: Five boys with DMD median age 15y (12-16) who were independently ambulatory. Median 10m walk speed prior to their first fracture was 8 sec (7-17.37) and 4 of 5 were less than the 9 seconds predictive of 2 year ambulation retention. Three of the cases had a single incident causing fracture; the remaining cases had 2 and 3 incidents respectively representing a total of 8 fractures 6 of which were surgically stabilized. RESULTS: Following the first fracture, all 5 subjects regained some form of ambulation. Three patients regained independent ambulation and 2 with hand held support or contact guard. Two subjects went on to have additional falls with associated fracture. No patient regained the ability to rise from the floor and only one of the 5 regained the ability to climb steps and all demonstrated a decline in walking speed. CONCLUSION: Prompt orthopedic intervention, early mobility, and intensive rehabilitation even in the end stage ambulatory patient, were factors in helping preserve function in these patients with dystrophinopathies.

DksA plays an essential role in regulating the virulence of Borrelia burgdorferi.

The RNA polymerase-binding protein DksA, together with the alarmone nucleotides (p)ppGpp, mediates the stringent response to nutrient starvation in Borrelia burgdorferi. To date, the contribution of DksA to B. burgdorferi infection remains unknown. We report here that DksA is essential for B. burgdorferi to infect a mammalian host. dksA expression was highly induced during infection. Moreover, a dksA-deficient mutant was incapable of infecting mice. The mutant displayed growth defects when cultured in vitro and resistance to osmotic pressure was markedly reduced. These phenotypes were fully restored to those of the wild type when dksA mutation was complemented. We further showed that DksA controlled the expression of virulence-associated lipoprotein OspC, likely via the central alternative sigma factor RpoS. Synthesis of RpoS was abolished in the dksA mutant, but rpoS transcription remained unaffected. Additionally, we found that the expression of clpX, clpA, clpP, and clpP2 was significantly increased in the mutant, suggesting that DksA may post-transcriptionally regulate rpoS expression via its effect on ClpXP and/or ClpAP proteases. These combined data demonstrate that DksA regulates B. burgdorferi virulence at least partially through its influence on RpoS and OspC. This study thus elucidates that, in addition to function as a stringent response regulator, DksA promotes the transcription and/or translation of genes contributing to B. burgdorferi infectivity.

The Lon-1 Protease Is Required by Borrelia burgdorferi To Infect the Mammalian Host.

Borrelia burgdorferi encodes a functional homolog of canonical Lon protease termed Lon-2. In addition, B. burgdorferi encodes a second Lon homolog called Lon-1. Recent studies suggest that Lon-1 may function differently from the prototypical Lon protease. However, the function of Lon-1 in B. burgdorferi biology remains virtually unknown. Particularly, the contribution of Lon-1 to B. burgdorferi fitness and infection remains hitherto unexplored. Herein, we show that Lon-1 plays a critical role for the infection of B. burgdorferi in a mammalian host. We found that lon-1 was highly expressed during animal infection, implying an important function of this protein in bacterial infection. We further generated a lon-1 deletion mutant and an isogenic complemented strain. Relative to that of the wild-type strain, the infectivity of the mutant was severely attenuated in a murine infection model. Our data also showed that the mutant displayed growth defects in regular BSK-II medium. Furthermore, bacterial resistance to osmotic stress was markedly reduced when lon-1 was inactivated. When exposed to tert-butyl hydroperoxide, survival of the lon-1 mutant was impaired. In addition, production of several virulence factors, such as BosR, RpoS, and OspC, was elevated in the mutant. These phenotypes were restored when the lon-1 mutation was complemented. Finally, we created a lon-1(S714A) mutant and found that this mutant failed to infect mice, suggesting that the proteolytic activity of Lon-1 is essential for bacterial infection. Taken together, these results demonstrate that Lon-1 is required by B. burgdorferi to infect animal hosts and to cope with environmental stresses.

Supramolecular design in 2D covalent organic frameworks.

2D covalent organic frameworks (COFs) are a class of porous polymers with highly crystalline structures and tunable function. The structure of a 2D-COF consists of two dimensional sheets held together through covalent bonds which are then stacked together through non-covalent forces. Since their first report, the synthesis of new COFs has relied mostly on imparting functionality to the monomer structures through covalent modification, or through the use of new thermodynamically controlled covalent bond forming methods. This tutorial review will discuss recent efforts to use supramolecular design to leverage the non-covalent forces between COF monomers and sheets to improve their properties and function. The importance of supramolecular interactions in COFs to their mechanisms of formation and overall structure will also be covered.

The Lon-2 protease of Borrelia burgdorferi is critical for infection in the mammalian host.

Borrelia burgdorferi encodes a functional homolog of canonical Lon protease termed Lon-2. To date, the contribution of Lon-2 to B. burgdorferi fitness and infection remains unexplored. Herein, we showed that expression of lon-2 was highly induced during animal infection, suggesting that Lon-2 is important for B. burgdorferi infection. We further generated a lon-2 deletion mutant. Compared with that of wild-type (WT) strain, the infectivity of the mutant was severely attenuated in a murine infection model. Although no growth defect was observed for the mutant in normal BSK-II medium, resistance of the lon-2 mutant to osmotic stress was markedly reduced. In addition, when exposed to tert-Butyl hydroperoxide, survival of the lon-2 mutant was impaired. In addition, we found that the protein levels of RpoS and RpoS-dependent OspC were decreased in the mutant. All these phenotypes were restored to WT or near-WT levels when lon-2 mutation was complemented in cis. Taken together, these results demonstrate that Lon-2 is critical for B. burgdorferi to establish infection and to cope with environmental stresses. This study provides a foundation for further uncovering the direct link between the dual roles of Lon-2 in protein quality control and bacterial pathogenesis.

Hierarchical Co-Assembly Enhanced Direct Ink Writing.

Integrating intelligent molecular systems into 3D printing materials and transforming their molecular functions to the macroscale with controlled superstructures will unleash great potential for the development of smart materials. Compared to macromolecular 3D printing materials, self-assembled small-molecule-based 3D printing materials are very rare owing to the difficulties of facilitating 3D printability as well as preserving their molecular functions macroscopically. Herein, we report a general approach for the integration of functional small molecules into 3D printing materials for direct ink writing through the introduction of a supramolecular template. A variety of inorganic and organic small-molecule-based inks were 3D-printed, and their superstructures were refined by post-printing hierarchical co-assembly. Through spatial and temporal control of individual molecular events from the nano- to the macroscale, fine-tuned macroscale features were successfully installed in the monoliths.

Computational and Experimental Studies on the Effects of Monomer Planarity on Covalent Organic Framework Formation.

We report the synthesis of one new boronate ester-based covalent organic framework (COF) and two new covalent organic polymers (COPs) made with fluoranthene-containing monomers and hexahydroxytriphenylene. The structure of the monomer heavily influences whether this material forms a highly ordered mesoporous material (COF) or an amorphous, microporous material (COP). The synthesis of the fluoranthene monomers was carried out using a divergent strategy that allows for systematic structural variation and the ability to conduct a careful structure-function study. We found that small structural variations in the monomers dramatically affected the crystallinity, surface area, pore structure, and luminescence properties of the polymers. While each of the monomers contains the same fluoranthene core, the resultant pore sizes range from microporous (10 Å) to mesoporous (37 Å), with surface areas ranging from ∼500 to 1200 m2/g. To help explain how these small structural differences can have such a large effect, we carried out a series of molecular dynamics simulations on the polymers to obtain information with atomic-scale resolution on how the monomer structure affects non-covalent COF layer stacking.

An Elastic Hydrogen-Bonded Cross-Linked Organic Framework for Effective Iodine Capture in Water.

A crystalline microporous hydrogen-bonded cross-linked organic framework has been developed through covalent photo-cross-linking of molecular monomers that are assembled in a crystalline state. The elastic framework expands its void space to adsorb iodine rapidly with a high uptake capacity in an aqueous environment as well as recovering its crystalline form after the release of iodine.

Design Principles for Covalent Organic Frameworks in Energy Storage Applications.

Covalent organic frameworks (COFs) are an exciting class of porous materials that have been explored as energy-storage materials for more than a decade. This review discusses efforts to develop these materials for applications in gas and electrical power storage. Some of the design strategies for developing the gas sorption properties of COFs and mechanistic studies on their formation are also discussed.

Gender differences in Parkinson's disease depression.

INTRODUCTION: 30-40% of patients with Parkinson's disease (PD) experience depression during their illness; identifying subtypes of depression and groups at risk remains a challenge in routine clinical care. One avenue that remains underexplored is the gender-specific profiles manifested in PD depression. We sought to explore this in a large sample of clinical PD patients. METHODS: 307 patient records at a tertiary referral centre were reviewed for clinical and demographic factors. We used recursive partitioning to determine which items on the Beck Depression Inventory (BDI) were most useful in differentiating patients who scored in the depressed range (≥14) from those who scored in the non-depressed range (≤13). We also used recursive partitioning to identify those BDI items that were most effective in differentiating depressed from non-depressed patients in both genders. RESULTS: We were able to identify a subset of items on the BDI that were most useful in partitioning depressed from non-depressed in the entire cohort. Partitioning of men and women with PD depression relied on different key BDI items, melancholy featuring prominently in women, while the more classical factors associated with depression in PD (apathy and loss of libido) featured more prominently in men. CONCLUSION: Unique factors not previously identified as core features of depression in PD were found most useful in partitioning depressed women from non-depressed women. This raises the possibility that a female-specific depressive profile has been under-appreciated in past work. Additional studies are required to discern how this may impact future research, diagnosis and treatment.

Enhanced Structural Organization in Covalent Organic Frameworks Through Fluorination.

Here, we report a structure-function study of imine covalent organic frameworks (COFs) comparing a series of novel fluorine-containing monomers to their non-fluorinated analogues. We found that the fluorine-containing monomers produced 2D-COFs with not only greatly improved surface areas (over 2000 m2 g-1 compared to 760 m2 g-1 for the non-fluorinated analogue), but also with improved crystallinity and larger, more defined pore diameters. We then studied the formation of these COFs under varying reaction times and temperatures to obtain a greater insight into their mechanism of formation.

Electrochemically active porous organic polymers based on corannulene.

For the first time, porous organic polymers (POPs) based on the smallest buckybowl, corannulene (BB-POPs) have been synthesized. Three POPs were synthesised via Sonogashira co-polymerization of 1,2,5,6-tetrabromocorannulene and alkyne linkers. BB-POP-3 exhibits the highest surface area (SABET = 560 m2 g-1) and CO2 adsorption of 11.7 wt%, while they retain the redox properties of corannulene.

Template-Directed Synthesis of Porous and Protective Core-Shell Bionanoparticles.

Metal-organic frameworks (MOFs) are promising high surface area coordination polymers with tunable pore structures and functionality; however, a lack of good size and morphological control over the as-prepared MOFs has persisted as an issue in their application. Herein, we show how a robust protein template, tobacco mosaic virus (TMV), can be used to regulate the size and shape of as-fabricated MOF materials. We were able to obtain discrete rod-shaped TMV@MOF core-shell hybrids with good uniformity, and their diameters could be tuned by adjusting the synthetic conditions, which can also significantly impact the stability of the core-shell composite. More interestingly, the virus particle underneath the MOF shell can be chemically modified using a standard bioconjugation reaction, showing mass transportation within the MOF shell.

Design Paradigm Utilizing Reversible Diels-Alder Reactions to Enhance the Mechanical Properties of 3D Printed Materials.

A design paradigm is demonstrated that enables new functional 3D printed materials made by fused filament fabrication (FFF) utilizing a thermally reversible dynamic covalent Diels-Alder reaction to dramatically improve both strength and toughness via self-healing mechanisms. To achieve this, we used as a mending agent a partially cross-linked terpolymer consisting of furan-maleimide Diels-Alder (fmDA) adducts that exhibit reversibility at temperatures typically used for FFF printing. When this mending agent is blended with commercially available polylactic acid (PLA) and printed, the resulting materials demonstrate an increase in the interfilament adhesion strength along the z-axis of up to 130%, with ultimate tensile strength increasing from 10 MPa in neat PLA to 24 MPa in fmDA-enhanced PLA. Toughness in the z-axis aligned prints increases by up to 460% from 0.05 MJ/m(3) for unmodified PLA to 0.28 MJ/m(3) for the remendable PLA. Importantly, it is demonstrated that a thermally reversible cross-linking paradigm based on the furan-maleimide Diels-Alder (fmDA) reaction can be more broadly applied to engineer property enhancements and remending abilities to a host of other 3D printable materials with superior mechanical properties.