Steroid Drugs Inhibit Bacterial Respiratory Oxidases and Are Lethal Toward Methicillin-Resistant Staphylococcus aureus.

BACKGROUND: Cytochrome bd complexes are respiratory oxidases found exclusively in prokaryotes that are important during infection for numerous bacterial pathogens. METHODS: In silico docking was employed to screen approved drugs for their ability to bind to the quinol site of Escherichia coli cytochrome bd-I. Respiratory inhibition was assessed with oxygen electrodes using membranes isolated from E. coli and methicillin-resistant Staphylococcus aureus strains expressing single respiratory oxidases (ie, cytochromes bd, bo', or aa3). Growth/viability assays were used to measure bacteriostatic and bactericidal effects. RESULTS: The steroid drugs ethinylestradiol and quinestrol inhibited E. coli bd-I activity with median inhibitory concentration (IC50) values of 47 ± 28.9 µg/mL (158 ± 97.2 µM) and 0.2 ± 0.04 µg/mL (0.5 ± 0.1 µM), respectively. Quinestrol inhibited growth of an E. coli "bd-I only" strain with an IC50 of 0.06 ± 0.02 µg/mL (0.2 ± 0.07 µM). Growth of an S. aureus "bd only" strain was inhibited by quinestrol with an IC50 of 2.2 ± 0.43 µg/mL (6.0 ± 1.2 µM). Quinestrol exhibited potent bactericidal effects against S. aureus but not E. coli. CONCLUSIONS: Quinestrol inhibits cytochrome bd in E. coli and S. aureus membranes and inhibits the growth of both species, yet is only bactericidal toward S. aureus.

The nitric oxide paradox: antimicrobial and inhibitor of antibiotic efficacy.

It is well-known that antibiotics target energy-consuming processes and a significant body of research now supports the conclusion that the metabolic state of bacteria can have a profound impact upon the efficacy of antibiotics. Several articles implicate bacterial energetics and the respiratory inhibitor nitric oxide (NO) in this process, although pinpointing the precise mechanism for how NO can diminish the potency of a range of antibiotics through modulating bacterial energy metabolism has proved challenging. Herein, we introduce the role of NO during infection, consider known links between NO and antibiotic efficacy, and discuss potential mechanisms via which NO present at the site of infection could mediate these effects through controlling bacterial energetics. This perspective article highlights an important relationship between NO and antibiotic action that has largely been overlooked and outlines future considerations for the development of new drugs and therapies that target bacterial energy metabolism.

Patient-reported outcome measures in physical therapy practice for neck pain: an overview of reviews.

BACKGROUND: Understanding which patient-reported outcome measures are being collected and utilized in clinical practice and research for patients with neck pain will help to inform recommendations for a core set of measures that provide value to patients and clinicians during diagnosis, clinical decision-making, goal setting and evaluation of responsiveness to treatment. Therefore, the aim of this study was to conduct a review of systematic reviews using a qualitative synthesis on the use of patient-reported outcome measures (PROMs) for patients presenting with neck pain to physical therapy. METHODS: An electronic search of systematic reviews and guideline publications was performed using MEDLINE (OVID), Embase (Elsevier), CINAHL Complete (EBSCOhost), and Web of Science (Clarivate) databases to identify reviews that evaluated physical therapy interventions or interventions commonly performed by a physical therapist for individuals with neck pain and included at least one patient-reported outcome measure. The frequency and variability in which the outcome measures were reported among the studies in the review and the constructs for which they measured were evaluated. The evaluation of a core set of outcome measures was assessed. Risk of bias and quality assessment was performed using A Measurement Tool to Assess systematic Reviews 2. RESULTS: Of the initial 7,003 articles, a total of 37 studies were included in the final review. Thirty-one PROMs were represented within the 37 reviews with eleven patient-reported outcome measures in three or more reviews. The eleven PROMs assessed the constructs of disability, pain intensity, psychosocial factors and quality of life. The greatest variability was found amongst individual measures assessing psychosocial factors. Assessment of psychosocial factors was the least represented construct in the included studies. Overall, the most frequently utilized patient reported outcome measures were the Neck Disability Index, Visual Analog Scale, and Numeric Pain Rating Scale. The most frequently used measures evaluating the constructs of disability, pain intensity, quality of life and psychosocial functioning included the Neck Disability Index, Visual Analog Scale, Short-Form-36 health survey and Fear Avoidance Belief Questionnaire respectively. Overall risk of bias and quality assessment confidence levels ranged from critically low (2 studies), low (12 studies), moderate (8 studies), and high (15 studies). CONCLUSION: This study identified a core set of patient-reported outcome measures that represented the constructs of disability, pain intensity and quality of life. This review recommends the collection and use of the Neck Disability Index and the Numeric Pain Rating Scale or Visual Analog Scale. Recommendation for a QoL measure needs to be considered in the context of available resources and administrative burden. Further research is needed to confidently recommend a QoL and psychosocial measure for patients presenting with neck pain. Other measures that were not included in this review but should be further evaluated for patients with neck pain are the Patient Reported Outcomes Measurement Information System (PROMIS) Physical function, PROMIS Pain Interference and the Optimal Screening for Prediction of Referral and Outcome Yellow Flag (OSPRO-YF) tool.

Measures of sleep disturbance are not routinely captured in trials for chronic low back pain: a systematic scoping review of 282 trials.

STUDY OBJECTIVES: To investigate the extent to which sleep measures are reported in intervention trials for chronic low back pain. METHODS: A systematic scoping review was conducted. Ovid MEDLINE, Cochrane CENTRAL, and CINAHL were queried for trials published between January 2010 and December 2022 using keywords related to chronic low back pain. Two reviewers screened and reviewed abstracts and full texts for eligibility criteria and extracted data. Randomized intervention trials with the aim to treat pain or disability related to chronic low back pain in adults were included. Data were pooled and synthesized from trials that included a measure of sleep. RESULTS: Two hundred eighty-two trials conducted in 40 different countries were included in the final review. Twenty-six trials (9.2%) assessed any sleep measure, and 13 (4.6%) collected a formal sleep disturbance measure at multiple time points. Three trials analyzed the mediating effects of sleep disturbance on pain. Reporting of sleep measures was no better in more recently published trials; trials published in 2010 (22%; n = 2/9) and 2022 (23%; n = 3/13) had the highest reporting rates. CONCLUSIONS: The poor adherence to guideline recommendations for capturing measures of sleep quality or disturbance limits clinicians' and researchers' understanding of how sleep may influence treatment effects for chronic low back pain. There is an opportunity to improve the understanding of the relationship between sleep and pain with improved collection and reporting of sleep disturbance measures. CITATION: Neilson BD, Dickerson C, Young JL, Shepherd MH, Rhon DI. Measures of sleep disturbance are not routinely captured in trials for chronic low back pain: a systematic scoping review of 282 trials. J Clin Sleep Med. 2023;19(11):1961-1970.

Controlling the structure of supramolecular fibre formation for benzothiazole based hydrogels with antimicrobial activity against methicillin resistant Staphylococcus aureus.

Antimicrobial resistance is one of the greatest threats to human health. Gram-positive methicillin resistant Staphylococcus aureus (MRSA), in both its planktonic and biofilm form, is of particular concern. Herein we identify the hydrogelation properties for a series of intrinsically fluorescent, structurally related supramolecular self-associating amphiphiles and determine their efficacy against both planktonic and biofilm forms of MRSA. To further explore the potential translation of this hydrogel technology for real-world applications, the toxicity of the amphiphiles was determined against the eukaryotic multicellular model organism, Caenorhabditis elegans. Due to the intrinsic fluorescent nature of these supramolecular amphiphiles, material characterisation of their molecular self-associating properties included; comparative optical density plate reader assays, rheometry and widefield fluorescence microscopy. This enabled determination of amphiphile structure and hydrogel sol dependence on resultant fibre formation.

The novel two-component system AmsSR governs alternative metabolic pathway usage in Acinetobacter baumannii.

In this study, we identify a novel two-component system in Acinetobacter baumannii (herein named AmsSR for regulator of alternative metabolic systems) only present in select gammaproteobacterial and betaproteobacterial species. Bioinformatic analysis revealed that the histidine kinase, AmsS, contains 14 predicted N-terminal transmembrane domains and harbors a hybrid histidine kinase arrangement in its C-terminus. Transcriptional analysis revealed the proton ionophore CCCP selectively induces P amsSR expression. Disruption of amsSR resulted in decreased intracellular pH and increased depolarization of cytoplasmic membranes. Transcriptome profiling revealed a major reordering of metabolic circuits upon amsR disruption, with energy generation pathways typically used by bacteria growing in limited oxygen being favored. Interestingly, we observed enhanced growth rates for mutant strains in the presence of glucose, which led to overproduction of pyruvate. To mitigate the toxic effects of carbon overflow, we noted acetate overproduction in amsSR-null strains, resulting from a hyperactive Pta-AckA pathway. Additionally, due to altered expression of key metabolic genes, amsSR mutants favor an incomplete TCA cycle, relying heavily on an overactive glyoxylate shunt. This metabolic reordering overproduces NADH, which is not oxidized by the ETC; components of which were significantly downregulated upon amsSR disruption. As a result, the mutants almost exclusively rely on substrate phosphorylation for ATP production, and consequently display reduced oxygen consumption in the presence of glucose. Collectively, our data suggests that disruption of amsSR affects the function of the aerobic respiratory chain, impacting the energy status of the cell, which in turn upregulates alternative metabolic and energy generation pathways.

Dissecting the conformational complexity and mechanism of a bacterial heme transporter.

Iron-bound cyclic tetrapyrroles (hemes) are redox-active cofactors in bioenergetic enzymes. However, the mechanisms of heme transport and insertion into respiratory chain complexes remain unclear. Here, we used cellular, biochemical, structural and computational methods to characterize the structure and function of the heterodimeric bacterial ABC transporter CydDC. We provide multi-level evidence that CydDC is a heme transporter required for functional maturation of cytochrome bd, a pharmaceutically relevant drug target. Our systematic single-particle cryogenic-electron microscopy approach combined with atomistic molecular dynamics simulations provides detailed insight into the conformational landscape of CydDC during substrate binding and occlusion. Our simulations reveal that heme binds laterally from the membrane space to the transmembrane region of CydDC, enabled by a highly asymmetrical inward-facing CydDC conformation. During the binding process, heme propionates interact with positively charged residues on the surface and later in the substrate-binding pocket of the transporter, causing the heme orientation to rotate 180°.

Leveraging the Short-Term Benefits of Manual Therapy which Includes Exercise Over Exercise Therapy Alone Appears Justified for Knee Osteoarthritis.

Letter to the Editor-in-Chief in response to JOSPT article "The Benefits of Adding Manual Therapy to Exercise Therapy for Improving Pain and Function in Patients with Knee or Hip Osteoarthritis: A Systematic Review with Meta-analysis" by Runge et al. J Orthop Sports Phys Ther 2023;53(1):49-50. doi:10.2519/jospt.2023.0201.

A mini-review: environmental and metabolic factors affecting aminoglycoside efficacy.

Following the discovery of streptomycin from Streptomyces griseus in the 1940s by Selman Waksman and colleagues, aminoglycosides were first used to treat tuberculosis and then numerous derivatives have since been used to combat a wide variety of bacterial infections. These bactericidal antibiotics were used as first-line treatments for several decades but were largely replaced by ß-lactams and fluoroquinolones in the 1980s, although widespread emergence of antibiotic-resistance has led to renewed interest in aminoglycosides. The primary site of action for aminoglycosides is the 30 S ribosomal subunit where they disrupt protein translation, which contributes to widespread cellular damage through a number of secondary effects including rapid uptake of aminoglycosides via elevated proton-motive force (PMF), membrane damage and breakdown, oxidative stress, and hyperpolarisation of the membrane. Several factors associated with aminoglycoside entry have been shown to impact upon bacterial killing, and more recent work has revealed a complex relationship between metabolic states and the efficacy of different aminoglycosides. Hence, it is imperative to consider the environmental conditions and bacterial physiology and how this can impact upon aminoglycoside entry and potency. This mini-review seeks to discuss recent advances in this area and how this might affect the future use of aminoglycosides.

The Evolution of Nitric Oxide Function: From Reactivity in the Prebiotic Earth to Examples of Biological Roles and Therapeutic Applications.

Nitric oxide was once considered to be of marginal interest to the biological sciences and medicine; however, there is now wide recognition, but not yet a comprehensive understanding, of its functions and effects. NO is a reactive, toxic free radical with numerous biological targets, especially metal ions. However, NO and its reaction products also play key roles as reductant and oxidant in biological redox processes, in signal transduction, immunity and infection, as well as other roles. Consequently, it can be sensed, metabolized and modified in biological systems. Here, we present a brief overview of the chemistry and biology of NO-in particular, its origins in geological time and in contemporary biology, its toxic consequences and its critical biological functions. Given that NO, with its intrinsic reactivity, appeared in the early Earth's atmosphere before the evolution of complex lifeforms, we speculate that the potential for toxicity preceded biological function. To examine this hypothesis, we consider the nature of non-biological and biological targets of NO, the evolution of biological mechanisms for NO detoxification, and how living organisms generate this multifunctional gas.

The Blind Men, the Elephant, and the Continuing Education Course: Why Higher Standards Are Needed in Physical Therapist Professional Development.

SYNOPSIS: Evidence-based practice and implementing clinical practice guidelines are familiar themes for musculoskeletal rehabilitation clinicians. Yet, many clinicians continue to eschew recommended treatments. One explanation could be that physical therapists largely rely on continuing education courses-not research reports or standardized postprofessional education-to learn new treatments and update their practice patterns. However, continuing education courses in physical therapy have a much less rigorous review process, and interventions taught in these courses often conflict with high-quality evidence. The lack of rigor in continuing education may contribute to unwarranted variability in practice, which is a major threat to physical therapy. The current continuing competence paradigm in the United States, of which continuing education is a part, needs an overhaul to ensure clinicians learn current best evidence. Now is the time for change in professional development. We offer 3 suggestions to improve the current system of continuing competence in physical therapy. J Orthop Sports Phys Ther 2022;52(10):642-646. Epub: 27 July 2022. doi:10.2519/jospt.2022.11377.

Relationship Between Attitudes and Beliefs About Sleep, Sleep Disturbance, and Pain Interference in Patients With Spinal Pain.

OBJECTIVES: Sleep impairments are a strong predictor of pain, making sleep a potential interest when treating patients with spine pain. Typical beliefs about the importance of sleep in patients seeking care for spinal pain are unknown. The purpose of this study was to describe the beliefs and attitudes about sleep in patients seeking care for spinal pain and to examine the relationships between dysfunctional beliefs and attitudes about sleep (DBAS), disordered sleep, and pain interference. MATERIALS AND METHODS: This cross-sectional study included patients presenting to physical therapy with spine pain. Participants completed questionnaires including demographics, medical history, pain interference (pain, enjoyment, and general activity), DBAS-16, and sleep-related impairment (Patient-Reported Outcome Measurement Information System). Correlations were calculated between DBAS-16 scores and measures of sleep quality/quantity, and a generalized linear model was used to investigate the predictive ability of DBAS-16 scores on pain interference. RESULTS: The mean DBAS-16 score was 4.22 (SD=2.03), with 52.5% of participants having DBAS. There was a strong relationship between DBAS-16 and Patient-Reported Outcome Measurement Information System ( rs =0.7; P <0.001). For every point higher score on the DBAS-16, pain interference scores increased by approximately half a point (B=0.46; 95% CI 0.33, 0.59, 1.80; P <0.001). DISCUSSION: These results highlight a strong relationship between beliefs and attitudes about sleep and measures of sleep quality/quantity and a linear association with pain interference scores. These findings provide a rationale for targeting beliefs and attitudes about sleep when managing pain-related symptoms in patients seeking care for spine pain.

The polyene antifungal candicidin is selectively packaged into membrane vesicles in Streptomyces S4.

In recent years, much attention has been focused on the biogenesis, engineering and utilisation of outer membrane vesicles (OMVs) in Gram-negative bacteria in a range of environments and niches. While the precise mechanism of biogenesis is unknown, it is focused on the modification of the Gram-negative cell wall to facilitate blebbing at sites of weakness in and around the characteristically thin peptidoglycan layer within the periplasm. Here, we investigate the biogenesis of membrane vesicles (MVs) in the Gram-positive organism Streptomyces albus S4 (Seipke et al. J Bacteriol 193:4270-4271, 2011 and Fazal et al. Antonie Van Leeuwenhoek 113:511-520, 2020). The S. albus S4 strain is an antifungal (candicidin and antimycin) producing organism that was isolated from attine ants (Barke et al. BMC Biol 8:109, 2010). The biogenesis and characterisation of S. albus S4 MVs is demonstrated using the wild-type (WT) and mutant strains ΔantC (no antimycin production) ΔfscC (no candicidin production) and ΔantC ΔfscC (produces neither antimycin nor candicidin). Here, we have shown that the S. albus S4 strain produces MVs and that these are comprised of both specific protein profiles and secondary metabolites, with a clear demonstration of the ability to selectively package one antifungal (candicidin) but not the other (antimycin).

The influence of manual therapy dosing on outcomes in patients with hip osteoarthritis: a systematic review.

OBJECTIVE: To 1) Determine if specific dosing parameters of manual therapy are related to improved pain, disability, and quality of life outcomes in patients with hip osteoarthritis and 2) to provide recommendations for optimal manual therapy dosing based on our findings. DESIGN: A systematic review of randomized controlled trials from the PubMed, CINAHL, and OVID databases that used manual therapy interventions to treat hip osteoarthritis was performed. Three reviewers assessed the risk of bias for included studies and extracted relevant outcome data based on predetermined criteria. Baseline and follow-up means and standard deviations for outcome measures were used to calculate effect sizes for within and between-group differences. RESULTS: Ten studies were included in the final analyses totaling 768 participants, and half were graded as high risk of bias. Trends emerged: 1) large effect sizes were seen using long-axis distraction, mobilization and thrust manipulation, 2) mobilization with movement showed large effects for pain and range of motion, and (3) small effects were associated with graded mobilization. Durations of 10 to 30 minutes per session, and frequency 2-3 times per week for 2-6 weeks were the most common dosing parameters. CONCLUSIONS: There were varied effect sizes associated with pain, function, and quality of life for both thrust and non-thrust mobilizations, and mobilization with movement into hip flexion and internal rotation. Due to the heterogeneity of MT dosage, it is difficult to recommend a specific manual therapy dosage for those with hip osteoarthritis.

The effectiveness of post-professional physical therapist training in the treatment of chronic low back pain using a propensity score approach with machine learning.

RATIONALE: Low back pain (LBP) is a leading cause of disability in the United States creating substantial hardships through negative social, financial, and health effects. Chronic low back pain (CLBP) accounted for above half of patients treated in physical therapy (PT) clinics for LBP. However, research shows small benefit from PT in CLBP treatment. Preliminary evidence suggests clinician-level training variables may affect outcomes, but requires further investigation to determine whether patients with CLBP benefit from treatment by providers with post-professional training. This study examined the relationship between clinician training levels and patient-reported outcomes in CLBP treatment. METHODS: Physical therapies were surveyed using a large patient outcome assessment system to determine and categorise them by level of post-professional education. To account for the possibility that clinicians with higher levels of training are referred more-complex patients, a machine learning approach was used to identify predictive variables for clinician group, then to construct propensity scores to account for differences between groups. Differences in functional status score change among pooled data were analysed using linear models adjusted for propensity scores. RESULTS: There were no clinically meaningful differences in patient outcomes when comparing clinician post-professional training level. The propensity score method proved to be a valuable way to account for differences at baseline between groups. CONCLUSION: Post-professional training does not appear to contribute to improved patient outcomes in the treatment of CLBP. This study demonstrates that propensity score analysis can be used to ensure that differences observed are true and not due to differences at baseline.

Proton motive force underpins respiration-mediated potentiation of aminoglycoside lethality in pathogenic Escherichia coli.

It is well known that loss of aerobic respiration in Gram-negative bacteria can diminish the efficacy of a variety of bactericidal antibiotics, which has lead to subsequent demonstrations that the formation of reactive oxygen species (ROS) and the proton motive force (PMF) can both play a role in antibiotic toxicity. The susceptibility of Gram-negative bacteria to aminoglycoside antibiotics, particularly gentamicin, has previously been linked to both the production of ROS and the rate of antibiotic uptake that is mediated by the PMF, although the relative contributions of ROS and PMF to aminoglycoside toxicity has remained poorly understood. Herein, gentamicin was shown to elicit a very modest increase in ROS levels in an aerobically grown Escherichia coli clinical isolate. The well-characterised uncoupler 2,4-dinitrophenol (DNP) was used to disrupt the PMF, which resulted in a significant decrease in gentamicin lethality towards E. coli. DNP did not significantly alter respiratory oxygen consumption, supporting the hypothesis that this uncoupler does not increase ROS production via elevated respiratory oxidase activity. These observations support the hypothesis that maintenance of PMF rather than induction of ROS production underpins the mechanism for how the respiratory chain potentiates the toxicity of aminoglycosides. This was further supported by the demonstration that the uncoupler DNP elicits a dramatic decrease in gentamicin lethality under anaerobic conditions. Together, these data strongly suggest that maintenance of the PMF is the dominant mechanism for the respiratory chain in potentiating the toxic effects of aminoglycosides.

Deletion of glyceraldehyde-3-phosphate dehydrogenase (gapN) in Clostridium saccharoperbutylacetonicum N1-4(HMT) using CLEAVE™ increases the ATP pool and accelerates solvent production.

The development and advent of mutagenesis tools for solventogenic clostridial species in recent years has allowed for the increased refinement of industrially relevant strains. In this study we have utilised CLEAVE™, a CRISPR/Cas genome editing system developed by Green Biologics Ltd., to engineer a strain of Clostridium saccharoperbutylacetonicum N1-4(HMT) with potentially useful solvents titres and energy metabolism. As one of two enzymes responsible for the conversion of glyceraldehyde-3-phosphate (GAP) to 3-phosphoglyceric acid in glycolysis, it was hypothesised that deletion of gapN would increase ATP and NADH production that could in turn improve solvent production. Herein, whole genome sequencing has been used to evaluate CLEAVE™ and the successful knockout of gapN, demonstrating a clean knockout with no other detectable variations from the wild type sequence. Elevated solvent levels were detected during the first 24 h of batch fermentation, indicating an earlier shift to solventogenesis. A 2.4-fold increase in ATP concentration was observed, and quantitation of NAD(P)H derivatives revealed a more reducing cytoplasm for the gapN strain. These findings expand our understanding of clostridium carbon metabolism and report a new approach to optimising biofuel production.

Parallel bioreactor system for accessible and reproducible anaerobic culture.

When working with anaerobic bacteria it is important to have the capability to perform parallel bioreactor growth experiments that are both controllable and reproducible, although capital and consumables costs for commercially available systems are often prohibitively high. Hence, a three-vessel parallel bioreactor system was designed and constructed that has the capabilities for batch and fed batch processes and can also be set up for continuous culture at a fraction of the cost of commercial systems. This system carries over many of the same functionalities of those systems with a higher price point of entry, including in-line monitoring of temperature, pH, and redox poise. To validate the performance of this system Clostridium saccharoperbutylacetonicum was grown under conditions that promote ABE fermentation, an established industrial process used to produce the solvents acetone, butanol and ethanol. Measurements of cell density, pH, and redox poise all confirmed reproducible culture conditions for these parallel vessels, and solvent quantitation via GCMS verified consistent metabolic activities for the separate cultures. In future, this system will be of interest to researchers that require high performance parallel fermentation platforms but where commercial systems are not accessible.