Antibiotic tolerance among clinical isolates: mechanisms, detection, prevalence, and significance.

SUMMARYAntibiotic treatment failures in the absence of resistance are not uncommon. Recently, attention has grown around the phenomenon of antibiotic tolerance, an underappreciated contributor to recalcitrant infections first detected in the 1970s. Tolerance describes the ability of a bacterial population to survive transient exposure to an otherwise lethal concentration of antibiotic without exhibiting resistance. With advances in genomics, we are gaining a better understanding of the molecular mechanisms behind tolerance, and several studies have sought to examine the clinical prevalence of tolerance. Attempts have also been made to assess the clinical significance of tolerance through in vivo infection models and prospective/retrospective clinical studies. Here, we review the data available on the molecular mechanisms, detection, prevalence, and clinical significance of genotypic tolerance that span ~50 years. We discuss the need for standardized methodology and interpretation criteria for tolerance detection and the impact that methodological inconsistencies have on our ability to accurately assess the scale of the problem. In terms of the clinical significance of tolerance, studies suggest that tolerance contributes to worse outcomes for patients (e.g., higher mortality, prolonged hospitalization), but historical data from animal models are varied. Furthermore, we lack the necessary information to effectively treat tolerant infections. Overall, while the tolerance field is gaining much-needed traction, the underlying clinical significance of tolerance that underpins all tolerance research is still far from clear and requires attention.

Perforated duodenal diverticulum treated conservatively.

Perforation is the rarest complication of duodenal diverticulum but also one of its most serious complications. Mortality rate was reported to up to 30%. Clinical diagnosis is usually vague and non-specific. High clinical suspicion is important as rapid deterioration is likely. Consensus regarding management is lacking and currently guided by a small series of case reviews. Surgical treatment is historically the standard therapeutic option. However, more current literature suggests improvement in patients with non-surgical management. In this paper, we discuss a case of a patient in her early 60s who had a perforated duodenal diverticulum that was treated conservatively. It highlights the importance of a good history and a CT scan to help with diagnosis. Close clinical observation is essential to detect disease progression. A step-up approach to clinical deterioration with either percutaneous drainage or surgical management should be considered if the patient does not respond to conservative treatment.

Investigation of the hospital pharmacy profession in Europe.

OBJECTIVE: From 1995, the European Association of Hospital Pharmacists (EAHP) has regularly investigated the progress of the hospital pharmacy profession in Europe, and identified key barriers and drivers of this. The most recent 'Investigation of the Hospital Pharmacy Profession in Europe' was conducted from November 2022 to March 2023. METHODS: The online questionnaire was sent to all hospital pharmacies in EAHP member countries. The investigation was drafted using the same questions as the 2015 baseline survey. Where possible and relevant, responses were compared with the data from previous surveys that monitored the implementation of the EAHP statements. Keele University, Centre for Medicines Optimisation, School of Pharmacy and Bioengineering, UK analysed the data. RESULTS: The overall number of responses was 653, with a better response rate of 19% compared with 14% in 2018 statements survey. The findings indicated that participating hospital pharmacies have similar characteristics to previous surveys. Section 1 (Introductory statements and governance), section 2 (Selection, procurement and distribution), section 3 (Production and compounding), section 5 (Patient safety and quality assurance) questions were generally answered positively, with results ranging from 52% to 90%. However, results for section 4 (Clinical pharmacy services) returned lower levels of positivity, with responses from 8 of the 15 questions being less than 60%. When asked what is preventing hospital pharmacists from achieving implementation of these activities, most answers were limited capacity, not considered to be a priority by managers, or other healthcare professionals do this. The last section focused on self-assessment and action planning, with fewer than 50% of positive responses; COVID-19 preparedness and vaccines with mixed positive and negative responses. Furthermore, implementation of the falsified medicines directive impacted the medication handling processes in 50% or more of the answers. Regarding sustainability, the majority (59%) of respondents felt a greater focus should be on sustainability from an organisational or management perspective. CONCLUSION: Results offer valuable insights into the hospital pharmacy profession throughout Europe. While there have been improvements in certain areas, challenges remain, particularly in implementing clinical pharmacy services. The findings provide a foundation for further dialogue, advocacy, and strategic planning to advance the role of hospital pharmacists and enhance patient care in Europe's healthcare systems.

Targeting 14-3-3θ-mediated TDP-43 pathology in amyotrophic lateral sclerosis and frontotemporal dementia mice.

Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are characterized by cytoplasmic deposition of the nuclear TAR-binding protein 43 (TDP-43). Although cytoplasmic re-localization of TDP-43 is a key event in the pathogenesis of ALS/FTD, the underlying mechanisms remain unknown. Here, we identified a non-canonical interaction between 14-3-3θ and TDP-43, which regulates nuclear-cytoplasmic shuttling. Neuronal 14-3-3θ levels were increased in sporadic ALS and FTD with TDP-43 pathology. Pathogenic TDP-43 showed increased interaction with 14-3-3θ, resulting in cytoplasmic accumulation, insolubility, phosphorylation, and fragmentation of TDP-43, resembling pathological changes in disease. Harnessing this increased affinity of 14-3-3θ for pathogenic TDP-43, we devised a gene therapy vector targeting TDP-43 pathology, which mitigated functional deficits and neurodegeneration in different ALS/FTD mouse models expressing mutant or non-mutant TDP-43, including when already symptomatic at the time of treatment. Our study identified 14-3-3θ as a mediator of cytoplasmic TDP-43 localization with implications for ALS/FTD pathogenesis and therapy.

Prevalence of Skeletal Fluorosis in Northern Tanzania: A Follow-Up Study.

OBJECTIVES: Skeletal fluorosis is a metabolic bone disease caused by excessive exposure to fluoride, predominantly through contamination of drinking water. This study aimed to identify all cases of skeletal fluorosis in Tindigani village situated in Northern Tanzania. This was done following changes in drinking water sources after a previous prevalence study in 2009 in this population. METHODS: In a door-to-door cross-sectional study of Tindigani village, a sample of residents was assessed for skeletal fluorosis and dental fluorosis. Diagnosis of skeletal fluorosis was based on pre-defined angles of deformity of the lower limbs. Dental fluorosis was diagnosed and graded using the Thylstrup and Fejerskov Index. Samples from current drinking water sources underwent fluoride analysis. RESULTS: Tindigani village had a population of 1,944 individuals. Of the 1,532 individuals who were screened, 45 had skeletal fluorosis, giving a prevalence of 3.3% (95% CI=2.4, 4.3). Dental fluorosis was present in 82.5% of those examined (95% CI=79.8, 85.3). Dental fluorosis was present in all individuals with skeletal fluorosis and at higher grades than in the rest of the population. Drinking water samples were collected from 28 sources. These included piped, surface, well, and borehole water sources. Fluoride concentrations ranged from 0.45-38.59 mg/L of fluoride. CONCLUSIONS: Skeletal fluorosis is an ongoing but preventable health problem in the current population. The delivery of sustainable low fluoride piped water to this community would be of clear health benefit. This has been addressed at a local level.

Genomic, epigenomic, and transcriptomic signatures of prostate cancer between African American and European American patients.

Prostate cancer is the second most common cancer in men in the United States, and racial disparities are greatly observed in the disease. Specifically, African American (AA) patients have 60% higher incidence and mortality rates, in addition to higher grade and stage prostate tumors, than European American (EA) patients. In order to narrow the gap between clinical outcomes for these two populations, genetic and molecular signatures contributing to this disparity have been characterized. Over the past decade, profiles of prostate tumor samples from different ethnic groups have been developed using molecular and functional assays coupled with next generation sequencing or microarrays. Comparative genome-wide analyses of genomic, epigenomic, and transcriptomic profiles from prostate tumor samples have uncovered potential race-specific mutations, copy number alterations, DNA methylation, and gene expression patterns. In this study, we reviewed over 20 published studies that examined the aforementioned molecular contributions to racial disparities in AA and EA prostate cancer patients. The reviewed genomic studies revealed mutations, deletions, amplifications, duplications, or fusion genes differentially enriched in AA patients relative to EA patients. Commonly reported genomic alterations included mutations or copy number alterations of FOXA1, KMT2D, SPOP, MYC, PTEN, TP53, ZFHX3, and the TMPRSS2-ERG fusion. The reviewed epigenomic studies identified that CpG sites near the promoters of PMEPA1, RARB, SNRPN, and TIMP3 genes were differentially methylated between AA and EA patients. Lastly, the reviewed transcriptomic studies identified genes (e.g. CCL4, CHRM3, CRYBB2, CXCR4, GALR1, GSTM3, SPINK1) and signaling pathways dysregulated between AA and EA patients. The most frequently found dysregulated pathways were involved in immune and inflammatory responses and neuroactive ligand signaling. Overall, we observed that the genomic, epigenomic, and transcriptomic alterations evaluated between AA and EA prostate cancer patients varied between studies, highlighting the impact of using different methods and sample sizes. The reported genomic, epigenomic, and transcriptomic alterations do not only uncover molecular mechanisms of tumorigenesis but also provide researchers and clinicians valuable resources to identify novel biomarkers and treatment modalities to improve the disparity of clinical outcomes between AA and EA patients.

A Door-to-Door Prevalence Survey of Dental Fluorosis in Tindigani, a Village in the Hai District of Northern Tanzania.

Fluoride in excess of the World Health Organisation limit of 1.5mg/L in drinking water can cause dental fluorosis (DF) in developing teeth. DF is a significant problem in the Hai District of Northern Tanzania, where there is limited access to safe piped water and groundwater is high in fluoride. A door-to-door prevalence survey of residents of Tindigani village was undertaken to assess current prevalence and severity of DF in the Hai District, and the effectiveness of previous interventions to promote low-fluoride drinking water, following a prevalence survey in 2009. DF was graded by trained assessors, utilising dental photography, and drinking water sources were sampled for chemical analysis. DF was endemic in the 563 people assessed, with a prevalence of 79.4% (CI=76.1-82.7%). Prevalence and severity were found to be higher in permanent teeth than deciduous teeth. Fluoride concentrations in non-piped water sources ranged from 2.5-38.6mg/L. Despite more households reporting the use of low-fluoride, piped water sources, compared to 2009 (82.8% versus 62%), DF remains a significant problem in Tindigani and other such communities where low-fluoride drinking water is not easily and reliably accessible. Policy makers must prioritise reliable access to low-fluoride water, especially for children as their permanent dentition develops.

Escherichia coli recombinant expression of SARS-CoV-2 protein fragments.

We have developed a method for the inexpensive, high-level expression of antigenic protein fragments of SARS-CoV-2 proteins in Escherichia coli. Our approach uses the thermophilic family 9 carbohydrate-binding module (CBM9) as an N-terminal carrier protein and affinity tag. The CBM9 module was joined to SARS-CoV-2 protein fragments via a flexible proline-threonine linker, which proved to be resistant to E. coli proteases. Two CBM9-spike protein fragment fusion proteins and one CBM9-nucleocapsid fragment fusion protein largely resisted protease degradation, while most of the CBM9 fusion proteins were degraded at some site in the SARS-CoV-2 protein fragment. All of the fusion proteins were highly expressed in E. coli and the CBM9-ID-H1 fusion protein was shown to yield 122 mg/L of purified product. Three purified CBM9-SARS-CoV-2 fusion proteins were tested and found to bind antibodies directed to the appropriate SARS-CoV-2 antigenic regions. The largest intact CBM9 fusion protein, CBM9-ID-H1, incorporates spike protein amino acids 540-588, which is a conserved region overlapping and C-terminal to the receptor binding domain that is widely recognized by human convalescent sera and contains a putative protective epitope.

It's time to talk about the M word.

Menopause is often seen as a taboo subject within the workplace, yet it affects around half of the population. Within dentistry, as of 2019, 92.6% of dental care professionals and 50.9% of dentists were women. The subject is one which is affecting the dental profession but appears to have not been discussed, with no literature on the effects that menopause can have on members of the dental team. Outside of dentistry, employers are starting to realise the effects that menopause can have on colleagues, friends and family.

Development and psychometric testing of the self-regulatory questionnaire for lung cancer screening (SRQ-LCS).

BACKGROUND: Research implicates psychological factors in low uptake of lung cancer screening. We developed and psychometrically tested a standardised measure of these psychological determinants in preparation for a prospective, longitudinal cohort study of screening uptake. METHODS: Leventhal's Common-Sense Model of Self-Regulation of Health and Illness provided the theoretical framework to generate the initial item pool. Items were refined during expert review and cognitive interviews which tested for face validity, redundancy, acceptability and comprehensibility. An online survey piloted the refined pool with 1500 current and former (quit ≤ 15 years) smokers aged 55-80. The response distributions, internal reliability and factor structure determined the final retained constructs. Regression analyses examined these constructs' associations with screening intention, smoking status and demographics. RESULTS: The final measure included seven factor-derived subscales (consequences, personal control, treatment control, illness coherence, emotional representation, behavioural response and appraisal, risk perception) with Cronbach's alphas ranging from 0.59 to 0.91 and four single-item questions (response efficacy for smoking cessation, treatment intention, perceived stigma and lung cancer survival). Most constructs were associated with smoking status and screening intention (p's < .05). CONCLUSIONS: The Self-Regulatory Questionnaire for Lung Cancer Screening (SRQ-LCS) is an acceptable, reliable and valid measure for investigating the psychological determinants of screening uptake.

Watch and learn? A systematic review comparing oral health educational videos with written patient information aimed at parents/carers or children.

Introduction Childhood caries remains a major UK health issue. The COVID-19 pandemic has necessitated rapid introduction of innovative practice to minimise footfall into dental clinics. Video-based oral health education could help promote oral health advice remotely and has been included in national guidance. This systematic review evaluates the impact of video education versus written patient leaflets on knowledge and oral health behaviours in parents/carers or children.Method PubMed and Medline were searched. In total, 47 articles were identified and relevance assessed by examining titles and abstracts. Seven full-text articles were assessed and reference lists manually screened for additional publications. Three studies met the inclusion criteria.Results Only one study assessed participant knowledge gain and found no statistical significance between scores at baseline and after video, leaflet or hygienist-led education. For oral health behaviour change, one study demonstrated no significant difference in plaque score improvements between video and leaflet groups. The other showed significant improvements in plaque and bleeding scores for both leaflet and video groups compared to the control.Conclusion This review identified too few studies with heterogeneity to make conclusions on the impact of written versusvideo oral health education. A standardised outcome measurement tool is needed to evaluate the impact of current educational videos on oral health behaviour and knowledge. There is potential for video to educate the masses as we navigate through this pandemic.

Pilot randomised controlled trial of the Risk Acceptance Ladder (RAL) as a tool for targeting health communications.

BACKGROUND: Improving adherence to self-protective behaviours is a public health priority. We aimed to assess the potential effectiveness and ease of use of an online version of the Risk Acceptance Ladder (RAL) in promoting help-seeking for cigarette smoking, excessive alcohol consumption, insufficient physical activity, or low fruit and vegetable consumption. METHODS: 843 UK adults were recruited, of whom 602 engaged in at least one risky behaviour. Those with no immediate plans to change (n = 171) completed a behaviour specific RAL. Participants were randomised to one of two conditions; a short message congruent (on-target, n = 73) or incongruent (off-target, n = 98) with their RAL response. Performance of the RAL was assessed by participants' ability to select an applicable RAL item and reported ease of use of the RAL. Effectiveness was assessed by whether or not participants clicked a link to receive information about changing their target behaviour. RESULTS: Two thirds (68.9%, 95% CI = 61.8%-75.3%) of participants were able to select an applicable RAL item that corresponded to what they believed would need to change in order to alter their target behaviour, with 64.9% (95% CI = 57.5%-71.7%) reporting that it was easy to select one option. Compared with the off-target group, participants allocated to the on-target group had greater odds of clicking on the link to receive information (31.5% vs 19.4%; OR = 2.07, 95% CI = 1.01-4.26). CONCLUSION: The Risk Acceptance Ladder may have utility as a tool for tailoring messages to prompt initial steps to engaging in self-protective behaviours.

Role of EphA4 in Mediating Motor Neuron Death in MND.

Motor neuron disease (MND) comprises a group of fatal neurodegenerative diseases with no effective cure. As progressive motor neuron cell death is one of pathological characteristics of MND, molecules which protect these cells are attractive therapeutic targets. Accumulating evidence indicates that EphA4 activation is involved in MND pathogenesis, and inhibition of EphA4 improves functional outcomes. However, the underlying mechanism of EphA4's function in MND is unclear. In this review, we first present results to demonstrate that EphA4 signalling acts directly on motor neurons to cause cell death. We then review the three most likely mechanisms underlying this effect.

Cognitive impairment in syphilis: Does treatment based on cerebrospinal fluid analysis improve outcome?

BACKGROUND: Individuals with previous syphilis may experience cognitive impairment. The goal of this study was to determine if those at high risk for laboratory-defined neurosyphilis are cognitively impaired, and whether treatment based on cerebrospinal fluid (CSF) findings results in better outcomes. METHODS: Participants had a new syphilis diagnosis, serum RPR titer ≥ 1:32 or peripheral blood CD4+ T cells ≤ 350/ul (in persons living with HIV) and did not endorse neurological symptoms. They underwent computerized cognitive assessment with the CogState. Thirty-two were randomized to either undergo lumbar puncture (LP) or to not undergo LP and 14 underwent LP; 64 were not randomized and 48 opted to undergo LP. RESULTS: Demographics, cognitive complaints and cognitive impairment did not differ between randomized and nonrandomized participants. Two-thirds were cognitively impaired, and impairment was not more common in those with cognitive complaints. The adjusted odds of increased severity of impairment were 3.8 times greater in those with CSF pleocytosis compared to those without. Time to cognitive normalization, improvement or decline did not differ between those who did not undergo LP and those who underwent LP and whose treatment was based on CSF analysis. Taking into account pre-treatment cognitive impairment, the risk of cognitive decline was lower in those with CSF pleocytosis treated for neurosyphilis compared to those without CSF pleocytosis not treated for neurosyphilis, (HR 0.24 (95% CI 0.07-0.88], p = 0.03). CONCLUSION: In individuals at high risk for laboratory-defined neurosyphilis, cognitive complaints are not a good indicator of cognitive impairment. Severity of cognitive impairment was greater in those with CSF pleocytosis. Identification and treatment of those with neurosyphilis may mitigate subsequent cognitive decline.

Balancing the dental boards.

The gender balance on boards is an important issue because any imbalance represents gender inequality and is not acceptable. We describe data that we have gathered on the current balance of the UK dental boards and then outline potential ways forward to address any imbalance.

Unbiased Label-Free Quantitative Proteomics of Cells Expressing Amyotrophic Lateral Sclerosis (ALS) Mutations in CCNF Reveals Activation of the Apoptosis Pathway: A Workflow to Screen Pathogenic Gene Mutations.

The past decade has seen a rapid acceleration in the discovery of new genetic causes of ALS, with more than 20 putative ALS-causing genes now cited. These genes encode proteins that cover a diverse range of molecular functions, including free radical scavenging (e.g., SOD1), regulation of RNA homeostasis (e.g., TDP-43 and FUS), and protein degradation through the ubiquitin-proteasome system (e.g., ubiquilin-2 and cyclin F) and autophagy (TBK1 and sequestosome-1/p62). It is likely that the various initial triggers of disease (either genetic, environmental and/or gene-environment interaction) must converge upon a common set of molecular pathways that underlie ALS pathogenesis. Given the complexity, it is not surprising that a catalog of molecular pathways and proteostasis dysfunctions have been linked to ALS. One of the challenges in ALS research is determining, at the early stage of discovery, whether a new gene mutation is indeed disease-specific, and if it is linked to signaling pathways that trigger neuronal cell death. We have established a proof-of-concept proteogenomic workflow to assess new gene mutations, using CCNF (cyclin F) as an example, in cell culture models to screen whether potential gene candidates fit the criteria of activating apoptosis. This can provide an informative and time-efficient output that can be extended further for validation in a variety of in vitro and in vivo models and/or for mechanistic studies. As a proof-of-concept, we expressed cyclin F mutations (K97R, S195R, S509P, R574Q, S621G) in HEK293 cells for label-free quantitative proteomics that bioinformatically predicted activation of the neuronal cell death pathways, which was validated by immunoblot analysis. Proteomic analysis of induced pluripotent stem cells (iPSCs) derived from patient fibroblasts bearing the S621G mutation showed the same activation of these pathways providing compelling evidence for these candidate gene mutations to be strong candidates for further validation and mechanistic studies (such as E3 enzymatic activity assays, protein-protein and protein-substrate studies, and neuronal apoptosis and aberrant branching measurements in zebrafish). Our proteogenomics approach has great utility and provides a relatively high-throughput screening platform to explore candidate gene mutations for their propensity to cause neuronal cell death, which will guide a researcher for further experimental studies.

Syntaxins 6 and 8 facilitate tau into secretory pathways.

Tau pathology initiates in defined brain regions and is known to spread along neuronal connections as symptoms progress in Alzheimer's disease (AD) and other tauopathies. This spread requires the release of tau from donor cells, but the underlying molecular mechanisms remained unknown. Here, we established the interactome of the C-terminal tail region of tau and identified syntaxin 8 (STX8) as a mediator of tau release from cells. Similarly, we showed the syntaxin 6 (STX6), part of the same SNARE family as STX8 also facilitated tau release. STX6 was previously genetically linked to progressive supranuclear palsy (PSP), a tauopathy. Finally, we demonstrated that the transmembrane domain of STX6 is required and sufficient to mediate tau secretion. The differential role of STX6 and STX8 in alternative secretory pathways suggests the association of tau with different secretory processes. Taken together, both syntaxins, STX6 and STX8, may contribute to AD and PSP pathogenesis by mediating release of tau from cells and facilitating pathology spreading.

Treatment with Commonly Used Antiretroviral Drugs Induces a Type I/III Interferon Signature in the Gut in the Absence of HIV Infection.

Tenofovir disoproxil fumarate (TDF) and emtricitabine (FTC) are used for HIV treatment and prevention. Previously, we found that topical rectal tenofovir gel caused immunological changes in the mucosa. Here, we assess the effect of oral TDF/FTC in three HIV pre-exposure prophylaxis trials, two with gastrointestinal and one with cervicovaginal biopsies. TDF/FTC induces type I/III interferon-related (IFN I/III) genes in the gastrointestinal tract, but not blood, with strong correlations between the two independent rectal biopsy groups (Spearman r = 0.91) and between the rectum and duodenum (r = 0.81). Gene set testing also indicates stimulation of the type I/III pathways in the ectocervix and of cellular proliferation in the duodenum. mRNA sequencing, digital droplet PCR, proteomics, and immunofluorescence confirm IFN I/III pathway stimulation in the gastrointestinal tract. Thus, oral TDF/FTC stimulates an IFN I/III signature throughout the gut, which could increase antiviral efficacy but also cause chronic immune activation in HIV prevention and treatment settings.

Onset of hippocampal network aberration and memory deficits in P301S tau mice are associated with an early gene signature.

Hyperphosphorylation and deposition of tau in the brain characterizes frontotemporal dementia and Alzheimer's disease. Disease-associated mutations in the tau-encoding MAPT gene have enabled the generation of transgenic mouse models that recapitulate aspects of human neurodegenerative diseases, including tau hyperphosphorylation and neurofibrillary tangle formation. Here, we characterized the effects of transgenic P301S mutant human tau expression on neuronal network function in the murine hippocampus. Onset of progressive spatial learning deficits in P301S tau transgenic TAU58/2 mice were paralleled by long-term potentiation deficits and neuronal network aberrations during electrophysiological and EEG recordings. Gene-expression profiling just prior to onset of apparent deficits in TAU58/2 mice revealed a signature of immediate early genes that is consistent with neuronal network hypersynchronicity. We found that the increased immediate early gene activity was confined to neurons harbouring tau pathology, providing a cellular link between aberrant tau and network dysfunction. Taken together, our data suggest that tau pathology drives neuronal network dysfunction through hyperexcitation of individual, pathology-harbouring neurons, thereby contributing to memory deficits.