Microfluidic single-cell measurements of oxidative stress as a function of cell cycle position.

Single-cell measurements routinely demonstrate high levels of variation between cells, but fewer studies provide insight into the analytical and biological sources of this variation. This is particularly true of chemical cytometry, in which individual cells are lysed and their contents separated, compared to more established single-cell measurements of the genome and transcriptome. To characterize population-level variation and its sources, we analyzed oxidative stress levels in 1278 individual Dictyostelium discoideum cells as a function of exogenous stress level and cell cycle position. Cells were exposed to varying levels of oxidative stress via singlet oxygen generation using the photosensitizer Rose Bengal. Single-cell data reproduced the dose-response observed in ensemble measurements by CE-LIF, superimposed with high levels of heterogeneity. Through experiments and data analysis, we explored possible biological sources of this heterogeneity. No trend was observed between population variation and oxidative stress level, but cell cycle position was a major contributor to heterogeneity in oxidative stress. Cells synchronized to the same stage of cell division were less heterogeneous than unsynchronized cells (RSD of 37-51% vs 93%), and mitotic cells had higher levels of reactive oxygen species than interphase cells. While past research has proposed changes in cell size during the cell cycle as a source of biological noise, the measurements presented here use an internal standard to normalize for effects of cell volume, suggesting a more complex contribution of cell cycle to heterogeneity of oxidative stress.

Microfluidic single-cell analysis of oxidative stress in Dictyostelium discoideum.

Microfluidic chemical cytometry is a powerful technique for examining chemical contents of individual cells, but applications have focused on cells from multicellular organisms, especially mammals. We demonstrate the first use of microfluidic chemical cytometry to examine a unicellular organism, the social amoeba Dictyostelium discoideum. We used the reactive oxygen species indicator dichlorodihydrofluorescein diacetate to report on oxidative stress and controlled for variations in indicator loading and retention using carboxyfluorescein diacetate as an internal standard. After optimizing indicator concentration, we investigated the effect of peroxide treatment through single-cell measurements of 353 individual cells. The peak area ratio of dichlorofluorescein to carboxyfluorescein increased from 1.69 ± 0.89 for untreated cells to 5.19 ± 2.72 for cells treated with 40 mM hydrogen peroxide. Interestingly, the variance of the data also increased with oxidative stress. While preliminary, these results are consistent with the hypothesis that heterogeneous stress responses in unicellular organisms may be adaptive.

Virulence adaptations of Pseudomonas aeruginosa isolated from patients with non-cystic fibrosis bronchiectasis.

Pseudomonas aeruginosa is a major pathogen in chronic lung diseases such as cystic fibrosis (CF) and non-cystic fibrosis bronchiectasis (nCFB). Much of our understanding regarding infections in nCFB patients is extrapolated from findings in CF with little direct investigation on the adaptation of P. aeruginosa in nCFB patients. As such, we investigated whether the adaptation of P. aeruginosa was indeed similar between nCFB and CF. From our prospectively collected biobank, we identified 40 nCFB patients who had repeated P. aeruginosa isolates separated by ≥6 months and compared these to a control population of 28 CF patients. A total of 84 nCFB isolates [40 early (defined as the earliest isolate in the biobank) and 41 late (defined as the last available isolate in the biobank)] were compared to 83 CF isolates (39 early and 44 late). We assessed the isolates for protease, lipase and elastase production; mucoid phenotype; swarm and swim motility; biofilm production; and the presence of the lasR mutant phenotype. Overall, we observed phenotypic heterogeneity in both nCFB and CF isolates and found that P. aeruginosa adapted to the nCFB lung environment similarly to the way observed in CF isolates in terms of protease and elastase expression, motility and biofilm formation. However, significant differences between nCFB and CF isolates were observed in lipase expression, which may allude to distinct characteristics found in the lung environment of nCFB patients. We also sought to determine virulence potential over time in nCFB P. aeruginosa isolates and found that virulence decreased over time, similar to CF.

Twenty-five-year outbreak of Pseudomonas aeruginosa infecting individuals with cystic fibrosis: identification of the prairie epidemic strain.

Transmissible strains of Pseudomonas aeruginosa have been described for cystic fibrosis (CF) and may be associated with a worse prognosis. Using a comprehensive strain biobank spanning 3 decades, we sought to determine the prevalence and stability of chronic P. aeruginosa infection in an adult population. P. aeruginosa isolates from sputum samples collected at initial enrollment in our adult clinic and at the most recent clinic visit were examined by a combination of pulsed-field gel electrophoresis and multilocus sequence typing and compared against a collection of established transmissible and local non-CF bronchiectasis (nCFB) isolates. A total of 372 isolates from 107 patients, spanning 674 patient-years, including 66 patients with matched isolates from initial and final encounters, were screened. A novel clone with increased antibacterial resistance, termed the prairie epidemic strain (PES), was found in 29% (31/107 patients) of chronically infected patients referred from multiple prairie-based CF centers. This isolate was not found in those diagnosed with CF as adults or in a control population with nCFB. While 90% (60/66 patients) of patients had stable infection over a mean of 10.8 years, five patients experienced strain displacement of unique isolates, with PES occurring within 2 years of transitioning to adult care. PES has been present in our cohort since at least 1987, is unique to CF, generally establishes chronic infection during childhood, and has been found in patients at the time of transition of patients from multiple prairie-based CF clinics, suggesting broad endemicity. Studies are under way to evaluate the clinical implications of PES infection.

How can physicians improve medication adherence and outcomes in dermatological conditions?

INTRODUCTION: Medication non-adherence is a major contributor to suboptimal disease treatment across medical specialties and is a particular hurdle with topicals. While adherence is a patient behavior affected by many socioeconomic and health system factors, physicians can play an important role in encouraging good adherence. AREAS COVERED: We discuss methods for measuring adherence, including ethics of such research, provide select examples of dermatology-specific adherence studies, and conclude with physician-focused practices to improve patients' adherence. Articles were selected from a PubMed search spanning 2003 to 10 December 2023, using the following terms: 'dermatology,' 'medication,' 'treatment,' 'adherence,' 'compliance,' and 'intervention.' EXPERT OPINION: Poor adherence to treatment is a major cause of poor treatment outcomes. As the goal of medical care is to achieve successful treatment outcomes, encouraging good adherence may be as much a foundation of care as making the right diagnosis and prescribing the right treatment. Taking a doctor-centric perspective on reasons for non-adherence may be more productive than simply finding fault with the patient. Establishing trust and accountability is a foundation for good adherence; after establishing the provider-patient relationship, physicians can improve adherence by incorporating behavioral and counseling strategies, communicating through technology, and advocating for distribution of validated educational information.

Tapinarof cream for the topical treatment of plaque psoriasis in adults.

INTRODUCTION: Plaque psoriasis, a chronic immune-mediated skin disorder, is characterized by well-demarcated erythematous plaques with silvery scales. This condition stems from complex interactions between genetic predisposition, immune dysregulation, and environmental triggers. Tapinarof downregulates the cytokine IL-17, diminishes the inflammatory infiltrate, and provides antioxidant properties while enhancing the expression of skin barrier proteins. AREAS COVERED: This review begins by assessing tapinarof's mechanism in treating plaque psoriasis. Subsequently, it examines the effectiveness and safety of tapinarof 1% cream in adult patients. EXPERT OPINION: Tapinarof 1% cream, which works by activating the aryl hydrocarbon receptor, is an FDA-approved treatment for adult plaque psoriasis. This therapy introduces a novel, nonsteroidal method for addressing inflammation and skin barrier issues, potentially serving as an alternative to conventional treatments. The once-daily, convenient cream formulation and favorable safety profile may enhance patient adherence, which is often poor with topical treatments. Tapinarof also maintains disease clearance for a mean of 4 months after treatment cessation.

Adherence to Psoriasis Therapies.

Understanding the underlying causes of nonadherence among patients with psoriasis and adopting strategies to address these issues may allow providers to share responsibility and work alongside patients to overcome these barriers. The review explores patient adherence to different types of psoriasis treatment, suggestions for interventions to overcome barriers, and methods to promote adherence that have been published in the literature.

Validity of facial skin analysis pore detection: A comparative analysis.

BACKGROUND: Reliable, objective measures to assess facial characteristics would aid in the assessment of many dermatological treatments. Previous work utilized an iOS application-based artificial intelligence (AI) tool compared to the "gold standard" computer-based and a physician assessment on five skin metrics (British Journal of Dermatology, 2013, 169, 474). The AI tool had superior agreement for all skin metrics except pores and subsequently underwent an algorithm update for its pore detection system. AIMS: This comparative analysis assessed the performance of the updated AI tool's pore scores across all Fitzpatrick skin phototypes to determine whether the AI tool more accurately represents a dermatologist's assessment of pores. PATIENTS/METHODS: Frontal facing photographs in uniform lighting conditions were taken of each participant. Percentile scores were generated by each of the four self-learning models of the updated AI tool. The pore percentile scores generated by the original and updated AI tool were used to rate "worse" pores among participant pairs. These ratings were compared to pore assessments performed by a "gold-standard" device and a board-certified dermatologist. RESULTS: Compared to the original pore detection tool and the computer-based program, models A and D had the highest concordance with the physician's pore assessments for Fitzpatrick skin phototypes III-IV and V-VI, respectively. CONCLUSIONS: The AI tool's pores detection update was successful in its ability to accurately detect pores on all Fitzpatrick skin types, improving on the performance of the AI prior to the update. Responsibly developed AI tools that can accurately and reliably detect skin metrics across diverse Fitzpatrick skin types can facilitate dermatologic evaluation, individualize treatment, and determine treatment response.

Lethality and cooperation of Pseudomonas aeruginosa quorum-sensing mutants in Drosophila melanogaster infection models.

The virulence profiles of Pseudomonas aeruginosa quorum-sensing (QS) mutants were assessed in Drosophila melanogaster feeding and nicking infection models. Functional RhlIR and LasIR QS systems were required for killing in the fly feeding infection model but were not essential in the fly nicking infection model. Mixed infections between PAO1 and strains harbouring mutations in lasR, rhlI and lasI rhlI resulted in increased lethality in the fly feeding model compared with either isolate alone. These results suggested that the parental strain could cooperate with QS mutants in the Drosophila feeding infection model. Finally, the mixed infection between PAO1 and an rhlR mutant resulted in spiteful behaviour and reduced pathogenicity of the mixed culture.

Defining Expertise in Cancer Nursing Practice.

BACKGROUND: Thirty-five years ago, Benner defined an expert nurse as one who applies deep knowledge and experience across different contexts and clinical situations. Since that time, there has been little exploration of expertise in cancer nursing. OBJECTIVES: To explore and describe characteristics of expert cancer nurses and to consider whether Benner's typology of an expert nurse remains relevant in today's complex oncology settings. METHODS: An exploratory, descriptive study using audio-recorded focus group methodology was undertaken. Audio-recordings were transcribed, and an inductive thematic analysis approach applied to the data. Nurses also documented key characteristics of expert practice on Post-it notes to illustrate dominant characteristics. RESULTS: Twenty-four registered nurses from a comprehensive cancer center in Australia took part in 1 of 3 focus groups. Seven key themes were identified: knowledge, leadership, adaptability, communication, motivation, patient-centered care, organization, and culture. Key word cloud characteristics included knowledge, compassion, motivation, experience, and communication. CONCLUSIONS: Many of the expert characteristics identified in this study reflect traits common to other nursing specialty groups. Of particular relevance to cancer nurses was "adaptability," reflecting the complexity of contemporary cancer care and reaffirming Benner's definition of an expert nurse as one who can fluidly connect knowledge and experience to unfamiliar practice contexts. IMPLICATIONS FOR PRACTICE: Understanding characteristics of expert cancer nurses may help inform and support professional practice advancement and guide future research about select characteristics of expert cancer nurses to patient- and system-level outcomes.

Epidemiology and natural history of Pseudomonas aeruginosa airway infections in non-cystic fibrosis bronchiectasis.

The natural history and epidemiology of Pseudomonas aeruginosa infections in non-cystic fibrosis (non-CF) bronchiectasis is not well understood. As such it was our intention to determine the evolution of airway infection and the transmission potential of P. aeruginosa in patients with non-CF bronchiectasis. A longitudinal cohort study was conducted from 1986-2011 using a biobank of prospectively collected isolates from patients with non-CF bronchiectasis. Patients included were ≥18 years old and had ≥2 positive P. aeruginosa cultures over a minimum 6-month period. All isolates obtained at first and most recent clinical encounters, as well as during exacerbations, that were morphologically distinct on MacConkey agar were genotyped by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). A total of 203 isolates from 39 patients were analysed. These were compared to a large collection of globally epidemic and local CF strains, as well as non-CF isolates. We identified four patterns of infection in non-CF bronchiectasis including: 1) persistence of a single strain (n=26; 67%); 2) strain displacement (n=8; 20%); 3) temporary disruption (n=3; 8%); and 4) chaotic airway infection (n=2; 5%). Patterns of infection were not significant predictors of rates of lung function decline or progression to end-stage disease and acquisition of new strains did not associate with the occurrence of exacerbations. Rarely, non-CF bronchiectasis strains with similar pulsotypes were observed in CF and non-CF controls, but no CF epidemic strains were observed. While rare shared strains were observed in non-CF bronchiectasis, whole-genome sequencing refuted patient-patient transmission. We observed a higher incidence of strain-displacement in our patient cohort compared to those observed in CF studies, although this did not impact on outcomes.

Phenotypic and Genotypic Comparison of Epidemic and Non-Epidemic Strains of Pseudomonas aeruginosa from Individuals with Cystic Fibrosis.

Epidemic strains of Pseudomonas aeruginosa have been found worldwide among the cystic fibrosis (CF) patient population. Using pulse-field gel electrophoresis, the Prairie Epidemic Strain (PES) has recently been found in one-third of patients attending the Calgary Adult CF Clinic in Canada. Using multi-locus sequence typing, PES isolates from unrelated patients were found to consistently have ST192. Though most patients acquired PES prior to enrolling in the clinic, some patients were observed to experience strain replacement upon transitioning to the clinic whereby local non-epidemic P. aeruginosa isolates were displaced by PES. Here we genotypically and phenotypically compared PES to other P. aeruginosa epidemic strains (OES) found around the world as well as local non-epidemic CF P. aeruginosa isolates in order to characterize PES. Since some epidemic strains are associated with worse clinical outcomes, we assessed the pathogenic potential of PES to determine if these isolates are virulent, shared properties with OES, and if its phenotypic properties may offer a competitive advantage in displacing local non-epidemic isolates during strain replacement. As such, we conducted a comparative analysis using fourteen phenotypic traits, including virulence factor production, biofilm formation, planktonic growth, mucoidy, and antibiotic susceptibility to characterize PES, OES, and local non-epidemic isolates. We observed that PES and OES could be differentiated from local non-epidemic isolates based on biofilm growth with PES isolates being more mucoid. Pairwise comparisons indicated that PES produced significantly higher levels of proteases and formed better biofilms than OES but were more susceptible to antibiotic treatment. Amongst five patients experiencing strain replacement, we found that super-infecting PES produced lower levels of proteases and elastases but were more resistant to antibiotics compared to the displaced non-epidemic isolates. This comparative analysis is the first to be completed on a large scale between groups of epidemic and non-epidemic CF P. aeruginosa isolates.