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Buried under a flash avalanche while skiing, young married couple Jake and Zoe miraculously dig their way out only to discover themselves alone in an eerily silent, evacuated region and unable to contact the outside world.

Mycobacteria are members of the actinomycetes that grow by tip extension and lack apparent homologues of the known cell division regulators found in other rod-shaped bacteria. Previous work using static microscopy on dividing mycobacteria led to the hypothesis that these cells can grow and divide asymmetrically, and at a wide range of sizes, in contrast to the cell growth and division patterns observed in the model rod-shaped organisms. In this study, we test this hypothesis using live-cell time-lapse imaging of dividing Mycobacterium smegmatis labelled with fluorescent PBP1a, to probe peptidoglycan synthesis and label the cell septum. We demonstrate that the new septum is placed accurately at mid-cell, and that the asymmetric division observed is a result of differential growth from the cell tips, with a more than 2-fold difference in growth rate between fast and slow growing poles. We also show that the division site is not selected at a characteristic cell length, suggesting this is not an important cue during the mycobacterial cell cycle.

Introduction In 2020, surging cases of COVID‐19 meant that health services had to plan for crisis‐level triage. In the Australian Capital Territory, the Clinical Health Emergency Coordination Centre sought to develop a triage policy in collaboration with a range of consumer, carer and community groups. This study aims to map the collaborative development of the COVID‐19 ICU triage policy onto the principles of co‐production. Methods Interviews were conducted with facilitators, members of advocacy or consumer groups and clinicians who were involved in the development of the triage policy. Interviews were thematically analysed using both theory‐ and data‐driven approaches to, respectively, draw on the theoretical framework of co‐production, and to explore participants' perspectives relevant to but beyond the scope of this theoretical framework. Results The findings suggest that at each stage of the initiative, there were ways in which the principles of co‐production were met, and ways in which they were not met. One of the fundamental concerns that arose was about whether trying to solve a problem based on resources was compatible with a solution based on human rights. Conclusion Literature about co‐production has been critiqued for being limited to aspirational concerns, or implying co‐production is easily achievable. The current study contributes to existing research through the application of the theoretical framework of co‐production and exploring ways its aims were met and not met within a system‐level collaboration developing a high‐stakes health policy. Patient or Public Contribution This study has been conducted and written by researchers working from lived experience perspectives, and other researchers working from traditionally mainstream health disciplines, including psychology and medicine. Further, the study is about patient and public involvement in the development of a health policy. Thus it both embodies and is about non‐tokenistic collaboration between people with lived experience and other health professionals.

Correct chromosomal segregation, coordinated with cell division, is crucial for bacterial survival, but despite extensive studies, the mechanisms underlying this remain incompletely understood in mycobacteria. We report a detailed investigation of the dynamic interactions between ParA and ParB partitioning proteins in Mycobacterium smegmatis using microfluidics and time-lapse fluorescence microscopy to observe both proteins simultaneously. During growth and division, ParB presents as a focused fluorescent spot that subsequently splits in two. One focus moves towards a higher concentration of ParA at the new pole, while the other moves towards the old pole. We show ParB movement is in part an active process that does not rely on passive movement associated with cell growth. In some cells, another round of ParB segregation starts before cell division is complete, consistent with initiation of a second round of chromosome replication. ParA fluorescence distribution correlates with cell size, and in sister cells, the larger cell inherits a local peak of concentrated ParA, while the smaller sister inherits more homogeneously distributed protein. Cells which inherit more ParA grow faster than their sister cell, raising the question of whether inheritance of a local concentration of ParA provides a growth advantage. Alterations in levels of ParA and ParB were also found to disturb cell growth.

Background Two general approaches to prokaryotic live-cell imaging have been employed to date, growing bacteria on thin agar pads or growing bacteria in micro-channels. The methods using agar pads 'sandwich' the cells between the agar pad on the bottom and a glass cover slip on top, before sealing the cover slip. The advantages of this technique are that it is simple and relatively inexpensive to set up. However, once the cover slip is sealed, the environmental conditions cannot be manipulated. Furthermore, desiccation of the agar pad, and the growth of cells in a sealed environment where the oxygen concentration will be in gradual decline, may not permit longer term studies such as those required for the slower growing mycobacteria. Findings We report here a modified agar pad method where the cells are sandwiched between a cover slip on the bottom and an agar pad on top of the cover slip (rather than the reverse) and the cells viewed from below using an inverted microscope. This critical modification overcomes some of the current limitations with agar pad methods and was used to produce time-lapse images and movies of cell growth for Mycobacterium smegmatis and Mycobacterium bovis BCG. Conclusions This method offers improvement on the current agar pad methods in that long term live cell imaging studies can be performed and modification of the media during the experiment is permitted.

Diabetic retinopathy (DR) is a leading cause of blindness in adults worldwide. Early detection and intervention can prevent blindness; however, many patients do not receive their recommended annual diabetic eye examinations, primarily owing to limited access. To evaluate the safety and accuracy of an artificial intelligence (AI) system (the EyeArt Automated DR Detection System, version 2.1.0) in detecting both more-than-mild diabetic retinopathy (mtmDR) and vision-threatening diabetic retinopathy (vtDR). A prospective multicenter cross-sectional diagnostic study was preregistered (NCT03112005) and conducted from April 17, 2017, to May 30, 2018. A total of 942 individuals aged 18 years or older who had diabetes gave consent to participate at 15 primary care and eye care facilities. Data analysis was performed from February 14 to July 10, 2019. Retinal imaging for the autonomous AI system and Early Treatment Diabetic Retinopathy Study (ETDRS) reference standard determination. Primary outcome measures included the sensitivity and specificity of the AI system in identifying participants' eyes with mtmDR and/or vtDR by 2-field undilated fundus photography vs a rigorous clinical reference standard comprising reading center grading of 4 wide-field dilated images using the ETDRS severity scale. Secondary outcome measures included the evaluation of imageability, dilated-if-needed analysis, enrichment correction analysis, worst-case imputation, and safety outcomes. Of 942 consenting individuals, 893 patients (1786 eyes) met the inclusion criteria and completed the study protocol. The population included 449 men (50.3%). Mean (SD) participant age was 53.9 (15.2) years (median, 56; range, 18-88 years), 655 were White (73.3%), and 206 had type 1 diabetes (23.1%). Sensitivity and specificity of the AI system were high in detecting mtmDR (sensitivity: 95.5%; 95% CI, 92.4%-98.5% and specificity: 85.0%; 95% CI, 82.6%-87.4%) and vtDR (sensitivity: 95.1%; 95% CI, 90.1%-100% and specificity: 89.0%; 95% CI, 87.0%-91.1%) without dilation. Imageability was high without dilation, with the AI system able to grade 87.4% (95% CI, 85.2%-89.6%) of the eyes with reading center grades. When eyes with ungradable results were dilated per the protocol, the imageability improved to 97.4% (95% CI, 96.4%-98.5%), with the sensitivity and specificity being similar. After correcting for enrichment, the mtmDR specificity increased to 87.8% (95% CI, 86.3%-89.5%) and the sensitivity remained similar; for vtDR, both sensitivity (97.0%; 95% CI, 91.2%-100%) and specificity (90.1%; 95% CI, 89.4%-91.5%) improved. This prospective multicenter cross-sectional diagnostic study noted safety and accuracy with use of the EyeArt Automated DR Detection System in detecting both mtmDR and, for the first time, vtDR, without physician assistance. These findings suggest that improved access to accurate, reliable diabetic eye examinations may increase adherence to recommended annual screenings and allow for accelerated referral of patients identified as having vtDR.

Abstract Disclosure: E.M. Everett: None. R. Tiu: None. I. Melgoza: None. J. Graham: None. G. Villanueva: None. J. Menjivar Cruz: None. A. Rivas Deras: None. K. Udeze: None. J. Cushing: None. B. Escobar Barrios: None. J. Mendez: None. E. Shaw: None. C. Santana: None. B. Cobb: None. D. Polan: None. A. Naguib Guirguis: None. N. Ghaltakchyan: None. D. Chukwu: None. A. Manandhar: None. R. Hargrove: None. D. Cheng: None. M. Freeby: None. E. Ipp: None. T. Moin: None. Background: Diabetes technology, including continuous glucose monitors (CGMs) and insulin pumps, are associated with improved type 1 diabetes (T1D) outcomes compared to management with insulin injections and glucometer. However, prior studies have shown that only 40-60% of patients with T1D use diabetes technology, with even lower rates of 26-30% in safety-net settings. To address these gaps, the state of California expanded coverage policies between 2022-2023 to support guideline-concordant CGM and pump use among individuals with T1D; we evaluated diabetes technology use in two endocrinology clinics (one academic and one safety-net). Methods: We used 2024-2025 electronic health record (EHR) data from two large health systems, an academic hospital (AH) and a safety-net hospital (SNH). We first identified patients with T1D and then conducted manual chart reviews to assess insulin pump and CGM utilization. We used Chi-squared test to evaluate differences between sites and logistic regression to evaluate the association of glycemic control and any technology use. Results: Our analytic sample included 1,685 patients with T1D (SNH=186, AH=1,499) and 86% had available demographic data. The SNH had higher proportion of uninsured patients (8% vs. 1%, p<0.001) but lower proportions of younger patients (age <45; 55% vs 69%, p<0.001), male patients (45% vs. 51%, p<0.001), White patients (13% vs. 39%, p<0.001), and English-proficient patients (79% vs. 98%, p<0.001) compared to the AH. The majority of patients in both settings used at least one form of diabetes technology (SNH= 70%, AH= 91%, p<0.001); more patients used CGMs (SNH= 70%, AH= 90%, p<0.001) than insulin pumps (SNH=42%, AH=58%, p<0.001). Use of any diabetes technology was associated with significantly greater odds of achieving glycemic targets: patients using diabetes technology had a 2.3-fold higher odds of achieving an A1c <7% (OR 2.3, 95% CI 1.61-3.37), and nearly a 4-fold higher odds of achieving an A1c <8% (OR 3.94, 95% CI 1.87-5.34), compared to non-users. Conclusions: Our recent review of 2024-2025 EHR data shows that technology use among patients with T1D from two large real-world healthcare systems in California is substantially higher than previously reported averages—even in a safety-net setting. We also found that use of diabetes technology is associated with greater odds of achieving glycemic control. Our findings suggest that recent policy changes have helped expand access to CGMs and/or insulin pumps and that access to these technologies can play a critical role in achieving diabetes care goals in real-world settings. Presentation: Monday, July 14, 2025

Tuberculosis remains a global health problem, exacerbated by the increasing emergence of multi‐drug resistant strains. The identification of new drug targets and the discovery of new anti‐tuberculosis drugs is therefore a high priority. Although little is currently known about mycobacterial cell division, the process is essential for the survival and expansion of all bacterial species so may involve proteins that represent excellent drug targets. In this thesis, proven tools for the study of bacterial cell division such as live‐cell time‐lapse imaging and Fluorescent Repressor Operator System (FROS) were adapted for use in mycobacteria. Application of such techniques, fluorescent tagging of cell division proteins and deletion of parA in M. smegmatis helped to elucidate some interesting characteristics of mycobacterial cell division. In contrast to model organisms, live cell imaging and septal staining indicated that M. smegmatis can grow and divide asymmetrically and divides at a range of lengths suggesting a fundamentally different mechanism of division regulation. The chromosome was hypothesised to play a key role in cell division so was investigated further by labelling a specific chromosomal loci. The key finding was that M. smegmatis cells only contain 1 or 2 chromosomal copies and that regardless of cell length, the nucleoid occupies almost the entire intracellular space. To examine if the nucleoid organisation is important for cell division, a putative chromosome segregation gene parA was disrupted. The ΔparA mutant displayed a classic cell division phenotype characterised by the production of anuclear mini‐cells. The mechanism responsible for the ΔparA mutant phenotype was studied further by applying live cell imaging, FROS and expressing a ParA‐mCherry fusion protein. The data obtained from all work presented was collated and used to propose a novel model of bacterial cell division regulation applicable to mycobacteria where the nucleoid plays a central role and ParA is required to ensure correct nucleoid placement.

A short story is presented.