Detailed Instructions Research Articles

Response of Patients with Color Vision Defects to Worth 4-Dot and Duochrome Tests

Background: Individuals with congenital color vision defects (CVDs) are at greater risk of misidentifying colors, necessitating an investigation into their ability to distinguish colors accurately. This study aimed to assess how individuals with CVDs perceive the colors in the Worth four-dot (W4D) and duochrome tests. It also explored whether individuals with CVDs require more detailed instructions from optometrists and eye care providers during these tests. Method: Thirty-two participants with congenital CVDs were recruited for this study. Participants with ocular diseases were excluded based on a brief questionnaire. The participants underwent a W4D test to determine the colors of four circles and the duochrome test without using filters. Result: All the participants correctly identified the W4D colors, except for five participants (15.62%). Furthermore, among all the participants, two (one with deuteranopia and one with protanopia) failed to identify the colors correctly in the duochrome test (6.25%). Conclusions: The findings suggest that in the context of optometric assessment, it could be essential for optometrists to incorporate a preliminary inquiry into a patient’s ability to identify colors prior to performing diagnostic tests such as the W4D or duochrome tests. Incorrect responses from patients with CVDs might mislead optometrists regarding the proper outcomes of the test.

Evaluating the User Experience and Usability of the MINI Robot for Elderly Adults with Mild Dementia and Mild Cognitive Impairment: Insights and Recommendations

Introduction: In recent years, the integration of robotic systems into various aspects of daily life has become increasingly common. As these technologies continue to advance, ensuring user-friendly interfaces and seamless interactions becomes more essential. For social robots to genuinely provide lasting value to humans, a favourable user experience (UX) emerges as an essential prerequisite. This article aimed to evaluate the usability of the MINI robot, highlighting its strengths and areas for improvement based on user feedback and performance. Materials and Methods: In a controlled lab setting, a mixed-method qualitative study was conducted with ten individuals aged 65 and above diagnosed with mild dementia (MD) and mild cognitive impairment (MCI). Participants engaged in individual MINI robot interaction sessions, completing cognitive tasks as per written instructions. Video and audio recordings documented interactions, while post-session System Usability Scale (SUS) questionnaires quantified usability perception. Ethical guidelines were followed, ensuring informed consent, and the data underwent qualitative and quantitative analyses, contributing insights into the MINI robot’s usability for this demographic. Results: The study addresses the ongoing challenges that tasks present, especially for MD individuals, emphasizing the importance of user support. Most tasks require both verbal and physical interactions, indicating that MD individuals face challenges when switching response methods within subtasks. These complexities originate from the selection and use of response methods, including difficulties with voice recognition, tablet touch, and tactile sensors. These challenges persist across tasks, with individuals with MD struggling to comprehend task instructions and provide correct answers and individuals with MCI struggling to use response devices, often due to the limitations of the robot’s speech recognition. Technical shortcomings have been identified. The results of the SUS indicate positive perceptions, although there are lower ratings for instructor assistance and pre-use learning. The average SUS score of 68.3 places device usability in the “good” category. Conclusions: Our study examines the usability of the MINI robot, revealing strengths in quick learning, simple system and operation, and integration of features, while also highlighting areas for improvement. Careful design and modifications are essential for meaningful engagement with people with dementia. The robot could better benefit people with MD and MCI if clear, detailed instructions and instructor assistance were available.

Model-informed precision dosing of vancomycin in clinical practice: an intervention development study.

Current guidelines recommend dosing vancomycin based on the area under the concentration time curve (AUC) to maximise efficacy and minimise the risk of nephrotoxicity. The preferred approach to AUC-guided therapy is to apply model-informed precision dosing (MIPD). However, the adoption in clinical practice has been slow. We aimed to develop an intervention, including a standardised MIPD workflow and an implementation plan for vancomycin AUC-guided dosing, in a Swedish tertiary hospital. The intervention was developed in a framework-guided process. The design phase included stakeholder feedback (nurses, pharmacists, physicians), local data collection and feasibility testing of intervention components with parallel consideration of implementation aspects. The hypothesised relationships between the different components, implementation strategies and the mechanism of action resulting in expected outcomes were represented by a logic model. The final intervention consisted of a workflow for MIPD, with defined roles and responsibilities, as well as processes for data and information transfer. Details were provided in supportive documents; an instruction on therapeutic drug monitoring (TDM) sampling and documentation for nurses, and a detailed dosing software instruction for MIPD consultants and clinical pharmacists. Activities to facilitate implementation included the development of a local clinical routine for vancomycin dosing, staff training and recurring MIPD rounds. An intervention for MIPD, with an implementation plan for AUC-guided dosing of vancomycin, was developed for a tertiary hospital setting. The process can be used as guidance for other institutions with similar context wishing to initiate MIPD.

Exploring protein natural diversity in environmental microbiomes with DeepMetagenome.

Protein natural diversity offers a vast sequence space for protein engineering, and deep learning enables its detection from metagenomes/proteomes without prior assumptions. DeepMetagenome, a Python-based method, explores protein diversity through modules for training and analyzing sequence datasets. The deep learning model includes Embedding, Conv1D, LSTM, and Dense layers, with sequence feature analysis for data cleaning. Applied to metallothioneins from a database of over 146 million coding features, DeepMetagenome identified over 500 high-confidence metallothionein sequences, outperforming DIAMOND and CNN-based models. It showed stable performance compared to a Transformer-based model over 25 epochs. Among 23 synthesized sequences, 20 exhibited metal resistance. The tool also successfully explored the diversity of three additional protein families and is freely available on GitHub with detailed instructions.

Expressive 3D Facial Animation Generation Based on Local-to-Global Latent Diffusion.

3D Facial animations, crucial to augmented and mixed reality digital media, have evolved from mere aesthetic elements to potent storytelling media. Despite considerable progress in facial animation of neutral emotions, existing methods still struggle to capture the authenticity of emotions. This paper introduces a novel approach to capture fine facial expressions and generate facial animations using audio synchronization. Our method consists of two key components: First, the Local-to-global Latent Diffusion Model (LG-LDM) tailored for authentic facial expressions, which can integrate audio, time step, facial expressions, and other conditions towards possible encoding of emotionally rich yet latent features in response to possibly noisy raw audio signals. The core of LG-LDM is our carefully designed Facial Denoiser Model (FDM) for aligning the local-to-global animation feature with audio. Second, we redesign an Emotion-centric Vector Quantized-Variational AutoEncoder framework (EVQ-VAE) to finely decode the subtle differences under different emotions and reconstruct the final 3D facial geometry. Our work significantly contributes to the key challenges of emotionally realistic 3D facial animation for audio synchronization and enhances the immersive experience and emotional depth in augmented and mixed reality applications. We provide a reproducibility kit including our code, dataset, and detailed instructions for running the experiments. This kit is available at https://github.com/wangxuanx/Face-Diffusion-Model.

Smartphone Light Sensors as an Innovative Tool for Solar Irradiance Measurements.

In recent years, the teaching of experimental science and engineering has been revolutionized by the integration of smartphone sensors, which are widely used by a large portion of the population. Concurrently, interest in solar energy has surged. This raises the important question of how smartphone sensors can be harnessed to incorporate solar energy studies into undergraduate education. We provide comprehensive guidelines for using smartphone sensors in various conditions, along with detailed instructions on how to calibrate them with widely accessible clear-sky satellite data. This smartphone-based method is also compared with professional reference measurements to ensure consistency. This experiment can be easily conducted with most smartphones, basic materials, and a clear, open location over a few hours (methods). The findings demonstrate that smartphones, combined with simple resources, can accurately measure solar irradiance and support experiments on solar radiation physics, atmospheric interactions, and variations in solar energy across locations, cloud cover, and time scales. This approach provides a practical and accessible tool for studying solar energy, offering an innovative and engaging method for measuring solar resources.

A cost-effective protocol for single-cell RNA sequencing of human skin.

Single-cell RNA sequencing (scRNAseq) and flow cytometry studies in skin are methodologically complex and costly, limiting their accessibility to researchers worldwide. Ideally, RNA and protein-based analyses should be performed on the same lesion to obtain more comprehensive data. However, current protocols generally focus on either scRNAseq or flow cytometry of healthy skin. We present a novel label-free sample multiplexing strategy, building on the souporcell algorithm, which enables scRNAseq analysis of paired blood and skin samples. Additionally, we provide detailed instructions for simultaneous flow cytometry analysis from the same sample, with necessary adaptations for both healthy and inflamed skin specimens. This tissue multiplexing strategy mitigates technical batch effects and reduces costs by 2-4 times compared to existing protocols. We also demonstrate the effects of varying enzymatic incubation durations (1, 3, and 16 hours, with and without enzyme P) on flow cytometry outcomes. Comprehensive explanations of bioinformatic demultiplexing steps and a detailed step-by-step protocol of the entire experimental procedure are included. The protocol outlined in this article will make scRNAseq and flow cytometry analysis of skin samples more accessible to researchers, especially those new to these techniques.

Design and Implementation of Digital Integrated Circuit Based CPU

This paper presents a simplified 16-bit CPU design and its implementation using digital integrated circuits developed with Verilog hardware description language, while emphasizing simulation through the EDA tool. This design puts more emphasis on the critical roles played by the data path and the control unit, executing key instructions including arithmetic operations such as addition, subtraction, bitwise logic, and conditional jumps. This structurally modular CPU is both functioning and stable in nature, allowing support for future enhancements and enhancements as well, thereby serving as an effective model towards an understanding of the principles of CPU design. The execution of these basic instructions puts the core of the CPU under test and reaffirms its proper operation in respect of the detailed instruction set, while its extensibility design confirms further improvement possibilities for future complex instructions as well as finite-state machine optimization for the control unit. The present work not only proves the possibility of creating a fully working CPU, using readily available bare minimum resources to implement a prototype, but is also a breakthrough in future research with CPU architecture. The obtained results will shed light on CPU functionality optimization and extension, being the ground for building more powerful and effective processors in the future.

Exploring deceptions: cognitive strategies and dynamics in espionage

Abstract The paper examines deceptions employed during the Cold War with the purpose of exploring their underlying conceptions, mechanisms, and applications. It focuses on the interactions between the deceiver and the deceived, analyzing both sides’ perspectives. The examination centers on declassified guides, recently published as The Official C.I.A. Manual of Trickery and Deception, which contain detailed instructions concerning how spies should carry out their activities of deception. Three kinds of procedures were selected for analysis: behavior in public for men and women, group infiltration, and the use of secret recognition signals. To provide a practical visualization, the 2012 movie Argo, based on a real-life operation using the same principles, is complementarily analyzed. Merleau-Ponty’s approach to perception and sensation is considered along with Husserl’s concepts of sedimentation and lifeworld, while narrative semiotics and its framework guide the treatment of textuality and the approach to persuasion. The article conceives of sedimentation and contemplation of the lifeworld as mandatory components of a kind of deception that is primarily acted, reflecting upon how the spies’ interference affects the environment. The results indicate a common structure for deceptions that builds on what Greimas calls a “fiduciary contract.”

In Vitro Differentiation of Naive CD4+ T Cells into Pathogenic Th17 Cells in Mouse.

In vitro T cell differentiation techniques are essential for both functional and mechanistic investigations of CD4+ T cells. Pathogenic Th17 cells have been linked to a wide range of diseases in recent times, including multiple sclerosis (MS), rheumatoid arthritis, acute respiratory distress syndrome (ARDS), sepsis, and other autoimmune disorders. However, the currently known in vitro differentiation protocols have difficulty achieving high purity of pathogenic Th17 cells, with the induction efficiency often below 50%, which is a key challenge in in vitro experiments. In this protocol, we propose an enhanced in vitro culture and differentiation protocol for pathogenic Th17 cells, which is used to directly differentiate naive CD4+ T cells isolated from mouse spleens into pathogenic Th17 cells. This protocol provides detailed instructions on splenocyte isolation, purification of naive CD4+ T cells, and differentiation of pathogenic Th17 cells. Through this protocol, we can achieve a differentiation purity of approximately 90% for pathogenic Th17 cells, which meets the basic needs of many cellular experiments.

An Integrated Framework for Scientific and Ethical Review of Research Proposals

Research proposals must adhere to both scientific and ethical standards to ensure the integrity and impact of research endeavours. However, existing guidelines often fall short in providing comprehensive guidance for non-medical disciplines, requiring separate documents for scientific and ethical reviews, and lacking detailed instructions for specific steps such as risk-benefit analysis and the use of secondary data. This study addresses these challenges by developing a detailed, step-by-step guideline for preparing research proposals that seamlessly integrate scientific and ethical reviews. Through a qualitative research design, involving focus group discussions with ethics review committee members and key informant interviews with experts, combined with a comprehensive literature review, this study identified best practices and challenges in the current review process. The proposed guideline offers a structured approach to preparing research proposals, emphasizing clarity in research objectives, methodology, data analysis, and expected outcomes for scientific review. For ethical review, it provides detailed instructions on risk-benefit analysis, informed consent processes, privacy and confidentiality measures, and the ethical use of secondary or archival data. The guidelines were pilot-tested with researchers to ensure their usability and effectiveness, resulting in a refined tool that can standardize the proposal preparation process and improve the quality and integrity of research across various disciplines.

Systems Thinking Principles for Making Change

Traditionally, systems thinking support has relied on an ever-increasing plethora of systems tools, methods, and approaches. Arguably though, such support requires something different from, and more accessible than, detailed instruction on somewhat abstract laws and detailed principles and/or constitutive rules associated with conventional systems approaches or systems ‘tools of the trade’. For busy managers and decision makers working in often-stressful conditions, what is perhaps more valued are simple principles for enabling systems thinking in practice. Such principles should acknowledge and build on existing (multi)disciplinary skill sets and expertise, allowing for more meaningful interdisciplinary support amongst professions, as part of a nested transdisciplinary support for addressing wider social challenges. This monograph offers three principles of systems thinking in practice (STiP): relational STiP, perspective STiP, and adaptive STiP. They each have two sets of operational principles applicable to first-order and second-order practice, respectively. The three general principles are nested in an overriding principle of STiP as praxis (theory-informed action or thinking in practice) manifest in the need for being both systemic and systematic. The three principles represent a distilled expression of a systematic literacy of systems thinking, a literacy that speaks to the systemic sensibilities of Inter-relationships, Perspectives, and Boundaries (sometimes referred collectively as IPB), associated with any area of intervention. Drawing on metaphors of bricolage, conversation, and performance, and building on philosophical foundations of boundary critique, the three principles provide for a requisite systems literacy (as an emergent property of systemic sensibilities and systems thinking literacy) for enabling appropriate STiP capabilities to flourish when making a meaningful change.

Enhancing methods engineering education with a digital platform: usability and educational impact on industrial engineering students

IntroductionWork study aims to comprehend the potential of human work by assessing the duration of tasks and identifying methods for reducing them to enhance productivity and efficiency. On the other hand, methods engineering analyzes job processes to determine the most efficient and cost-effective techniques, resulting in improved operations. These areas are essential in the field of industrial engineering. Traditionally, they are taught using printed manuals and classroom-based instruction. Nevertheless, these conventional strategies frequently encounter difficulties engaging students and generating solid comprehension of the topics.MethodsThis study involved the development of a digital platform to improve methods engineering education. The platform adapts the curriculum of the methods engineering course offered at Universidad Panamericana. Each laboratory's practice includes detailed instructions and tasks for students to complete. Upon finishing, students submit their reports, which the professor then assesses. If the report meets the required standards, it is approved; otherwise, students must repeat the activities. An experience system serves as a tracker for course completion and keeps track of the student's progress.ResultsAn experimental group of 26 students who responded to Doll and Torkzadeh's end-user computer satisfaction survey tested the platform. Participants responded positively, particularly regarding the platform's accuracy (73.08% high satisfaction), ease of use (majority rated 5 for user-friendliness), and timeliness (majority satisfied with up-to-date information). However, the format dimension received mixed ratings, indicating areas for improvement. Additionally, participants appreciated the platform's ability to track progress and motivate the completion of laboratories, with 61.54% finding the content relevant and valuable for learning methods engineering concepts.DiscussionThe results suggest that integrating digital platforms into educational settings could significantly enhance educational settings, particularly in engineering courses where conventional approaches struggle to maintain student interest and engagement. Participants' positive responses support digital platforms' potential to complement and improve teaching methods. Future research will focus on integrating adaptive learning and generating micro-credentials to certify student expertise.

Measuring carbohydrate recognition profile of lectins on live cells using liquid glycan array (LiGA).

Glycans constitute a significant fraction of biomolecular diversity on cellular surfaces across all kingdoms of life. As the structure of glycans is not directly encoded by the organism's DNA, it is impossible to use high-throughput DNA technologies to study the role of cellular glycosylation or to understand how glycocalyx is recognized by glycan-binding proteins (GBPs). To address this gap, we recently described a liquid glycan array (LiGA) platform that allows profiling of glycan-GBP interactions on the surface of live cells in vitro and in vivo using next-generation sequencing. LiGA is a library of DNA-barcoded bacteriophages, where each clonal bacteriophage displays 5-1,500 copies of a glycan and the distinct DNA barcode inside each bacteriophage clone encodes the structure and density of the displayed glycans. Deep sequencing of the glycophages associated with live cells yields a glycan-binding profile of GBPs expressed on the surface of cells. This protocol provides detailed instructions for how to use LiGA to probe cell surface receptors and includes information on the preparation of glycophages, analysis by MALDI-TOF mass spectrometry, the assembly of a LiGA library and its deep sequencing. Using this protocol, we measure glycan-binding profiles of the immunomodulatory sialic acid-binding immunoglobulin-like lectins‑1, -2, -6, -7 and -9 expressed on the surface of different cell types. Compared with existing methods that require complex specialist equipment, this method allows users with basic molecular biology expertise to measure the precise glycan-binding profile of GBPs on the surface of any cell type expressing exogenous GBP within 2-3 d.

Disease trajectories before dementia: evidence from a large-scale community-based prospective study.

Systemic changes in multiple diseases may influence the onset of dementia. However, the specific temporality between exposure diseases and dementia remains uncertain. By characterising the full spectrum of temporal disease trajectories before dementia, this study aims to yield a global picture of precursor diseases to dementia and to provide detailed instructions for risk management and primary prevention of dementia. Using the multicentre, community-based prospective UK Biobank, we constructed disease trajectories before dementia utilising the phenome-wide association analysis, paired directional test and association quantification. Stratified disease trajectories were constructed by dementia subtypes, gender, age of diagnosis and Apolipoprotein E (ApoE) status, respectively. Our study population comprised 434 266 participants without baseline dementia and 4638 individuals with all-cause dementia. In total, 1253 diseases were extracted as potential components of the disease trajectory before dementia. We identified three clusters of disease trajectories preceding all-cause dementia, initiated by circulatory, metabolic and respiratory diseases occurring approximately 5-15 years before dementia. Cerebral infarction or chronic renal failure following chronic ischaemic heart disease was the specific trajectory before vascular dementia. Apolipoprotein E (ApoE) ε4 non-carriers exhibited more complex trajectories compared with carriers. Lipid metabolism disorders remained in the trajectories regardless of dementia subtypes, gender, age of diagnosis and ApoE status. This study provides a comprehensive view of the longitudinal disease trajectories before dementia and highlights the potential targets of midlife cardiometabolic dysfunction for dementia screening and prevention.

Integrating Real-Time Air Quality Monitoring, Ecological Momentary Assessment, and Spirometry to Evaluate Asthma Symptoms: Usability Study.

Individuals are exposed to a variety of indoor residential toxins including volatile organic compounds and particulates. In adults with asthma, such exposures are associated with asthma symptoms, asthma exacerbations, and decreased lung function. However, data on these exposures and asthma-related outcomes are generally collected at different times and not in real time. The integration of multiple platforms to collect real-time data on environmental exposure, asthma symptoms, and lung function has rarely been explored. This paper describes how adults with asthma perceive the acceptability and usability of three integrated devices: (1) residential indoor air quality monitor, (2) ecological momentary assessment (EMA) surveys delivered via a smartphone app, and (3) home spirometry, over 14 days. Participants (N=40) with uncontrolled asthma were mailed the Awair Omni indoor air quality monitor, ZEPHYRx home spirometer, and detailed instructions required for the in-home monitoring. The air quality monitor, spirometer, and EMA app were set up and tested during a videoconference or phone orientation with a research team member. Midway through the 14-day data collection period, participants completed an interview about the acceptability of the study devices or apps, instructional materials provided, and the setup process. At the end of the 14-day data collection period, participants completed a modified System Usability Scale. A random sample of 20 participants also completed a phone interview regarding the acceptability of the study and the impact of the study on their asthma. Participants ranged in age from 26 to 77 (mean 45, SD 13.5) years and were primarily female (n=36, 90%), White (n=26, 67%), college graduates (n=25, 66%), and residing in a single-family home (n=30, 75%). Most indicated that the air quality monitor (n=23, 58%), the EMA (n=20, 50%), and the spirometer (n=17, 43%) were easy to set up and use. Challenges with the EMA included repetitive surveys, surveys arriving during the night, and technical issues. While the home spirometer was identified as a plausible means to evaluate lung function in real time, the interpretation of the readings was unclear, and several participants reported side effects from home spirometer use. Overall, the acceptability of the study and the System Usability Scale scores were high. The study devices were highly acceptable and usable. Participant feedback was instrumental in identifying technical challenges that should be addressed in future studies.

Developing Canva-Based Sitekol on Collage Topic for First-Grade Public Elementary School Students

The study aims to create a learning media product using Canva for elementary school students to alleviate the lack of learning materials, which has been negatively impacting student academic performance. The research also seeks to evaluate the practicality and effectiveness of the learning media product. This study uses research and development with ADDIE design. The results of the analysis revealed that some students do not meet the minimum standards and require learning tools to improve their skills in cutting and sticking when studying collage topics. This research developed Sitekol media on collage topics in the design stage using Canva and Wordwall applications. The validity test results showed that Sitekol media received a 90% score from media experts for color appearance, image combinations, fonts, media layout, videos, and practical use. Material experts rated it at 95.3% for material aspects, levels of thinking, and supporting learning content. Students also found Canva-based Sitekol learning media to be beneficial as a teaching tool due to its accessibility, portability, convenience, interactive features, and detailed learning media instructions. The statistical tests conducted resulted in significant findings with high average values. Students responded very positively to the practicality of the media, giving an average score of 4.9 out of 5, indicating 98% of the ideal score. It can be concluded that using Canva-based Sitecol learning on grade 1 collage material exhibits practicality and validity as a learning medium.

Bowel preparation experiences and needs before follow-up colonoscopy in older adult postoperative colorectal cancer patients: A qualitative study.

The bowel preparation process prior to colonoscopy determines the quality of the bowel preparation, which in turn affects the quality of the colonoscopy. Colonoscopy is an essential procedure for postoperative follow-up monitoring of colorectal cancer (CRC) patients. Previous studies have shown that advanced age and a history of colorectal resection are both risk factors for inadequate bowel preparation. However, little attention has been paid to the bowel preparation experiences and needs of predominantly older adult postoperative CRC patients. To explore the experiences and needs of older adult postoperative CRC patients during bowel preparation for follow-up colonoscopy. Fifteen older adult postoperative CRC patients who underwent follow-up colonoscopy at a tertiary hospital in Shanghai were selected using purposive sampling from August 2023 to November 2023. The phenomenological method in qualitative research was employed to construct an interview outline and conduct semi-structured interviews with the patients. Colaizzi's seven-step analysis was utilized to organize, code, categorize, summarize, and verify the interview data. The results of this study were summarized into four themes and eight sub-themes: (1) Inadequate knowledge about bowel preparation; (2) Decreased physiological comfort during bowel preparation (gastrointestinal discomfort and sleep deprivation caused by bowel cleansing agents, and hunger caused by dietary restrictions; (3) Psychological changes during different stages of bowel preparation (pre-preparation: Fear and resistance due to previous experiences; during preparation: Irritation and helplessness caused by taking bowel cleansing agents, and post-preparation: Anxiety and worry while waiting for the colonoscopy); and (4) Needs related to bowel preparation (detailed instructions from healthcare professionals; more ideal bowel cleansing agents; and shortened waiting times for colonoscopy). Older adult postoperative CRC patients' knowledge of bowel preparation is not adequate, and they may encounter numerous difficulties and challenges during the process. Healthcare professionals should place great emphasis on providing instruction for their bowel preparation.

Using Mycobacterium smegmatis as a Bioindicator for Zinc-limited Growth Conditions in Mycobacteria.

Many bacteria build alternative ribosomes in Zn2+-limiting growth conditions by replacing Zn2+-binding ribosomal proteins with Zn2+-independent paralogs. Defining a system to study these alternative ribosomes has proven difficult because Zn2+ contamination in the laboratory is common. To address this issue, chelating agents are sometimes added to growth media, but this approach convolutes the biological response to gradual Zn2+limitation and is associated with ribosome hibernation. Here, detailed instructions are outlined for preparing media and seeding cultures for Zn2+-limited growth without adding chelators. Following this method, the model bacterium, Mycobacterium smegmatis, undergoes morphogenesis, which depends on alternative ribosomes. Because morphogenesis is tractable and only occurs in Zn2+-limiting conditions, M. smegmatis can be used as a bioindicator to verify biologically relevant growth conditions. Three bioindicator phenotypes (cell density, cell length, and coenzyme F420 fluorescence) that indicate Zn2+ limitation in the wild-type are described, and changes in these bioindicators for a deletion mutant that cannot build alternative ribosomes are outlined. Since trace Zn2+ contamination is difficult to control for each batch of media, and precise quantification of Zn2+ in each media preparation is overly burdensome, following this bioindicator phenotype is an accessible way to validate the preparation of Zn2+-limited growth media. To help identify proper conditions for Zn2+-limiting growth and alternative ribosome production, changes in the bioindicator phenotypes were profiled for Zn2+-contaminated or severely Zn2+-depleted preparations of Zn2+-limited media as well. Further details to achieve Zn2+-limiting growth and alternative ribosome production in M. tuberculosis are presented, along with the associated bioindicator phenotype. Overall, the detailed instructions and bioindicator phenotypes described here will help standardize the production of translationally active alternative ribosomes in mycobacteria.

Adverse Events and Efficacy of Acetazolamide in Meniere's Disease in a Vertigo Outpatient Clinic: A Retrospective Study.

Background Acetazolamide, a carbonic anhydrase inhibitor, is used in the treatment of several otolaryngological conditions, including Meniere's disease (MD). Despite its efficacy in reducing vertigo attacks, it is associated with a high rate of adverse events such as sensory disturbances, electrolyte imbalances, and liver dysfunction. The primary objective of this study is to evaluate the efficacy of acetazolamide in reducing vertigo attacks in MD patients. The secondary objective is to determine the incidence and severity of adverse drug events associated with acetazolamide treatment in this patient population. Methodology The study subjects were 70 patients who visited the vertigo outpatient clinic of our department from March 2019 to July 2024 and were diagnosed with definite or probable MD. Results A total of 15 cases had a history of acetazolamide prescription, with doses ranging from 125 to 750 mg/day. In total, 12 (80%) patients had symptoms of numbness in hands, feet, or other parts of the body, which caused discontinuation of medication after the initial prescription in five cases. Hypokalemia with liver dysfunction and hypokalemia with an open fracture of the distal end of the right radius due to staggering were observed in one patient each. In both cases of hypokalemia, Chinese herbal medicines containing licorice were prescribed in combination. Overall, 11 (73%) cases had a decrease in vertigo attacks. Hearing thresholds did not recover significantly after the prescription. Conclusions Although acetazolamide is effective in preventing vertigo attacks of MD, the rate of adverse events is high. Detailed instructions, careful dosage adjustment, periodic assessment of symptoms, and blood examinations for electrolyte and liver function are required.