Expression and functional analysis of a recombinant aquaporin Z from Antarctic Pseudomonas sp. AMS3.

Aquaporin (AQP) is a water channel protein from the family of transmembrane proteins which facilitates the movement of water across the cell membrane. It is ubiquitous in nature, however the understanding of the water transport mechanism, especially for AQPs in microbes adapted to low temperatures, remains limited. AQP also has been recognized for its ability to be used for water filtration, but knowledge of the biochemical features necessary for its potential applications in industrial processes has been lacking. Therefore, this research was conducted to express, extract, solubilize, purify, and study the functional adaptations of the aquaporin Z family from Pseudomonas sp. AMS3 via molecular approaches. In this study, AqpZ1 AMS3 was successfully subcloned and expressed in E. coli BL21 (DE3) as a recombinant protein. The AqpZ1 AMS3 gene was expressed under optimized conditions and the best optimized condition for the AQP was in 0.5 mM IPTG incubated at 25°C for 20 h induction time. A zwitterionic mild detergent [(3-cholamidopropyl) dimethylammonio]-1-propanesulfonate was the suitable surfactant for the protein solubilization. The protein was then purified via affinity chromatography. Liposome and proteoliposome was reconstituted to determine the particle size using dynamic light scattering. This information obtained from this psychrophilic AQP identified provides new insights into the structural adaptation of this protein at low temperatures and could be useful for low temperature application and molecular engineering purposes in the future.

Arterial Gas Embolism in Breath-Hold Diver.

An arterial gas embolism (AGE) is a potentially fatal complication of scuba diving that is related to insufficient exhalation during ascent. During breath-hold diving, an arterial gas embolism is unlikely because the volume of gas in the lungs generally cannot exceed the volume at the beginning of the dive. However, if a diver breathes from a gas source at any time during the dive, they are at risk for an AGE or other pulmonary overinflation syndromes (POIS). In this case report, a breath-hold diver suffered a suspected AGE due to rapidly ascending without exhalation following breathing from an air pocket at approximately 40 feet.

Translation and validation study of obstructive sleep apnoea (OSA-18) questionnaire into Bahasa Malaysia (MALAY OSA-18).

INTRODUCTION: According to the American Academy of Sleep Medicine, obstructive sleep apnoea (OSA) is sleep-related breathing disorder that involves a decrease or complete halt in airflow despite an ongoing effort to breathe. The OSA-18 questionnaire is a short and self-administered questionnaire to assess paediatric patients' symptoms and quality of life with obstructive sleep apnoea. MATERIALS AND METHODS: This cross-sectional study is conducted at the Otorhinolaryngology-Head and Neck Surgery (ORL-HNS) clinic of Hospital Universiti Sains Malaysia. The forward and backward translation of the OSA- 18 questionnaire into the Malay language (Malay OSA-18) was performed and tested for content and face validity. The questionnaire's internal validity and reliability were tested using Pearson's correlation, Cronbach α and inter-reliability coefficient tests. The psychometric properties (validity, reliability and reproducibility) were assessed. RESULTS: We observed 84 patients ranging from six months up to 12 years of age. The mean age was 8 years old, and 63.1% were male patients. Among the samples, 96.4% presented with palatine tonsillar enlargement, and 84.5% presented with adenoid tonsillar enlargement. Based on the questionnaire the patient's caregiver answered, Pearson's correlation demonstrated that all the symptom scales correlate and measure the same things. The Cronbach's α coefficient value for each symptom scale was acceptable, within 0.6-0.8. The total Cronbach's α coefficient value was 0.89. The test-retest evaluation was excellent, with the value of intraclass correlation (ICC) more than 0.90. CONCLUSION: The Malay version of the OSA-18 questionnaire is equivalent to the original English version. It is an effective tool to assess the paediatric OSA patient's symptoms and quality of life based on the obtained validity, reliability and reproducibility values. Therefore, it is recommended to be a screening tool in daily practice.

Spatiotemporal analysis and forecasting of air quality in the greater Dhaka region and assessment of a novel particulate matter filtration unit.

Bangladesh is one of the most polluted nations in the world, with an average Air Quality Index (AQI) of 161 in 2021; its capital, Dhaka, has the worst air quality of any major city in the world. The present study aims to analyze the spatiotemporal distribution of air quality indicators in the greater Dhaka region, forecast weekly AQI, and assess the performance of a novel particulate matter filtration unit in removing particulate matter. Air quality indicators remained highest during the dry season with an average of 128.5 µm/m3, while the lowest concentration was found in the monsoon season with an average of 19.096 µm/m3. Analysis revealed a statistically significant annual increasing trend of CO, which was associated with the growing number of brick kilns and usage of high-sulfur diesel. Except for the pre-monsoon AQI, concentrations of both seasonal and yearly AQI and PM2.5 showed decreasing trend, though predominantly insignificant, demonstrating the improvement in air quality. Prevailing winds influenced the seasonal distribution of tropospheric CO & NO2. The study also employed a seasonal autoregressive integrated moving average (ARIMA) model to forecast weekly AQI values. ARIMA (3,0,4) (3,1,3) at the 7-periodicity level performed best forecasting the AQI values among all developed models with low root mean square error (RMSE)-29.42 and mean absolute percentage error (MAPE)-13.11 values. The predicted AQI values suggested that the air quality would remain unhealthy for most weeks. The experimental simulation of the particulate matter filtration unit, designed in the shape of a road divider, generated substantial cyclonic motion while maintaining a very minimal pressure drop. In the real-world scenario, using only cyclonic separation and dry deposition, the suggested air filtration system removed 40%, 44%, and 42% of PM2.5, PM10, and TSP, respectively. Without employing filters, the device removed significant amounts of particulate matter, implying enormous potential to be used in the study area. The study could be useful for policy makers to improve urban air quality and public health in Bangladesh and in other developing countries.

Eosinophilic/ T cell chorionic vasculitis.

No abstract available.

Mental Health Status during COVID-19 Pandemic and its Relationship with Economic Hardship and Financial Threat among Rural Population in Sarawak, Malaysia.

Background During the COVID-19 pandemic, job and income loss, social isolation may aggravate mental health, particularly among the most vulnerable groups. Objective To assess the current mental health situation among the rural population in Samarahan district and determine the relationship between economic hardships, financial threats, and mental health status. Method This study was a cross-sectional study conducted among the 530 households in the Samarahan district. A multistage cluster sampling technique was used to select the participants in this study. Data were collected by face-to-face interview using a structured questionnaire. The mental health status was assessed by using a validated and translated DASS-21 scale. Data analysis was done by SPSS version 27.0. A p-value of ≤ .05 was considered statistically significant. Result Analysis showed that two-thirds of the respondents (66.6%) had no mental health problem. Meanwhile, 22.4% had anxiety, 1% had depression, and 0.19% had stress. Anxiety and depression accounted for 5.8% and stress and anxiety 1.3%. However, 2.64% had stress, anxiety, and depression. In bivariate analysis, age, monthly income, type of job, economic hardship, and expenditure difference appeared to be significant predictors of mental health problems (p < .05). Conclusion Depression, anxiety, and stress pose a significant threat to the rural population's health. Therefore, public health practitioners and policymakers need to address this to minimise the pandemic's impact on mental health and provide psychological support, particularly among the most affected group.

The effect of emergency department pharmacists on drug overuse and drug underuse in patients with an ADE-related hospitalisation: a controlled intervention study.

BACKGROUND: Drug overuse or drug underuse are the most common causes of adverse drug events and can lead to hospital admissions. Using clinical pharmacists in the emergency department may improve patient safety as they are specialised in recognising of adverse drug events and tackling drug overuse and drug underuse. This study tested the effect of an emergency department pharmacist on the number of medication changes for drug overuse and drug underuse taking place in patients with an adverse drug event-related hospitalisation following an emergency department visit. METHODS: A multicenter prospective non-randomized controlled intervention study was conducted in a university hospital and a general teaching hospital. Trained emergency department pharmacists included patients in the intervention group with a hospital admission related to an adverse drug event. The interdisciplinary intervention consisted of a pharmacist-led medication review, patient counselling regarding medication, and information transmission to general practitioners and community pharmacies after discharge. The control patients were also admitted after an emergency department visit and received the usual care. The primary outcome was the number of medication changes for drug overuse and drug underuse that took place during hospital admission and persisted 6 months thereafter. Poisson regression analysis was used to estimate the difference in these medication changes between the intervention group and the control group. RESULTS: A total of 216 patients were included (intervention group 104, control group 112). In the intervention group, 156 medication changes for drug overuse and drug underuse persisted 6 months after admission compared to 59 in the control group (adjusted rate ratio 1.22 [95%CI 1.01-1.49] p = 0.039). CONCLUSION: Emergency department pharmacists do contribute to reduction of drug overuse and drug underuse of medication in patients with a hospitalisation related to adverse drug events after an emergency department visit.

Assessment of POFA -Cementitious material as backfill barrier in DSRS borehole disposal: 226Ra confinement.

Disused Sealed Radioactive Sources (DSRS) borehole disposal is an innovative concept recommended by international atomic energy agency (IAEA) to improve the safety and security of the management end point for these sources. A green application of Palm Oil Fuel Ash (POFA) as a supplementary material for cementitious backfill barrier in DSRS borehole disposal facility is proposed. Samples with up to 50% POFA replacement complied with the mechanical and hydraulic performance requirements for backfill barriers in retrievable radioactive waste disposal facilities. The structures of one year old OPC and optimum OPC-POFA cement backfills were evaluated using FESEM, XRD, EDXRF, BET, and TGA and their 226 Ra confinement performances were assessed. 30% POFA replacement improved the geochemical conditions by reducing competitive Ca2+ release into the disposal environment. It enhanced 226Ra confinement performance independently on the amount of water intrusion or releases below 2% of 1 Ci source. The improved performance is attributed to the higher fraction of active sites of OPC-POFA backfill compared to that of OPC backfill. 226Ra sorption onto C-S-H is irreversible, spontaneous, endothermic, and independent on the degree of the surface filling. The provided experimental data and theoretical analysis proved the feasibility of this green use of POFA in reducing the radiological hazard of 226Ra.

Severe central airway stenosis and tracheomalacia in hunter syndrome.

Hunter Syndrome is a genetic disease characterized by deficiency of Iduronate-2-Sulfatase enzyme activity, resulting in accumulation of glycoaminoglycans in various organs including the central airways. We report a case of severe tracheomalacia and airway stenosis at Hospital Sultanah Aminah, Johor Bahru, Malaysia requiring mechanical ventilation in a middle aged gentleman who was previously undiagnosed of mucopolysaccharidosis. The patient underwent emergency tracheostomy for failed intubation, when he presented with shortness of breath and acute respiratory failure. A contrast-enhanced computed tomography of the neck and thorax revealed that the trachea distal to the tracheostomy tube had collapsed with narrowed right and left main bronchus. These findings were confirmed via direct visualization of the airway through a flexible bronchoscopy. Eventually, a tracheal stenting were performed to maintain the airway patency and assist in weaning off from mechanical ventilation. Further investigations to identify the aetiology of the central airway stenosis revealed elevated urinary glycoaminoglycans and the absence of iduronate-2-Sulfatase activity tested on dried blood spots, thus confirming the diagnosis of Hunter Syndrome. Managing mucopolysacharidosis with central airway obstruction requires multidisciplinary team effort in handling the difficult airway, anaesthesiology risk, potential comorbidities and providing genetic counselling.

Drone technology in maternal healthcare in Malaysia: A narrative review.

INTRODUCTION: The vast advancement of technology and breakthrough in high-tech disciplines created multiple areas of research activities, including the emergence of the medical drone. Malaysia, a rapidly developing country in Southeast Asia is on track to achieving high-income status. However, the stagnant growth of Malaysian maternal healthcare does not run parallel with the aspiration. This review paper assessed and reported narratively the current condition of maternal healthcare in Malaysia, the possible application of drones in improving the sector, exploring in detail several challenges, and providing recommendations for experts in studying the rising technological phenomena. MATERIALS AND METHODS: A literature search was done from June 2019 to November 2019 with restrictions to the English language. The search was performed in ScienceDirect, PubMed, and EMBASE databases, using a combination of search terms related to drones, Unmanned Aerial Vehicles (UAV), Unmanned Aerial Systems (UAS), maternal, obstetric, healthcare, medical products transportation and Malaysia. A discourse analysis followed and a narrative review was provided on this subject. RESULTS AND DISCUSSION: The validated ability of drones in the delivery of blood products is highlighted as a possible application in improving maternal healthcare in Malaysia, particularly in the state of Sabah. Five key challenges are identified: infrastructure, technicalities, regulations, expertise, and social acceptance. Future predictions of drone technology in healthcare were outlined with the suggestion of three principle arms of application. CONCLUSION: The usage of the medical drone in medical products transportation supports the objectives of WHO MDG 5 for Malaysian maternal health. A study on the impact of drones in reducing the maternal mortality ratio is recommended for further exploration.

Assessment of Cyanex 301 impregnated resin for its potential use to remove cobalt from aqueous solutions.

The feasibility of using Cyanex 301 impregnated onto biopolymer resin for removing cobalt from radioactive waste streams is investigated. The emulsion compositions for resin preparation were optimized to ensure compliance with nuclear grade resin specifications on particle size distribution and chemical stabilities using multi-variant analysis. The enhanced chemical stabilities of the resins in different solutions are attributed to the interaction between Cyanex 301 and the emulsifier with alginate via a reaction with the glycosidic bond. The structure and morphology of the three optimized samples were studied using FTIR, SEM, and TGA and their end-of-life cycle treatment was preliminarily investigated. Equilibrium sorption investigations revealed that the sorption is taking place via bi-layers onto two sites on the surface layer independently from the cobalt contamination level and the nature of these sites were characterized by determining the energy barriers and fraction of the weak and strong sites. Cobalt adsorption is chemical spontaneous endothermic reaction and the loaded material spectroscopic analysis and thermodynamics calculations referred to chemical coordination and hydrogen bonding. The enhanced radiological stabilities of the optimum resins at irradiation fields <100 KGy were explained and the relations between the loss of exchange capacity and the doses were quantified. The resins could be regenerated using 0.5 M HCl and the effect of the elution cycles on cobalt uptake was presented.

Personalised management of Chronic Obstructive Pulmonary Disease (COPD): Malaysian consensus algorithm for appropriate use of inhaled corticosteroid (ICS) in COPD patients.

BACKGROUND: Regarding the long-term safety issues with the use of inhaled corticosteroids (ICS) and the clinical predominance of dual bronchodilators in enhancing treatment outcomes in chronic obstructive pulmonary disease (COPD), ICS is no longer a "preferred therapy" according to the Global Initiative for Chronic Obstructive Lung Disease except on top of a dual bronchodilator. This has necessitated a change in the current therapy for many COPD patients. OBJECTIVE: To determine a standardised algorithm to reassess and personalise the treatment COPD patients based on the available evidence. METHODS: A consensus statement was agreed upon by a panel of pulmonologists in from 11 institutes in Malaysia whose members formed this consensus group. RESULTS: According to the consensus, which was unanimously adopted, all COPD patients who are currently receiving an ICS-based treatment should be reassessed based on the presence of co-existence of asthma or high eosinophil counts and frequency of moderate or severe exacerbations in the previous 12 months. When that the patients meet any of the aforementioned criteria, then the patient can continue taking ICS-based therapy. However, if the patients do not meet the criteria, then the treatment of patients need to be personalised based on whether the patient is currently receiving long-acting beta-agonists (LABA)/ICS or triple therapy. CONCLUSION: A flowchart of the consensus providing a guidance to Malaysian clinicians was elucidated based on evidences and international guidelines that identifies the right patients who should receive inhaled corticosteroids and enable to switch non ICS based therapies in patients less likely to benefit from such treatments.

Divining responder populations from survival data.

BACKGROUND: Biomarkers that predict treatment response are the foundation of precision medicine in clinical decision-making and have the potential to significantly improve the efficiency of clinical trials. Such biomarkers may be identified before clinical testing but many trials enroll unselected populations. We hypothesized that time-varying treatment effects in unselected trials may result from identifiable responder subpopulations that may have associated biomarkers. MATERIALS AND METHODS: We first simulated scenarios of clinical trials with biomarker populations of varying prevalence and prognostic and predictive associations to illustrate the impact of subgroup-specific effects on overall population estimates. To show a real-world example of time-dependent treatment effects resulting from a prognostic and predictive biomarker, we re-analyzed data from a published clinical trial (RTOG, Radiation Therapy Oncology Group, 9402). We then demonstrated a quantitative framework to fit survival data from clinical trials using statistical models incorporating known estimates of biomarker prevalence and prognostic value to prioritize predictive biomarker hypotheses. RESULTS: Our simulation studies demonstrate how biomarker subgroups that are both predictive and prognostic can manifest as time-dependent treatment effects in overall populations. RTOG 9402 provides a representative example where 1p/19q co-deletion and IDH mutation biomarker-specific effects led to time-varying treatment effects and a considerable deviation from proportional hazards in the overall trial population. Finally, using biomarker data from The Cancer Genome Atlas, we were able to generate statistical models that correctly identified and prioritized a commonly used biomarker through retrospective analysis of published clinical trial data. CONCLUSIONS: Biomarkers that are both predictive and prognostic can result in characteristic changes in survival results. Retrospectively analyzing survival data from clinical trials may highlight potential indications for which an underlying predictive biomarker may be found.

Optimizing the removal of strontium and cesium ions from binary solutions on magnetic nano-zeolite using response surface methodology (RSM) and artificial neural network (ANN).

The feasibility of using magnetic nano-zeolite (MNZ) to remove cesium and strontium from their binary corrosive solutions was investigated by considering the multi-variant/multi-objective nature of the process. RSM (Response Surface Methodology) and ANN (Artificial Neural Network) were used to model and optimize the removal system and assess sensitive parameters that can affect the process reliability. MNZ is characterized by its high surface area and cation exchange capacity and possesses good regeneration behavior for both elements using citric acid. Its stability is comparable to other sorbents in acidic media and the stability increases in alkaline media, where dissolution rate follow first order reaction on heterogeneous sites. MNZ removes both contaminants simultaneously with small tendency toward Cs, where MNZ is suggested for application in pre-treatment of highly contaminated alkaline solutions. The percentage removal, decontamination factors, and separation factors have different dependency on the effluent/process conditions; this dependency is the same for both contaminants. Sorption kinetics is initially controlled by external mass transfer through the boundaries then intra-particle diffusion dominates the reactions. The process sensitivity to pH changes is attributed to changes in structural elements -species distribution at the solid/aqueous interface. Cs+ and Sr+2 are exchanged with Na+ and H+, regardless the effluent pH value, and with Al and Fe cations at specific pH. Isosteric heat of sorption calculations indicated that the total heat needed to complete the reaction was considerably reduced by operating the process at optimized temperature.

Prevalence of Clostridium difficile infection and colonization in a tertiary hospital and elderly community of North-Eastern Peninsular Malaysia.

Little is known about Clostridium difficile infection (CDI) in Asia. The aims of our study were to explore (i) the prevalence, risk factors and molecular epidemiology of CDI and colonization in a tertiary academic hospital in North-Eastern Peninsular Malaysia; (ii) the rate of carriage of C. difficile among the elderly in the region; (iii) the awareness level of this infection among the hospital staffs and students. For stool samples collected from hospital inpatients with diarrhea (n = 76) and healthy community members (n = 138), C. difficile antigen and toxins were tested by enzyme immunoassay. Stool samples were subsequently analyzed by culture and molecular detection of toxin genes, and PCR ribotyping of isolates. To examine awareness among hospital staff and students, participants were asked to complete a self-administered questionnaire. For the hospital and community studies, the prevalence of non-toxigenic C. difficile colonization was 16% and 2%, respectively. The prevalence of CDI among hospital inpatients with diarrhea was 13%. Out of 22 C. difficile strains from hospital inpatients, the toxigenic ribotypes 043 and 017 were most common (both 14%). In univariate analysis, C. difficile colonization in hospital inpatients was significantly associated with greater duration of hospitalization and use of penicillin (both P < 0·05). Absence of these factors was a possible reason for low colonization in the community. Only 3% of 154 respondents answered all questions correctly in the awareness survey. C. difficile colonization is prevalent in a Malaysian hospital setting but not in the elderly community with little or no contact with hospitals. Awareness of CDI is alarmingly poor.

Primary Angioplasty for the Treatment of Acute ST Elevated Myocardial Infarction: Single Centre Experience.

Worldwide primary angioplasty is a recommended strategy of reperfusion in patient with acute myocardial infarction as because it ensures reperfusion of the infarct-related vessels more than 90% whereas, with thrombolytics it is only 60-70%. This retrospective observational study includes all patients treated with primary angioplasty at United Hospital from Between March 2007 to January 2011. Total 114 consecutive patients with acute myocardial infarction were treated with primary angioplasty included. Those presented beyond 12 hours of onset of chest pain, in cardiogenic shock, resuscitate and intubated before the procedure were excluded from the study. Majority (89%) of the patient were male, age was minimum 30 years and maximum 90 years, 41.5% were diabetics, 58.4% were hypertensive, 43.5% were dyslipidaemic, 17% were smoker, 29.3% with positive family history. Fifty seven percent patients presented with anterior MI , 42 % with inferior MI and 1% with lateral MI. Left anterior descending (LAD) is the most common vessel involved (57%), followed by Right coronary artery (RCA) 31%, Left circumflex artery (LCX) 8 %, Ramus 1.3% and Graft vessel 2.7%. Our door to balloon time was minimum 23 min, maximum 184 min. We used drug eluting stents for most of the patients, GP IIb- IIIa receptor blockers used in 50% cases and thrombus suction device were used when indicated. We faced complications like arrhythmias in 24%, hypotension in18%, no flow or slow flow in 45%, cardiac arrest in 3% and coronary perforation in 1%. Our overall survival was 97.9%. Primary angioplasty is an emerging area in context of our country. Many of the new centers start this novel strategy which helps to save many lives Primary angioplasty is feasible and safe method of reperfusion in patient with acute myocardial infarction in our center. With the help of our initial experience we can perform PAMI with confidence to those who can afford and who need most.

Spatially resolving valley quantum interference of a donor in silicon.

Electron and nuclear spins of donor ensembles in isotopically pure silicon experience a vacuum-like environment, giving them extraordinary coherence. However, in contrast to a real vacuum, electrons in silicon occupy quantum superpositions of valleys in momentum space. Addressable single-qubit and two-qubit operations in silicon require that qubits are placed near interfaces, modifying the valley degrees of freedom associated with these quantum superpositions and strongly influencing qubit relaxation and exchange processes. Yet to date, spectroscopic measurements have only probed wavefunctions indirectly, preventing direct experimental access to valley population, donor position and environment. Here we directly probe the probability density of single quantum states of individual subsurface donors, in real space and reciprocal space, using scanning tunnelling spectroscopy. We directly observe quantum mechanical valley interference patterns associated with linear superpositions of valleys in the donor ground state. The valley population is found to be within 5% of a bulk donor when 2.85 ± 0.45 nm from the interface, indicating that valley-perturbation-induced enhancement of spin relaxation will be negligible for depths greater than 3 nm. The observed valley interference will render two-qubit exchange gates sensitive to atomic-scale variations in positions of subsurface donors. Moreover, these results will also be of interest for emerging schemes proposing to encode information directly in valley polarization.

Research advisory board members' contributions and expectations in the USA.

This study focuses on the Implementation Community Collaborative Board (I-CCB) to identify members' anticipated contributions to and returns from the I-CCB; examine whether or not members achieved these contributions and returns over time; and explore barriers and facilitators that influenced accomplishments. Longitudinal study with repeated semi-structured in-depth interviews; baseline captured anticipated contributions and returns; 6- and 18-month follow-ups short- and longer-term achievements. We used content analysis to code/reduce text into variables, describe, count and compare categories. Participants anticipated involvement in I-CCB dynamics/governance and in research tasks/procedures. Anticipated returns included social support. Participants exerting influence on I-CCB's research agenda stayed the same over time. Participants conducting research doubled between follow-ups; those writing grant proposals increased by 50%. Participants receiving emotional support remained the same. Challenges: meetings steered by researchers; lack of time; use of jargon. Facilitators: outreaching to community; being affected by HIV; having overlapping identities/roles as researcher, service consumer and/or practitioner. Research partners can maximize facilitators, redress barriers and improve advisory board members' retention. Findings may help optimize the functioning of advisory boards worldwide.

Transcriptional regulation of cuticular protein glycine-rich13 gene expression in wing disc of Bombyx mori, Lepidoptera.

Cuticular protein genes are good models to study the molecular mechanisms of signaling by ecdysteroids, which regulate molting and metamorphosis in insects. The present research demonstrates on hormonal regulation and analysis of the regulatory sequences and transcription factors important for Bombyx mori cuticular protein glycine-rich13 (CPG13) gene expression. Expression of CPG13 was strong at prepupal stage in wing tissues of B. mori. CPG13 expression was induced by the addition of 20E, which was inhibited by cycloheximide in the wing disc. The upstream region of the CPG13 gene was analyzed using a transient reporter assay with a gene gun system and identified two BR-Z2 binding sites to be important cis-acting elements for the transcription activation of the luciferase reporter gene by an ecdysone pulse. Site-directed mutagenesis of these sites, followed by introduction into wing discs, significantly decreased the reporter activity. It was found that the regions carrying the binding sites for the ecdysone-responsive transcription factor BR-Z2 were responsible for the hormonal enhancement of the reporter gene activity in wing discs. Mutation of the BR-Z2 binding sites decreased the reporter activity suggesting that the BR-Z2 isoform can bind to the upstream region of the cuticle protein gene, CPG13 and activates its expression.