Hb Mizuho Case Report; Early Genomic Testing Facilitates a Life Changing Diagnosis.

Unstable variant hemoglobinopathies are an uncommon cause of hemolysis in the pediatric patient and may cause a delay in diagnosis if there is not a high index of suspicion. Hemoglobin (Hb) Mizuho is a rare unstable hemoglobinopathy caused by a pathogenic variant of the HBB gene with a severe phenotype. Here we report on the first known case of Hb Mizuho in Australia, presenting with features of acute and chronic hemolysis. The morphological features on blood film review, in conjunction with biochemical findings and other clinical features, did not immediately suggest an alternative diagnosis and a Next Generation Sequencing gene analysis approach was taken to investigate genes associated with red blood cell disorders and atypical uremic syndrome. The HBB Mizuho variant was detected and established the diagnosis. This report highlights the challenge of diagnosing Hb Mizuho on conventional testing and the need for early genomic testing to clarify a diagnosis.

Evidence-based geriatric knowledge among healthcare providers in Vietnam: adaptation, validation, and pilot of the knowledge about older patients quiz.

BACKGROUND: Vietnam's aging population is growing rapidly, but its health workforce's capacity to provide quality geriatric care is not clearly understood. We aimed to provide a cross-culturally relevant and validated instrument to assess evidence-based geriatric knowledge among healthcare providers in Vietnam. METHODS: We translated the Knowledge about Older Patients Quiz from English to Vietnamese using cross-cultural adaptation methods. We validated the translated version by evaluating its relevance to the Vietnamese context, as well as its semantic and technical equivalence. We fielded the translated instrument on a pilot sample of healthcare providers in Hanoi, Vietnam. RESULTS: The Vietnamese Knowledge about Older Patients Quiz (VKOP-Q) had excellent content validity (S-CVI/Ave) and translation equivalence (TS-CVI/Ave) of 0.94 and 0.92, respectively. The average VKOP-Q score was 54.2% (95% CI: 52.5-55.8) and ranged from 33.3 to 73.3% among 110 healthcare providers in the pilot study. Healthcare providers in the pilot study had low scores on questions related to the physiopathology of geriatric conditions, communication techniques with sensory impaired older adults, and differentiating age related changes from abnormal changes or symptoms. CONCLUSIONS: The VKOP-Q is a validated instrument to assess geriatric knowledge among healthcare providers in Vietnam. The level of geriatric knowledge among healthcare providers in the pilot study was unsatisfactory, which supports the need for further assessment of geriatric knowledge among a nationally representative sample of healthcare providers.

Patient-centered care and geriatric knowledge translation among healthcare providers in Vietnam: translation and validation of the patient-centered care measure.

BACKGROUND: People are living longer, and the majority of aging people reside in low- and middle-income countries (LMICs). However, inappropriate healthcare contributes to health disparities between populations of aging people and leads to care dependency and social isolation. Tools to assess and evaluate the effectiveness of quality improvement interventions for geriatric care in LMICs are limited. The aim of this study was to provide a validated and culturally relevant instrument to assess patient-centered care in Vietnam, where the population of aging people is growing rapidly. METHODS: The Patient-Centered Care (PCC) measure was translated from English to Vietnamese using forward-backward method. The PCC measure grouped activities into sub-domains of holistic, collaborative, and responsive care. A bilingual expert panel rated the cross-cultural relevance and translation equivalence of the instrument. We calculated Content Validity Indexing (CVI) scores at both the item (I-CVI) and scale (S-CVI/Ave) levels to evaluate the relevance of the Vietnamese PCC (VPCC) measure to geriatric care in the Vietnamese context. We piloted the translated instrument VPCC measure with 112 healthcare providers in Hanoi, Vietnam. Multiple logistic regression models were specified to test the a priori null hypothesis that geriatric knowledge is not different among healthcare providers with perception of high implementation compared with low implementation of PCC measures. RESULTS: On the item level, all 20 questions had excellent validity ratings. The VPCC had excellent content validity (S-CVI/Ave of 0.96) and translation equivalence (TS- CVI/Ave of 0.94). In the pilot study, the highest-rated PCC elements were the holistic provision of information and collaborative care, while the lowest-rated elements were the holistic attendance to patients' needs and responsive care. Attention to the psychosocial needs of aging people and poor coordination of care within and beyond the health system were the lowest-rated PCC activities. After controlling for healthcare provider characteristics, the odds of the perception of high implementation of collaborative care were increased by 21% for each increase in geriatric knowledge score. We fail to reject the null hypotheses for holistic care, responsive care and PCC. CONCLUSION: The VPCC is a validated instrument that may be utilized to systemically evaluate the practice of patient-centered geriatric care in Vietnam.

Centering culture in the treatment of opioid use disorder with American Indian and Alaska Native Communities: Contributions from a National Collaborative Board.

American Indian/Alaska Native (AI/AN) communities are disproportionally impacted by the opioid overdose epidemic. There remains a dearth of research evaluating methods for effectively implementing treatments for opioid use disorder (OUD) within these communities. We describe proceedings from a 2-day Collaborative Board (CB) meeting tasked with developing an implementation intervention for AI/AN clinical programs to improve the delivery of medications to treat OUD (MOUD). The CB was comprised of Elders, cultural leaders, providers, individuals with lived experience with OUD, and researchers from over 25 communities, organizations, and academic institutions. Conversations were audio-recorded, transcribed, and coded by two academic researchers with interpretation oversight provided by the CB. These proceedings provided a foundation for ongoing CB work and a frame for developing the program-level implementation intervention using a strength-based and holistic model of OUD recovery and wellbeing. Topics of discussion posed to the CB included engagement and recovery strategies, integration of extended family traditions, and addressing stigma and building trust with providers and clients. Integration of traditional healing practices, ceremonies, and other cultural practices was recommended. The importance of centering AI/AN culture and involving family were highlighted as priorities for the intervention.

Retrospective Cohort Study of Effects of the COVID-19 Pandemic on Tuberculosis Notifications, Vietnam, 2020.

We evaluated the effects of the coronavirus disease pandemic on diagnosis of and treatment for tuberculosis (TB) in Vietnam. We obtained quarterly notifications for TB and multidrug-resistant/rifampin-resistant (MDR/RR) TB from 2015-2020 and evaluated changes in monthly TB case notifications. We used an interrupted time series to assess the change in notifications and treatment outcomes. Overall, TB case notifications were 8% lower in 2020 than in 2019; MDR/RR TB notifications were 1% lower. TB case notifications decreased by 364 (95% CI -1,236 to 508) notifications per quarter and MDR/RR TB by 1 (95% CI -129 to 132) notification per quarter. The proportion of successful TB treatment outcomes decreased by 0.1% per quarter (95% CI -1.1% to 0.8%) in 2020 compared with previous years. Our study suggests that Vietnam was able to maintain its TB response in 2020, despite the pandemic.

Recovery from tachyphylaxis of TRPV1 coincides with recycling to the surface membrane.

The transient receptor potential vanilloid-1 (TRPV1) ion channel is essential for sensation of thermal and chemical pain. TRPV1 activation is accompanied by Ca2+-dependent desensitization; acute desensitization reflects rapid reduction in channel activity during stimulation, whereas tachyphylaxis denotes the diminution in TRPV1 responses to repetitive stimulation. Acute desensitization has been attributed to conformational changes of the TRPV1 channel; however, the mechanisms underlying the establishment of tachyphylaxis remain to be defined. Here, we report that the degree of whole-cell TRPV1 tachyphylaxis is regulated by the strength of inducing stimulation. Using light-sheet microscopy and pH-sensitive sensor pHluorin to follow TRPV1 endocytosis and exocytosis trafficking, we provide real-time information that tachyphylaxis of different degrees concurs with TRPV1 recycling to the plasma membrane in a proportional manner. This process controls TRPV1 surface expression level thereby the whole-cell nociceptive response. We further show that activity-gated TRPV1 trafficking associates with intracellular Ca2+ signals of distinct kinetics, and recruits recycling routes mediated by synaptotagmin 1 and 7, respectively. These results suggest that activity-dependent TRPV1 recycling contributes to the establishment of tachyphylaxis.

Masked face recognition with convolutional neural networks and local binary patterns.

Face recognition is one of the most common biometric authentication methods as its feasibility while convenient use. Recently, the COVID-19 pandemic is dramatically spreading throughout the world, which seriously leads to negative impacts on people's health and economy. Wearing masks in public settings is an effective way to prevent viruses from spreading. However, masked face recognition is a highly challenging task due to the lack of facial feature information. In this paper, we propose a method that takes advantage of the combination of deep learning and Local Binary Pattern (LBP) features to recognize the masked face by utilizing RetinaFace, a joint extra-supervised and self-supervised multi-task learning face detector that can deal with various scales of faces, as a fast yet effective encoder. In addition, we extract local binary pattern features from masked face's eye, forehead and eyebow areas and combine them with features learnt from RetinaFace into a unified framework for recognizing masked faces. In addition, we collected a dataset named COMASK20 from 300 subjects at our institution. In the experiment, we compared our proposed system with several state of the art face recognition methods on the published Essex dataset and our self-collected dataset COMASK20. With the recognition results of 87% f1-score on the COMASK20 dataset and 98% f1-score on the Essex dataset, these demonstrated that our proposed system outperforms Dlib and InsightFace, which has shown the effectiveness and suitability of the proposed method. The COMASK20 dataset is available on https://github.com/tuminguyen/COMASK20 for research purposes.

Multi-task learning neural networks for breath sound detection and classification in pervasive healthcare.

With the emergence of many grave Chronic obstructive pulmonary diseases (COPDs) and the COVID-19 pandemic, there is a need for timely detection of abnormal respiratory sounds, such as deep and heavy breaths. Although numerous efficient pervasive healthcare systems have been proposed for tracking patients, few studies have focused on these breaths. This paper presents a method that supports physicians in monitoring in-hospital and at-home patients by monitoring their breath. The proposed method is based on three deep neural networks in audio analysis: RNNoise for noise suppression, SincNet - Convolutional Neural Network, and Residual Bidirectional Long Short-Term Memory for breath sound analysis at edge devices and centralized servers, respectively. We also developed a pervasive system with two configurations: (i) an edge architecture for in-hospital patients; and (ii) a central architecture for at-home ones. Furthermore, a dataset, named BreathSet, was collected from 27 COPD patients being treated at three hospitals in Vietnam to verify our proposed method. The experimental results demonstrated that our system efficiently detected and classified breath sounds with F1-scores of 90% and 91% for the tiny model version on low-cost edge devices, and 90% and 95% for the full model version on central servers, respectively. The proposed system was successfully implemented at hospitals to help physicians in monitoring respiratory patients in real time.

Bovine extracellular vesicles contaminate human extracellular vesicles produced in cell culture conditioned medium when 'exosome-depleted serum' is utilised.

Extracellular vesicles (EVs) are important intercellular communication messengers. Half of the published studies in the field are in vitro cell culture based in which bovine serum in various concentrations and forms is used to facilitate the production of extracellular vesicles. 'Exosome depleted serum' is the type of bovine serum most widely used in the production of human EVs. Herein, we demonstrate that, despite the initial caution raised in 2014 about the persistence of bovine EVs, 'exosome depleted serum' was still used in 46% of publications on human or rodent EVs between 2015 and 2019. Using nanoparticle tracking analysis combined with detergent lysis of vesicles as well as bovine CD9 ELISA, we show that there were approximately 5.33 x 107/mL of bovine EVs remaining in the 'exosome depleted serum'. Importantly, the 'exosome depleted serum' was relatively enriched in small EVs by approximately 2.7-fold relative to the large EVs compared to that in the original serum. Specifically, the percentage of small EVs in total vesicles had increased from the original 48% in the serum before ultracentrifugation to 92% in the 'exosome depleted serum'. Furthermore, the pervasive bovine EVs carried over by the 'exosome depleted serum', even when the lowest concentration (0.5%) was used in cell culture, resulted in a significant contamination of human EVs in cell culture conditioned medium. Our findings indicate that the use 'exosome depleted serum' in cell culture-based studies may introduce artefacts into research examining the function of human and rodent EVs, in particular those involving EV miRNA. Thus, we appeal to the researchers in the EV field to seriously reconsider the practice of using 'exosome depleted serum' in the production of human and other mammalian EVs in vitro.

Central GABAA receptor mediates taurine-induced hypothermia and possibly reduces food intake in thermo-neutral chicks and regulates plasma metabolites in heat-exposed chicks.

The aim of this study was to examine the central action of taurine on body temperature and food intake in neonatal chicks under control thermoneutral temperature (CT) and high ambient temperature (HT). Intracerebroventricular injection of taurine caused dose-dependent hypothermia and reduced food intake under CT. The mRNA expression of the GABAA receptors, GABAAR-α1 and GABAAR-γ, but not that of GABABR, significantly decreased in the diencephalon after central injection of taurine. Subsequently, we found that picrotoxin, a GABAAR antagonist, attenuated taurine-induced hypothermia. Central taurine significantly decreased the brain concentrations of 3-methoxy-4-hydroxyphenylglycol, a major metabolite of norepinephrine; however, the concentrations of serotonin, dopamine, and the epinephrine metabolites, 3,4-hydroxyindoleacetic acid and homovanillic acid, were unchanged. Although hypothermia was not observed under HT after central injection of taurine, plasma glucose and uric acid levels were higher, and plasma sodium and calcium levels were lower, than those in chicks under CT. In conclusion, brain taurine may play a role in regulating body temperature and food intake in chicks through GABAAR. The changes in plasma metabolites under heat stress suggest that brain taurine may play an important role in maintaining homeostasis in chicks.

Icariin reduces bone loss in a Rankl-induced transgenic medaka (Oryzias latipes) model for osteoporosis.

Given the limitations and side effects of many synthetic drugs, natural products are an important alternative source for drugs and medications for many diseases. Icariin (ICA), one of the main flavonoids from plants of the Epimedium genus, has been shown to ameliorate osteoporosis and improve bone health in preclinical studies. Those studies have used different in vivo models, mostly rodents, but the underlying mechanisms remain unclear. The present study shows, for the first time, that ICA reduces bone damage in a Rankl-induced medaka fish (Oryzias latipes), a non-rodent osteoporosis model. Live imaging was previously performed in this model to characterize antiresorptive and bone-anabolic properties of drugs. Here, a new quantification method (IM ) was established based on the length of mineralized neural arches to quantify levels of bone mineralization damage and protection in early post-embryonic fish. This method was validated by quantification of three levels of bone damage in three independent Rankl fish lines, and by the determination of different degrees of severity of osteoporosis-like phenotypes in one Rankl line exposed to variable Rankl induction schemes. IM was also used to quantify the efficacy of alendronate and etidronate, two common anti-osteoporotic bisphosphonates, and revealed comparable bone protective effects for ICA and alendronate in this fish osteoporosis model. This study's data support the value of the medaka fish model for bone research and establish a method to screen for novel osteoprotective compounds.

Mapping astrocyte activity domains by light sheet imaging and spatio-temporal correlation screening.

Astrocytes are a major type of glial cell in the mammalian brain, essentially regulating neuronal development and function. Quantitative imaging represents an important approach to study astrocytic signaling in neural circuits. Focusing on astrocytic Ca2+ activity, a key pathway implicated in astrocye-neuron interaction, we here report a strategy combining fast light sheet fluorescence microscopy (LSFM) and correlative screening-based time series analysis, to map activity domains in astrocytes in living mammalian nerve tissue. Light sheet of micron-scale thickness enables wide-field optical sectioning to image astrocytes in acute mouse brain slices. Using both chemical and genetically encoded Ca2+ indicators, we demonstrate the complementary advantages of LSFM in mapping Ca2+ domains in astrocyte populations as compared to epifluorescence and two-photon microscopy. Our approach then revealed distinct kinetics of Ca2+ signals between cortical and hypothalamic astrocytes in resting conditions and following the activation of adrenergic G protein coupled receptor (GPCR). This observation highlights the activity heterogeneity across regionally distinct astrocyte populations, and indicates the potential of our method for investigating dynamic signals in astrocytes.

Trends of drowning mortality in Vietnam: evidence from the national injury mortality surveillance system.

OBJECTIVE: To describe the trends of drowning mortality in Vietnam over time and to identify socioeconomic characteristics associated with higher drowning mortality at the provincial level. METHODS: We analysed data from the Ministry of Health injury mortality surveillance system from 1 January 2009 to 31 December 2013. The surveillance covers more than 11 000 commune health centres in all provinces of Vietnam. For provincial population and socioeconomic characteristics, we extracted data from the National census 2009, the Population change and family planning surveys in 2011 and 2013. Multilevel linear models were used to identify provincial characteristics associated with higher mortality rates. RESULTS: Over the 5-year period between 2009 and 2013, 31 232 drowning deaths were reported, equivalent to a 5-year average of 6246 drowning deaths. During this period, drowning mortality rate decreased 7.2/100 000 to 6.9/100 000 (p=0.035). Of six major geographical regions, Northern midland, Central highland and Mekong delta were those with highest mortality rates. In all regions, children aged 1-4 years had the highest mortality rates, followed by those aged 5-9 and 10-14 years. At provincial level, having a coastline was not associated with higher mortality rate. Provinces with larger population size and greater proportion of poor households were statistically significantly associated with higher mortality rates (p=0.042 and 0.006, respectively). CONCLUSION: While some gains have been made in reducing drowning mortality, child deaths due to drowning in Vietnam remain alarmingly high. Targeted scale-up of known effective interventions such as child supervision and basic survival skills are needed for reducing child mortality due to drowning, particularly in socioeconomically disadvantaged provinces.

Enabling factors and barriers to the sustainability and scale-up of drowning reduction interventions in Vietnam: a qualitative assessment.

BACKGROUND: Vietnam has some of the highest rates of drowning deaths in the Western Pacific Region, particularly among children aged 19 years or younger. Several policies aimed at drowning prevention have been developed over the last decade; however, despite policy support, generally these have not been sustained beyond a pilot phase or have been limited to small geographical regions. The present study aims to explore barriers and facilitators for sustainability and scale-up of drowning prevention initiatives in Vietnam, identifying ways forward for future implementation. METHODS: This was a qualitative study using semistructured indepth interviews with key stakeholders (n=12) engaged in drowning prevention in Vietnam. The Framework Method was used to analyse the data drawing on Schell's theoretical framework for public health programme sustainability. The Framework Method is most commonly used for the thematic analysis of semistructured interview transcripts, particularly as the data were fairly homogeneous. RESULTS: Four key factors were identified that facilitated implementation of drowning reduction activities in Vietnam. Strong political support at all levels, underpinned by policy; effective partnerships with the community; widespread communication; and programme adaptation to local contexts and application of innovative approaches, for example, strengthening organisational capacity in limited resource settings, were the key enablers. Barriers include the instability of the funding sources; inadequate programme evaluations to generate evidence of effectiveness and lack of consistent and timely data collection; and insufficient strategic planning for long-term implementation of drowning prevention interventions. CONCLUSION: Ensuring the sustainability and scale-up of drowning prevention programmes in Vietnam requires a continued focus on enablers such as on community engagement, communication activities and partnership approaches, and importantly concerted efforts to mobilise resources for continued long-term funding, improvements in planning and intersectoral coordination, and ensuring that future programmes are robustly evaluated for effectiveness.

Induction of Antitumor Immunity by Exosomes Isolated from Cryopreserved Cord Blood Monocyte-Derived Dendritic Cells.

(1) Background: Dendritic cell (DC) vaccination has shown outstanding achievements in cancer treatment, although it still has some adverse side effects. Vaccination with DC-derived exosomes has been thought to overcome the side effects of the parental DCs. (2) Method: We performed the experiments to check the ability of cryopreserved umbilical cord blood mononuclear cell-derived DCs (cryo CBMDCs) and their exosomes to prime allogeneic T cell proliferation and allogeneic peripheral blood mononuclear cell (alloPBMCs) cytotoxicity against A549 lung cancer cells. (3) Results: We found that both lung tumor cell lysate-pulsed DCs and their exosomes could induce allogeneic T cell proliferation. Moreover, alloPBMCs primed with tumor cell lysate-pulsed DCs and their exosomes have a greater cytotoxic activity against A549 cells compared to unprimed cells and cells primed with unpulsed DCs and their exosomes. (4) Conclusion: Tumor cell lysate-pulsed DCs and their exosomes should be considered to develop into a novel immunotherapeutic strategy-e.g., vaccines-for patients with lung cancer. Our results also suggested that cryo umbilical cord blood mononuclear cells source, which is a readily and available source, is effective for generation of allogeneic DCs and their exosomes will be material for vaccinating against cancer.

Removing Hydrogen Sulfide Contamination in Biogas Produced from Animal Wastes.

Hydrogen sulfide (HS) contamination in biogas produced from animal wastes limits its use to cooking and precludes it from being used for heating, lighting, or electricity generation. This limitation results in the release to the atmosphere of between 3 and 51% of total biogas produced. Biogas contains 50 to 70% methane (CH), a potent greenhouse gas that contributes to global warming. This study aimed to develop a cost-effective HS filtering system using local materials rich in iron as iron oxide (FeO), which reacts readily with HS and forms adsorbed iron sulfide (FeS) when gas is passed through it. Here we tested the performance of seven New Zealand soils and sand, each at five different gas flow rates (59, 74, 94, 129, and 189 mL min). We found that three materials (allophanic soil, brown soil, and black sand) had stable HS removal efficiencies close to 100% at all gas flow rates, followed by typic sand (89-99%), raw sand (76-99%), acidic sand (48-89%), and podzol soil (58-87%). These results show that inexpensive and simple filters to remove HS from biogas can be made using local soils. Used soil in the filters can then be easily regenerated by exposure to the atmosphere and reused to achieve sustained HS removal efficiency.

An acute increase in water temperature can increase free amino acid concentrations in the blood, brain, liver, and muscle in goldfish (Carassius auratus).

Water temperature directly affects the body temperature in fish, so increasing water temperatures in oceans and rivers will lead to increases in fish body temperatures. Whilst a range of responses of fish to increases in water temperature have been measured, amino acid metabolism in a fish under high water temperature (HT) conditions has not been investigated. The aim of this study was to determine the effects of an acute increase in water temperature on oxygen consumption, plasma cortisol concentrations, and free amino acid concentrations in plasma and several tissues in goldfish (Carassius auratus). Oxygen consumption and plasma cortisol concentrations were increased in goldfish exposed to HT (30 ± 1 °C) for 200 min compared with goldfish at a control water temperature (CT 17 ± 1 °C). Oxygen consumption and plasma cortisol concentrations in both groups of fish combined were positively correlated. When goldfish were exposed to HT for 300 min oxygen consumption and plasma concentrations of 15 free amino acids were increased compared with goldish at CT. Concentrations of several free amino acids were increased to varying extents in the brain, liver, and muscle tissues. In conclusion, an acute increase in water temperature affected amino acid metabolism differently in the brain, liver, and muscle tissues. Goldfish will be a useful species for further studies of the possible roles of various amino acids in the brain, muscle, and liver during acute increases in water temperature in fish.

Health-related quality of life and recovery patterns among hospitalised injury patients in Vietnam.

PURPOSE: To measure post-injury health-related quality of life (HRQoL) among hospitalised injury patients following discharge and to identify factor associated with lower HRQoL over time. METHOD: A prospective cohort study was conducted from January 2010 to October 2011 in Thai Binh province, Vietnam. Participants were 18 years or older, hospitalised for at least 1 day and normally residing in Thai Binh province. Data on demographic and injury characteristics were collected during their hospital stay. Data on HRQoL, measured by the Health Utilities Index mark 3 (HUI3), were collected in the participants' home at 1, 2, 4 and 12 months after their hospital discharge. Generalised estimating equation model was selected to examine the association of demographic and injury characteristics with HRQoL which were repeatedly measured over time. RESULTS: HUI3 multi-attribute score was lowest at the first follow-up (0.52 for males, 0.28 for females). The scores significantly improved over time (increment of 0.08 or greater, p values < 0.001). Ambulation and Pain were single-attributes with scores significantly increased over time (increment of 0.05 or greater, p values < 0.001). Factors identified to be associated with lower HUI3 scores were older age (p values < 0.02 or less), more severe injury (p values = 0.05 or less) and having other illnesses (p values < 0.01). CONCLUSION: The study provides important, and previously unreported, estimates of post-injury HRQoL in Vietnam. Even for those having the lowest levels of severity, the loss in HRQoL is significant, highlighting the importance of efforts on injury prevention and rehabilitation services.

Alcohol Warning Label Awareness and Attention: A Multi-method Study.

AIMS: Evaluation of alcohol warning labels requires careful consideration ensuring that research captures more than awareness given that labels may not be prominent enough to attract attention. This study investigates attention of current in market alcohol warning labels and examines whether attention can be enhanced through theoretically informed design. Attention scores obtained through self-report methods are compared to objective measures (eye-tracking). METHODS: A multi-method experimental design was used delivering four conditions, namely control, colour, size and colour and size. The first study (n = 559) involved a self-report survey to measure attention. The second study (n = 87) utilized eye-tracking to measure fixation count and duration and time to first fixation. Analysis of Variance (ANOVA) was utilized. RESULTS: Eye-tracking identified that 60% of participants looked at the current in market alcohol warning label while 81% looked at the optimized design (larger and red). In line with observed attention self-reported attention increased for the optimized design. CONCLUSIONS: The current study casts doubt on dominant practices (largely self-report), which have been used to evaluate alcohol warning labels. Awareness cannot be used to assess warning label effectiveness in isolation in cases where attention does not occur 100% of the time. Mixed methods permit objective data collection methodologies to be triangulated with surveys to assess warning label effectiveness. SHORT SUMMARY: Attention should be incorporated as a measure in warning label effectiveness evaluations. Colour and size changes to the existing Australian warning labels aided by theoretically informed design increased attention.