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A comparison of super-resolution microscopy techniques for imaging tightly packed microcolonies of an obligate intracellular bacterium.
Conventional optical microscopy imaging of obligate intracellular bacteria is hampered by the small size of bacterial cells, tight clustering exhibited by some bacterial species and challenges relating to labelling such as background from host cells, a lack of validated reagents, and a lack of tools for genetic manipulation. In this study, we imaged intracellular bacteria from the species Orientia tsutsugamushi (Ot) using five different fluorescence microscopy techniques: standard confocal, Airyscan confocal, instant Structured Illumination Microscopy (iSIM), three-dimensional Structured Illumination Microscopy (3D-SIM) and Stimulated Emission Depletion Microscopy (STED). We compared the ability of each to resolve bacterial cells in intracellular clumps in the lateral (xy) axis, using full width half-maximum (FWHM) measurements of a labelled outer membrane protein (ScaA) and the ability to detect small, outer membrane vesicles external to the cells. Comparing the techniques readily available to us (above), 3D-SIM microscopy, in combination with the shortest-wavelength dyes, was found overall to give the best lateral resolution. We next compared the ability of each technique to sufficiently resolve bacteria in the axial (z) direction and found 3D-STED to be the most successful method for this. We then combined this 3D-STED approach with a custom 3D cell segmentation and analysis pipeline using the open-source, deep learning software, Cellpose to segment the cells and subsequently the commercial software Imaris to analyse their 3D shape and size. Using this combination, we demonstrated differences in bacterial shape, but not their size, when grown in different mammalian cell lines. Overall, we compare the advantages and disadvantages of different super-resolution microscopy techniques for imaging this cytoplasmic obligate intracellular bacterium based on the specific research question being addressed.
Addressing the gap in health data management skills: an online self-guided course for researchers and health professionals.
BackgroundThe healthcare sector is rapidly evolving with the rise of digital technology and data-driven decision-making. However, traditional medical education has yet to fully integrate training on managing health-related information, resulting in a significant skills gap among medical and research professionals. This gap is pronounced in low- and middle-income countries (LMICs), where data privacy concerns and inadequate infrastructure hinder efforts to utilise and share health data.AimsTo address this gap, we developed an online, modular course aimed at providing foundational skills on capturing, storing and sharing health data.MethodsThe course was developed using the ADDIE(Analyze, Design, Develop, Implement, Evaluate) instructional design model. A needs assessment workshop involving 25 global health proffesionals identified key training gaps which informed the curriculum's development. A multidisciplinary team from six institutions developed the modules. The course was piloted in a face-to-face setting with 37 participants and later adapted for online delivery via the Global Health Network platform. We evaluated the course using Level 1 of Kirkpatrick's model for training evaluation.ResultsSix foundational modules were developed: Introduction to Data Management, Data Quality, Data Repositories, Ethics of Data Sharing, Data Governance, and Costing for Data Management. Between December 2020 and April 2024, 6,384 individuals from 90 countries completed the course. Of these, 32% were from Africa, 15% from Asia, 16% from South/Central America and the Caribbean, and 24% from Europe. Summative evaluations, based on voluntary post-module surveys, demonstrated high relevance to participants' learning needs (96.6%) and strong intentions to apply the skills gained (88.3%). Key motivators for enrollment included the course's free access, relevance to professional or academic needs, and trust in the organizations and authors behind the content.ConclusionsThe high enrolment and broad geographical reach demonstrates the potential of online training as a cost-effective tool to equip health practitioners and researchers with data literacy skills. Future evaluations will assess its impact on participants' knowledge, behavior, and data-sharing and reuse practices.
Asparagine availability controls germinal center B cell homeostasis.
The rapid proliferation of germinal center (GC) B cells requires metabolic reprogramming to meet energy demands, yet these metabolic processes are poorly understood. By integrating metabolomic and transcriptomic profiling of GC B cells, we identified that asparagine (Asn) metabolism was highly up-regulated and essential for B cell function. Asparagine synthetase (ASNS) was up-regulated after B cell activation through the integrated stress response sensor GCN2. Conditional deletion of Asns in B cells impaired survival and proliferation in low Asn conditions. Removal of environmental Asn by asparaginase or dietary restriction compromised the GC reaction, impairing affinity maturation and the humoral response to influenza infection. Furthermore, metabolic adaptation to the absence of Asn required ASNS, and oxidative phosphorylation, mitochondrial homeostasis, and synthesis of nucleotides were particularly sensitive to Asn deprivation. These findings demonstrate that Asn metabolism acts as a key regulator of B cell function and GC homeostasis.
Rethinking village malaria workers in Cambodia: Perspectives from the communities, programme managers, and international stakeholders.
Since the early 2000s, malaria cases in Cambodia have declined steadily. Village malaria workers (VMWs) have played a critical role in reducing malaria transmission and progress towards malaria elimination. To prevent malaria re-establishment, however, implementation strategies need to consider carefully the changing healthcare needs in the communities as well as challenges to, and opportunities for, programme adaptation. We conducted in-depth interviews with a diverse range of stakeholders involved in the planning or implementation of the VMW programme in Cambodia, to elicit their views and experiences about health priorities in the communities, the public health value of VMWs and community-based care, and prospects for future programme development. Respondents included managers and implementers involved in the VMW programme at the central and provincial level (n = 9), technical officers at international agencies in Cambodia (n = 7), international stakeholders in non-governmental and research organisations based in Cambodia or other countries in the region (n = 5), as well as VMWs (n = 10), and community members (n = 16) in six endemic communes of Kravanh District, Pursat Province. In Kravanh, we also conducted four focus group discussions with 19 community members who had previous experience of malaria. The qualitative dataset was analysed using a thematic approach. VMWs, particularly mobile malaria workers tasked with active case detection among forest workers, were deemed necessary to maintain effective malaria control. However, there was a clear demand in the communities for additional services including treatment for common illnesses, monitoring of blood pressure and blood sugar levels, and relief of general symptoms through medication, such as for fever, headache, and stomach pain. Programme managers and international stakeholders agreed that the VMW programme needs a rethinking of the current implementation model to ensure continued uptake, relevance, and motivation of VMWs. Suggestions for add-on activities included adoption of new tests for febrile illnesses such as dengue and chikungunya, and screening for the prevention and monitoring of non-communicable diseases. There was emphasis on the needs for more sustainable financing mechanisms and integration with the existing community health infrastructure. The potential expansion of VMW services will benefit from the continued involvement of external donors and partners for technical and financial support. However, the implementation strategy should consider since the outset opportunities for enhanced local ownership and health system integration. To maintain domestic political momentum and access new potential sources of domestic funding, further programme development should align with national health priorities and the ongoing process of administrative decentralisation, while being responsive to changing public health needs within the communities.
Triage processes in primary, secondary, and tertiary health care facilities in the Kathmandu Valley, Nepal: a mixed-methods study.
BackgroundIn healthcare facilities, an efficient triage system is critical to optimize patient care. The main objective of this study was to explore the triage processes and practices in three different tiers of healthcare facilities in the Kathmandu Valley, Nepal.MethodsA mixed-methods approach in this study comprised observations and interviews in ten healthcare settings across primary care centers (PHC; n = 6), secondary care centers (SHC; n = 3), and tertiary care hospital (n = 1). Data were collected from June to November 2023. Semi-structured interviews were conducted among patients (n = 30) including survey questionnaires among 144 healthcare workers (HCWs) focused on triage. The qualitative data were analyzed using Interpretative Phenomenological Analysis and quantitative data were analyzed to explore the median score on the consistent practice of triage based on the Likert scale.ResultsPHCs had designated space for triage with less equipped emergency services and outpatient departments (OPDs) and received severely ill patients rarely. Although prioritizing critical patients and prompt care was part of the services, there was a lack of triage protocols with more than half of the HCWs (56.3%; 36/64) from the tertiary hospital reporting the availability of triage guidelines compared to SHCs (28.1%; 9/32) and PHCs (6.3%; 3/48). The majority of HCWs from the tertiary level recognized triage's effectiveness in reducing time lag, and prioritizing patients. Tertiary level had the consistent use of triage (94%; 60/64) compared to only around two-thirds in SHCs (66%; 19/29) and PHCs (62%; 28/45). Patients often attended PHC services for routine check-ups and were motivated by health insurance, affordability, free medicines, referral cards, and proximity. In the SHC, there was a well-equipped emergency department (ED) with specific guidelines, but its use was infrequent. Patients were unaware of the triage process and its utility. In all settings, while most HCWs had a basic knowledge of triage, some were not confident due to limited exposure to the triage process and training. Many HCWs reported the need for triage-related training and its' consistent implementation.ConclusionsConsistent utilization of triage protocols, coupled with improved infrastructure, resource allocation, and training for healthcare workers is critical for the optimization of triage processes in healthcare settings in the Kathmandu Valley, Nepal.
Modeling the within-host dynamics of Plasmodium vivax hypnozoite activation: An analysis of the SPf66 vaccine trial.
Plasmodium vivax parasites can lie dormant in the liver as hypnozoites, activating weeks to months after sporozoite inoculation to cause relapsing malarial illness. It is not known what biological processes govern hypnozoite activation. We use longitudinal data from the most detailed cohort study ever conducted in an area where both Plasmodium falciparum and P. vivax were endemic to fit a simple within-host mathematical model of P. vivax hypnozoite activation. 1,344 children living on the Thailand-Myanmar border were followed daily for 21 mo. There were 2,504 vivax and 1,164 falciparum malaria symptomatic episodes recorded over 1988 person-years. The model assumes that hypnozoites activate independently at a constant rate ("exponential clock model"). When this model was embedded in a stochastic framework for repeated infectious mosquito bites, with seasonality inferred from the incidence of clinical falciparum malaria episodes, it explained the observed temporal patterns of multiple (up to 13) recurrent vivax malaria episodes. Under this model, we estimate the mean dormancy period for a single hypnozoite to be 6 mo (i.e., a half-life of 4 mo). We use the calibrated within-host model to characterize population-level overdispersion in the risk of relapse, and assess the potential utility of a serological test for radical cure in low transmission settings. We show that mefloquine treatment of falciparum malaria eliminates early vivax relapses; and that there are substantially more P. vivax recurrences than expected under the model following artesunate monotherapy treatment for falciparum malaria. These results suggest that hypnozoites can be activated by symptomatic malarial illness.
ACORN (A Clinically-Oriented Antimicrobial Resistance Surveillance Network) II: protocol for case based antimicrobial resistance surveillance
Background: Antimicrobial resistance surveillance is essential for empiric antibiotic prescribing, infection prevention and control policies and to drive novel antibiotic discovery. However, most existing surveillance systems are isolate-based without supporting patient-based clinical data, and not widely implemented especially in low- and middle-income countries (LMICs). Methods: A Clinically-Oriented Antimicrobial Resistance Surveillance Network (ACORN) II is a large-scale multicentre protocol which builds on the WHO Global Antimicrobial Resistance and Use Surveillance System to estimate syndromic and pathogen outcomes along with associated health economic costs. ACORN-healthcare associated infection (ACORN-HAI) is an extension study which focuses on healthcare-associated bloodstream infections and ventilator-associated pneumonia. Our main aim is to implement an efficient clinically-oriented antimicrobial resistance surveillance system, which can be incorporated as part of routine workflow in hospitals in LMICs. These surveillance systems include hospitalised patients of any age with clinically compatible acute community-acquired or healthcare-associated bacterial infection syndromes, and who were prescribed parenteral antibiotics. Diagnostic stewardship activities will be implemented to optimise microbiology culture specimen collection practices. Basic patient characteristics, clinician diagnosis, empiric treatment, infection severity and risk factors for HAI are recorded on enrolment and during 28-day follow-up. An R Shiny application can be used offline and online for merging clinical and microbiology data, and generating collated reports to inform local antibiotic stewardship and infection control policies. Discussion: ACORN II is a comprehensive antimicrobial resistance surveillance activity which advocates pragmatic implementation and prioritises improving local diagnostic and antibiotic prescribing practices through patient-centred data collection. These data can be rapidly communicated to local physicians and infection prevention and control teams. Relative ease of data collection promotes sustainability and maximises participation and scalability. With ACORN-HAI as an example, ACORN II has the capacity to accommodate extensions to investigate further specific questions of interest.
Case Report: Soft tissue infection with Burkholderia thailandensis capsular variant: case report from the Lao PDR
Background Burkholderia thailandensis is an environmental bacteria closely related to Burkholderia pseudomallei that rarely causes infection in humans. Some environmental isolates have shown to express a capsular polysaccharide known as B. thailandensis capsular variant (BTCV), but human infection has not previously been reported. Although B. thailandenisis has been identified in environmental samples in Laos before, there have not been any human cases reported. Case A 44-year-old man presented to a district hospital in Laos with a short history of fever and pain in his left foot. Physical examination identified a deep soft-tissue abscess in his left foot and an elevated white blood count. A deep pus sample was taken and melioidosis was suspected from preliminary laboratory tests. The patient was initially started on cloxacillin, ceftriaxone and metronidazole, and was then changed to ceftazidime treatment following local melioidosis treatment guidelines. Laboratory methods A deep pus sample was sent to Mahosot Hospital microbiology laboratory where a mixed infection was identified including Burkholderia sp. Conventional identification tests and API 20NE were inconclusive, and the B. pseudomallei-specific latex agglutination was positive. The isolate then underwent a Burkholderia species specific PCR which identified the isolate as B. thailandensis. The isolate was sent for sequencing on the Illumina NovaSeq 6000 system and multi-locus sequence typing analysis identified the isolate had the same sequence type (ST696) as B. thailandensis E555, a strain which expresses a B. pseudomallei-like capsular polysaccharide. Conclusion This is the first report of human infection with B. thailandensis in Laos, and the first report of any human infection with the B. thailandensis capsular variant. Due to the potential for laboratory tests to incorrectly identify this bacteria, staff in endemic areas for B. thailandensis and B. pseudomallei should be aware and ensure that appropriate confirmatory methods are used to differentiate between the species.
Case Report: Soft tissue infection with Burkholderia thailandensis capsular variant: case report from the Lao PDR
Background Burkholderia thailandensis is an environmental bacteria closely related to Burkholderia pseudomallei that rarely causes infection in humans. Some environmental isolates have shown to express a capsular polysaccharide known as B. thailandensis capsular variant (BTCV), but human infection has not previously been reported. Although B. thailandednisis has been identified in environmental samples in Laos before, there have not been any human cases reported. Case A 44-year-old man presented to a district hospital in Laos with a short history of fever and pain in his left foot. Physical examination identified a deep soft-tissue abscess in his left foot and an elevated white blood count. A deep pus sample was taken and melioidosis was suspected from preliminary laboratory tests. The patient was initially started on cloxacillin, ceftriaxone and metronidazole, and was then changed to ceftazidime treatment following local melioidosis treatment guidelines. Laboratory methods A deep pus sample was sent to Mahosot Hospital microbiology laboratory where a mixed infection was identified including Burkholderia sp. Conventional identification tests and API 20NE were inconclusive, and the B. pseudomallei-specific latex agglutination was positive. The isolate then underwent a Burkholderia species specific PCR which identified the isolate as B. thailandensis. The isolate was sent for sequencing on the Illumina NovaSeq 6000 system and multi-locus sequence typing analysis identified the isolate had the same sequence type (ST696) as B. thailandensis E555, a strain which expresses a B. pseudomallei-like capsular polysaccharide. Conclusion This is the first report of human infection with B. thailandensis in Laos, and the first report of any human infection with the B. thailandensis capsular variant. Due to the potential for laboratory tests to incorrectly identify this bacteria, staff in endemic areas for B. thailandensis and B. pseudomallei should be aware and ensure that appropriate confirmatory methods are used to differentiate between the species.
A Clinically Oriented antimicrobial Resistance surveillance Network (ACORN): pilot implementation in three countries in Southeast Asia, 2019-2020
Background: Case-based surveillance of antimicrobial resistance (AMR) provides more actionable data than isolate- or sample-based surveillance. We developed A Clinically Oriented antimicrobial Resistance surveillance Network (ACORN) as a lightweight but comprehensive platform, in which we combine clinical data collection with diagnostic stewardship, microbiological data collection and visualisation of the linked clinical-microbiology dataset. Data are compatible with WHO GLASS surveillance and can be stratified by syndrome and other metadata. Summary metrics can be visualised and fed back directly for clinical decision-making and to inform local treatment guidelines and national policy. Methods: An ACORN pilot was implemented in three hospitals in Southeast Asia (1 paediatric, 2 general) to collect clinical and microbiological data from patients with community- or hospital-acquired pneumonia, sepsis, or meningitis. The implementation package included tools to capture site and laboratory capacity information, guidelines on diagnostic stewardship, and a web-based data visualisation and analysis platform. Results: Between December 2019 and October 2020, 2294 patients were enrolled with 2464 discrete infection episodes (1786 community-acquired, 518 healthcare-associated and 160 hospital-acquired). Overall, 28-day mortality was 8.7%. Third generation cephalosporin resistance was identified in 54.2% (39/72) of E. coli and 38.7% (12/31) of K. pneumoniae isolates. Almost a quarter of S. aureus isolates were methicillin resistant (23.0%, 14/61). 290/2464 episodes could be linked to a pathogen, highlighting the level of enrolment required to achieve an acceptable volume of isolate data. However, the combination with clinical metadata allowed for more nuanced interpretation and immediate feedback of results. Conclusions: ACORN was technically feasible to implement and acceptable at site level. With minor changes from lessons learned during the pilot ACORN is now being scaled up and implemented in 15 hospitals in 9 low- and middle-income countries to generate sufficient case-based data to determine incidence, outcomes, and susceptibility of target pathogens among patients with infectious syndromes.
A hill tribe community advisory board in Northern Thailand: lessons learned one year on.
Northern Thailand and its neighbouring regions are home to several minority ethnic groups known as hill tribes, each with their own language and customs. Hill tribe communities live mostly in remote agricultural communities, face barriers in accessing health, and have a lower socio-economic status compared to the main Thai ethnic group. Due to their increased risk of infectious diseases, they are often participants in our research projects.To make sure our work is in line with the interests of hill tribe communities and respects their beliefs and customs, we set up a hill tribe community advisory board. We consult the members before, during, and after our projects are carried out. This manuscript recounts how we set up the community advisory board and our reflections following one year of activities. Our experience strongly supports engaging with community advisory boards when working with minority ethnic groups in lower and middle-income settings. In particular, we found that over time, as researchers and members familiarise with one another and their respective environments, exchanges gain meaning and benefits increase, stressing the advantages of long-term collaborations over short or project-based ones.
Comparison of WHO versus national COVID-19 therapeutic guidelines across the world: not exactly a perfect match
BackgroundThe COVID-19 pandemic affected all WHO member states. We compared and contrasted the COVID-19 treatment guidelines of each member state with the WHO COVID-19 therapeutic guidelines.MethodsMinistries of Health or accessed National Infectious Disease websites and other relevant bodies and experts were contacted to obtain national guidelines (NGs) for COVID-19 treatment. NGs were included only if they delineated specific pharmacological treatments for COVID-19, which were stratified by disease severity. We conducted a retrospective review using the adapted Reporting Checklist for Public Versions of Guidelines (RIGHT-PVG) survey checklist and a derived comparative metric based on the WHO guidelines was performed.ResultsCOVID-19 therapeutics NGs could be obtained from 109 of the 194 WHO member states. There was considerable variation in guidelines and in disease severity stratifications. Therapeutic recommendations in many NGs differed substantially from the WHO guidelines. Overall in late 2022, 93% of NGs were recommending at least one treatment which had proved to be ineffective in large randomised trials, and was not recommended by WHO. Corticosteroids were not recommended in severe disease in nearly 10% of NGs despite overwhelming evidence of their benefit. NGs from countries with low-resource settings showed the greatest divergence when stratified by gross domestic product per year, Human Development Index and the Global Health Security Index.DiscussionOur study is limited to NGs that were readily accessible, and it does not reflect the availability of recommended medicines in the field. Three years after the start of the SARS-CoV-2 pandemic, available COVID-19 NGs vary substantially in their therapeutic recommendations, often differ from the WHO guidelines, and commonly recommend ineffective, unaffordable or unavailable medicines.
Greater preservation of SARS-CoV-2 neutralising antibody responses following the ChAdOx1-S (AZD1222) vaccine compared with mRNA vaccines in haematopoietic cell transplant recipients.
Whilst SARS-CoV-2 mRNA vaccines generate high neutralising antibodies (nAb) in most individuals, haematopoietic stem cell transplant (HSCT) and chimeric antigen receptor T-cell (CAR-T) recipients respond poorly. HSCT/CAR-T treatment ablates existing immune memory, with recipients requiring revaccination analogous to being vaccine naive. An optimal revaccination strategy for this cohort has not been defined. Factors predicting immunogenicity following three ancestral SARS-CoV-2 vaccines were assessed in 198 HSCT/CAR-T recipients and 96 healthcare workers (HCWs) recruited to multicentre studies. Only 25% of HSCT/CAR-T recipients generated nAbs following one dose, with titres 167-fold and 7-fold lower than that in HCWs after the first and second doses, respectively. Lower post-second dose nAb titres were associated with older age, rituximab use, and previous HSCT. ChAdOx1-S recipients were more likely to generate nAbs compared with mRNA vaccines, with titres comparable to HCWs. In contrast, nAbs were significantly lower in HSCT/CAR-T recipients than HCWs after mRNA vaccination. The poor first-dose immunogenicity in HSCT/CAR-T recipients suggests a minimum licensed dosing interval could limit the period of vulnerability following HSCT/CAR-T. The relative preservation of nAbs with ChAdOx1-S vaccination highlights the importance of evaluating alternative platforms to mRNA vaccination within this highly vulnerable clinical cohort.
Outbreaks of COVID-19 in a tuberculosis treatment sanatorium on the Thailand-Myanmar border: a retrospective cohort analysis.
BackgroundTuberculosis (TB) is a chronic condition, with overlapping symptoms to those of coronavirus disease 2019 (COVID-19). There has been inconsistent evidence on whether TB is a predisposing factor for developing severe COVID-19. The aim of this report is to explore whether TB influences the severity of COVID-19.MethodsCOVID-19 cases at two TB sanatoria on the Thailand-Myanmar border were reviewed. Demographic, clinical and laboratory data including TB treatment and co-morbidities, were analyzed. Characteristics and COVID-19 clinical outcomes were compared between two groups of patients: TB and those without TB (the caretakers and the medical personnel). Multivariable ordered logistic regression was conducted to compare the risk of severe COVID-19 between the two groups.ResultsBetween September 2021 and March 2022, 161 COVID-19 cases were diagnosed. Over half of the COVID-19 patients were infected with TB (n= 104, 64.6%), and the rest were not (n=57, 35.4%). The median (interquartile range) age was 48 (33.5-57.0) and 27 (23-33) years in the TB and in the non-TB COVID-19 patients, respectively. Before COVID-19 infection, 67.1% (106/158) of patients had received at least one dose of COVID-19 vaccine. The median cycle threshold value at diagnosis was not different between TB (18.5, IQR 16.1-32.3) and non-TB patients (18.8, 15.1-30.0). Fever, gastrointestinal symptoms and ageusia were more common in non-TB patients. Six patients (3.8%, 6/156) all from the TB group became severe of which five (3.2%, 5/156) required oxygen therapy. One TB patient died (1/104, 0.96%) of lung cancer. After adjustment for potential confounders, the final clinical severity was not different between the two groups (adjusted odds ratio 1.40, 95% confidence interval 0.16-12.39).ConclusionsTB was not associated with severe outcomes in the two TB sanatoria. The high uptake of COVID-19 vaccination and active screening could have impacted on disease progression and prevented unfavorable outcomes.
Informed consent and risk communication challenges in antimicrobial clinical trials: a scoping review.
OBJECTIVES: Randomised trials for the management of drug-resistant infections are challenging to conduct as target patient populations often lack decision-making capacity, and enrolment windows are typically short. Improving informed consent and risk communication in these trials is especially crucial for protecting patient interests and maximising trial efficiency. This study aimed to understand challenges in risk communication and informed consent in antimicrobial clinical trials. DESIGN: Scoping review. DATA SOURCES: Searches were conducted in Embase, Medline, CINAHL and Web of Science Core for peer-reviewed English articles that were published from January 2000 to April 2023. ELIGIBILITY CRITERIA: Included articles were empirical studies or expert opinions that sought experts', patients' or representatives' opinions on informed consent in the context of clinical trials involving antibiotic/anti-infective agents. DATA EXTRACTION AND SYNTHESIS: Abstract screening, full-text review, data extraction and evidence rating were performed by two independent reviewers. Extracted data were summarised and reported qualitatively based on common themes. A total of 2330 records were retrieved, and 29 articles were included in the review. RESULTS: Half of the articles involving medical experts and one-third involving patients and representatives reported that full comprehension by patients and representatives was challenging or not achievable. Healthcare providers and consent takers were crucial for the quality of informed consent. The level of trust consent givers placed on healthcare providers had a critical influence on the consent rate. Emotional distress was pervasive among patients/representatives. CONCLUSION: The findings indicate that strengthening consent takers' communication skills in providing emotional support to patients and their representatives may improve informed consent. More research is needed to understand informed consent in low-income and middle-income and non-English-speaking countries.
Methaemoglobin as a surrogate marker of primaquine antihypnozoite activity in Plasmodium vivax malaria: A systematic review and individual patient data meta-analysis
Background The 8-aminoquinolines, primaquine and tafenoquine, are the only available drugs for the radical cure of Plasmodium vivax hypnozoites. Previous evidence suggests that there is dose-dependent 8-aminoquinoline induced methaemoglobinaemia and that higher methaemoglobin concentrations are associated with a lower risk of P. vivax recurrence. We undertook a systematic review and individual patient data meta-analysis to examine the utility of methaemoglobin as a population-level surrogate endpoint for 8-aminoquinoline antihypnozoite activity to prevent P. vivax recurrence. Methods and findings We conducted a systematic search of Medline, Embase, Web of Science, and the Cochrane Library, from 1 January 2000 to 29 September 2022, inclusive, of prospective clinical efficacy studies of acute, uncomplicated P. vivax malaria mono-infections treated with radical curative doses of primaquine. The day 7 methaemoglobin concentration was the primary surrogate outcome of interest. The primary clinical outcome was the time to first P. vivax recurrence between day 7 and day 120 after enrolment. We used multivariable Cox proportional-hazards regression with site random-effects to characterise the time to first recurrence as a function of the day 7 methaemoglobin percentage (log base 2 transformed), adjusted for the partner schizonticidal drug, the primaquine regimen duration as a proxy for the total primaquine dose (mg base/kg), the daily primaquine dose (mg/kg), and other factors. The systematic review protocol was registered with PROSPERO (CRD42023345956). We identified 219 P. vivax efficacy studies, of which 8 provided relevant individual-level data from patients treated with primaquine; all were randomised, parallel arm clinical trials assessed as having low or moderate risk of bias. In the primary analysis data set, there were 1,747 patients with normal glucose-6-phosphate dehydrogenase (G6PD) activity enrolled from 24 study sites across 8 different countries (Indonesia, Brazil, Vietnam, Thailand, Peru, Colombia, Ethiopia, and India). We observed an increasing dose-response relationship between the daily weight-adjusted primaquine dose and day 7 methaemoglobin level. For a given primaquine dose regimen, an observed doubling in day 7 methaemoglobin percentage was associated with an estimated 30% reduction in the risk of P. vivax recurrence (adjusted hazard ratio = 0.70; 95% confidence interval [CI] [0.57, 0.86]; p = 0.0005). These pooled estimates were largely consistent across the study sites. Using day 7 methaemoglobin as a surrogate endpoint for recurrence would reduce required sample sizes by approximately 40%. Study limitations include the inability to distinguish between recrudescence, reinfection, and relapse in P. vivax recurrences. Conclusions For a given primaquine regimen, higher methaemoglobin on day 7 was associated with a reduced risk of P. vivax recurrence. Under our proposed causal model, this justifies the use of methaemoglobin as a population-level surrogate endpoint for primaquine antihypnozoite activity in patients with P. vivax malaria who have normal G6PD activity.