Adherence to Anti-Malarial Treatment in Malaria Endemic Areas of Bangladesh.

Ensuring adherence to antimalarial treatment is crucial for achieving a radical cure and elimination of malaria, especially in hard-to-reach areas. We conducted this study to assess the current scenario of drug adherence in four endemic sub-districts of Bangladesh. Among 110 enrolled participants, 70% were mono-infected with Plasmodium falciparum and the remaining 30% with P. vivax. The overall treatment adherence frequency was 92.7% (95% CI: 83.0-96.3%). A total of eight participants were found to be nonadherent to treatment and all of them were from Bandarban. Level of nonadherence was equally observed in two age groups: 11-17 and 18+ years. However, male participants (n = 6) were found to be more nonadherent than females (n = 2). Among 7.3% with nonadherence to treatment, a single participant with P. falciparum mono-infection refused to take medication and became nonadherent. Remaining participants stated that they were feeling well and going to work, thus leaving treatment course uncompleted. Although overall compliance with malaria medication seems good, a gradual increase in noncompliance to P. vivax malaria treatment suggests that the National Malaria Elimination Program must be enhanced and monitored to fulfil the projected malaria elimination goal before 2030 from Bangladesh.

Primaquine radical cure in patients with Plasmodium falciparum malaria in areas co-endemic for P falciparum and Plasmodium vivax (PRIMA): a multicentre, open-label, superiority randomised controlled trial.

BACKGROUND: In areas co-endemic for Plasmodium vivax and Plasmodium falciparum there is an increased risk of P vivax parasitaemia following P falciparum malaria. Radical cure is currently only recommended for patients presenting with P vivax malaria. Expanding the indication for radical cure to patients presenting with P falciparum malaria could reduce their risk of subsequent P vivax parasitaemia. METHODS: We did a multicentre, open-label, superiority randomised controlled trial in five health clinics in Bangladesh, Indonesia, and Ethiopia. In Bangladesh and Indonesia, patients were excluded if they were younger than 1 year, whereas in Ethiopia patients were excluded if they were younger than 18 years. Patients with uncomplicated P falciparum monoinfection who had fever or a history of fever in the 48 h preceding clinic visit were eligible for enrolment and were required to have a glucose-6-dehydrogenase (G6PD) activity of 70% or greater. Patients received blood schizontocidal treatment (artemether-lumefantrine in Ethiopia and Bangladesh and dihydroartemisinin-piperaquine in Indonesia) and were randomly assigned (1:1) to receive either high-dose short-course oral primaquine (intervention arm; total dose 7 mg/kg over 7 days) or standard care (standard care arm; single dose oral primaquine of 0·25 mg/kg). Random assignment was done by an independent statistician in blocks of eight by use of sealed envelopes. All randomly assigned and eligible patients were included in the primary and safety analyses. The per-protocol analysis excluded those who did not complete treatment or had substantial protocol violations. The primary endpoint was the incidence risk of P vivax parasitaemia on day 63. This trial is registered at ClinicalTrials.gov, NCT03916003. FINDINGS: Between Aug 18, 2019, and March 14, 2022, a total of 500 patients were enrolled and randomly assigned, and 495 eligible patients were included in the intention-to-treat analysis (246 intervention and 249 control). The incidence risk of P vivax parasitaemia at day 63 was 11·0% (95% CI 7·5-15·9) in the standard care arm compared with 2·5% (1·0-5·9) in the intervention arm (hazard ratio 0·20, 95% CI 0·08-0·51; p=0·0009). The effect size differed with blood schizontocidal treatment and site. Routine symptom reporting on day 2 and day 7 were similar between groups. In the first 42 days, there were a total of four primaquine-related adverse events reported in the standard care arm and 26 in the intervention arm; 132 (92%) of all 143 adverse events were mild. There were two serious adverse events in the intervention arm, which were considered unrelated to the study drug. None of the patients developed severe anaemia (defined as haemoglobin <5 g/dL). INTERPRETATION: In patients with a G6PD activity of 70% or greater, high-dose short-course primaquine was safe and relatively well tolerated and reduced the risk of subsequent P vivax parasitaemia within 63 days by five fold. Universal radical cure therefore potentially offers substantial clinical, public health, and operational benefits, but these benefits will vary with endemic setting. FUNDING: Australian Academy of Science Regional Collaborations Program, Bill & Melinda Gates Foundation, and National Health and Medical Research Council.

Adapting international clinical trials during COVID-19 and beyond.

BACKGROUND: The COVID-19 pandemic and resulting restrictions, particularly travel restrictions, have had significant impact on the conduct of global clinical trials. Our clinical trials programme, which relied on in-person visits for training, monitoring and capacity building across nine low- and middle-income countries, had to adapt to those unprecedented operational challenges. We report the adaptation of our working model with a focus on the operational areas of training, monitoring and cross-site collaboration. THE NEW WORKING MODEL: Adaptations include changing training strategies from in-person site visits with three or four team members to a multi-pronged virtual approach, with generic online training for good clinical practice, the development of a library of study-specific training videos, and interactive virtual training sessions, including practical laboratory-focused training sessions. We also report changes from in-person monitoring to remote monitoring as well as the development of a more localized network of clinical trial monitors to support hybrid models with in-person and remote monitoring depending on identified risks at each site. We established a virtual network across different trial and study sites with the objective to further build capacity for good clinical practice-compliant antimalarial trials and foster cross-country and cross-study site collaboration. CONCLUSION: The forced adaptation of these new strategies has come with advantages that we did not envisage initially. This includes improved, more frequent engagement through the established network with opportunities for increased south-to-south support and a substantially reduced carbon footprint and budget savings. Our new approach is challenging for study sites with limited prior experience but this can be overcome with hybrid models. Capacity building for laboratory-based work remains difficult using a virtual environment. The changes to our working model are likely to last, even after the end of the pandemic, providing a more sustainable and equitable approach to our research.

Direct Nasal Swab for Rapid Test and Saliva as an Alternative Biological Sample for RT-PCR in COVID-19 Diagnosis.

Accurate and early diagnoses are prerequisites for prompt treatment. For coronavirus disease 2019 (COVID-19), it is even more crucial. Currently, choice of methods include rapid diagnostic tests and reverse transcription polymerase chain reaction (RT-PCR) using samples mostly of respiratory origin and sometimes saliva. We evaluated two rapid diagnostic tests with three specimen types using viral transport medium (VTM) containing naso-oropharyngeal (NOP) swabs, direct nasal and direct nasopharyngeal (NP) samples from 428 prospective patients. We also performed RT-PCR for 428 NOP VTM and 316 saliva samples to compare results. The sensitivity of the SD Biosensor Standard Q COVID-19 antigen (Ag) test kit drastically raised from an average of 65.55% (NOP VTM) to 85.25% (direct nasal samples), while RT-PCR was the gold standard. For the CareStart kit, the sensitivity was almost similar for direct NP swabs; the average was 84.57%. The specificities were ≥95% for both SD Biosensor Standard Q and CareStart COVID-19 Ag tests in all platforms. The kits were also able to detect patients with different variants as well. Alternatively, RT-PCR results from saliva and NOP VTM samples showed high sensitivities of 96.45% and 95.48% with respect to each other as standard. The overall results demonstrated high performance of the rapid tests, indicating the suitability for regular surveillance at clinical facilities when using direct nasal or direct NP samples rather than NOP VTM. Additionally, the analysis also signifies not showed that RT-PCR of saliva can be used as an choice of method to RT-PCR of NOP VTM, providing an easier, non-invasive sample collection method. IMPORTANCE There are several methods for the diagnosis of coronavirus disease 2019 (COVID-19), and the choice of methods depends mostly on the resources and level of sensitivity required by the user and health care providers. Still, reverse transcription polymerase chain reaction (RT-PCR) has been chosen as the best method using direct naso-oropharyngeal swabs. There are also other methods of fast detection, such as rapid diagnostic tests (RDTs), which offer result within 15 to 20 min and have become quite popular for self-testing and in the clinical setting. The major drawback of the currently used RT-PCR method is compliance, as it may cause irritation, and patients often refuse to test in such a way. RDTs, although inexpensive, suffer from low sensitivity due to technical issues. In this article, we propose saliva as a noninvasive source for RT-PCR samples and evaluate various specimen types at different times after infection for the best possible output from COVID-19 rapid tests.

Health hazards related to using masks and/or personal protective equipment among physicians working in public hospitals in Dhaka: A cross-sectional study.

BACKGROUND: Wearing masks or personal protective equipment (PPE) has become an integral part of the occupational life of physicians due to the coronavirus disease 2019 (COVID-19) pandemic. Most physicians have been developing various health hazards related to the use of different protective gears. This study aimed to determine the burden and spectrum of various health hazards associated with using masks or PPE and their associated risk factors. METHODS: This cross-sectional survey was conducted in Dhaka Medical College from March 01-May 30, 2021, among physicians from different public hospitals in Dhaka, Bangladesh. We analyzed the responses of 506 physicians who completed case record forms through Google forms or hard copies. FINDINGS: The mean (SD) age of the respondents was 35.4 [7.7], and 69.4% were men. Approximately 40% were using full PPE, and 55% were using N-95 masks. A total of 489 (96.6%) patients experienced at least one health hazard. The reported severe health hazards were syncope, severe dyspnea, severe chest pain, and anaphylaxis. Headache, dizziness, mood irritation, chest pain, excessive sweating, panic attack, and permanent facial disfigurement were the minor health hazards reported. Extended periods of work in the COVID-19-unit, reuse of masks, diabetes, obesity, and mental stress were risk factors for dyspnea. The risk factors for headaches were female sex, diabetes, and previous primary headaches. Furthermore, female sex and reusing masks for an extended period (> 6 h) were risk factors for facial disfigurement. The risk factors for excessive sweating were female sex and additional evening office practice for an extended period. CONCLUSIONS: Healthcare workers experienced several occupational hazards after using masks and PPE. Therefore, an appropriate policy is required to reduce such risks.

Heterogeneity in prevalence of subclinical Plasmodium falciparum and Plasmodium vivax infections but no parasite genomic clustering in the Chittagong Hill Tracts, Bangladesh.

BACKGROUND: Malaria remains endemic in Bangladesh, with the majority of cases occurring in forested, mountainous region in the Chittagong Hill Tracts (CHT). This area is home to Bengali and diverse groups of indigenous people (Pahari) residing largely in mono-ethnic villages. METHODS: 1002 individuals of the 9 most prominent Pahari and the Bengali population were randomly selected and screened by RDT and qPCR. Parasites were genotyped by msp2 and deep sequencing of 5 amplicons (ama1-D3, cpmp, cpp, csp, and msp7) for Plasmodium falciparum (n = 20), and by microsatellite (MS) typing of ten loci and amplicon sequencing of msp1 for Plasmodium vivax (n = 21). Population structure was analysed using STRUCTURE software. Identity-by-state (IBS) was calculated as a measure of parasite relatedness and used to generate relatedness networks. RESULTS: The prevalence of P. falciparum and P. vivax infection was 0.7% by RDT (P. falciparum 6/1002; P. vivax 0/1002, mixed: 1/1002) and 4% by qPCR (P. falciparum 21/1002; P. vivax 16/1002, mixed: 5/1002). Infections were highly clustered, with 64% (27/42) of infections occurring in only two Pahari groups, the Khumi and Mro. Diversity was high; expected heterozygosity was 0.93 for P. falciparum and 0.81 for P. vivax. 85.7% (18/21) of P. vivax and 25% (5/20) of P. falciparum infections were polyclonal. No population structure was evident for either species, suggesting high transmission and gene flow among Pahari groups. CONCLUSIONS: High subclinical infection prevalence and genetic diversity mirror ongoing transmission. Control activities should be specifically directed to Pahari groups at greatest risk.

Variation in Glucose-6-Phosphate Dehydrogenase activity following acute malaria.

Primaquine and tafenoquine are the only licensed drugs with activity against Plasmodium vivax hypnozoites but cause haemolysis in patients with glucose-6-phosphate dehydrogenase (G6PD) deficiency. Malaria also causes haemolysis, leading to the replacement of older erythrocytes with low G6PD activity by reticulocytes and young erythrocytes with higher activity. Aim of this study was to assess the impact of acute malaria on G6PD activity. Selected patients with uncomplicated malaria were recruited in Bangladesh (n = 87), Indonesia (n = 75), and Ethiopia (n = 173); G6PD activity was measured at the initial presentation with malaria and a median of 176 days later (range 140 to 998) in the absence of malaria. Among selected participants (deficient participants preferentially enrolled in Bangladesh but not at other sites) G6PD activity fell between malaria and follow up by 79.1% (95%CI: 40.4 to 117.8) in 6 participants classified as deficient (<30% activity), 43.7% (95%CI: 34.2 to 53.1) in 39 individuals with intermediate activity (30% to <70%), and by 4.5% (95%CI: 1.4 to 7.6) in 290 G6PD normal (≥70%) participants. In Bangladesh and Indonesia G6PD activity was significantly higher during acute malaria than when the same individuals were retested during follow up (40.9% (95%CI: 33.4-48.1) and 7.4% (95%CI: 0.2 to 14.6) respectively), whereas in Ethiopia G6PD activity was 3.6% (95%CI: -1.0 to -6.1) lower during acute malaria. The change in G6PD activity was apparent in patients presenting with either P. vivax or P. falciparum infection. Overall, 66.7% (4/6) severely deficient participants and 87.2% (34/39) with intermediate deficiency had normal activities when presenting with malaria. These findings suggest that G6PD activity rises significantly and at clinically relevant levels during acute malaria. Prospective case-control studies are warranted to confirm the degree to which the predicted population attributable risks of drug induced haemolysis is lower than would be predicted from cross sectional surveys.

Reducing the risk of Plasmodium vivax after falciparum infections in co-endemic areas-a randomized controlled trial (PRIMA).

BACKGROUND: Plasmodium vivax forms dormant liver stages that can reactivate weeks or months following an acute infection. Recurrent infections are often associated with a febrile illness and can cause a cumulative risk of severe anaemia, direct and indirect mortality, and onward transmission of the parasite. There is an increased risk of P. vivax parasitaemia following falciparum malaria suggesting a rationale for universal use of radically curative treatment in patients with P. falciparum malaria even in the absence of detectable P. vivax parasitaemia in areas that are co-endemic for both species. METHODS: This is a multicentre, health care facility-based, randomized, controlled, open-label trial in Bangladesh, Indonesia and Ethiopia. Patients with uncomplicated falciparum malaria, G6PD activity of ≥70% of the adjusted male median (AMM) and haemoglobin levels ≥8g/dl are recruited into the study and randomized to either receive standard schizonticidal treatment plus 7-day high dose primaquine (total dose 7mg/kg) or standard care in a 1:1 ratio. Patients are followed up weekly until day 63. The primary endpoint is the incidence risk of any P. vivax parasitemia on day 63. Secondary endpoints include incidence risk on day 63 of symptomatic P. vivax malaria and the risk of any P. falciparum parasitaemia. Secondary safety outcomes include the proportion of adverse events and serious adverse events, the incidence risk of severe anaemia (Hb<5g/dl and <7g/dl) and/or the risk for blood transfusion, the incidence risk of ≥ 25% fall in haemoglobin with and without haemoglobinuria, and the incidence risk of ≥ 25% fall in haemoglobin to under 7g/dl with and without haemoglobinuria. DISCUSSION: This study evaluates the potential benefit of a universal radical cure for both P. vivax and P. falciparum in different endemic locations. If found safe and effective universal radical cure could represent a cost-effective approach to clear otherwise unrecognised P. vivax infections and hence accelerate P. vivax elimination. TRIAL REGISTRATION: NCT03916003 . Registered on 12 April 2019.

Solar geoengineering could redistribute malaria risk in developing countries.

Solar geoengineering is often framed as a stopgap measure to decrease the magnitude, impacts, and injustice of climate change. However, the benefits or costs of geoengineering for human health are largely unknown. We project how geoengineering could impact malaria risk by comparing current transmission suitability and populations-at-risk under moderate and high greenhouse gas emissions scenarios (Representative Concentration Pathways 4.5 and 8.5) with and without geoengineering. We show that if geoengineering deployment cools the tropics, it could help protect high elevation populations in eastern Africa from malaria encroachment, but could increase transmission in lowland sub-Saharan Africa and southern Asia. Compared to extreme warming, we find that by 2070, geoengineering would nullify a projected reduction of nearly one billion people at risk of malaria. Our results indicate that geoengineering strategies designed to offset warming are not guaranteed to unilaterally improve health outcomes, and could produce regional trade-offs among Global South countries that are often excluded from geoengineering conversations.

Clinical Evaluation of a New Antigen-Based COVID-19 Rapid Diagnostic Test from Symptomatic Patients.

Accurate diagnosis at the right moment is the prerequisite for treatment of any disease. Failure to correctly diagnose a disease can result in highly detrimental effects, unmistakably a crucial factor during the COVID-19 pandemic. RT-PCR is the gold standard for COVID-19 detection while there are other test procedures available, such as LAMP, X-Ray, and ELISA. However, these tests are expensive, require sophisticated equipment and a highly trained workforce, and multiple hours or even days are often required to obtain the test results. A rapid and cheap detection system can thus render a solution to the screening system on a larger scale and be added as an aid to the current detection processes. Recently, some rapid antigen-based COVID-19 tests devices have been developed and commercialized. In this study, we evaluated the clinical performance of a new rapid detection device (OnSite® COVID-19 Ag Rapid Test by CTK Biotech Inc., Poway, CA, USA) on COVID-19 symptomatic patients (n = 380). The overall sensitivity and specificity were 91.0% (95% CI: 84.8-95.3%) and 99.2% (95% CI: 97.1-99.9), against gold standard RT-PCR. The kit was capable of detecting patients even after 06 days of onset of symptoms and the sensitivity can be maximized to 98% in samples with an average RT-PCR Ct ≤ 26.48, demonstrating a high potential of the kit for clinical diagnosis of symptomatic patients in healthcare facilities.

Glucose-6-phosphate dehydrogenase activity in individuals with and without malaria: Analysis of clinical trial, cross-sectional and case-control data from Bangladesh.

BACKGROUND: Glucose-6-phosphate dehydrogenase (G6PD) activity is dependent upon G6PD genotype and age of the red blood cell (RBC) population, with younger RBCs having higher activity. Peripheral parasitemia with Plasmodium spp. induces hemolysis, replacing older RBCs with younger cells with higher G6PD activity. This study aimed to assess whether G6PD activity varies between individuals with and without malaria or a history of malaria. METHODS AND FINDINGS: Individuals living in the Chittagong Hill Tracts of Bangladesh were enrolled into 3 complementary studies: (i) a prospective, single-arm clinical efficacy trial of patients (n = 175) with uncomplicated malaria done between 2014 and 2015, (ii) a cross-sectional survey done between 2015 and 2016 (n = 999), and (iii) a matched case-control study of aparasitemic individuals with and without a history of malaria done in 2020 (n = 506). G6PD activity was compared between individuals with and without malaria diagnosed by microscopy, rapid diagnostic test (RDT), or polymerase chain reaction (PCR), and in aparasitemic participants with and without a history of malaria. In the cross-sectional survey and clinical trial, 15.5% (182/1,174) of participants had peripheral parasitemia detected by microscopy or RDT, 3.1% (36/1,174) were positive by PCR only, and 81.4% (956/1,174) were aparasitemic. Aparasitemic individuals had significantly lower G6PD activity (median 6.9 U/g Hb, IQR 5.2-8.6) than those with peripheral parasitemia detected by microscopy or RDT (7.9 U/g Hb, IQR 6.6-9.8, p < 0.001), but G6PD activity similar to those with parasitemia detected by PCR alone (submicroscopic parasitemia) (6.1 U/g Hb, IQR 4.8-8.6, p = 0.312). In total, 7.7% (14/182) of patients with malaria had G6PD activity < 70% compared to 25.0% (248/992) of participants with submicroscopic or no parasitemia (odds ratio [OR] 0.25, 95% CI 0.14-0.44, p < 0.001). In the case-control study, the median G6PD activity was 10.3 U/g Hb (IQR 8.8-12.2) in 253 patients with a history of malaria and 10.2 U/g Hb (IQR 8.7-11.8) in 253 individuals without a history of malaria (p = 0.323). The proportion of individuals with G6PD activity < 70% was 11.5% (29/253) in the cases and 15.4% (39/253) in the controls (OR 0.7, 95% CI 0.41-1.23, p = 0.192). Limitations of the study included the non-contemporaneous nature of the clinical trial and cross-sectional survey. CONCLUSIONS: Patients with acute malaria had significantly higher G6PD activity than individuals without malaria, and this could not be accounted for by a protective effect of G6PD deficiency. G6PD-deficient patients with malaria may have higher than expected G6PD enzyme activity and an attenuated risk of primaquine-induced hemolysis compared to the risk when not infected.

Persistence of Markers of Chloroquine Resistance in Plasmodium falciparum in Bangladesh.

The control of malaria, in terms of drug resistance, remains a significant global challenge, with Bangladesh, a malaria-endemic country, being no exception. The aim of this study was to explore antimalarial resistance in Bangladesh by molecular analysis of Plasmodium falciparum chloroquine resistance transporter (pfcrt) and P. falciparum multidrug resistance transporter 1 (pfmdr1) genetic markers of P. falciparum. Samples were obtained from uncomplicated malaria patients between 2009 and 2014 from six malaria-endemic districts. Based on parasite transmission intensity, the endemic districts were divided into high-transmission (Chittagong Hill Tracts [CHT]) and low-transmission (non-CHT) regions. Falciparum malaria-positive isolates were genotyped for K76T of the pfcrt gene, and N86Y and Y184F of the pfmdr1 gene: in total, 262 P. falciparum clinical isolates were analyzed. In CHT areas, the prevalence of polymorphisms was 70.6% for 76T, 14.4% for 86Y, and 7.8% for 184F. In non-CHT areas, 76T and 86Y mutations were found in 78.0% and 19.5% of the samples, respectively, whereas no 184F mutations were observed. We compared our data with previous similar molecular observations, which shows a significant decrease in pfcrt 76T mutation prevalence. No pfmdr1 amplification was observed in any of the samples suggesting an unaltered susceptibility to amino alcohol drugs such as mefloquine and lumefantrine. This study provides an updated assessment of the current status of pfcrt and pfmdr1 gene mutations in Bangladesh, and suggests there is persistent high prevalence of markers of resistance to aminoquinoline drugs.

A five-day course of ivermectin for the treatment of COVID-19 may reduce the duration of illness.

Ivermectin, a US Food and Drug Administration-approved anti-parasitic agent, was found to inhibit severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) replication in vitro. A randomized, double-blind, placebo-controlled trial was conducted to determine the rapidity of viral clearance and safety of ivermectin among adult SARS-CoV-2 patients. The trial included 72 hospitalized patients in Dhaka, Bangladesh, who were assigned to one of three groups: oral ivermectin alone (12 mg once daily for 5 days), oral ivermectin in combination with doxycycline (12 mg ivermectin single dose and 200 mg doxycycline on day 1, followed by 100 mg every 12 h for the next 4 days), and a placebo control group. Clinical symptoms of fever, cough, and sore throat were comparable among the three groups. Virological clearance was earlier in the 5-day ivermectin treatment arm when compared to the placebo group (9.7 days vs 12.7 days; p = 0.02), but this was not the case for the ivermectin + doxycycline arm (11.5 days; p = 0.27). There were no severe adverse drug events recorded in the study. A 5-day course of ivermectin was found to be safe and effective in treating adult patients with mild COVID-19. Larger trials will be needed to confirm these preliminary findings.

Wide range of G6PD activities found among ethnic groups of the Chittagong Hill Tracts, Bangladesh.

The proportion of Plasmodium vivax malaria among all malarias is increasing worldwide. Treatment with 8-aminoquinolines remain the only radical cure. However, 8-aminoquinolines can cause severe hemolysis in glucose-6-phosphate dehydrogenase (G6PD) deficient patients. The population of the multi-ethnic Chittagong Hill Tracts (CHT) carry the highest malaria burden within Bangladesh. As in many countries the national treatment guidelines recommend 8-aminoquinoline based radical cure without routine G6PD deficiency (G6PDd) testing to guide treatment. Aim of this study was to determine the need for routine testing within a multi-ethnic population by assessing the prevalence of G6PDd among the local population. Participants from 11 ethnicities were randomly selected and malaria status was assessed by microscopy, rapid diagnostic test (RDT) and polymerase chain reaction (PCR). G6PD status was determined by spectrophotometry and G6PD genotyping. The adjusted male median (AMM) was defined as 100% G6PD activity, participants were categorized as G6PD deficient (<30% activity), G6PD intermediate (30% to 70% activity) or G6PD normal (>70% activity). Median G6PD activities between ethnicities were compared and the association between G6PD activity and malaria status was assessed. 1002 participants were enrolled and tested for malaria. G6PD activity was measured by spectrophotometry in 999 participants and host G6PD genotyping undertaken in 323 participants. Seven participants (0.7%) had peripheral parasitaemia detected by microscopy or RDT and 42 by PCR (4.2%). Among 106 participants (32.8%) with confirmed genotype, 99 (93.4%) had the Mahidol variant. The AMM was 7.03U/gHb with 90 (9.0%) G6PD deficient participants and 133 (13.3%) with intermediate G6PD activity. Median G6PD activity differed significantly between ethnicities (p<0.001), proportions of G6PD deficient individuals ranged from 2% to 26% but did not differ between participants with and without malaria. The high G6PDd prevalence and significant variation between ethnicities suggest routine G6PDd testing to guide 8-aminoquinoline based radical in the CHT and comparable settings.

Field evaluation of quantitative point of care diagnostics to measure glucose-6-phosphate dehydrogenase activity.

BACKGROUND: Glucose-6-Phosphate dehydrogenase (G6PD) deficiency is the most common enzymopathy worldwide, no reliable bedside diagnostic tests to quantify G6PD activity exist. This study evaluated two novel quantitative G6PD diagnostics. METHODS: Participants with known G6PD activity were enrolled in Bangladesh. G6PD activity was measured by spectrophotometry, Biosensor (BS; AccessBio/CareStart, USA) and STANDARD G6PD (SG; SDBiosensor, ROK). G6PD activity was measured repeatedly in a subset of samples stored at room temperature and 4°C. RESULTS: 158 participants were enrolled, 152 samples tested by BS, 108 samples by SG and 102 samples were tested by all three methods. In comparison to spectrophotometry BS had sensitivity and specificity of 72% (95%CI: 53-86) and 100% (95%CI: 97-100) at 30% cut off respectively, while SG had a sensitivity of 100% (95%CI: 88-100) and specificity of 97% (95%CI: 91-99) at the same cut off. The sensitivity and specificity at 70% cut off activity were 71% (95%CI: 59-82) and 98% (95%CI, 92-100) respectively for BS and 89% (95%CI: 77-96) and 93% (95%CI: 83-98) respectively for SG. When an optimal cut-off was applied the sensitivity of the BS at 70 cut off rose to 91% [95%CI: 80-96] and specificity to 82% [95%CI: 83-89]; a diagnostic accuracy comparable to that of the SG (p = 0.879). G6PD activity dropped significantly (-0.31U/gHb, 95%CI: -0.61 to -0.01, p = 0.022) within 24 hours in samples stored at room temperature, but did not fall below 90% of baseline activity until day 13 (-0.87U/gHb, 95%CI: (-1.11 to -0.62), p<0.001). CONCLUSION: BS and SG are the first quantitative diagnostics to measure G6PD activity reliably at the bedside and represent suitable alternatives to spectrophotometry in resource poor settings. If samples are stored at 4°C, G6PD activity can be measured reliably for at least 7 days after sample collection.

A Comparison of Three Quantitative Methods to Estimate G6PD Activity in the Chittagong Hill Tracts, Bangladesh.

BACKGROUND: Glucose-6-phosphate-dehydrogenase-deficiency (G6PDd) is a major risk factor for primaquine-induced haemolysis. There is a need for improved point-of-care and laboratory-based G6PD diagnostics to unsure safe use of primaquine. METHODS: G6PD activities of participants in a cross-sectional survey in Bangladesh were assessed using two novel quantitative assays, the modified WST-8 test and the CareStart™ G6PD Biosensor (Access Bio), The results were compared with a gold standard UV spectrophotometry assay (Randox). The handheld CareStart™ Hb instrument (Access Bio) is designed to be a companion instrument to the CareStart™ G6PD biosensor, and its performance was compared to the well-validated HemoCue™ method. All quantitative G6PD results were normalized with the HemoCue™ result. RESULTS: A total of 1002 individuals were enrolled. The adjusted male median (AMM) derived by spectrophotometry was 7.03 U/g Hb (interquartile range (IQR): 5.38-8.69), by WST-8 was 7.03 U/g Hb (IQR: 5.22-8.16) and by Biosensor was 8.61 U/g Hb (IQR: 6.71-10.08). The AMM between spectrophotometry and WST-8 did not differ (p = 1.0) but differed significantly between spectrophotometry and Biosensor (p<0.01). Both, WST-8 and Biosensor were correlated with spectrophotometry (rs = 0.5 and rs = 0.4, both p<0.001). The mean difference in G6PD activity was -0.12 U/g Hb (95% limit of agreement (95% LoA): -5.45 to 5.20) between spectrophotometry and WST-8 and -1.74U/g Hb (95% LoA: -7.63 to 4.23) between spectrophotometry and Biosensor. The WST-8 identified 55.1% (49/89) and the Biosensor 19.1% (17/89) of individuals with G6PD activity <30% by spectrophotometry. Areas under the ROC curve did not differ significantly for the WST-8 and Biosensor irrespective of the cut-off activity applied (all p>0.05). Sensitivity and specificity for detecting G6PD activity <30% was 0.55 (95% confidence interval (95%CI): 0.44-0.66) and 0.98 (95%CI: 0.97-0.99) respectively for the WST-8 and 0.19 (95%CI: 0.12-0.29) and 0.99 (95%CI: 0.98-0.99) respectively for the Biosensor. Hb concentrations measured by HemoCue™ and CareStart™ Hb were strongly correlated (rs = 0.8, p<0.001, mean difference = 0.09 g Hb/dL, 95% LoA: -2.15 to 2.34). CONCLUSION: WST-8 and the CareStart™ G6PD Biosensor represent advances in G6PD diagnostics in resource poor settings, but will require further development before clinical deployment. The CareStart™ Hb instrument produced a precise measure of haemoglobin concentration.

Reduction in malaria prevalence and increase in malaria awareness in endemic districts of Bangladesh.

BACKGROUND: Malaria is endemic in 13 districts of Bangladesh. A baseline malaria prevalence survey across the endemic districts of Bangladesh was conducted in 2007, when the prevalence was reported around 39.7 per 1000 population. After two rounds of Global Fund to Fight AIDS, Tuberculosis and Malaria (GFATM)-funded intervention by the National Malaria Control Programme (NMCP) and a BRAC-led NGO consortium, a follow-up survey was conducted across the malaria-endemic districts of Bangladesh to measure the change in prevalence rate and in people's knowledge of malaria. METHODS: The survey was carried out from August to November 2013 in 70 upazilas (sub-districts) of 13 malaria-endemic districts of Bangladesh, following the same multi-stage cluster sampling design and the same number of households enrolled during the baseline prevalence survey in 2007, to collect 9750 randomly selected blood samples. For on-the-spot diagnosis of malaria, a rapid diagnostic test was used. The household head or eldest person available was interviewed using a pre-coded structured questionnaire to collect data on the knowledge and awareness of malaria in the household. RESULTS: Based on a weighted calculation, the overall malaria prevalence was found to be 1.41 per 1000 population. The proportion of Plasmodium falciparum mono-infection was 77.78% while both Plasmodium vivax mono-infection and mixed infection of the two species were found to be 11.11%. Bandarban had the highest prevalence (6.67 per 1000 population). Knowledge of malaria signs, symptoms and mode of transmission were higher in the follow-up survey (97.26%) than the baseline survey. Use of bed nets for prevention of malaria was found to be high (90.15%) at respondent level. People's knowledge of selected parameters increased significantly during the follow-up survey compared to the baseline survey conducted in 2007. CONCLUSIONS: A reduced prevalence rate of malaria and increased level of knowledge were observed in the present malaria prevalence survey in Bangladesh.

G6PD Deficiency and Antimalarial Efficacy for Uncomplicated Malaria in Bangladesh: A Prospective Observational Study.

BACKGROUND: The Bangladeshi national treatment guidelines for uncomplicated malaria follow WHO recommendations but without G6PD testing prior to primaquine administration. A prospective observational study was conducted to assess the efficacy of the current antimalarial policy. METHODS: Patients with uncomplicated malaria, confirmed by microscopy, attending a health care facility in the Chittagong Hill Tracts, Bangladesh, were treated with artemether-lumefantrine (days 0-2) plus single dose primaquine (0.75mg/kg on day2) for P. falciparum infections, or with chloroquine (days 0-2) plus 14 days primaquine (3.5mg/kg total over 14 days) for P. vivax infections. Hb was measured on days 0, 2 and 9 in all patients and also on days 16 and 30 in patients with P. vivax infection. Participants were followed for 30 days. The study was registered with the clinical trials website (NCT02389374). RESULTS: Between September 2014 and February 2015 a total of 181 patients were enrolled (64% P. falciparum, 30% P. vivax and 6% mixed infections). Median parasite clearance times were 22.0 (Interquartile Range, IQR: 15.2-27.3) hours for P. falciparum, 20.0 (IQR: 9.5-22.7) hours for P. vivax and 16.6 (IQR: 10.0-46.0) hours for mixed infections. All participants were afebrile within 48 hours, two patients with P. falciparum infection remained parasitemic at 48 hours. No patient had recurrent parasitaemia within 30 days. Adjusted male median G6PD activity was 7.82U/gHb. One male participant (1/174) had severe G6PD deficiency (<10% activity), five participants (5/174) had mild G6PD deficiency (10-60% activity). The Hb nadir occurred on day 2 prior to primaquine treatment in P. falciparum and P. vivax infected patients; mean fractional fall in Hb was -8.8% (95%CI -6.7% to -11.0%) and -7.4% (95%CI: -4.5 to -10.4%) respectively. CONCLUSION: The current antimalarial policy remains effective. The prevalence of G6PD deficiency was low. Main contribution to haemolysis in G6PD normal individuals was attributable to acute malaria rather than primaquine administration. TRIAL REGISTRATION: ClinicalTrials.gov NCT02389374.

Prevalence and diversity of avian haematozoan parasites in wetlands of bangladesh.

The parasites of genera Haemoproteus, Plasmodium, and Leucocytozoon are well-known avian haematozoa and can cause declined productivity and high mortality in wild birds. The objective of the study was to record the prevalence of haematozoan parasites in a wide range of wetland birds in Bangladesh. Six species of Haemoproteus, seven species of Plasmodium, one unidentified species of Leucocytozoon, and one unidentified microfilaria of the genus Paronchocerca were found. Data on the morphology, size, hosts, prevalence, and infection intensity of the parasites are provided. The overall prevalence among the birds was 29.5% (95 out of 322 birds). Of those, 13.2% (42 of 319) of birds were infected with Haemoproteus spp., 15.1% with Plasmodium spp. (48 of 319) and 0.6% with Leucocytozoon spp. (2 of 319). Two birds were positive for both Haemoproteus sp. and Plasmodium sp. A single resident bird, Ardeola grayii, was found positive for an unidentified microfilaria. Prevalence of infection varied significantly among different bird families. Wild birds of Bangladesh carry several types of haematozoan parasites. Further investigation with a larger sample size is necessary to estimate more accurately the prevalence of haematozoan parasites among wild birds as well as domestic ducks for better understanding of the disease ecology.