Single Low Dose Primaquine (0.25mg/kg) Does Not Cause Clinically Significant Haemolysis in G6PD Deficient Subjects

Background

Primaquine is the only drug consistently effective against mature gametocytes of Plasmodium falciparum. The transmission blocking dose of primaquine previously recommended was 0.75mg/kg (adult dose 45mg) but its deployment was limited because of concerns over haemolytic effects in patients with glucose-6-phosphate dehydrogenase (G6PD) deficiency. G6PD deficiency is an inherited X-linked enzymatic defect that affects an estimated 400 million people around the world with high frequencies (15–20%) in populations living in malarious areas. To reduce transmission in low transmission settings and facilitate elimination of P. falciparum, the World Health Organization now recommends adding a single dose of 0.25mg/kg (adult dose 15mg) to Artemisinin-based Combination Therapies (ACTs) without G6PD testing. Direct evidence of the safety of this low dose is lacking. Adverse events and haemoglobin variations after this treatment were assessed in both G6PD normal and deficient subjects in the context of targeted malaria elimination in a malaria endemic area on the North-Western Myanmar-Thailand border where prevalence of G6PD deficiency (Mahidol variant) approximates 15%.

Methods and Findings

The tolerability and safety of primaquine (single dose 0.25 mg base/kg) combined with dihydroartemisinin-piperaquine (DHA-PPQ) given three times at monthly intervals was assessed in 819 subjects. Haemoglobin concentrations were estimated over the six months preceding the ACT + primaquine rounds of mass drug administration. G6PD deficiency was assessed with a phenotypic test and genotyping was performed in male subjects with deficient phenotypes and in all females. Fractional haemoglobin changes in relation to G6PD phenotype and genotype and primaquine round were assessed using linear mixed-effects models. No adverse events related to primaquine were reported during the trial. Mean fractional haemoglobin changes after each primaquine treatment in G6PD deficient subjects (-5.0%, -4.2% and -4.7%) were greater than in G6PD normal subjects (0.3%, -0.8 and -1.7%) but were clinically insignificant. Fractional drops in haemoglobin concentration larger than 25% following single dose primaquine were observed in 1.8% of the population but were asymptomatic.

Conclusions

The single low dose (0.25mg/kg) of primaquine is clinically well tolerated and can be used safely without prior G6PD testing in populations with high prevalence of G6PD deficiency. The present evidence supports a broader use of low dose primaquine without G6PD testing for the treatment and elimination of falciparum malaria.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT01872702","term_id":"NCT01872702"}}NCT01872702     

Perioperative Myocardial Injury after Adult Heart Transplant: Determinants and Prognostic Value

MethodsData on 362 consecutive recipients (mean age: 47.8±13.7, 20.2% female, 18.2% diabetics, 22.1% with previous cardiac operations, 27.6% hospitalized, 84.9±29.4 ml/min preoperative glomerular filtration rate) were analyzed using multivariable logistic regression modeling. Target outcomes were determinants of troponin release, early graft failure (EGF), acute kidney injury (AKI) and operative death.ResultsMean cTnI release measured 24 hours after transplant was 10.9±11.6 μg/L. Overall hospital mortality was 10.8%, EGF 10.5%, and AKI was 12.2%. cTnI release>10 μg/L proved an independent predictor of EGF (OR 2.2; 95% CI, 1.06–4.6) and AKI (OR 1.031; 95% CI, 1.001-1.064). EGF, in turn, proved a determinant of hospital mortality. Risk factors for cTnI>10 μg/L release were: status 2B (OR 0.35; 95% CI, 0.18-0.69, protective), duration of the ischemic period (OR 1.006; 95% CI, 1.001-1.011), previous cardiac operation (OR 2.9; 95% CI, 1.67-5.0), and left ventricular hypertrophy (OR 3.3; 95% CI, 1.9-5.6).ConclusionsMyocardial enzyme leakage clearly emerged as an epiphenomenon of more complicated clinical course. The complex interplay between surgical procedure features, graft characteristics and recipient end-organ function highlights cTnI release as a risk marker of graft failure and acute kidney injury. The search for optimal myocardial preservation is still an issue.     

Morphological and molecular characterization of healthy human ascending aorta

Knowledge of the characteristics of the normal human aorta has been constrained by lack of data on fresh aortic tissue, especially from healthy individuals. In this study, the gene expression and morphological characteristics of the thoracic ascending aorta (AA) of healthy organ donors have been evaluated, with the aim of providing reference data for the analysis of pathological AAs. We analysed by RT-PCR the differential expression of mRNAs coding for myocardin, smoothelin, alpha-smooth muscle actin (alpha-SMA) and the ED-A isoform of fibronectin (ED-A FN) in AA specimens from donors, integrating the results with immunohistochemical analysis of the same targets. Morphological and morphometric characteristics of the AAs were also evaluated. In order to account for possible regional variations in wall structure, the convexity of the aortic profile was compared to the concavity. No differences in gene expression occurred for any of the target genes between the concavity and the convexity of AAs. Immunohistochemistry revealed a different distribution of total FN and of its ED-A isoform in the media and in the intima. Smoothelin is expressed by the majority of cells in the media, with some positive cells also in the intima. Alpha-SMA is expressed in all the tunicae. Immunohistochemistry also revealed in the convexity of 50% of AAs the presence of discrete areas in the subadventital media with altered structure and cell morphology and with altered gene expression, resulting positive for ED-A FN and alpha-SMA, but not for smoothelin, indicating the occurrence of early lesions also in macroscopically healthy AAs.     

Assessment of therapeutic responses to gametocytocidal drugs in Plasmodium falciparum malaria

Background

Effective mating between laboratory-reared males and wild females is paramount to the success of vector control strategies aiming to decrease disease transmission via the release of sterile or genetically modified male mosquitoes. However mosquito colonization and laboratory maintenance have the potential to negatively affect male genotypic and phenotypic quality through inbreeding and selection, which in turn can decrease male mating competitiveness in the field. To date, very little is known about the impact of those evolutionary forces on the reproductive biology of mosquito colonies and how they ultimately affect male reproductive fitness.

Methods

Here several male reproductive physiological traits likely to be affected by inbreeding and selection following colonization and laboratory rearing were examined. Sperm length, and accessory gland and testes size were compared in male progeny from field-collected females and laboratory strains of Anopheles gambiae sensu stricto colonized from one to over 25 years ago. These traits were also compared in the parental and sequentially derived, genetically modified strains produced using a two-phase genetic transformation system. Finally, genetic crosses were performed between strains in order to distinguish the effects of inbreeding and selection on reproductive traits.

Results

Sperm length was found to steadily decrease with the age of mosquito colonies but was recovered in refreshed strains and crosses between inbred strains therefore incriminating inbreeding costs. In contrast, testes size progressively increased with colony age, whilst accessory gland size quickly decreased in males from colonies of all ages. The lack of heterosis in response to crossing and strain refreshing in the latter two reproductive traits suggests selection for insectary conditions.

Conclusions

These results show that inbreeding and selection differentially affect reproductive traits in laboratory strains overtime and that heterotic ‘supermales’ could be used to rescue some male reproductive characteristics. Further experiments are needed to establish the exact relationship between sperm length, accessory gland and testes size, and male reproductive success in the laboratory and field settings.     

Haemoglobin C and S role in acquired immunity against Plasmodium falciparum malaria

A recently proposed mechanism of protection for haemoglobin C (HbC; beta6Glu-->Lys) links an abnormal display of PfEMP1, an antigen involved in malaria pathogenesis, on the surface of HbC infected erythrocytes together with the observation of reduced cytoadhesion of parasitized erythrocytes and impaired rosetting in vitro. We investigated the impact of this hypothesis on the development of acquired immunity against Plasmodium falciparum variant surface antigens (VSA) encoding PfEMP1 in HbC in comparison with HbA and HbS carriers of Burkina Faso. We measured: i) total IgG against a single VSA, A4U, and against a panel of VSA from severe malaria cases in human sera from urban and rural areas of Burkina Faso of different haemoglobin genotypes (CC, AC, AS, SC, SS); ii) total IgG against recombinant proteins of P. falciparum asexual sporozoite, blood stage antigens, and parasite schizont extract; iii) total IgG against tetanus toxoid. Results showed that the reported abnormal cell-surface display of PfEMP1 on HbC infected erythrocytes observed in vitro is not associated to lower anti- PfEMP1 response in vivo. Higher immune response against the VSA panel and malaria antigens were observed in all adaptive genotypes containing at least one allelic variant HbC or HbS in the low transmission urban area whereas no differences were detected in the high transmission rural area. In both contexts the response against tetanus toxoid was not influenced by the beta-globin genotype. These findings suggest that both HbC and HbS affect the early development of naturally acquired immunity against malaria. The enhanced immune reactivity in both HbC and HbS carriers supports the hypothesis that the protection against malaria of these adaptive genotypes might be at least partially mediated by acquired immunity against malaria.     

High Risk of Severe Anaemia after Chlorproguanil-Dapsone+Artesunate Antimalarial Treatment in Patients with G6PD (A-) Deficiency

Background

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common inherited human enzyme defect. This deficiency provides some protection from clinical malaria, but it can also cause haemolysis after administration of drugs with oxidant properties.

Methods

The safety of chlorproguanil-dapsone+artesunate (CD+A) and amodiaquine+sulphadoxine-pyrimethamine (AQ+SP) for the treatment of uncomplicated P. falciparum malaria was evaluated according to G6PD deficiency in a secondary analysis of an open-label, randomized clinical trial [1]. 702 children, treated with CD+A or AQ+SP and followed for 28 days after treatment were genotyped for G6PD A- deficiency.

Findings

In the first 4 days following CD+A treatment, mean haematocrit declined on average 1.94% (95% CI 1.54 to 2.33) and 1.05% per day (95% CI 0.95 to 1.15) respectively in patients with G6PD deficiency and normal patients; a mean reduction of 1.3% per day was observed among patients who received AQ+SP regardless of G6PD status (95% CI 1.25 to 1.45). Patients with G6PD deficiency recipients of CD+A had significantly lower haematocrit than the other groups until day 7 (p = 0.04). In total, 10 patients had severe post-treatment haemolysis requiring blood transfusion. Patients with G6PD deficiency showed a higher risk of severe anaemia following treatment with CD+A (RR = 10.2; 95% CI 1.8 to 59.3) or AQ+SP (RR = 5.6; 95% CI 1.0 to 32.7).

Conclusions

CD+A showed a poor safety profile in individuals with G6PD deficiency most likely as a result of dapsone induced haemolysis. Screening for G6PD deficiency before drug administration of potentially pro-oxidants drugs, like dapsone-containing combinations, although seldom available, is necessary.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00461578","term_id":"NCT00461578"}}NCT00461578     

Haemoglobin C and S in natural selection against Plasmodium falciparum malaria: a plethora or a single shared adaptive mechanism?

Conclusive evidence exists on the protective role against clinical Plasmodium falciparum malaria of Haemoglobin S (beta 6Glu-->Val) and HbC (HbC; beta 6Glu-->Lys), both occurring in sub-Saharan Africa. However, the mechanism/s of the protection exerted remain/s debated for both haemoglobin variants, HbC and HbS. Recently, an abnormal display of PfEMP1, an antigen involved in malaria pathogenesis, was reported on HbAC and HbCC infected erythrocytes that showed reduced cytoadhesion and impaired rosetting in vitro. On this basis it has been proposed that HbC protection might be attributed to the reduced PfEMP1-mediated adherence of parasitized erythrocytes in the microvasculature. Furthermore, impaired cytoadherence was observed in HbS carriers suggesting for the first time a convergence in the protection mechanism of these two haemoglobin variants. We investigated the impact of this hypothesis on the development of acquired immunity against P. falciparum variant surface antigens (VSA) encoding PfEMP1 in HbC and HbS carriers in comparison with HbA of Burkina Faso. Higher immune response against a VSA panel and several malaria antigens were observed in all adaptive genotypes containing at least one allelic variant HbC or HbS in the low transmission urban area whereas no differences were detected in the high transmission rural area. In both contexts the response against tetanus toxoid was not influenced by the beta-globin genotype. Thus, these findings suggest that both HbC and HbS affect the early development of naturally acquired immunity against malaria. We reviewed the hypothesized mechanisms so far proposed in light of these recent results.     

Analysis of Extracellular Superoxide Dismutase and Akt in Ascending Aortic Aneurysm With Tricuspid or Bicuspid Aortic Valve

Ascending aortic aneurysm (AsAA) is a consequence of medial degeneration (MD), deriving from apoptotic loss of smooth muscle cells (SMC) and fragmentation of elastin and collagen fibers. Alterations of extracellular matrix structure and protein composition, typical of medial degeneration, can modulate intracellular pathways. In this study we examined the relevance of extracellular superoxide dismutase (SOD3) and Akt in AsAA pathogenesis, evaluating their tissue distribution and protein levels in ascending aortic tissues from controls (n=6), patients affected by AsAA associated to tricuspid aortic valve (TAV, n=9) or bicuspid aortic valve (BAV, n=9). The results showed a significant reduction of SOD3, phospho-Akt and Akt protein levels in AsAA tissues from patients with BAV, compared to controls, whereas the differences observed between controls and patients with TAV were not significant. The decreased levels of SOD3 and Akt in BAV aortic tissues are associated with decreased Erk1/Erk2 phosphorylation and MMP-9 levels increase. The authors suggest a role of decreased SOD3 protein levels in the progression of AsAA with BAV and a link between ECM modifications of aortic media layer and impaired Erk1/Erk2 and Akt signaling in the late stages of the aortopathy associated with BAV.Key words: Ascending aortic aneurysm, bicuspid aortic valve, medial degeneration, smooth muscle cells, extracellular superoxide dismutase, Akt, tri-cuspid aortic valve     

Validation of G6PD Point-of-Care Tests among Healthy Volunteers in Yangon,Myanmar

Primaquine and other 8-amnoquinoline based anti-malarials can cause haemolysis in subjects with glucose-6-phosphate dehydrogenase (G6PD) deficiency. Correct diagnosis of G6PD status in patients is crucial for safe treatment of both relapsing stages of Plasmodium vivax and transmitting forms of Plasmodium falciparum. Lack of suitable point-of-care tests has hampered a much needed wide use of primaquine for malaria elimination. In this study we have assessed the performances of two qualitative tests, the fluorescent spot test (FST) and the G6PD CareStart test (CST), against the gold standard quantitative spectrophotometric assay in a population of 1000 random adult healthy volunteers living in Yangon, Myanmar. The prevalence of G6PD deficiency in the Bamar, Karen and in the whole sample set was 6.6% (10.1% in males), 9.2% (21.0% in males) and 6.8% (11.1% in males) respectively. The FST and CST showed comparable performances with sensitivity over 95% and specificity over 90%, however for cases with severe G6PD activity the FTS had improved performance. If used with a conservative interpretation of the signal, the CareStart test has the potential to be used in the field and, by allowing a wider use of primaquine, to help malaria elimination.