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BackgroundThe influence of the CYPs (cytochrome P-450) in the success of antimalarial therapy remains uncertain. In this study, the association of CYP2D6, CYP2C19 and CYP3A4 polymorphisms and predicted phenotypes with malaria recurrence was investigated.MethodsAfter diagnosis of vivax malaria, individuals treated at a reference center in Manaus were followed up for 180 days. Patients were separated into two groups: a recurrence group and a non-recurrence group. Genotyping of CYP2D6, CYP2C19 and CYP3A4 was performed using a TaqMan™ assay and real-time PCR.FindingsThe frequencies of decreased-function and normal-function alleles and phenotypes for all CYPs were similar between the groups, except for the CYP2D6*2xN allele (p=0.047) and the CYP2D6 gUM phenotype (p=0.057), which were more frequent in individuals without recurrence. Despite this, the CYP2D6, CYP2C19 and CYP3A4 genotypes had no association with an increased risk of recurrence. CYPs polymorphisms also had no influence in parasite clearance, neither in the time nor the number of recurrence episodes. MAINConclusionThis prospective cohort study demonstrated that CYP2D6, CYP2C19 and CYP3A4 polymorphisms have no influence on malaria recurrence. Nonetheless, our findings suggest that the CYP2D6 predicted ultrarapid phenotype was less susceptible to recurrence, and that patients with the CYP2D6 gUM phenotype are less susceptible to primaquine failure. Additional investigation of pharmacogenetics and pharmacokinetics are needed before implementing CYP analysis to better orientate individualized radical treatment of vivax malaria in reference centers that treat patients with multiple recurrences.

Background: Early recurrence of Plasmodium vivax is a challenge for malaria control in the field, particularly because this species is associated with lower parasitemia, which hinders diagnosis and monitoring through blood smear testing. Early recurrences, defined as the persistence of parasites in the peripheral blood despite adequate drug dosages, may arise from resistance to chloroquine. The objective of the study was to estimate early recurrence of P. vivax in the Brazilian Amazon by using a highly-sensitive detection method, in this case, PCR. Methods: An ultra-sensitive qPCR that targeted mitochondrial DNA was used to compare a standard qPCR that targeted 18S rDNA to detect early recurrence of P. vivax in very low densities in samples from patients treated with chloroquine. Results: Out of a total of 312 cases, 29 samples (9.3%) were characterized as recurrences, from which 3.2% (10/312) were only detected through ultra-sensitive qPCR testing. Conclusions: Studies that report the detection of P. vivax early recurrences using light microscopy may severely underestimate their true incidence.

Cytochrome P450 (CYP) enzymes are involved in the biotransformation of chloroquine (CQ), but the role of the different profiles of metabolism of this drug in relation to Plasmodium vivax recurrences has not been properly investigated. To investigate the influence of the CYP genotypes associated with CQ metabolism on the rates of P. vivax early recurrences, a case-control study was carried out. The cases included patients presenting with an early recurrence (CQ-recurrent individuals), defined as a recurrence during the first 28 days after initial infection and plasma concentrations of CQ plus desethylchloroquine (DCQ; the major CQ metabolite) higher than 100 ng/ml. A control group with no parasite recurrence over the follow-up (the CQ-responsive group) was also included. CQ and DCQ plasma levels were measured on day 28. CQ-metabolizing CYP (CYP2C8, CYP3A4, and CYP3A5) genotypes were determined by real-time PCR. An ex vivo study was conducted to verify the efficacy of CQ and DCQ against P. vivax isolates. The frequency of alleles associated with normal and slow metabolism was similar between the cases and the controls for the CYP2C8 (odds ratio [OR] = 1.45, 95% confidence interval [CI] = 0.51 to 4.14, P = 0.570), CYP3A4 (OR = 2.38, 95% CI = 0.92 to 6.19, P = 0.105), and CYP3A5 (OR = 4.17, 95% CI = 0.79 to 22.04, P = 1.038) genes. DCQ levels were higher than CQ levels, regardless of the genotype. Regarding the DCQ/CQ ratio, there was no difference between groups or between those patients who had a normal genotype and those patients who had a mutant genotype. DCQ and CQ showed similar efficacy ex vivo CYP genotypes had no influence on early recurrence rates. The similar efficacy of CQ and DCQ ex vivo could explain the absence of therapeutic failure, despite the presence of alleles associated with slow metabolism.

Cytochrome P450 (CYP) enzymes are involved in the biotransformation of chloroquine (CQ), but the role of the different profiles of metabolism of this drug in relation to Plasmodium vivax recurrences has not been properly investigated. To investigate the influence of the CYP genotypes associated with CQ metabolism on the rates of P. vivax early recurrences, a case-control study was carried out. The cases included patients presenting with an early recurrence (CQ-recurrent individuals), defined as a recurrence during the first 28 days after initial infection and plasma concentrations of CQ plus desethylchloroquine (DCQ; the major CQ metabolite) higher than 100 ng/ml. A control group with no parasite recurrence over the follow-up (the CQ-responsive group) was also included. CQ and DCQ plasma levels were measured on day 28. CQ-metabolizing CYP (CYP2C8, CYP3A4, and CYP3A5) genotypes were determined by real-time PCR. An ex vivo study was conducted to verify the efficacy of CQ and DCQ against P. vivax isolates. The frequency of alleles associated with normal and slow metabolism was similar between the cases and the controls for the CYP2C8 (odds ratio [OR] = 1.45, 95% confidence interval [CI] = 0.51 to 4.14, P = 0.570), CYP3A4 (OR = 2.38, 95% CI = 0.92 to 6.19, P = 0.105), and CYP3A5 (OR = 4.17, 95% CI = 0.79 to 22.04, P = 1.038) genes. DCQ levels were higher than CQ levels, regardless of the genotype. Regarding the DCQ/CQ ratio, there was no difference between groups or between those patients who had a normal genotype and those patients who had a mutant genotype. DCQ and CQ showed similar efficacy ex vivo. CYP genotypes had no influence on early recurrence rates. The similar efficacy of CQ and DCQ ex vivo could explain the absence of therapeutic failure, despite the presence of alleles associated with slow metabolism.

Background Glucose-6-phosphate dehydrogenase (G6PD) deficiency greatly hinders Plasmodium vivax malaria radical cure and further elimination due to 8-aminoquinolines-associated hemolysis. Although the deleterious health effects of primaquine in G6PD deficient individuals have been known for over 50 years, G6PD testing is not routinely performed before primaquine treatment in most P. vivax endemic areas. Method/Principal findings The qualitative CareStart G6PD screening test was implemented in 12 malaria treatment units (MTUs) in the municipality of Rio Preto da Eva, Western Brazilian Amazon, a malaria endemic area, between February 2019 and early January 2020. Training materials were developed and validated; evaluations were conducted on the effectiveness of training health care professionals (HCPs) to perform the test, the interpretation and reliability of routine testing performed by HCPs, and perceptions of HCPs and patients. Most HCPs were unaware of G6PD deficiency and primaquine-related adverse effects. Most of 110 HCPs trained (86/110, 78%) were able to correctly perform the G6PD test after a single 4-hour training session. The test performed by HCPs during implementation showed 100.0% (4/4) sensitivity and 68.1% (62/91) specificity in identifying G6PD deficient patients as compared to a point-of-care quantitative test (Standard G6PD). Conclusions/Significance G6PD screening using the qualitative CareStart G6PD test performed by HCPs in MTUs of an endemic area showed high sensitivity and concerning low specificity. The amount of false G6PD deficiency detected led to substantial loss of opportunities for radical cure., Author summary Glucose 6-phosphate dehydrogenase deficiency (G6PDd) has greatly impacted the treatment of Plasmodium vivax malaria because of the red blood cell destruction in what is known as hemolysis. Primaquine, used to clear dormant liver parasites that cause relapses of the disease, is a well-known trigger that may lead to life-threatening complications in patients with this condition. Although there are several G6PDd diagnostic tests available to guide the decision of weekly or daily primaquine treatment, they are not yet routinely used: questions on how, when, where and who is going to perform the test remain unanswered. This study revealed that, although G6PDd was not previously known by most of the healthcare workers, they were able to perform the test after a single training session. The test performed well in the field, differentiating patients that cannot use daily primaquine from the others, but some expected limitations require further action to be taken into consideration. This research provides an important overall understanding that may aid policy makers in the process of recommending proven interventions, such as G6PDd screening, to implement them pragmatically.