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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. Ihsan Fadilah and colleagues conduct an individual patient data meta-analysis examining the utility and validity of blood methaemoglobin as a surrogate endpoint for recurrent vivax malaria. Author summary: Why was this study done?: Plasmodium vivax causes recurrent malaria due to dormant liver stage parasites. The pro-drugs primaquine and tafenoquine are the only available treatments to prevent relapsing malaria, but their optimal dosing remains unclear. The active metabolites of primaquine and tafenoquine cause predictable increases in blood methaemoglobin; these same metabolites also kill liver parasites. This suggests that increases in blood methaemoglobin could be used as a population-level surrogate endpoint for liver stage parasite killing. What did the researchers do and find?: We conducted an individual patient data meta-analysis examining the utility and validity of blood methaemoglobin as a surrogate endpoint for recurrent vivax malaria. We systematically reviewed and pooled the available methaemoglobin and clinical data from patients with P. vivax malaria treated with primaquine over the last 20 years. We fit statistical models to these data to quantify the relationship between day 7 methaemoglobin levels and the risk of recurrent P. vivax malaria. We observed an increasing dose-response relationship between weight-adjusted primaquine dose and methaemoglobin levels measured on day 7. We found that for a fixed primaquine dose regimen, a doubling in day 7 methaemoglobin was associated with an estimated 30% reduction in the risk of vivax recurrence. Using day 7 methaemoglobin levels as a surrogate endpoint for recurrent malaria could reduce sample sizes needed in future studies by about 40% and reduce follow-up duration by 94%. What do these findings mean?: Patients with higher methaemoglobin levels a week after starting primaquine treatment appear to have a lower risk of recurrence. Day 7 methaemoglobin levels can potentially be used as a proxy for later vivax recurrence in exploratory trials, making it more efficient (e.g., fewer volunteers and resources required) to determine whether new drugs or regimens are effective. Study limitations include the inability to determine whether P. vivax recurrences are due to treatment failure, reinfection, or relapse. [ABSTRACT FROM AUTHOR]

Background: Treatment guidance for children and older adult patients affected by cutaneous leishmaniasis (CL) is unclear due to limited representation of these groups in clinical trials. Methods: We conducted a collaborative retrospective study to describe the effectiveness and safety of antileishmanial treatments in children ≤ 10 and adults ≥ 60 years of age, treated between 2014 and 2018 in ten CL referral centers in Latin America. Results: 2,037 clinical records were assessed for eligibility. Of them, the main reason for non-inclusion was lack of data on treatment follow-up and therapeutic response (182/242, 75% of children and 179/468, 38% of adults). Data on 1,325 eligible CL patients (736 children and 589 older adults) were analyzed. In both age groups, disease presentation was mild, with a median number of lesions of one (IQR: 1–2) and median lesion diameter of less than 3 cm. Less than 50% of the patients had data for two or more follow-up visits post-treatment (being only 28% in pediatric patients). Systemic antimonials were the most common monotherapy regimen in both age groups (590/736, 80.2% of children and 308/589, 52.3% of older adults) with overall cure rates of 54.6% (95% CI: 50.5–58.6%) and 68.2% (95% CI: 62.6–73.4%), respectively. Other treatments used include miltefosine, amphotericin B, intralesional antimonials, and pentamidine. Adverse reactions related to the main treatment were experienced in 11.9% (86/722) of children versus 38.4% (206/537) of older adults. Most adverse reactions were of mild intensity. Conclusion: Our findings support the need for greater availability and use of alternatives to systemic antimonials, particularly local therapies, and development of strategies to improve patient follow-up across the region, with special attention to pediatric populations. Author summary: Cutaneous leishmaniasis (CL) is a vector-borne disease caused by a parasite called Leishmania that affects more than 1 million people worldwide each year and mostly causes chronic skin ulcers. The first line treatment continues to be systemic antimonials, despite drawbacks such as numerous daily injections and toxic side effects. Some groups, including children and older adults, are usually not included in clinical trials and, therefore, specific treatment guidance for them is uncertain due to the scarcity of robust evidence. RedeLEISH, a Latin American network of leishmaniasis researchers, conducted a retrospective study of the medical records of children ≤ 10 years of age and adults ≥ 60 years of age, treated between 2014 and 2018 in ten CL referral centers in the region, with the aim of learning about and documenting the effectiveness and tolerability of routine antileishmanial treatments, and ultimately contributing to improving the management of these patient groups. Our findings, based on 1,325 eligible patients' records (589 adults ≥ 60 years of age and 736 children ≤ 10 years of age), showed that in both age groups the disease presents predominantly in a mild form, but that the most commonly used treatment, systemic antimonials, has limited effectiveness. This supports and encourages greater use of local therapies, and other treatment alternatives in these special populations. The results of this study also highlight the need to develop strategies for better follow-up of routine CL treatment and determination of effectiveness, as lack of such information was the principal criterion for exclusion of patients screened for eligibility, and less than 50% of eligible patients had information on two or more follow-up visits. [ABSTRACT FROM AUTHOR]

Background: The landscape of malaria transmission in the Peruvian Amazon is temporally and spatially heterogeneous, presenting different micro-geographies with particular epidemiologies. Most cases are asymptomatic and escape routine malaria surveillance based on light microscopy (LM). Following the implementation of control programs in this region, new approaches to stratify transmission and direct efforts at an individual and community level are needed. Antibody responses to serological exposure markers (SEM) to Plasmodium vivax have proven diagnostic performance to identify people exposed in the previous 9 months. Methodology: We measured antibody responses against 8 SEM to identify recently exposed people and determine the transmission dynamics of P. vivax in peri-urban (Iquitos) and riverine (Mazán) communities of Loreto, communities that have seen significant recent reductions in malaria transmission. Socio-demographic, geo-reference, LM and qPCR diagnosis data were collected from two cross-sectional surveys. Spatial and multilevel analyses were implemented to describe the distribution of seropositive cases and the risk factors associated with exposure to P. vivax. Principal findings: Low local transmission was detected by qPCR in both Iquitos (5.3%) and Mazán (2.7%); however, seroprevalence indicated a higher level of (past) exposure to P. vivax in Mazán (56.5%) than Iquitos (38.2%). Age and being male were factors associated with high odds of being seropositive in both sites. Higher antibody levels were found in individuals >15 years old. The persistence of long-lived antibodies in these individuals could overestimate the detection of recent exposure. Antibody levels in younger populations (<15 years old) could be a better indicator of recent exposure to P. vivax. Conclusions: The large number of current and past infections detected by SEMs allows for detailed local epidemiological analyses, in contrast to data from qPCR prevalence surveys which did not produce statistically significant associations. Serological surveillance will be increasingly important in the Peruvian Amazon as malaria transmission is reduced by continued control and elimination efforts. Author summary: A challenge for eliminating Plasmodium vivax is its relapsing nature that can contribute up to 80% of the P. vivax cases. The detection of this source of transmission remains crucial for focused interventions in settings attempting to achieve elimination. Antibody responses to serological exposure markers to P. vivax have been validated to detect recent exposure and likely hypnozoite carriers in low transmission settings. To evaluate the benefits of adding serology to the detection of infection in areas where malaria transmission is rapidly reduced, we measured the prevalence of P. vivax by qPCR and serology in two sites of the Peruvian Amazon. This study shows that antibody responses to P. vivax antigens can detect more recently exposed people than molecular testing, providing more statistical power for surveillance and critical additional epidemiological knowledge for regions attempting to eliminate malaria transmission. [ABSTRACT FROM AUTHOR]

Background: Systemic pentavalent antimonials, mainly meglumine antimoniate, continue to be the first-choice drugs for treatment of cutaneous leishmaniasis (CL) despite their toxicity, difficulty of administration and high cost. In the search for therapeutic alternatives, combining two treatment interventions has emerged as a potential alternative to either reduce the use of antimonials with the associated toxicities, or to increase efficacy. Here, we report the results of a recently completed trial assessing the efficacy and safety of a combination of thermotherapy (TT) plus a short course of miltefosine (MLT) for the treatment of uncomplicated CL in Colombia and Peru. Methods: A multicenter, randomized, evaluator-blinded, phase II, controled clinical trial was conducted. Adult volunteers with a parasitologically confirmed diagnosis of uncomplicated CL were randomly allocated to receive either a single session of TT or a combination of TT plus a short course of MLT (3 weeks). Therapeutic response outcomes and safety were assessed. Results: 130 subjects were included in the study, of whom 64 were randomly assigned to the TT arm and 66 to the TT + MLT arm. Cure at 3 months' follow-up was achieved in 57.8% (n = 37) and 80.3% (n = 53) in the TT and TT + MLT groups, respectively, in the intention to treat analysis. The TT + MLT regimen was better that TT alone (p = 0.0055). The presence of vesicles at the site of heat application was the most common adverse event reported associated with the use of TT; while vomiting (31.8%) and elevation of liver enzymes (28.8%) were the most frequent adverse events reported associated with the use of MLT. Conclusion: The combination of TT plus a short course of MLT was shown to be significantly better than TT alone for the treatment of uncomplicated CL in the New World. Trial registration: Registered in clinicaltrials.gov NCT02687971. Author summary: Cutaneous leishmaniasis is a disease caused by Leishmania parasites transmitted by sandflies. It is a complex and neglected disease. Generally, it manifests as ulcers, nodules, or other lesions, mainly on the face and extremities. An estimated 0.7–1 million new cases occur annually in around 88 countries in South and Central America, Africa, Europe, the Middle East, and Asia, with the appearance of new outbreaks in different regions of the world. For more than 70 years, pentavalent antimonials have been the first line treatment, despite their toxicity, varying efficacy and need of administration of painful injections for 20 days or more. There is limited availability of effective and safe treatment alternatives and the strength of scientific evidence on the efficacy and safety of the different schemes is variable, limiting the control of the disease from a therapeutic approach. Hence the need for effective, safe, low-cost treatment alternatives with non-invasive routes of administration. This article reports the results of a clinical trial aiming to evaluate a combined therapy with two therapeutic alternatives (miltefosine and thermotherapy) approved for the treatment of cutaneous leishmaniasis, comparing it with thermotherapy alone. In this study, the combined therapy was significantly better than the monotherapy in managing cutaneous leishmaniasis. [ABSTRACT FROM AUTHOR]

American Tegumentary Leishmaniasis (ATL) is an endemic and neglected disease of South America. Here, mucosal leishmaniasis (ML) disproportionately affects up to 20% of subjects with current or previous localised cutaneous leishmaniasis (LCL). Preclinical and clinical reports have implicated the Leishmania RNA virus-1 (LRV1) as a possible determinant of progression to ML and other severe manifestations such as extensive cutaneous and mucosal disease and treatment failure and relapse. However, these associations were not consistently found in other observational studies and are exclusively based on cross-sectional designs. In the present study, 56 subjects with confirmed ATL were assessed and followed out for 24-months post-treatment. Lesion biopsy specimens were processed for molecular detection and quantification of Leishmania parasites, species identification, and LRV1 detection. Among individuals presenting LRV1 positive lesions, 40% harboured metastatic phenotypes; comparatively 58.1% of patients with LRV1 negative lesions harboured metastatic phenotypes (p = 0.299). We found treatment failure (p = 0.575) and frequency of severe metastatic phenotypes (p = 0.667) to be similarly independent of the LRV1. Parasite loads did not differ according to the LRV1 status (p = 0.330), nor did Leishmanin skin induration size (p = 0.907) or histopathologic patterns (p = 0.780). This study did not find clinical, parasitological, or immunological evidence supporting the hypothesis that LRV1 is a significant determinant of the pathobiology of ATL. Author summary: The Leishmania RNA virus-1 (LRV1) has been implicated as a possible modulator agent in the pathogenesis of leishmaniasis. In-vivo and in-vitro studies have depicted specific mechanisms of how LRV1 could lead to metastasis. Clinical studies and epidemiological evidence have both supported and rejected the hypothesis that LRV1 is a relevant determinant of progression, treatment failure and clinical severity of American Tegumentary Leishmaniasis (ATL). This lack of consistency between preclinical and clinical reports requires further longitudinal studies to clarify the role of LRV1 in ATL. Due to the complex nature of ATL, as other frequent human diseases, these studies should tackle multiple determinants of pathogenicity, including LRV1 status, parasite features, immune status, and prevalent comorbidities affecting individuals in endemic settings. Also, critical methodological aspects allowing for the reliable identification and quantification of LRV1 should be guaranteed. [ABSTRACT FROM AUTHOR]

Background: Antibody responses as serological markers of Plasmodium vivax infection have been shown to correlate with exposure, but little is known about the other factors that affect antibody responses in naturally infected people from endemic settings. To address this question, we studied IgG responses to novel serological exposure markers (SEMs) of P. vivax in three settings with different transmission intensity. Methodology: We validated a panel of 34 SEMs in a Peruvian cohort with up to three years' longitudinal follow-up using a multiplex platform and compared results to data from cohorts in Thailand and Brazil. Linear regression models were used to characterize the association between antibody responses and age, the number of detected blood-stage infections during follow-up, and time since previous infection. Receiver Operating Characteristic (ROC) analysis was used to test the performance of SEMs to identify P. vivax infections in the previous 9 months. Principal findings: Antibody titers were associated with age, the number of blood-stage infections, and time since previous P. vivax infection in all three study sites. The association between antibody titers and time since previous P. vivax infection was stronger in the low transmission settings of Thailand and Brazil compared to the higher transmission setting in Peru. Of the SEMs tested, antibody responses to RBP2b had the highest performance for classifying recent exposure in all sites, with area under the ROC curve (AUC) = 0.83 in Thailand, AUC = 0.79 in Brazil, and AUC = 0.68 in Peru. Conclusions: In low transmission settings, P. vivax SEMs can accurately identify individuals with recent blood-stage infections. In higher transmission settings, the accuracy of this approach diminishes substantially. We recommend using P. vivax SEMs in low transmission settings pursuing malaria elimination, but they are likely to be less effective in high transmission settings focused on malaria control. Author summary: Plasmodium vivax still poses a threat in many countries due to its ability to cause recurrent infections. Key to achieving the goal of malaria elimination is the ability to quickly detect and treat carriers of relapsing parasites. Failing to identify this transmission reservoir will hinder progress towards malaria elimination. Recently, novel serological markers of recent exposure to P. vivax (SEM) have been developed and validated in low transmission settings. It is still poorly understood what factors affect the antibody response to these markers when evaluated in contrasting endemic contexts. To determine the factors that influence the antibody response to SEM, we compared the antibody levels in three sites with different transmission intensity: Thailand (low), Brazil (moderate) and Peru (high). In this study, we found that transmission intensity plays a key role in the acquisition of the antibody repertoire to P. vivax. In highly endemic sites, it is likely that immunological memory resulting from a constant and sustained exposure will impact the performance of SEMs to detect individuals with recent exposure to P. vivax. In summary, SEMs that perform well in low transmission sites do not perform as well in high transmission regions. [ABSTRACT FROM AUTHOR]

Despite efforts made over decades by the Peruvian government to eliminate malaria, Plasmodium vivax remains a challenge for public health decision-makers in the country. The uneven distribution of its incidence, plus its complex pattern of dispersion, has made ineffective control measures based on global information that lack the necessary detail to understand transmission fully. In this sense, population genetic tools can complement current surveillance. This study describes the genetic diversity and population structure from September 2012 to March 2015 in three geographically distant settlements, Cahuide (CAH), Lupuna (LUP) and Santa Emilia (STE), located in the Peruvian Amazon. A total 777 P. vivax mono-infections, out of 3264, were genotyped. Among study areas, LUP showed 19.7% of polyclonal infections, and its genetic diversity (Hexp) was 0.544. Temporal analysis showed a significant increment of polyclonal infections and Hexp, and the introduction and persistence of a new parasite population since March 2013. In STE, 40.1% of infections were polyclonal, with Hexp = 0.596. The presence of four genetic clusters without signals of clonal expansion and infections with lower parasite densities compared against the other two areas were also found. At least four parasite populations were present in CAH in 2012, where, after June 2014, malaria cases decreased from 213 to 61, concomitant with a decrease in polyclonal infections (from 0.286 to 0.18), and expectedly variable Hexp. Strong signals of gene flow were present in the study areas and wide geographic distribution of highly diverse parasite populations were found. This study suggests that movement of malaria parasites by human reservoirs connects geographically distant malaria transmission areas in the Peruvian Amazon. The maintenance of high levels of parasite genetic diversity through human mobility is a critical barrier to malaria elimination in this region. Author summary: Plasmodium vivax transmission is heterogeneous and discontinuous in the Peruvian Amazon. Such heterogeneity is the result of factors that include, but are not restricted to, the environment, public policies, and characteristics of the parasite, the vector, and human activities. All these factors make P. vivax transmission resilient to interventions. In order to achieve the goals of control and local elimination, P. vivax surveillance must inform how those factors sustain disease transmission in order to focalize and synchronize control strategies. In this study, we implemented molecular surveillance complemented with population genetic tools in the areas of Cahuide, Lupuna, and Santa Emilia located in the Peruvian Amazon. In particular, we characterize the transmission and the parasite genetic variation in these sites from September 2012 to March 2015. The changes in parasite diversity, the wide geographic dispersion of parasite subpopulation and the introduction of a new parasite clone or subpopulation in Lupuna documented in this study suggest that connectivity among the different endemic areas, likely due to human mobility, sustains disease transmission in the region hindering the success of control measures. This information must be considered in the design of current control strategies. [ABSTRACT FROM AUTHOR]

Background: Climate change is affecting food systems globally, with implications for food security, nutrition, and the health of human populations. There are limited data characterizing the current and future consequences of climate change on local food security for populations already experiencing poor nutritional indicators. Indigenous Amazonian populations have a high reported prevalence of nutritional deficiencies. This paper characterizes the food system of the Shawi of the Peruvian Amazon, climatic and non-climatic drivers of their food security vulnerability to climate change, and identifies potential maladaptation trajectories. Methods and findings: Semi-structured interviews with key informants (n = 24), three photovoice workshops (n = 17 individuals), transect walks (n = 2), a food calendar exercise, and two community dissemination meetings (n = 30 individuals), were conducted within two Shawi communities in Balsapuerto District in the Peruvian Loreto region between June and September of 2014. The Shawi food system was based on three main food sub-systems (forest, farming and externally-sourced). Shawi reported collective, gendered, and emotional notions related to their food system activities. Climatic and non-climatic drivers of food security vulnerability among Shawi participants acted at proximal and distal levels, and mutually reinforced key maladaptation trajectories, including: 1) a growing population and natural resource degradation coupled with limited opportunities to increase incomes, and 2) a desire for education and deforestation reinforced by governmental social and food interventions. Conclusion: A series of maladaptive trajectories have the potential to increase social and nutritional inequities for the Shawi. Transformational food security adaptation should include consideration of Indigenous perceptions and priorities, and should be part of Peruvian food and socioeconomic development policies. [ABSTRACT FROM AUTHOR]

Background: Tegumentary leishmaniasis (TL) is a disease of skin and/or mucosal tissues caused by Leishmania parasites. TL patients may concurrently carry other pathogens, which may influence the clinical outcome of TL. Methodology and principal findings: This review focuses on the frequency of TL coinfections in human populations, interactions between Leishmania and other pathogens in animal models and human subjects, and implications of TL coinfections for clinical practice. For the purpose of this review, TL is defined as all forms of cutaneous (localised, disseminated, or diffuse) and mucocutaneous leishmaniasis. Human immunodeficiency virus (HIV) coinfection, superinfection with skin bacteria, and skin manifestations of visceral leishmaniasis are not included. We searched MEDLINE and other databases and included 73 records: 21 experimental studies in animals and 52 studies about human subjects (mainly cross-sectional and case studies). Several reports describe the frequency of Trypanosoma cruzi coinfection in TL patients in Argentina (about 41%) and the frequency of helminthiasis in TL patients in Brazil (15% to 88%). Different hypotheses have been explored about mechanisms of interaction between different microorganisms, but no clear answers emerge. Such interactions may involve innate immunity coupled with regulatory networks that affect quality and quantity of acquired immune responses. Diagnostic problems may occur when concurrent infections cause similar lesions (e.g., TL and leprosy), when different pathogens are present in the same lesions (e.g., Leishmania and Sporothrix schenckii), or when similarities between phylogenetically close pathogens affect accuracy of diagnostic tests (e.g., serology for leishmaniasis and Chagas disease). Some coinfections (e.g., helminthiasis) appear to reduce the effectiveness of antileishmanial treatment, and drug combinations may cause cumulative adverse effects. Conclusions and significance: In patients with TL, coinfection is frequent, it can lead to diagnostic errors and delays, and it can influence the effectiveness and safety of treatment. More research is needed to unravel how coinfections interfere with the pathogenesis of TL. [ABSTRACT FROM AUTHOR]

Introduction: Progress with the treatment of cutaneous leishmaniasis (CL) has been hampered by inconsistent methodologies used to assess treatment effects. A sizable number of trials conducted over the years has generated only weak evidence backing current treatment recommendations, as shown by systematic reviews on old-world and new-world CL (OWCL and NWCL). Materials and methods: Using a previously published guidance paper on CL treatment trial methodology as the reference, consensus was sought on key parameters including core eligibility and outcome measures, among OWCL (7 countries, 10 trial sites) and NWCL (7 countries, 11 trial sites) during two separate meetings. Results: Findings and level of consensus within and between OWCL and NWCL sites are presented and discussed. In addition, CL trial site characteristics and capacities are summarized. Conclusions: The consensus reached allows standardization of future clinical research across OWCL and NWCL sites. We encourage CL researchers to adopt and adapt as required the proposed parameters and outcomes in their future trials and provide feedback on their experience. The expertise afforded between the two sets of clinical sites provides the basis for a powerful consortium with potential for extensive, standardized assessment of interventions for CL and faster approval of candidate treatments. [ABSTRACT FROM AUTHOR]

Malaria transmission requires that Anopheles mosquitoes ingest Plasmodium gametocyte stages circulating in the human bloodstream. In the context of malaria elimination, understanding the epidemiology of gametocytes relative to all Plasmodium infections and the contribution of asymptomatic and sub-microscopic parasite carriers to the gametocyte reservoir is necessary, especially in low endemic settings with predominance of P.vivax. A 13-month longitudinal study was conducted in two communities (n = 1935 individuals) of Loreto Department, Peru, with five active screenings for Plasmodium infections and gametocyte stages by quantitative real-time PCR (qPCR) and reverse transcription (RT)-qPCR, respectively. Parasite prevalence by qPCR was 7.2% for P.vivax (n = 520/7235; range by survey 6.0%-8.1%) and 3.2% for P.falciparum (n = 235/7235; range by survey 0.4%-7.7%). Sub-microscopic infections accounted for 73.5% of P.vivax (range by survey 60%-89%) and almost the totality of P.falciparum cases. Gametocytes were found in 28.4% P.vivax infections (range by survey 18.7%-34.1%), with a peak of 61.5% in one community at the start of the transmission season. About 59.8% of all P.vivax gametocyte carriers were asymptomatic and 31.9% were sub-microscopic. Age patterns for gametocyte prevalence paralleled asexual stage infections and peaked among 15–25 year old individuals. Asexual parasite density was found to be the strongest predictor for P.vivax gametocyte presence in longitudinal multivariate analysis (odds ratio 2.33 [95% confidence interval 1.96, 2.78]; P<0.001). Despite significant differences in seasonality patterns and P.vivax prevalence found at the local scale, sub-microscopic and asymptomatic infections predominate and contribute significantly to the gametocyte reservoir in different communities of the Peruvian Amazon. Control and elimination campaigns need sensitive tools to detect all infections that escape routine malaria surveillance, which may contribute to maintain transmission in the region. [ABSTRACT FROM AUTHOR]

Background: Characterizing the parasite dynamics and population structure provides useful information to understand the dynamic of transmission and to better target control interventions. Despite considerable efforts for its control, vivax malaria remains a major health problem in Peru. In this study, we have explored the population genetics of Plasmodium vivax isolates from Iquitos, the main city in the Peruvian Amazon, and 25 neighbouring peri-urban as well as rural villages along the Iquitos-Nauta Road. Methodology/ Results: From April to December 2008, 292 P. vivax isolates were collected and successfully genotyped using 14 neutral microsatellites. Analysis of the molecular data revealed a similar proportion of monoclonal and polyclonal infections in urban areas, while in rural areas monoclonal infections were predominant (p = 0.002). Multiplicity of infection was higher in urban (MOI = 1.5–2) compared to rural areas (MOI = 1) (p = 0.003). The level of genetic diversity was similar in all areas (He = 0.66–0.76, p = 0.32) though genetic differentiation between areas was substantial (PHIPT = 0.17, p<0.0001). Principal coordinate analysis showed a marked differentiation between parasites from urban and rural areas. Linkage disequilibrium was detected in all the areas ( = 0.08–0.49, for all p<0.0001). Gene flow among the areas was stablished through Bayesian analysis of migration models. Recent bottleneck events were detected in 4 areas and a recent parasite expansion in one of the isolated areas. In total, 87 unique haplotypes grouped in 2 or 3 genetic clusters described a sub-structured parasite population. Conclusion/Significance: Our study shows a sub-structured parasite population with clonal propagation, with most of its components recently affected by bottleneck events. Iquitos city is the main source of parasite spreading for all the peripheral study areas. The routes of transmission and gene flow and the reduction of the parasite population described are important from the public health perspective as well for the formulation of future control policies. [ABSTRACT FROM AUTHOR]

Background: With low and markedly seasonal malaria transmission, increasingly sensitive tools for better stratifying the risk of infection and targeting control interventions are needed. A cross-sectional survey to characterize the current malaria transmission patterns, identify hotspots, and detect recent changes using parasitological and serological measures was conducted in three sites of the Peruvian Amazon. Material and Methods: After full census of the study population, 651 participants were interviewed, clinically examined and had a blood sample taken for the detection of malaria parasites (microscopy and PCR) and antibodies against P. vivax (PvMSP119, PvAMA1) and P. falciparum (PfGLURP, PfAMA1) antigens by ELISA. Risk factors for malaria infection (positive PCR) and malaria exposure (seropositivity) were assessed by multivariate survey logistic regression models. Age-specific seroprevalence was analyzed using a reversible catalytic conversion model based on maximum likelihood for generating seroconversion rates (SCR, λ). SaTScan was used to detect spatial clusters of serology-positive individuals within each site. Results: The overall parasite prevalence by PCR was low, i.e. 3.9% for P. vivax and 6.7% for P. falciparum, while the seroprevalence was substantially higher, 33.6% for P. vivax and 22.0% for P. falciparum, with major differences between study sites. Age and location (site) were significantly associated with P. vivax exposure; while location, age and outdoor occupation were associated with P. falciparum exposure. P. falciparum seroprevalence curves showed a stable transmission throughout time, while for P. vivax transmission was better described by a model with two SCRs. The spatial analysis identified well-defined clusters of P. falciparum seropositive individuals in two sites, while it detected only a very small cluster of P. vivax exposure. Conclusion: The use of a single parasitological and serological malaria survey has proven to be an efficient and accurate method to characterize the species specific heterogeneity in malaria transmission at micro-geographical level as well as to identify recent changes in transmission. [ABSTRACT FROM AUTHOR]

Background: Cutaneous leishmaniasis (CL) is a skin disease caused by the protozoan parasite Leishmania. Few studies have assessed the influence of the sample collection site within the ulcer and the sampling method on the sensitivity of parasitological and molecular diagnostic techniques for CL. Sensitivity of the technique can be dependent upon the load and distribution of Leishmania amastigotes in the lesion. Methodology/Principal Findings: We applied a quantitative real-time PCR (qPCR) assay for Leishmania (Viannia) minicircle kinetoplast DNA (kDNA) detection and parasite load quantification in biopsy and scraping samples obtained from 3 sites within each ulcer (border, base, and center) as well as in cytology brush specimens taken from the ulcer base and center. A total of 248 lesion samples from 31 patients with laboratory confirmed CL of recent onset (≤3 months) were evaluated. The kDNA-qPCR detected Leishmania DNA in 97.6% (242/248) of the examined samples. Median parasite loads were significantly higher in the ulcer base and center than in the border in biopsies (P<0.0001) and scrapings (P = 0.0002). There was no significant difference in parasite load between the ulcer base and center (P = 0.80, 0.43, and 0.07 for biopsy, scraping, and cytology brush specimens, respectively). The parasite load varied significantly by sampling method: in the ulcer base and center, the descending order for the parasite load levels in samples was: cytology brushes, scrapings, and biopsies (P<0.0001); in the ulcer border, scrapings had higher parasite load than biopsies (P<0.0001). There was no difference in parasite load according to L. braziliensis and L. peruviana infections (P = 0.4). Conclusion/Significance: Our results suggest an uneven distribution of Leishmania amastigotes in acute CL ulcers, with higher parasite loads in the ulcer base and center, which has implications for bedside collection of diagnostic specimens. The use of scrapings and cytology brushes is recommended instead of the more invasive biopsy. [ABSTRACT FROM AUTHOR]

Thermotherapy is an accepted alternative therapy for new-world cutaneous leishmaniasis, but current heat-delivery modalities are too costly to be made widely available to endemic populations. We adapted a low-cost heat pack named the HECT-CL device that delivers safe, reliable, and renewable conduction heat. 25 patients with cutaneous leishmaniasis completed treatment with the device at an initial temperature of 52°C±2°C for 3 minutes to each lesion, repeated daily for 7 days, and were followed up for 6 months by direct observation. The overall definitive clinical cure rate was 60%. Concurrently, 13 patients meeting minimally significant exclusion criteria received identical compassionate use treatment with a cumulative definitive cure rate of 68.4%, 75% for those who had experienced CL relapse after prior antimonial treatment. Therapy was well tolerated. Reversible second-degree burns occurred in two patients and no bacterial super-infections were observed. HECT-CL is a promising treatment and deserves further study to verify its safety and efficacy as adjuvant and mono- therapy. Author Summary: American cutaneous leishmaniasis is an endemic parasitic disease in Peru, with more than a reported thousand cases per year associated with significant disabilities and economic impact. In comparison to old-world cutaneous Leishmaniasis where the infection generally heals on its own, American cutaneous Leishmaniasis is a chronic skin disease and its treatment can be expensive, difficult to administer, and associated with significant adverse events. We developed a heat treatment using a novel and low-technology device named Hand-held Exothermic Crystallization Thermotherapy for Cutaneous Leishmaniasis (HECT-CL). This device produces a stable thermal reaction ranging from 50–54°C which is considered therapeutic and has enormous benefits for its application in real world conditions, as it is extremely low cost, simple to use, and highly tolerable in patients receiving this treatment. Our pilot evaluation found cure rates close to those reported with the standard antimonial treatment and likely higher in cases of relapsing disease. Additionally, fewer severe adverse events were observed than reported with the use of other currently available heat therapies. Our objective is to show that HECT-CL therapy could be instituted in rural and real situations with limited health care infrastructure and with less cost and fewer side effects than standard antimonial treatment. [ABSTRACT FROM AUTHOR]

In South America, various species of Leishmania are endemic and cause New World tegumentary leishmaniasis (NWTL). The correct identification of these species is critical for adequate clinical management and surveillance activities. We developed a real-time polymerase chain reaction (PCR) assay and evaluated its diagnostic performance using 64 archived parasite isolates and 192 prospectively identified samples collected from individuals with suspected leishmaniasis enrolled at two reference clinics in Lima, Peru. The real-time PCR assay was able to detect a single parasite and provided unambiguous melting peaks for five Leishmania species of the Viannia subgenus that are highly prevalent in South America: L. (V.) braziliensis, L. (V.) panamensis, L. (V.) guyanensis, L. (V.) peruviana and L. (V.) lainsoni. Using kinetoplastid DNA-based PCR as a gold standard, the real-time PCR had sensitivity and specificity values of 92% and 77%, respectively, which were significantly higher than those of conventional tests such as microscopy, culture and the leishmanin skin test (LST). In addition, the real-time PCR identified 147 different clinical samples at the species level, providing an overall agreement of 100% when compared to multilocus sequence typing (MLST) data performed on a subset of these samples. Furthermore, the real-time PCR was three times faster and five times less expensive when compared to PCR - MLST for species identification from clinical specimens. In summary, this new assay represents a cost-effective and reliable alternative for the identification of the main species causing NWTL in South America. Author Summary: Leishmaniasis is a neglected disease with more than two million new human infections annually worldwide. Tegumentary leishmaniasis, cutaneous and mucocutaneous, is mainly caused by five Leishmania species of the Viannia complex in South America. Different species can cause disease with similar symptoms but have dissimilar prognoses and may need different therapeutic regimens. Identification of Leishmania species traditionally relies on the multilocus enzyme electrophoresis (MLEE) assay, but it can only be applied to culture-positive samples and takes at least six weeks of intense laboratory work. A reliable and rapid assay for species identification can be a valuable tool. Molecular assays are the fastest and most accurate way to identify the etiological agents causing leishmaniasis. This paper describes a novel real-time PCR assay for identification of the five main species that cause tegumentary leishmaniasis in the New World. The assay correctly identified each of these five species of Leishmania directly from clinical samples. Because of its reliability, speed and simplicity, this assay could be used for species identification in routine laboratory diagnosis of leishmaniasis in endemic regions. [ABSTRACT FROM AUTHOR]

Background: Traditional methods of detecting Leishmania from cutaneous lesions involve invasive diagnostic procedures, such as scrapings, which cause discomfort, require technical expertise, and carry risks of invasive procedures. We compared the performance of 2 novel, molecular-based non-invasive methods for the diagnosis of cutaneous leishmaniasis (CL). Methods: Consecutive patients presenting to the Leishmania Clinic at the Hospital Nacional Cayetano Heredia were enrolled. PCR was performed on filter paper lesion impressions (FPLIs), cytology brushes, and lancets for detection of Leishmania DNA. Smears from lesion scrapings and leishmanin skin test were also performed. Outcome measures were sensitivity and specificity. Composite reference standard was any 2 of 5 tests positive. Species identification was performed by PCR assays of positive specimens. Results: Ninety patients with 129 lesions were enrolled, 117 of which fulfilled reference criteria for a diagnosis of CL. Of these 117 lesions, 113 were positive by PCR of lancets used for lesion scrapings versus 116 by PCR of FPLIs (p = 0.930) or 116 by PCR of cytology brushes (p = 0.930). Sensitivity and specificity of PCR on lancets were 96.6% [95% CI 93.3-99.9%] and 100%, respectively. Sensitivity and specificity of FPLI PCR were 99.1% [95% CI 97.4-100%] and 100%, respectively. Sensitivity and specificity of cytology brush PCR were 99.1% [95% CI 97.4-100%] and 100%, respectively. Giemsa-stained lesion smear and leishmanin skin test had inferior sensitivities at 47.9% [95% CI 38.9-57.0%] and 82.3% [95% CI 73.9-90.7%], respectively, compared to PCR of invasive or non-invasive specimens (p<0.001). Conclusions: Cytology brush PCR constitutes a sensitive and specific alternative to traditional diagnostic assays performed on invasive specimens such as lesion scrapings. It performs comparatively to non-invasive FPLI PCR. This novel, rapid, and well-tolerated method has the potential for widespread use in the field and in pediatric populations where traditional specimen collection is difficult. [ABSTRACT FROM AUTHOR]

Background: Increased rates for failure in leishmaniasis antimony treatment have been recently recognized worldwide. Although several risk factors have been identified there is no clinical score to predict antimony therapy failure of cutaneous leishmaniasis. Methods: A case control study was conducted in Peru from 2001 to 2004. 171 patients were treated with pentavalent antimony and followed up to at least 6 months to determine cure or failure. Only patients with ulcerative cutaneous leishmaniasis (N = 87) were considered for data analysis. Epidemiological, demographical, clinical and laboratory data were analyzed to identify risk factors for treatment failure. Two prognostic scores for antimonial treatment failure were tested for sensitivity and specificity to predict antimony therapy failure by comparison with treatment outcome. Results: Among 87 antimony-treated patients, 18 (21%) failed the treatment and 69 (79%) were cured. A novel risk factor for treatment failure was identified: presence of concomitant distant lesions. Patients presenting concomitant-distant lesions showed a 30.5-fold increase in the risk of treatment failure compared to other patients. The best prognostic score for antimonial treatment failure showed a sensitivity of 77.78% and specificity of 95.52% to predict antimony therapy failure. Conclusions: A prognostic score including a novel risk factor was able to predict antimonial treatment failure in cutaneous leishmaniasis with high specificity and sensitivity. This prognostic score presents practical advantages as it relies on clinical and epidemiological characteristics, easily obtained by physicians or health workers, and makes it a promising clinical tool that needs to be validated before their use for developing countries. Author Summary: The manuscript is relevant because of the finding of a new risk factor for chemotherapy failure and the development of a prognosis score for cutaneous leishmaniasis. The proportion of patients that have multiple lesions in American Tegumentary Leishmaniasis (ATL) is considerable. Publications and our experience permit to estimate that they represent around 20% of the affected population from the Amazon basin with cutaneous lesions. In addition, about 1/3 of them would correspond to the concomitant distant lesions category, the novel risk factor identified with a very high odds ratio (20–30) associated. Such numbers merit study of concomitant distant ulcers category on its own, not only because of clinical management implications, but also to search for factors that are contributing to chemotherapy failure. Finally, the simple equation proposed in the manuscript can be easily adapted to smart phone technologies. Similar prognosis equations are scarce for other pathologies and do not exist for Cutaneous Leishmaniasis at all. The simplicity of this tool should be followed by subsequent epidemiologic studies in other ATL endemic regions. [ABSTRACT FROM AUTHOR]

Background: Traditional methods of diagnosing mucosal leishmaniasis (ML), such as biopsy with histopathology, are insensitive and require collection of an invasive diagnostic specimen. Methods: We compared standard invasive procedures including biopsy histopathology, biopsy PCR, and leishmanin skin test (LST) to a novel, non-invasive, cytology-brush based PCR for the diagnosis of ML in Lima, Peru. Consensus reference standard was 2/4 tests positive, and outcome measures were sensitivity and specificity. Leishmania species identification was performed by PCR-based assays of positive specimens. Results: Twenty-eight patients were enrolled, 23 of whom fulfilled criteria for a diagnosis of ML. Sensitivity and specificity of biopsy with histopathology were 21.7% [95% CI 4.9-38.5%] and 100%; 69.6% [95% CI 50.8-88.4%] and 100% for LST; 95.7% [95% CI 87.4-100%] and 100% for biopsy PCR; and 95.7% [95% CI 87.4-100%] and 90% [95% CI 71.4-100%] for cytology brush PCR using both CervisoftH and HistobrushH cervical cytology brushes. Represented species identified by PCR-RFLP included: L. (V). braziliensis (n = 4), and L. (V). peruviana (n = 3). Conclusions: Use of commercial grade cytology brush PCR for diagnosis of ML is sensitive, rapid, well tolerated, and carries none of the risks of invasive diagnostic procedures such as biopsy. Further optimization is required for adequate species identification. Further evaluation of this method in field and other settings is warranted. [ABSTRACT FROM AUTHOR]

Background: Most of the Leishmania genome is reported to be constitutively expressed during the life cycle of the parasite, with a few regulated genes. Inter-species comparative transcriptomics evidenced a low number of species-specific differences related to differentially distributed genes or the differential regulation of conserved genes. It is of uppermost importance to ensure that the observed differences are indeed species-specific and not simply specific of the strains selected for representing the species. The relevance of this concern is illustrated by current study. Methodology/Principal Findings: We selected 5 clinical isolates of L. braziliensis characterized by their diversity of clinical and in vitro phenotypes. Real-time quantitative PCR was performed on promastigote and amastigote life stages to assess gene expression profiles at seven time points covering the whole life cycle. We tested 12 genes encoding proteins with roles in transport, thiol-based redox metabolism, cellular reduction, RNA poly(A)-tail metabolism, cytoskeleton function and ribosomal function. The general trend of expression profiles showed that regulation of gene expression essentially occurs around the stationary phase of promastigotes. However, the genes involved in this phenomenon appeared to vary significantly among the isolates considered. Conclusion/Significance: Our results clearly illustrate the unique character of each isolate in terms of gene expression dynamics. Results obtained on an individual strain are not necessarily representative of a given species. Therefore, extreme care should be taken when comparing the profiles of different species and extrapolating functional differences between them. Author Summary: Leishmania is a group of parasites (Protozoa, Trypanosomatidae) responsible for a wide spectrum of clinical forms. Among the factors explaining this phenotypic polymorphism, parasite features are important contributors. One approach to identify them consists in characterizing the gene expression profiles throughout the life cycle. In a recent study, the transcriptome of 3 Leishmania species was compared and this revealed species-specific differences, albeit in a low number. A key issue, however, is to ensure that the observed differences are indeed species-specific and not specific of the strains selected for representing the species. In order to illustrate the relevance of this concern, we analyzed here the gene expression profiles of 5 clinical isolates of L. braziliensis at seven time points of the life cycle. Our results clearly illustrate the unique character of each isolate in terms of gene expression dynamics: one Leishmania strain is not necessarily representative of a given species. [ABSTRACT FROM AUTHOR]

Background: There is an increasing body of literature reporting treatment failure of the currently recommended radical treatment of Plasmodium vivax infections. As P. vivax is the main malaria species outside the African continent, emerging tolerance to its radical treatment regime could have major consequences in countries like Peru, where 80% of malaria cases are due to P. vivax. Here we describe the results of a 1-year longitudinal follow up of 51 confirmed P. vivax patients living around Iquitos, Peruvian Amazon, and treated according to the Peruvian national guidelines. Methodology: Each month a blood sample for microscopy and later genotyping was systematically collected. Recent exposure to infection was estimated by detecting antibodies against the P. vivax circumsporozoite protein (CSP) and all PCR confirmed P. vivax infections were genotyped with 16 polymorphic microsatellites. Results: During a 1-year period, 84 recurrent infections, 22 positive also by microscopy, were identified, with a median survival time to first recurrent infection of 203 days. Most of them (71%) were asymptomatic; in 13 patients the infection persisted undetected by microscopy for several consecutive months. The genotype of mostly recurrent infections differed from that at day 0 while fewer differences were seen between the recurrent infections. The average expected heterozygosity was 0.56. There was strong linkage disequilibrium (IA s = 0.29, p<1.10-4) that remained also when analyzing only the unique haplotypes, suggesting common inbreeding. Conclusion: In Peru, the P. vivax recurrent infections were common and displayed a high turnover of parasite genotypes compared to day 0. Plasmodium vivax patients, even when treated according to the national guidelines, may still represent an important parasite reservoir that can maintain transmission. Any elimination effort should consider such a hidden reservoir. [ABSTRACT FROM AUTHOR]

Background: Current therapies for cutaneous leishmaniasis are limited by poor efficacy, long-term course of treatment, and the development of resistance. We evaluated if pentavalent antimony (an anti-parasitic drug) combined with imiquimod (an immunomodulator) was more effective than pentavalent antimony alone in patients who had not previously been treated. Methods: A randomized double-blind clinical trial involving 80 cutaneous leishmaniasis patients was conducted in Peru. The study subjects were recruited in Lima and Cusco (20 experimental and 20 control subjects at each site). Experimental arm: Standard dose of pentavalent antimony plus 5% imiquimod cream applied to each lesion three times per week for 20 days. Control arm: Standard dose of pentavalent antimony plus placebo (vehicle cream) applied as above. The primary outcome was cure defined as complete re-epithelization with no inflammation assessed during the 12 months post-treatment period. Results: Of the 80 subjects enrolled, 75 completed the study. The overall cure rate at the 12-month follow-up for the intention-to-treat analysis was 75% (30/40) in the experimental arm and 58% (23/40) in the control arm (p = 0.098). Subgroup analyses suggested that combination treatment benefits were most often observed at the Cusco site, where L. braziliensis is the prevalent species. Over the study period, only one adverse event (rash) was recorded, in the experimental arm. Conclusion: The combination treatment of imiquimod plus pentavalent antimony performed better than placebo plus pentavalent antimony, but the difference was not statistically significant. Trial Registration: Clinical Trials.gov NCT00257530 Author Summary: Neglected tropical diseases (NTDs) are a group of tropical infections including trypanosomiasis, filariasis, schistosomiasis, onchocerciasis, leishmaniasis and other such diseases of poverty. Of the classic neglected diseases, leishmaniasis has among the highest level of morbidity and mortality. Infection with Leishmania parasites causes severe disease in humans, including fatal visceral leishmaniasis and cutaneous leishmaniasis resulting in severe scarring, often in the face. This is a difficult infection to treat because the current therapies are generally poorly effective. The present study carried out a placebo-controlled, double-blinded study to investigated whether a combined therapy with imiquimod plus pentavalent antimony was superior to the standard therapy of pentavalent antimony alone as a first-line treatment for cutaneous leishmaniasis in Peru. A higher cure rate with the combination therapy was observed, but could not be conclusively proven. [ABSTRACT FROM AUTHOR]

Background: While coverage of long-lasting insecticide-treated nets (LLIN) has steadily increased, a growing number of studies report gaps between net ownership and use. We conducted a mixed-methods social science study assessing the importance of net preference and use after Olyset® LLINs were distributed through a mass campaign in rural communities surrounding Iquitos, the capital city of the Amazonian region of Peru., Methods: The study was conducted in the catchment area of the Paujil and Cahuide Health Centres (San Juan district) between July 2007 and November 2008. During a first qualitative phase, participant observation and in-depth interviews collected information on key determinants for net preference and use. In a second quantitative phase, a survey among recently confirmed malaria patients evaluated the acceptability and use of both LLINs and traditional nets, and a case control study assessed the association between net preference/use and housing structure (open vs. closed houses)., Results: A total of 10 communities were selected for the anthropological fieldwork and 228 households participated in the quantitative studies. In the study area, bed nets are considered part of the housing structure and are therefore required to fulfil specific architectural and social functions, such as providing privacy and shelter, which the newly distributed Olyset® LLINs ultimately did not. The LLINs' failure to meet these criteria could mainly be attributed to their large mesh size, transparency and perceived ineffectiveness to protect against mosquitoes and other insects, resulting in 63.3% of households not using any of the distributed LLINs. Notably, LLIN usage was significantly lower in houses with no interior or exterior walls (35.2%) than in those with walls (73.8%) (OR = 5.2, 95CI [2.2; 12.3], p<0.001)., Conclusion: Net preference can interfere with optimal LLIN use. In order to improve the number of effective days of LLIN protection per dollar spent, appropriate quantitative and qualitative methods for collecting information on net preference should be developed before any LLIN procurement decision is made.

Background: Multi-drug resistant falciparum malaria is an important health problem in the Peruvian Amazon region. We carried out a randomised open label clinical trial comparing mefloquine-artesunate, the current first line treatment in this region, with dihydroartemisinin-piperaquine., Methods and Findings: Between July 2003 and July 2005, 522 patients with P. falciparum uncomplicated malaria were recruited, randomized (260 with mefloquine-artesunate and 262 with dihydroartemisinin-piperaquine), treated and followed up for 63 days. PCR-adjusted adequate clinical and parasitological response, estimated by Kaplan Meier survival and Per Protocol analysis, was extremely high for both drugs (99.6% for mefloquine-artesunate and 98.4% and for dihydroartemisinin-piperaquine) (RR: 0.99, 95%CI [0.97-1.01], Fisher Exact p = 0.21). All recrudescences were late parasitological failures. Overall, gametocytes were cleared faster in the mefloquine-artesunate group (28 vs 35 days) and new gametocytes tended to appear more frequently in patients treated with dihydroartemisinin-piperaquine (day 7: 8 (3.6%) vs 2 (0.9%), RR: 3.84, 95%CI [0.82-17.87]). Adverse events such as anxiety and insomnia were significantly more frequent in the mefloquine-artesunate group, both in adults and children., Conclusion: Dihydroartemisinin-piperaquine is as effective as mefloquine-artesunate in treating uncomplicated P. falciparum malaria but it is better tolerated and more affordable than mefloquine-artesunate (US$1.0 versus US$18.65 on the local market). Therefore, it should be considered as a potential candidate for the first line treatment of P. falciparum malaria in Peru., Trial Registration: ClinicalTrials.gov NCT00373607.