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Malaria-infected individuals often harbor mixtures of genetically distinct parasite genotypes. We studied intra-host dynamics of parasite genotypes co-infecting asymptomatic adults in an area of intense malaria transmission in Chikhwawa, Malawi. Serial blood samples (5 ml) were collected over seven consecutive days from 25 adults with asymptomatic Plasmodium falciparum malaria and analyzed to determine whether a single peripheral blood sample accurately captures within-host parasite diversity. Blood samples from three of the participants were also analyzed by limiting dilution cloning and SNP genotyping of the parasite clones isolated to examine both the number and relatedness of co-infecting parasite haplotypes. We observed rapid turnover of co-infecting parasite genotypes in 88% of the individuals sampled (n = 22) such that the genetic composition of parasites infecting these individuals changed dramatically over the course of seven days of follow up. Nineteen of the 25 individuals sampled (76%) carried multiple parasite genotypes at baseline. Analysis of serial blood samples from three of the individuals revealed that they harbored 6, 12 and 17 distinct parasite haplotypes respectively. Approximately 70% of parasite haplotypes recovered from the three extensively sampled individuals were unrelated (proportion of shared alleles <83.3%) and were deemed to have primarily arisen from superinfection (inoculation of unrelated parasite haplotypes through multiple mosquito bites). The rest were related at the half-sib level or greater and were deemed to have been inoculated into individual human hosts via parasite co-transmission from single mosquito bites. These findings add further to the growing weight of evidence indicating that a single blood sample poorly captures within-host parasite diversity and underscore the importance of repeated blood sampling to accurately capture within-host parasite ecology. Our data also demonstrate a more pronounced role for parasite co-transmission in generating within-host parasite diversity in high transmission settings than previously assumed. Taken together, these findings have important implications for understanding the evolution of drug resistance, malaria transmission, parasite virulence, allocation of gametocyte sex ratios and acquisition of malaria immunity., (Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.)

Genotyping of allelic variants of Plasmodium falciparum merozoite surface proteins 1 and 2 ( msp-1 and msp-2 ), and the glutamate-rich protein is the gold standard for distinguishing reinfections from recrudescences in antimalarial drug trials. We compared performance of the recently developed 24-single-nucleotide polymorphism (SNP) Barcoding Assay against msp-1 and msp-2 genotyping in a cluster-randomized effectiveness trial of artemether-lumefantrine and dihydroartemisinin-piperaquine in Malawi. Rates of recrudescence and reinfection estimated by the two methods did not differ significantly (Fisher's exact test; P = 0.887 and P = 0.768, respectively). There was a strong agreement between the two methods in predicting treatment outcomes and resolving the genetic complexity of malaria infections in this setting. These results support the use of this SNP assay as an alternative method for correcting antimalarial efficacy/effectiveness data.

Single-cell genomics is a powerful tool for determining the genetic architecture of complex communities of unicellular organisms. In areas of high transmission, malaria patients are often challenged by the activities of multiple Plasmodium falciparum lineages, which can potentiate pathology, spread drug resistance loci, and also complicate most genetic analysis. Single-cell sequencing of P. falciparum would be key to understanding infection complexity, though efforts are hampered by the extreme nucleotide composition of its genome (∼80% AT-rich). To counter the low coverage achieved in previous studies, we targeted DNA-rich late-stage parasites by Fluorescence-Activated Cell Sorting and whole genome sequencing. Our method routinely generates accurate, near-complete capture of the 23 Mb P. falciparum genome (mean breadth of coverage 90.7%) at high efficiency. Data from 48 single-cell genomes derived from a polyclonal infection sampled in Chikhwawa, Malawi allowed for unambiguous determination of haplotype diversity and recent meiotic events, information that will aid public health efforts., (© The Author(s) 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

Context:  Heat injury is a significant threat to military trainees. Different methods of heat mitigation are in use across military units. Mist fans are 1 of several methods used in the hot and humid climate of Fort Benning, Georgia., Objectives:  To determine if (1) the mist fan or the cooling towel effectively lowered participant core temperature in the humid environment found at Fort Benning and (2) the mist fan or the cooling towel presented additional physiologic or safety benefits or detriments when used in this environment., Design:  Randomized controlled clinical trial., Setting:  Laboratory environmental chamber., Patients or Other Participants:  Thirty-five physically active men aged 19 to 35 years., Intervention(s):  (1) Mist fan, (2) commercial cooling towel, (3) passive-cooling (no intervention) control. All treatments lasted 20 minutes. Participants ran on a treadmill at 60% V̇o 2 max., Main Outcome Measure(s):  Rectal core temperature, heart rate, thermal comfort, perceived temperature, perceived wetness, and blood pressure., Results:  Average core temperature increased during 20 minutes of cooling (F 1,28 = 64.76, P < .001, η p 2 = 0.70), regardless of group (F 1,28 = 3.41, P = .08, η p 2 = 0.11) or condition (F 1,28 < 1.0). Core temperature, heart rate, and blood pressure did not differ among the 3 conditions. Perceived temperature during 20 minutes of cooling decreased (F 1,30 = 141.19, P < .001, η p 2 = 0.83) regardless of group or condition. Perceived temperature was lower with the mist-fan treatment than with the control treatment (F 1,15 = 7.38, P = .02, η p 2 = 0.32). The mist-fan group perceived themselves to be cooler even at elevated core temperatures., Conclusions:  The mist fan and cooling towel were both ineffective at lowering core temperature. Core temperature continued to increase after exercise in all groups. The mist fan produced feelings of coolness while the core temperature remained elevated, possibly increasing the risk of heat illness.