Precision symptom phenotyping identifies early clinical and proteomic predictors of distinct COVID-19 sequelae.

BACKGROUND: Post-COVID conditions (PCC) are difficult to characterize, diagnose, predict, and treat due to overlapping symptoms and poorly understood pathology. Identifying inflammatory profiles may improve clinical prognostication and trial endpoints. METHODS: 1,988 SARS-CoV-2 positive U.S. Military Health System beneficiaries with quantitative post-COVID symptom scores were included in this analysis. Among participants who reported moderate-to-severe symptoms on surveys collected 6-months post-SARS-CoV-2 infection, principal component analysis (PCA) followed by K-means clustering identified distinct clusters of symptoms. RESULTS: Three symptom-based clusters were identified: a sensory cluster (loss of smell and/or taste), a fatigue/difficulty thinking cluster, and a difficulty breathing/exercise intolerance cluster. Individuals within the sensory cluster were all outpatients during their initial COVID-19 presentation. The difficulty breathing cluster had a higher likelihood of obesity and COVID-19 hospitalization compared to those with no/mild symptoms at 6-months post-infection. Multinomial regression linked early post-infection D-dimer and IL-1RA elevation to fatigue/difficulty thinking, and elevated ICAM-1 concentrations to sensory symptoms. CONCLUSIONS: We identified three distinct symptom-based PCC phenotypes with specific clinical risk factors and early post-infection inflammatory predictors. With further validation and characterization, this framework may allow more precise classification of PCC cases and potentially improve the diagnosis, prognostication, and treatment of PCC.

Time measurement in insect photoperiodism: external and internal coincidence.

The identity and nature of the photoperiodic photoreceptors are now quite well known, as is the nature of the endocrine regulation of the resulting diapauses. The central problem of time measurement-how the photoperiodic clock differentiates long from short days-however, is still obscure, known only from whole-animal experiments and abstract models, although it is clearly a function of the insect circadian system. This review describes some of these experiments in terms of oscillator entrainment and two widely applicable photoperiodic clock models, external and internal coincidence, mainly using data from experiments on flesh flies (Sarcophaga spp) and the parasitic wasp, Nasonia vitripennis.

In vitro activity of rhinacanthin analogues against drug resistant Plasmodium falciparum isolates from Northeast Thailand.

BACKGROUND: New anti-malarial drugs are needed urgently to address the increasing challenges of drug-resistant falciparum malaria. Two rhinacanthin analogues containing a naphthoquinone moiety resembling atovaquone showed promising in-vitro activity against a P. falciparum laboratory reference strain (K1). The anti-malarial activity of these 2 compounds was further evaluated for P. falciparum field isolates from an area of multi-drug resistance in Northeast Thailand. METHODS: Using a pLDH enzyme-linked immunosorbent assay, four P. falciparum isolates from Northeast Thailand in 2018 were tested for in vitro sensitivity to the two synthetic rhinacanthin analogues 1 and 2 as well as established anti-malarials. Mutations in the P. falciparum cytochrome b gene, a marker for atovaquone (ATQ) resistance, were genotyped in all four field isolates as well as 100 other clinical isolates from the same area using PCR-artificial Restriction Fragment Length Polymorphisms. Pfkelch13 mutations, a marker for artemisinin (ART) resistance, were also examined in all isolates. RESULTS: The 50% inhibitory concentrations (IC50) of P. falciparum field isolates for rhinacanthin analogue 1 was 321.9-791.1 nM (median = 403.1 nM). Parasites were more sensitive to analogue 2: IC50 48.6-63.3 nM (median = 52.2 nM). Similar results were obtained against P. falciparum reference laboratory strains 3D7 and W2. The ART-resistant IPC-5202 laboratory strain was more sensitive to these compounds with a median IC50 45.9 and 3.3 nM for rhinacanthin analogues 1 and 2, respectively. The ATQ-resistant C2B laboratory strain showed high-grade resistance towards both compounds (IC50 > 15,000 nM), and there was a strong positive correlation between the IC50 values for these compounds and ATQ (r = 0.83-0.97, P < 0.001). There were no P. falciparum cytochrome b mutations observed in the field isolates, indicating that P. falciparum isolates from this area remained ATQ-sensitive. Pfkelch13 mutations and the ring-stage survival assay confirmed that most isolates were resistant to ART. CONCLUSIONS: Two rhinacanthin analogues showed parasiticidal activity against multi-drug resistant P. falciparum isolates, although less potent than ATQ. Rhinacanthin analogue 2 was more potent than analogue 1, and can be a lead compound for further optimization as an anti-malarial in areas with multidrug resistance.

Undiagnosing St Joan: She Does Not Need a Medical or Psychiatric Diagnosis.

ABSTRACT: This article traces the history of Joan of Arc through her brief life that includes leading an army in defense of France at the age of 17 and ending with her death at the stake at the age of 19. In her activities, St Joan reported that she was guided by voices and visions in which she communicated with venerated spiritual figures such as St Michael and St Margaret. Questions have arisen about the nature of these experiences, and various medical and psychiatric diagnoses have been offered by contemporary experts. In our effort to evaluate the diagnostic proposals, we have examined the incidence of voices and visions in the Middle Ages, and we have followed that with a review of nonpathologic voice-hearing in our own era. We then move on to an analysis of some proposed medical and psychiatric diagnoses, all of which we find unconvincing. With this background, we argue that St Joan does not warrant a medical or psychiatric diagnosis. Such a conclusion, however, leaves us with another issue, that of Joan's achievements. How do we understand an adolescent being able to lead an army? Addressing this question proves more difficult than deciding whether St Joan warrants a diagnosis. In addition to her achievements in the war against Britain, Joan of Arc stands out as both the most documented person in Western civilization up until her era, and as the only person who has been both condemned and canonized by the Catholic Church.

Measurements of 5,6-Orthoquinone, a Surrogate for the Presumed Active Primaquine Metabolite 5-Hydroxyprimaquine, in the Urine of Cambodian Adults.

The active metabolites of primaquine, in particular 5-hydroxyprimaquine, likely responsible for the clearance of dormant hypnozoites, are produced through the hepatic CYP450 2D6 (CYP2D6) enzymatic pathway. With the inherent instability of 5-hydroxyprimaquine, a stable surrogate, 5,6-orthoquinone, can now be detected and measured in the urine as part of primaquine pharmacokinetic studies. This study performed CYP450 2D6 genotyping and primaquine pharmacokinetic testing, to include urine 5,6-orthoquinone, in 27 healthy adult Cambodians, as a preliminary step to prepare for future clinical studies assessing primaquine efficacy for Plasmodium vivax infections. The CYP2D6 *10 reduced activity allele was found in 57% of volunteers, and the CYP2D6 genotypes were dominated by *1/*10 (33%) and *10/*10 (30%). Predicted phenotypes were evenly split between Normal Metabolizer (NM) and Intermediate Metabolizer (IM) except for one volunteer with a gene duplication and unclear phenotype, classifying as either IM or NM. Median plasma primaquine (PQ) area under the curve (AUC) was lower in the NM group (460 h*ng/mL) compared to the IM group (561 h*ng/mL), although not statistically significant. Similar to what has been found in the US study, no 5,6-orthoquinone was detected in the plasma. The urine creatinine-corrected 5,6-orthoquinone AUC in the NM group was almost three times higher than in the IM group, with peak measurements (Tmax) at 4 h. Although there is variation among individuals, future studies examining the relationship between the levels of urine 5,6-orthoquinone and primaquine radical cure efficacy could result in a metabolism biomarker predictive of radical cure.

Single-Dose Tafenoquine to Prevent Relapse of Plasmodium vivax Malaria.

BACKGROUND: Treatment of Plasmodium vivax malaria requires the clearing of asexual parasites, but relapse can be prevented only if dormant hypnozoites are cleared from the liver (a treatment termed "radical cure"). Tafenoquine is a single-dose 8-aminoquinoline that has recently been registered for the radical cure of P. vivax. METHODS: This multicenter, double-blind, double-dummy, parallel group, randomized, placebo-controlled trial was conducted in Ethiopia, Peru, Brazil, Cambodia, Thailand, and the Philippines. We enrolled 522 patients with microscopically confirmed P. vivax infection (>100 to <100,000 parasites per microliter) and normal glucose-6-phosphate dehydrogenase (G6PD) activity (with normal activity defined as ≥70% of the median value determined at each trial site among 36 healthy male volunteers who were otherwise not involved in the trial). All patients received a 3-day course of chloroquine (total dose of 1500 mg). In addition, patients were assigned to receive a single 300-mg dose of tafenoquine on day 1 or 2 (260 patients), placebo (133 patients), or a 15-mg dose of primaquine once daily for 14 days (129 patients). The primary outcome was the Kaplan-Meier estimated percentage of patients who were free from recurrence at 6 months, defined as P. vivax clearance without recurrent parasitemia. RESULTS: In the intention-to-treat population, the percentage of patients who were free from recurrence at 6 months was 62.4% in the tafenoquine group (95% confidence interval [CI], 54.9 to 69.0), 27.7% in the placebo group (95% CI, 19.6 to 36.6), and 69.6% in the primaquine group (95% CI, 60.2 to 77.1). The hazard ratio for the risk of recurrence was 0.30 (95% CI, 0.22 to 0.40) with tafenoquine as compared with placebo (P<0.001) and 0.26 (95% CI, 0.18 to 0.39) with primaquine as compared with placebo (P<0.001). Tafenoquine was associated with asymptomatic declines in hemoglobin levels, which resolved without intervention. CONCLUSIONS: Single-dose tafenoquine resulted in a significantly lower risk of P. vivax recurrence than placebo in patients with phenotypically normal G6PD activity. (Funded by GlaxoSmithKline and Medicines for Malaria Venture; DETECTIVE ClinicalTrials.gov number, NCT01376167 .).

Efficacy of three anti-malarial regimens for uncomplicated Plasmodium falciparum malaria in Cambodia, 2009-2011: a randomized controlled trial and brief review.

BACKGROUND: Anti-malarial resistance remains an important public health challenge in Cambodia. The effectiveness of three therapies for uncomplicated falciparum malaria was evaluated in Oddar Meanchey province in Northern Cambodia from 2009 to 2011. METHODS: In this randomized, open-label, parallel group-controlled trial, 211 subjects at least 5 years old with uncomplicated falciparum malaria were treated with 3 days of directly observed therapy: 63 received artesunate-mefloquine (AS/MQ), 77 received dihydroartemisinin-piperaquine (DHA/PPQ), and 71 received atovaquone-proguanil (ATQ/PG). The subjects were followed for 42 days or until recurrent parasitaemia. Genotyping of msp1, msp2, and glurp among individual parasite isolates distinguished recrudescence from reinfection. Pfmdr1 copy number was measured by real-time PCR and half-maximal parasite inhibitory concentrations (IC50) were measured in vitro by 48-h isotopic hypoxanthine incorporation assay. RESULTS: The per-protocol PCR-adjusted efficacy (95% confidence interval) at 42 days was 80.6% (70.8-90.5%) for AS/MQ, 97.2% (93.3-100%) for DHA/PPQ, and 92.9% (86.1-99.6%) for ATQ/PG. On day 3, 57.9% remained parasitaemic in the AS/MQ and DHA/PPQ arms. At baseline, 46.9% had microscopic Plasmodium falciparum gametocytaemia. Both recurrences in the DHA/PPQ arm lost Pfmdr1 copy number amplification at recrudescence. All four recurrences in the ATQ/PG arm were wild-type for cytochrome bc1. One subject withdrew from the ATQ/PG arm due to drug allergy. CONCLUSIONS: This study was conducted at the epicentre of substantial multi-drug resistance that emerged soon thereafter. Occurring early in the national transition from AS/MQ to DHA/PPQ, both DHA/PPQ and ATQ/PG had acceptable efficacy against uncomplicated falciparum malaria. However, efficacy of AS/MQ was only 80% with apparent mefloquine resistance based on elevated Pfmdr1 copy number and IC50. By 2009, there was already significant evidence of artemisinin resistance not previously reported at the Northern Cambodia-Thai border. This study suggests the basis for early development of significant DHA/PPQ failures within 3 years of introduction. Artemisinin resistance likely occurred on the Northern border concurrently with that reported along the Western border in Pailin. Trial registration This legacy trial was conducted prior to International Committee of Medical Journal Editors' requirements for preregistration on ClinicalTrials.gov. The full protocol has been provided.

Distribution and Temporal Dynamics of Plasmodium falciparum Chloroquine Resistance Transporter Mutations Associated With Piperaquine Resistance in Northern Cambodia.

BACKGROUND: Newly emerged mutations within the Plasmodium falciparum chloroquine resistance transporter (PfCRT) can confer piperaquine resistance in the absence of amplified plasmepsin II (pfpm2). In this study, we estimated the prevalence of co-circulating piperaquine resistance mutations in P. falciparum isolates collected in northern Cambodia from 2009 to 2017. METHODS: The sequence of pfcrt was determined for 410 P. falciparum isolates using PacBio amplicon sequencing or whole genome sequencing. Quantitative polymerase chain reaction was used to estimate pfpm2 and pfmdr1 copy number. RESULTS: Newly emerged PfCRT mutations increased in prevalence after the change to dihydroartemisinin-piperaquine in 2010, with >98% of parasites harboring these mutations by 2017. After 2014, the prevalence of PfCRT F145I declined, being outcompeted by parasites with less resistant, but more fit PfCRT alleles. After the change to artesunate-mefloquine, the prevalence of parasites with amplified pfpm2 decreased, with nearly half of piperaquine-resistant PfCRT mutants having single-copy pfpm2. CONCLUSIONS: The large proportion of PfCRT mutants that lack pfpm2 amplification emphasizes the importance of including PfCRT mutations as part of molecular surveillance for piperaquine resistance in this region. Likewise, it is critical to monitor for amplified pfmdr1 in these PfCRT mutants, as increased mefloquine pressure could lead to mutants resistant to both drugs.

Selection of Cytochrome b Mutants Is Rare among Plasmodium falciparum Patients Failing Treatment with Atovaquone-Proguanil in Cambodia.

Atovaquone-proguanil remains effective against multidrug-resistant Plasmodium falciparum in Southeast Asia, but resistance is mediated by a single point mutation in cytochrome b (cytb) that can arise during treatment. Among 14 atovaquone-proguanil treatment failures in a clinical trial in Cambodia, only one recrudescence harbored the cytb mutation Y268C. Deep sequencing did not detect the mutation at baseline or in the first 3 days of treatment, suggesting that it arose de novo Further sequencing across cytb similarly found no low-frequency cytb mutations that were up-selected from baseline to recrudescence. Copy number amplification in dihydroorotate dehydrogenase (DHODH) and cytb as markers of atovaquone tolerance was also absent. Cytb mutation played a minor role in atovaquone-proguanil treatment failures in an active comparator clinical trial.

Regenerative rehabilitation of catastrophic extremity injury in military conflicts and a review of recent developmental efforts.

AIM OF THE REVIEW: This review aims to describe the current state of regenerative rehabilitation of severe military extremity injuries, and promising new therapies on the horizon. DISCUSSION: The nature of warfare is rapidly shifting with information operations, autonomous weapons, and the threat of full-scale peer adversary conflicts threatening to create contested environments with delayed medical evacuation to definitive care. More destructive weapons will lead to more devastating injuries, creating new challenges for limb repair and restoration. Current paradigms of delayed rehabilitation following initial stabilization, damage control surgery, and prolonged antibiotic therapy will need to shift. Advances in regenerative medicine technologies offer the possibility of treatment along the continuum of care. Regenerative rehabilitation will begin at the point of injury and require a holistic, organ-systems approach. CONCLUSIONS: Both technological improvements and a rapidly advancing understanding of injury pathophysiology will contribute to improved limb-salvage outcomes, and shift the calculus away from early limb amputation.

Determining the Temperature Dependency of Biodegradation Kinetics for 34 Hydrocarbons while Avoiding Chemical and Microbial Confounding Factors.

Biodegradation kinetics data are keystone for evaluating the environmental persistence and risk of chemicals. Biodegradation kinetics depend highly on the prevailing temperature, which influences microbial community structures, metabolic rates, and chemical availability. There is a lack of high-quality comparative biodegradation kinetics data that are determined at different test temperatures but with the same microbial inoculum and chemical availability. The present study was designed to determine the effect of test temperature on the biodegradation kinetics of hydrocarbons while avoiding confounding factors. We used inocula from a Northern river (2.7 °C) and a Central European river (12.5 °C). Aqueous stock solutions containing 45 individual hydrocarbons were generated by passive dosing and added to river water containing the native microorganisms. Compound-specific biodegradation kinetics were then determined at 2.7, 12, and 20 °C based on substrate depletion. Main findings comprise the following: (1) Degradation half-times (DegT50) of 34 test chemicals were determined at different test temperatures and were largely consistent with the Arrhenius equation (activation energy, 65.4 kJ/mol). (2) Differences in biodegradation kinetics between tested isomers were rather limited. (3) The recent lowering of standard test temperature from 20 to 12 °C results typically in a doubling of DegT50 values and can lead to a stricter persistency assessment.

An Exploration of Sedentary Behavior Patterns in Community-Dwelling People With Stroke: A Cluster-Based Analysis.

BACKGROUND AND PURPOSE: Long periods of daily sedentary time, particularly accumulated in long uninterrupted bouts, are a risk factor for cardiovascular disease. People with stroke are at high risk of recurrent events and prolonged sedentary time may increase this risk. We aimed to explore how people with stroke distribute their periods of sedentary behavior, which factors influence this distribution, and whether sedentary behavior clusters can be distinguished? METHODS: This was a secondary analysis of original accelerometry data from adults with stroke living in the community. We conducted data-driven clustering analyses to identify unique accumulation patterns of sedentary time across participants, followed by multinomial logistical regression to determine the association between the clusters, and the total amount of sedentary time, age, gender, body mass index (BMI), walking speed, and wake time. RESULTS: Participants in the highest quartile of total sedentary time accumulated a significantly higher proportion of their sedentary time in prolonged bouts (P < 0.001). Six unique accumulation patterns were identified, all of which were characterized by high sedentary time. Total sedentary time, age, gender, BMI, and walking speed were significantly associated with the probability of a person being in a specific accumulation pattern cluster, P < 0.001 - P = 0.002. DISCUSSION AND CONCLUSIONS: Although unique accumulation patterns were identified, there is not just one accumulation pattern for high sedentary time. This suggests that interventions to reduce sedentary time must be individually tailored.Video Abstract available for more insight from the authors (see the Video Supplemental Digital Content 1, available at: http://links.lww.com/JNPT/A343).

Interventions for reducing sedentary behaviour in people with stroke.

BACKGROUND: Stroke survivors are often physically inactive as well as sedentary,and may sit for long periods of time each day. This increases cardiometabolic risk and has impacts on physical and other functions. Interventions to reduce or interrupt periods of sedentary time, as well as to increase physical activity after stroke, could reduce the risk of secondary cardiovascular events and mortality during life after stroke. OBJECTIVES: To determine whether interventions designed to reduce sedentary behaviour after stroke, or interventions with the potential to do so, can reduce the risk of death or secondary vascular events, modify cardiovascular risk, and reduce sedentary behaviour. SEARCH METHODS: In December 2019, we searched the Cochrane Stroke Trials Register, CENTRAL, MEDLINE, Embase, CINAHL, PsycINFO, Conference Proceedings Citation Index, and PEDro. We also searched registers of ongoing trials, screened reference lists, and contacted experts in the field. SELECTION CRITERIA: Randomised trials comparing interventions to reduce sedentary time with usual care, no intervention, or waiting-list control, attention control, sham intervention or adjunct intervention. We also included interventions intended to fragment or interrupt periods of sedentary behaviour. DATA COLLECTION AND ANALYSIS: Two review authors independently selected studies and performed 'Risk of bias' assessments. We analyzed data using random-effects meta-analyses and assessed the certainty of the evidence with the GRADE approach. MAIN RESULTS: We included 10 studies with 753 people with stroke. Five studies used physical activity interventions, four studies used a multicomponent lifestyle intervention, and one study used an intervention to reduce and interrupt sedentary behaviour. In all studies, the risk of bias was high or unclear in two or more domains. Nine studies had high risk of bias in at least one domain. The interventions did not increase or reduce deaths (risk difference (RD) 0.00, 95% confidence interval (CI) -0.02 to 0.03; 10 studies, 753 participants; low-certainty evidence), the incidence of recurrent cardiovascular or cerebrovascular events (RD -0.01, 95% CI -0.04 to 0.01; 10 studies, 753 participants; low-certainty evidence), the incidence of falls (and injuries) (RD 0.00, 95% CI -0.02 to 0.02; 10 studies, 753 participants; low-certainty evidence), or incidence of other adverse events (moderate-certainty evidence). Interventions did not increase or reduce the amount of sedentary behaviour time (mean difference (MD) +0.13 hours/day, 95% CI -0.42 to 0.68; 7 studies, 300 participants; very low-certainty evidence). There were too few data to examine effects on patterns of sedentary behaviour. The effect of interventions on cardiometabolic risk factors allowed very limited meta-analysis. AUTHORS' CONCLUSIONS: Sedentary behaviour research in stroke seems important, yet the evidence is currently incomplete, and we found no evidence for beneficial effects. Current World Health Organization (WHO) guidelines recommend reducing the amount of sedentary time in people with disabilities, in general. The evidence is currently not strong enough to guide practice on how best to reduce sedentariness specifically in people with stroke. More high-quality randomised trials are needed, particularly involving participants with mobility limitations. Trials should include longer-term interventions specifically targeted at reducing time spent sedentary, risk factor outcomes, objective measures of sedentary behaviour (and physical activity), and long-term follow-up.

Differential Association of Spirituality and Religiosity With Rumination: Implications for the Treatment of Depression.

ABSTRACT: Recent studies have shown that religiosity (R) is associated with lower rates of depression, whereas spirituality (S) is associated with higher rates. Rumination has also been associated with higher rates of depression. Some have hypothesized that rumination mediates the differential association of religiosity and spirituality with depression. We empirically test this hypothesis in a longitudinal, multigenerational sample through associations between rumination and depression, R/S and depression, and R/S and rumination. Cross-sectionally, total rumination scores were predicted by spirituality (standardized ß = 0.13; 95% confidence interval [CI], 0.00-0.26), with subscale (reflection, depression, and brooding) standardized betas ranging from 0.11 to 0.15 (95% CI, -0.03 to -0.29). Cross-sectionally, rumination was not predicted by religiosity. Longitudinally, and consistent with previous findings, religiosity, but not spirituality, predicted reduced depressive symptoms (standardized ß = -0.3; 95% CI, -0.58 to -0.01). The association between spirituality and rumination was driven by millennials. Psychotherapies that target rumination for depression might therefore be especially effective in the millennial demographic.

Implementation and evaluation of a novel, unofficial, trainee-organized hospital addiction medicine consultation service.

Background: To evaluate a novel, unofficial, trainee-organized, hospital addiction medicine consultation service (AMCS), we aimed to assess whether it was (1) acceptable to hospital providers and patients, (2) feasible to organize and deliver, and (3) impacted patient care. Methods: We performed a retrospective descriptive study of all AMCS consultations over the first 16 months. We determined acceptability via the number of referrals received from admitting services, and the proportion of referred patients who consented to consultation. We evaluated feasibility via continuation/growth of the service over time, and the proportion of referrals successfully completed before hospital discharge. As most referrals related to opioid use disorder, we determined impact through the proportion of eligible patients offered and initiated on opioid agonist therapy (OAT) in hospital, and the proportion of patients who filled their outpatient prescription or attended their first visit with their outpatient OAT prescriber. Results: The unofficial AMCS grew to involve six hospital-based residents and five supervising community-based addiction physicians. The service received 59 referrals, primarily related to injection opioid use, for 50 unique patients from 12 different admitting services. 90% of patients were seen before discharge, and 98% agreed to addiction medicine consultation. Among 34 patients with active moderate-severe opioid use disorder who were not already on OAT, 82% initiated OAT in hospital and 89% of these patients continued after discharge. Conclusions: Established in response to identified gaps in patient care and learning opportunities, a novel, unofficial, trainee-organized AMCS was acceptable, feasible, and positively impacted patient care over the first 16 months. This trainee-organized, unofficial AMCS could be used as a model for other hospitals that do not yet have an official AMCS.

Efficient Transmission of Mixed Plasmodium falciparum/vivax Infections From Humans to Mosquitoes.

BACKGROUND: In Southeast Asia, people are often coinfected with different species of malaria (Plasmodium falciparum [Pf] and Plasmodium vivax [Pv]) as well as with multiple clones of the same species. Whether particular species or clones within mixed infections are more readily transmitted to mosquitoes remains unknown. METHODS: Laboratory-reared Anopheles dirus were fed on blood from 119 Pf-infected Cambodian adults, with 5950 dissected to evaluate for transmitted infection. Among 12 persons who infected mosquitoes, polymerase chain reaction and amplicon deep sequencing were used to track species and clone-specific transmission to mosquitoes. RESULTS: Seven of 12 persons that infected mosquitoes harbored mixed Pf/Pv infection. Among these 7 persons, all transmitted Pv with 2 transmitting both Pf and Pv, leading to Pf/Pv coinfection in 21% of infected mosquitoes. Up to 4 clones of each species were detected within persons. Shifts in clone frequency were detected during transmission. However, in general, all parasite clones in humans were transmitted to mosquitoes, with individual mosquitoes frequently carrying multiple transmitted clones. CONCLUSIONS: Malaria diversity in human hosts was maintained in the parasite populations recovered from mosquitoes fed on their blood. However, in persons with mixed Pf/Pv malaria, Pv appears to be transmitted more readily, in association with more prevalent patent gametocytemia.

Dormancy, Diapause, and the Role of the Circadian System in Insect Photoperiodism.

Whole-animal experiments devised to investigate possible association between photoperiodic time measurement and the circadian system (Bünning's hypothesis) are compared with more recent molecular investigations of circadian clock genes. In Sarcophaga argyrostoma and some other species, experimental cycles of light and darkness revealed a photoperiodic oscillator, set to constant phase at dusk and measuring night length repeatedly during extended periods of darkness. In some species, however, extreme dampening revealed an unrepetitive (i.e., hourglass-like) response. Rhythms of clock gene transcript abundance may also show similar phase relationships to the light cycle, and gene silencing of important clock genes indicates that they play a crucial role in photoperiodism either alone or in concert. However, the multiplicity of peripheral oscillators in the insect circadian system indicates that more complex mechanisms might also be important.

Correction: Quantification of glucose-6-phosphate dehydrogenase activity by spectrophotometry: A systematic review and meta-analysis.

[This corrects the article DOI: 10.1371/journal.pmed.1003084.].

Quantification of glucose-6-phosphate dehydrogenase activity by spectrophotometry: A systematic review and meta-analysis.

BACKGROUND: The radical cure of Plasmodium vivax and P. ovale requires treatment with primaquine or tafenoquine to clear dormant liver stages. Either drug can induce haemolysis in individuals with glucose-6-phosphate dehydrogenase (G6PD) deficiency, necessitating screening. The reference diagnostic method for G6PD activity is ultraviolet (UV) spectrophotometry; however, a universal G6PD activity threshold above which these drugs can be safely administered is not yet defined. Our study aimed to quantify assay-based variation in G6PD spectrophotometry and to explore the diagnostic implications of applying a universal threshold. METHODS AND FINDINGS: Individual-level data were pooled from studies that used G6PD spectrophotometry. Studies were identified via PubMed search (25 April 2018) and unpublished contributions from contacted authors (PROSPERO: CRD42019121414). Studies were excluded if they assessed only individuals with known haematological conditions, were family studies, or had insufficient details. Studies of malaria patients were included but analysed separately. Included studies were assessed for risk of bias using an adapted form of the Quality Assessment of Diagnostic Accuracy Studies-2 (QUADAS-2) tool. Repeatability and intra- and interlaboratory variability in G6PD activity measurements were compared between studies and pooled across the dataset. A universal threshold for G6PD deficiency was derived, and its diagnostic performance was compared to site-specific thresholds. Study participants (n = 15,811) were aged between 0 and 86 years, and 44.4% (7,083) were women. Median (range) activity of G6PD normal (G6PDn) control samples was 10.0 U/g Hb (6.3-14.0) for the Trinity assay and 8.3 U/g Hb (6.8-15.6) for the Randox assay. G6PD activity distributions varied significantly between studies. For the 13 studies that used the Trinity assay, the adjusted male median (AMM; a standardised metric of 100% G6PD activity) varied from 5.7 to 12.6 U/g Hb (p < 0.001). Assay precision varied between laboratories, as assessed by variance in control measurements (from 0.1 to 1.5 U/g Hb; p < 0.001) and study-wise mean coefficient of variation (CV) of replicate measures (from 1.6% to 14.9%; p < 0.001). A universal threshold of 100% G6PD activity was defined as 9.4 U/g Hb, yielding diagnostic thresholds of 6.6 U/g Hb (70% activity) and 2.8 U/g Hb (30% activity). These thresholds diagnosed individuals with less than 30% G6PD activity with study-wise sensitivity from 89% (95% CI: 81%-94%) to 100% (95% CI: 96%-100%) and specificity from 96% (95% CI: 89%-99%) to 100% (100%-100%). However, when considering intermediate deficiency (<70% G6PD activity), sensitivity fell to a minimum of 64% (95% CI: 52%-75%) and specificity to 35% (95% CI: 24%-46%). Our ability to identify underlying factors associated with study-level heterogeneity was limited by the lack of availability of covariate data and diverse study contexts and methodologies. CONCLUSIONS: Our findings indicate that there is substantial variation in G6PD measurements by spectrophotometry between sites. This is likely due to variability in laboratory methods, with possible contribution of unmeasured population factors. While an assay-specific, universal quantitative threshold offers robust diagnosis at the 30% level, inter-study variability impedes performance of universal thresholds at the 70% level. Caution is advised in comparing findings based on absolute G6PD activity measurements across studies. Novel handheld quantitative G6PD diagnostics may allow greater standardisation in the future.

The risk of Plasmodium vivax parasitaemia after P. falciparum malaria: An individual patient data meta-analysis from the WorldWide Antimalarial Resistance Network.

BACKGROUND: There is a high risk of Plasmodium vivax parasitaemia following treatment of falciparum malaria. Our study aimed to quantify this risk and the associated determinants using an individual patient data meta-analysis in order to identify populations in which a policy of universal radical cure, combining artemisinin-based combination therapy (ACT) with a hypnozoitocidal antimalarial drug, would be beneficial. METHODS AND FINDINGS: A systematic review of Medline, Embase, Web of Science, and the Cochrane Database of Systematic Reviews identified efficacy studies of uncomplicated falciparum malaria treated with ACT that were undertaken in regions coendemic for P. vivax between 1 January 1960 and 5 January 2018. Data from eligible studies were pooled using standardised methodology. The risk of P. vivax parasitaemia at days 42 and 63 and associated risk factors were investigated by multivariable Cox regression analyses. Study quality was assessed using a tool developed by the Joanna Briggs Institute. The study was registered in the International Prospective Register of Systematic Reviews (PROSPERO: CRD42018097400). In total, 42 studies enrolling 15,341 patients were included in the analysis, including 30 randomised controlled trials and 12 cohort studies. Overall, 14,146 (92.2%) patients had P. falciparum monoinfection and 1,195 (7.8%) mixed infection with P. falciparum and P. vivax. The median age was 17.0 years (interquartile range [IQR] = 9.0-29.0 years; range = 0-80 years), with 1,584 (10.3%) patients younger than 5 years. 2,711 (17.7%) patients were treated with artemether-lumefantrine (AL, 13 studies), 651 (4.2%) with artesunate-amodiaquine (AA, 6 studies), 7,340 (47.8%) with artesunate-mefloquine (AM, 25 studies), and 4,639 (30.2%) with dihydroartemisinin-piperaquine (DP, 16 studies). 14,537 patients (94.8%) were enrolled from the Asia-Pacific region, 684 (4.5%) from the Americas, and 120 (0.8%) from Africa. At day 42, the cumulative risk of vivax parasitaemia following treatment of P. falciparum was 31.1% (95% CI 28.9-33.4) after AL, 14.1% (95% CI 10.8-18.3) after AA, 7.4% (95% CI 6.7-8.1) after AM, and 4.5% (95% CI 3.9-5.3) after DP. By day 63, the risks had risen to 39.9% (95% CI 36.6-43.3), 42.4% (95% CI 34.7-51.2), 22.8% (95% CI 21.2-24.4), and 12.8% (95% CI 11.4-14.5), respectively. In multivariable analyses, the highest rate of P. vivax parasitaemia over 42 days of follow-up was in patients residing in areas of short relapse periodicity (adjusted hazard ratio [AHR] = 6.2, 95% CI 2.0-19.5; p = 0.002); patients treated with AL (AHR = 6.2, 95% CI 4.6-8.5; p < 0.001), AA (AHR = 2.3, 95% CI 1.4-3.7; p = 0.001), or AM (AHR = 1.4, 95% CI 1.0-1.9; p = 0.028) compared with DP; and patients who did not clear their initial parasitaemia within 2 days (AHR = 1.8, 95% CI 1.4-2.3; p < 0.001). The analysis was limited by heterogeneity between study populations and lack of data from very low transmission settings. Study quality was high. CONCLUSIONS: In this meta-analysis, we found a high risk of P. vivax parasitaemia after treatment of P. falciparum malaria that varied significantly between studies. These P. vivax infections are likely attributable to relapses that could be prevented with radical cure including a hypnozoitocidal agent; however, the benefits of such a novel strategy will vary considerably between geographical areas.