Plasmodium falciparum infection in humans and mosquitoes influence natural Anopheline biting behavior and transmission.

The human infectious reservoir of Plasmodium falciparum is governed by transmission efficiency during vector-human contact and mosquito biting preferences. Understanding biting bias in a natural setting can help target interventions to interrupt transmission. In a 15-month cohort in western Kenya, we detected P. falciparum in indoor-resting Anopheles and human blood samples by qPCR and matched mosquito bloodmeals to cohort participants using short-tandem repeat genotyping. Using risk factor analyses and discrete choice models, we assessed mosquito biting behavior with respect to parasite transmission. Biting was highly unequal; 20% of people received 86% of bites. Biting rates were higher on males (biting rate ratio (BRR): 1.68; CI: 1.28-2.19), children 5-15 years (BRR: 1.49; CI: 1.13-1.98), and P. falciparum-infected individuals (BRR: 1.25; CI: 1.01-1.55). In aggregate, P. falciparum-infected school-age (5-15 years) boys accounted for 50% of bites potentially leading to onward transmission and had an entomological inoculation rate 6.4x higher than any other group. Additionally, infectious mosquitoes were nearly 3x more likely than non-infectious mosquitoes to bite P. falciparum-infected individuals (relative risk ratio 2.76, 95% CI 1.65-4.61). Thus, persistent P. falciparum transmission was characterized by disproportionate onward transmission from school-age boys and by the preference of infected mosquitoes to feed upon infected people.

Novel bioinformatic methods and machine learning approaches reveal candidate biomarkers of the intensity and timing of past exposure to Plasmodium falciparum.

Accurately quantifying the burden of malaria over time is an important goal of malaria surveillance efforts and can enable effective targeting and evaluation of interventions. Malaria surveillance methods capture active or recent infections which poses several challenges to achieving malaria surveillance goals. In high transmission settings, asymptomatic infections are common and therefore accurate measurement of malaria burden demands active surveillance; in low transmission regions where infections are rare accurate surveillance requires sampling large subsets of the population; and in any context monitoring malaria burden over time necessitates serial sampling. Antibody responses to Plasmodium falciparum parasites persist after infection and therefore measuring antibodies has the potential to overcome several of the current obstacles to accurate malaria surveillance. Identifying which antibody responses are markers of the timing and intensity of past exposure to P. falciparum remains challenging, particularly among adults who tend to be re-exposed multiple times over the course of their lifetime and therefore have similarly high antibody responses to many Plasmodium antigens. A previous analysis of 479 serum samples from individuals in three regions in southern Africa with different historical levels of P. falciparum malaria transmission (high, intermediate, and low) revealed regional differences in antibody responses to P. falciparum antigens among children under 5 years of age. Using a novel bioinformatic pipeline optimized for protein microarrays that minimizes between-sample technical variation, we used antibody responses to Plasmodium antigens as predictors in random forest models to classify samples from adults into these three regions of differing historical malaria transmission with high accuracy (AUC = 0.99). Many of the most important antigens for classification in these models do not overlap with previously published results and are therefore novel candidate markers for the timing and intensity of past exposure to P. falciparum. Measuring antibody responses to these antigens could lead to improved malaria surveillance.

Declining Age-Specific Seroprevalence and Seroconversion Rates in Plasmodium falciparum from 2009 to 2018 Documents Progress toward Malaria Elimination in Southern Zambia.

Obtaining accurate malaria surveillance data is challenging in low-transmission settings because large sample sizes are required to estimate incidence and prevalence precisely. Serology is an additional tool to document progress toward malaria elimination. An enzyme immunoassay to Plasmodium falciparum lysate was used to estimate age-specific seroprevalence among residents of southern Zambia, where malaria transmission has declined to pre-elimination levels during the past two decades. Plasma was eluted from 3,362 dried blood spots collected during five cross-sectional surveys conducted between 2009 and 2012, and again in 2018. Annual seroconversion rates (SCRs), an estimate of the force of infection, were calculated using a reversible catalytic model. The SCR decreased by two thirds from a level of approximately 0.15/year in 2009 and 2010 to approximately 0.05/year in 2011 and 2012, and then decreased 5-fold to 0.01/year by 2018, demonstrating the utility of serology in documenting progress toward elimination.

A Bayesian hierarchical model for signal extraction from protein microarrays.

Protein microarrays are a promising technology that measure protein levels in serum or plasma samples. Due to their high technical variability and high variation in protein levels across serum samples in any population, directly answering biological questions of interest using protein microarray measurements is challenging. Analyzing preprocessed data and within-sample ranks of protein levels can mitigate the impact of between-sample variation. As for any analysis, ranks are sensitive to preprocessing, but loss function based ranks that accommodate major structural relations and components of uncertainty are very effective. Bayesian modeling with full posterior distributions for quantities of interest produce the most effective ranks. Such Bayesian models have been developed for other assays, for example, DNA microarrays, but modeling assumptions for these assays are not appropriate for protein microarrays. Consequently, we develop and evaluate a Bayesian model to extract the full posterior distribution of normalized protein levels and associated ranks for protein microarrays, and show that it fits well to data from two studies that use protein microarrays produced by different manufacturing processes. We validate the model via simulation and demonstrate the downstream impact of using estimates from this model to obtain optimal ranks.

Drug Prescriptions Requiring Compounding at a Canadian University Affiliated Pediatric Hospital: A Cross-Sectional Study.

Despite ongoing international efforts, many drugs administered to children must be compounded from dosage forms designed for adults because they remain unavailable in commercial formulations that suit their needs. Even though oral drug compounding is common in pediatrics, the extent of this practice has not been well described in recent years. This cross-sectional and retrospective study was conducted at a Canadian university-affiliated, 484-bed, tertiary care pediatric hospital and its rehabilitation centre on two randomly selected days. A total of 606 hospitalized children with 5465 prescriptions were included. Overall, compounded drugs for enteral administration (CDEA) represented 13% of all prescriptions (enteral and parenteral) and 23% of prescriptions for enteral administration. Of the 390 prescribed drugs, 122 required compounding. CDEA were mostly liquids (n = 478 [67%]) and mainly included drugs of the central nervous (35%), cardiovascular (21%), and gastro-intestinal (12%) systems. Nearly half (N = 298 [49%]) of children had at least one CDEA prescribed in their medical file. Many CDEA are available as commercial products in other jurisdictions. Collaboration is needed between all stakeholders to make these drugs available to Canadian children.

Exploring how space, time, and sampling impact our ability to measure genetic structure across Plasmodium falciparum populations.

A primary use of malaria parasite genomics is identifying highly related infections to quantify epidemiological, spatial, or temporal factors associated with patterns of transmission. For example, spatial clustering of highly related parasites can indicate foci of transmission and temporal differences in relatedness can serve as evidence for changes in transmission over time. However, for infections in settings of moderate to high endemicity, understanding patterns of relatedness is compromised by complex infections, overall high forces of infection, and a highly diverse parasite population. It is not clear how much these factors limit the utility of using genomic data to better understand transmission in these settings. In particular, further investigation is required to determine which patterns of relatedness we expect to see with high quality, densely sampled genomic data in a high transmission setting and how these observations change under different study designs, missingness, and biases in sample collection. Here we investigate two identity-by-state measures of relatedness and apply them to amplicon deep sequencing data collected as part of a longitudinal cohort in Western Kenya that has previously been analysed to identify individual-factors associated with sharing parasites with infected mosquitoes. With these data we use permutation tests, to evaluate several hypotheses about spatiotemporal patterns of relatedness compared to a null distribution. We observe evidence of temporal structure, but not of fine-scale spatial structure in the cohort data. To explore factors associated with the lack of spatial structure in these data, we construct a series of simplified simulation scenarios using an agent based model calibrated to entomological, epidemiological and genomic data from this cohort study to investigate whether the lack of spatial structure observed in the cohort could be due to inherent power limitations of this analytical method. We further investigate how our hypothesis testing behaves under different sampling schemes, levels of completely random and systematic missingness, and different transmission intensities.

From paediatric formulations development to access: Advances made and remaining challenges.

Developing suitable paediatric formulations and ensuring access to them by the greatest number of the 2.2 billion children worldwide are equally important to provide optimal pharmacotherapy. This review focuses on the progress made over the last two decades with paediatric oral formulations with respect to evidence for acceptability and dosing flexibility of liquid and solid oral dosage forms. It also discusses the clinical needs for, and the access to, paediatric formulations for existing authorised medicines. A significant body of new knowledge now supports the acceptability of solid oral dosage forms in children, resulting in an increasing number of medicines commercialised as multiparticulates, including minitablets that are starting to be brought to market. However, there are gaps with these formulations that deserve more research. Even though efforts have been made to identify medicines in need of age-appropriate formulations, there is no common priority list shared internationally. Such prioritisation would help to develop paediatric formulations with the greatest potential for providing a health benefit to children worldwide. In addition, available data highlight that paediatric formulation access is fragmented and unequal, with commercialisation of suitable paediatric formulations too often limited to some countries/regions. We propose actions to better align decisions during the development of paediatric formulations and promote a more globalised approach to facilitate registration pathways between different jurisdictions. Furthermore, discussions about alignment between approval, pricing and reimbursement processes should also happen, leaving working in siloes behind us. It is time for adults to start thinking outside the box for children.

A pre-processing pipeline to quantify, visualize, and reduce technical variation in protein microarray studies.

Technical variation, or variation from non-biological sources, is present in most laboratory assays. Correcting for this variation enables analysts to extract a biological signal that informs questions of interest. However, each assay has different sources and levels of technical variation, and the choice of correction methods can impact downstream analyses. Compared to similar assays such as DNA microarrays, relatively few methods have been developed and evaluated for protein microarrays, a versatile tool for measuring levels of various proteins in serum samples. Here, we propose a pre-processing pipeline to correct for some common sources of technical variation in protein microarrays. The pipeline builds upon an existing normalization method by using controls to reduce technical variation. We evaluate our method using data from two protein microarray studies and by simulation. We demonstrate that pre-processing choices impact the fluorescent-intensity based ranks of proteins, which in turn, impact downstream analysis.

Development and dissemination of infectious disease dynamic transmission models during the COVID-19 pandemic: what can we learn from other pathogens and how can we move forward?

The current COVID-19 pandemic has resulted in the unprecedented development and integration of infectious disease dynamic transmission models into policy making and public health practice. Models offer a systematic way to investigate transmission dynamics and produce short-term and long-term predictions that explicitly integrate assumptions about biological, behavioural, and epidemiological processes that affect disease transmission, burden, and surveillance. Models have been valuable tools during the COVID-19 pandemic and other infectious disease outbreaks, able to generate possible trajectories of disease burden, evaluate the effectiveness of intervention strategies, and estimate key transmission variables. Particularly given the rapid pace of model development, evaluation, and integration with decision making in emergency situations, it is necessary to understand the benefits and pitfalls of transmission models. We review and highlight key aspects of the history of infectious disease dynamic models, the role of rigorous testing and evaluation, the integration with data, and the successful application of models to guide public health. Rather than being an expansive history of infectious disease models, this Review focuses on how the integration of modelling can continue to be advanced through policy and practice in appropriate and conscientious ways to support the current pandemic response.

Providing Suitable Pediatric Formulations for Canadian Children: A Call for Action.

BACKGROUND: Many medications given to children have no commercially available, age-appropriate formulations. This leads to manipulation of dosage forms designed for adults (compounding), which can result in an increased risk of dosing errors and adverse events, lack of medication adherence because of taste issues, and suboptimal dosing with therapeutic failure. OBJECTIVES: To determine which drugs required compounding for oral administration to children in a Canadian hospital and, for each compounded drug, to determine whether it was available as licensed oral pediatric formulations in the United States or the European Union. METHODS: Drugs requiring compounded liquid formulations for oral administration, dispensed from January 1 to December 31, 2015, at a Canadian university-affiliated tertiary pediatric hospital, and prepared in a quantity exceeding 0.5 L per year, were retrospectively identified. The online drug databases of Health Canada, the US Food and Drug Administration, the European Medicines Agency (EMA), and the UK Medicines and Healthcare Products Regulatory Agency were searched to determine the availability of child-friendly oral formulations for these drugs. The regulatory status in each jurisdiction was also compared. For licensed formulations with potential concerns about excipient safety, EMA guidelines for sorbitol, propylene glycol, ethanol, and sodium benzoate were used to determine pediatric suitability. RESULTS: Of the 56 compounded drugs investigated, 27 (48%) had a suitable commercialized child-friendly formulation available outside Canada. Overall, these drugs had been on the Canadian market for a median of 35 years, and almost half (27 [48%]) had a pediatric indication in Canada. CONCLUSIONS: Canada is lagging behind the United States and the European Union in ensuring availability of and access to suitable pediatric formulations. Potential explanations for this gap include small market size, regulatory uncertainties, and reimbursement shortcomings. Steps must be taken to implement pediatric-sensitive regulations and incentives, as well as reimbursement policies, to address these unmet needs.

CONTEXTE: Plusieurs médicaments administrés aux enfants ne sont pas disponibles commercialement sous une forme pharmaceutique adaptée à leur âge. Ceci entraîne une manipulation des formes destinées aux adultes (préparation magistrale) et peut conduire à une augmentation du risque d'erreurs de dosage et d'effets indésirables, un manque d'observance médicamenteuse secondairement à des problèmes de goût, et un dosage sous-optimal associé à des échecs thérapeutiques. OBJECTIFS: Définir les médicaments qui exigent une préparation magistrale pour être administrés par voie orale aux enfants dans un hôpital canadien et, pour chaque médicament faisant l'objet d'une préparation magistrale, déterminer s'il est disponible sous une forme pharmaceutique orale autorisée pour les enfants aux États-Unis ou dans l'Union européene. MÉTHODES: Les médicaments nécessitant des préparations magistrales liquides pour administration orale, distribués entre le 1er janvier et le 31 décembre 2015 dans un hôpital de soins pédiatriques tertiaires affilié à une université canadienne et dont la quantité préparée était supérieure à 0.5 L par an, ont été déterminés rétrospectivement. Les bases de données en ligne de médicaments de Santé Canada, de la Food and Drug Administration américaine, de l'Agence européenne des médicaments (AEM) et de la Medicines and Healthcare Products Regulatory Agency (Royaume-Uni) ont été interrogées pour déterminer la disponibilité de formes pharmaceutiques orales adaptées aux enfants pour ces médicaments. Le statut réglementaire de chaque pays a également fait l'objet d'une comparaison. Pour les formes pharmaceutiques autorisées présentant des problèmes potentiels d'innocuité des excipients, les directives de l'AEM concernant le sorbitol, le propylène glycol, l'éthanol et le benzoate de sodium ont servi à déterminer si un usage pédiatrique était acceptable. RÉSULTATS: Des 56 médicaments étudiés faisant l'objet d'une préparation magistrale, 27 (48 %) avaient une forme pharmaceutique commercialisée adaptée aux enfants en dehors du Canada. Au total, ces médicaments sont sur le marché canadien depuis une médiane de 35 ans et près de la moitié (27 [48 %]) ont une indication pédiatrique au Canada. CONCLUSIONS: Le Canada accuse un retard par rapport aux États-Unis et à l'Union européenne quant à la disponibilité et à l'accès à des formes pharmaceutiques adéquates pour les enfants. La petite taille du marché, les incertitudes en matière réglementaire et les lacunes concernant le remboursement pourraient notamment expliquer cet écart. Il est nécessaire de prendre des mesures pour mettre en place des réglementations et des incitatifs ainsi que des politiques de remboursement axés sur les enfants pour répondre à ces besoins criants.

High Plasmodium falciparum genetic diversity and temporal stability despite control efforts in high transmission settings along the international border between Zambia and the Democratic Republic of the Congo.

BACKGROUND: While the utility of parasite genotyping for malaria elimination has been extensively documented in low to moderate transmission settings, it has been less well-characterized in holoendemic regions. High malaria burden settings have received renewed attention acknowledging their critical role in malaria elimination. Defining the role for parasite genomics in driving these high burden settings towards elimination will enhance future control programme planning. METHODS: Amplicon deep sequencing was used to characterize parasite population genetic diversity at polymorphic Plasmodium falciparum loci, Pfama1 and Pfcsp, at two timepoints in June-July 2016 and January-March 2017 in a high transmission region along the international border between Luapula Province, Zambia and Haut-Katanga Province, the Democratic Republic of the Congo (DRC). RESULTS: High genetic diversity was observed across both seasons and in both countries. No evidence of population structure was observed between parasite populations on either side of the border, suggesting that this region may be one contiguous transmission zone. Despite a decline in parasite prevalence at the sampling locations in Haut-Katanga Province, no genetic signatures of a population bottleneck were detected, suggesting that larger declines in transmission may be required to reduce parasite genetic diversity. Analysing rare variants may be a suitable alternative approach for detecting epidemiologically important genetic signatures in highly diverse populations; however, the challenge is distinguishing true signals from potential artifacts introduced by small sample sizes. CONCLUSIONS: Continuing to explore and document the utility of various parasite genotyping approaches for understanding malaria transmission in holoendemic settings will be valuable to future control and elimination programmes, empowering evidence-based selection of tools and methods to address pertinent questions, thus enabling more efficient resource allocation.

Analyzing Prokaryotic RNA-Seq Data: A Case Study Identifying Holo-Fur Regulated Genes in Campylobacter jejuni.

In recent years, RNA-seq has become an important method in the process of measuring gene expression in various cells and organisms. This chapter will detail all the bioinformatic steps that should be undertaken to determine differentially expressed genes from a typical RNA-seq experiment. Each step will be clearly explained in "non-bioinformatic" terminology so that readers embarking on RNA-seq analysis will be able to understand the rationale and reasoning behind each step. Moreover, the exact command lines used to process the data will be presented along with a description of the various flags and commands.