Impact of changing drug treatment and malaria endemicity on the heritability of malaria phenotypes in a longitudinal family-based cohort study

Despite considerable success of genome wide association (GWA) studies in identifying causal variants for many human diseases, their success in unraveling the genetic basis to complex diseases has been more mitigated. Pathogen population structure may impact upon the infectious phenotype, especially with the intense short-term selective pressure that drug treatment exerts on pathogens. Rigorous analysis that accounts for repeated measures and disentangles the influence of genetic and environmental factors must be performed. Attempts should be made to consider whether pathogen diversity will impact upon host genetic responses to infection.We analyzed the heritability of two Plasmodium falciparum phenotypes, the number of clinical malaria episodes (PFA) and the proportion of these episodes positive for gametocytes (Pfgam), in a family-based cohort followed for 19 years, during which time there were four successive drug treatment regimes, with documented appearance of drug resistance. Repeated measures and variance components analyses were performed with fixed environmental, additive genetic, intra-individual and maternal effects for each drug period. Whilst there was a significant additive genetic effect underlying PFA during the first drug period of study, this was lost in subsequent periods. There was no additive genetic effect for Pfgam. The intra-individual effect increased significantly in the chloroquine period.The loss of an additive genetic effect following novel drug treatment may result in significant loss of power to detect genes in a GWA study. Prior genetic analysis must be a pre-requisite for more detailed GWA studies. The temporal changes in the individual genetic and the intra-individual estimates are consistent with those expected if there were specific host-parasite interactions. The complex basis to the human response to malaria parasite infection likely includes dominance/epistatic genetic effects encompassed within the intra-individual variance component. Evaluating their role in influencing the outcome of infection through host genotype by parasite genotype interactions warrants research effort.     

Expression Patterns and Potential Biological Roles of Dip2a

Disconnected (disco)-interacting protein 2 homolog A is a member of the DIP2 protein family encoded by Dip2a gene. Dip2a expression pattern has never been systematically studied. Functions of Dip2a in embryonic development and adult are not known. To investigate Dip2a gene expression and function in embryo and adult, a Dip2a-LacZ mouse model was generated by insertion of β-Gal cDNA after Dip2a promoter using CRISPR/Cas9 technology. Dip2a-LacZ mouse was designed to be a lacZ reporter mouse as well as a Dip2a knockout mouse. Heterozygous mice were used to study endogenous Dip2a expression and homozygotes to study DIP2A-associated structure and function. LacZ staining indicated that Dip2a is broadly expressed in neuronal, reproductive and vascular tissues, as well as in heart, kidney, liver and lung. Results demonstrate that Dip2a is expressed in ectoderm-derived tissues in developing embryos. Adult tissues showed rich staining in neurons, mesenchymal, endothelial, smooth muscle cells and cardiomyocytes by cell types. The expression pattern highly overlaps with FSTL1 and supports previous report that DIP2A to be potential receptor of FSTL1 and its protective roles of cardiomyocytes. Broad and intense embryonic and adult expression of Dip2a has implied their multiple structural and physiological roles.     

Severe bacterial neonatal infections in Madagascar,Senegal, and Cambodia: A multicentric community-based cohort study

BackgroundSevere bacterial infections (SBIs) are a leading cause of neonatal deaths in low- and middle-income countries (LMICs). However, most data came from hospitals, which do not include neonates who did not seek care or were treated outside the hospital. Studies from the community are scarce, and few among those available were conducted with high-quality microbiological techniques. The burden of SBI at the community level is therefore largely unknown. We aimed here to describe the incidence, etiology, risk factors, and antibiotic resistance profiles of community-acquired neonatal SBI in 3 LMICs.Methods and findingsThe BIRDY study is a prospective multicentric community-based mother and child cohort study and was conducted in both urban and rural areas in Madagascar (2012 to 2018), Cambodia (2014 to 2018), and Senegal (2014 to 2018). All pregnant women within a geographically defined population were identified and enrolled. Their neonates were actively followed from birth to 28 days to document all episodes of SBI. A total of 3,858 pregnant women (2,273 (58.9%) in Madagascar, 814 (21.1%) in Cambodia, and 771 (20.0%) in Senegal) were enrolled in the study, and, of these, 31.2% were primigravidae. Women enrolled in the urban sites represented 39.6% (900/2,273), 45.5% (370/814), and 61.9% (477/771), and those enrolled in the rural sites represented 60.4% (1,373/2,273), 54.5% (444/814), and 38.1% (294/771) of the total in Madagascar, Cambodia, and Senegal, respectively. Among the 3,688 recruited newborns, 49.6% were male and 8.7% were low birth weight (LBW). The incidence of possible severe bacterial infection (pSBI; clinical diagnosis based on WHO guidelines of the Integrated Management of Childhood Illness) was 196.3 [95% confidence interval (CI) 176.5 to 218.2], 110.1 [88.3 to 137.3], and 78.3 [59.5 to 103] per 1,000 live births in Madagascar, Cambodia, and Senegal, respectively. The incidence of pSBI differed between urban and rural sites in all study countries. In Madagascar, we estimated an incidence of 161.0 pSBI per 1,000 live births [133.5 to 194] in the urban site and 219.0 [192.6 to 249.1] pSBI per 1,000 live births in the rural site (p = 0.008). In Cambodia, estimated incidences were 141.1 [105.4 to 189.0] and 85.3 [61.0 to 119.4] pSBI per 1,000 live births in urban and rural sites, respectively (p = 0.025), while in Senegal, we estimated 103.6 [76.0 to 141.2] pSBI and 41.5 [23.0 to 75.0] pSBI per 1,000 live births in urban and rural sites, respectively (p = 0.006). The incidences of culture-confirmed SBI were 15.2 [10.6 to 21.8], 6.5 [2.7 to 15.6], and 10.2 [4.8 to 21.3] per 1,000 live births in Madagascar, Cambodia, and Senegal, respectively, with no difference between urban and rural sites in each country. The great majority of early-onset infections occurred during the first 3 days of life (72.7%). The 3 main pathogens isolated were Klebsiella spp. (11/45, 24.4%), Escherichia coli (10/45, 22.2%), and Staphylococcus spp. (11/45, 24.4%). Among the 13 gram-positive isolates, 5 were resistant to gentamicin, and, among the 29 gram-negative isolates, 13 were resistant to gentamicin, with only 1 E. coli out of 10 sensitive to ampicillin. Almost one-third of the isolates were resistant to both first-line drugs recommended for the management of neonatal sepsis (ampicillin and gentamicin). Overall, 38 deaths occurred among neonates with SBI (possible and culture-confirmed SBI together). LBW and foul-smelling amniotic fluid at delivery were common risk factors for early pSBI in all 3 countries. A main limitation of the study was the lack of samples from a significant proportion of infants with pBSI including 35 neonatal deaths. Without these samples, bacterial infection and resistance profiles could not be confirmed.ConclusionsIn this study, we observed a high incidence of neonatal SBI, particularly in the first 3 days of life, in the community of 3 LMICs. The current treatment for the management of neonatal infection is hindered by antimicrobial resistance. Our findings suggest that microbiological diagnosis of SBI remains a challenge in these settings and support more research on causes of neonatal death and the implementation of early interventions (e.g., follow-up of at-risk newborns during the first days of life) to decrease the burden of neonatal SBI and associated mortality and help achieve Sustainable Development Goal 3.

In a community-based, prospective cohort study, Bich-Tram Huynh and colleagues investigate the incidence and factors associated with several bacterial infections among neonates in rural and urban areas of three low-middle income countries.     

Is there NO help for leptin?

Since the initial identification of leptin as the product of the ob gene in 1994, the signaling pathways by which this hormone alters cell physiology have been the subject of extensive investigations. The fact that leptin can induce nitric oxide (NO) production was first demonstrated in studies of the pituitary gland and pancreatic islets. A large number of additional studies further showed that this adipokine stimulates NO synthesis in multiple tissues. This review article discusses the role of leptin in NO production and its pathophysiological consequences. The role of this gaseous messenger in cell physiology depends on the cell type, the concentration of NO and the duration of exposure. It can be either a potent oxidant or a protector of cell integrity against the formation of reactive oxygen species. Leptin plays two opposing roles on arterial pressure. It exerts a hypertensive effect due to sympathetic activation and a vasorelaxant effect due to NO production. This adipokine acts via NO to produce pro-inflammatory factors in cartilage pathology, potentially contributing to an increased risk for osteoarthritis. Another well-documented role of leptin-induced NO, acting either directly or via the hypothalamus, concerns lipid metabolism in muscle and adipose tissue. In adipocytes, the direct and rapid action of leptin is to activate the nitric oxide synthase III, which favors lipolysis. In contrast, in the long-term, leptin reduces lipolysis. However, both in the short-term and in the long-term, glyceroneogenesis and its key enzyme, the cytosolic phosphoenolpyruvatecarboxykinase (PEPCK-C), are down-regulated by the adipokine, thus favoring fatty acid release. Hence, leptin-induced NO production plays a crucial role in fatty acid metabolism in adipose tissue. The resulting effects are to prevent lipid storage and to improve energy expenditure, with possible improvements of the obese state and its associated diseases.     

Saccharomyces cerevisiae and Candida tropicalis as starter cultures for the alcoholic fermentation of tchapalo, a traditional sorghum beer

Starter cultures of Candida tropicalis and Saccharomyces cerevisiae isolated from tchapalo were tested in pure culture and co-culture of four ratios [2:1, 25:4, 1:4, 2:3 (cells/cells)] for their ability to ferment sorghum wort to produce tchapalo. All the starters showed means growth rate between 0.043 and 0.101 h^?1. Only C. tropicalis in pure culture showed growth rate lower than that of S. cerevisiae in single culture. During fermentation, according to total soluble solids depletion, yeast starters could be grouped in four different profiles. But in the beer produced, total soluble solids contents were statistically identical. The lowest values were obtained with co-culture C. tropicalis + S. cerevisiae in the ratios of 2:1 and 2:3. Starter cultures with large ratio of C. tropicalis produced a higher organic acids and 2-butanone than S. cerevisiae in pure culture. However, co-culture C. tropicalis + S. cerevisiae (2:1) was the alone starter which produced higher ethanol than S. cerevisiae in pure culture. The beers produced with C. tropicalis + S. cerevisiae (25:4), C. tropicalis + S. cerevisiae (1:4) and C. tropicalis were widely different from those produced with the others starter cultures.     

An exhaustive, non-euclidean, non-parametric data mining tool for unraveling the complexity of biological systems--novel insights into malaria

Complex, high-dimensional data sets pose significant analytical challenges in the post-genomic era. Such data sets are not exclusive to genetic analyses and are also pertinent to epidemiology. There has been considerable effort to develop hypothesis-free data mining and machine learning methodologies. However, current methodologies lack exhaustivity and general applicability. Here we use a novel non-parametric, non-euclidean data mining tool, HyperCube®, to explore exhaustively a complex epidemiological malaria data set by searching for over density of events in m-dimensional space. Hotspots of over density correspond to strings of variables, rules, that determine, in this case, the occurrence of Plasmodium falciparum clinical malaria episodes. The data set contained 46,837 outcome events from 1,653 individuals and 34 explanatory variables. The best predictive rule contained 1,689 events from 148 individuals and was defined as: individuals present during 1992–2003, aged 1–5 years old, having hemoglobin AA, and having had previous Plasmodium malariae malaria parasite infection ≤10 times. These individuals had 3.71 times more P. falciparum clinical malaria episodes than the general population. We validated the rule in two different cohorts. We compared and contrasted the HyperCube® rule with the rules using variables identified by both traditional statistical methods and non-parametric regression tree methods. In addition, we tried all possible sub-stratified quantitative variables. No other model with equal or greater representativity gave a higher Relative Risk. Although three of the four variables in the rule were intuitive, the effect of number of P. malariae episodes was not. HyperCube® efficiently sub-stratified quantitative variables to optimize the rule and was able to identify interactions among the variables, tasks not easy to perform using standard data mining methods. Search of local over density in m-dimensional space, explained by easily interpretable rules, is thus seemingly ideal for generating hypotheses for large datasets to unravel the complexity inherent in biological systems.     

Time Series Analysis of Trends in Malaria Cases and Deaths at Hospitals and the Effect of Antimalarial Interventions, 2001–2011, Ethiopia

Background

The Government of Ethiopia and its partners have deployed artemisinin-based combination therapies (ACT) since 2004 and long-lasting insecticidal nets (LLINs) since 2005. Malaria interventions and trends in malaria cases and deaths were assessed at hospitals in malaria transmission areas during 2001–2011.

Methods

Regional LLINs distribution records were used to estimate the proportion of the population-at-risk protected by LLINs. Hospital records were reviewed to estimate ACT availability. Time-series analysis was applied to data from 41 hospitals in malaria risk areas to assess trends of malaria cases and deaths during pre-intervention (2001–2005) and post-interventions (2006–2011) periods.

Findings

The proportion of the population-at-risk potentially protected by LLINs increased to 51% in 2011. The proportion of facilities with ACTs in stock exceeded 87% during 2006–2011. Among all ages, confirmed malaria cases in 2011 declined by 66% (95% confidence interval [CI], 44–79%) and SPR by 37% (CI, 20%–51%) compared to the level predicted by pre-intervention trends. In children under 5 years of age, malaria admissions and deaths fell by 81% (CI, 47%–94%) and 73% (CI, 48%–86%) respectively. Optimal breakpoint of the trendlines occurred between January and June 2006, consistent with the timing of malaria interventions. Over the same period, non-malaria cases and deaths either increased or remained unchanged, the number of malaria diagnostic tests performed reflected the decline in malaria cases, and rainfall remained at levels supportive of malaria transmission.

Conclusions

Malaria cases and deaths in Ethiopian hospitals decreased substantially during 2006–2011 in conjunction with scale-up of malaria interventions. The decrease could not be accounted for by changes in hospital visits, malaria diagnostic testing or rainfall. However, given the history of variable malaria transmission in Ethiopia, more data would be required to exclude the possibility that the decrease is due to other factors.     

Shifts in Mycobacterial Populations and Emerging Drug-Resistance in West and Central Africa

In this study, we retrospectively analysed a total of 605 clinical isolates from six West or Central African countries (Benin, Cameroon, Central African Republic, Guinea-Conakry, Niger and Senegal). Besides spoligotyping to assign isolates to ancient and modern mycobacterial lineages, we conducted phenotypic drug-susceptibility-testing for each isolate for the four first-line drugs. We showed that phylogenetically modern Mycobacterium tuberculosis strains are more likely associated with drug resistance than ancient strains and predict that the currently ongoing replacement of the endemic ancient by a modern mycobacterial population in West/Central Africa might result in increased drug resistance in the sub-region.     

Rapid nitration of adipocyte phosphoenolpyruvate carboxykinase by leptin reduces glyceroneogenesis and induces fatty acid release

Fatty acid (FA) release from white adipose tissue (WAT) is the result of the balance between triglyceride breakdown and FA re-esterification. The latter relies on the induction of cytosolic phosphoenolpyruvate carboxykinase (PEPCK-C), the key enzyme for glyceroneogenesis. We previously demonstrated that long-term (18 h) leptin treatment of rat epididymal WAT explants reduced glyceroneogenesis through nitric oxide (NO)-induced decrease in PEPCK-C expression. We investigated the effect of a short-term leptin treatment (2 h) on PEPCK-C expression and glyceroneogenesis in relation to NO production. We demonstrate that in WAT explants, leptin-induced NO synthase III (NOS III) phosphorylation was associated with reduced PEPCK-C level and glyceroneogenesis, leading to FA release, while PEPCK-C gene expression remained unaffected. These effects were absent in WAT explants from leptin receptor-deficient Zucker rat. Immunoprecipitation and western blot experiments showed that the leptin-induced decrease in PEPCK-C level was correlated with an increase in PEPCK-C nitration. All these effects were abolished by the NOS inhibitor Nω-nitro-L-arginine methyl ester and mimicked by the NO donor S-nitroso-N-acetyl-DL penicillamine. We propose a mechanism in which leptin activates NOS III and induces NO that nitrates PEPCK-C to reduce its level and glyceroneogenesis, therefore limiting FA re-esterification in WAT.