Preeclampsia is the most important cause of maternal mortality in developing countries. A comprehensive prenatal care program including bio-psychosocial components was developed and introduced at a national level in Colombia. We report on the trends in maternal mortality rates and their related causes before and after implementation of this program.
General and specific maternal mortality rates were monitored for nine years (1998-2006). An interrupted time-series analysis was performed with monthly data on cases of maternal mortality that compared trends and changes in national mortality rates and the impact of these changes attributable to the introduction of a bio-psychosocial model. Multivariate analyses were performed to evaluate correlations between the interventions.
Five years after (2002 - 2006) its introduction the general maternal mortality rate was significantly reduced to 23% (OR=0.77, CI 95% 0.71-0.82).The implementation of BPSM also reduced the incidence of preeclampsia in 22% (OR= 0.78, CI 95% 0.67-0.88), as also the labor complications by hemorrhage in 25% (OR=0.75, CI 95% 0.59-0.90) associated with the implementation of red code. The other causes of maternal mortality did not reveal significant changes. Biomedical, nutritional, psychosocial assessments, and other individual interventions in prenatal care were not correlated to maternal mortality (p= 0.112); however, together as a model we observed a significant association (p= 0.042).
General maternal mortality was reduced after the implementation of a comprehensive national prenatal care program. Is important the evaluation of this program in others populations.
Preeclampsia; maternal mortality; population study; bio-psychosocial model
Most commonly used malaria diagnostic tests, including microscopy and antigen-detecting rapid tests, cannot reliably detect low-density infections which are frequent in low transmission settings. Molecular methods such as polymerase chain reaction (PCR) are highly sensitive but remain too laborious for field deployment. In this study, the applicability of a malaria diagnosis kit based on loop-mediated isothermal amplification (mLAMP) was assessed in malaria endemic areas of Colombia with Plasmodium vivax predominance.
First, a passive case detection (PCD) study on 278 febrile patients recruited in Tierralta (department of Cordoba) was conducted to assess the diagnostic performance of the mLAMP method. Second, an active case detection (ACD) study on 980 volunteers was conducted in 10 sentinel sites with different epidemiological profiles. Whole blood samples were processed for microscopic and mLAMP diagnosis. Additionally RT-PCR and nested RT-PCR were used as reference tests. In the PCD study, P. falciparum accounted for 23.9% and P. vivax for 76.1% of the infections and no cases of mixed-infections were identified. Microscopy sensitivity for P. falciparum and P. vivax were 100% and 86.1%, respectively. mLAMP sensitivity for P. falciparum and P. vivax was 100% and 91.4%, respectively. In the ACD study, mLAMP detected 65 times more cases than microscopy. A high proportion (98.0%) of the infections detected by mLAMP was from volunteers without symptoms.
mLAMP sensitivity and specificity were comparable to RT-PCR. LAMP was significantly superior to microscopy and in P. vivax low-endemicity settings and under minimum infrastructure conditions, it displayed sensitivity and specificity similar to that of single-well RT-PCR for detection of both P. falciparum and P. vivax infections. Here, the dramatically increased detection of asymptomatic malaria infections by mLAMP demonstrates the usefulness of this new tool for diagnosis, surveillance, and screening in elimination strategies.
The ability to detect and treat asymptomatic infections will be fundamental to eliminate malaria. This requires highly sensitive screening tests close enough to cases to enable rapid treatment. Very low parasitemias can be detected by molecular methods such as PCR; however, these techniques require considerable training and are restricted to reference laboratories. A new field-stable diagnostic kit for malaria based on loop-mediated isothermal DNA amplification (LAMP) is now commercially available. This LAMP kit is able to detect down to 1 parasite/µl of blood in less than 1 hour. In order to evaluate the feasibility of this LAMP kit as a tool for the detection of asymptomatic malaria in malaria endemic areas of Colombia with Plasmodium vivax predominance, we conducted field studies using the LAMP kit implemented in remote settings. We found that LAMP sensitivity and specificity were comparable to RT-PCR for detection of both P. falciparum and P. vivax infections and dramatically increased detection of asymptomatic malaria infections. This simple detection method for very low parasitemia raises opportunities and new strategies for malaria elimination.
Discoidin domain receptor 2 (DDR2) is a receptor tyrosine kinase involved in a variety of cellular response pathways, including regulation of cell growth, proliferation and motility. Using a newly developed platform to identify the signaling pathway/molecular target of natural products, we identified a family of alkaloid natural products, discoipyrroles A–D (1–4), from Bacillus hunanensis that inhibit the DDR2 signaling pathway. The structure of 1–4, determined by detailed 2D NMR methods and confirmed by X-ray crystallographic analysis has an unusual 3H-benzo[ d]pyrrolo][1,3]oxazine-3,5-dione core. Discoipyrroles A–D potently inhibit DDR2 dependent migration of BR5 fibroblasts and show selective cytotoxicity to DDR2 mutant cell lung cancer cell lines (IC50 120–400 nM). Examination of the biosynthesis has led to the conclusion that the discoipyrroles are formed through a non-enzymatic process, leading to a one-pot total synthesis of 1.
A challenge for biomedical research is the development of pharmaceuticals that appropriately target disease mechanisms. Natural products can be a rich source of bioactive chemicals for medicinal applications but can act through unknown mechanisms and can be difficult to produce or obtain. To address these challenges, we developed a new marine-derived, renewable natural products resource and a method for linking bioactive derivatives of this library to the proteins and biological processes that they target in cells. We used cell-based screening and computational analysis to match gene expression signatures produced by natural products to those produced by siRNA and synthetic microRNA libraries. With this strategy, we matched proteins and microRNAs with diverse biological processes and also identified putative protein targets and mechanisms of action for several previously undescribed marine-derived natural products. We confirmed mechanistic relationships for selected short-interfering RNAs, microRNAs, and compounds with functional roles in autophagy, chemotaxis mediated by discoidin domain receptor 2, or activation of the kinase AKT. Thus, this approach may be an effective method for screening new drugs while simultaneously identifying their targets.
Significant progress has been recently achieved in the development of Plasmodium vivax challenge infections in humans, which are essential for vaccine and drug testing. With the goal of accelerating clinical development of malaria vaccines, the outcome of infections experimentally induced in naïve and semi-immune volunteers by infected mosquito bites was compared.
Seven malaria-naïve and nine semi-immune Colombian adults (n = 16) were subjected to the bites of 2–4 P. vivax sporozoite-infected Anopheles mosquitoes. Parasitemia levels, malaria clinical manifestations, and immune responses were assessed and compared.
All volunteers developed infections as confirmed by microscopy and RT-qPCR. No significant difference in the pre-patent period (mean 12.5 and 12.8 days for malaria-naïve and malaria-exposed, respectively) was observed but naïve volunteers developed classical malaria signs and symptoms, while semi-immune volunteers displayed minor or no symptoms at the day of diagnosis. A malaria-naïve volunteer developed a transient low submicroscopic parasitemia that cured spontaneously. Infection induced an increase in specific antibody levels in both groups.
Sporozoite infectious challenge was safe and reproducible in semi-immune and naïve volunteers. This model will provide information for simultaneous comparison of the protective efficacy of P. vivax vaccines in naïve and semi-immune volunteers under controlled conditions and would accelerate P. vivax vaccine development.
Protein α-helical coiled coil structures that elicit antibody responses, which block critical functions of medically important microorganisms, represent a means for vaccine development. By using bioinformatics algorithms, a total of 50 antigens with α-helical coiled coil motifs orthologous to Plasmodium falciparum were identified in the P. vivax genome. The peptides identified in silico were chemically synthesized; circular dichroism studies indicated partial or high α-helical content. Antigenicity was evaluated using human sera samples from malaria-endemic areas of Colombia and Papua New Guinea. Eight of these fragments were selected and used to assess immunogenicity in BALB/c mice. ELISA assays indicated strong reactivity of serum samples from individuals residing in malaria-endemic regions and sera of immunized mice, with the α-helical coiled coil structures. In addition, ex vivo production of IFN-γ by murine mononuclear cells confirmed the immunogenicity of these structures and the presence of T-cell epitopes in the peptide sequences. Moreover, sera of mice immunized with four of the eight antigens recognized native proteins on blood-stage P. vivax parasites, and antigenic cross-reactivity with three of the peptides was observed when reacted with both the P. falciparum orthologous fragments and whole parasites. Results here point to the α-helical coiled coil peptides as possible P. vivax malaria vaccine candidates as were observed for P. falciparum. Fragments selected here warrant further study in humans and non-human primate models to assess their protective efficacy as single components or assembled as hybrid linear epitopes.
Although Colombia has witnessed an important decrease in malaria transmission, the disease remains a public health problem with an estimated ~10 million people currently living in areas with malaria risk and ~61,000 cases reported in 2012. This study aimed to determine and compare the level of knowledge, attitudes and practices (KAP) about malaria in three endemic communities of Colombia to provide the knowledge framework for development of new intervention strategies for malaria elimination.
A cross-sectional KAP survey was conducted in the municipalities of Tierralta, Buenaventura and Tumaco, categorized according to high risk (HR) and moderate risk (MR) based on the annual parasite index (API). Surveys were managed using REDCap and analysed using MATLAB and GraphPad Prism.
A total of 267 residents, mostly women (74%) were surveyed. Although no differences were observed on the knowledge of classical malaria symptoms between HR and MR regions, significant differences were found in knowledge and attitudes about transmission mechanisms, anti-malarial use and malaria diagnosis. Most responders in both regions (93.5% in MR, and 94.3% in HR areas) indicated use of insecticide-treated nets (ITNs) to protect themselves from malaria, and 75.5% of responders in HR indicated they did nothing to prevent malaria transmission outdoors. Despite a high level of knowledge in the study regions, significant gaps persisted relating to practices. Self-medication and poor adherence to treatment, as well as lack of both indoor and outdoor vector control measures, were significantly associated with higher malaria risk.
Although significant efforts are currently being made by the Ministry of Health to use community education as one of the main components of the control strategy, these generic education programmes may not be applicable to all endemic regions of Colombia given the substantial geographic, ethnic and cultural diversity.
The development of periodontal disease and dental caries is influenced by several factors, such as microorganisms of bacterial biofilm or commensal bacteria in the mouth. These microorganisms trigger inflammatory and immune responses in the host. Currently, medicinal plants are treatment options for these oral diseases. Mammea americana extracts have reported antimicrobial effects against several microorganisms. Nevertheless, this effect is unknown against oral bacteria. Therefore, the aim of this study was to evaluate the antibacterial effect of M. americana extract against Porphyromonas gingivalis and Streptococcus mutans. For this, an experimental study was conducted. Ethanolic extract was obtained from seeds of M. americana (one oil phase and one ethanolic phase). The strains of Porphyromonas gingivalis ATCC 33277 and Streptococcus mutans ATCC 25175 were exposed to this extract to evaluate its antibacterial effect. Antibacterial activity was observed with the two phases of M. americana extract on P. gingivalis and S. mutans with lower MICs (minimum inhibitory concentration). Also, bactericidal and bacteriostatic activity was detected against S. mutans, depending on the concentration of the extract, while on M. americana extract presented only bacteriostatic activity against P. gingivalis. These findings provide important and promising information allowing for further exploration in the future.
Introduction. Cytoreductive surgery (CS) with hyperthermic intraperitoneal chemotherapy (HIPEC) is a new approach for peritoneal carcinomatosis. However, high rates of complications are associated with CS and HIPEC due to treatment complexity; that is why some patients need stabilization and surveillance for complications in the intensive care unit. Objective. This study analyzed that ICU stay is necessary after HIPEC. Methods. 39 patients with peritoneal carcinomatosis were treated according to strict selection criteria with CS and HIPEC, with closed technique, and the chemotherapy administered were cisplatin 25 mg/m2/L and mitomycin C 3.3 mg/m2/L for 90-minutes at 40.5°C. Results. 26 (67%) of the 39 patients were transferred to the ICU. Major postoperative complications were seen in 14/26 patients (53%). The mean time on surgical procedures was 7.06 hours (range 5−9 hours). The mean blood loss was 939 ml (range 100–3700 ml). The mean time stay in the ICU was 2.7 days. Conclusion. CS with HIPEC for the treatment of PC results in low mortality and high morbidity. Therefore, ICU stay directly following HIPEC should not be standardized, but should preferably be based on the extent or resections performed and individual patient characteristics and risk factors. Late complications were comparable to those reported after large abdominal surgery without HIPEC.
Massive implementation of malaria diagnostics in low-resource countries is regarded as a pivotal strategy in control and elimination efforts. Although malaria rapid diagnostic tests (RDTs) are considered a viable alternative, there are still obstacles to the widespread implementation of this strategy, such as reporting constraints and lack of proper quality assurance of RDT-based programmes at point-of-care (POC).
A prospective cohort of patients, seeking routine care for febrile episodes at health centres in malaria-endemic areas of Colombia, was used to assess the diagnostic performance of a device based on smartphone technology (Deki ReaderTM, former codename “GenZero”), that assists users at POC to process RDTs. After informed consent, patients were enrolled into the study and blood samples were collected for thick blood smear (TBS) and RDT. The RDT results were interpreted by both visual inspection and Deki Reader device and concordance between visual and device interpretation was measured. Microscopy corrected by real-time polymerase chain reaction (PCR) and microscopy were “gold standard” tests to assess the diagnostic performance.
In total, 1,807 patients were enrolled at seven health centres in malaria-endemic areas of Colombia. Thirty-three Plasmodium falciparum and 100 Plasmodium vivax cases were positive by corrected microscopy. Both sensitivity and specificity were 93.9% (95% CI 69.7-95.2) and 98.7% (95% CI 98.5-99.4) for P. falciparum, and 98.0% (95% CI 90.3-98.9) and 97.9% (95% CI 97.1-98.5) for P. vivax. Percentage concordance between visual and device interpretation of RDT was 98.5% and 99.0% for P. vivax and P. falciparum, respectively.The RDT, when compared to TBS, showed high sensitivity and specificity for P. falciparum in both visual and device interpretation, and good overall diagnostic performance for P. vivax. Comparison between PCR as gold standard and visual and device interpretation showed acceptable overall performance for both species.
The diagnostic performance of the Deki Reader was comparable to visual interpretation of RDTs (without significant differences) for both malaria species. Providing standardized automated interpretation of RDTs at POC in remote areas, in addition to almost real-time reporting of cases and enabling quality control would greatly benefit large-scale implementation of RDT-based malaria diagnostic programmes.
DekiReader; Malaria rapid diagnostic test; PCR; mHealth
AIM: To investigate the impact of primary reperfusion therapy (RT) on early and late mortality in acute right ventricular infarction (RVI).
METHODS: RVI patients (n = 679) were prospectively classified as without right ventricular failure (RVF) (class A, n = 425, 64%), with RVF (class B, n = 158, 24%) or with cardiogenic shock (CS) (class C, n = 96, 12%). Of the 679 patients, 148 (21.7%) were considered to be eligible for thrombolytic therapy (TT) and 351 (51.6%) for primary percutaneous coronary intervention (PPCI). TIMI 3-flow by TT was achieved for A, B and C RVI class in 65%, 64% and 0%, respectively and with PPCI in 93%, 91% and 87%, respectively.
RESULTS: For class A without RT, the mortality rate was 7.9%, with TT was reduced to 4.4% (P < 0.01) and with PPCI to 3.2% (P < 0.01). Considering TT vs PPCI, PPCI was superior (P < 0.05). For class B without RT the mortality was 27%, decreased to 13% with TT (P < 0.01) and to 8.3% with PPCI (P < 0.01). In a TT and PPCI comparison, PPCI was superior (P < 0.01). For class C without RT the in-hospital mortality was 80%, with TT was 100% and with PPCI, the rate decreased to 44% (P < 0.01). At 8 years, the mortality rate without RT for class A was 32%, for class B was 48% and for class C was 85%. When PPCI was successful, the long-term mortality was lower than previously reported for the 3 RVI classes (A: 21%, B: 38%, C: 70%; P < 0.001).
CONCLUSION: PPCI is superior to TT and reduces short/long-term mortality for all RVI categories. RVI CS patients should be encouraged to undergo PPCI at a specialized center.
Right ventricular infarction; Reperfusion therapy; Ventricular failure; Cardiogenic shock; Morbidity; Mortality
The development of pre-erythrocytic Plasmodium vivax vaccines is hindered by the lack of in vitro culture systems or experimental rodent models. To help bypass these roadblocks, we exploited the fact that naturally exposed Fy− individuals who lack the Duffy blood antigen (Fy) receptor are less likely to develop blood-stage infections; therefore, they preferentially develop immune responses to pre-erythrocytic–stage parasites, whereas Fy+ individuals experience both liver- and blood-stage infections and develop immune responses to both pre-erythrocytic and erythrocytic parasites. We screened 60 endemic sera from P. vivax-exposed Fy+ or Fy− donors against a protein microarray containing 91 P. vivax proteins with P. falciparum orthologs that were up-regulated in sporozoites. Antibodies against 10 P. vivax antigens were identified in sera from P. vivax-exposed individuals but not unexposed controls. This technology has promising implications in the discovery of potential vaccine candidates against P. vivax malaria.
Malaria is a disease induced by parasites of the Plasmodium genus, which are transmitted by Anopheles mosquitoes and represents a great socio-economic burden Worldwide. Plasmodium vivax is the second species of malaria Worldwide, but it is the most prevalent in Latin America and other regions of the planet. It is currently considered that vaccines represent a cost-effective strategy for controlling transmissible diseases and could complement other malaria control measures; however, the chemical and immunological complexity of the parasite has hindered development of effective vaccines. Recent availability of several genomes of Plasmodium species, as well as bioinformatic tools are allowing the selection of large numbers of proteins and analysis of their immune potential. Herein, we review recently developed strategies for discovery of novel antigens with potential for malaria vaccine development.
malaria; vaccines; Plasmodium; Antigens; peptides; recombinant proteins
Latin America contributes 1 to 1.2 million clinical malaria cases to the global malaria burden of about 300 million per year. In 21 malaria endemic countries, the population at risk in this region represents less than 10% of the total population exposed worldwide. Factors such as rapid deforestation, inadequate agricultural practices, climate change, political instability, and both increasing parasite drug resistance and vector resistance to insecticides contribute to malaria transmission. Recently, several malaria endemic countries have experienced a significant reduction in numbers of malaria cases. This is most likely due to actions taken by National Malaria Control Programs (NMCP) with the support from international funding agencies. We describe here the research strategies and activities to be undertaken by the Centro Latino Americano de Investigación en Malaria (CLAIM), a new research center established for the non-Amazonian region of Latin America by the National Institute of Allergy and Infectious Diseases (NIAID). Throughout a network of countries in the region, initially including Colombia, Guatemala, Panama, and Peru, CLAIM will address major gaps in our understanding of changing malaria epidemiology, vector biology and control, and clinical malaria mainly due to Plasmodium vivax. In close partnership with NMCPs, CLAIM seeks to conduct research on how and why malaria is decreasing in many countries of the region as a basis for developing and implementing new strategies that will accelerate malaria elimination.
malaria; Plasmodium falciparum; Plasmodium vivax; Anopheles mosquitoes; vector control; epidemiology; malaria elimination; malaria pathogenesis; non-Amazon regions; Latin America
Approximately 170 million inhabitants of the American continent live at risk of malaria transmission. Although the continent’s contribution to the global malaria burden is small, at least 1 to 1.2 million malaria cases are reported annually. Sixty per cent of the malaria cases occur in Brazil and the other 40% are distributed in 20 other countries of Central and South America. Plasmodium vivax is the predominant species (74.2 %) followed by P. falciparum (25.7 %) and P. malariae (0.1%), and no less than 10 Anopheles species have been identified as primary or secondary malaria vectors. Rapid deforestation and agricultural practices are directly related to increases in Anopheles species diversity and abundance, as well as in the number of malaria cases. Additionally, climate changes profoundly affect malaria transmission and are responsible for malaria epidemics in some regions of South America. Parasite drug resistance is increasing, but due to bio-geographic barriers there is extraordinary genetic differentiation of parasites with limited dispersion. Although the clinical spectrum ranges from uncomplicated to severe malaria cases, due to the generally low to middle transmission intensity, features such as severe anemia, cerebral malaria and other complications appear to be less frequent than in other endemic regions and asymptomatic infections are a common feature. Although the National Malaria Control Programs (NMCP) of different countries differ in their control activities these are all directed to reduce morbidity and mortality by using strategies like health promotion, vector control and impregnate bed nets among others. Recently, international initiatives such as the Malaria Control Program in Andean-country Border Regions (PAMAFRO) (implemented by the Andean Organism for Health (ORAS) and sponsored by The Global Fund to Fight AIDS, Tuberculosis and Malaria (GFATM)) and The Amazon Network for the Surveillance of Antimalarial Drug Resistance (RAVREDA) (sponsored by the Pan American Health Organization/World Health Organization (PAHO/WHO) and several other partners), have made great investments for malaria control in the region. We describe here the current status of malaria in a non-Amazonian region comprising several countries of South and Central America participating in the Centro Latino Americano de Investigación en Malaria (CLAIM), an International Center of Excellence for Malaria Research (ICEMR) sponsored by the National Institutes of Health’s (NIH) National Institute of Allergy and Infectious Diseases (NIAID).
malaria; Plasmodium falciparum; Plasmodium vivax; malaria elimination; epidemiology; Latin America
Armenia, situated between the Black and Caspian Seas, lies at the junction of Turkey, Iran, Georgia, Azerbaijan and former Mesopotamia. This geographic position made it a potential contact zone between Eastern and Western civilizations. In this investigation, we assess Y-chromosomal diversity in four geographically distinct populations that represent the extent of historical Armenia. We find a striking prominence of haplogroups previously implicated with the Agricultural Revolution in the Near East, including the J2a-M410-, R1b1b1*-L23-, G2a-P15- and J1-M267-derived lineages. Given that the Last Glacial Maximum event in the Armenian plateau occured a few millennia before the Neolithic era, we envision a scenario in which its repopulation was achieved mainly by the arrival of farmers from the Fertile Crescent temporally coincident with the initial inception of farming in Greece. However, we detect very restricted genetic affinities with Europe that suggest any later cultural diffusions from Armenia to Europe were not associated with substantial amounts of paternal gene flow, despite the presence of closely related Indo-European languages in both Armenia and Southeast Europe.
Armenia; phylogenetics; Y-chromosome; SNPs; neolithic
Several studies have reported the following as determining factors for the adoption of healthy lifestyles among undergraduate students: gender, socioeconomic level, prior lifestyles, environment, parental lifestyles and health status, career choice, and healthy support networks. However, these factors are influenced by students’ knowledge about healthy lifestyles.
We will carry out a randomized trial in a sample of 280 new undergraduate students at the National Autonomous University of Mexico’s Faculty of Higher Studies-Zaragoza (FES-Zaragoza, UNAM). There will be an experimental group (n = 140), comprising 20 students from each of the seven university departments (careers); these students will receive training as university student health promoters through an e-learning course. This course will allow the topics necessary for such promoters to be reviewed. There will be a control group (n = 140), comprising 20 students from each of the seven departments (careers); these students will not undergo the training. Later, the students who comply satisfactorily with the e-learning course will replicate the course to 10 of their classmates. A healthy-lifestyle questionnaire will be given to all the participants, and the parameters established in the self-care card will be recorded before and after the training. The study variables are as follows: (i) independent variable—compliance with the e-learning course; (ii) dependent variables—lifestyles changes prior to the educative intervention (including healthy eating, physical activity, and addiction prevention) and parameters related to health status established in self-care (including weight, body mass index, waist circumference, and hip circumference). Data will be analyzed using Student’s t test and logistic regression analysis odds ratios with 95% confidence intervals. The analysis of the open answers will be carried out with ATLAS. ti 5.5 software.
Health promotion among university students should incorporate options that are feasible for and attractive to students. Thus, as proposed in the present protocol, e-learning courses offer excellent possibilities because they allow students to program their learning in their available time without affecting their academic studies.
University health promoter; University student health promoter; Self-care; Health promotion
This report describes the baseline characteristics of patients in the Reduction of Events with Darbepoetin alfa in Heart Failure trial (RED-HF) which is testing the hypothesis that anaemia correction with darbepoetin alfa will reduce the composite endpoint of death from any cause or hospital admission for worsening heart failure, and improve other outcomes.
Methods and results
Key demographic, clinical, and laboratory findings, along with baseline treatment, are reported and compared with those of patients in other recent clinical trials in heart failure. Compared with other recent trials, RED-HF enrolled more elderly [mean age 70 (SD 11.4) years], female (41%), and black (9%) patients. RED-HF patients more often had diabetes (46%) and renal impairment (72% had an estimated glomerular filtration rate <60 mL/min/1.73 m2). Patients in RED-HF had heart failure of longer duration [5.3 (5.4) years], worse NYHA class (35% II, 63% III, and 2% IV), and more signs of congestion. Mean EF was 30% (6.8%). RED-HF patients were well treated at randomization, and pharmacological therapy at baseline was broadly similar to that of other recent trials, taking account of study-specific inclusion/exclusion criteria. Median (interquartile range) haemoglobin at baseline was 112 (106–117) g/L.
The anaemic patients enrolled in RED-HF were older, moderately to markedly symptomatic, and had extensive co-morbidity.
Heart failure; Anaemia
The micropropagation is a powerful tool to scale up plants of economical and agronomical importance, enhancing crop productivity. However, a small but growing body of evidence suggests that epigenetic mechanisms, such as DNA methylation and histone modifications, can be affected under the in vitro conditions characteristic of micropropagation. Here, we tested whether the adaptation to different in vitro systems (Magenta boxes and Bioreactors) modified epigenetically different clones of Agave fourcroydes and A. angustifolia. Furthermore, we assessed whether these epigenetic changes affect the regulatory expression of KNOTTED1-like HOMEOBOX (KNOX) transcription factors.
To gain a better understanding of epigenetic changes during in vitro and ex vitro conditions in Agave fourcroydes and A. angustifolia, we analyzed global DNA methylation, as well as different histone modification marks, in two different systems: semisolid in Magenta boxes (M) and temporary immersion in modular Bioreactors (B). No significant difference was found in DNA methylation in A. fourcroydes grown in either M or B. However, when A. fourcroydes was compared with A. angustifolia, there was a two-fold difference in DNA methylation between the species, independent of the in vitro system used. Furthermore, we detected an absence or a low amount of the repressive mark H3K9me2 in ex vitro conditions in plants that were cultured earlier either in M or B. Moreover, the expression of AtqKNOX1 and AtqKNOX2, on A. fourcroydes and A. angustifolia clones, is affected during in vitro conditions. Therefore, we used Chromatin ImmunoPrecipitation (ChIP) to know whether these genes were epigenetically regulated. In the case of AtqKNOX1, the H3K4me3 and H3K9me2 were affected during in vitro conditions in comparison with AtqKNOX2.
Agave clones plants with higher DNA methylation during in vitro conditions were better adapted to ex vitro conditions. In addition, A. fourcroydes and A. angustifolia clones displayed differential expression of the KNOX1 gene during in vitro conditions, which is epigenetically regulated by the H3K4me3 and H3K9me2 marks. The finding of an epigenetic regulation in key developmental genes will make it important in future studies to identify factors that help to find climate-resistant micropropagated plants.
Epigenetics; In vitro; Histone methylation; Agave; KNOX genes
At less than 90 Mbp, the tiny nuclear genome of the carnivorous bladderwort plant Utricularia is an attractive model system for studying molecular evolutionary processes leading to genome miniaturization. Recently, we reported that expression of genes encoding DNA repair and reactive oxygen species (ROS) detoxification enzymes is highest in Utricularia traps, and we argued that ROS mutagenic action correlates with the high nucleotide substitution rates observed in the Utricularia plastid, mitochondrial, and nuclear genomes. Here, we extend our analysis of 100 nuclear genes from Utricularia and related asterid eudicots to examine nucleotide substitution biases and their potential correlation with ROS-induced DNA lesions. We discovered an unusual bias toward GC nucleotides, most prominently in transition substitutions at the third position of codons, which are presumably silent with respect to adaptation. Given the general tendency of biased gene conversion to drive GC bias, and of ROS to induce double strand breaks requiring recombinational repair, we propose that some of the unusual features of the bladderwort and its genome may be more reflective of these nonadaptive processes than of natural selection.
Carnivorous plant; DNA mutation; GC bias; Gene conversion; transcriptome
A safe and reproducible Plasmodium vivax infectious challenge method is required to evaluate the efficacy of malaria vaccine candidates. Seventeen healthy Duffy (+) and five Duffy (−) subjects were randomly allocated into three (A–C) groups and were exposed to the bites of 2–4 Anopheles albimanus mosquitoes infected with Plasmodium vivax derived from three donors. Duffy (−) subjects were included as controls for each group. Clinical manifestations of malaria and parasitemia were monitored beginning 7 days post-challenge. All Duffy (+) volunteers developed patent malaria infection within 16 days after challenge. Prepatent period determined by thick smear, was longer for Group A (median 14.5 d) than for Groups B and C (median 10 d/each). Infected volunteers recovered rapidly after treatment with no serious adverse events. The bite of as low as two P. vivax-infected mosquitoes provides safe and reliable infections in malaria-naive volunteers, suitable for assessing antimalarial and vaccine efficacy trials.
We assessed the safety, tolerability, and immunogenicity of a mixture of three synthetic peptides derived from the Plasmodium vivax circumsporozoite protein formulated in Montanide ISA 720 or Montanide ISA 51. Forty healthy malaria-naive volunteers were allocated to five experimental groups (A–E): four groups (A–D) were immunized intramuscularly with 50 and 100 μg/dose injections of a mixture of N, R, and C peptides formulated in the two different adjuvants at 0, 2, and 4 months and one group was administered placebo. Vaccines were immunogenic, safe, well tolerated, and no serious adverse events related to the vaccine occurred. Seroconversion occurred in > 90% of the vaccines and antibodies recognized the sporozoite protein on immunofluorescent antibody test. Vaccines in Montanide ISA 51 showed a higher sporozoite protein recognition and interferon production. Results encourage further testing of the vaccine protective efficacy.
Plasmodium vivax circumsporozoite (CS) protein is a leading malaria vaccine candidate previously assessed in animals and humans. Here, combinations of three synthetic polypeptides corresponding to amino (N), central repeat (R), and carboxyl (C) regions of the CS protein formulated in Montanide ISA 720 or Montanide ISA 51 adjuvants were assessed for immunogenicity in rodents and primates. BALB/c mice and Aotus monkeys were divided into test and control groups and were immunized three times with doses of 50 and 100 μg of vaccine or placebo. Antigen-specific antimalarial antibodies were determined by enzyme-linked immunosorbent assay, immunofluorescent antibody test, and IFN-γ responses by enzyme-linked immunosorbent spot (ELIspot). Both vaccine formulations were highly immunogenic in both species. Mice developed better antibody responses against C and R polypeptides, whereas the N polypeptide was more immunogenic in monkeys. Anti-peptide antibodies remained detectable for several months and recognized native proteins on sporozoites. Differences between Montanide ISA 720 and Montanide ISA 51 formulations were not significant.
Vaccine development for Plasmodium vivax malaria is underway. A model to assess the protective efficacy of vaccine candidates in humans is urgently needed. Given the lack of continuous P. vivax cultures, we developed a system to infect Anopheles albimanus mosquitoes using blood from P. vivax-infected patients and determined parameters for challenge of malaria-naive volunteers by mosquito bite. Absence of co-infections in parasitized blood was confirmed by tests consistent with blood bank screening. A total of 119 experiments were conducted using batches of 900–4,500 mosquitoes fed by an artificial membrane feeding method. Optimal conditions for mosquito probing and infection were determined. Presence of oocyst and sporozoites were assessed on Days 7–8 and 14–15, respectively, and conditions to choose batches of infected mosquitoes for sporozoite challenge were established. Procedures to infect volunteers took a 2-hour period including verification of inoculum dose. Anopheles albimanus mosquitoes represent a valuable resource for P. vivax sporozoite challenge of volunteers.
Plasmodium vivax circumsporozoite (CS) protein is a leading malaria vaccine candidate. We describe the characterization of specific immune responses induced in 21 malaria-naive volunteers vaccinated with long synthetic peptides derived from the CS protein formulated in Montanide ISA 720. Both antibody- and cell-mediated immune responses were analyzed. Antibodies were predominantly of IgG1 and IgG3 isotypes, recognized parasite proteins on the immunofluorescent antibody test, and partially blocked sporozoite invasion of hepatoma cell lines in vitro. Peripheral blood mononuclear cells from most volunteers (94%) showed IFN-γ production in vitro upon stimulation with both long signal peptide and short peptides containing CD8+ T-cell epitopes. The relatively limited sample size did not allow conclusions about HLA associations with the immune responses observed. In summary, the inherent safety and tolerability together with strong antibody responses, invasion blocking activity, and the IFN-γ production induced by these vaccine candidates warrants further testing in a phase II clinical trial.