H2A.Z Demarcates Intergenic Regions of the Plasmodium falciparum Epigenome That Are Dynamically Marked by H3K9ac and H3K4me3

Epigenetic regulatory mechanisms and their enzymes are promising targets for malaria therapeutic intervention; however, the epigenetic component of gene expression in P. falciparum is poorly understood. Dynamic or stable association of epigenetic marks with genomic features provides important clues about their function and helps to understand how histone variants/modifications are used for indexing the Plasmodium epigenome. We describe a novel, linear amplification method for next-generation sequencing (NGS) that allows unbiased analysis of the extremely AT-rich Plasmodium genome. We used this method for high resolution, genome-wide analysis of a histone H2A variant, H2A.Z and two histone H3 marks throughout parasite intraerythrocytic development. Unlike in other organisms, H2A.Z is a constant, ubiquitous feature of euchromatic intergenic regions throughout the intraerythrocytic cycle. The almost perfect colocalisation of H2A.Z with H3K9ac and H3K4me3 suggests that these marks are preferentially deposited on H2A.Z-containing nucleosomes. By performing RNA-seq on 8 time-points, we show that acetylation of H3K9 at promoter regions correlates very well with the transcriptional status whereas H3K4me3 appears to have stage-specific regulation, being low at early stages, peaking at trophozoite stage, but does not closely follow changes in gene expression. Our improved NGS library preparation procedure provides a foundation to exploit the malaria epigenome in detail. Furthermore, our findings place H2A.Z at the cradle of P. falciparum epigenetic regulation by stably defining intergenic regions and providing a platform for dynamic assembly of epigenetic and other transcription related complexes

Genome of the Avirulent Human-Infective Trypanosome—Trypanosoma rangeli

Background: Trypanosoma rangeli is a hemoflagellate protozoan parasite infecting humans and other wild and domestic mammals across Central and South America. It does not cause human disease, but it can be mistaken for the etiologic agent of Chagas disease, Trypanosoma cruzi. We have sequenced the T. rangeli genome to provide new tools for elucidating the distinct and intriguing biology of this species and the key pathways related to interaction with its arthropod and mammalian hosts.  Methodology/Principal Findings: The T. rangeli haploid genome is ,24 Mb in length, and is the smallest and least repetitive trypanosomatid genome sequenced thus far. This parasite genome has shorter subtelomeric sequences compared to those of T. cruzi and T. brucei; displays intraspecific karyotype variability and lacks minichromosomes. Of the predicted 7,613 protein coding sequences, functional annotations could be determined for 2,415, while 5,043 are hypothetical proteins, some with evidence of protein expression. 7,101 genes (93%) are shared with other trypanosomatids that infect humans. An ortholog of the dcl2 gene involved in the T. brucei RNAi pathway was found in T. rangeli, but the RNAi machinery is non-functional since the other genes in this pathway are pseudogenized. T. rangeli is highly susceptible to oxidative stress, a phenotype that may be explained by a smaller number of anti-oxidant defense enzymes and heatshock proteins.  Conclusions/Significance: Phylogenetic comparison of nuclear and mitochondrial genes indicates that T. rangeli and T. cruzi are equidistant from T. brucei. In addition to revealing new aspects of trypanosome co-evolution within the vertebrate and invertebrate hosts, comparative genomic analysis with pathogenic trypanosomatids provides valuable new information that can be further explored with the aim of developing better diagnostic tools and/or therapeutic targets

Polarizable Water Model for the Coarse-Grained MARTINI Force Field

Coarse-grained (CG) simulations have become an essential tool to study a large variety of biomolecular processes, exploring temporal and spatial scales inaccessible to traditional models of atomistic resolution. One of the major simplifications of CG models is the representation of the solvent, which is either implicit or modeled explicitly as a van der Waals particle. The effect of polarization, and thus a proper screening of interactions depending on the local environment, is absent. Given the important role of water as a ubiquitous solvent in biological systems, its treatment is crucial to the properties derived from simulation studies. Here, we parameterize a polarizable coarse-grained water model to be used in combination with the CG MARTINI force field. Using a three-bead model to represent four water molecules, we show that the orientational polarizability of real water can be effectively accounted for. This has the consequence that the dielectric screening of bulk water is reproduced. At the same time, we parameterized our new water model such that bulk water density and oil/water partitioning data remain at the same level of accuracy as for the standard MARTINI force field. We apply the new model to two cases for which current CG force fields are inadequate. First, we address the transport of ions across a lipid membrane. The computed potential of mean force shows that the ions now naturally feel the change in dielectric medium when moving from the high dielectric aqueous phase toward the low dielectric membrane interior. In the second application we consider the electroporation process of both an oil slab and a lipid bilayer. The electrostatic field drives the formation of water filled pores in both cases, following a similar mechanism as seen with atomistically detailed models

Cell-Autonomous Requirement for Rx Function in the Mammalian Retina and Posterior Pituitary

Rx is a paired-like homeobox gene that is required for vertebrate eye formation. Mice lacking Rx function do not develop eyes or the posterior pituitary. To determine whether Rx is required cell autonomously in these tissues, we generated embryonic chimeras consisting of wild type and Rx−/− cells. We found that in the eye, Rx-deficient cells cannot participate in the formation of the neuroretina, retina pigment epithelium and the distal part of the optic stalk. In addition, in the ventral forebrain, Rx function is required cell autonomously for the formation of the posterior pituitary. Interestingly, Rx−/− and wild type cells segregate before the morphogenesis of these two tissues begins. Our observations suggest that Rx function is not only required for the morphogenesis of the retina and posterior pituitary, but also prior to morphogenesis, for the sorting out of cells to form distinct fields of retinal/pituitary cells

A922 Sequential measurement of 1 hour creatinine clearance (1-CRCL) in critically ill patients at risk of acute kidney injury (AKI)

Meeting abstrac

Riociguat treatment in patients with chronic thromboembolic pulmonary hypertension: Final safety data from the EXPERT registry

Objective: The soluble guanylate cyclase stimulator riociguat is approved for the treatment of adult patients with pulmonary arterial hypertension (PAH) and inoperable or persistent/recurrent chronic thromboembolic pulmonary hypertension (CTEPH) following Phase

Non-standard errors

In statistics, samples are drawn from a population in a data-generating process (DGP). Standard errors measure the uncertainty in estimates of population parameters. In science, evidence is generated to test hypotheses in an evidence generating process (EGP). We claim that EGP variation across researchers adds uncertainty: Non-standard errors (NSEs). We study NSEs by letting 164 teams test the same hypotheses on the same data. NSEs turn out to be sizable, but smaller for better reproducible or higher rated research. Adding peer-review stages reduces NSEs. We further find that this type of uncertainty is underestimated by participants

Clinical aptitude of family physicians to detect familial dysfunction in family medical units from Guadalajara, Mexico [Aptitud clínica de los médicos familiares en la identificación de la disfunción familiar en unidades de medicina familiar de Guadalajara, México]

Introduction: Familles function as dynamic systems, where the different members stimulate each other to achieve common objectives. Family development is thus conceived as a chain of changes, in form and function, which follow evolution stages. The balance of positive and negative forces inside the family is translated into an evolution, and so the ability to respond to internal or external changes is vital to avoid discrepancies or clashes between the tasks and roles of the different family members. In this sense, family physicians must be able to identify any potential dysfunction or difficulty inside the family, and to facilitate the compatibility of tasks in order to reestablish the continuity and good functioning of the family. The family physicians' clinical aptitude is made up of a series of abilities intended to identify any signs and symptoms of dysfunction. They must also be skilled in using auxiliary resources for the diagnosis and treatment of all these. Such an aptitude is measured by a structured and validated instrument. Material and methods: This is an observational, prospective and comparative research of a 450 family physicians sample from 23 first level family health care medical clinics from the National Social Security Institute (Instituto Mexicano del Seguro Social: IMSS). All family physicians working at the clinics were included, excepting those who were at the time on vacation, worked the night shift, attended the ER, were absent or refused to participate. Clinical aptitude for family dysfunction was measured in three areas: 1. Identification of risk factors for family dysfunction; 2. Diagnosis with an integral point of view and 3. Proposal, identification and guidance, which describes a physician's ability to judge decisions taken on case reports and to propose alternative actions. Other variables taken into account were sex, age, specialty, years of experience, shift, clinic and type of contract. Instruments. The instrument was designed to integrate theory and practice. It was conformed by real case reports, which were condensed, divided in sections, and followed by a series of questions with three possible answers: "true", "false" or "I don't know". In total, there were 187 questions, 94 of which were true and 93, false. Correct answers accounted for one point, while incorrect ones rested one point; "I don't know" answers had no effect on the results. There were 42 lines to explore risk factors; 24 to explore the use of diagnostic resources; 19 to explore the use of therapeutic resources; 36 to evaluate a physician's knowledge of family sociology; 42 to assess family psychology, and 24 designed to evaluate proposal abilities. It was all validated and standardized with a group of post-graduate medical residents in Family Medicine from Mexico City. The Richardson K index was 0.90. Clinical aptitude was measured using an ordinal scale, where a random level "1 was defined by 140 points. A descriptive and inferential statistical analysis was used with median, percentage, Mann-Whitney's and Kruskal-Wallis' tests. All this was then processed with the EPI INFO-6 and SPSS Plus software packages. Ethical considerations. This is a risk-free research, as established in the Health Research section of the Mexican Health Law. Nevertheless, a signed acceptance form was required from all participants. Results: Table 1 shows the general characteristics of the study sample. In turn, table 2 presents clinical aptitude to identify family dysfunction, sorted by clinic. Clinics B and D had, respectively, a median of 105 and 102, with similar ranges. There were no statistically signicant differences among the sub-indexes of each clinic. The diagnosis median was higher than that for guidance. Table 3 reveals a high level of clinical aptitude in 3% of the physicians, an intermediate level in 25%, a low level in 58%, and a random-defined level in 14%. There were no significant differences when clinical aptitude was correlated with sex, shift, type of contract, specialty and years of experience. Discussion. The main objective of a family dysfunction diagnosis is to reestablish the normal flow of a family's vital cycle with the support of a specialized physician. It has been reported that clinical aptitude measurement is useful to discriminate and establish the aptitude level of experiment and non-experiment physicians with the aim of creating educational opportunities. A slight advantage, with no significant difference, was appreciated in physicians who attended patients in their offices, which suggests they are in a better position to gain a higher level of trust from their patients. This is due to the fact that they attend a regular set group of patients assigned to their offices. Since the education of family physicians is aimed at offering an integral care to families, family dysfunction recognition is essential. Results also suggest a non-significant advantage from family physicians with curricular education (not all family physicians working at the IMSS have a degree in Family Medicine). Experienced physicians (10 to 19 years on the job) showed another non-significant advantage, which pointed to the value of clinical practice. This is a powerful reason to promote continuous educational programs for family physicians. Family physicians who worked the morning shift showed a non-statistical advantage over their afternoon shift counterparts. This could be explained by the fact that educational and other institutional activities are more likely to take place in the morning. The educational model of family physicians should promote the physicians' involvement in understanding how to become active elements in gathering their own knowledge. Such a model should promote physicians' initiatives for the development of an experience based on constructive critic. The current health care model is mainly focused on a biological interpretation of the health-disease process. However, this is only a partial approach which prevents the implementation of an integral clinical practice. From our research, we expect changes in institutional health care orientation and a reframing of the curricula of general and family physicians'. Although the acquisition of clinical aptitude requires the physicians' experience and involvement in developing their own knowledge, our results do not reflect this ideal condition. This is due to the low percentage of clinical aptitude, which correlates with an evident inability for research and interpretation. Half of the physicians were capable of elaborating diagnostic hypothesis and two thirds of them showed a adequate use of diagnostic resources, such as clinical tests, functional family diagnostic instruments and a guide to conform an integral family workup. All these should be useful educational tools to establish the social functions diagnosis of a family's members, together with their formal and informal roles and their importance in the health-disease process. Guidance requires the ability to judge decisions taken by other professionals and make suggestions for alternative actions in case reports. This latter skill includes the use of therapeutic resources for only less than half of the physicians know how to properly use these resources. The use of instruments to measure aptitude, competency and work performance is a growing practice in continuous education and human resources formation. Even though these instruments are capable of discriminating high clinical aptitude, they cannot be used to account for this non-significant advantage, because educational activities are conceived as the consumption of information and not as the acquisition of it from each one's experience. Overall, 58% of the family physicians showed a low level of clinical aptitude. Such a result reflects a poor ability to integrate daily experience