Radial spoke protein 9 is necessary for axoneme assembly in Plasmodium but not in trypanosomatid parasites.

Flagella are important for eukaryote cell motility, including in sperm, and are vital for life cycle progression of many unicellular eukaryotic pathogens. The '9+2' axoneme in most motile flagella comprises nine outer doublet and two central-pair singlet microtubules. T-shaped radial spokes protrude from the outer doublets towards the central pair and are necessary for effective beating. We asked whether there were radial spoke adaptations associated with parasite lineage-specific properties in apicomplexans and trypanosomatids. Following an orthologue search for experimentally uncharacterised radial spoke proteins (RSPs), we identified and analysed RSP9. Trypanosoma brucei and Leishmania mexicana have an extensive RSP complement, including two divergent RSP9 orthologues, necessary for flagellar beating and swimming. Detailed structural analysis showed that neither orthologue is needed for axoneme assembly in Leishmania. In contrast, Plasmodium has a reduced set of RSPs including a single RSP9 orthologue, deletion of which in Plasmodium berghei leads to failure of axoneme formation, failed male gamete release, greatly reduced fertilisation and inefficient life cycle progression in the mosquito. This indicates contrasting selection pressures on axoneme complexity, likely linked to the different mode of assembly of trypanosomatid versus Plasmodium flagella.

Lab-on-Valve Automated and Miniaturized Assessment of Nanoparticle Concentration Based on Light-Scattering.

Nanoparticles (NPs) concentration directly impacts the dose delivered to target tissues by nanocarriers. The evaluation of this parameter is required during NPs developmental and quality control stages, for setting dose-response correlations and for evaluating the reproducibility of the manufacturing process. Still, faster and simpler procedures, dismissing skilled operators and post-analysis conversions are needed to quantify NPs for research and quality control operations, and to support result validation. Herein, a miniaturized automated ensemble method to measure NPs concentration was established under the lab-on-valve (LOV) mesofluidic platform. Automatic NPs sampling and delivery to the LOV detection unit were set by flow programming. NPs concentration measurements were based on the decrease in the light transmitted to the detector due to the light scattered by NPs when passing through the optical path. Each analysis was accomplished in 2 min, rendering a determination throughput of 30 h-1 (6 samples h-1 for n = 5) and only requiring 30 µL (≈0.03 g) of NPs suspension. Measurements were performed on polymeric NPs, as these represent one of the major classes of NPs under development for drug-delivery aims. Determinations for polystyrene NPs (of 100, 200, and 500 nm) and for NPs made of PEGylated poly-d,l-lactide-co-glycolide (PEG-PLGA, a biocompatible FDA-approved polymer) were accomplished within 108-1012 particles mL-1 range, depending on the NPs size and composition. NPs size and concentration were maintained during analysis, as verified for NPs eluted from the LOV by particle tracking analysis (PTA). Moreover, concentration measurements for PEG-PLGA NPs loaded with an anti-inflammatory drug, methotrexate (MTX), after their incubation in simulated gastric and intestinal fluids were successfully achieved (recovery values of 102-115%, as confirmed by PTA), showing the suitability of the proposed method to support the development of polymeric NPs targeting intestinal delivery.

Rapid and sustainable HPLC method for the determination of uremic toxins in human plasma samples.

Protein-bound uremic toxins, mainly indoxyl sulfate (3-INDS), p-cresol sulfate (pCS), and indole-3-acetic acid (3-IAA) but also phenol (Pol) and p-cresol (pC), are progressively accumulated during chronic kidney disease (CKD). Their accurate measurement in biomatrices is demanded for timely diagnosis and adoption of appropriate therapeutic measures. Multianalyte methods allowing the establishment of a uremic metabolite profile are still missing. Hence, the aim of this work was to develop a rapid and sensitive method based on high-performance liquid chromatography with fluorescence detection for the simultaneous quantification of Pol, 3-IAA, pC, 3-INDS, and pCS in human plasma. Separation was attained in 12 min, using a monolithic C18 column and isocratic elution with acetonitrile and phosphate buffer containing an ion-pairing reagent, at a flow rate of 2 mL min-1. Standards were prepared in plasma and quantification was performed using the background subtraction approach. LOQ values were ≤ 0.2 µg mL-1 for all analytes except for pCS (LOQ of 2 µg mL-1). The method proved to be accurate (93.5-112%) and precise (CV ≤ 14.3%). The multianalyte application of the method, associated to a reduced sample volume (50 µL), a less toxic internal standard (eugenol) in comparison to the previously applied 2,6-dimethylphenol and 4-ethylphenol, and a green extraction solvent (ethanol), resulted in the AGREE score of 0.62 which is in line with the recent trend of green and sustainable analytical chemistry. The validated method was successfully applied to the analysis of plasma samples from control subjects exhibiting normal levels of uremic toxins and CKD patients presenting significantly higher levels of 3-IAA, pC, 3-INDS, and pCS that can be further investigated as biomarkers of disease progression.

Cryo-XPS for Surface Characterization of Nanomedicines.

Nanoparticles used for medical applications commonly possess coatings or surface functionalities intended to provide specific behavior in vivo, for example, the use of PEG to provide stealth properties. Direct, quantitative measurement of the surface chemistry and composition of such systems in a hydrated environment has thus far not been demonstrated, yet such measurements are of great importance for the development of nanomedicine systems. Here we demonstrate the first use of cryo-XPS for the measurement of two PEG-functionalized nanomedicines: a polymeric drug delivery system and a lipid nanoparticle mRNA carrier. The observed differences between cryo-XPS and standard XPS measurements indicate the potential of cryo-XPS for providing quantitative measurements of such nanoparticle systems in hydrated conditions.

Vital role for Plasmodium berghei Kinesin8B in axoneme assembly during male gamete formation and mosquito transmission.

Sexual development is an essential phase in the Plasmodium life cycle, where male gametogenesis is an unusual and extraordinarily rapid process. It produces 8 haploid motile microgametes, from a microgametocyte within 15 minutes. Its unique achievement lies in linking the assembly of 8 axonemes in the cytoplasm to the three rounds of intranuclear genome replication, forming motile microgametes, which are expelled in a process called exflagellation. Surprisingly little is known about the actors involved in these processes. We are interested in kinesins, molecular motors that could play potential roles in male gametogenesis. We have undertaken a functional characterization in Plasmodium berghei of kinesin-8B (PbKIN8B) expressed specifically in male gametocytes and gametes. By generating Pbkin8B-gfp parasites, we show that PbKIN8B is specifically expressed during male gametogenesis and is associated with the axoneme. We created a ΔPbkin8B knockout cell line and analysed the consequences of the absence of PbKIN8B on male gametogenesis. We show that the ability to produce sexually differentiated gametocytes is not affected in ΔPbkin8B parasites and that the 3 rounds of genome replication occur normally. Nevertheless, the development to free motile microgametes is halted and the life cycle is interrupted in vivo. Ultrastructural analysis revealed that intranuclear mitoses are unaffected whereas cytoplasmic microtubules, although assembled in doublets and elongated, fail to assemble in the normal axonemal '9+2' structure and become motile. Absence of a functional axoneme prevented microgamete assembly and release from the microgametocyte, severely reducing infection of the mosquito vector. This is the first functional study of a kinesin involved in male gametogenesis. These results reveal a previously unknown role for PbKIN8B in male gametogenesis, providing new insights into Plasmodium flagellar formation.

FGF signaling enforces cardiac chamber identity in the developing ventricle.

Atrial and ventricular cardiac chambers behave as distinct subunits with unique morphological, electrophysiological and contractile properties. Despite the importance of chamber-specific features, chamber fate assignments remain relatively plastic, even after differentiation is underway. In zebrafish, Nkx transcription factors are essential for the maintenance of ventricular characteristics, but the signaling pathways that operate upstream of Nkx factors in this context are not well understood. Here, we show that FGF signaling plays an essential part in enforcing ventricular identity. Loss of FGF signaling results in a gradual accumulation of atrial cells, a corresponding loss of ventricular cells, and the appearance of ectopic atrial gene expression within the ventricle. These phenotypes reflect important roles for FGF signaling in promoting ventricular traits, both in early-differentiating cells that form the initial ventricle and in late-differentiating cells that append to its arterial pole. Moreover, we find that FGF signaling functions upstream of Nkx genes to inhibit ectopic atrial gene expression. Together, our data suggest a model in which sustained FGF signaling acts to suppress cardiomyocyte plasticity and to preserve the integrity of the ventricular chamber.

Insights on Ultrafiltration-Based Separation for the Purification and Quantification of Methotrexate in Nanocarriers.

The evaluation of encapsulation efficiency is a regulatory requirement for the characterization of drug delivery systems. However, the difficulties in efficiently separating nanomedicines from the free drug may compromise the achievement of accurate determinations. Herein, ultrafiltration was exploited as a separative strategy towards the evaluation of methotrexate (MTX) encapsulation efficiency in nanostructured lipid carriers and polymeric nanoparticles. The effect of experimental conditions such as pH and the amount of surfactant present in the ultrafiltration media was addressed aiming at the selection of suitable conditions for the effective purification of nanocarriers. MTX-loaded nanoparticles were then submitted to ultrafiltration and the portions remaining in the upper compartment of the filtering device and in the ultrafiltrate were collected and analyzed by HPLC-UV using a reversed-phase (C18) monolithic column. A short centrifugation time (5 min) was suitable for establishing the amount of encapsulated MTX in nanostructured lipid carriers, based on the assumption that the free MTX concentration was the same in the upper compartment and in the ultrafiltrate. The defined conditions allowed the efficient separation of nanocarriers from the free drug, with recoveries of >85% even when nanoparticles were present in cell culture media and in pig skin surrogate from permeation assays.

CCBE1 is required for coronary vessel development and proper coronary artery stem formation in the mouse heart.

BACKGROUND: Proper coronary vasculature development is essential for late-embryonic and adult heart function. The developmental regulation of coronary embryogenesis is complex and includes the coordinated activity of multiple signaling pathways. CCBE1 plays an important role during lymphangiogenesis, enhancing VEGF-C signaling, which is also required for coronary vasculature formation. However, whether CCBE1 plays a similar role during coronary vasculature development is still unknown. Here, we investigate the coronary vasculature development in Ccbe1 mutant embryos. RESULTS: We show that Ccbe1 is expressed in the epicardium, like Vegf-c, and also in the sinus venosus (SV) at the stages of its contribution to coronary vasculature formation. We also report that absence of CCBE1 in cardiac tissue inhibited coronary growth that sprouts from the SV endocardium at the dorsal cardiac wall. This disruption of coronary formation correlates with abnormal processing of VEGF-C propeptides, suggesting VEGF-C-dependent signaling alteration. Moreover, Ccbe1 loss-of-function leads to the development of defective dorsal and ventral intramyocardial vessels. We also demonstrate that Ccbe1 mutants display delayed and mispatterned coronary artery (CA) stem formation. CONCLUSIONS: CCBE1 is essential for coronary vessel formation, independent of their embryonic origin, and is also necessary for peritruncal vessel growth and proper CA stem patterning. Developmental Dynamics 247:1135-1145, 2018. © 2018 Wiley Periodicals, Inc.

Diagnosis of sleep apnea in patients with stable chronic heart failure using a portable sleep test diagnostic device.

PURPOSE: ApneaLink is a portable device for the screening of sleep apnea, a prevalent and underdiagnosed comorbidity in heart failure patients. A prospective cross-sectional study in patients with chronic heart failure was carried out to assess the sensitivity and specificity of apnea-hypopnea index (AHI) measurements using ApneaLink against the standard polysomnography test. METHODS: Adult patients with a prior hospitalization in an acute heart failure hospital unit were recruited for the study. All participants were tested for sleep apnea using ApneaLink and polysomnography simultaneously during an overnight stay at a sleep laboratory. Global sleep apnea was evaluated according to the AHI, which was analyzed and compared. Subpopulation comparison based on ejection fraction was not realized due to population size. RESULTS: Thirty-five patients with stable chronic heart failure completed the study (mean age 70.9 ± 10.5 years and body mass index 30.0 ± 4.7 kg/m2). Two patients were excluded due to insufficient study duration. ApneaLink had a sensitivity greater than 80% for all AHI measurements, and a specificity greater than 80% for all AHI measurements, except for AHI ≥ 5 events/h (61.5%). The results showed higher sensitivities and specificities at AHI values of ≥ 10 events/h (sensitivity 81.3% and specificity 84.2%) and ≥ 15 events/h (sensitivity 83.3% and specificity 91.3%). Correlation analysis showed that AHI measurements using ApneaLink and polysomnography had a strong and significant correlation (r = 0.794; P < 0.001). CONCLUSIONS: Our results suggest that ApneaLink could be used in clinical practice to identify heart failure patients with high (AHI ≥ 15 events/h) and low (AHI < 5 events/h) probability of having sleep apnea, sparing the need for a diagnostic polysomnography and thus potentially impacting prognosis by providing a more cost-effective and timely diagnosis of this non-cardiac comorbidity.

Systematic review of pediatric health outcomes associated with childhood adversity.

BACKGROUND: Early detection of and intervention in childhood adversity has powerful potential to improve the health and well-being of children. A systematic review was conducted to better understand the pediatric health outcomes associated with childhood adversity. METHODS: PubMed, PsycArticles, and CINAHL were searched for relevant articles. Longitudinal studies examining various adverse childhood experiences and biological health outcomes occurring prior to age 20 were selected. Mental and behavioral health outcomes were excluded, as were physical health outcomes that were a direct result of adversity (i.e. abusive head trauma). Data were extracted and risk of bias was assessed by 2 independent reviewers. RESULTS: After identifying 15940 records, 35 studies were included in this review. Selected studies indicated that exposure to childhood adversity was associated with delays in cognitive development, asthma, infection, somatic complaints, and sleep disruption. Studies on household dysfunction reported an effect on weight during early childhood, and studies on maltreatment reported an effect on weight during adolescence. Maternal mental health issues were associated with elevated cortisol levels, and maltreatment was associated with blunted cortisol levels in childhood. Furthermore, exposure to childhood adversity was associated with alterations of immune and inflammatory response and stress-related accelerated telomere erosion. CONCLUSION: Childhood adversity affects brain development and multiple body systems, and the physiologic manifestations can be detectable in childhood. A history of childhood adversity should be considered in the differential diagnosis of developmental delay, asthma, recurrent infections requiring hospitalization, somatic complaints, and sleep disruption. The variability in children's response to adversity suggests complex underlying mechanisms and poses a challenge in the development of uniform diagnostic guidelines. More large longitudinal studies are needed to better understand how adversity, its timing and severity, and the presence of individual genetic, epigenetic, and protective factors affects children's health and development.

Sng1 associates with Nce102 to regulate the yeast Pkh-Ypk signalling module in response to sphingolipid status.

All cells are delimited by biological membranes, which are consequently a primary target of stress-induced damage. Cold alters membrane functionality by decreasing lipid fluidity and the activity of membrane proteins. In Saccharomyces cerevisiae, evidence links sphingolipid homeostasis and membrane phospholipid asymmetry to the activity of the Ypk1/2 proteins, the yeast orthologous of the mammalian SGK1-3 kinases. Their regulation is mediated by different protein kinases, including the PDK1 orthologous Pkh1/2p, and requires the function of protein effectors, among them Nce102p, a component of the sphingolipid sensor machinery. Nevertheless, the mechanisms and the actors involved in Pkh/Ypk regulation remain poorly defined. Here, we demonstrate that Sng1, a transmembrane protein, is an effector of the Pkh/Ypk module and identify the phospholipid asymmetry as key for yeast cold adaptation. Overexpression of SNG1 impairs phospholipid flipping, reduces reactive oxygen species (ROS) and improves, in a Pkh-dependent manner, yeast growth in myriocin-treated cells, suggesting that excess Sng1p stimulates the Pkh/Ypk signalling. Furthermore, we link these effects to the association of Sng1p with Nce102p. Indeed, we found that Sng1p interacts with Nce102p both physically and genetically. Moreover, mutant nce102∆ sng1∆ cells show features of impaired Pkh/Ypk signalling, including increased ROS accumulation, reduced life span and defects in Pkh/Ypk-controlled regulatory pathways. Finally, myriocin-induced hyperphosphorylation of Ypk1p and Orm2p, which controls sphingolipid homeostasis, does not occur in nce102∆ sng1∆ cells. Hence, both Nce102p and Sng1p participate in a regulatory circuit that controls the activity of the Pkh/Ypk module and their function is required in response to sphingolipid status.

Proposal for a unified classification system and nomenclature of lagoviruses.

Lagoviruses belong to the Caliciviridae family. They were first recognized as highly pathogenic viruses of the European rabbit (Oryctolagus cuniculus) and European brown hare (Lepus europaeus) that emerged in the 1970-1980s, namely, rabbit haemorrhagic disease virus (RHDV) and European brown hare syndrome virus (EBHSV), according to the host species from which they had been first detected. However, the diversity of lagoviruses has recently expanded to include new related viruses with varying pathogenicity, geographic distribution and host ranges. Together with the frequent recombination observed amongst circulating viruses, there is a clear need to establish precise guidelines for classifying and naming lagovirus strains. Therefore, here we propose a new nomenclature based on phylogenetic relationships. In this new nomenclature, a single species of lagovirus would be recognized and called Lagovirus europaeus. The species would be divided into two genogroups that correspond to RHDV- and EBHSV-related viruses, respectively. Genogroups could be subdivided into genotypes, which could themselves be subdivided into phylogenetically well-supported variants. Based on available sequences, pairwise distance cutoffs have been defined, but with the accumulation of new sequences these cutoffs may need to be revised. We propose that an international working group could coordinate the nomenclature of lagoviruses and any proposals for revision.

Phakic Intraocular Lens Implantation: Refractive Outcome and Safety in Patients with Anterior Chamber Depth between 2.8 and 3.0 versus ≥3.0 mm.

PURPOSE: To compare endothelial cell (EC) variation after anterior chamber phakic intraocular lens (AC-pIOL) implantation in highly myopic patients with a preoperative anterior chamber depth (ACD) between 2.8 and 3.0 versus ≥3.0 mm. METHODS: A total of 280 eyes submitted to primary AC-pIOL implantation were analyzed. Pre- and postoperative values for uncorrected distance visual acuity, corrected distance visual acuity, spherical equivalent, ACD (endothelial surface), and EC count were collected. The eyes were divided into 2 groups: group A - ACD between 2.8 and 3.0 mm; group B - ACD ≥3.0 mm. Mean global EC loss (ECL) and loss for each ACD group, according to pIOL type, were analyzed. RESULTS: Significant improvement of the spherical equivalent (-11.38 ± 4.57 vs. -0.49 ± 0.79; p = 0.000) and a significant decrease in EC density (2,810.95 ± 343.88 vs. 2,584.09 ± 374.88 cells/mm2; p = 0.000) were noted. The mean annual ECL was -2.19 ± 3.97%. Regarding group A (n = 80), a mean annual ECL of -2.06 ± 3.88% was registered, higher for the Acrysof Cachet® subtype, while group B (n = 200) showed -2.25 ± 4.01% ECL, higher for the Verisyse® subtype. There was no significant difference between the groups (p = 0.96). CONCLUSIONS: AC-pIOL implantation significantly improves the spherical equivalent in myopic patients. The mean annual ECL after pIOL implantation was higher in the larger ACD group, but this value was not statistically significant. A 2.8-mm ACD value seems to be a safe cutoff for AC-pIOL implantation.

Social and economic value of Portuguese community pharmacies in health care.

BACKGROUND: Community pharmacies are major contributors to health care systems across the world. Several studies have been conducted to evaluate community pharmacies services in health care. The purpose of this study was to estimate the social and economic benefits of current and potential future community pharmacies services provided by pharmacists in health care in Portugal. METHODS: The social and economic value of community pharmacies services was estimated through a decision-model. Model inputs included effectiveness data, quality of life (QoL) and health resource consumption, obtained though literature review and adapted to Portuguese reality by an expert panel. The estimated economic value was the result of non-remunerated pharmaceutical services plus health resource consumption potentially avoided. Social and economic value of community pharmacies services derives from the comparison of two scenarios: "with service" versus "without service". RESULTS: It is estimated that current community pharmacies services in Portugal provide a gain in QoL of 8.3% and an economic value of 879.6 million euros (M€), including 342.1 M€ in non-remunerated pharmaceutical services and 448.1 M€ in avoided expense with health resource consumption. Potential future community pharmacies services may provide an additional increase of 6.9% in QoL and be associated with an economic value of 144.8 M€: 120.3 M€ in non-remunerated services and 24.5 M€ in potential savings with health resource consumption. CONCLUSIONS: Community pharmacies services provide considerable benefit in QoL and economic value. An increase range of services including a greater integration in primary and secondary care, among other transversal services, may add further social and economic value to the society.

An essential role of the basal body protein SAS-6 in Plasmodium male gamete development and malaria transmission.

Gametocytes are the sole Plasmodium parasite stages that infect mosquitoes; therefore development of functional gametes is required for malaria transmission. Flagellum assembly of the Plasmodium male gamete differs from that of most other eukaryotes in that it is intracytoplasmic but retains a key conserved feature: axonemes assemble from basal bodies. The centriole/basal body protein SAS-6 normally regulates assembly and duplication of these organelles and its depletion causes severe flagellar/ciliary abnormalities in a diverse array of eukaryotes. Since basal body and flagellum assembly are intimately coupled to male gamete development in Plasmodium, we hypothesized that SAS-6 disruption may cause gametogenesis defects and perturb transmission. We show that Plasmodium berghei sas6 knockouts display severely abnormal male gametogenesis presenting reduced basal body numbers, axonemal assembly defects and abnormal nuclear allocation. The defects in gametogenesis reduce fertilization and render Pbsas6 knockouts less infectious to mosquitoes. Additionally, we show that lack of Pbsas6 blocks transmission from mosquito to vertebrate host, revealing an additional yet undefined role in ookinete to sporulating oocysts transition. These findings underscore the vulnerability of the basal body/SAS-6 to malaria transmission blocking interventions.

Fully automatic flow-based device for monitoring of drug permeation across a cell monolayer.

A novel flow-programming setup based on the sequential injection principle is herein proposed for on-line monitoring of temporal events in cell permeation studies. The permeation unit consists of a Franz cell with its basolateral compartment mixed under mechanical agitation and thermostated at 37 °C. The apical compartment is replaced by commercially available Transwell inserts with a precultivated cell monolayer. The transport of drug substances across epithelial cells genetically modified with the P-glycoprotein membrane transporter (MDCKII-MDR1) is monitored on-line using rhodamine 123 as a fluorescent marker. The permeation kinetics of the marker is obtained in a fully automated mode by sampling minute volumes of solution from the basolateral compartment in short intervals (10 min) up to 4 h. The effect of a P-glycoprotein transporter inhibitor, verapamil as a model drug, on the efficiency of the marker transport across the cell monolayer is thoroughly investigated. The analytical features of the proposed flow method for cell permeation studies in real time are critically compared against conventional batch-wise procedures and microfluidic devices.

Imaging-based high-throughput screening assay to identify new molecules with transmission-blocking potential against Plasmodium falciparum female gamete formation.

In response to a call for the global eradication of malaria, drug discovery has recently been extended to identify compounds that prevent the onward transmission of the parasite, which is mediated by Plasmodium falciparum stage V gametocytes. Lately, metabolic activity has been used in vitro as a surrogate for gametocyte viability; however, as gametocytes remain relatively quiescent at this stage, their ability to undergo onward development (gamete formation) may be a better measure of their functional viability. During gamete formation, female gametocytes undergo profound morphological changes and express translationally repressed mRNA. By assessing female gamete cell surface expression of one such repressed protein, Pfs25, as the readout for female gametocyte functional viability, we developed an imaging-based high-throughput screening (HTS) assay to identify transmission-blocking compounds. This assay, designated the P. falciparum female gametocyte activation assay (FGAA), was scaled up to a high-throughput format (Z' factor, 0.7 ± 0.1) and subsequently validated using a selection of 50 known antimalarials from diverse chemical families. Only a few of these agents showed submicromolar 50% inhibitory concentrations in the assay: thiostrepton, methylene blue, and some endoperoxides. To determine the best conditions for HTS, a robustness test was performed with a selection of the GlaxoSmithKline Tres Cantos Antimalarial Set (TCAMS) and the final screening conditions for this library were determined to be a 2 µM concentration and 48 h of incubation with gametocytes. The P. falciparum FGAA has been proven to be a robust HTS assay faithful to Plasmodium transmission-stage cell biology, and it is an innovative useful tool for antimalarial drug discovery which aims to identify new molecules with transmission-blocking potential.

Sexuality Education Websites for Adolescents: A Framework-Based Content Analysis.

The web has unique potential for adolescents seeking comprehensive sexual health information. As such, it is important to understand the nature, scope, and readability of the content and messaging provided by sexuality educational websites. We conducted a content analysis of 14 sexuality education websites for adolescents, based on the 7 essential components (sexual and reproductive health and HIV, relationships, sexual rights and sexual citizenship, pleasure, violence, diversity, and gender) of the International Planned Parenthood Framework for Comprehensive Sexuality Education. A majority of content across all sites focused on sexual and reproductive health and HIV, particularly pregnancy and STI prevention, and other information about STIs and HIV. No other topic comprised more than 10% of content coverage across a majority of sites. The authors found little discussion of gender issues, sexual rights, sexual diversity, or sexual violence. Most sites provided brief references to sexual pleasure, generally moderated with cautionary words. Language used implied a heterosexual female audience. Reading levels for most sites were above the 9th-grade level, with several at the college level. These findings have implications for enhancing online sexuality education and broadening the coverage of essential topics.

Proteomic analysis of the Plasmodium male gamete reveals the key role for glycolysis in flagellar motility.

BACKGROUND: Gametogenesis and fertilization play crucial roles in malaria transmission. While male gametes are thought to be amongst the simplest eukaryotic cells and are proven targets of transmission blocking immunity, little is known about their molecular organization. For example, the pathway of energy metabolism that power motility, a feature that facilitates gamete encounter and fertilization, is unknown. METHODS: Plasmodium berghei microgametes were purified and analysed by whole-cell proteomic analysis for the first time. Data are available via ProteomeXchange with identifier PXD001163. RESULTS: 615 proteins were recovered, they included all male gamete proteins described thus far. Amongst them were the 11 enzymes of the glycolytic pathway. The hexose transporter was localized to the gamete plasma membrane and it was shown that microgamete motility can be suppressed effectively by inhibitors of this transporter and of the glycolytic pathway. CONCLUSIONS: This study describes the first whole-cell proteomic analysis of the malaria male gamete. It identifies glycolysis as the likely exclusive source of energy for flagellar beat, and provides new insights in original features of Plasmodium flagellar organization.

Detection of RHDV strains in the Iberian hare (Lepus granatensis): earliest evidence of rabbit lagovirus cross-species infection.

Rabbit hemorrhagic disease virus (RHDV) is a highly lethal Lagovirus, family Caliciviridae, that threatens European rabbits (Oryctolagus cuniculus). Although a related virus severely affects hares, cross-species infection was only recently described for new variant RHDV in Cape hares (Lepus capensis mediterraneus). We sequenced two strains from dead Iberian hares (Lepus granatensis) collected in the 1990s in Portugal. Clinical signs were compatible with a Lagovirus infection. Phylogenetic analysis of the complete capsid gene positioned them in the RHDV genogroup that circulated on the Iberian Peninsula at that time. This is the earliest evidence of RHDV affecting a species other than European rabbits.