Complement receptor 3 is required for maximum in vitro trogocytic killing of the parasite Trichomonas vaginalis by human neutrophil-like cells.

Trichomonas vaginalis (Tv) is a parasite that causes trichomoniasis, a prevalent sexually-transmitted infection. Neutrophils are found at the site of infection, and can rapidly kill the parasite in vitro, using trogocytosis. However, the specific molecular players in neutrophil killing of Tv are unknown. Here, we show that complement proteins play a role in Tv killing by human neutrophil-like cells (NLCs). Using CRISPR/Cas9, we generated NLCs deficient in each of three complement receptors (CRs) known to be expressed on human neutrophils: CR1, CR3, and CR4. Using in vitro trogocytosis assays, we found that CR3, but not CR1 or CR4 is required for maximum trogocytosis of the parasite by NLCs, with NLCs lacking CR3 demonstrating ~40% reduction in trogocytosis, on average. We also observed a reduction in NLC killing of Tv in CR3 knockout, but not CR1 or CR4 knockout NLCs. On average, NLCs lacking CR3 had ~50% reduction in killing activity. We also used a parallel approach of pre-incubating NLCs with blocking antibodies against CR3, which similarly reduced NLC killing of parasites. These data support a model in which Tv is opsonized by the complement protein iC3b, and bound by neutrophil CR3 receptor, to facilitate trogocytic killing of the parasite.

Aerodynamic Size-Dependent Collection and Inactivation of Virus-Laden Aerosol Particles in an Electrostatic Precipitator.

Electrostatic precipitators (ESPs) may enable high particle collection efficiency with minimal pressure drop in HVAC systems. However, studies of pathogen collection and inactivation in ESPs at medium to higher flow rates are limited. Here, a single-stage, wire-plate ESP operated at flow rates of 51 and 85 m3 h-1 was used to study the removal of virus-laden aerosol particles for three different airborne viruses: (1) bovine coronavirus (BCoV), (2) influenza A virus (IAV), and (3) porcine reproductive and respiratory virus (PRRSV). Size-resolved measurements of collection efficiency were obtained using Andersen cascade impactors (ACI) sampling upstream and downstream of the ESP. All measurements were analyzed based on three distinctive but complementary methods: (1) fluorimetry to assess physical collection, (2) RT-qPCR to assess viral RNA concentrations and (3) virus titration to assess virus viability. In general, log reductions by virus titration were highest followed by those from RT-qPCR, and last fluorimetry, suggesting that a portion of virus may be potentially inactivated in flight in the ESP. An effective migration (deposition) velocity ranging from 3.10 to 10.05 cm s-1 was also determined using the spatially resolved measurements of virus collection on the ESP plates.

Identification of a novel base J binding protein complex involved in RNA polymerase II transcription termination in trypanosomes.

Base J, ß-D-glucosyl-hydroxymethyluracil, is a modification of thymine DNA base involved in RNA Polymerase (Pol) II transcription termination in kinetoplastid protozoa. Little is understood regarding how specific thymine residues are targeted for J-modification or the mechanism of J regulated transcription termination. To identify proteins involved in J-synthesis, we expressed a tagged version of the J-glucosyltransferase (JGT) in Leishmania tarentolae, and identified four co-purified proteins by mass spectrometry: protein phosphatase (PP1), a homolog of Wdr82, a potential PP1 regulatory protein (PNUTS) and a protein containing a J-DNA binding domain (named JBP3). Gel shift studies indicate JBP3 is a J-DNA binding protein. Reciprocal tagging, co-IP and sucrose gradient analyses indicate PP1, JGT, JBP3, Wdr82 and PNUTS form a multimeric complex in kinetoplastids, similar to the mammalian PTW/PP1 complex involved in transcription termination via PP1 mediated dephosphorylation of Pol II. Using RNAi and analysis of Pol II termination by RNA-seq and RT-PCR, we demonstrate that ablation of PNUTS, JBP3 and Wdr82 lead to defects in Pol II termination at the 3'-end of polycistronic gene arrays in Trypanosoma brucei. Mutants also contain increased antisense RNA levels upstream of transcription start sites, suggesting an additional role of the complex in regulating termination of bi-directional transcription. In addition, PNUTS loss causes derepression of silent Variant Surface Glycoprotein genes involved in host immune evasion. Our results suggest a novel mechanistic link between base J and Pol II polycistronic transcription termination in kinetoplastids.

The importance of the urea cycle and its relationships to polyamine metabolism during ammonium stress in Medicago truncatula.

The ornithine-urea cycle (urea cycle) makes a significant contribution to the metabolic responses of lower photosynthetic eukaryotes to episodes of high nitrogen availability. In this study, we compared the role of the plant urea cycle and its relationships to polyamine metabolism in ammonium-fed and nitrate-fed Medicago truncatula plants. High ammonium resulted in the accumulation of ammonium and pathway intermediates, particularly glutamine, arginine, ornithine, and putrescine. Arginine decarboxylase activity was decreased in roots, suggesting that the ornithine decarboxylase-dependent production of putrescine was important in situations of ammonium stress. The activity of copper amine oxidase, which releases ammonium from putrescine, was significantly decreased in both shoots and roots. In addition, physiological concentrations of ammonium inhibited copper amine oxidase activity in in vitro assays, supporting the conclusion that high ammonium accumulation favors putrescine synthesis. Moreover, early supplementation of plants with putrescine avoided ammonium toxicity. The levels of transcripts encoding urea-cycle-related proteins were increased and transcripts involved in polyamine catabolism were decreased under high ammonium concentrations. We conclude that the urea cycle and associated polyamine metabolism function as important protective mechanisms limiting ammonium toxicity in M. truncatula. These findings demonstrate the relevance of the urea cycle to polyamine metabolism in higher plants.

By force of habit: Self-trapping in a dynamical utility landscape.

Historically, rational choice theory has focused on the utility maximization principle to describe how individuals make choices. In reality, there is a computational cost related to exploring the universe of available choices and it is often not clear whether we are truly maximizing an underlying utility function. In particular, memory effects and habit formation may dominate over utility maximization. We propose a stylized model with a history-dependent utility function, where the utility associated to each choice is increased when that choice has been made in the past, with a certain decaying memory kernel. We show that self-reinforcing effects can cause the agent to get stuck with a choice by sheer force of habit. We discuss the special nature of the transition between free exploration of the space of choice and self-trapping. We find, in particular, that the trapping time distribution is precisely a Zipf law at the transition, and that the self-trapped phase exhibits super-aging behavior.

Leishmaniasis in Norway Rats in Sewers, Barcelona, Spain.

We detected Leishmania infantum in 98 Norway rats (Rattus norvegicus) trapped in parks and sewers of Barcelona, Spain. The 84 rats from the sewers showed a prevalence of 33.3% and up to 2,272 estimated parasites. These results, in the most abundant potential reservoir in cities, is of public health concern.

Volatile compounds other than CO2 emitted by different microorganisms promote distinct posttranscriptionally regulated responses in plants.

A "box-in-box" cocultivation system was used to investigate plant responses to microbial volatile compounds (VCs) and to evaluate the contributions of organic and inorganic VCs (VOCs and VICs, respectively) to these responses. Arabidopsis plants were exposed to VCs emitted by adjacent Alternaria alternata and Penicillium aurantiogriseum cultures, with and without charcoal filtration. No VOCs were detected in the headspace of growth chambers containing fungal cultures with charcoal filters. However, these growth chambers exhibited elevated CO2 and bioactive CO and NO headspace concentrations. Independently of charcoal filtration, VCs from both fungal phytopathogens promoted growth and distinct developmental changes. Plants cultured at CO2 levels observed in growth boxes containing fungal cultures were identical to those cultured at ambient CO2 . Plants exposed to charcoal-filtered fungal VCs, nonfiltered VCs, or superelevated CO2 levels exhibited transcriptional changes resembling those induced by increased irradiance. Thus, in the "box-in-box" system, (a) fungal VICs other than CO2 and/or VOCs not detected by our analytical systems strongly influence the plants' responses to fungal VCs, (b) different microorganisms release VCs with distinct action potentials, (c) transcriptional changes in VC-exposed plants are mainly due to enhanced photosynthesis signaling, and (d) regulation of some plant responses to fungal VCs is primarily posttranscriptional.

Drought tolerance response of high-yielding soybean varieties to mild drought: physiological and photochemical adjustments.

Soybean is a crop of agronomic importance that requires adequate watering during its growth to achieve high production. In this study, we determined physiological, photochemical and metabolic differences in five soybean varieties selected from the parental lines of a nested association mapping population during mild drought. These varieties have been described as high yielding (NE3001, HY1; LD01-5907, HY2) or drought tolerant (PI518751; HYD1; PI398881, HYD2). Nevertheless, there has been little research on the physiological traits that sustain their high productivity under water-limited conditions. The results indicate that high-yielding varieties under drought cope with the shortage of water by enhancing their photoprotective defences and invest in growth and productivity, linked to a higher intrinsic water use efficiency. This is the case of the variety N-3001 (HY1), with a tolerance strategy involving a faster transition into the reproductive stage to avoid the drought period. The present study highlights the role of the physiological and biochemical adjustments of various soybean varieties to cope with water-limited conditions. Moreover, the obtained results underscore the fact that the high phenotypic plasticity among soybean phenotypes should be exploited to compensate for the low genetic variability of this species when selecting plant productivity in constrained environments.

Evaluation of the cost-effectiveness of the treatment of uncomplicated severe acute malnutrition by lady health workers as compared to an outpatient therapeutic feeding programme in Sindh Province, Pakistan.

BACKGROUND: Due to the limited evidence of the cost-effectiveness of Community Health Workers (CHW) delivering treatment for severe acute malnutrition (SAM), there is a need to better understand the costs incurred by both implementing institutions and beneficiary households. This study assessed the costs and cost-effectiveness of treatment for cases of SAM without complications delivered by government-employed Lady Health Workers (LHWs) and complemented with non-governmental organisation (NGO) delivered outpatient facility-based care compared with NGO delivered outpatient facility-based care only alongside a two-arm randomised controlled trial conducted in Sindh Province, Pakistan. METHODS: An activity-based cost model was used, employing a societal perspective to include costs incurred by beneficiaries and the wider community. Costs were estimated through accounting records, interviews and informal group discussions. Cost-effectiveness was assessed for each arm relative to no intervention, and incrementally between the two interventions, providing information on both absolute and relative costs and effects. RESULTS: The cost per child recovered in outpatient facility-based care was similar to LHW-delivered care, at 363 USD and 382 USD respectively. An additional 146 USD was spent per additional child recovered by outpatient facilities compared to LHWs. Results of sensitivity analyses indicated considerable uncertainty in which strategy was most cost-effective due to small differences in cost and recovery rates between arms. The cost to the beneficiary household of outpatient facility-based care was double that of LHW-delivered care. CONCLUSIONS: Outpatient facility-based care was found to be slightly more cost-effective compared to LHW-delivered care, despite the potential for cost-effectiveness of CHWs managing SAM being demonstrated in other settings. The similarity of cost-effectiveness outcomes between the two models resulted in uncertainty as to which strategy was the most cost-effective. Similarity of costs and effectiveness between models suggests that whether it is appropriate to engage LHWs in substituting or complementing outpatient facilities may depend on population needs, including coverage and accessibility of existing services, rather than be purely a consideration of cost. Future research should assess the cost-effectiveness of LHW-delivered care when delivered solely by the government. TRIAL REGISTRATION: NCT03043352 , ClinicalTrials.gov. Retrospectively registered.

Cost-effectiveness of the treatment of uncomplicated severe acute malnutrition by community health workers compared to treatment provided at an outpatient facility in rural Mali.

BACKGROUND: The Malian Nutrition Division of the Ministry of Health and Action Against Hunger tested the feasibility of integrating treatment of severe acute malnutrition (SAM) into the existing Integrated Community Case Management package delivered by community health workers (CHWs). This study assessed costs and cost-effectiveness of CHW-delivered care compared to outpatient facility-based care. METHODS: Activity-based costing methods were used, and a societal perspective employed to include all relevant costs incurred by institutions, beneficiaries and communities. The intervention and control arm enrolled different numbers of children so a modelled scenario sensitivity analysis was conducted to assess the cost-effectiveness of the two arms, assuming equal numbers of children enrolled. RESULTS: In the base case, with unequal numbers of children in each arm, for CHW-delivered care, the cost per child treated was 244 USD and cost per child recovered was 259 USD. Outpatient facility-based care was less cost-effective at 442 USD per child and 501 USD per child recovered. The conclusions of the analysis changed in the modelled scenario sensitivity analysis, with outpatient facility-based care being marginally more cost-effective (cost per child treated is 188 USD, cost per child recovered is 214 USD), compared to CHW-delivered care. This suggests that achieving good coverage is a key factor influencing cost-effectiveness of CHWs delivering treatment for SAM in this setting. Per week of treatment, households receiving CHW-delivered care spent half of the time receiving treatment and three times less money compared with those receiving treatment from the outpatient facility. CONCLUSIONS: This study supports existing evidence that the delivery of treatment by CHWs is a cost-effective intervention, provided that good coverage is achieved. A major benefit of this strategy was the lower cost incurred by the beneficiary household when treatment is available in the community. Further research is needed on the implementation costs that would be incurred by the government to increase the operability of these results.

Quality of care of treatment for uncomplicated severe acute malnutrition provided by lady health workers in Pakistan.

OBJECTIVE: To assess the quality of care provided by lady health workers (LHW) managing cases of uncomplicated severe acute malnutrition (SAM) in the community. DESIGN: Cross-sectional quality-of-care study. SETTING: The feasibility of the implementation of screening and treatment for uncomplicated SAM in the community by LHW was tested in Sindh Province, Pakistan. An observational, clinical prospective multicentre cohort study compared the LHW-delivered care with the existing outpatient health facility model. SUBJECTS: LHW implementing treatment for uncomplicated SAM in the community. RESULTS: Oedema was diagnosed conducted correctly for 87·5 % of children; weight and mid upper-arm circumference were measured correctly for 60·0 % and 57·4 % of children, respectively. The appetite test was conducted correctly for 42·0 % of cases. Of all cases of SAM without complications assessed during the study, 68·0 % received the correct medical and nutrition treatment. The proportion of cases that received the correct medical and nutrition treatment and key counselling messages was 4·0 %. CONCLUSIONS: This quality-of-care study supports existing evidence that LHW are able to identify uncomplicated SAM, and a majority can provide appropriate nutrition and medical treatment in the community. However, the findings also show that their ability to provide the complete package with an acceptable level of care is not assured. Additional evidence on the impact of supervision and training on the quality of SAM treatment and counselling provided by LHW to children with SAM is required. The study has also shown that, as in other sectors, it is essential that operational challenges are addressed in a timely manner and that implementers receive appropriate levels of support, if SAM is to be treated successfully in the community.

Quality of care for treatment of uncomplicated severe acute malnutrition delivered by community health workers in a rural area of Mali.

An alternative Community-based Management of Acute Malnutrition model with community health workers (CHWs) delivering treatment for uncomplicated severe acute malnutrition (SAM) was piloted in Mali. The capacity of the CHWs to evaluate, classify, and treat cases of uncomplicated SAM, to provide nutritional counselling to caretakers of children receiving treatment for SAM, malaria, pneumonia or diarrhoea and to correctly refer cases of complicated SAM, was assessed. This was done using direct observation by trained enumerators of the management of SAM cases using checklists, re-diagnosing the cases admitted for treatment and reviewing admissions cards and registers. One hundred twenty-five cases, assessed and treated by the CHWs, were observed. The majority of children were correctly assessed for the presence of major clinical signs (cough, diarrhoea, fever, and vomiting; 97.6%), and similarly most children were checked for the presence of danger signs (95.2%). Mid-upper arm circumference was correctly assessed in 96.8% of children and oedema was correctly assessed in 78.4% (The composite indicator, which includes all essential tasks to provide high-quality treatment, was achieved in 79.5% of cases. This paper concludes that well-trained and supervised CHWs are capable of managing cases of uncomplicated SAM. This suggests that such a strategy is an opportunity to increase access to quality treatment in Mali for SAM cases. However, further evidence is required to ensure that this level of care can be achieved at scale.

Nitric oxide induces the alternative oxidase pathway in Arabidopsis seedlings deprived of inorganic phosphate.

Phosphate starvation compromises electron flow through the cytochrome pathway of the mitochondrial electron transport chain, and plants commonly respond to phosphate deprivation by increasing flow through the alternative oxidase (AOX). To test whether this response is linked to the increase in nitric oxide (NO) production that also increases under phosphate starvation, Arabidopsis thaliana seedlings were grown for 15 d on media containing either 0 or 1mM inorganic phosphate. The effects of the phosphate supply on growth, the production of NO, respiration, the AOX level and the production of superoxide were compared for wild-type (WT) seedlings and the nitrate reductase double mutant nia. Phosphate deprivation increased NO production in WT roots, and the AOX level and the capacity of the alternative pathway to consume electrons in WT seedlings; whereas the same treatment failed to stimulate NO production and AOX expression in the nia mutant, and the plants had an altered growth phenotype. The NO donor S-nitrosoglutathione rescued the growth phenotype of the nia mutants under phosphate deprivation to some extent, and it also increased the respiratory capacity of AOX. It is concluded that NO is required for the induction of the AOX pathway when seedlings are grown under phosphate-limiting conditions.

Segmental fat-free and fat mass measurements by bioelectrical impedance analysis in 2,224 healthy Spanish women aged 18-85 years.

OBJECTIVES: This study provides updated data on body composition in adult Spanish women. METHODS: We considered data, including height and weight, from a survey conducted on a total of 4,013 adult women between 2009 and 2010. A subgroup of 2,224 women completed a bioelectrical body impedance analysis of body composition using a Tanita Body Composition Analyzer (Model no. BF-418). RESULTS: Total fat mass (FM) gradually increased between 18 and 74 years of age and decreased thereafter. FM increased in both legs between 65 and 74 years of age (5.69 ± 1.4 Kg and 5.66 ± 1.4 Kg for right and left legs, respectively) and decreased thereafter. FM in the right arm increased until 44 years of age (1.22 ± 2.6 Kg), decreased to 1.19 ± 0.5 Kg between 45 and 54 years of age, and increased to 1.54 ± 0.63 from 65 to 74 years of age. FM in the left arm increased constantly until it reached a peak of 1.63 ± 0.6 Kg between 65 and 74 years of age and decreased thereafter. FM increased in the trunk throughout life (peaks at 13.27±3.8 Kg) until subjects reached between 65 and 74 years of age. Fat free mass increased until 44 years of age (42.42 ± 4.17 Kg) and decreased thereafter. The prevalence of overweight/obesity significantly increased with age in the entire sample. CONCLUSIONS: Our results indicate that age-related increases in weight were at least partially due to increased adiposity.

Quantitative ultrasound measurements of the calcaneus and hand phalanges in elderly Spanish men: relationship with peripheral bone mineral density of the hand phalanges.

OBJECTIVES: The aims of this pilot study were to describe quantitative ultrasound (US) measurements and peripheral bone mineral density (BMD) of the hand phalanges on dual-energy x-ray absorptiometry and to examine the correlations between them in elderly Spanish men. METHODS: We studied 199 healthy men (mean age ± SD, 73.31 ± 5.10 years). The participants were not taking any medications, and they reported no diseases, including diseases that are associated with abnormalities in mineral metabolism. Phalangeal and calcaneal quantitative US measurements and phalangeal BMD measurements were performed in all participants. RESULTS: A bivariate correlation analysis showed no association between quantitative US assessments at the phalanges or the calcaneus (P = .409). After adjustment for potential confounders, the correlation between phalangeal BMD and phalangeal quantitative US measurements was r = 0.417 (P < .0001), and the correlation for calcaneal quantitative US was r = 0.26 (P = .001). Further adjustment by percentage of body fat increased quantitative US correlations with phalangeal BMD: r = 0.450 (P < .0001) at the phalanges; r = 0.291 (P = .001) at the calcaneus. CONCLUSIONS: There is a small correlation between quantitative US measurements at the calcaneus and phalangeal BMD that increases to a moderate level with quantitative US measurements at the phalanges in elderly Spanish men.

Defining the Effects of PKC Modulator HIV Latency-Reversing Agents on Natural Killer Cells.

Background: Latency reversing agents (LRAs) such as protein kinase C (PKC) modulators can reduce rebound-competent HIV reservoirs in small animal models. Furthermore, administration of natural killer (NK) cells following LRA treatment improves this reservoir reduction. It is currently unknown why the combination of a PKC modulator and NK cells is so potent and whether exposure to PKC modulators may augment NK cell function in some way. Methods: Primary human NK cells were treated with PKC modulators (bryostatin-1, prostratin, or the designed, synthetic bryostatin-1 analog SUW133), and evaluated by examining expression of activation markers by flow cytometry, analyzing transcriptomic profiles by RNA sequencing, measuring cytotoxicity by co-culturing with K562 cells, assessing cytokine production by Luminex assay, and examining the ability of cytokines and secreted factors to independently reverse HIV latency by co-culturing with Jurkat-Latency (J-Lat) cells. Results: PKC modulators increased expression of proteins involved in NK cell activation. Transcriptomic profiles from PKC-treated NK cells displayed signatures of cellular activation and enrichment of genes associated with the NFκB pathway. NK cell cytotoxicity was unaffected by prostratin but significantly decreased by bryostatin-1 and SUW133. Cytokines from PKC-stimulated NK cells did not induce latency reversal in J-Lat cell lines. Conclusions: Although PKC modulators have some significant effects on NK cells, their contribution in "kick and kill" strategies is likely due to upregulating HIV expression in CD4+ T cells, not directly enhancing the effector functions of NK cells. This suggests that PKC modulators are primarily augmenting the "kick" rather than the "kill" arm of this HIV cure approach.

A new oxidative pathway of nitric oxide production from oximes in plants.

Nitric oxide (NO) is an essential reactive oxygen species and a signal molecule in plants. Although several studies have proposed the occurrence of oxidative NO production, only reductive routes for NO production, such as the nitrate (NO-3) -upper-reductase pathway, have been evidenced to date in land plants. However, plants grown axenically with ammonium as the sole source of nitrogen exhibit contents of nitrite and NO3-, evidencing the existence of a metabolic pathway for oxidative production of NO. We hypothesized that oximes, such as indole-3-acetaldoxime (IAOx), a precursor to indole-3-acetic acid, are intermediate oxidation products in NO synthesis. We detected the production of NO from IAOx and other oximes catalyzed by peroxidase (POD) enzyme using both 4-amino-5-methylamino-2',7'-difluorescein fluorescence and chemiluminescence. Flavins stimulated the reaction, while superoxide dismutase inhibited it. Interestingly, mouse NO synthase can also use IAOx to produce NO at a lower rate than POD. We provided a full mechanism for POD-dependent NO production from IAOx consistent with the experimental data and supported by density functional theory calculations. We showed that the addition of IAOx to extracts from Medicago truncatula increased the in vitro production of NO, while in vivo supplementation of IAOx and other oximes increased the number of lateral roots, as shown for NO donors, and a more than 10-fold increase in IAOx dehydratase expression. Furthermore, we found that in vivo supplementation of IAOx increased NO production in Arabidopsis thaliana wild-type plants, while prx33-34 mutant plants, defective in POD33-34, had reduced production. Our data show that the release of NO by IAOx, as well as its auxinic effect, explain the superroot phenotype. Collectively, our study reveals that plants produce NO utilizing diverse molecules such as oximes, POD, and flavins, which are widely distributed in the plant kingdom, thus introducing a long-awaited oxidative pathway to NO production in plants. This knowledge has essential implications for understanding signaling in biological systems.