Entomological surveillance following a long-lasting insecticidal net universal coverage campaign in Midwestern Uganda.

BACKGROUND: A universal coverage campaign (UCC) with long-lasting insecticidal nets (LLINs) was implemented in four districts in Midwestern Uganda in 2009-2010. Entomological surveys were carried out to monitor changes in vector density, behaviour and malaria transmission following this intervention. METHODS: Anopheles mosquitoes were collected using CDC light traps quarterly and human landing catch twice a year in four sites. Collections were done at baseline before the campaign and over a three-year period following the campaign. Plasmodium falciparum circumsporozoite enzyme-linked immunosorbent assays were performed. A subset of anophelines were molecularly identified to species, and kdr L1014S frequencies were determined. RESULTS: The prevailing malaria vector in three sites was Anopheles gambiae s.l. (>97 %), with An. funestus s.l. being present in low numbers only. An. gambiae s.s. dominated (> 95 %) over An. arabiensis within A. gambiae s.l. In the remaining site, all three vector species were observed, although their relative densities varied among seasons and years. Vector densities were low in the year following the UCC but increased over time. Vector infectivity was 3.2 % at baseline and 1.8 % three years post-distribution (p = 0.001). The daily entomological inoculation rate (EIR) in 2012 varied between 0.0-0.98 for the different sites compared to a baseline EIR that was between 0.0-5.8 in 2009. There was no indication of a change in indoor feeding times, and both An. gambiae s.l. and An. funestus s.l. continued to feed primarily after midnight with vectors being active until the early morning. Kdr L1014S frequencies were already high at baseline (53-85 %) but increased significantly in all sites over time. CONCLUSIONS: The entomological surveys indicate that there was a reduction in transmission intensity coinciding with an increase in use of LLINs and other antimalarial interventions in areas of high malaria transmission. There was no change in feeding behaviour, and human-vector contact occurred indoors and primarily after midnight constantly throughout the study. Although the study was not designed to evaluate the effectiveness of the intervention compared to areas with no such intervention, the reduction in transmission occurred in an area with previously stable malaria, which seems to indicate a substantial contribution of the increased LLIN coverage.

When should we thrombolyse patients with pulmonary embolism? A systematic review of the literature.

The early mortality in pulmonary embolism (PE) is largely predicted by the associated cardiovascular response, with progressive right ventricular failure, hypotension, shock, and circulatory arrest being associated with increasing mortality. Thrombolysis may improve the prognosis of PE associated with these varying degrees of circulatory collapse, but has no place in the treatment of small emboli with no cardiovascular compromise, as it carries a significant risk of haemorrhage. This review sets out to guide the emergency physician in deciding which patients with PE may benefit from thrombolysis.

Observational study of prehospital delays in patients with chest pain.

OBJECTIVE: To define and measure patient reported prehospital delay in presentation to the emergency department with chest pain and identify simple strategies that may reduce this delay. The authors investigated the null hypothesis that the patients choice of service to call for acute medical help has no effect on the timing of thrombolysis. METHOD: A prospective observational study of prehospital times and events was undertaken on a target population of patients presenting with acute chest pain attributable to an acute coronary syndrome over a three month period. RESULTS: Patients who decided to call the ambulance service were compared with patients who contacted any other service. Most patients who contact non-ambulance services are seen by general practitioners. The prehospital system time for 121 patients who chose to call the ambulance service first was significantly shorter than for 96 patients who chose to call another service (median 57 min v 107 min; p<0.001). Of the 42 patients thrombolysed in the emergency department, those who chose to call the ambulance service had significantly shorter prehospital system times (number 21 v 21; median 44 v 69 min; p<0.001). Overall time from pain onset to initiation of thrombolysis was significantly longer in the group of patients who called a non-ambulance service first (median 130 min v 248 min; p=0.005). CONCLUSIONS: Patient with acute ischaemic chest pain who call their general practice instead of the ambulance service are likely to have delayed thrombolysis. This is likely to result in increased mortality. The most beneficial current approach is for general practices to divert all patients with possible ischaemic chest pain onset within 12 hours direct to the ambulance service.

ROMEO: a rapid rule out strategy for low risk chest pain. Does it work in a UK emergency department?

AIMS: To examine the feasibility of using the ROMEO (rule out myocardial events on "obs" ward) pathway for low risk patients with chest pain in a UK emergency department. METHODS: A prospective study was undertaken to determine outcomes for the first 100 patients entering the pathway (from May to Oct 1999). Serum troponin levels, serial ECG recordings, exercise test result, total length of stay, and final diagnoses were reviewed. Patients were telephoned after discharge to inquire about persisting or recurrent pain, and further investigations after completing the ROMEO pathway. RESULTS: 82 of 100 (82%) had myocardial damage excluded by serum troponin assay. Sixty two of 82 (76%) of these completed exercise tolerance testing (ETT). Fifty seven of 62 (92%) ETTs were negative. Twenty of 82 (26%) did not undergo ETT because of mobility problems, recent ETT, or if considered very low probability of cardiac pain on consultant review. Five of 100 (5%) had an increased initial troponin and five of 100 (5%) had an increased 12 hour troponin. These patients were referred for admission under the general physicians. Seven of 100 (7%) were referred for other reasons (late ECG changes, continuing or worsening pain). One patient self discharged. Length of stay varied because of changes to arrangements for ETT. The median time for all patients over the period studied was 23 hours. All patients were discharged within an hour of a negative ETT. FOLLOW UP RESULTS: 67 of 74 (91%) eligible patients were contacted by telephone. Forty six of 67 (69%) had no further pain, attendances, or GP consultations. Six of 67 (9%) had further cardiological investigation or treatment. CONCLUSIONS: A rapid rule out strategy such as the ROMEO pathway is feasible in the UK healthcare setting and provides standardised and consistent evaluation.

Regulation of plant NR activity by reversible phosphorylation, 14-3-3 proteins and proteolysis.

This review highlights progress in dissecting how plant nitrate reductase (NR) activity is regulated by Ca2+, protein kinases, protein kinase kinases, protein phosphatases, 14-3-3 proteins and protease(s). The signalling components that regulate NR have also been discovered to target other enzymes of metabolism, vesicle trafficking and cellular signalling. Extracellular sugars exert a major impact on the 14-3-3-binding status and stability of many target proteins, including NR in plants, whereas other stimuli affect the regulation of some targets and not others. We thus begin to see how selective or global switches in cellular behaviour are triggered by regulatory networks in response to different environmental stimuli. Surprisingly, the question of how changes in NR activity actually affect the rate of nitrate assimilation is turning out to be a tough problem.

Outline strategy for malaria control in complex emergencies

It presents the actions necessary to develop and implement a strategy for effective malaria control in complex emergencies

Outline strategy for malaria control in complex emergencies

It presents the actions necessary to develop and implement a strategy for effective malaria control in complex emergencies.

14-3-3s regulate global cleavage of their diverse binding partners in sugar-starved Arabidopsis cells.

Despite 14-3-3 proteins being implicated in the control of the eukaryotic cell cycle, metabolism, cell signalling and survival, little is known about the global regulation or functions of the phosphorylation-dependent binding of 14-3-3s to diverse target proteins. We identified Arabidopsis cytosolic proteins that bound 14-3-3s in competition with a 14-3-3-binding phosphopeptide, including nitrate reductase, glyceraldehyde- 3-phosphate dehydrogenase, a calcium-dependent protein kinase, sucrose-phosphate synthase (SPS) and glutamyl-tRNA synthetase. Remarkably, in cells starved of sugars or fed with non-metabolizable glucose analogues, all 14-3-3 binding was lost and the target proteins were selectively cleaved into proteolytic fragments. 14-3-3 binding reappeared after several hours of re-feeding with sugars. Starvation-induced degradation was blocked by 5-amino imidazole-4-carboxamide riboside (which is converted to an AMP-mimetic) or the protease inhibitor MG132 (Cbz-leu-leu-leucinal). Extracts of sugar-starved (but not sugar-fed) Arabidopsis cells contained an ATP-independent, MG132-sensitive, neutral protease that cleaved Arabidopsis SPS, and the mammalian 14-3-3-regulated transcription factor, FKHR. Cleavage of SPS and phosphorylated FKHR in vitro was blocked by binding to 14-3-3s. The finding that 14-3-3s participate in a nutrient-sensing pathway controlling cleavage of many targets may underlie the effects of these proteins on plant development.

The kidney is an important site for in vivo phenylalanine-to-tyrosine conversion in adult humans: A metabolic role of the kidney.

Synthesis of Tyr in the human body occurs by hydroxylation of the indispensable amino acid Phe. Until now, it was believed that in humans, this process was restricted to the liver, but we provide compelling evidence of production of Tyr from Phe in the kidney. To determine whether the human kidney produces Tyr, we measured Tyr balance, the Tyr appearance rate, and the Phe-to-Tyr conversion in 12 healthy human subjects by using [(15)N]Phe and [(2)H(4)]Tyr as tracers. Renal plasma flow was measured by using paraaminohippurate, and sampling from the femoral artery and renal veins was performed. The results were compared with those obtained in 12 control subjects undergoing hepatic vein catheterization and infusion of identical tracers. In all 12 subjects, there was a net uptake of Phe by the kidney (2.2 +/- 1.2 micromol/min), whereas Tyr was released (5.3 +/- 1.5 micromol/min). In contrast, there was a net uptake of both Phe (9.5 +/- 1.2 micromol/min) and Tyr (14.3 +/- 1.3 micromol/min) by the splanchnic bed. Phe conversion to Tyr occurred at a rate of 5.2 +/- 1.2 micromol/min in kidney and 3.0 +/- 0.7 micromol/min in the splanchnic bed. The kidney contributed a substantial amount of Tyr to the systemic circulation where the splanchnic bed was a net remover of Tyr. Our results demonstrate that the kidney is the major donor of Tyr to the systemic circulation by its conversion of Phe to Tyr. This observation may have important clinical implications for patients with both renal and hepatic disease, who may be at risk of Phe overloading and Tyr deficiency, and it should be considered when parenteral or enteral nutrients are administered rich in Phe and low in Tyr.

Combination therapy for malaria in Africa: hype or hope?

The development of resistance to drugs poses one of the greatest threats to malaria control. In Africa, the efficacy of readily affordable antimalarial drugs is declining rapidly, while highly efficacious drugs tend to be too expensive. Cost-effective strategies are needed to extend the useful life spans of antimalarial drugs. Observations in South-East Asia on combination therapy with artemisinin derivatives and mefloquine indicate that the development of resistance to both components is slowed down. This suggests the possibility of a solution to the problem of drug resistance in Africa, where, however, there are major obstacles in the way of deploying combination therapy effectively. The rates of transmission are relatively high, a large proportion of asymptomatic infection occurs in semi-immune persons, the use of drugs is frequently inappropriate and ill-informed, there is a general lack of laboratory diagnoses, and public health systems in sub-Saharan Africa are generally weak. Furthermore, the cost of combination therapy is comparatively high. We review combination therapy as used in South-East Asia and outline the problems that have to be overcome in order to adopt it successfully in sub-Saharan Africa.

Generation and analysis of 25 Mb of genomic DNA from the pufferfish Fugu rubripes by sequence scanning.

We have generated and analyzed >50,000 shotgun clones from 1059 Fugu cosmid clones. All sequences have been minimally edited and searched against protein and DNA databases. These data are all displayed on a searchable, publicly available web site at. With an average of 50 reads per cosmid, this is virtually nonredundant sequence skimming, covering 30%-50% of each clone. This essentially random data set covers nearly 25 Mb (>6%) of the Fugu genome and forms the basis of a series of whole genome analyses which address questions regarding gene density and distribution in the Fugu genome and the similarity between Fugu and mammalian genes. The Fugu genome, with eight times less DNA but a similar gene repertoire, is ideally suited to this type of study because most cosmids contain more than one identifiable gene. General features of the genome are also discussed. We have made some estimation of the syntenic relationship between mammals and Fugu and looked at the efficacy of ORF prediction from short, unedited Fugu genomic sequences. Comparative DNA sequence analyses are an essential tool in the functional interpretation of complex vertebrate genomes. This project highlights the utility of using the Fugu genome in this kind of study.

Microcystin affinity purification of plant protein phosphatases: PP1C, PP5 and a regulatory A-subunit of PP2A.

Proteins of approximately 35, 55 and 65kDa were purified from cauliflower extracts by microcystin-Sepharose chromatography and identified by amino acid sequencing as plant forms of protein (serine/threonine) phosphatase 1 (PP1) catalytic subunit, PP5 and a regulatory A-subunit of PP2A, respectively. Peptides that corresponded both to the tetratricopeptide (TPR) repeat and catalytic domains of PP5 were identified. Similar to mammalian PP5,the casein phosphatase activity of plant PP5 was activated >10-fold by arachidonic acid, with half-maximal stimulation occurring at approximately 100 microM lipid.

Isolation and characterisation of the retinoic acid receptor-alpha gene in the Japanese pufferfish, F. rubripes.

Nuclear hormone receptors (NRs) are ligand-inducible transcription factors that mediate critical functions in many species. The majority of novel NRs have hitherto been cloned from cDNA libraries by virtue of their homology to previously identified receptors. In this study, we validate a genomic DNA-based approach to isolating NRs by cloning the retinoic acid receptor-alpha (RARalpha) gene from the genome of the Japanese pufferfish, Fugu rubripes. The fRARalpha gene is more compact than its human and murine counterparts and demonstrates a highly conserved genomic organisation and amino acid sequence, generating two isoforms (fRARalpha1 and fRARalpha2) with divergent aminoterminal domains. In addition, a conserved regulatory element containing a retinoic acid response element was identified upstream of the fRARalpha2-specific exon, implying that retinoid induction of this isoform is evolutionarily conserved and critical to its function in vivo. We propose two uses for the Fugu genome in the study of NRs: the isolation of novel NRs that exhibit restricted spatio-temporal expression from genomic DNA and the identification of evolutionarily conserved promoter or intragenic regulatory DNA elements.

Arterial KIC as marker of liver and muscle intracellular leucine pools in healthy and type 1 diabetic humans.

In human protein turnover studies with isotopically labeled leucine (Leu) as a tracer, plasma ketoisocaproate (KIC) enrichment is extensively used as a surrogate measure of intracellular leucine enrichment. To test how accurately arterial ketoisocaproate (A-KIC) represents leucine isotopic enrichment in the hepatic (HV) and femoral veins (FV), which drain liver and muscle beds, we measured Leu and KIC enrichments in samples collected from HV, FV, and femoral artery (A) in 24 control and 6 type I diabetic subjects after a primed, continuous infusion of L-[1-(13)C,(15)N]-Leu. Studies were performed during insulin deprivation or insulin replacement in the diabetic group, whereas the effect of normal saline or three different doses of insulin infusion (0.25, 0.50, and 1 mU. kg(-1). min(-1)) were assessed in healthy controls. The ratios of baseline isotopic enrichments of A-KIC to HV Leu and FV Leu were 0.93 +/- 0.01 and 0.94 +/- 0.02, respectively, in normal subjects and 1.07 +/- 0.04 and 1.05 +/- 0.03, respectively, in diabetic subjects (P < 0.01, diabetic vs. normal subjects). Insulin did not change A-KIC-to-HV Leu ratios in either group, but the A-KIC-to-FV Leu ratio decreased during insulin infusion in normal subjects (P < 0.05). In conclusion, A-KIC represents a reliable surrogate measure of HV Leu enrichment at different levels of circulating insulin in humans. The present data support the use of A-KIC as a surrogate precursor pool for hepatic protein synthesis.

Whole body protein kinetics using Phe and Tyr tracers: an evaluation of the accuracy of approximated flux values.

Phenylalanine (Phe) kinetics are increasingly used in studies of amino acid kinetics, because the metabolic fate of Phe is limited to incorporation into protein (protein synthesis, Sp) and catabolism via hydroxylation (Qpt) to tyrosine (Tyr). Besides an infusion of labeled Phe to measure Phe flux (Qp), a priming dose of Tyr and an independent Tyr tracer are used to measure Tyr flux (Qt) and Qpt. Alternatively, Qt, Qpt, and Sp can be approximated by using equations, based on Phe and Tyr concentrations in body proteins, that eliminate the need for a Tyr tracer. To evaluate the accuracy of this approach, data were obtained from 12 type I diabetic patients and 24 nondiabetic control subjects who were studied with the full complement of tracers both with and without insulin infusion. Sp approximations closely matched measured values in both groups (mean difference <2%, all values <5%), but the agreement was poor for Qpt (error range = -8 to +43%) and Qt (error range -22 to +41%). Insulin status had no effect on these comparisons. The lower approximation error for Sp vs. Qpt is due to the small contribution ( approximately 10%) of Qpt to Qp. Approximation error for Qpt (r > 0.99) can be explained by variability in the ratio of Tyr to Phe coming from protein breakdown, (Qt - Qpt)/Qp. Ideally, all fluxes should be directly measured, but these data suggest that whole body Sp can be approximated with an acceptably small margin of error. However, the same equations do not yield reliably accurate values for Qpt or Qt.