Use of hub cutters and the volume of sharp waste and occurrence of needle-stick injuries during 2011 mass immunization campaigns against yellow fever in Ghana: a cohort study.

BACKGROUND: Current WHO best infection control practices for injections do not address the use of hub cutters due to insufficient evidence on safety and efficacy. OBJECTIVE: To assess the impact of the use of hub cutters on 1) the frequency of needle-stick injuries (NSIs) and other blood exposures among workers and 2) the volume of sharps waste in a mass vaccination campaign setting. METHODS: During yellow fever vaccination in Ghana, we conducted a cohort study on the use of hub cutters. We compared two groups---one group using hub cutters and a control group---for the occurrences of NSIs and the volume of sharp waste produced. RESULTS: In the control arm, vaccinators used 284 482 syringes in 825 vaccination sessions. In the group using hub cutter, vaccinators used 397 079 syringes in 1599 sessions. Among vaccinators, the rate of NSI was not significantly (p=0.14) different between the hub cutter users (0.15/10 000 syringes) and the control group (0.04/10 000). Factors such as workload, lack of organization and pressure seemed to have influence the occurrence of NSIs. With all the limitations of the work, the volume of sharp waste per 10 000 syringes was 0.24 m(3) in the hub cutter users and 0.41 m(3) in the control group---a reduction of 41.2%. Vaccinators found hub cutters easy to use and safe. Use of hub cutter was not associated with increased duration of work. CONCLUSION: The use of hub cutters did not increase the risk of NSIs. More training is needed to facilitate its implementation in mass campaign setting.

Identification and characterization of a mitochondrial thioredoxin system in plants.

Plants possess two well described thioredoxin systems: a cytoplasmic system including several thioredoxins and an NADPH-dependent thioredoxin reductase and a specific chloroplastic system characterized by a ferredoxin-dependent thioredoxin reductase. On the basis of biochemical activities, plants also are supposed to have a mitochondrial thioredoxin system as described in yeast and mammals, but no gene encoding plant mitochondrial thioredoxin or thioredoxin reductase has been identified yet. We report the characterization of a plant thioredoxin system located in mitochondria. Arabidopsis thaliana genome sequencing has revealed numerous thioredoxin genes among which we have identified AtTRX-o1, a gene encoding a thioredoxin with a potential mitochondrial transit peptide. AtTRX-o1 and a second gene, AtTRX-o2, define, on the basis of the sequence and intron positions, a new thioredoxin type up to now specific to plants. We also have characterized AtNTRA, a gene encoding a protein highly similar to the previously described cytosolic NADPH-dependent thioredoxin reductase AtNTRB but with a putative presequence for import into mitochondria. Western blot analysis of A. thaliana subcellular and submitochondrial fractions and in vitro import experiments show that AtTRX-o1 and AtNTRA are targeted to the mitochondrial matrix through their cleavable N-terminal signal. The two proteins truncated to the estimated mature forms were produced in Escherichia coli; AtTRX-o1 efficiently reduces insulin in the presence of DTT and is reduced efficiently by AtNTRA and NADPH. Therefore, the thioredoxin and the NADPH-dependent thioredoxin reductase described here are proposed to constitute a functional plant mitochondrial thioredoxin system.

Keeping clean water clean in a Malawi refugee camp: a randomized intervention trial.

OBJECTIVE: This study was undertaken to assess the ability of a water container with a cover and a spout to prevent household contamination of water in a Malawian refugee camp. METHODS: A randomized trial was conducted in a refugee population that had experienced repeated outbreaks of cholera and diarrhoea and where contamination of water in the home was found to be a significant cause of cholera. Four hundred Mozambican refugee households were systematically identified and followed over a 4-month period, one fourth of the households were randomly assigned to exclusively use the improved container for water collection. FINDINGS: Water flowing from the source wells had little or no microbial contamination although the water collectors quickly contaminated their water, primarily through contact with their hands. Analysis of water samples demonstrated that there was a 69% reduction in the geometric mean of faecal coliform levels in household water and 31% less diarrhoeal disease (P = 0.06) in children under 5 years of age among the group using the improved bucket. Regression models examining diarrhoea among under 5-year-olds confirmed the protective effect of the bucket and found that visible faeces in the family latrine and the presence of animals were significantly associated with an increased diarrhoeal incidence in children. CONCLUSION: Household contamination of drinking-water significantly contributed to diarrhoea in this population. Proper chlorination is a less expensive and more effective means of water quality protection in comparison with the improved bucket, but was unpopular and rarely utilized by the camp inhabitants.

In vivo characterization of a thioredoxin h target protein defines a new peroxiredoxin family.

Disruption of the two thioredoxin genes in yeast dramatically affects cell viability and growth. Expression of Arabidopsis thioredoxin AtTRX3 in the Saccharomyces thioredoxin Delta strain EMY63 restores a wild-type cell cycle, the ability to grow on methionine sulfoxide, and H2O2 tolerance. In order to isolate thioredoxin targets related to these phenotypes, we prepared a C35S (Escherichia coli numbering) thioredoxin mutant to stabilize the intermediate disulfide bridged complex and we added a polyhistidine N-terminal extension in order to purify the complex rapidly. Expression of this mutant thioredoxin in the wild-type yeast induces a reduced tolerance to H2O2, but only limited change in the cell cycle and no change in methionine sulfoxide utilization. Expression in the Delta thioredoxin strain EMY63 allowed us to isolate a complex of the thioredoxin with YLR109, an abundant yeast protein related to PMP20, a peroxisomal protein of Candida. No function has so far been attributed to this protein or to the other numerous homologues described in plants, animals, fungi, and prokaryotes. On the basis of the complementation and of low similarity with peroxiredoxins, we produced YLR109 and one of its Arabidopsis homologues in E. coli to test their peroxiredoxins activity. We demonstrate that both recombinant proteins present a thioredoxin-dependent peroxidase activity in vitro. The possible functions of this new peroxiredoxin family are discussed.

[Patient information and medical responsibility]. / Information du patient et responsabilité médicale.

The duty of every physician to inform his patient, the failure of which can engage his responsibility, can be essentially considered from the following four angles: a duty to inform: why? a duty to inform: what duty? a duty to inform: how? a duty to inform: for whom?

Intron position as an evolutionary marker of thioredoxins and thioredoxin domains.

In contrast to prokaryotes, which typically possess one thioredoxin gene per genome, three different thioredoxin types have been described in higher plants. All are encoded by nuclear genes, but thioredoxins m and f are chloroplastic while thioredoxins h have no transit peptide and are probably cytoplasmic. We have cloned and sequenced Arabidopsis thaliana genomic fragments encoding the five previously described thioredoxins h, as well as a sixth gene encoding a new thioredoxin h. In spite of the high divergence of the sequences, five of them possess two introns at positions identical to the previously sequenced tobacco thioredoxin h gene, while a single one has only the first intron. The recently published sequence of Chlamydomonas thioredoxin h shows three introns, two at the same positions as in higher plants. This strongly suggests a common origin for all cytoplasmic thioredoxins of plants and green algae. In addition, we have cloned and sequenced pea DNA genomic fragments encoding thioredoxins m and f. The thioredoxin m sequence shows only one intron between the regions encoding the transit peptide and the mature protein, supporting the prokaryotic origin of this sequence and suggesting that its association with the transit peptide has been facilitated by exon shuffling. In contrast, the thioredoxin f sequence shows two introns, one at the same position as an intron in various plant and animal thioredoxins and the second at the same position as an intron in thioredoxin domains of disulfide isomerases. This strongly supports the hypothesis of a eukaryotic origin for chloroplastic thioredoxin f.

Growth-related gene expression in Nicotiana tabacum mesophyll protoplasts.

Eight cDNAs whose genes are more strongly expressed in suspension cells in growth phase than in stationary phase and at a low level in mature leaves have been isolated. The corresponding mRNAs are abundantly accumulated in young plant organs and in germinating seeds but are almost undetectable in mature plant tissues and dry seeds. Six of these cDNAs were characterized by comparison of nucleotide and protein sequences to the EMBL and SWISSPROT databanks. These eight growth-related genes are expressed in protoplasts isolated from Nicotiana tabacum mesophyll cells shortly after preparation (4 h). Two of them are expressed in freshly isolated protoplasts (early genes), while the other six are detected after 4 h of culture (late genes). Seven are more abundantly expressed in protoplasts than in growing plant organs while one growth-related gene is weakly expressed in protoplasts, as is the histone H4 gene. They seem to be induced in protoplasts by a synergistic effect of wounding and maceration. Sustained expression of the early genes is dependent on the presence of sucrose in the culture medium.

The Nicotiana tabacum genome encodes two cytoplasmic thioredoxin genes which are differently expressed.

A Nicotiana tabacum thioredoxin h gene (EMBL Accession No. Z11803) encoding a new thioredoxin (called h2) was isolated using thioredoxin h1 cDNA (X58527), and represents the first thioredoxin h gene isolated from a higher plant. It encodes a polypeptide of 118 amino acids with the conserved thioredoxin active site Trp-Cys-Gly-Pro-Cys. This gene comprises two introns which have lengths of 1071 and 147 bp respectively, and three exons which encode peptides of 29, 41 and 48 amino acids, respectively. This thioredoxin h shows 66% identity with the amino acid sequence of thioredoxin h1 (X58527) and only around 35% with the choroplastic thioredoxins. The two thioredoxins, h1 and h2, do not have any signal peptides and are most probably cytoplasmic. Using the 3' regions of the mRNAs, two probes specific for thioredoxins h1 and h2 have been prepared. Southern blot analysis shows that thioredoxin sequences are present in only two genomic EcoRI fragments: a 3.3 kb fragment encodes h1 and a 4.5 kb fragment encodes h2. Analysis of the ancestors of the allotetraploid N. tabacum shows that thioredoxin h2 is present in N. sylvestris and N. tomentosiformis but that thioredoxin h1 is absent from both putative ancestors. Thus, the thioredoxin h1 gene has probably been recently introduced in to N. tabacum as a gene of agronomic importance, or linked to such genes. Northern blot analysis shows that both genes are expressed in N. tabacum, mostly in organs or tissues that contain growing cells.(ABSTRACT TRUNCATED AT 250 WORDS)

mRNAs newly synthesized by tobacco mesophyll protoplasts are wound-inducible.

We have used 2-dimensional (2D) non-equilibrium pH gradient gel electrophoresis (NEPHGE) of in vitro synthesized proteins and northern hybridization with labelled cDNAs coding for three pathogenesis related (P.R.) proteins, to analyze the shift in mRNA content induced by the isolation and culture of tobacco mesophyll protoplasts. The in vitro protein pattern of mRNAs from freshly isolated protoplasts is characterized by the absence of most leaf spots and the appearance of 19 new spots. After 6 hours of culture, the mRNAs coding for the P.R. proteins become detectable and after 12 hours the protoplasts contain an mRNA population almost typical of callus cells. The different steps involved in the isolation and culture of protoplasts were analysed. Cutting off the leaf and sterilization do not change the mRNA set. In contrast, the mechanical injury applied to the leaf in order to facilitate the penetration of the enzymatic mixture induces a modification of the mRNA content identical to that resulting from protoplast isolation. Wounding is the essential event inducing dedifferentiation. Varying the culture medium and conditions leads to only limited modifications of the mRNA pattern. These results are discussed on the basis of present knowledge of the reaction of the plant to wounding and we suggest that wound healing callus and in vitro callus correspond to the same differentiation state.

Tobacco Mesophyll Protoplasts Synthesize 1,3-beta-Glucanase, Chitinases, and "Osmotins" during in Vitro Culture.

Tobacco (Nicotiana tabacum) mesophyll protoplasts synthesize six basic proteins (a, a', a(1), b, b', and c) which are undetectable in the leaf and whose synthesis is reduced by auxin (Y Meyer, L Aspart, Y Chartier [1984] Plant Physiol 75: 1027-1033). Polypeptides a, a', and a(1) were shown to have similar mobilities on two-dimensional electrophoresis as one 1,3-beta-glucanase and two chitinases from tobacco mosaic virus-infected leaves. In immunoblotting experiments, polypeptide a was recognized by specific antibodies raised against the 1,3-beta-glucanase and a' and a(1) reacted with anti-chitinase antibodies. Similarly, b and b' comigrated with osmotin and its neutral counterpart, two proteins characteristic of salt-adapted tobacco cells, and reacted with anti-osmotin antibodies. In addition it has been shown that 1,3-beta-glucanase and chitinase activities increased at the same time as a, a', and a(1) accumulated in cultivated protoplasts. Finally, polypeptide c was also detected in tobacco mosaic virus-infected leaves but could not be identified as any of the pathogenesis-related proteins characterized so far in tobacco. Thus, cultivated tobacco protoplasts synthesize and accumulate typical stress proteins.

Preparation by two-dimensional electrophoresis of proteins for antibody production: antibodies against proteins whose synthesis is reduced by auxin in tobacco mesophyll protoplasts.

Using phenol extraction from tobacco callus, we have prepared extracts with a high protein content. These proteins were separated in cylindrical non-equilibrium pH gradient gels and visualized by dipping in sodium dodecyl sulfate (SDS)-containing solution. Three gel sections, each containing proteins previously detected as abundantly synthesized in tobacco mesophyll protoplasts and whose synthesis is reduced by auxin application, were excised from each gel and collected. These proteins were further separated on slab SDS gels and protein bands were excised after Coomassie Brilliant Blue R-250 staining and used to inject three rabbits. After one booster, highly specific antibodies were detected in their sera by ELISA and immunoblotting. Using these sera we have confirmed that the corresponding proteins are identical in callus and mesophyll protoplast and demonstrated that they are abundantly accumulated in tobacco roots but are undetectable in aerial organs and seeds.

Auxin reduces the synthesis of major vacuolar proteins in tobacco mesophyl protoplast.

We have established that polypeptides whose synthesis is reduced by 2,4-dichlorophenoxyacetic acid during in vitro culture of tobacco mesophyll protoplasts are secreted into the vacuole where they constitute the bulk of labeled proteins. In addition, these proteins continue to be synthesized in protoplast-derived cultured cells and their synthesis is strictly correlated with the size of the cell, i.e. with vacuolar size.

Auxin-induced regulation of protein synthesis in tobacco mesophyll protoplasts cultivated in vitro: I. Characteristics of auxin-sensitive proteins.

The presence of auxin (2,4-D), in the culture medium of tobacco (Nicotiana tabacum var Maryland) mesophyll protoplasts is necessary both for cell wall regeneration and for passage of the cells from phase G(0) to phase G(1) of the cell cycle. Among about 250 proteins synthesized by protoplasts and characterized by their migration in a two-dimensional electrophoresis gel, 2,4-dichlorophenoxyacetic acid affects the synthesis of 11.Nine proteins are synthesized at a reduced level in the presence of the hormone, of which three are rapidly labeled and short-lived, while the others, which are long-lived, become detectable only after 2 hours of radioactive labeling, suggesting that they undergo slow posttranslational maturation. These nine proteins are proline-rich but the proline radicals are not strongly hydroxylated. The synthesis of these proteins is no longer inhibited by auxin if dichlorobenzonitril, a weed-killer which inhibits cell wall reformation of tobacco protoplasts, is added to the culture medium.Two proteins are only synthesized if protoplasts are cultivated in an auxin-containing medium. These polypeptides are rapidly labeled, and are long-lived. The inhibition of cell wall reformation by dichlorobenzonitril does not modify their synthesis.These results suggest that proteins whose synthesis is reduced by auxin are related to cell wall reformation and that they do not play a role in the induction of the cell cycle. In contrast, proteins whose synthesis is stimulated in the presence of auxin are good candidates for a role in the induction of the cell cycle.

Auxin-Induced Regulation of Protein Synthesis in Tobacco Mesophyll Protoplasts Cultivated In Vitro: II. Time Course and Level of Auxin Control.

When protoplasts, previously cultivated in a medium lacking auxin, are transferred to complete medium, proteins whose synthesis is stimulated by the hormone become detectable after about 30 minutes and reach a constant level 2 to 4 hours after the beginning of hormonal treatment. In contrast, proteins whose level of synthesis is reduced by auxin, are only affected after 6 hours of treatment.Short radioactive labelings in deficient medium followed by chases in complete medium show that auxin does not interfere with posttranslational processes.Analysis of in vitro translation products of protoplast RNA shows that the time courses of auxin effects on protein synthesis and mRNA accumulation are perfectly superimposable. This allows us to exclude the possibility that auxin affects the translation process, but indicates that this hormone acts by regulating the concentration of the auxin-sensitive protein mRNAs.

Long-lived and short-lived heat-shock proteins in tobacco mesophyll protoplasts.

We have studied modifications in the pattern of proteins synthesized by tobacco (Nicotiana tabacum var Maryland) mesophyll protoplasts when they are transferred from 25 degrees C to 40 degrees C. The synthesis of one group of proteins is practically unaffected by the heat shock. On the other hand, the synthesis of most other 25 degrees C proteins is greatly reduced, while specific heat-shock proteins appear: 17 stable, neutral, major proteins, which are synthesized throughout the culture period at the higher temperature and which correspond to those observed in other organisms, and two basic proteins with a short lifetime and which are synthesized only during the first 2 hours of heat shock. We suggest that these latter proteins are regulatory peptides which intervene in the inhibition of 25 degrees C syntheses.

Hormonal Control of Mitotic Development in Tobacco Protoplasts: TWO-DIMENSIONAL DISTRIBUTION OF NEWLY-SYNTHESIZED PROTEINS.

Two-dimensional separation of proteins newly synthesized by tobacco mesophyll protoplasts cultivated in vitro allows us to detect, reproducibly, 257 spots. The pattern is extremely stable throughout the three days of culture, the intensity of only 24 spots varying during this time. The absence of cytokinin (N(6)-benzyladenine) in the culture medium prohibits entry into S phase but does not modify the pattern, indicating that none of the observed proteins is specifically synthesized in S, G(2), or M phases. The presence of 2,4-dichlorophenoxyacetic acid is necessary for the mitotic development of protoplasts. It induces the appearance of one protein, increases the level of another, and reduces that of eight others. All proteins sensitive to auxin belong to the group of proteins the levels of which vary during culture.