Characterization of two temperature-sensitive mutants of Sigma virus (authors transl)

Summary Temperature-sensitive mutants have been isolated from Drosophila Sigma virus, a Rhabdovirus inducing CO₂ sensitivity in Drosophila melanogaster. We have studied the decay of infectious centers at non permissive temperature. The proportion of destroyed infectious centers is the same for the wild type, ts⁺, and for ts9. On the opposite, it is more important for ts 4. Temperature-sensitive function of ts 4 appears necessary to the viral genome replication. With the three clones, ts⁺, ts 4 and ts 9, we have obtained stabilized Drosophila females able to transmit Sigma virus to their whole progeny. We have tried to see in each case, if stabilized flies could transmit the virus to their progeny at non permissive temperature. Flies stabilized with ts⁺ and ts 9 can, flies stabilized with ts 4 cannot. Therefore two categories of mutants are defined: those that are transmitted hereditarily. at non permissive temperature, and not blocked in genome replication. Those that are blocked in genome replication and not transmitted. When the virus cannot replicate, the divisions in the germ line cells dilute the viral genomes. The consequence will be a real healing of germ line cells, and then a break in hereditary transmission by stabilized flies. All the results with temperature-sensitive mutants are coherent with this hypothesis.

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Étude de mutants thermosensibles du virus Sigma

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Mémoire présenté par F. Gros     

Activated T cells complicate the identification of regulatory T cells in rheumatoid arthritis

Most cell surface markers for CD4⁺CD25⁺ regulatory T cells (Tregs) are also expressed by activated non-regulatory T cells. Recently, CD127 down-regulation was found to identify functional Tregs in healthy individuals, but there are no data from patients with inflammatory conditions. We examined peripheral blood mononuclear cells (PBMC) from rheumatoid arthritis patients with active inflammation and from healthy controls, and found that CD4⁺ T cells contained an equal proportion of CD25⁺CD127⁻/low cells in both groups. In patients, not all these cells expressed intracellular FOXP3. Upon activation by anti-CD3/anti-CD28, PBMC rapidly down-regulated CD127, while FOXP3 up-regulation was transitory and occurred in fewer cells. The activated cells were not anergic to restimulation and had no suppressive effects. The distinct kinetics indicate that the FOXP3⁻CD127⁻/low cells in rheumatoid arthritis patients most likely represent activated non-regulatory T cells. This complicates the use of CD127 for identification of Tregs in inflammatory diseases.     

The ubiquitin-proteasome and the mitochondria-associated apoptotic pathways are sequentially downregulated during recovery after immobilization-induced muscle atrophy

Immobilization produces morphological, physiological, and biochemical alterations in skeletal muscle leading to muscle atrophy and long periods of recovery. Muscle atrophy during disuse results from an imbalance between protein synthesis and proteolysis but also between apoptosis and regeneration processes. This work aimed to characterize the mechanisms underlying muscle atrophy and recovery following immobilization by studying the regulation of the mitochondria-associated apoptotic and the ubiquitin-proteasome-dependent proteolytic pathways. Animals were subjected to hindlimb immobilization for 4-8 days (I4 to I8) and allowed to recover after cast removal for 10-40 days (R10 to R40). Soleus and gastrocnemius muscles atrophied from I4 to I8 to a greater extent than extensor digitorum longus and tibialis anterior muscles. Gastrocnemius muscle atrophy was first stabilized at R10 before being progressively reduced until R40. Polyubiquitinated proteins accumulated from I4, whereas the increased ubiquitination rates and chymotrypsin-like activity of the proteasome were detectable from I6 to I8. Apoptosome and caspase-3 or -9 activities increased at I6 and I8, respectively. The ubiquitin-proteasome-dependent pathway was normalized early when muscle stops to atrophy (R10). By contrast, the mitochondria-associated apoptotic pathway was first downregulated below basal levels when muscle started to recover at R15 and completely normalized at R20. Myf 5 protein levels decreased from I4 to I8 and were normalized at R10. Altogether, our results suggest a two-stage process in which the ubiquitin-proteasome pathway is rapidly up- and downregulated when muscle atrophies and recovers, respectively, whereas apoptotic processes may be involved in the late stages of atrophy and recovery.     

Agreement among health care professionals in diagnosing case Vignette-based surgical site infections

Objective

To assess agreement in diagnosing surgical site infection (SSI) among healthcare professionals involved in SSI surveillance.

Methods

Case-vignette study done in 2009 in 140 healthcare professionals from seven specialties (20 in each specialty, Anesthesiologists, Surgeons, Public health specialists, Infection control physicians, Infection control nurses, Infectious diseases specialists, Microbiologists) in 29 University and 36 non-University hospitals in France. We developed 40 case-vignettes based on cardiac and gastrointestinal surgery patients with suspected SSI. Each participant scored six randomly assigned case-vignettes before and after reading the SSI definition on an online secure relational database. The intraclass correlation coefficient (ICC) was used to assess agreement regarding SSI diagnosis on a seven-point Likert scale and the kappa coefficient to assess agreement for superficial or deep SSI on a three-point scale.

Results

Based on a consensus, SSI was present in 21 of 40 vignettes (52.5%). Intraspecialty agreement for SSI diagnosis ranged across specialties from 0.15 (95% confidence interval, 0.00–0.59) (anesthesiologists and infection control nurses) to 0.73 (0.32–0.90) (infectious diseases specialists). Reading the SSI definition improved agreement in the specialties with poor initial agreement. Intraspecialty agreement for superficial or deep SSI ranged from 0.10 (−0.19–0.38) to 0.54 (0.25–0.83) (surgeons) and increased after reading the SSI definition only among the infection control nurses from 0.10 (−0.19–0.38) to 0.41 (−0.09–0.72). Interspecialty agreement for SSI diagnosis was 0.36 (0.22–0.54) and increased to 0.47 (0.31–0.64) after reading the SSI definition.

Conclusion

Among healthcare professionals evaluating case-vignettes for possible surgical site infection, there was large disagreement in diagnosis that varied both between and within specialties.     

Maternal hypertension induces tissue-specific modulations of the apelinergic system in the fetoplacental unit in rat

Apelin and its receptor APJ are expressed in fetal tissues but their function and regulation remain largely unknown. In rat, maternal treatment with a nitric oxide synthase inhibitor inducing hypertension was used to investigate apelin plasma levels in mother/fetus pairs and on the gene expression level of the apelin/APJ system in fetal tissues and placenta. At term, plasma levels of apelin were not modulated but APJ expression was increased in placenta and lung but reduced in heart. Apelin expression was increased only in the heart. We postulate that the apelinergic system may control fetal growth and cardiovascular functions in utero.     

Dynamics of G6PD activity in patients receiving weekly primaquine for therapy of Plasmodium vivax malaria

BackgroundAcute Plasmodium vivax malaria is associated with haemolysis, bone marrow suppression, reticulocytopenia, and post-treatment reticulocytosis leading to haemoglobin recovery. Little is known how malaria affects glucose-6-phosphate dehydrogenase (G6PD) activity and whether changes in activity when patients present may lead qualitative tests, like the fluorescent spot test (FST), to misdiagnose G6PD deficient (G6PDd) patients as G6PD normal (G6PDn). Giving primaquine or tafenoquine to such patients could result in severe haemolysis.MethodsWe investigated the G6PD genotype, G6PD enzyme activity over time and the baseline FST phenotype in Cambodians with acute P. vivax malaria treated with 3-day dihydroartemisinin piperaquine and weekly primaquine, 0·75 mg/kg x8 doses.ResultsOf 75 recruited patients (males 63), aged 5–63 years (median 24), 15 were G6PDd males (14 Viangchan, 1 Canton), 3 were G6PD Viangchan heterozygous females, and 57 were G6PDn; 6 patients had α/β-thalassaemia and 26 had HbE.Median (range) Day0 G6PD activities were 0·85 U/g Hb (0·10–1·36) and 11·4 U/g Hb (6·67–16·78) in G6PDd and G6PDn patients, respectively, rising significantly to 1·45 (0·36–5·54, p<0.01) and 12·0 (8·1–17·4, p = 0.04) U/g Hb on Day7, then falling to ~Day0 values by Day56. Day0 G6PD activity did not correlate (p = 0.28) with the Day0 reticulocyte counts but both correlated over time. The FST diagnosed correctly 17/18 G6PDd patients, misclassifying one heterozygous female as G6PDn.ConclusionsIn Cambodia, acute P. vivax malaria did not elevate G6PD activities in our small sample of G6PDd patients to levels that would result in a false normal qualitative test. Low G6PDd enzyme activity at disease presentation increases upon parasite clearance, parallel to reticulocytosis. More work is needed in G6PDd heterozygous females to ascertain the effect of P. vivax on their G6PD activities.Trial registrationThe trial was registered (ACTRN12613000003774) with the Australia New Zealand Clinical trials (https://www.anzctr.org.au/Trial/Registration/TrialReview.aspx?id=363399&isReview=true).     

Novel Virus Influenza A (H1N1sw) in South-Eastern France,April-August 2009

Background

In April 2009, the first cases of pandemic (H1N1)-2009 influenza [H1N1sw] virus were detected in France. Virological surveillance was undertaken in reference laboratories of the seven French Defence Zones.

Methodology/Principal Findings

We report results of virological analyses performed in the Public Hospitals of Marseille during the first months of the outbreak. (i) Nasal swabs were tested using rapid influenza diagnostic test (RIDT) and two RT-PCR assays. Epidemiological characteristics of the 99 first suspected cases were analyzed, including detection of influenza virus and 18 other respiratory viruses. During three months, a total of 1,815 patients were tested (including 236 patients infected H1N1sw virus) and distribution in age groups and results of RIDT were analyzed. (ii) 600 sera received before April 2009 and randomly selected from in-patients were tested by a standard hemagglutination inhibition assay for antibody to the novel H1N1sw virus. (iii) One early (May 2009) and one late (July 2009) viral isolates were characterized by sequencing the complete hemagglutinine and neuraminidase genes. (iiii) Epidemiological characteristics of a cluster of cases that occurred in July 2009 in a summer camp were analyzed.

Conclusions/Significance

This study presents new virological and epidemiological data regarding infection by the pandemic A/H1N1 virus in Europe. Distribution in age groups was found to be similar to that previously reported for seasonal H1N1. The first seroprevalence data made available for a European population suggest a previous exposure of individuals over 40 years old to influenza viruses antigenically related to the pandemic (H1N1)-2009 virus. Genomic analysis indicates that strains harbouring a new amino-acid pattern in the neuraminidase gene appeared secondarily and tended to supplant the first strains. Finally, in contrast with previous reports, our data support the use of RIDT for the detection of infection in children, especially in the context of the investigation of grouped cases.     

Formyl peptide receptors: a promiscuous subfamily of G protein-coupled receptors controlling immune responses

The formyl peptide receptor (FPR) family is involved in host defence against pathogens, but also in sensing internal molecules that may constitute signals of cellular dysfunction. It includes three subtypes in human and other primates. FPR responds to formyl peptides derived from bacterial and mitochondrial proteins. FPRL1 displays a large array of exogenous and endogenous ligands, including the chemokine variant sCKβ8-1, the neuroprotective peptide humanin, and lipoxin A4. Two high affinity agonists (F2L and humanin) were recently described for FPRL2. In mouse, eight FPR-related receptors have been described. Fpr1 is the ortholog of human FPR, while fpr2 appears to share many ligands with human FPRL1. Altogether, the physiological role of the FPR family is still incompletely understood, due in part to the large variety of ligands, the redundancy with other chemoattractant agents, and the lack of clear orthologs between human and mouse receptors. Newly developed tools will allow to study further this family of receptors.     

Hyperhydricity of Prunus avium shoots cultured on gelrite: a controlled stress response.

Hyperhydricity is a physiological disorder frequently affecting shoots vegetatively propagated in vitro. Hyperhydric shoots are characterised by a translucent aspect due to a chlorophyll deficiency, a not very developed cell wall and a high water content. Hyperhydricity of Prunus avium shoots was expressed in vitro in one multiplication cycle by replacing the gelling agent agar (normal shoots: NS) by gelrite (hyperhydric shoots: HS). P. avium shoots evolving towards the hyperhydric state produced higher amounts of ethylene, polyamines (PAs) and proline, which are substances considered as stress markers. A higher activity of glutathione peroxidase (GPX; EC 1.11.1.9), involved in organic hydroperoxide elimination, suggested an increased production of these compounds in HS. The unchanged free fatty acid composition indicated no HS membrane damages compared to NS. The ploidy level of HS nuclei was not affected, but the bigger size and the lower percentage of nuclei during the S phase suggested a slowing down of the cell cycle. The results argued for a stress response of the HS, but no signs of oxidative damages of lipid membrane and nucleus were observed. The discussion points out paradoxical results in a classical analysis of stress and suggests an alternative way of defense mechanisms in HS, involving homeostatic regulation and controlled degradation processes to maintain integrity and vital functions of the cell.