MiaB protein from Thermotoga maritima. Characterization of an extremely thermophilic tRNA-methylthiotransferase

In Escherichia coli, the MiaB protein catalyzes the methylthiolation of N-6-isopentenyl adenosine in tRNAs, the last reaction step during biosynthesis of 2-methylthio-N-6-isopentenyl adenosine (ms2i6A-37). For the first time the thermophilic bacterium Thermotoga maritima is shown here to contain such a MiaB tRNA-modifying enzyme, named MiaBTm, and to synthesize ms2i6A-37 as demonstrated by an analysis of modified nucleosides from tRNA hydrolysates. The corresponding gene (TM0653) was identified by sequence similarity to the miaB gene cloned and expressed in E. coli. MiaBTm was purified to homogeneity and thoroughly characterized by biochemical and spectroscopic methods. It is a monomer of 443 residues with a molecular mass of 50,710 kilodaltons. Its amino acid sequence shares the CysXXX-CysXXCys sequence with MiaB from E. coli as well as with biotin synthase and lipoate synthase. This sequence was shown to be essential for chelation of an iron-sulfur center and for activity in these enzymes. As isolated, MiaBTm contains both iron and sulfide and an apoprotein form can coordinate up to 4 iron and 4 sulfur atoms per polypeptide chain. UV-visible absorption, resonance Raman, variable temperature magnetic circular dichroism, and EPR spectroscopy of MiaBTm indicate the presence of a [4Fe-4S]+2/+1 cluster under reducing and anaerobic conditions, whereas [3Fe-4S]+1 and [2Fe-2S]+2 forms are generated under aerobic conditions. The redox potential of the [4Fe-4S]+2/+1 transition is -495 +/- 10 mV (versus the normal hydrogen electrode). Finally, the expression of MiaBTm from T. maritima in an E. coli mutant strain lacking functional miaB gene allowed production of ms2i6A-37. These results provide further information on the enzymes involved in methylthiolation of tRNAs.     

Sulphated heteropolysaccharides from Padina pavonia

Extraction with hydrochloric acid (pH 2.5) of the brown alga Padina pavonia afforded water-soluble and water-insoluble polysaccharides comprising D-glucuronic acid, L-fucose, D-xylose, D-mannose, D-glucose and D- galactose residues. The water-soluble polysaccharide was fractionated by using ethanol, and cetylpyridinium chloride and by chromatography on DEAE-cellulose. A neutral laminaran-like glucan, a sulphated heteropolysaccharide composed of the aforementioned sugars and a protein moiety were obtained. The isolated heteropolysaccharide showed high anticoagulation activity.     

Differentiation of Fanconi anemia and aplastic anemia using mitomycin C test in Tunisia

Fanconi anemia (FA) is a recessive chromosomal instability syndrome that is clinically characterized by multiple symptoms. Chromosome breakage hypersensitivity to alkylating agents is the gold standard test for FA diagnosis. In this study, we provide a detailed laboratory protocol for accurate assessment of FA diagnosis based on mitomycin C (MMC) test. Induced chromosomal breakage study was successful in 171 out of 205 aplastic anemia (AA) patients. According to the sensitivity of MMC at 50 ng/ml, 38 patients (22.22%) were diagnosed as affected and 132 patients (77.17%) as unaffected. Somatic mosaicism was suspected in an 11-year-old patient with a FA phenotype. Twenty-six siblings of FA patients were also evaluated and five of them (19.23%) were diagnosed as FA. From this study, a standard protocol for diagnosis of FA was developed. It is routinely used as a diagnostic test of FA in Tunisia.     

Oral Ingestion of Transgenic RIDL Ae. aegypti Larvae Has No Negative Effect on Two Predator Toxorhynchites Species

Dengue is the most important mosquito-borne viral disease. No specific treatment or vaccine is currently available; traditional vector control methods can rarely achieve adequate control. Recently, the RIDL (Release of Insect carrying Dominant Lethality) approach has been developed, based on the sterile insect technique, in which genetically engineered ‘sterile’ homozygous RIDL male insects are released to mate wild females; the offspring inherit a copy of the RIDL construct and die. A RIDL strain of the dengue mosquito, Aedes aegypti, OX513A, expresses a fluorescent marker gene for identification (DsRed2) and a protein (tTAV) that causes the offspring to die. We examined whether these proteins could adversely affect predators that may feed on the insect. Aedes aegypti is a peri-domestic mosquito that typically breeds in small, rain-water-filled containers and has no specific predators. Toxorhynchites larvae feed on small aquatic organisms and are easily reared in the laboratory where they can be fed exclusively on mosquito larvae. To evaluate the effect of a predator feeding on a diet of RIDL insects, OX513A Ae. aegypti larvae were fed to two different species of Toxorhynchites (Tx. splendens and Tx. amboinensis) and effects on life table parameters of all life stages were compared to being fed on wild type larvae. No significant negative effect was observed on any life table parameter studied; this outcome and the benign nature of the expressed proteins (tTAV and DsRed2) indicate that Ae. aegypti OX513A RIDL strain is unlikely to have any adverse effects on predators in the environment.     

Effect of organic and mineral fertilizers on N-use by wheat under different irrigation frequencies

A field trial was established in Errachidia, southern Morocco, to investigate the interaction between wheat residue management and mineral 15N-labelled ammonium sulphate, under different irrigation treatments, applied to wheat (Triticum durum var. Karim). In treatments I1, I2, I3 and I4, plots were irrigated every 10, 15, 21 and 30 days. Each plot contained three sub-plots that received three fertilization treatments: T1 received 42 kg N ha-1 of ammonium sulphate before seedling, 42 kg N ha-1 of ammonium sulphate labelled with 9.764 at % 15N excess at tillering and 84 N kg ha-1 of ammonium sulphate at flowering; T2 received 42 kg N ha-1 of ammonium sulphate labelled with 9.764 at % 15N excess at seedling, 42 kg N ha-1 at tillering and 42 kg N ha-1 at flowering; T3 received 4800 kg ha-1 of wheat residue labelled with 1.504 at % 15N excess and 42 kg N ha-1 of ammonium sulphate before seedling and 42 kg N ha-1 of ammonium sulphate at flowering. Nitrogen fertilization with 168 kg N ha-1 did no significantly increase grain and straw yields in comparison to the 126 kg N ha-1 application. The combination of the organic input and supplementary application of mineral fertilizer N has been found as a more attractive management option. For all irrigation treatments, the % recovery of N in the whole plant was higher in plants that received 15N at tillering (63%, 49% respectively for irrigation intervals between 10 and 30 d) than in plants that received 15N just after seeding (28% for irrigation each 10- and 30-d intervals). For the irrigation treatment each 10 and 15 days, the 15N was mainly recovered by the grain for all fertilization treatments, whereas for irrigation treatment each 30 days, the grain and straw recovered nearly equal amounts of fertilizer. For grain and straw of wheat, nitrogen in the plant derived from the fertilizer was low, while most of the N was derived from the soil for all irrigation and fertilization treatments. The % nitrogen in the plant derived from the fertilizer values showed no significant difference between the different plant parts. The results suggested a dominant influence of moisture availability on the fertilizer N uptake by wheat. Under dry conditions the losses of N can be allotted to denitrification and volatilisation.     

Characterization and gene mapping of a chlorophyll-deficient mutant clm1 of Triticum monococcum L.

Diploid wheat Triticum monococcum L. is a model plant for wheat functional genomics. Chlorophyll-deficient mutant (clm1) was identified during manual screening of the ethyl methanesulphonate (EMS)-treated M₂ progenies of T. monococcum accession pau14087 in the field. The clm1 mutant, due to significantly decreased chlorophyll content compared with the wild-type (WT), exhibited pale yellow leaves which slowly recovered to green before flowering. The clm1 mutant showed early flowering, reduced number of tillers, trichome length and density, and different shape as compared with the WT. At the same time, clm1 mutant culm had more chlorophyll-containing parenchymatous tissues compared to WT, presumably to absorb more sunlight for photosynthesis. Genetic analysis indicated that the clm1 mutation was monogenic recessive. The clm1 mutant was mapped between Xgwm473 and Xwmc96 SSR markers, with genetic distances of 2.1 and 2.6 cM, respectively, on the 7A^mL chromosome.     

Effet du climat et de l’exposition sur la dynamique des populations de la cochenille violette,Parlatoria oleae Colvée (Hemiptera : Diaspididae), en conditions arides

Résumé

Ce travail a pour objectif l’étude de quelques aspects bio-écologiques de la cochenille violette, Parlatoria oleae Colvée 1880, bio-agresseur des cultures de l’Olivier en régions arides. Le suivi du cycle biologique ainsi que la démo-écologie de ce ravageur ont été réalisés grâce à des dénombrements périodiques des populations sur les différents organes de l’arbre (méthodes de Vasseur &; Schvester) de décembre 2010 à décembre 2011 dans la région d’Ain Touta (nord-est algérien). L’espèce y a montré deux générations par an : une génération printanière se développant entre avril et juillet et une génération automnale évoluant entre août et octobre. La ponte débuta en avril et s’échelonna jusqu‘à la fin septembre. L’exposition nord est la plus favorable à cette diaspine qui y trouve des conditions microclimatiques optimales pour son développement. La ponte moyenne est de 8 à 9 ?ufs par femelle. L’analyse statistique de l’effet des conditions climatiques étudiées (températures minimale, maximale et moyenne ; précipitations, gelée et indice d’aridité De Martone) sur les effectifs des différents stades, montre une grande variabilité d’un stade à un autre. L’analyse statistique établie révèle également que les effectifs de l’espèce présentent des variations très hautement significatives selon l’orientation dans l’arbre d’olive colonisé.     

PTPN22 gene polymorphism in Egyptian alopecia areata patients and its impact on response to diphencyprone immunotherapy

PTPN22 1858C>T gene polymorphism has been associated with several autoimmune disorders including alopecia areata. The aim of the current study was to investigate the effect of the inherited genetic polymorphism 1858C>T of PTPN22 gene on the predisposition to severe forms of alopecia areata and its effect on the response to DPC treatment. To achieve our aim, PTPN22 1858C>T genotyping was performed by PCR-based restricted fragment length polymorphism (PCR-RFLP) analysis. The study included 103 Egyptian patients with extensive alopecia areata treated by DPC. Hundred healthy age and sex matched blood donors were included in the current study as a control group. Results of genotyping showed that PTPN22 CT and TT mutant genotypes were significantly higher in AA patients compared to controls and conferred increase risk of AA (OR = 2.601, 95% CI = 1.081–6.255). Statistical comparison between AA patients with wild and mutant genotypes revealed that the duration of the illness was significantly longer in those harboring the mutant genotypes. Moreover, the association of other autoimmune diseases as atopy and diabetes mellitus was higher in patients with mutant genotypes. Furthermore, PTPN22 1858C>T genetic polymorphism did not affect the patients' response to DPC immunotherapy.