Characterization of the carboxysomal carbonic anhydrase CsoSCA from Halothiobacillus neapolitanus

In cyanobacteria and many chemolithotrophic bacteria, the CO(2)-fixing enzyme ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) is sequestered into polyhedral protein bodies called carboxysomes. The carboxysome is believed to function as a microcompartment that enhances the catalytic efficacy of RubisCO by providing the enzyme with its substrate, CO(2), through the action of the shell protein CsoSCA, which is a novel carbonic anhydrase. In the work reported here, the biochemical properties of purified, recombinant CsoSCA were studied, and the catalytic characteristics of the carbonic anhydrase for the CO(2) hydration and bicarbonate dehydration reactions were compared with those of intact and ruptured carboxysomes. The low apparent catalytic rates measured for CsoSCA in intact carboxysomes suggest that the protein shell acts as a barrier for the CO(2) that has been produced by CsoSCA through directional dehydration of cytoplasmic bicarbonate. This CO(2) trap provides the sequestered RubisCO with ample substrate for efficient fixation and constitutes a means by which microcompartmentalization enhances the catalytic efficiency of this enzyme.     

Nitrous oxide and methane fluxes of a pristine slope mire in the German National Park Harz Mountains

Pristine peatlands covered by Histosols (bogs and fens) with high water table and a restricted oxygen (O₂) availability are known to have low emissions of nitrous oxide (N₂O) but may be a significant source for atmospheric methane (CH₄) which are both important greenhouse gases. For the first time N₂O and CH₄ fluxes of a pristine slope mire in the German Harz Mountains have been monitored. Previously reported peatlands are characterised by anaerobic conditions due to high water table levels. Slope mires monitored here receive O₂ through slope water inflow. Gas fluxes have been monitored deploying closed chamber method on a central non-forested area and a forested area at the periphery of the slope mire. By means of groundwater piezometers water table levels, ammonium and nitrate contents as well as hydro-chemical variables like oxygen content and redox potential of the mire pore water have been concurrently measured with trace gas fluxes at both monitoring sites of the slope mire. The slope mire took up small amounts of atmospheric methane at a rate of −0.02 ± 0.01 kg C ha⁻¹ year⁻¹ revealing no significant difference between the forested and non-forested site. Higher uptake rates were observed during low water table level. In contrast to pristine peatlands influx of oxygen containing pore water into slope mire does limit reduction processes and resultant CH₄ emission. N₂O fluxes of the forested and non-forested sites of the slope mire did not differ and amounted to 0.25 ± 0.44 kg N ha⁻¹ year⁻¹. Higher emissions were observed at low water table levels and during thawing periods. In spite of favourable conditions N₂O fluxes of the slope mire have been comparable to those of pristine peatlands.     

Amplified immunohistochemical detection of PrPsc in animal transmissible spongiform encephalopathies using streptomycin.

Due to its sensitivity, immunohistochemistry (IHC) of abnormal prion protein (PrPsc) is used to study experimental and natural cases of transmissible spongiform encephalopathies (TSEs) such as Creutzfeldt-Jakob disease in humans or scrapie and bovine spongiform encephalopathy (BSE) in animals. The limits of detection are particularly critical when PrPsc IHC is used for diagnostic purposes. In this article, we describe for the first time the use of streptomycin sulfate in IHC, providing a novel original and easy way to amplify specifically PrPsc immunohistochemical detection in natural cases of BSE and scrapie, as well as in experimental TSEs in mice models using two different PrP antibodies.     

Trehalose-6-phosphate-mediated phenotypic change in Acinetobacter baumannii

The stress protectant trehalose is synthesized in Acinetobacter baumannii from UPD-glucose and glucose-6-phosphase via the OtsA/OtsB pathway. Previous studies proved that deletion of otsB led to a decreased virulence, the inability to grow at 45°C and a slight reduction of growth at high salinities indicating that trehalose is the cause of these phenotypes. We have questioned this conclusion by producing ∆otsA and ∆otsBA mutants and studying their phenotypes. Only deletion of otsB, but not deletion of otsA or otsBA, led to growth impairments at high salt and high temperature. The intracellular concentrations of trehalose and trehalose-6-phosphate were measured by NMR or enzymatic assay. Interestingly, none of the mutants accumulated trehalose any more but the ∆otsB mutant with its defect in trehalose-6-phosphate phosphatase activity accumulated trehalose-6-phosphate. Moreover, expression of otsA in a ∆otsB background under conditions where trehalose synthesis is not induced led to growth inhibition and the accumulation of trehalose-6-phosphate. Our results demonstrate that trehalose-6-phosphate affects multiple physiological activities in A. baumannii ATCC 19606.     

Evolutionary significance of the variation in acoustic communication of a cryptic nocturnal primate radiation (Microcebus spp.)

Acoustic phenotypic variation is of major importance for speciation and the evolution of species diversity. Whereas selective and stochastic forces shaping the acoustic divergence of signaling systems are well studied in insects, frogs, and birds, knowledge on the processes driving acoustic phenotypic evolution in mammals is limited. We quantified the acoustic variation of a call type exchanged during agonistic encounters across eight distinct species of the smallest‐bodied nocturnal primate radiation, the Malagasy mouse lemurs. The species live in two different habitats (dry forest vs. humid forest), differ in geographic distance to each other, and belong to four distinct phylogenetic clades within the genus. Genetically defined species were discriminated reliably on the phenotypic level based on their acoustic distinctiveness in a discriminant function analysis. Acoustic variation was explained by genetic distance, whereas differences in morphology, forest type, or geographic distance had no effect. The strong impact of genetics was supported by a correlation between acoustic and genetic distance and the high agreement in branching pattern between the acoustic and molecular phylogenetic trees. In sum, stochastic factors such as genetic drift best explained acoustic diversification in a social communication call of mouse lemurs.     

Two calcium‐binding chaperones from the fat body of the Colorado potato beetle,Leptinotarsa decemlineata (Coleoptera: Chrysomelidae) involved in diapause

Molecular chaperones are crucial for the correct folding of newly synthesized polypeptides, in particular, under stress conditions. Various studies have revealed the involvement of molecular chaperones, such as heat shock proteins, in diapause maintenance and starvation; however, the role of other chaperones in diapause and starvation relatively is unknown. In the current study, we identified two lectin‐type chaperones with calcium affinity, a calreticulin (LdCrT) and a calnexin (LdCnX), that were present in the fat body of the Colorado potato beetle, Leptinotarsa decemlineata (Coleoptera: Chrysomelidae) during diapause. Both proteins possessed an N‐globular domain, a P‐arm domain, and a highly charged C‐terminal domain, while an additional transmembrane domain was present in LdCnX. Phylogenetic analysis revealed distinction at the order level. Both genes were expressed in multiple tissues in larval and adult stages, and constitutively throughout development, though a starvation response was detected only for LdCrT. In females, diapause‐related expression analysis in the whole body revealed an upregulation of both genes by post‐diapause, but a downregulation by diapause only for LdCrT. By contrast, males revealed no alteration in their diapause‐related expression pattern in the entire body for both genes. Fat body‐specific expression analysis of both genes in relation to diapause revealed the same expression pattern with no alteration in females and downregulation in males by post‐diapause. This study suggests that calcium‐binding chaperones play similar and possibly gender‐specific roles during diapause.     

Ultra‐Conserved Elements and morphology reciprocally illuminate conflicting phylogenetic hypotheses in Chalcididae (Hymenoptera,Chalcidoidea)

Recent technical advances combined with novel computational approaches have promised the acceleration of our understanding of the tree of life. However, when it comes to hyperdiverse and poorly known groups of invertebrates, studies are still scarce. As published phylogenies will be rarely challenged by future taxonomists, careful attention must be paid to potential analytical bias. We present the first molecular phylogenetic hypothesis for the family Chalcididae, a group of parasitoid wasps, with a representative sampling (144 ingroups and seven outgroups) that covers all described subfamilies and tribes, and 82% of the known genera. Analyses of 538 Ultra‐Conserved Elements (UCEs) with supermatrix (RAx ML and IQTREE) and gene tree reconciliation approaches (ASTRAL, ASTRID) resulted in highly supported topologies in overall agreement with morphology but reveal conflicting topologies for some of the deepest nodes. To resolve these conflicts, we explored the phylogenetic tree space with clustering and gene genealogy interrogation methods, analyzed marker and taxon properties that could bias inferences and performed a thorough morphological analysis (130 characters encoded for 40 taxa representative of the diversity). This joint analysis reveals that UCEs enable attainment of resolution between ancestry and convergent/divergent evolution when morphology is not informative enough, but also shows that a systematic exploration of bias with different analytical methods and a careful analysis of morphological features is required to prevent publication of artifactual results. We highlight a GC content bias for maximum‐likelihood approaches, an artifactual mid‐point rooting of the ASTRAL tree and a deleterious effect of high percentage of missing data (>85% missing UCEs) on gene tree reconciliation methods. Based on the results we propose a new classification of the family into eight subfamilies and ten tribes that lay the foundation for future studies on the evolutionary history of Chalcididae.