MALDI‐TOF MS identification of microbiota associated with pest insect Diabrotica speciosa

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Fine litterfall in the Brazilian Atlantic Forest

Litterfall is an essential component of tropical forest productivity, transferring nutrients from the vegetation back to soils. Here, we summarize the data from 105 estimates of fine litterfall production from 45 sites in the Atlantic Forest domain, including two types of forests, evergreen and seasonal, and two successional stages, secondary and old growth. The overall litterfall average was 8.0 ± 2.5 Mg/ha. Litterfall was significantly in higher seasonal forests than in evergreen forests and in old growth versus secondary forests. Leaves were the major component of litterfall, contributing 68 percent to the total. The second most important component was branches, contributing 22 percent, followed by reproductive organs (flowers and fruits), at 6 percent. Accurate measurements of tropical forest productivity are crucial for estimating their role in sequestering atmospheric carbon, and we suggest some ways to standardize litterfall sampling to obtain better estimates.     

Learning brain dynamics for decoding and predicting individual differences

Insights from functional Magnetic Resonance Imaging (fMRI), as well as recordings of large numbers of neurons, reveal that many cognitive, emotional, and motor functions depend on the multivariate interactions of brain signals. To decode brain dynamics, we propose an architecture based on recurrent neural networks to uncover distributed spatiotemporal signatures. We demonstrate the potential of the approach using human fMRI data during movie-watching data and a continuous experimental paradigm. The model was able to learn spatiotemporal patterns that supported 15-way movie-clip classification (∼90%) at the level of brain regions, and binary classification of experimental conditions (∼60%) at the level of voxels. The model was also able to learn individual differences in measures of fluid intelligence and verbal IQ at levels comparable to that of existing techniques. We propose a dimensionality reduction approach that uncovers low-dimensional trajectories and captures essential informational (i.e., classification related) properties of brain dynamics. Finally, saliency maps and lesion analysis were employed to characterize brain-region/voxel importance, and uncovered how dynamic but consistent changes in fMRI activation influenced decoding performance. When applied at the level of voxels, our framework implements a dynamic version of multivariate pattern analysis. Our approach provides a framework for visualizing, analyzing, and discovering dynamic spatially distributed brain representations during naturalistic conditions.     

Candidatus Rickettsia andeanae,a spotted fever group agent infecting Amblyomma parvum ticks in two Brazilian biomes

Adult ticks of the species Amblyomma parvum were collected from the vegetation in the Pantanal biome (state of Mato Grosso do Sul) and from horses in the Cerrado biome (state of Piauí) in Brazil. The ticks were individually tested for rickettsial infection via polymerase chain reaction (PCR) targeting three rickettsial genes, gltA, ompA and ompB. Overall, 63.5% (40/63) and 66.7% (2/3) of A. parvum ticks from Pantanal and Cerrado, respectively, contained rickettsial DNA, which were all confirmed by DNA sequencing to be 100% identical to the corresponding fragments of the gltA, ompA and ompB genes of Candidatus Rickettsia andeanae. This report is the first to describe Ca. R. andeanae in Brazil.     

In house reverse membrane hybridisation assay versus GenoType MTBDRplus and their performance to detect mutations in the genes rpoB,katG and inhA

Drug-resistant tuberculosis (TB) threatens global TB control and is a major public health concern in several countries. We therefore developed a multiplex assay (LINE-TB/MDR) that is able to identify the most frequent mutations related to rifampicin (RMP) and isoniazid (INH) resistance. The assay is based on multiplex polymerase chain reaction, membrane hybridisation and colorimetric detection targeting of rpoB and katG genes, as well as the inhA promoter, which are all known to carry specific mutations associated with multidrug-resistant TB (MDR-TB). The assay was validated on a reference panel of 108 M. tuberculosis isolates that were characterised by the proportion method and by DNA sequencing of the targets. When comparing the performance of LINE-TB/MDR with DNA sequencing, the sensitivity, specificity and agreement were 100%, 100% and 100%, respectively, for RMP and 77.6%, 90.6% and 88.9%, respectively, for INH. Using drug sensibility testing as a reference standard, the performance of LINE-TB/MDR regarding sensitivity, specificity and agreement was 100%, 100% and 100% (95%), respectively, for RMP and 77%, 100% and 88.7% (82.2-95.1), respectively, for INH. LINE-TB/MDR was compared with GenoType MTBDRplus for 65 isolates, resulting in an agreement of 93.6% (86.7-97.5) for RIF and 87.4% (84.3-96.2) for INH. LINE-TB/MDR warrants further clinical validation and may be an affordable alternative for MDR-TB diagnosis.     

Glutathione Reductase-Mediated Synthesis of Tellurium-Containing Nanostructures Exhibiting Antibacterial Properties

Tellurium, a metalloid belonging to group 16 of the periodic table, displays very interesting physical and chemical properties and lately has attracted significant attention for its use in nanotechnology. In this context, the use of microorganisms for synthesizing nanostructures emerges as an eco-friendly and exciting approach compared to their chemical synthesis. To generate Te-containing nanostructures, bacteria enzymatically reduce tellurite to elemental tellurium. In this work, using a classic biochemical approach, we looked for a novel tellurite reductase from the Antarctic bacterium Pseudomonas sp. strain BNF22 and used it to generate tellurium-containing nanostructures. A new tellurite reductase was identified as glutathione reductase, which was subsequently overproduced in Escherichia coli. The characterization of this enzyme showed that it is an NADPH-dependent tellurite reductase, with optimum reducing activity at 30°C and pH 9.0. Finally, the enzyme was able to generate Te-containing nanostructures, about 68 nm in size, which exhibit interesting antibacterial properties against E. coli, with no apparent cytotoxicity against eukaryotic cells.     

Crystal structure determination and dynamic studies of Mycobacterium tuberculosis Cytidine deaminase in complex with products

Cytidine deaminase (CDA) is a key enzyme in the pyrimidine salvage pathway. It is involved in the hydrolytic deamination of cytidine or 2′-deoxycytidine to uridine or 2′-deoxyuridine, respectively. Here we report the crystal structures of Mycobacterium tuberculosis CDA (MtCDA) in complex with uridine (2.4 Å resolution) and deoxyuridine (1.9 Å resolution). Molecular dynamics (MD) simulation was performed to analyze the physically relevant motions involved in the protein–ligand recognition process, showing that structural flexibility of some protein regions are important to product binding. In addition, MD simulations allowed the analysis of the stability of tetrameric MtCDA structure. These findings open-up the possibility to use MtCDA as a target in future studies aiming to the rational design of new inhibitor of MtCDA-catalyzed chemical reaction with potential anti-proliferative activity on cell growth of M. tuberculosis, the major causative agent of tuberculosis.     

On the chemical mechanism of succinic semialdehyde dehydrogenase (GabD1) from Mycobacterium tuberculosis

Succinic semialdehyde dehydrogenases (SSADHs) are ubiquitous enzymes that catalyze the NAD(P)⁺-coupled oxidation of succinic semialdehyde (SSA) to succinate, the last step of the γ-aminobutyrate shunt. Mycobacterium tuberculosis encodes two paralogous SSADHs (gabD1 and gabD2). Here, we describe the first mechanistic characterization of GabD1, using steady-state kinetics, pH-rate profiles, ¹H NMR, and kinetic isotope effects. Our results confirmed SSA and NADP⁺ as substrates and demonstrated that a divalent metal, such as Mg²⁺, linearizes the time course. pH-rate studies failed to identify any ionizable groups with pK_a between 5.5 and 10 involved in substrate binding or rate-limiting chemistry. Primary deuterium, solvent and multiple kinetic isotope effects revealed that nucleophilic addition to SSA is very fast, followed by a modestly rate-limiting hydride transfer and fast thioester hydrolysis. Proton inventory studies revealed that a single proton is associated with the solvent-sensitive rate-limiting step. Together, these results suggest that product dissociation and/or conformational changes linked to it are rate-limiting. Using structural information for the human homolog enzyme and ¹H NMR, we further established that nucleophilic attack takes place at the Si face of SSA, generating a thiohemiacetal with S stereochemistry. Deuteride transfer to the Pro-R position in NADP⁺ generates the thioester intermediate and [4A-²H, 4B-¹H] NADPH. A chemical mechanism based on these data and the structural information available is proposed.     

Dual diaminopimelate biosynthesis pathways in Bacteroides fragilis and Clostridium thermocellum

Bacteroides fragilis and Clostridium thermocellum were recently found to synthesize diaminopimelate (DAP) by way of LL-DAP aminotransferase. Both species also contain an ortholog of meso-diaminopimelate dehydrogenase (Ddh), suggesting that they may have redundant pathways for DAP biosynthesis. The B. fragilis Ddh ortholog shows low homology with other examples of Ddh and this species belongs to a phylum, the Bacteriodetes, not previously known to contain this enzyme. By contrast, the C. thermocellum ortholog is well conserved with known examples of Ddh. Using in vitro and in vivo assays both the B. fragilis and C. thermocellum enzymes were found to be authentic examples of Ddh, displaying kinetic properties typical of this enzyme. The result indicates that B. fragilis contains a sequence diverged form of Ddh. Phylogenomic analysis of the microbial genome database revealed that 77% of species with a Ddh ortholog also contain a second pathway for DAP biosynthesis suggesting that Ddh evolved as an ancillary mechanism for DAP biosynthesis.