Impact of land‐use change to Jatropha bioenergy plantations on biomass and soil carbon stocks: a field study in Mali

Small‐scale Jatropha cultivation and biodiesel production have the potential of contributing to local development, energy security, and greenhouse gas (GHG) mitigation. In recent years however, the GHG mitigation potential of biofuel crops is heavily disputed due to the occurrence of a carbon debt, caused by CO₂ emissions from biomass and soil after land‐use change (LUC). Most published carbon footprint studies of Jatropha report modeled results based on a very limited database. In particular, little empirical data exist on the effects of Jatropha on biomass and soil C stocks. In this study, we used field data to quantify these C pools in three land uses in Mali, that is, Jatropha plantations, annual cropland, and fallow land, to estimate both the Jatropha C debt and its C sequestration potential. Four‐year‐old Jatropha plantations hold on average 2.3 Mg C ha^?1 in their above‐ and belowground woody biomass, which is considerably lower compared to results from other regions. This can be explained by the adverse growing conditions and poor local management. No significant soil organic carbon (SOC) sequestration could be demonstrated after 4 years of cultivation. While the conversion of cropland to Jatropha does not entail significant C losses, the replacement of fallow land results in an average C debt of 34.7 Mg C ha^?1, mainly caused by biomass removal (73%). Retaining native savannah woodland trees on the field during LUC and improved crop management focusing on SOC conservation can play an important role in reducing Jatropha's C debt. Although planting Jatropha on degraded, carbon‐poor cropland results in a limited C debt, the low biomass production, and seed yield attained on these lands reduce Jatropha's potential to sequester C and replace fossil fuels. Therefore, future research should mainly focus on increasing Jatropha's crop productivity in these degraded lands.     

Discrimination of bacteriophage infected cells using locked nucleic acid fluorescent in situ hybridization (LNA-FISH)

Bacteriophage–host interaction studies in biofilm structures are still challenging due to the technical limitations of traditional methods. The aim of this study was to provide a direct fluorescence in situ hybridization (FISH) method based on locked nucleic acid (LNA) probes, which targets the phage replication phase, allowing the study of population dynamics during infection. Bacteriophages specific for two biofilm-forming bacteria, Pseudomonas aeruginosa and Acinetobacter, were selected. Four LNA probes were designed and optimized for phage-specific detection and for bacterial counterstaining. To validate the method, LNA-FISH counts were compared with the traditional plaque forming unit (PFU) technique. To visualize the progression of phage infection within a biofilm, colony-biofilms were formed and infected with bacteriophages. A good correlation (r = 0.707) was observed between LNA-FISH and PFU techniques. In biofilm structures, LNA-FISH provided a good discrimination of the infected cells and also allowed the assessment of the spatial distribution of infected and non-infected populations.     

Have the cake and eat it: Optimizing nondestructive DNA metabarcoding of macroinvertebrate samples for freshwater biomonitoring

DNA metabarcoding can contribute to improving cost‐effectiveness and accuracy of biological assessments of aquatic ecosystems, but significant optimization and standardization efforts are still required to mainstream its application into biomonitoring programmes. In assessments based on freshwater macroinvertebrates, a key challenge is that DNA is often extracted from cleaned, sorted and homogenized bulk samples, which is time‐consuming and may be incompatible with sample preservation requirements of regulatory agencies. Here, we optimize and evaluate metabarcoding procedures based on DNA recovered from 96% ethanol used to preserve field samples and thus including potential PCR inhibitors and nontarget organisms. We sampled macroinvertebrates at five sites and subsampled the preservative ethanol at 1 to 14 days thereafter. DNA was extracted using column‐based enzymatic (TISSUE) or mechanic (SOIL) protocols, or with a new magnetic‐based enzymatic protocol (BEAD), and a 313‐bp COI fragment was amplified. Metabarcoding detected at least 200 macroinvertebrate taxa, including most taxa detected through morphology and for which there was a reference barcode. Better results were obtained with BEAD than SOIL or TISSUE, and with subsamples taken 7–14 than 1–7 days after sampling, in terms of DNA concentration and integrity, taxa diversity and matching between metabarcoding and morphology. Most variation in community composition was explained by differences among sites, with small but significant contributions of subsampling day and extraction method, and negligible contributions of extraction and PCR replication. Our methods enhance reliability of preservative ethanol as a potential source of DNA for macroinvertebrate metabarcoding, with a strong potential application in freshwater biomonitoring.     

Analysis of a new flavodiiron core structural arrangement in Flv1-ΔFlR protein from Synechocystis sp. PCC6803

Flavodiiron proteins (FDPs) play key roles in biological response mechanisms against oxygen and/or nitric oxide; in particular they are present in oxygenic phototrophs (including cyanobacteria and gymnosperms). Two conserved domains define the core of this family of proteins: a N-terminal metallo-β-lactamase-like domain followed by a C-terminal flavodoxin-like one, containing the catalytic diiron centre and a FMN cofactor, respectively. Members of the FDP family may present extra modules in the C-terminus, and were classified into several classes according to their distribution and composition. The cyanobacterium Synechocystis sp. PCC6803 contains four Class C FDPs (Flv1-4) that include at the C-terminus an additional NAD(P)H:flavin oxidoreductase (FlR) domain. Two of them (Flv3 and Flv4) have the canonical diiron ligands (Class C, Type 1), while the other two (Flv1 and Flv2) present different residues in that region (Class C, Type 2). Most phototrophs, either Bacterial or Eukaryal, contain at least two FDP genes, each encoding for one of those two types. Crystals of the Flv1 two core domains (Flv1-ΔFlR), without the C-terminal NAD(P)H:flavin oxidoreductase extension, were obtained and the structure was determined. Its pseudo diiron site contains non-canonical basic and neutral residues, and showed anion moieties, instead. The presented structure revealed for the first time the structure of the two-domain core of a Class C-Type 2 FDP.     

Extracellular lipid droplets promote hemozoin crystallization in the gut of the blood fluke Schistosoma mansoni

Hemozoin (Hz) is a heme crystal produced upon hemoglobin digestion as the main mechanism of heme disposal in several hematophagous organisms. Here, we show that, in the helminth Schistosoma mansoni, Hz formation occurs in extracellular lipid droplets (LDs). Transmission electron microscopy of adult worms revealed the presence of numerous electron-lucent round structures similar to LDs in gut lumen, where multicrystalline Hz assemblies were found associated to their surfaces. Female regurgitates promoted Hz formation in vitro in reactions partially inhibited by boiling. Fractionation of regurgitates showed that Hz crystallization activity was essentially concentrated on lower density fractions, which have small amounts of pre-formed Hz crystals, suggesting that hydrophilic-hydrophobic interfaces, and not Hz itself, play a key catalytic role in Hz formation in S. mansoni. Thus, these data demonstrate that LDs present in the gut lumen of S. mansoni support Hz formation possibly by allowing association of heme to the lipid-water interface of these structures.     

Morphogenesis control in Candida albicans and Candida dubliniensis through signaling molecules produced by planktonic and biofilm cells

Morphogenesis control by chemical signaling molecules is beginning to be highlighted in Candida biology. The present study focuses on morphogenic compounds produced in situ by Candida albicans and Candida dubliniensis during planktonic and biofilm growth that may at least partially substantiate the effect promoted by supernatants in morphogenesis. For both species, planktonic versus biofilm supernatants were analyzed by headspace-solid-phase microextraction and gas chromatography-mass spectrometry. Both planktonic cells and biofilm supernatants of C. albicans and C. dubliniensis contained isoamyl alcohol, 2-phenylethanol, 1-dodecanol, E-nerolidol, and E,E-farnesol. Alcohol secretion profiles were species, culture mode, and growth time specific. The addition of exogenous alcohols to the cultures of both species inhibited the morphological transition from the yeast to the filamentous form by up to 50%. The physiological role of these alcohols was put to evidence by comparing the effects of a 96-h cultured supernatant with synthetic mixtures containing isoamyl alcohol, 2-phenylethanol, E-nerolidol, and E,E-farnesol at concentrations determined herein. All synthetic mixtures elicited a morphological effect similar to that observed for the corresponding supernatants when used to treat C. albicans and C. dubliniensis cultures, except for the effect of the 96-h C. dubliniensis planktonic supernatant culture on C. albicans. Overall, these results reveal a group of alcohol extracellular signaling molecules that are biologically active with C. albicans and C. dubliniensis morphogenesis.     

A new marine prasinophyte genus alternates between a flagellate and a dominant benthic stage with microrhizoids for adhesion

Prasinophytes (Chlorophyta) are a diverse, paraphyletic group of planktonic microalgae for which benthic species are largely unknown. Here, we report a sand‐dwelling, marine prasinophyte with several novel features observed in clonal cultures established from numerous locations around Australia. The new genus and species, which we name Microrhizoidea pickettheapsiorum (Mamiellophyceae), alternates between a benthic palmelloid colony, where cell division occurs, and a planktonic flagellate. Flagellates are short lived, settle and quickly resorb their flagella, the basal bodies then nucleate novel tubular appendages, termed “microrhizoids”, that lack an axoneme and function to anchor benthic cells to the substratum. To our knowledge, microrhizoids have not been observed in any other green alga or protist, are slightly smaller in diameter than flagella, generally contain nine microtubules, are long (3–5 times the length of flagella) and are not encased in scales. Following settlement, cell divisions result in a loose, palmelloid colony, each cell connected to the substratum by two microrhizoids. Flagellates are round to bean‐shaped with two long, slightly uneven flagella. Both benthic cells and flagellates, along with their flagella, are encased in thin scales. Phylogenies based on the complete chloroplast genome of Microrhizoidea show that it is clearly a member of the Mamiellophyceae, most closely related to Dolichomastix tenuilepsis. More taxon‐rich phylogenetic analyses of the 18S rRNA gene, including metabarcodes from the Tara Oceans and Ocean Sampling Day projects, confidently show the distinctive nature of Microrhizoidea, and that the described biodiversity of the Mamiellophyceae is a fraction of its real biodiversity. The discovery of a largely benthic prasinophyte changes our perspective on this group of algae and, along with the observation of other potential benthic lineages in environmental sequences, illustrates that benthic habitats can be a rich ground for algal biodiscovery.     

Modulation of the central carbon metabolism of Corynebacterium glutamicum improves malonyl-CoA availability and increases plant polyphenol synthesis

In recent years microorganisms have been engineered towards synthesizing interesting plant polyphenols such as flavonoids and stilbenes from glucose. Currently, the low endogenous supply of malonyl-CoA, indispensable for plant polyphenol synthesis, impedes high product titers. Usually, limited malonyl-CoA availability during plant polyphenol production is avoided by supplementing fatty acid synthesis-inhibiting antibiotics such as cerulenin, which are known to increase the intracellular malonyl-CoA pool as a side effect. Motivated by the goal of microbial polyphenol synthesis being independent of such expensive additives, we used rational metabolic engineering approaches to modulate regulation of fatty acid synthesis and flux into the tricarboxylic acid cycle (TCA cycle) in Corynebacterium glutamicum strains capable of flavonoid and stilbene synthesis. Initial experiments showed that sole overexpression of genes coding for the native malonyl-CoA-forming acetyl-CoA carboxylase is not sufficient for increasing polyphenol production in C. glutamicum. Hence, the intracellular acetyl-CoA availability was also increased by reducing the flux into the TCA cycle through reduction of citrate synthase activity. In defined cultivation medium, the constructed C. glutamicum strains accumulated 24 mg·L ⁻¹ (0.088 mM) naringenin or 112 mg·L ⁻¹ (0.49 mM) resveratrol from glucose without supplementation of phenylpropanoid precursor molecules or any inhibitors of fatty acid synthesis.