Diatoms synthesize sterols by inclusion of animal and fungal genes in the plant pathway

Diatoms are ubiquitous microalgae that have developed remarkable metabolic plasticity and gene diversification. Here we report the first elucidation of the complete biosynthesis of sterols in the lineage. The study has been carried out on the bloom-forming species Skeletonema marinoi and Cyclotella cryptica that synthesise an ensemble of sterols with chemotypes of animals (cholesterol and desmosterol), plants (dihydrobrassicasterol and 24-methylene cholesterol), algae (fucosterol) and marine invertebrates (clionasterol). In both species, sterols derive from mevalonate through cyclization of squalene to cycloartenol by cycloartenol synthase. The pathway anticipates synthesis of cholesterol by enzymes of the phytosterol route in plants, as recently reported in Solanaceae. Major divergences stem from reduction of Δ24(28) and Δ24(25) double bonds which, in diatoms, are apparently dependent on sterol reductases of fungi, algae and animals. Phylogenetic comparison revealed a good level of similarity between the sterol biosynthetic genes of S. marinoi and C. cryptica with those in the genomes of the other diatoms sequenced so far

CO2-Induced Transcriptional Reorganization: Molecular Basis of Capnophillic Lactic Fermentation in Thermotoga neapolitana

Capnophilic lactic fermentation (CLF) is a novel anaplerotic pathway able to convert sugars to lactic acid (LA) and hydrogen using CO2 as carbon enhancer in the hyperthermophilic bacterium Thermotoga neapolitana. In order to give further insights into CLF metabolic networks, we investigated the transcriptional modification induced by CO2 using a RNA-seq approach. Transcriptomic analysis revealed 1601 differentially expressed genes (DEGs) in an enriched CO2 atmosphere over a total of 1938 genes of the T. neapolitana genome. Transcription of PFOR and LDH genes belonging to the CLF pathway was up-regulated by CO2 together with 6-phosphogluconolactonase (6PGL) and 6-phosphogluconate dehydratase (EDD) of the Entner–Doudoroff (ED) pathway. The transcriptomic study also revealed up-regulation of genes coding for the flavin-based enzymes NADH-dependent reduced ferredoxin:NADP oxidoreductase (NFN) and NAD-ferredoxin oxidoreductase (RNF) that control supply of reduced ferredoxin and NADH and allow energy conservation-based sodium translocation through the cell membrane. These results support the hypothesis that CO2 induces rearrangement of the central carbon metabolism together with activation of mechanisms that increase availability of the reducing equivalents that are necessary to sustain CLF. In this view, this study reports a first rationale of the molecular basis of CLF in T. neapolitana and provides a list of target genes for the biotechnological implementation of this process

Probing the therapeutic potential of marine phyla by spe extraction

The marine environment is potentially a prolific source of small molecules with significant biological activities. In recent years, the development of new chromatographic phases and the progress in cell and molecular techniques have facilitated the search for marine natural products (MNPs) as novel pharmacophores and enhanced the success rate in the selection of new potential drug candidates. However, most of this exploration has so far been driven by anticancer research and has been limited to a reduced number of taxonomic groups. In this article, we report a test study on the screening potential of an in-house library of natural small molecules composed of 285 samples derived from 57 marine organisms that were chosen from among the major eukaryotic phyla so far represented in studies on bioactive MNPs. Both the extracts and SPE fractions of these organisms were simultaneously submitted to three different bioassays—two phenotypic and one enzymatic—for cytotoxic, antidiabetic, and antibacterial activity. On the whole, the screening of the MNP library selected 11 potential hits, but the distribution of the biological results showed that SPE fractionation increased the positive score regardless of the taxonomic group. In many cases, activity could be detected only in the enriched fractions after the elimination of the bulky effect due to salts. On a statistical basis, sponges and molluscs were confirmed to be the most significant source of cytotoxic and antimicrobial products, but other phyla were found to be effective with the other therapeutic target

Oxylipin Diversity in the Diatom Family Leptocylindraceae Reveals DHA Derivatives in Marine Diatoms

Marine planktonic organisms, such as diatoms, are prospective sources of novel bioactive metabolites. Oxygenated derivatives of fatty acids, generally referred to as oxylipins, in diatoms comprise a highly diverse and complex family of secondary metabolites. These molecules have recently been implicated in several biological processes including intra- and inter-cellular signaling as well as in defense against biotic stressors and grazers. Here, we analyze the production and diversity of C20 and C22 non-volatile oxylipins in five species of the family Leptocylindraceae, which constitute a basal clade in the diatom phylogeny. We report the presence of species-specific lipoxygenase activity and oxylipin patterns, providing the first demonstration of enzymatic production of docosahexaenoic acid derivatives in marine diatoms. The differences observed in lipoxygenase pathways among the species investigated broadly reflected the relationships observed with phylogenetic markers, thus providing functional support to the taxonomic diversity of the individual species

Capnophilic Lactic Fermentation from <i>Thermotoga neapolitana</i>: A Resourceful Pathway to Obtain Almost Enantiopure L-lactic Acid

The industrial production of lactic acid (LA) is mainly based on bacterial fermentation. This process can result in enantiopure or racemic mixture according to the producing organism. Between the enantiomers, L-lactic acid shows superior market value. Recently, we reported a novel anaplerotic pathway called capnophilic lactic fermentation (CLF) that produces a high concentration of LA by fermentation of sugar in the anaerobic thermophilic bacterium Thermotoga neapolitana. The aim of this work was the identification of the enantiomeric characterization of the LA produced by T. neapolitana and identification of the lactate dehydrogenase in T. neapolitana (TnLDH) and related bacteria of the order Thermotogales. Chemical derivatization and GC/MS analysis were applied to define the stereochemistry of LA from T. neapolitana. A bioinformatics study on TnLDH was carried out for the characterization of the enzyme. Chemical analysis showed a 95.2% enantiomeric excess of L-LA produced by T. neapolitana. A phylogenetic approach clearly clustered the TnLDH together with the L-LDH from lactic acid bacteria. We report for the first time that T. neapolitana is able to produce almost enantiopure L-lactic acid. The result was confirmed by bioinformatics analysis on TnLDH, which is a member of the L-LDH sub-family

Performance of Reduced Titanium Oxide and Boron Doped Diamond as anodes in hyperthermophilic bioelectrochemical systems

This work investigates Reduced Titanium Oxide (RTO) in comparison with Carbon Cloth (CC) and commercial Boron Doped Diamond (BDD) as anodes in hyperthermophilic bioelectrochemical systems operating at 80°C by Thermotoga neapolitana. Two samples of RTO were synthesized by plasma electrolytic oxidation (PEO) of titanium plates and subsequent electrochemical reduction. Electrochemical performance of CC, BDD, and RTO are tested by performing cyclic voltammetry in the anodic region (0-1V, 50 mV/s), in abiotic and biotic conditions. The surface of colonized materials is observed by SEM microscopy. Results show that bacteria fast settle on all tested material, significantly affecting their electrochemical conductivity. The integration of voltammetric cycles reveals that biofilm generates capacitive effects on the anodic surfaces, particularly evident in RTO, less in CC and absent in BDD. Charge densities provided by capacitive response of RTO and CC are of the order of 5.58 and 0.77 mC/cm2, respectively

Effects of agitation speed and gas recirculation on hydrogen supersaturation and dark fermentation by Thermotoga neapolitana

Thermotoga neapolitana is capable to produce bio-hydrogen at high yields in hyperthermophilic dark fermentation from a large variety of substrates including organic wastes. However, in dark fermentation hydrogen concentration is one of the main inhibitors affecting the hydrogen yield and production rate. In this study, the effect of agitation speed and recirculation of the produced biogas on dissolved hydrogen concentration and its impact on hydrogen production by Thermotoga neapolitana were investigated. In 2 L batch bioassays using 1% inoculum (v/v), an increase of the agitation speed from 300 to 500 rpm led to a decrease of the dissolved hydrogen concentration from 17.0 ± 2.3 to 7.4 ± 0.7 mL/L. As a result, the hydrogen production rate and the glucose consumption rate increased from 54 ± 13 to 112 ± 22 mL H2/L/h and from 3.6 ± 0.4 to 4.7 ± 0.3 mmol glucose/L/h, respectively. In a second stage, the biomass concentration was increased to 0.8 g initial cell dry weight /L (equal to 200% inoculum v/v), applying agitation exclusively (low agitation “LA” - 300 rpm; high agitation “HA” - 500 rpm) and agitation combined with gas recirculation “GaR” (“LA+GaR” - 300 rpm; “HA+GaR” - 500 rpm). Gas recirculation increased the hydrogen production rate (ml H2/L/h) from 235 ± 35 (LA) and 535 ± 35 (HA) to 850 ± 71 (LA+GaR) and 813 ± 18 (HA+GaR). Concomitantly, the hydrogen yield (mol H2/ mol glucose) was increased from 3.0 ± 0.0 (LA) and 3.2 ± 0.1 (HA) to 3.5 ± 0.2 (LA+GaR) and 3.3 ± 0.1 (HA+GaR), respectively. At the same time, the fermentation time was reduced from 11 (LA) and 6 h (HA) to 4 h (LA+GaR; HA+GaR). The previous results confirm that the dissolved hydrogen plays a crucial role in dark fermentation hydrogen production even under hyperthermophilic stirred conditions. At higher biomass concentrations, the increase of the agitation speed increased the hydrogen yield and production rate by approximately 9 and 128% (HA/LA). In comparison, the application of the biogas recirculation increased the hydrogen yield and production rate by approximately 17 and 262% (LA+GaR/LA). The increase of agitation speed had a little effect when gas recirculation was applied, as indicated by the similar results achieved under LA+GaR and HA+GaR conditions. The present study confirms that gas recirculation is a simple and effective method to eliminate hydrogen supersaturation, considerably accelerating the hydrogen production and enhancing the yield

The different behaviour of

Thermotoga neapolitana is a hyperthermophilic bacterium that can metabolize glucose and several organic wastes in hydrogen and lactate at a temperature of 80°C. Their high performance in producing hydrogen at so high a temperature as 80°C suggests a potential energy application of them where hydrogen is an important element of the process. In this view, experimentation of a T.neapolitana strain is carried out in double-chamber electrochemical systems. The aim is to explore the interaction of these bacteria with the anode and the cathode, stressing their capability to survive in presence of a polarized electrode which can drastically change the pH of the media. A culture enriched of 5 g/L of glucose, under CO2 pressure (80 °C) was used to fill both the anodic and cathodic compartments of the electrochemical system, applying a voltage of 1.5 V between the anode and the cathode. The test lasted ten days. Results clearly indicate that bacteria colonize both electrodes, but the glucose metabolism is completely inhibited in the anodic compartments. On the contrary, metabolism is stimulated in the cathodic compartment. Bacteria are alive on the electrodes in the pH interval of 3 - 9