Sergey Gavrilov

Characterization of Melioribacter roseus gen. nov., sp. nov., a novel facultatively anaerobic thermophilic cellulolytic bacterium from the class Ignavibacteria, and a proposal of a novel bacterial phylum Ignavibacteriae

A novel moderately thermophilic, facultatively anaerobic chemoorganotrophic bacterium strain P3M-2(T) was isolated from a microbial mat developing on the wooden surface of a chute under the flow of hot water (46°C) coming out of a 2775-m-deep oil exploration well (Tomsk region, Russia). Strain P3M-2(T) is a moderate thermophile and facultative anaerobe growing on mono-, di- or polysaccharides by aerobic respiration, fermentation or by reducing diverse electron acceptors [nitrite, Fe(III), As(V)]. Its closest cultivated relative (90.8% rRNA gene sequence identity) is Ignavibacterium album, the only chemoorganotrophic member of the phylum Chlorobi. New genus and species Melioribacter roseus are proposed for isolate P3M-2(T) . Together with I. album, the new organism represents the class Ignavibacteria assigned to the phylum Chlorobi. The revealed group includes a variety of uncultured environmental clones, the 16S rRNA gene sequences of some of which have been previously attributed to the candidate division ZB1. Phylogenetic analysis of M. roseus and I. album based on their 23S rRNA and RecA sequences confirmed that these two organisms could represent an even deeper, phylum-level lineage. Hence, we propose a new phylum Ignavibacteriae within the Bacteroidetes-Chlorobi group with a sole class Ignavibacteria, two families Ignavibacteriaceae and Melioribacteraceae and two species I. album and M. roseus. This proposal correlates with chemotaxonomic data and phenotypic differences of both organisms from other cultured representatives of Chlorobi. The most essential differences, supported by the analyses of complete genomes of both organisms, are motility, facultatively anaerobic and obligately organotrophic mode of life, the absence of chlorosomes and the apparent inability to grow phototrophically.

Library-less synthesis for static CMOS combinational logic circuits

Traditional synthesis techniques optimize CMOS circuits in two phases: i) logic minimization and ii) library mapping phase. Typically, the structures and the sizes of the gates in the library are chosen to yield good synthesis results over many blocks or even for an entire chip. Consequently this approach precludes an optimal design of individual blocks which may need custom structures. The authors present a new transistor level technique that optimizes CMOS circuits both structurally and size-wise. The technique is independent of a library and hence can explore a design space much larger than that possible due to gate level optimization. Results demonstrate a significant improvement in circuit performance of the resynthesized circuits.

Comparative Metagenomics of Eight Geographically Remote Terrestrial Hot Springs

Hot springs are natural habitats for thermophilic Archaea and Bacteria. In this paper, we present the metagenomic analysis of eight globally distributed terrestrial hot springs from China, Iceland, Italy, Russia, and the USA with a temperature range between 61 and 92 ∘C and pH between 1.8 and 7. A comparison of the biodiversity and community composition generally showed a decrease in biodiversity with increasing temperature and decreasing pH. Another important factor shaping microbial diversity of the studied sites was the abundance of organic substrates. Several species of the Crenarchaeal order Thermoprotei were detected, whereas no single bacterial species was found in all samples, suggesting a better adaptation of certain archaeal species to different thermophilic environments. Two hot springs show high abundance of Acidithiobacillus, supporting the idea of a true thermophilic Acidithiobacillus species that can thrive in hyperthermophilic environments. Depending on the sample, up to 58 % of sequencing reads could not be assigned to a known phylum, reinforcing the fact that a large number of microorganisms in nature, including those thriving in hot environments remain to be isolated and characterized.

Genomic Analysis of Melioribacter roseus, Facultatively Anaerobic Organotrophic Bacterium Representing a Novel Deep Lineage within Bacteriodetes/Chlorobi Group

Melioribacter roseus is a moderately thermophilic facultatively anaerobic organotrophic bacterium representing a novel deep branch within Bacteriodetes/Chlorobi group. To better understand the metabolic capabilities and possible ecological functions of M. roseus and get insights into the evolutionary history of this bacterial lineage, we sequenced the genome of the type strain P3M-2T. A total of 2838 open reading frames was predicted from its 3.30 Mb genome. The whole proteome analysis supported phylum-level classification of M. roseus since most of the predicted proteins had closest matches in Bacteriodetes, Proteobacteria, Chlorobi, Firmicutes and deeply-branching bacterium Caldithrix abyssi, rather than in one particular phylum. Consistent with the ability of the bacterium to grow on complex carbohydrates, the genome analysis revealed more than one hundred glycoside hydrolases, glycoside transferases, polysaccharide lyases and carbohydrate esterases. The reconstructed central metabolism revealed pathways enabling the fermentation of complex organic substrates, as well as their complete oxidation through aerobic and anaerobic respiration. Genes encoding the photosynthetic and nitrogen-fixation machinery of green sulfur bacteria, as well as key enzymes of autotrophic carbon fixation pathways, were not identified. The M. roseus genome supports its affiliation to a novel phylum Ignavibateriae, representing the first step on the evolutionary pathway from heterotrophic ancestors of Bacteriodetes/Chlorobi group towards anaerobic photoautotrophic Chlorobi.

Fe(III) Oxide Reduction by a Gram-positive Thermophile: Physiological Mechanisms for Dissimilatory Reduction of Poorly Crystalline Fe(III) Oxide by a Thermophilic Gram-positive Bacterium Carboxydothermus ferrireducens

Physiological strategies driving the reduction of poorly crystalline Fe(III) oxide by the thermophilic Gram-positive dissimilatory Fe(III)-reducing bacterium C. ferrireducens were evaluated. Direct cell-to-mineral contact appears to be the major physiological strategy for ferrihydrite reduction. This strategy is promoted by cell surface-associated c-type cytochromes, and the extracellular electron transfer to ferrihydrite is linked to energy generation via a membrane-bound electron transport chain. The involvement of pili-like appendages in ferrihydrite reduction has been detected for the first time in a thermophilic microorganism. A supplementary strategy for the utilization of a siderophore (DFO) in dissimilatory ferrihydrite reduction has also been characterized.

The Geoglobus acetivorans Genome: Fe(III) Reduction, Acetate Utilization, Autotrophic Growth, and Degradation of Aromatic Compounds in a Hyperthermophilic Archaeon

ABSTRACT Geoglobus acetivorans is a hyperthermophilic anaerobic euryarchaeon of the order Archaeoglobales isolated from deep-sea hydrothermal vents. A unique physiological feature of the members of the genus Geoglobus is their obligate dependence on Fe(III) reduction, which plays an important role in the geochemistry of hydrothermal systems. The features of this organism and its complete 1,860,815-bp genome sequence are described in this report. Genome analysis revealed pathways enabling oxidation of molecular hydrogen, proteinaceous substrates, fatty acids, aromatic compounds, n-alkanes, and organic acids, including acetate, through anaerobic respiration linked to Fe(III) reduction. Consistent with the inability of G. acetivorans to grow on carbohydrates, the modified Embden-Meyerhof pathway encoded by the genome is incomplete. Autotrophic CO2 fixation is enabled by the Wood-Ljungdahl pathway. Reduction of insoluble poorly crystalline Fe(III) oxide depends on the transfer of electrons from the quinone pool to multiheme c-type cytochromes exposed on the cell surface. Direct contact of the cells and Fe(III) oxide particles could be facilitated by pilus-like appendages. Genome analysis indicated the presence of metabolic pathways for anaerobic degradation of aromatic compounds and n-alkanes, although an ability of G. acetivorans to grow on these substrates was not observed in laboratory experiments. Overall, our results suggest that Geoglobus species could play an important role in microbial communities of deep-sea hydrothermal vents as lithoautotrophic producers. An additional role as decomposers would close the biogeochemical cycle of carbon through complete mineralization of various organic compounds via Fe(III) respiration.

Characterization of Membrane-Bound Fe(III)-EDTA Reductase Activities of the Thermophilic Gram-Positive Dissimilatory Iron- Reducing Bacterium Thermoterrabacterium ferrireducens*

Whole-cell suspensions of T. ferrireducens reduced Fe(III) citrate, Fe(III)-EDTA, and ferrihydrite with glycerol as an electron donor. After cell disruption, the highest activity was registered with Fe(III)-EDTA as the electron acceptor and NADH or NADPH as electron donors. About 80% of the NAD(P)H-dependent Fe(III)-EDTA reductase activities were associated with the membrane fraction of the cells. Treatment of the membranes with lauryl maltoside led to complete solubilization of the NADH-dependent and 70% solubilization of the NADPH-dependent Fe(III)-EDTA reductase activities. After purification by ion-exchange chromatography, the NADH-dependent activity was concentrated 8-fold, and the NADPH-dependent activity was concentrated 11-fold, with a yield of about 10% for both activities. The Fe(III)-EDTA-reducing enzyme complex included c-type cytochromes and a protein with a molecular mass of ca. 115 kDa, consisting of two polypeptides. This is the first description of membrane-bound Fe(III)-reducing oxidoreductase activities from a grampositive dissimilatory Fe(III)-reducing bacterium.

False-noise analysis using resolution method

High-performance digital circuits are facing increasingly severe noise problems due to cross-coupled noise injection. Traditionally, noise analysis tools use the conservative assumption that all neighbors of a net can switch simultaneously, thereby producing the worst-case noise on a net. However, due to the logic correlations in the circuit, this worst-case noise may not be realizable, resulting in a so-called false noise failure. Since the problem has been shown to be NP-hard in general, exact solutions to this problem are not possible. In this paper, we therefore propose a new heuristic to eliminate false noise failures based on the resolution method. It is shown that multi-variable logic relations can be computed directly from a transistor level description. Based on these generated logic relations, a characteristic ROBDD for a signal net and its neighboring nets is constructed. This ROBDD is then used to determine the set of neighboring nets that result in the maximum realizable noise on the net. The proposed approach was implemented and tested on industrial circuits. The results demonstrate the effectiveness of the approach to eliminate false noise failures.

Delay noise pessimism reduction by logic correlations

High-performance digital circuits are facing increasingly severe signal integrity problems due to crosstalk noise and therefore the state-of-the-art static timing analysis (STA) methods consider crosstalk-induced delay variation. Current noise-aware STA methods compute noise-induced delay uncertainty for each net independently and annotate appropriate delay changes of nets onto data paths and associated clock paths to determine timing violations. Since delay changes in individual nets contribute cumulatively to delay changes of paths, even small amounts of pessimism in noise computation of nets can add up to produce large timing violations for paths, which may be unrealistic. Unlike glitch noise analysis where noise often attenuates during propagation, quality of delay noise analysis is severely affected by any pessimism in noise estimation and can unnecessarily cost valuable silicon and design resources for fixing unreal violations. In this paper, we propose a method to reduce pessimism in noise-aware STA by considering signal correlations of all nets associated with an entire timing path simultaneously, in a path-based approach. We first present an exact algorithm based on the branch-and-bound technique and then extend it with several heuristic techniques so that very large industrial designs can be analyzed efficiently. These techniques, which are implemented in an industrial crosstalk noise analysis tool, show as much as 75% reduction in the computed path delay variations.

Physiology of Organotrophic and Lithotrophic Growth of the Thermophilic Iron-Reducing Bacteria Thermoterrabacterium ferrireducens and Thermoanaerobacter siderophilus

Growth physiology of the iron-reducing bacteria Thermoterrabacterium ferrireducens and Thermoanaerobacter siderophilus was investigated. The stimulation of the organotrophic growth of T. ferrireducens and T. siderophilusin the presence of Fe(III) was shown to be due to the utilization of ferric iron as an electron acceptor in catabolic processes and not to the effect exerted on the metabolism by Fe(II) or by changes in the redox potential. It was established that Fe(III) reduction in T. ferrireducens is not a detoxication strategy. In T. siderophilus, this process is carried out to alleviate the inhibitory effect of hydrogen. T. ferrireducens was shown to be capable of lithoautotrophic growth with molecular hydrogen as an electron donor and amorphous ferric oxide as an electron acceptor, in the absence of any organic substances. The minimum threshold of H2 consumption was 3 × 10–5 vol % of H2. The presence of CO dehydrogenase activity in T. ferrireducens suggests that CO2 fixation in this organism involves the anaerobic acetyl-CoA pathway. T. siderophilus failed to grow under lithoautotrophic conditions. The fact that T. ferrireducens contains c-type cytochromes and T. siderophilus lacks them confirms the operation of different mechanisms of ferric iron reduction in these species.