Refining and illuminating acetogenic Eubacterium strains for reclassification and metabolic engineering.

BACKGROUND: The genus Eubacterium is quite diverse and includes several acetogenic strains capable of fermenting C1-substrates into valuable products. Especially, Eubacterium limosum and closely related strains attract attention not only for their capability to ferment C1 gases and liquids, but also due to their ability to produce butyrate. Apart from its well-elucidated metabolism, E. limosum is also genetically accessible, which makes it an interesting candidate to be an industrial biocatalyst. RESULTS: In this study, we examined genomic, phylogenetic, and physiologic features of E. limosum and the closest related species E. callanderi as well as E. maltosivorans. We sequenced the genomes of the six Eubacterium strains 'FD' (DSM 3662T), 'Marburg' (DSM 3468), '2A' (DSM 2593), '11A' (DSM 2594), 'G14' (DSM 107592), and '32' (DSM 20517) and subsequently compared these with previously available genomes of the E. limosum type strain (DSM 20543T) as well as the strains 'B2', 'KIST612', 'YI' (DSM 105863T), and 'SA11'. This comparison revealed a close relationship between all eleven Eubacterium strains, forming three distinct clades: E. limosum, E. callanderi, and E. maltosivorans. Moreover, we identified the gene clusters responsible for methanol utilization as well as genes mediating chain elongation in all analyzed strains. Subsequent growth experiments revealed that strains of all three clades can convert methanol and produce acetate, butyrate, and hexanoate via reverse ß-oxidation. Additionally, we used a harmonized electroporation protocol and successfully transformed eight of these Eubacterium strains to enable recombinant plasmid-based expression of the gene encoding the fluorescence-activating and absorption shifting tag (FAST). Engineered Eubacterium strains were verified regarding their FAST-mediated fluorescence at a single-cell level using a flow cytometry approach. Eventually, strains 'FD' (DSM 3662T), '2A' (DSM 2593), '11A' (DSM 2594), and '32' (DSM 20517) were genetically engineered for the first time. CONCLUSION: Strains of E. limosum, E. callanderi, and E. maltosivorans are outstanding candidates as biocatalysts for anaerobic C1-substrate conversion into valuable biocommodities. A large variety of strains is genetically accessible using a harmonized electroporation protocol, and FAST can serve as a reliable fluorescent reporter protein to characterize genetically engineered cells. In total eleven strains have been assigned to distinct clades, providing a clear and updated classification. Thus, the description of respective Eubacterium species has been emended, improved, aligned, and is requested to be implemented in respective databases.

Spurious Symmetry Enhancement in Linear Spin Wave Theory and Interaction-Induced Topology in Magnons.

Linear spin wave theory (LSWT) is the standard technique to compute the spectra of magnetic excitations in quantum materials. In this Letter, we show that LSWT, even under ordinary circumstances, may fail to implement the symmetries of the underlying ordered magnetic Hamiltonian leading to spurious degeneracies. In common with pseudo-Goldstone modes in cases of quantum order by disorder these degeneracies tend to be lifted by magnon-magnon interactions. We show how, instead, the correct symmetries may be restored at the level of LSWT. In the process we give examples, supported by nonperturbative matrix product based time evolution calculations, where symmetry dictates topological features but where LSWT fails to implement them. We also comment on possible spin split magnons in MnF_{2} and similar rutiles by analogy to recently proposed altermagnets.

Zero-Frequency Chiral Magnonic Edge States Protected by Nonequilibrium Topology.

Topological bosonic excitations must, in contrast to their fermionic counterparts, appear at finite energies. This is a key challenge for magnons, as it prevents straightforward excitation and detection of topologically protected magnonic edge states and their use in magnonic devices. In this Letter, we show that in a nonequilibrium state, in which the magnetization is pointing against the external magnetic field, the topologically protected chiral edge states in a magnon Chern insulator can be lowered to zero frequency, making them directly accessible by existing experimental techniques. We discuss the spin-orbit torque required to stabilize this nonequilibrium state, and show explicitly using numerical Landau-Lifshitz-Gilbert simulations that the edge states can be excited with a microwave field. Finally, we consider a propagating spin wave spectroscopy experiment, and demonstrate that the edge states can be directly detected.

Lactate based caproate production with Clostridium drakei and process control of Acetobacterium woodii via lactate dependent in situ electrolysis.

Syngas fermentation processes with acetogens represent a promising process for the reduction of CO2 emissions alongside bulk chemical production. However, to fully realize this potential the thermodynamic limits of acetogens need to be considered when designing a fermentation process. An adjustable supply of H2 as electron donor plays a key role in autotrophic product formation. In this study an anaerobic laboratory scale continuously stirred tank reactor was equipped with an All-in-One electrode allowing for in-situ H2 generation via electrolysis. Furthermore, this system was coupled to online lactate measurements to control the co-culture of a recombinant lactate-producing Acetobacterium woodii strain and a lactate-consuming Clostridium drakei strain to produce caproate. When C. drakei was grown in batch cultivations with lactate as substrate, 1.6 g·L-1 caproate were produced. Furthermore, lactate production of the A. woodii mutant strain could manually be stopped and reinitiated by controlling the electrolysis. Applying this automated process control, lactate production of the A. woodii mutant strain could be halted to achieve a steady lactate concentration. In a co-culture experiment with the A. woodii mutant strain and the C. drakei strain, the automated process control was able to dynamically react to changing lactate concentrations and adjust H2 formation respectively. This study confirms the potential of C. drakei as medium chain fatty acid producer in a lactate-mediated, autotrophic co-cultivation with an engineered A. woodii strain. Moreover, the monitoring and control strategy presented in this study reinforces the case for autotrophically produced lactate as a transfer metabolite in defined co-cultivations for value-added chemical production.

Novel synthetic co-culture of Acetobacterium woodii and Clostridium drakei using CO2 and in situ generated H2 for the production of caproic acid via lactic acid.

Acetobacterium woodii is known to produce mainly acetate from CO2 and H2, but the production of higher value chemicals is desired for the bioeconomy. Using chain-elongating bacteria, synthetic co-cultures have the potential to produce longer-chained products such as caproic acid. In this study, we present first results for a successful autotrophic co-cultivation of A. woodii mutants and a Clostridium drakei wild-type strain in a stirred-tank bioreactor for the production of caproic acid from CO2 and H2 via the intermediate lactic acid. For autotrophic lactate production, a recombinant A. woodii strain with a deleted Lct-dehydrogenase complex, which is encoded by the lctBCD genes, and an inserted D-lactate dehydrogenase (LdhD) originating from Leuconostoc mesenteroides, was used. Hydrogen for the process was supplied using an All-in-One electrode for in situ water electrolysis. Lactate concentrations as high as 0.5 g L-1 were achieved with the AiO-electrode, whereas 8.1 g L-1 lactate were produced with direct H2 sparging in a stirred-tank bioreactor. Hydrogen limitation was identified in the AiO process. However, with cathode surface area enlargement or numbering-up of the electrode and on-demand hydrogen generation, this process has great potential for a true carbon-negative production of value chemicals from CO2.

Thermal Hall Effect of Magnons in Collinear Antiferromagnetic Insulators: Signatures of Magnetic and Topological Phase Transitions.

We demonstrate theoretically that the thermal Hall effect of magnons in collinear antiferromagnetic insulators is an indicator of magnetic and topological phase transitions in the magnon spectrum. The transversal heat current of magnons caused by a thermal gradient is calculated for an antiferromagnet on a honeycomb lattice. An applied magnetic field drives the system from the antiferromagnetic phase via a spin-flop phase into the field-polarized phase. In addition to these magnetic phase transitions, we find topological phase transitions within the spin-flop phase. Both types of transitions manifest themselves in prominent and distinguishing features in the thermal conductivity, which changes by several orders of magnitude. The variation of temperature provides a tool to discern experimentally the two types of phase transitions. We include numerical results for the van der Waals magnet MnPS_{3}.

Autotrophic lactate production from H2 + CO2 using recombinant and fluorescent FAST-tagged Acetobacterium woodii strains.

Lactate has various uses as industrial platform chemical, poly-lactic acid precursor or feedstock for anaerobic co-cultivations. The aim of this study was to construct and characterise Acetobacterium woodii strains capable of autotrophic lactate production. Therefore, the lctBCD genes, encoding the native Lct dehydrogenase complex, responsible for lactate consumption, were knocked out. Subsequently, a gene encoding a D-lactate dehydrogenase (LDHD) originating from Leuconostoc mesenteroides was expressed in A. woodii, either under the control of the anhydrotetracycline-inducible promoter Ptet or under the lactose-inducible promoter PbgaL. Moreover, LDHD was N-terminally fused to the oxygen-independent fluorescence-activating and absorption-shifting tag (FAST) and expressed in respective A. woodii strains. Cells that produced the LDHD fusion protein were capable of lactate production of up to 18.8 mM in autotrophic batch experiments using H2 + CO2 as energy and carbon source. Furthermore, cells showed a clear and bright fluorescence during exponential growth, as well as in the stationary phase after induction, mediated by the N-terminal FAST. Flow cytometry at the single-cell level revealed phenotypic heterogeneities for cells expressing the FAST-tagged LDHD fusion protein. This study shows that FAST provides a new reporter tool to quickly analyze gene expression over the course of growth experiments of A. woodii. Consequently, fluorescence-based reporters allow for faster and more targeted optimization of production strains.Key points •Autotrophic lactate production was achieved with A. woodii. •FAST functions as fluorescent marker protein in A. woodii. •Fluorescence measurements on single-cell level revealed population heterogeneity.

Orbital Magnetic Moment of Magnons.

In experiments and their interpretation usually the spin magnetic moment of magnons is considered. In this Letter, we identify a complementing orbital magnetic moment of magnons brought about by spin-orbit coupling. Our microscopic theory uncovers that spin magnetization M^{S} and orbital magnetization M^{O} are independent quantities; they are not necessarily collinear. Even when the total spin moment is compensated due to antiferromagnetism, M^{O} may be nonzero. This scenario of orbital weak ferromagnetism is realized in paradigmatic kagome antiferromagnets with Dzyaloshinskii-Moriya interaction. We demonstrate that magnets exhibiting a magnonic orbital moment are omnipresent and propose transport experiments for probing it.

Tunable Magnon Weyl Points in Ferromagnetic Pyrochlores.

The dispersion relations of magnons in ferromagnetic pyrochlores with Dzyaloshinskii-Moriya interaction are shown to possess Weyl points, i. e., pairs of topologically nontrivial crossings of two magnon branches with opposite topological charge. As a consequence of their topological nature, their projections onto a surface are connected by magnon arcs, thereby resembling closely Fermi arcs of electronic Weyl semimetals. On top of this, the positions of the Weyl points in reciprocal space can be tuned widely by an external magnetic field: rotated within the surface plane, the Weyl points and magnon arcs are rotated as well; tilting the magnetic field out of plane shifts the Weyl points toward the center Γ[over ¯] of the surface Brillouin zone. The theory is valid for the class of ferromagnetic pyrochlores, i. e., three-dimensional extensions of topological magnon insulators on kagome lattices. In this Letter, we focus on the (111) surface, identify candidates of established ferromagnetic pyrochlores which apply to the considered spin model, and suggest experiments for the detection of the topological features.