Using Co-Culture to Functionalize Clostridium Fermentation.

Clostridium fermentations have been developed for producing butanol and other value-added chemicals, but their development is constrained by some limitations, such as relatively high substrate cost and the need to maintain an anaerobic condition. Recently, co-culture is emerging as a popular way to address these limitations by introducing a partner strain with Clostridium. Generally speaking, the co-culture strategy enables the use of a cheaper substrate, maintains the growth of Clostridium without any anaerobic treatment, improves product yields, and/or widens the product spectrum. Herein, we review recent developments of co-culture strategies involving Clostridium species according to their partner stains' functions with representative examples. We also discuss research challenges that need to be addressed for the future development of Clostridium co-cultures.

Aerobic acetone-butanol-isopropanol (ABI) fermentation through a co-culture of Clostridium beijerinckii G117 and recombinant Bacillus subtilis 1A1.

An engineered B. subtilis 1A1 strain (BsADH2) expressing a secondary alcohol dehydrogenase (CpSADH) was co-cultured with C. beijerinckii G117 under an aerobic condition. During the fermentation on glucose, B. subtilis BsADH2 depleted oxygen in culture media completely and created an anaerobic environment for C. beijerinckii G117, an obligate anaerobe, to grow. Meanwhile, lactate produced by B. subtilis BsADH2 was re-assimilated by C. beijerinckii G117. In return, acetone produced by C. beijerinckii G117 was reduced into isopropanol by B. subtilis BsADH2 via expressing the CpSADH, which helped maintain the redox balance of the engineered B. subtilis. In the symbiotic system consisting of two strains, 1.7 â€‹g/L of acetone, 4.8 â€‹g/L of butanol, and 0.9 â€‹g/L of isopropanol (with an isopropanol/acetone ratio of 0.53) was produced from 60 â€‹g/L of glucose. This symbiotic system also worked when oxygen was supplied to the culture, although less isopropanol was produced (0.9 â€‹g/L of acetone, 4.9 â€‹g/L of butanol, and 0.2 â€‹g/L of isopropanol). The isopropanol titer was increased substantially to 2.5 â€‹g/L when we increased the inoculum size of B. subtilis BsADH2 and optimized other process parameters. With the Bacillus-Clostridium co-culture, switching from the original acetone-butanol (AB) fermentation to an aerobic acetone-butanol-isopropanol (ABI) fermentation can be easily achieved without genetic engineering of Clostridium. This strategy of employing a recombinant Bacillus to co-culture with Clostridium should be potentially useful to modify traditional acetone-butanol-ethanol fermentation for the production of other value-added chemicals.

Production of isopropyl and butyl esters by Clostridium mono-culture and co-culture.

Production of esters from the acetone-butanol-ethanol (ABE) fermentation by Clostridium often focuses on butyl butyrate, leaving acetone as an undesired product. Addition of butyrate is also often needed because ABE fermentation does not produce enough butyrate. Here we addressed the problems using Clostridium beijerinckii BGS1 that preferred to produce isopropanol instead of acetone, and co-culturing it with Clostridium tyrobutyricum ATCC 25,755 that produced butyrate. Unlike acetone, isopropanol could be converted into ester using lipase and acids. C. tyrobutyricum ATCC 25,755 produced acids at pH 6, while C. beijerinckii BGS1 produced mainly solvents at the same pH. When the two strains were co-cultured, more butyrate was produced, leading to a higher titer of esters than the mono-culture of C. beijerinckii BGS1. As the first study reporting the production of isopropyl butyrate from the Clostridium fermentation, this study highlighted the potential use of lipase and co-culture strategy in ester production.

Publisher Correction: Intensity-dependent modulation of optically active signals in a chiral metamaterial.

This corrects the article DOI: 10.1038/ncomms14602.

Intensity-dependent modulation of optically active signals in a chiral metamaterial.

Chiral media exhibit optical phenomena that provide distinctive responses from opposite circular polarizations. The disparity between these responses can be optimized by structurally engineering absorptive materials into chiral nanopatterns to form metamaterials that provide gigantic chiroptical resonances. To fully leverage the innate duality of chiral metamaterials for future optical technologies, it is essential to make such chiroptical responses tunable via external means. Here we report an optical metamaterial with tailored chiroptical effects in the nonlinear regime, which exhibits a pronounced shift in its circular dichroism spectrum under a modest level of excitation power. Strong nonlinear optical rotation is observed at key spectral locations, with an intensity-induced change of 14° in the polarization rotation from a metamaterial thickness of less than λ/7. The modulation of chiroptical responses by manipulation of input powers incident on chiral metamaterials offers potential for active optics such as all-optical switching and light modulation.

Electrically Tunable Harmonic Generation of Light from Plasmonic Structures in Electrolytes.

An emerging trend in plasmonics is to exploit nanostructured metals as a self-contained electrooptic platform with simultaneously supported electrical and optical functions. When it comes to nonlinear optics, this dual electrical and optical functionality offers an exciting potential to enable electrically controlled wave mixing processes in various nanometallic systems. Here we demonstrate tunable nonlinear generation of light enabled by an electrically active plasmonic crystal in aqueous electrolytic solutions. A modulation depth of ∼150%/V is observed in the second-harmonic signal, thanks to the light concentrating ability of the resonant plasmonic structure as well as the voltage-assisted charge accumulation on the metallic surfaces. The hybrid plasmonic-electrolyte system demonstrated in this work offers the exciting new potential to use plasmonic nanostructures for voltage-controlled nonlinear signal generation and in situ biochemical sensing in an aqueous environment.

An Active Metamaterial Platform for Chiral Responsive Optoelectronics.

Chiral-selective non-linear optics and optoelectronic signal generation are demonstrated in an electrically active photonic metamaterial. The metamaterial reveals significant chiroptical responses in both harmonic generation and the photon drag effect, correlated to the resonance behavior in the linear regime. The multifunctional chiral metamaterial with dual electrical and optical functionality enables transduction of chiroptical responses to electrical signals for integrated photonics.

Backward phase-matching for nonlinear optical generation in negative-index materials.

Metamaterials have enabled the realization of unconventional electromagnetic properties not found in nature, which provokes us to rethink the established rules of optics in both the linear and nonlinear regimes. One of the most intriguing phenomena in nonlinear metamaterials is 'backward phase-matching', which describes counter-propagating fundamental and harmonic waves in a negative-index medium. Predicted nearly a decade ago, this process is still awaiting a definitive experimental confirmation at optical frequencies. Here, we report optical measurements showing backward phase-matching by exploiting two distinct modes in a nonlinear plasmonic waveguide, where the real parts of the mode refractive indices are 3.4 and -3.4 for the fundamental and the harmonic waves respectively. The observed peak conversion efficiency at the excitation wavelength of ∼780 nm indicates the fulfilment of the phase-matching condition of k(2ω) = 2k(ω) and n(2ω) = -n(ω), where the coherent harmonic wave emerges along a direction opposite to that of the incoming fundamental light.

Metamaterials enable chiral-selective enhancement of two-photon luminescence from quantum emitters.

The amplification of chirally modified, non-linear signals from quantum emitters is demonstrated by manipulating the geometric chirality of resonant plasmonic nanostructures. The chiral center of the metamaterial is opened and emitters occupy this light-confining and chirally sensitive region. Non-linear emission signals are enhanced by 40× that of the emitters not embedded in the metamaterial and display a 3× contrast for the opposite circular polarization.

Electrifying photonic metamaterials for tunable nonlinear optics.

Metamaterials have not only enabled unprecedented flexibility in producing unconventional optical properties that are not found in nature, they have also provided exciting potential to create customized nonlinear media with high-order properties correlated to linear behaviour. Two particularly compelling directions are active metamaterials, whose optical properties can be purposely tailored by external stimuli in a reversible manner, and nonlinear metamaterials, which enable intensity-dependent frequency conversion of light waves. Here, by exploring the interaction of these two directions, we leverage the electrical and optical functions simultaneously supported in nanostructured metals and demonstrate electrically controlled nonlinear optical processes from a metamaterial. Both second harmonic generation and optical rectification, enhanced by the resonance behaviour in the metamaterial absorber, are modulated externally with applied voltage signals. Our results reveal an opportunity to exploit optical metamaterials as self-contained, dynamic electro-optic systems with intrinsically embedded electrical functions and optical nonlinearities.

Nonlinear imaging and spectroscopy of chiral metamaterials.

A chiral metamaterial produces both distinguishable linear and non-linear resonant features when probed with left and right circularly polarized light. The material demonstrates a linear transmission contrast of 0.5 between left and right circular polarizations and a 20× contrast between second-harmonic responses from the two incident polarizations. Non-linear and linear response images probed with circularly polarized light show strongly defined contrast.

Giant chiral optical response from a twisted-arc metamaterial.

We demonstrate enormously strong chiral effects from a photonic metamaterial consisting of an array of dual-layer twisted-arcs with a total thickness of ∼ λ/6. Experimental results reveal a circular dichroism of ∼ 0.35 in the absolute value and a maximum polarization rotation of ∼ 305°/λ in a near-infrared wavelength region. A transmission of greater than 50% is achieved at the frequency where the polarization rotation peaks. Retrieved parameters from measured quantities further indicate an actual optical activity of 76° per λ and a difference of 0.42 in the indices of refraction for the two circularly polarized waves of opposite handedness.

Scattering reduction at near-infrared frequencies using plasmonic nanostructures.

Novel fabrication, detection and analysis approaches were employed to experimentally demonstrate scattering reduction by a plasmonic nanostructure operating at 1550 nm. The nanostructure consisted of a silicon nanorod surrounded by a plasmonic metamaterial cover comprised of eight gold nanowires and was fabricated by a combination of electron beam lithography, focused ion beam milling and dry and wet etching. The optical standing wave pattern of the device in the near-field was obtained using heterodyne near-field scanning optical microscopy. It was found that the spatial curvature of the interference fringes of the optical standing wave pattern was directly related to the scattering reduction of the device. The experiments were in excellent agreement with the theoretical predictions and suggested that the device reduced the scattering by 9.5 dB when compared to a bare silicon nanorod of diameter 240 nm and by 6 dB when compared to a bare silicon nanorod of diameter 160 nm.

Nanorod orientation dependence of tunable Fano resonance in plasmonic nanorod heptamers.

We present orientation dependence and tuning of Fano resonances in plasmonic gold nanorod heptamer nanostructures. The heptamer structures were formed by surrounding a central circular nanoparticle by six satellite nanorods in a hexagonal arrangement and the heptamer structures were then embedded in a thin flexible membrane. The Fano resonance was found to depend on the orientation of the nanorods within the heptamer and for a given orientation, the Fano resonance was dynamically tuned and its symmetry lowered by application of uniaxial mechanical stress. The expected spectral features were obtained by simulations and verified by experiments. The spectral features of the heptamer structures were further characterized by using detailed group theoretical analysis. We show that both sub-structural and mechanical tuning could be employed to engineer the Fano resonances in complex plasmonic molecules.

Dynamic tuning and symmetry lowering of Fano resonance in plasmonic nanostructure.

We present dynamic tuning and symmetry lowering of Fano resonances in gold heptamers accomplished by applying uniaxial mechanical stress. The flexible heptamer structure was obtained by embedding the seven-gold-nanocylinder complex in a polydimethylsiloxane membrane. Under uniaxial stress, the Fano resonance exhibited opposite spectral shifts for the two orthogonal polarizations parallel and perpendicular to the mechanical stress. Furthermore, a new resonance was observed for polarization parallel to the mechanical stress but not for the perpendicular polarization. The experimental results showed good agreement with the numerical simulations. A detailed group theoretical analysis showed that the symmetry lowering caused by the mechanical stress not only splits the originally degenerate mode but also modifies the originally optically inactive mode into an optically active mode, which then interacts strongly with a closely spaced mode and exhibits anticrossing behavior. The symmetry tuning enabled by applying mechanical stress is a simple and efficient way to engineer the nature of coupled plasmon resonances in complex nanostructures. The mechanically tunable plasmonic nanostructures also provide an excellent platform for dynamically tunable nanophotonic devices such as tunable filters and sensors.

Thermo-optically tunable silicon photonic crystal light modulator.

We designed, fabricated, and characterized a thermo-optically tunable compact (10 µm × 10 µm) silicon photonic crystal (PhC) light modulator that operates at around 1.55 µm for TE polarization. The operational principle of the device is the modulation of the cutoff frequency in a silicon-based line defect PhC. The cutoff frequency is shifted because of the thermo-optic tuning of the silicon refractive index, which is realized by localized heating on the PhC. The modulator is formed by a triangular lattice array of cylindrical air holes on a silicon-on-insulator wafer. Optical characterization was carried out, and the result clearly showed thermo-optic tuning of the cutoff frequency at around 1.55 µm.

Cell encapsulation and oxygenation in nanoporous microcontainers.

With strides in stem cell biology, cell engineering and molecular therapy, the transplantation of cells to produce therapeutic molecules endogenously is an attractive and achievable alternative to the use of exogenous drugs. The encapsulation of such cell transplants in semi-permeable, nanoporous constructs is often required to protect them from immune attack and to prevent their proliferation in the host. However, effective graft immunoisolation has been mostly elusive owing to the absence of a high-throughput method to create precisely controlled, high-aspect-ratio nanopores. To address the clinical need for effective cell encapsulation and immunoisolation, we devised a biocompatible cell-encapsulating microcontainer and a method to create highly anisotropic nanopores in the microcontainer's surface. To evaluate the efficacy of these nanopores in oxygenating the encapsulated cells, we engineered 9L rat glioma cells to bioluminesce under hypoxic conditions. The methods described above should aid in evaluating the long term survival and efficacy of cellular grafts.