Dichotomous feedback: a signal sequestration-based feedback mechanism for biocontroller design

We introduce a new design framework for implementing negative feedback regulation in synthetic biology, which we term ‘dichotomous feedback’. Our approach is different from current methods, in that it sequesters existing fluxes in the process to be controlled, and in this way takes advantage of the process’s architecture to design the control law. This signal sequestration mechanism appears in many natural biological systems and can potentially be easier to realize than ‘molecular sequestration’ and other comparison motifs that are nowadays common in biomolecular feedback control design. The loop is closed by linking the strength of signal sequestration to the process output. Our feedback regulation mechanism is motivated by two-component signalling systems, where a second response regulator could be competing with the natural response regulator thus sequestering kinase activity. Here, dichotomous feedback is established by increasing the concentration of the second response regulator as the level of the output of the natural process increases. Extensive analysis demonstrates how this type of feedback shapes the signal response, attenuates intrinsic noise while increasing robustness and reducing crosstalk.     

Contribution of different bitter-sensitive taste cells to feeding inhibition in a caterpillar (Manduca sexta).

Many compounds that taste bitter to humans also inhibit feeding in insects. Caterpillars (e.g., Manduca sexta) detect these compounds with a few bitter-sensitive taste cells. This study examined the role of these taste cells in feeding inhibition. Behavioral studies demonstrated that 3 bitter compounds (caffeine, salicin, and aristolochic acid) all inhibited feeding rapidly in Manduca sena. Electrophysiological studies revealed that each pair of bitter-sensitive taste cell differs in responsiveness to the bitter compounds. Ablation studies indicated that (a) those pairs of bitter-sensitive taste cells that responded vigorously to a particular bitter compound were sufficient to inhibit feeding on diets containing the same compound, but that (b) no pair of bitter-sensitive taste cells was necessary for inhibiting feeding. Thus, the different pairs of bitter-sensitive taste cells appear to make partially redundant contributions to feeding inhibition. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

High nitrification rate at low pH in a fluidized bed reactor with chalk as the biofilm carrier.

A typical steady state bulk pH of about 5 was established in a nitrifying fluidized bed with chalk as the only buffer agent. In spite of the low pH, high rate nitrification was observed with the nitrification kinetic parameters in the chalk reactor similar to those of biological reactors operating at pH>7. Various methods were used to determine the reasons for high rate nitrification at such low pH including (i) determination of bacterial species, (ii) microsensor measurements in the biofilm, and (iii) comparison of nitrification performance at low pH with a non-chalk fluidized bed reactor. Fluorescence in situ hybridization (FISH) using existing 16S rRNA-targeted oligonucleotide probes showed common nitrifying bacteria in the low pH chalk reactor. The prevalent nitrifying bacteria were identified in the Nitrosomonas oligotropha, Nitrosomonas europeae/eutropha, Nitrosospira and Nitrospira related groups, all well known nitrifiers. Microelectrode measurements showed that the pH in the biofilm was low and similar to that of the bulk pH. Finally, reactor performance using a non-chalk biofilm carrier (sintered glass) with the same bacterial inoculum also showed high rate nitrification below pH 5. The results suggest that inhibition of nitrification at low pH is highly overestimated.     

Changes in ammonia oxidiser population during transition to low pH in a biofilm reactor starting with Nitrosomonas europaea.

Recent experiments in our laboratory using both biofilm and suspended biomass reactors have demonstrated high rate nitrification at low pH with known autotrophic nitrifying bacteria originating from wastewater treatment plants refuting previous assumptions that nitrification is significantly inhibited at low pH. Since much of the earlier microbiological work regarding ammonia oxidising bacteria (AOB) physiology was carried out using Nitrosomonas europaea, this model bacterium's capability for high rate nitrification at low pH in a continuous biofilm reactor was tested. A biofilm reactor filled with sintered glass particles was inoculated with a pure culture of N. europaea. The reactor was first operated to high nitrification rates under conditions favourable to N. europaea (pH > 7; high ammonium concentrations). To eliminate inhibitory concentrations of nitrite at low pH, an enriched culture of Nitrospira (a nitrite oxidising bacterium) was then added. The transition from neutral to acidic conditions was attempted by sharply lowering the nitrification rate and by using a feeding solution containing insufficient buffer for complete nitrification. As opposed to other successful transitions, the pH in the N. europaea/Nitrospira reactor initially dropped only slightly and maintained pH > 6 for over two weeks. The reactor reached pH 4.5 only after four weeks. FISH results showed that while the percent of AOB and Nitrospira to eubacteria remained relatively constant at 51.1 +/- 8.2% and 40.8 +/- 6.4%, respectively, the AOB community changed completely in 60 days from 100% N. europaea to 100% Nitrosomonas oligotropha. Even though N. oligotropha was not intentionally introduced into the reactor, it is apparently much better adapted to conditions of low pH.     

Pt nanoparticles sputter-deposited on TiO2/MWCNT composites prepared by atomic layer deposition: Improved electrocatalytic activity towards the oxygen reduction reaction and durability in acid media

Acid-treated multi-walled carbon nanotubes (MWCNTs) were decorated with TiO₂ using the atomic layer deposition (ALD) technique followed by uniform distribution of platinum nanoparticles (PtNPs) through magnetron sputtering. Surface analyses were performed by scanning electron microscopy (SEM), scanning transmission electron microscopy (STEM) and X-ray photoelectron spectroscopy (XPS). Electrochemical decontamination and characterization of the Pt-TiO₂/MWCNT electrodes were carried out by CO stripping followed by cyclic voltammetry in acid media. The oxygen reduction reaction (ORR) was studied in O₂-saturated 0.05 M H₂SO₄ solution using the rotating disk electrode (RDE) method. Durability of the prepared catalysts was examined by repetitive potential cycling. Electrochemical data obtained was analyzed and compared to that of the commercial Pt/C catalyst. It was revealed that the Pt-TiO₂/MWCNT catalysts possess higher ORR activity and better durability as compared to that of the commercial Pt/C.     

Characterization of asymmetric rhombohedral twin in epitaxial α-Cr2O3 thin films by X-ray and electron diffraction

Epitaxial chromium oxide (α-Cr₂O₃) films grown by atomic layer deposition at 375 °C from CrO₂Cl₂ and CH₃OH on (1 1¯ 0 2) oriented α-Al₂O₃ have been studied by reflection high-energy electron diffraction (RHEED), X-ray diffraction (XRD) and X-ray reflection (XRR). The thickness of the films ranged from 10 to 310 nm, and the average growth rate was 0.1 nm per deposition cycle. According to the XRD analysis, the orientation relationship in thinner films was (1 1¯ 0 2)[1 1 0]Cr₂O₃ || (1 1¯ 0 2)[1 1 0]Al₂O₃. Confirmed by the RHEED and XRD analyses, (1¯ 1 0 2) became the preferred growth plane at the thicknesses above 40 nm. This change has been interpreted as the appearance of an asymmetric rhombohedral twin with the orientation relationship between the layers (1¯ 1 0 2)[1 1 0]top || (1 1¯ 0 2)[1 1 0]bottom and (1¯ 1 0 2)[1 1¯ 1]top || (1 1¯ 0 2)[1¯ 1 1]bottom. The match of the anion and cation sublattices of both layers was characterized in terms of the structural model of the twin interface.     

Parametrized model reduction based on semidefinite programming

A parametrized model in addition to the control and state-space variables depends on time-independent design parameters, which essentially define a family of models. The goal of parametrized model reduction is to approximate this family of models. In this paper, a reduction method for linear time-invariant (LTI) parametrized models is presented, which constitutes the development of a recently proposed reduction approach. Reduced order models are computed based on the finite number of frequency response samples of the full order model. This method uses a semidefinite relaxation, while enforcing stability on the reduced order model for all values of parameters of interest. As a main theoretical statement, the relaxation gap (the ratio between upper and lower bounds) is derived, which validates the relaxation. The proposed method is flexible in adding extra constraints (e.g., passivity can be enforced on reduced order models) and modifying the objective function (e.g., frequency weights can be added to the minimization criterion). The performance of the method is validated on a numerical example.     

Chalk as the carrier for nitrifying biofilm in a fluidized bed reactor

A fluidized bed reactor for nitrification with chalk as the biomass carrier and the sole buffer agent was studied. Chalk dissolution in the reactor was found to follow the stoichiometric ratio of 1 mole of CaCO3 dissolved for each mole of NH4+ oxidized. Three batches of chalk, each one having a different dissolution rate, were used to replace the dissolved chalk. The three dissolution rates resulted in three different steady state pH levels in the reactor (4.7-6.6) and nitrification rates. Nitrification was found to be limited by either the chalk dissolution rate or dissolved oxygen concentration depending on the type of chalk used. A maximal nitrification rate of 1.44 g NH4(+)-N/l reactor.d was observed. The average cell yield was 0.1 g cells/g N oxidized, similar to the cell yield during reactor start-up when the pH was 7. The specific ammonium oxidation rates varied between 0.08 and 0.15 mg NH4(+)-N oxidized/mg protein.h, values which are in the reported range for nitrification at pH 7 to 8. Oxygen update rate (OUR) results indicated that the major mechanism responsible for the high nitrification rate observed in the reactor operating at low pH seems to be the favorable microenvironment provided by the chalk.     

Simultaneous removal of atrazine and nitrate using a biological granulated activated carbon (BGAC) reactor

The objective of this research was to characterize the performance of granulated activated carbon (GAC) as a carrier for Pseudomonas ADP in a non‐sterile continuous fluidized bed reactor for atrazine degradation under anoxic conditions. The GAC was compared with two non‐adsorbing carriers: non‐adsorbing carbon particles (‘Baker product’) having the same surface area available for biofilm growth as the GAC, and sintered glass beads. The initial atrazine degradation efficiency was higher than 90% in the reactors with the non‐adsorbing carriers, but deteriorated to 20% with time due to contamination by foreign denitrifying bacteria. In contrast, no deterioration was observed in the biological granulated activated carbon (BGAC) reactor. A maximal atrazine volumetric and specific degradation rate of 0.820 ± 0.052 g atrazine dm^?3 day^?1 and 1.7 ± 0.4 g atrazine g^?1 protein day^?1 respectively were observed in the BGAC reactor. Concurrent atrazine biodegradation and desorption from the carrier was shown and an effluent concentration of 0.002 mg dm^?3 (below the EPA standard) was achieved in the BGAC reactor. The advantages of the BGAC reactor over the non‐adsorbing carrier reactors can probably be explained by the adsorption–desorption mechanism providing favorable microenvironmental conditions for atrazine–degrading bacteria. Copyright © 2004 Society of Chemical Industry     

The effect of CO2 concentration on a nitrifying chalk reactor

The effect of CO2 concentration on nitrification rate was studied in a fluidized bed reactor using chalk (solid calcium carbonate) as the biomass carrier and buffering agent. Using one chalk type and uniform particle size, carbon dioxide was found to limit the nitrification rate in the reactor at concentrations up to 0.3 mmol l(-1). At this concentration the nitrification rate was about 2.5-2.7g NH4+-Nl reactor(-1) d(-1). The pH established in the reactor varied between 4.5 and 5.5, remarkably with lower pH obtained remarked at higher nitrification rates. Kinetic parameters for nitrification rate with CO2 as the rate limiting substrate were determined: a Michaelis-Menten constant, Km, of 0.013 mmol l(-1) CO2 and a maximum ammonium oxidation rate of 2.33g NH4+-Nl reactor(-1) d(-1).