Synthetic Cultures Research Articles

Engineering Microbial Consortia as Living Materials: Advances and Prospectives.

The field of Engineered Living Materials (ELMs) integrates engineered living organisms into natural biomaterials to achieve diverse objectives. Multiorganism consortia, prevalent in both naturally occurring and synthetic microbial cultures, exhibit complex functionalities and interrelationships, extending the scope of what can be achieved with individual engineered bacterial strains. However, the ELMs comprising microbial consortia are still in the developmental stage. In this Review, we introduce two strategies for designing ELMs constituted of microbial consortia: a top-down strategy, which involves characterizing microbial interactions and mimicking and reconstructing natural ecosystems, and a bottom-up strategy, which entails the rational design of synthetic consortia and their assembly with material substrates to achieve user-defined functions. Next, we summarize technologies from synthetic biology that facilitate the efficient engineering of microbial consortia for performing tasks more complex than those that can be done with single bacterial strains. Finally, we discuss essential challenges and future perspectives for microbial consortia-based ELMs.

Yarrowia lipolytica CMGB32 Biosurfactants Produced Using n-Hexadecane: Developing Strategies for Environmental Remediation

The yeast Yarrowia lipolytica degrades petroleum compounds, including alkanes, via the monoterminal oxidation pathway, the hydrophobic carbon substrate assimilation is mediated by biosurfactants, and extracellular amphiphilic molecules are produced by the yeast cell. This study focuses on the ability of the strain Y. lipolytica CMGB32 to degrade n-hexadecane by producing biosurfactants with high potential for bioremediation. The hydrocarbon-degrading potential of the yeast strain was observed via a 2,6-dichlorophenolindophenol (DCPIP) test in Bushnell–Hass medium with 1% n-hexadecane, and cell hydrophobicity was expressed as microbial adhesion to hydrocarbons (MATH). Biosurfactant production on yeast peptone (YP) with 1% n-hexadecane was estimated after 72 h using the emulsification index (E24%) against toluene. Crude biosurfactant (cell-free broth) stability tests were performed at different temperatures (4 °C, 70 °C) and NaCl concentrations (2–10%). The effects of a biosurfactant on synthetic wastewater remediation comprised the growth curves (OD measurements) of natural heavy metal degrader Rhodotorula mucilaginosa, determination of nutrients (spectrophotometrically), physico-chemical parameters, and removal capacity of lead and cadmium ions (via inductively coupled plasma mass spectrometry—ICP-MS). The antimicrobial and anti-adherence activities of 20 mg/mL and 40 mg/mL of the biosurfactant against pathogenic Candida krusei strains involved growth observations and the crystal violet microtiter method. The DCPIP decolorization occurred after six days, corresponding to the maximum growth phase of the Y. lipolytica culture. After 72 h, the cells presented high hydrophobicity (82.61% MATH) and stable biosurfactant production (E24% 47%). The crude biosurfactant (5%) increased the growth of R. mucilaginosa strains cultivated on synthetic wastewater cultures contaminated with Pb2+ and Cd2+, increased the conductivity and COD (86%) of the samples, and determined Pb2+ (66%) and Cd2+ (42%) ions reduction. The concentrated biosurfactant inhibited C. krusei growth (70%) and biofilm adherence. In conclusion, Y. lipolytica CMGB32 shows important potential for development of biosurfactant-based technologies for the remediation of heavy-metal- and emerging pathogen-contaminated wastewaters.

Metabolite cross-feeding enables concomitant catabolism of chlorinated methanes and chlorinated ethenes in synthetic microbial assemblies.

Isolate studies have been a cornerstone for unraveling metabolic pathways and phenotypical (functional) features. Biogeochemical processes in natural and engineered ecosystems are generally performed by more than a single microbe and often rely on mutualistic interactions. We demonstrate the rational bottom-up design of synthetic, interdependent co-cultures to achieve concomitant utilization of chlorinated methanes as electron donors and organohalogens as electron acceptors. Specialized anaerobes conserve energy from the catabolic conversion of chloromethane or dichloromethane to formate, H2, and acetate, compounds that the organohalide-respiring bacterium Dehalogenimonas etheniformans strain GP requires to utilize cis-1,2-dichloroethenene and vinyl chloride as electron acceptors. Organism-specific qPCR enumeration matched the growth of individual dechlorinators to the respective functional (i.e. dechlorination) traits. The metabolite cross-feeding in the synthetic (co-)cultures enables concomitant utilization of chlorinated methanes (i.e. chloromethane and dichloromethane) and chlorinated ethenes (i.e. cis-1,2-dichloroethenene and vinyl chloride) without the addition of an external electron donor (i.e. formate and H2). The findings illustrate that naturally occurring chlorinated C1 compounds can sustain anaerobic food webs, an observation with implications for the development of interdependent, mutualistic communities, the sustenance of microbial life in oligotrophic and energy-deprived environments, and the fate of chloromethane/dichloromethane and chlorinated electron acceptors (e.g. chlorinated ethenes) in pristine environments and commingled contaminant plumes.

Enabling high throughput drug discovery in 3D cell cultures through a novel bioprinting workflow.

Advanced three dimensional cell culture techniques have been adopted in many laboratories to better model in vivo tissue by recapitulating multi-cellular architecture and the presence of extracellular matrix features. We describe here a 3D cell culture platform in a small molecule screening workflow that uses traditional biomarker and intracellular kinase end point assay readouts. By combining the high throughput bioprinter RASTRUM with the high throughput screening assay AlphaLISA, we demonstrate the utility of the protocol in 3D synthetic hydrogel cultures with breast cancer (MDA-MB-231 and MCF-7) and fibroblast cells. To establish and validate the workflow, we treated the breast cancer cultures with doxorubicin, while fibroblast cultures were stimulated with the pro-inflammatory lipopolysaccharide. 3D and 2D MDA-MB-231 cultures were equally susceptible to doxorubicin treatment, while showing opposite ERK phosphorylation changes. Doxorubicin readily entered embedded MCF-7 spheroids and markedly reduced intracellular GSK3β phosphorylation. Furthermore, quantifying extracellular interleukin 6 levels showed a very similar activation profile for fibroblasts in 2D and 3D cultures, with 3D fibroblast networks being more resistant against the immune challenge. Through these validation experiments we demonstrate the full compatibility of the bioprinted 3D cell cultures with several widely-used 2D culture assays. The efficiency of the workflow, minimal culture handling, and applicability of traditional screening assays, demonstrates that advanced encapsulated 3D cell cultures can be used in 2D cell culture screening workflows, while providing a more holistic view on cell biology to increase the predictability to in vivo drug response.

Corn stover conversion into bioethanol and xylitol through an integral bioprocess: Kinetic study and modelling

BackgroundCorn stover residue is an important lignocellulosic material with a biochemical composition that arouses an interest in transforming into bioethanol and high added value product, such as e.g., xylitol. MethodsHydrolysis experiments were carried out in a semi-pilot reactor of 12 L capacity. Fermentation experiments were conducted in a 1 L batch bioreactor capacity. Detailed kinetic study of the Candida guilliermondii NRC 5578 growth was carried out. Fermentation and the xylitol production process were modelled. Significant FindingsThe used crude corn stover residue was composed of 11.1 % of lignin, 27.1 % of hemicellulose, 32.6 % cellulose, 24.5 % pentosanes, and 33.0 % of α-cellulose. Reconditioning liquid fractions of prehydrolysate and hydrolysate at 1.4 % (w/v) of H2SO4 were determined for the prehydrolysate 5.31 g/L, 9.07 g/L, 3.07 g/L, 11.2 g/L, 3.24 g/L, 3.89 and 2.56 g/L for pentoses, hexoses, arabinose, xylose, mannose, glucose, and fructose, respectively, and for hydrolysate 15.2 g/L of glucose. The higher values of maximum specific growth rate were registered for control experiments and compared to prehydrolysated and hydrolysated cultures. The highest ethanol formation rates of 2.29 g/(g L) and 0.117 g/(g L) in synthetic and hydrolysated cultures, respectively, were registered when 7 g/L glucose was used as carbon source.

Dynamical robustness of collective neuronal activity upon targeted damage in interdependent networks

In the last decades, the availability of data about the structure of social, technological and biological systems has provided important insights on the mechanisms governing their correct functioning and robustness. These mechanisms are grounded on the complex backbone of interactions among the constituents of the system, which include both topological and dynamical aspects. Here, we analyze interdependent networks composed of two layers of interacting neuronal units and explore their robustness when these synthetic cultures are subjected to damage in the form of either targeted attack or failure. Our results show that the functionality of these networks does not decrease monotonically with damage but, on the contrary, they are able to increase their level of activity when the experienced damage is sufficiently strong.

Differential detection of zika virus based on PCR

Tropical countries are highly prone to infectious diseases such as the one caused by zika virus. Infection by zika is clinically and epidemiologically highly relevant. For example, when women are infected by zika during the first trimester of pregnancy, the child incurs a high risk of microcephaly and acute neurological syndromes. In adults, the virus is associated with the Guillain-Barré syndrome and other disorders. The worldwide emergency caused by zika in 2013/14 demonstrated the need for rapid and accurate diagnostic tools for the virus. Current diagnostic methods include virus isolation, serological tests, and molecular assays. However, virus isolation requires labor-intensive and time-consuming cell culture; serological detection suffers from cross-reactivity caused by previous exposure to homologous arboviruses that cause symptoms like those caused by zika, while molecular tools commonly are not designed for differential zika detection. This work reports on developing a specific molecular detection method based on phylogenetically conserved primers designed for the specific diagnosis of the zika virus. The zika primers were systematically selected through a rigorous bioinformatic analysis and demonstrated the capability to be highly specific. We tested our primers on synthetic DNA, cell cultures and samples from patients infected with zika, dengue and chikungunya and found that they detected zika with specificity high enough for differential virus diagnosis.

Pure cultures for synthetic culture development: Next level municipal waste treatment for polyhydroxyalkanoates production

Polyhydroxyalkanoates (PHAs), as bio-based plastics, promise a transition from petroleum products to green and sustainable alternatives. However, their commercial production is yet impeded by high production costs. In this study, we assessed synthetic culture in mono and co-culture modes for bacterial PHA production. It was demonstrated that volatile fatty acids (VFAs) derived from food waste and primary sludge are cheap carbon sources for maintaining high production yields in the synthetic cultures. The maximum obtained PHA was 77.54 ± 5.67% of cell dried weight (CDW) (1.723 g/L) from Cupriavidus necator and 54.9 ± 3.66% of CDW (1.088 g/L) from Burkholderia cepacia. The acquired results are comparable to those in literature using sugar substrates. Comparatively, lower PHA productions were obtained from the co-cultivations ranging between 36-45 CDW% (0.39–0.48 g/L). Meanwhile, the 3-hydroxyvalerate content in the biopolymers were increased up to 21%.

Volatile fatty acid production from semi-synthetic milk processing wastewater under alkali pH: The pearls and pitfalls of microbial culture.

Volatile fatty acids (VFA) are one of the most promising sustainable and environmentally friendly bioproduct owing to their wide usage area and high market demand. For this reason, in this study, the evaluation of VFA production from pure and mixed bacterial cultures was aimed. Three different mixed cultures with C. aceticum, C. butyricum and P. acidipropionici as pure cultures were used for inoculation of milk processing wastewater fermentation under pH 10 for 15days. The mixed culture fermentation had the highest VFA production efficiency whereas the highest amount of acetic, butyric and propionic acid productions were obtained by C. aceticum, C. butyricum and P. acidipropionici, respectively. Also, the mixed cultures demonstrated faster pH regulation and acclimation than the pure cultures tested. Therefore, development of synthetic cultures may offer a useful approach to produce VFA mixtures with one-dominant acid type and with high production efficiency.

Impact of targeted attack on the spontaneous activity in spatial and biologically-inspired neuronal networks.

We study the structural and dynamical consequences of damage in spatial neuronal networks. Inspired by real in vitro networks, we construct directed networks embedded in a two-dimensional space and follow biological rules for designing the wiring of the system. As a result, synthetic cultures display strong metric correlations similar to those observed in real experiments. In its turn, neuronal dynamics is incorporated through the Izhikevich model adopting the parameters derived from observation in real cultures. We consider two scenarios for damage, targeted attacks on those neurons with the highest out-degree and random failures. By analyzing the evolution of both the giant connected component and the dynamical patterns of the neurons as nodes are removed, we observe that network activity halts for a removal of 50% of the nodes in targeted attacks, much lower than the 70% node removal required in the case of random failures. Notably, the decrease of neuronal activity is not gradual. Both damage scenarios portray "boosts" of activity just before full silencing that are not present in equivalent random (Erdös-Rényi) graphs. These boosts correspond to small, spatially compact subnetworks that are able to maintain high levels of activity. Since these subnetworks are absent in the equivalent random graphs, we hypothesize that metric correlations facilitate the existence of local circuits sufficiently integrated to maintain activity, shaping an intrinsic mechanism for resilience.

Comparison of the behaviour of Brettanomyces bruxellensis strain LAMAP L2480 growing in authentic and synthetic wines.

Brettanomyces bruxellensis is the main microorganism responsible for the production of off-flavours in wine. Studies have been carried out in synthetic cultures using p-coumaric acid for the production of vinyl and ethylphenols. The results obtained have been extrapolated to authentic wine, but there is no evidence that this correlation will be correct. We studied the behaviour of B. bruxellensis native strain LAMAP L2480 in authentic wine and in a synthetic medium with a chemical composition similar to the authentic wine used in this study (basal synthetic wine + pH, ethanol and hydroxycinnamic acid concentrations of commercial wine). In some assays, B. bruxellensis has been studied using media containing 100 mg L(-1) p-coumaric acid, so we also used the same concentration added to the authentic and synthetic wines. The microorganism showed better growth in authentic wine, regardless of the presence of p-coumaric acid. In the case of synthetic wine, the addition of p-coumaric acid caused a delay in yeast growth and an increase in the production of volatile phenols. The coumarate decarboxylase activity did not show any difference regardless of the media and the presence of p-coumaric acid. Vinylphenol reductase showed higher activity when a higher concentration of p-coumaric acid was added in synthetic wine, but no change was observed in authentic wine.

Learning to Overcome Cultural Conflict through Engaging with Intelligent Agents in Synthetic Cultures

Providing opportunities for children to engage with intercultural learning has frequently focused on exposure to the ritual, celebrations and festivals of cultures, with the view that such experiences will result in greater acceptance of cultural differences. Intercultural conflict is often avoided, bringing as it does particular pedagogical, ethical and political dilemmas of which cultures we place in conflict in the multicultural classroom. In this paper we discuss an alternative approach, providing children with an interactive learning experience with synthetic cultures and characters. The agent architecture developed to enable intelligent agents to exhibit culturally appropriate affect and behaviours is outlined. MIXER, an experiential learning application developed for 9–11 year old children on intercultural conflict is described, highlighting the learning goals and approaches. A school-based evaluation of MIXER with 144 UK children is presented. Children demonstrated high levels of comprehension, engagement and enjoyment of MIXER, with MIXER contributing to near and far transfer, supporting children’s cognitive, emotional and behavioural learning and stimulating discussion and debate about how to resolve conflict.

Toward a Model for Intercultural Communication in Simulations

The growing need for intercultural literacy in an increasingly interconnected and computer-mediated world contrasts with the dearth of investigation in best practices when designing simulations aimed at improving intercultural communication. Synthetic cultures inspired by real-world cultural traits, problem-based learning, and a social constructivist theoretical base represent core components of immersive learning environments designed to improve intercultural literacy. Through an analysis of the literature, a model is proposed primarily to promote discussion and debate about intercultural simulations. The Model for Intercultural Communication in Simulations asserts that along with those components, the Experiential Learning Cycle (ELC), natural decision making, and third place thinking form an approach in understanding how intercultural simulations should be designed. The Model for Intercultural Communication in Simulations challenges the learner to move beyond known paradigms and encourages simulation and e-learning designers to employ socially situated contexts. Accordingly, within the context of the model, a culminating evaluation of an online intercultural training simulation ( ARGONAUTONLINE) follows the introduction of the model.

Intercultural sensitization with synthetic cultures: Evaluation of a computer-based multimedia learning tool

Abstract The purpose of the present study was to evaluate a new computer-based multimedia learning tool for intercultural sensitization that is based on synthetic cultures. The learning tool named “Culture Awareness Training” (CAT) is targeted at learners with few intercultural experiences, and its aim is to sensitize participants to cultural influences on behavior and to intercultural differences, and to enhance perspective taking. Two studies were conducted to evaluate the effectiveness of the CAT. Study 1 ( N = 107) showed that participation in the CAT led to higher intercultural awareness and higher intercultural interest compared to a control group. In Study 2 ( N = 46), the CAT was compared to a simulation game for intercultural sensitization. Participants were students shortly before their stay abroad. Results showed that both the CAT and the classical simulation game enhanced participants’ cultural awareness.

Glyoxal/glycolaldehyde: a redox system involved in malolactic fermentation of wine.

To verify the presence of glycolaldehyde in wine resulting from reduction of glyoxal after malolactic fermentation, sterilized solutions of synthetic cultures were inoculated with a lactic bacterium of the type Oenococcus oeni. Fermentation was also carried out on solutions with glyoxal added. The resulting glycolaldehyde concentrations turned out to be associated with the amounts of glyoxal present, and glyoxal was seen to decrease as glycolaldehyde increased. In addition, it was observed that glyoxal principally forms from breakdown of sugars and that reduction to glycolaldehyde is mainly promoted by bacterial activity. Finally, the ability of glycolaldehyde to induce browning of (+)-catechin in a model wine system was verified and turned out to be about 10 times higher than that of ascorbic acid.

You Must Have Been at A Different Meeting: Enacting Culture Clash in The International Office of the Future

This article introduces a new concept for exploring the design of IT infrastructure in transnational organizations. This concept is simulation gaming that uses so- called synthetic national culture profiles. The paper describes a game in which a telecommunication architecture was designed for a multinational company. It concludes that synthetic cultures are a powerful tool for generating awareness about the profound potential effects of differences in national culture within multinationals.

Synthetic Cultures: Intercultural Learning Through Simulation Games

This article introduces the concept of synthetic cultures. These are role profiles for enacting dimensions of national culture. The full profiles are given for 10 synthetic cultures, based on Gert Hofstede’s dimensions of national culture. They can be used for training or simulation/gaming. Examples of their application are presented. Apart from examples of existing games, suggestions are made for creating custom simulations that use synthetic cultures. Experiences of players are mentioned, both in face-to-face and in electronically mediated settings. Special attention is paid to debriefing issues. The article concludes with practical advice to prospective game leaders.

The future of wiggling things

Art is revealed to be a religion. Ecology swells beyond its scientific bulwarks, spawning subjective ecology. Wigglism, an ethic of nurturing vital confluences, slithers up out of this essay and surrounds the reader. To understand the emerging digital culture, which has occurred largely outside the purview of ordained “art” circles, we should avoid the verbal tentacles sucking our spirits into an older school of thought which has entered so late, and so grudgingly, into the electronic age. Risk entanglement in the youthful and daring culture which has been percolating in the shadows of the status quo: a generation of hackers, hip hoppers, ravers, and web‐activists. Through this groundswell of fresh cultural insight we might begin to abandon the narrow view of culture and civilisation as a privileged and separate sector of human reality. We might actually hazard a humanist heresy — to exercise a hyper‐green mode of being which interacts with, folds into, and merges with living systems — inclusive of local, popular, interspecies, and even synthetic cultures. We might even expand the definition of ecology to explore a kind of ecological subjectivity, an approach to the world the author calls “Wigglism”. Wigglism engages an ecosystem of feedback loops extending both inside and outside the very wiggle of being.

Formation of endomycorrhizas by an achlorophyllous orchid, Erythrorchis ochobiensis, and Auricularia polytricha

To test the mycorrhizal function of heterobasidiomycetous fungi on achlorophyllous orchids and to examine the symbiotic fungal range of a myco-heterotrophic orchid,Erythrorchis ochobiensis, synthetic cultures of the orchid seed were carried out withAuricularia polytricha isolates from Japan and Mexico. After three and a half mo of incubation, 57.0–70.7% of seeds germinated but none of them showed further growth. When cultured on peat moss at 25°C, the germination rate was 8.7% in the presence of Mexican isolate and 18.0% in the presence of Japanese isolate. Some germinated seeds developed into protocorms, and several seeds incubated with the Mexican isolate developed into plantlets after 5 mo. Pelotons were observed in the cells of protocorms and roots. The results indicated that some heterobasidiomycetous fungi could form endomycorrhizas with a myco-heterotrophic orchid. The results also showed that the symbiont ofE. ochobiensis extends, at least experimentally, to Heterobasidiomycetes. The variances of germination rate and seedling growth were suggested to be affected by the difference of isolates and culture conditions.

Culture-centered counseling and interviewing skills: a practical guide

Culture Centeredness The Multicultural Three-Step The Cultural Grid and Culture Systems Four Synthetic Cultures Culture-Centered Skill Development Preparing to Learn Culture-Centered Skills Basic Attending Skills Paraphrasing, Summarizing, and Encouraging Skills Question-Asking Skills Reflecting Feeling and Reflecting Meaning Confrontation and Mediation Skills Focusing, Directing, and Interpreting Skills References Index