Induction of a dwarf phenotype with IBH1 may enable increased production of plant‐made pharmaceuticals in plant factory conditions

Year‐round production in a contained, environmentally controlled ‘plant factory’ may provide a cost‐effective method to produce pharmaceuticals and other high‐value products. However, cost‐effective production may require substantial modification of the host plant phenotype; for example, using dwarf plants can enable the growth of more plants in a given volume by allowing more plants per shelf and enabling more shelves to be stacked vertically. We show here that the expression of the chimeric repressor for Arabidopsis AtIBH1 (P35S:AtIBH1SRDX) in transgenic tobacco plants (Nicotiana tabacum) induces a dwarf phenotype, with reduced cell size. We estimate that, in a given volume of cultivation space, we can grow five times more AtIBH1SRDX plants than wild‐type plants. Although, the AtIBH1SRDX plants also showed reduced biomass compared with wild‐type plants, they produced about four times more biomass per unit of cultivation volume. To test whether the dwarf phenotype affects the production of recombinant proteins, we expressed the genes for anti‐hepatitis B virus antibodies (anti‐HBs) in tobacco plants and found that the production of anti‐HBs per unit fresh weight did not significantly differ between wild‐type and AtIBH1SRDX plants. These data indicate that P35S:AtIBH1SRDX plants produced about fourfold more antibody per unit of cultivation volume, compared with wild type. Our results indicate that AtIBH1SRDX provides a useful tool for the modification of plant phenotype for cost‐effective production of high‐value products by stably transformed plants in plant factory conditions.     

A new agenda for blue agave landraces: food,energy and tequila

Agave tequilana Weber (Rigidae, Agavaceae), blue agave, is a native Mexican plant that has been associated with tequila since the 17th century. The tequila industry has matured over time and now has a geographical indication (Denominación de Origen; DOT). The tequila industry has grown substantially in the last 15 years (19.82% annual increase between 1995 and 2008), resulting in an increase in agave production and associated residue (leaves) and bagasse that can be used for second‐generation biofuels. At a time when the biofuel industry is undergoing unprecedented changes, with diversified demand and predictions of increased competitiveness, this paper presents a review of agave landraces that have been affected by tequila production but may be beneficial for a biofuel industry. Conventional botanical studies have revealed domestication syndromes in races related to blue agave (‘azul listado’, ‘sigüín’ and ‘pata de mula’) specifically for production of fructans in the plant core as would be expected in mezcal agaves (including those used for tequila). Some others, such as the ‘moraleño’ and ‘bermejo’ cultivars (Sisalanae) show domestication syndrome only in the fibers, while others, such as ‘chato,’A. americana L. subtilis (Americanae) show domestication syndrome in fructans and fibers and ‘zopilote,’A. rhodacantha (Rigidae) a relatively low domestication syndrome. No specimens of the cultivars named ‘mano larga’, ‘mano anchaque’ and ‘cucharo’ were found in the Tequila Region of Origin (Western Mexico). The genetic resources from landraces ignored by the tequila industry may be valuable for both ethanol production and conservation.     

Hosting the plant cells in vitro: recent trends in bioreactors

Biotechnological production of high-value metabolites and therapeutic proteins by plant in vitro systems has been considered as an attractive alternative of classical technologies. Numerous proof-of-concept studies have illustrated the feasibility of scaling up plant in vitro system-based processes while keeping their biosynthetic potential. Moreover, several commercial processes have been established so far. Though the progress on the field is still limited, in the recent years several bioreactor configurations has been developed (e.g., so-called single-use bioreactors) and successfully adapted for growing plant cells in vitro. This review highlights recent progress and limitations in the bioreactors for plant cells and outlines future perspectives for wider industrialization of plant in vitro systems as “green cell factories” for sustainable production of value-added molecules.     

‘Shit Happens’: The Selling of Risk in Extreme Sport

This article details the particular commodification of those high‐risk, highadrenalin activities known collectively as ‘extreme sports’. A variety of commercial operators now offer relative sporting neophytes the chance to take part in mountaineering, snow boarding or canyonning adventures that are billed as being ‘high thrill, low risk’. It is the way in which the risk and danger involved in these activities is discursively managed that is of particular interest for this article. The argument developed is that in selling extremity through a range of primarily tourist‐oriented commercial avenues, the very real prospect of death and injury has been stripped from the activity itself. To elaborate this position, this article draws on several sporting disasters, including the much publicised, ill‐fated ascent of Mount Everest in 1996, and the Interlaken canyonning disaster of 1999, as well as the burgeoning literary and media genre—the made‐for‐Hollywood ‘adventure saga’.     

Potential of plants to produce recombinant protein products

Plants have great potential as photosynthetic factories to produce pharmaceutically important and commercially valuable biomedicines and industrial proteins at low cost. The U.S. Food and Drug Administration (U.S. FDA) has approved the drug Elelyso (taliglucerase alfa) produced by carrot cells for treatment of type 1 Gaucher’s disease in 2012. The commercial potential of biomedicines produced by molecular farming has dramatically improved due to the success of an experimental drug called ZMapp, which has immunological activity in Ebola patients. A cocktail of three monoclonal antibodies was produced in tobacco (Nicotiana benthamiana) plants (Chen and Davis 2016). At present, very few drugs made by this technology have been approved by worldwide authorities such as the U.S. FDA. However, plants have been proposed as a novel paradigm for commercial production of proteins over the next decade. In recent years, leading researchers on molecular farming have given more priority to the area of animal-free therapeutic proteins such as parenteral and oral vaccines. Although plant-based platforms have considerable advantages over traditional systems such as bacterial and animal systems, there are several obstacles to commercial-scale production, especially with regards to improving the quality and quantity of plant-produced biologics and industrial materials. One of the biggest barriers to commercialization of this technology is the intense scrutiny of these new plant varieties by regulatory agencies and the public as well as the high costs associated with their regulatory approval.     

Bioreactors for plant cells: hardware configuration and internal environment optimization as tools for wider commercialization

Mass production of value-added molecules (including native and heterologous therapeutic proteins and enzymes) by plant cell culture has been demonstrated as an efficient alternative to classical technologies [i.e. natural harvest and chemical (semi)synthesis]. Numerous proof-of-concept studies have demonstrated the feasibility of scaling up plant cell culture-based processes (most notably to produce paclitaxel) and several commercial processes have been established so far. The choice of a suitable bioreactor design (or modification of an existing commercially available reactor) and the optimization of its internal environment have been proven as powerful tools toward successful mass production of desired molecules. This review highlights recent progress (mostly in the last 5 years) in hardware configuration and optimization of bioreactor culture conditions for suspended plant cells.     

Winter hardiness of Miscanthus (I): Overwintering ability and yield of new Miscanthus ×giganteus genotypes in Illinois and Arkansas

Miscanthus ×giganteus (M×g) is an important bioenergy feedstock crop. However, biomass production of Miscanthus has been largely limited to one sterile triploid cultivar, M×g ‘1993‐1780’, which we demonstrate can have insufficient overwintering ability in temperate regions with cold winters. Key objectives for Miscanthus breeding include greater biomass yield and better adaptation to different production environments than M×g ‘1993‐1780’. In this study, we evaluated 13 M×g genotypes, including ‘1993‐1780’, in replicated field trials conducted for three years at Urbana, IL; Dixon Springs, IL; and Jonesboro, AR. Entries were phenotyped for first‐winter overwintering ability and plant hardiness (ratio of new tillers to old), yield in years 2 and 3, and first heading date, plant height, and culm number in years 1 and 2. We observed substantial variation for overwintering ability and biomass yield among the M×g genotypes tested and identified ones with better overwintering ability and/or higher biomass yield than ‘1993‐1780’. Most entries at Urbana were damaged during the first winter, whereas few or no entries were damaged at Dixon Springs or Jonesboro. However, M×g ‘Nagara’ was entirely undamaged during the first winter and produced high biomass yields at Urbana (19.7 Mg/ha in year 2 and 20.9 Mg/ha in year 3), whereas M×g ‘1993‐1780’ exhibited an overwintering loss of 29%, had severely damaged survivors (hardiness score of 25%), and reduced biomass yield (8.1 Mg/ha in year 2 and 16.2 Mg/ha in year 3), indicating that M×g ‘Nagara’ could be a better choice in hardiness zone 5 (average annual minimum air temperature of ?23.3 to ?28.9°C) or lower. In Dixon Springs, where M×g ‘1993‐1780’ was undamaged by the first winter, it yielded highest among all the entries (21.6 Mg/ha in year 3), though not significantly higher than M×g ‘Nagara’ (18.2 Mg/ha in year 3).     

Microbial alkane production for jet fuel industry: motivation,state of the art and perspectives

Bio‐jet fuel has attracted a lot of interest in recent years and has become a focus for aircraft and engine manufacturers, oil companies, governments and researchers. Given the global concern about environmental issues and the instability of oil market, bio‐jet fuel has been identified as a promising way to reduce the greenhouse gas emissions from the aviation industry, while also promoting energy security. Although a number of bio‐jet fuel sources have been approved for manufacture, their commercialization and entry into the market is still a far way away. In this review, we provide an overview of the drivers for intensified research into bio‐jet fuel technologies, the type of chemical compounds found in bio‐jet fuel preparations and the current state of related pre‐commercial technologies. The biosynthesis of hydrocarbons is one of the most promising approaches for bio‐jet fuel production, and thus we provide a detailed analysis of recent advances in the microbial biosynthesis of hydrocarbons (with a focus on alkanes). Finally, we explore the latest developments and their implications for the future of research into bio‐jet fuel technologies.     

Urban Space and the Mediation of Political Action in Nepal: Local Television,Ritual Processions and Political Violence as Technologies of Enchantment

This paper examines how political identities in the town of Tansen in the central western district of Palpa, Nepal, are mediated by contrasting forms of cultural and material practice: religious and secular processions and programs made by a local, cable‐television production organisation. These practices and their materiality are conceptualised as ‘technologies of enchantment’ (Gell 1992) through which political culture is made manifest in urban space. Paradigmatically ‘modern’ and ‘traditional’ technologies are juxtaposed in order to analyse the different ways that political action is embodied within the community. The loss of life in Tansen and the destruction of buildings associated with these practices in the course of the 10‐year Maoist insurgency provide a tragic confirmation of the conclusions reached in this paper.     

Validation of leaf and microbial pectinases: commercial launching of a new platform technology

Almost all current genetically modified plant commercial products are derived from seeds. The first protein product made in leaves for commercial use is reported here. Leaf pectinases are validated here with eight liquid commercial microbial enzyme products for textile or juice industry applications. Leaf pectinases are functional in broad pH/temperature ranges as crude leaf extracts, while most commercial enzyme products showed significant loss at alkaline pH or higher temperature, essential for various textile applications. In contrast to commercial liquid enzymes requiring cold storage/transportation, leaf pectinase powder was stored up to 16 months at ambient temperature without loss of enzyme activity. Commercial pectinase products showed much higher enzyme protein PAGE than crude leaf extracts with comparable enzyme activity without protease inhibitors. Natural cotton fibre does not absorb water due to hydrophobic nature of waxes and pectins. After bioscouring with pectinase, measurement of contact‐angle water droplet absorption by the FAMAS videos showed 33 or 63 (leaf pectinase), 61 or 64 (commercial pectinase) milliseconds , well below the 10‐second industry requirements. First marker‐free lettuce plants expressing pectinases were also created by removal of the antibiotic resistance aadA gene. Leaf pectinase powder efficiently clarified orange juice pulp similar to several microbial enzyme products. Commercial pilot scale biomass production of tobacco leaves expressing different pectinases showed that hydroponic growth at Fraunhofer yielded 10 times lower leaf biomass per plant than soil‐grown plants in the greenhouse. Pectinase enzyme yield from the greenhouse plants was double that of Fraunhofer. Thus, this leaf‐production platform offers a novel, low‐cost approach for enzyme production by elimination of fermentation, purification, concentration, formulation and cold chain.     

High‐yield secretion of recombinant proteins expressed in tobacco cell culture with a designer glycopeptide tag: Process development

Low‐yield protein production remains the most significant economic hurdle with plant cell culture technology. Fusions of recombinant proteins with hydroxyproline‐O‐glycosylated designer glycopeptide tags have consistently boosted secreted protein yields. This prompted us to study the process development of this technology aiming to achieve productivity levels necessary for commercial viability. We used a tobacco BY‐2 cell culture expressing EGFP as fusion with a glycopeptide tag comprised of 32 repeat of ”Ser‐Pro“ dipeptide, or (SP)₃₂, to study cell growth and protein secretion, culture scale‐up, and establishment of perfusion cultures for continuous production. The BY‐2 cells accumulated low levels of cell biomass (～7.5 g DW/L) in Schenk & Hildebrandt medium, but secreted high yields of (SP)₃₂‐tagged EGFP (125 mg/L). Protein productivity of the cell culture has been stable for 6.0 years. The BY‐2 cells cultured in a 5‐L bioreactor similarly produced high secreted protein yield at 131 mg/L. Successful operation of a cell perfusion culture for 30 days was achieved under the perfusion rate of 0.25 and 0.5 day^?1, generating a protein volumetric productivity of 17.6 and 28.9 mg/day/L, respectively. This research demonstrates the great potential of the designer glycopeptide technology for use in commercial production of valuable proteins with plant cell cultures.     

THE LEBANESE PHYSICIAN: A PUBLIC'S VIEWPOINT

A physician's lack of humanity is a general complaint in public surveys. The physician‐patient relationship is viewed by the public as being reduced to a business relationship where the patient feels that she is merely a ‘client’ and the physician a healthcare ‘practitioner’ instead of a ‘care giver’. This public perception is not a phenomenon that is peculiar to Lebanon. Yet, the problem has been increasing over the years to the extent that patients feel that physicians are becoming inhumane and business oriented. While this might not characterize all physicians of the 21^st century, this might be true of at least some. Responses were collected from a study that was undertaken based on a questionnaire distributed to a pool of 650 participants from different geographical areas and different social and educational backgrounds in Lebanon. Participants were all older than18 years and mentally competent. None were physicians. The questionnaire was open‐ended and initially piloted among a random sample. The physician traits most desired by the public were found to be: moral traits (41%), interpersonal traits (36%), scientific traits (19%) and other (4%). The most unwanted traits/behaviours were a lack of interpersonal traits (57%), a lack of moral traits (40%) and a lack of scientific skills (3%). The physician‐patient relationship was perceived, in general, as being a flawed one. What can be done to remedy the image of the Lebanese physician that has been projected in the minds of the patients and the public at large? Nine major recommendations are presented.     

Microalgal industry in China: challenges and prospects

Over the past 15 years, China has become the major producer of microalgal biomass in the world. Spirulina (Arthrospira) is the largest microalgal product by tonnage and value, followed by Chlorella, Dunaliella, and Haematococcus, the four main microalgae grown commercially. China’s production is estimated at about two-thirds of global microalgae biomass of which roughly 90 % is sold for human consumption as human nutritional products (‘nutraceuticals’), with smaller markets in animal feeds mainly for marine aquaculture. Research is also ongoing in China, as in the rest of the world, for other high-value as well as commodity microalgal products, from pharmaceuticals to biofuels and CO₂ capture and utilization. This paper briefly reviews the main challenges and potential solutions for expanding commercial microalgae production in China and the markets for microalgae products. The Chinese Microalgae Industry Alliance (CMIA), a network founded by Chinese microalgae researchers and commercial enterprises, supports this industry by promoting improved safety and quality standards, and advancement of technologies that can innovate and increase the markets for microalgal products. Microalgae are a growing source of human nutritional products and could become a future source of sustainable commodities, from foods and feeds, to, possibly, fuels and fertilizers.