Life‐cycle and cost of goods assessment of fed‐batch and perfusion‐based manufacturing processes for mAbs

Life‐cycle assessment (LCA) is an environmental assessment tool that quantifies the environmental impact associated with a product or a process (e.g., water consumption, energy requirements, and solid waste generation). While LCA is a standard approach in many commercial industries, its application has not been exploited widely in the bioprocessing sector. To contribute toward the design of more cost‐efficient, robust and environmentally‐friendly manufacturing process for monoclonal antibodies (mAbs), a framework consisting of an LCA and economic analysis combined with a sensitivity analysis of manufacturing process parameters and a production scale‐up study is presented. The efficiency of the framework is demonstrated using a comparative study of the two most commonly used upstream configurations for mAb manufacture, namely fed‐batch (FB) and perfusion‐based processes. Results obtained by the framework are presented using a range of visualization tools, and indicate that a standard perfusion process (with a pooling duration of 4 days) has similar cost of goods than a FB process but a larger environmental footprint because it consumed 35% more water, demanded 17% more energy, and emitted 17% more CO₂ than the FB process. Water consumption was the most important impact category, especially when scaling‐up the processes, as energy was required to produce process water and water‐for‐injection, while CO₂ was emitted from energy generation. The sensitivity analysis revealed that the perfusion process can be made more environmentally‐friendly than the FB process if the pooling duration is extended to 8 days. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:1324–1335, 2016     

Life‐cycle variation,including female production by virgin females in Frankliniella occidentalis (Thysanoptera: Thripidae)

The influence of three different temperatures on developmental time and sex ratio was investigated in the bisexual Thysanoptera species Frankliniella occidentalis. Increasing temperatures decreased developmental time and induced a more female biased sex ratio. Remarkably, there are second instars with a prolonged developmental time requiring the same number of hours as the shortest developmental time from egg to adult. Arrhenotokous reproduction in this species is based on haplodiploidy, with virgin females producing male offspring exclusively. However, at all three temperatures tested, about 0.5% of offspring from unfertilized eggs were females. The presence of Wolbachia could not be detected in Western flower thrips and can be excluded as influencing reproduction in this species.     

A life‐cycle assessment model for zero emission neighborhoods

Buildings represent a critical piece of a low‐carbon future, and their long lifetime necessitates urgent adoption of state‐of‐the‐art performance standards to avoid significant lock‐in risk regarding long‐lasting technology solution choices. Buildings, mobility, and energy systems are closely linked, and assessing their nexus by aiming for Zero Emission Neighborhoods (ZENs) provides a unique chance to contribute to climate change mitigation. We conducted a life‐cycle assessment of a Norwegian ZEN and designed four scenarios to test the influence of the house size, household size, and energy used and produced in the buildings as well as mobility patterns. We ran our scenarios with different levels of decarbonization of the electricity mix over a period of 60 years. Our results show the importance of the operational phases of both the buildings and mobility in the neighborhood's construction, and its decline over time induced by the decarbonization of the electricity mix. At the neighborhood end‐of‐life, embodied emissions then become responsible for the majority of the emissions when the electricity mix is decarbonized. The choice of functional unit is decisive, and we thus argue for the use of a primary functional unit “per neighborhood,” and a second “per person.” The use of a “per m²” functional unit is misleading as it does not give credits to the precautionary use of floor area. To best mitigate climate change, climate‐positive behaviors should be combined with energy efficiency standards that incorporate embodied energy, and absolute threshold should be combined with behavioral changes.     

Life‐history traits of Tubastraea coccinea: Reproduction,development, and larval competence

The sun coral Tubastraea coccinea Lesson, 1829 (Dendrophylliidae) is a widely distributed shallow‐water scleractinian that has extended its range to non‐native habitats in recent decades. With its rapid spread, this coral is now one of the main invasive species in Brazil. Its high invasive capability is related to opportunistic characteristics, including several reproductive strategies that have allowed it to disperse rapidly and widely. To better understand the reproductive biology of T. coccinea and aid in developing management strategies for invaded areas, we investigated aspects of its reproductive performance and life cycle, including the effects of colony size, seawater temperature and salinity, and lunar periodicity on offspring production and larval metamorphosis competence. A total of 18,139 offspring were released in different developmental stages, mainly from the larger colonies, which also produced larvae with longer competence periods. The main reproductive peak occurred during the First Quarter and New Moon phases and was highest in water temperatures around 26°C. Together, these results help to explain the rapid expansion of T. coccinea into non‐native habitats such as the Caribbean and southwestern Atlantic, and will inform actions of the recent Brazilian National Plan for the prevention, eradication, control, and monitoring of sun corals.     

Effects of culture media, temperature, pH, and light on growth, sporulation, germination, and bioherbicidal efficacy of Phoma exigua, a potential biological control agent for salal (Gaultheria shallon)

In order to evaluate the potential use of Phoma exigua isolate PFC 2705 (PFC2705) as a biological control agent for salal (Gaultheria shallon), effect of cultural and environmental parameters on growth, conidia production, and pathogenicity of P. exigua were characterized in studies conducted under laboratory and greenhouse conditions. Within a range of 5-30°C, the optimum growth and germination temperature range was 20-25°C. The effect of pH on mycelial growth and conidial germination was not significant from pH 5 to 10. Fluorescent light significantly enhanced sporulation of the fungus on most agar media tested, yet was not necessary for growth. The type of culture media significantly affected mycelium growth, sporulation, and conidia germination. Age of mycelia used as inoculum affected the disease severity on salal. PFC2705 suppressed the growth of mature salal plant by inciting lesions on leaves, branch tips, and axillary buds and caused 56% death of the total biomass above ground. Characteristics such as easy inoculum production, wide range of growth environments, and high infectivity on salal increased the potential of P. exigua as a biocontrol agent for management of salal.     

RopGEF2 is involved in ABA‐suppression of seed germination and post‐germination growth of Arabidopsis

The involvement of Rho of Plants (ROP) GTPases in abscisic acid (ABA) signalling in Arabidopsis has been demonstrated in many studies. However, the roles of RopGEFs (Rop guanine nucleotide exchange factors), which modulate ROP activities in ABA signalling, are poorly understood. Here, we demonstrate that RopGEF2 may play a negative role in ABA‐suppressed seed germination and post‐germination growth. We show that disruption of RopGEF2 enhances sensitivity to exogenous ABA in seed germination assays and that RopGEF2pro‐GUS is mainly expressed in developing embryos and germinating seeds. Interestingly, YFP‐RopGEF2 is located in both the cytoplasmic region and in mitochondria. Notably, the PRONE2 (plant‐specific ROP nucleotide exchanger 2) domain of RopGEF2 is detected in mitochondria, whereas the N‐terminus of RopGEF2 is shown to be in the cytosol. After ABA treatment, degradation of RopGEF2 is triggered in the cytosol through the ubiquitin‐26S proteasome system. The binding of RopGEF2 to ROP2, ROP6 or ROP10, which has been demonstrated to be involved in ABA signalling, not only alters the localization of RopGEF2 but also enables RopGEF2 to escape degradation in the cell. Thus, in this study, we deduce a sophisticated mechanism of ABA‐mediated RopGEF2‐ROP signalling, which potentially implicates the inactivation of ROPs in responsiveness to ABA.     

Design and calibration of a simplified river-water quality model including heterotrophic bacteria and chemical parameters

Abstract The results obtained in the design and calibration of a deterministic water quality model from the Llobregat River (Spain) are presented. The water quality indicators studied are dissolved oxygen, biochemical oxygen demand, ammonia, nitrite, nitrate, phosphate, and heterotrophic bacteria. The factors which describe the rates of the transformation processes are of two types: first-order and Michaelis-Menten kinetics. A decomposition process is applied to the model for calibration. The inclusion in the model of heterotrophic bacteria shows good predictive capacity to describe the behaviour of the biological processes in the river. The proposed model gives a correct evaluation of the indicators studied, and may be used in water quality management.     

Life‐history strategy,with ctenidial and pallial larval brooding,of the troglodytic ‘living fossil’ Congeria kusceri (Bivalvia: Dreissenidae) from the subterranean Dinaric Alpine karst of Croatia

Among freshwater bivalves, the brooding of embryos and larvae within the maternal ctenidia is well known. Exceptions to this generalization are the non‐brooding freshwater and estuarine species of Dreissena and Mytilopsis, respectively. It was reported that the freshwater troglodytic cousin, Congeria kusceri Bole, 1962, of these dreissenids does not brood either. It is herein demonstrated that C. kusceri undergoes one reproductive cycle each year. Sexes are separate, with an early male and later female bias. A small percentage (2.14%) of individuals is hermaphroditic. The gonads mature over summer from May to November. Spawning commences in September, when females release mature oocytes into their ctenidia and inhale sperm from mature males. Here the oocytes are fertilized, and develop within interfilamentary marsupia. Ctenidial tissues glandularize, and may provide a source of maternal nutrition for the embryos. At the late prodissoconch‐1 or prodissoconch‐2 stage (PR2, ~220 μm), larvae are released into the infrabranchial chamber via a birth channel along the outer edge of the ventral marginal food groove of both inner demibranchs. Here, they are brooded further in mantle pouches located beneath the inhalant siphon. Subsequently, after the PR2 stage (nepioconch/dissoconch), they are released from the inhalant siphon and assume an independent life as crawling juveniles. Such juveniles may be found amongst clusters of adults. Not only is C. kusceri unique amongst the Dreissenidae in possessing the capacity to brood internally fertilized ova, but it is also exceptional amongst the Bivalvia in possessing the described methods of brooding and birth. Explanations for both lie in its troglodytic lifestyle, decadal length longevity and habitat: that of byssal attachment to the hard surfaces of underground freshwater rivers, caves, pits, and sinkholes in the Tethyan arc of the Dinaric karst. Internal fertilization of a few large yolky eggs, lecithotrophic larvae, ctenidial brooding, and secondary pallial parental care represent relatively recent, Late Miocene, evolutionary adaptations from a Tethyan lentic ancestor.     

Defining the importance of including transient ecosystem responses to simulate C‐cycle dynamics in a global change model

The ability of plant species to migrate is one of the critical issues in assessing accurately the future response of the terrestrial biosphere to climate change. This ability is confined by both natural and human‐induced changes in land cover. In this paper we present land‐cover and Carbon (C) cycle models designed to simulate the biospheric consequences of different types of land‐cover changes. These models, imbedded in the larger integrated assessment model IMAGE 2, were used to demonstrate the importance of considering spatial aspects for global C‐cycle modelling. A gradual‐migration, an unlimited‐migration and a no‐migration case were compared to show the range of possible consequences. Major differences between these cases were simulated for land‐cover patterns and the carbon budget. A large geographical variation in the biospheric response was also simulated. The strongest response was simulated in high‐latitude regions, especially for the migration cases in which land‐cover changes were permitted. In low‐latitudes regions the differences between the migration cases were smaller, mainly due to the effects of land‐use changes. The geographical variation among, and the different responses, the migration cases clearly demonstrate how essential it is to assess biospheric responses to climate change and land use simultaneously. Moreover, it also shows the urgent need for enhanced understanding of spatial and temporal dynamics of the biospheric responses.     

The Ethics of Continued Life‐Sustaining Treatment for those Diagnosed as Brain‐dead

Given the long‐standing controversy about whether the brain‐dead should be considered alive in an irreversible coma or dead despite displaying apparent signs of life, the ethical and policy issues posed when family members insist on continued treatment are not as simple as commentators have claimed. In this article, we consider the kind of policy that should be adopted to manage a family's insistence that their brain‐dead loved one continues to receive supportive care. We argue that while it would be ethically inappropriate to continue to devote scarce acute care resources to such patients in a hospital setting, it may not be ethically inappropriate for patients to receive these resources in certain other settings. Thus, if a family insists on continuing to care for their brain‐dead loved at their home, we should not, from a policy perspective, interfere with the family's wishes. We also argue that healthcare professionals should make some effort to facilitate the transfer of brain‐dead patients to these other settings when families insist on continued treatment despite being informed about the lack of any potential for recovery of consciousness. Our arguments are strengthened by the fact that patients in a persistent vegetative state, who, when correctly diagnosed, also have no potential for recovery of consciousness, are routinely transferred from hospitals to nursing homes or long‐term care facilities where they continue to be ventilated, tube fed and to receive other supportive care. We also briefly explore the question of who should be responsible for the costs of such treatment at the long‐term care facility.     

Bet‐hedging and germination in the Australian arid zone shrub Acacia ligulata

The diaspore of the Australian arid zone shrub Acacia ligulata is dispersed by birds and ants. To investigate the benefits of providing a dispersal structure attractive to both groups, we compared the germination response and viability of seeds eaten by birds, handled by ants or collected from trees to simulated precursors of germination: scarification, fire and rainfall were simulated. Seed germination and viability were related to the degree of preheating disturbance to the seed coat. Heating increased the germinability of seeds not scarified or eaten by birds. In the absence of heating, ingestion by birds increased germinability. Heating increased the mortality of seeds. Our results suggest that ingestion of seeds by birds may break seed dormancy and hence enable some seeds to germinate soon after dispersal. Alternatively, seeds not eaten by birds are likely to remain dormant until sufficiently scarified by soil or stimulated by fire. Consequently, in areas such as the Simpson Desert, A. ligulata may be able to use a range of seedling establishment ‘windows’ provided by monsoon rains, post‐fire environments and unseasonal winter rains, and also spread the risk of unsuccessful seedling establishment by retaining dormant seeds in the seedbank.     

A review of thermochemical upgrading of pyrolysis bio‐oil: Techno‐economic analysis,life cycle assessment,and technology readiness

Technologies for upgrading fast pyrolysis bio‐oil to drop‐in fuels and coproducts are under development and show promise for decarbonizing energy supply for transportation and chemicals markets. The successful commercialization of these fuels and the technologies deployed to produce them depend on production costs, scalability, and yield. To meet environmental regulations, pyrolysis‐based biofuels need to adhere to life cycle greenhouse gas intensity standards relative to their petroleum‐based counterparts. We review literature on fast pyrolysis bio‐oil upgrading and explore key metrics that influence their commercial viability through life cycle assessment (LCA) and techno‐economic analysis (TEA) methods together with technology readiness level (TRL) evaluation. We investigate the trade‐offs among economic, environmental, and technological metrics derived from these methods for individual technologies as a means of understanding their nearness to commercialization. Although the technologies reviewed have not attained commercial investment, some have been pilot tested. Predicting the projected performance at scale‐up through models can, with industrial experience, guide decision‐making to competitively meet energy policy goals. LCA and TEA methods that ensure consistent and reproducible models at a given TRL are needed to compare alternative technologies. This study highlights the importance of integrated analysis of multiple economic, environmental, and technological metrics for understanding performance prospects and barriers among early stage technologies.     

CO2 emissions from land‐use change affected more by nitrogen cycle,than by the choice of land‐cover data

The high uncertainty in land‐based CO₂ fluxes estimates is thought to be mainly due to uncertainty in not only quantifying historical changes among forests, croplands, and grassland, but also due to different processes included in calculation methods. Inclusion of a nitrogen (N) cycle in models is fairly recent and strongly affects carbon (C) fluxes. In this study, for the first time, we use a model with C and N dynamics with three distinct historical reconstructions of land‐use and land‐use change (LULUC) to quantify LULUC emissions and uncertainty that includes the integrated effects of not only climate and CO₂ but also N. The modeled global average emissions including N dynamics for the 1980s, 1990s, and 2000–2005 were 1.8 ± 0.2, 1.7 ± 0.2, and 1.4 ± 0.2 GtC yr^?1, respectively, (mean and range across LULUC data sets). The emissions from tropics were 0.8 ± 0.2, 0.8 ± 0.2, and 0.7 ± 0.3 GtC yr^?1, and the non tropics were 1.1 ± 0.5, 0.9 ± 0.2, and 0.7 ± 0.1 GtC yr^?1. Compared to previous studies that did not include N dynamics, modeled net LULUC emissions were higher, particularly in the non tropics. In the model, N limitation reduces regrowth rates of vegetation in temperate areas resulting in higher net emissions. Our results indicate that exclusion of N dynamics leads to an underestimation of LULUC emissions by around 70% in the non tropics, 10% in the tropics, and 40% globally in the 1990s. The differences due to inclusion/exclusion of the N cycle of 0.1 GtC yr^?1 in the tropics, 0.6 GtC yr^?1 in the non tropics, and 0.7 GtC yr^?1 globally (mean across land‐cover data sets) in the 1990s were greater than differences due to the land‐cover data in the non tropics and globally (0.2 GtC yr^?1). While land‐cover information is improving with satellite and inventory data, this study indicates the importance of accounting for different processes, in particular the N cycle.     

Orchid-associated bacteria produce indole-3-acetic acid,promote seed germination,and increase their microbial yield in response to exogenous auxin

Germination of orchid seeds is a complex process. In this paper we focus on interactions between the host-plant and its bacterial partners via indole-3-acetic acid (IAA). Originally isolated from the roots of the epiphytic orchid Dendrobium moschatum, the strains of Rhizobium, Microbacterium, Sphingomonas, and Mycobacterium genera were among the most active IAA producers. Addition of exogenous tryptophan significantly enhanced auxin formation both in mineral and complex media. The presence of IAA and indole-3-acetaldehyde was confirmed by HPLC. Indole-3-pyruvic and indole-3-lactic acids were also detected in supernatants of culture filtrates of Sphingomonas sp., Rhizobium sp., and Microbacterium sp., while indole-3-acetamide was identified only in Mycobacterium sp. Some concentration- and strain-dependent effects of exogenous IAA on bacterial development were also established. Treatment of the cultures with 10 and 100 μg/ml of auxin resulted in an increase in microbial yield. None of the investigated strains was able to utilize IAA as a source of carbon and energy. Furthermore, inoculation of D. moschatum seeds with Sphingomonas sp. and Mycobacterium sp. resulted in considerable enhancement of orchid seeds germination. This growth-promoting activity was observed in the absence of any plant growth stimulators or mycorrhizal fungi, usually required for orchid germination.