Antibody production using a ciliate generates unusual antibody glycoforms displaying enhanced cell-killing activity

Antibody glycosylation is a key parameter in the optimization of antibody therapeutics. Here, we describe the production of the anti-cancer monoclonal antibody rituximab in the unicellular ciliate, Tetrahymena thermophila. The resulting antibody demonstrated enhanced antibody-dependent cell-mediated cytotoxicity, which we attribute to unusual N-linked glycosylation. Detailed chromatographic and mass spectrometric analysis revealed afucosylated, oligomannose-type glycans, which, as a whole, displayed isomeric structures that deviate from the typical human counterparts, but whose branches were equivalent to fragments of metabolic intermediates observed in human glycoproteins. From the analysis of deposited crystal structures, we predict that the ciliate glycans adopt protein-carbohydrate interactions with the Fc domain that closely mimic those of native complex-type glycans. In addition, terminal glucose structures were identified that match biosynthetic precursors of human glycosylation. Our results suggest that ciliate-based expression systems offer a route to large-scale production of monoclonal antibodies exhibiting glycosylation that imparts enhanced cell killing activity.     

Effects of Ration and Temperature on Reproduction of a Semelparous and an Iteroparous Triclad

Summary

This paper compares aspects of the reproduction of a semelparous triclad, Dendrocoelum lacteum, with those of an iteroparous triclad, Dugesia polychroa. Cocoon output was less sensitive to ration and possibly temperature for the former as compared with the latter species. Cocoons of D. lacteum were on average bigger and contained more hatchlings than those of D. polychroa. The sizes of hatchlings did not differ between species, but for D. lacteum only, the hatchlings emerging from late cocoons were significantly larger, and less numerous, than those emerging from earlier cocoons. Possible reasons for these differences are considered.     

Applying Weight-of-Evidence in Retrospective Ecological Risk Assessment When Quantitative Data Are Limited

Retrospective ecological risk assessment attempts to identify likely causal agents to explain adverse effects that have occurred in ecological targets. It can never be decisive since it is post hoc and usually based on limited evidence that is rarely very quantitative. It can, nevertheless, be made more transparent, systematic, and logical, and less subjective. Based on human health epidemiological criteria we develop an approach that moves from systematic consideration of seven basic questions to assigning a likelihood of involvement of putative agents. The questions are: 1. Is there evidence that the target is or has been exposed to the agent? 2. Is there evidence for correlation between adverse effects in the target and exposure to the agent either in time or in space? 3. Do the measured or predicted environmental concentrations exceed quality criteria for water, sediment or body burden? 4. Have the results from controlled experiments in the field or laboratory led to the same effect? 5. Has removal of the agent led to amelioration of effects in the target? 6. Is there an effect in the target known to be specifically caused by exposure to the agent? 7. Does the proposed causal relationship make sense logically and scientifically? We identify 15 common scenarios of answers to the questions and illustrate the approach by reference to three real-world case studies (decline in benthos in a tropical marine bay, decline in fisheries in a temperate sea, decline in marine mollusc populations). The primary challenge in retrospective risk assessment is to make best use of the available evidence to develop rational management strategies and/or guide additional analyses to gain further evidence about likely agents as causes of observed harm.     

Species Sensitivity Distributions Revisited: A Critical Appraisal

We revisit the assumptions associated with the derivation and application of species sensitivity distributions (SSDs). Our questions are (1) Do SSDs clarify or obscure the setting of ecological effects thresholds for risk assessment? and (2) Do SSDs reduce or introduce uncertainty into risk assessment? Our conclusions are that if we could determine a community sensitivity distribution, this would provide a better estimate of an ecologically relevant effects threshold and therefore be an improvement for risk assessment. However, the distributions generated are typically based on haphazard collections of species and endpoints and by adjusting these to reflect more realistic trophic structures we show that effects thresholds can be shifted but in a direction and to an extent that is not predictable. Despite claims that the SSD approach uses all available data to assess effects, we demonstrate that in certain frequently used applications only a small fraction of the species going into the SSD determine the effects threshold. If the SSD approach is to lead to better risk assessments, improvements are needed in how the theory is put into practice. This requires careful definition of the risk assessment targets and of the species and endpoints selected for use in generating SSDs.     

The Daphnia bioassay: a critique

Daphnia magna is used widely as a standard ecotoxicological indicator organism, and protocols exist for its use in assessing the toxicity of substances under acute and chronic experimental conditions. Problems exist in repeatability of such bioassays between laboratories. Sources of variation are identified using a simple quantitative genetics model. Presenting specific examples, we conclude that these problems are tractable, but only if the genotype and culture conditions prior to and during tests are strictly controlled.     

Importance of fungi in the diet of Gammarus pulex and Asellus aquaticus I: feeding strategies

The importance of fungi in the trophic biology of the freshwater detritivores Gammarus pulex and Asellus aquaticus was investigated. Inspection of leaves used in feeding trials indicated that whereas A. aquaticus scrapes at the leaf surface, G. pulex bites through the leaf material. Both species discriminated between fungal mycelia, fungally colonized and uncolonized leaf material but, although A. aquaticus selectively consumed fungal mycelia, G. pulex fed preferentially on leaf material. Fungi appear to be an important food source for A. aquaticus and selection of food material was positively correlated with fungal biomass. In contrast, for G. pulex, fungi appear to be more important as modifiers of leaf material. However, no significant correlations were found between food preference and any of the leaf modifications measured.     

The application of bioassays in the resolution of environmental problems; past,present and future

Literature on bioassays for freshwater systems has been reviewed (between 1979 and 1987) and classified into studies concerned with prediction and assessment and, within these categories, into studies concerned with single- and multi-species bioassays. Changing trends in the response criteria and types of organisms used in the predictive tests are judged against results from a similar review carried out in 1979. This leads to the conclusion that though there may have been changes in detail, bioassays have remained surprisingly unchanged in general features over this time. The relative merits of, and relationship between single- and multi-species studies for both predicting and assessing the biological impact of toxicants are discussed. The conclusion is that some bioassays have more severe problems than others, but a concern with all of them is that responses observed in particular systems may not be relevant in general. The possibility of developing a general theoretical infrastructure for bioassays that addresses this problem is considered.     

Adding Value to Ecological Risk Assessment with Population Modeling

Current measures used to estimate the risks of toxic chemicals are not relevant to the goals of the environmental protection process, and thus ecological risk assessment (ERA) is not used as extensively as it should be as a basis for cost-effective management of environmental resources. Appropriate population models can provide a powerful basis for expressing ecological risks that better inform the environmental management process and thus that are more likely to be used by managers. Here we provide at least five reasons why population modeling should play an important role in bridging the gap between what we measure and what we want to protect. We then describe six actions needed for its implementation into management-relevant ERA.