An Integrated Approach to Crop Genetic Improvement

The balance between the supply and demand of the major food crops is fragile,fueling concerns for long-term global food security.The rising population,increasing wealth and a proliferation of nonfood uses(e.g.bioenergy) has led to growing demands on agriculture,while increased production is limited by greater urbanization,and the degradation of land.Furthermore,global climate change with increasing temperatures and lower,more erratic rainfall is projected to decrease agricultural yields.There is a predicted need to increase food production by at least 70% by 2050 and therefore an urgent need to develop novel and integrated approaches,incorporating high-throughput phenotyping that will both increaseproduction per unit area and simultaneously improve the resource use efficiency of crops.Yield potential,yield stability,nutrient and water use are all complex multigenic traits and while there is genetic variability,their complexity makes such traits difficult to breed for directly.Nevertheless molecular plant breeding has the potential to deliver substantial improvements,once the component traits and the genes underlying these traits have been identified.In addition,interactions between the individual traits must also be taken into account,a demand that is difficult to fulfill with traditional screening approaches.Identified traits will be incorporated into new cultivars using conventional or biotechnological tools.In order to better understand the relationship between genotype,component traits,and environment over time,a multidisciplinary approach must be adopted to both understand the underlying processes and identify candidate genes,QTLs and traits that can be used to develop improved crops.     

A Statistical Approach to Identify Ancient Template DNA

One of the key problems in the study of ancient DNA is that of authenticating sequences obtained from PCR amplifications of highly degraded samples. Contamination of ancient samples and postmortem damage to endogenous DNA templates are the major obstacles facing researchers in this task. In particular, the authentication of sequences obtained from ancient human remains is thought by many to be rather challenging. We propose a novel approach, based on the c statistic, that can be employed to help identify the sequence motif of an endogenous template, based on a sample of sequences that reflect the nucleotide composition of individual template molecules obtained from ancient tissues (such as cloned products from a PCR amplification). The c statistic exploits as information the most common form of postmortem damage observed among clone sequences in ancient DNA studies, namely, lesion-induced substitutions caused by cytosine deamination events. Analyses of simulated sets of templates with miscoding lesions and real sets of clone sequences from the literature indicate that the c-based approach is highly effective in identifying endogenous sequence motifs, even when they are not present among the sampled clones. The proposed approach is likely to be of general use to researchers working with DNA from ancient tissues, particularly from human remains, where authentication of results has been most challenging. [Reviewing Editor: Dr. Magnus Nordborg]     

Ecosystem Approach to Community Health Planning in Ghana

An ecosystem approach to community health seeks to investigate human health concerns from an ecological perspective. The goal is to improve the health of community members by instituting sustainable ecosystem management strategies that will preserve the health of both the ecosystem and its inhabitants. This article reports on a study in a rural community in Ghana that employed an ecosystem approach to community health planning. Adopting a participatory action research approach, a research team comprised of representatives from various local government departments collaborated with local residents to identify major community health problems, their underlying causes, and possible intervention strategies. The findings illustrate the usefulness of the ecosystem approach to mobilizing community-based interventions to addressing environmental determinants of human health.     

A Statistical Approach to Provide Individualized Privacy for Surveys

In this paper we propose an instrument for collecting sensitive data that allows for each participant to customize the amount of information that she is comfortable revealing. Current methods adopt a uniform approach where all subjects are afforded the same privacy guarantees; however, privacy is a highly subjective property with intermediate points between total disclosure and non-disclosure: each respondent has a different criterion regarding the sensitivity of a particular topic. The method we propose empowers respondents in this respect while still allowing for the discovery of interesting findings through the application of well-known inferential procedures.     

A Statistical Approach to the Definition of Bacterial Species

A statistical method is proposed for recognition of a bacterial species or for differentiation of two groups of bacterial strains. Comparison between two groups is done by the “t”-test. When the mean S-values (similarity value) of two groups are A and B, and the mean S-value for all possible combinations between strains of both groups is S, a condition necessary for defining the two groups as different species is to demonstrate the existence of equations A > S and B > S. Unless this condition is fulfilled, the two groups should be considered unseparable. The condition necessary for recognition of two groups as one species is to demonstrate the existence of equations A:= S and B = S. A few examples of the test were shown using the mycobacteria, and it was suggested that this statistical method is useful in the recognition of a species.     

Enhanced Weight Loss With Pramlintide/Metreleptin: An Integrated Neurohormonal Approach to Obesity Pharmacotherapy

The neurohormonal control of body weight involves a complex interplay between long‐term adiposity signals (e.g., leptin), and short‐term satiation signals (e.g., amylin). In diet‐induced obese (DIO) rodents, amylin/leptin combination treatment led to marked, synergistic, fat‐specific weight loss. To evaluate the weight‐lowering effect of combined amylin/leptin agonism (with pramlintide/metreleptin) in human obesity, a 24‐week, randomized, double‐blind, active‐drug‐controlled, proof‐of‐concept study was conducted in obese or overweight subjects (N = 177; 63% female; 39 ± 8 years; BMI 32.0 ± 2.1 kg/m²; 93.3 ± 13.2 kg; mean ± s.d.). After a 4‐week lead‐in period with pramlintide (180 µg b.i.d. for 2 weeks, 360 µg b.i.d. thereafter) and diet (40% calorie deficit), subjects achieving 2–8% weight loss were randomized 1:2:2 to 20 weeks of treatment with metreleptin (5 mg b.i.d.), pramlintide (360 µg b.i.d.), or pramlintide/metreleptin (360 µg/5 mg b.i.d.). Combination treatment with pramlintide/metreleptin led to significantly greater weight loss from enrollment to week 20 (?12.7 ± 0.9%; least squares mean ± s.e.) than treatment with pramlintide (?8.4 ± 0.9%; P < 0.001) or metreleptin (?8.2 ± 1.3%; P < 0.01) alone (evaluable, N = 93). The greater reduction in body weight was significant as early as week 4, and weight loss continued throughout the study, without evidence of a plateau. The most common adverse events with pramlintide/metreleptin were injection site events and nausea, which were mostly mild to moderate and decreased over time. These results support further development of pramlintide/metreleptin as a novel, integrated neurohormonal approach to obesity pharmacotherapy.     

Minireview: Nomenclature and Classification of Transmitter Receptors: an Integrated Approach

Abstract

This short assay is an attempt to rationalize an integrated approach to transmitter receptor nomenclature and classification based on three criteria: structural information (gene structure/amino acid sequence), operational¹ information (pharmacological/ ligand-defined/recognitory) and transductional information (receptor-effector coupling) are proposed to be used when considering the naming of existing or newly discovered receptors. It should be recognized that none of these criteria has primacy and that in an ideal situation, as much information as feasible on these three aspects should be collected before the naming of a receptor can be agreed upon.     

The Molecular Architecture of Native BBSome Obtained by an Integrated Structural Approach

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Planning for Restoration: A Decision Analysis Approach to Prioritization

Ecological restoration often relies on the use of expert opinion to make management decisions in the face of uncertainty. The quantification of expert opinion can be difficult, especially when more than one expert is consulted and experts are not in agreement. Decision analysis can provide a framework to systematically deconstruct a complex problem and provide greater objectivity to restoration decisions. We utilized decision analysis techniques to identify restoration objectives and to quantify expert opinions to prioritize restoration activities at 112 prairie openings in the Edge of Appalachia Preserve in southern Ohio, U.S.A. We first created an objectives hierarchy to model how decision‐makers decide which prairies to manage. We then determined how to measure each component of the hierarchy and sampled all prairies for percent woody cover, geology, indicator species index (an index of plant species richness), slope, aspect, and distance to nearest prairie. We modeled seven different experts’ preferences for managing prairies with varying values for each of these ecological measures. We then interviewed the same decision‐makers to determine relative weights for each component of the objectives hierarchy using trade‐off analysis. By combining the weights, preference relationships, and sampling data, we were able to rank each prairie and management unit based on its management priority. Experts had similar preferences except for the measure of distance to nearest prairie. We found that decision‐makers gave different weights to each of the different components of the hierarchy. Generally, experts weighted percent woody cover, indicator species index, and geology more highly than slope, aspect, and distance to nearest prairie. Despite these differences, priorities for management, once all factors were weighted and combined, were similar.     

A Risk-Based Approach to Management of Leachables Utilizing Statistical Analysis of Extractables

To incorporate quality by design concepts into the management of leachables, an emphasis is often put on understanding the extractable profile for the materials of construction for manufacturing disposables, container-closure, or delivery systems. Component manufacturing processes may also impact the extractable profile. An approach was developed to (1) identify critical components that may be sources of leachables, (2) enable an understanding of manufacturing process factors that affect extractable profiles, (3) determine if quantitative models can be developed that predict the effect of those key factors, and (4) evaluate the practical impact of the key factors on the product. A risk evaluation for an inhalation product identified injection molding as a key process. Designed experiments were performed to evaluate the impact of molding process parameters on the extractable profile from an ABS inhaler component. Statistical analysis of the resulting GC chromatographic profiles identified processing factors that were correlated with peak levels in the extractable profiles. The combination of statistically significant molding process parameters was different for different types of extractable compounds. ANOVA models were used to obtain optimal process settings and predict extractable levels for a selected number of compounds. The proposed paradigm may be applied to evaluate the impact of material composition and processing parameters on extractable profiles and utilized to manage product leachables early in the development process and throughout the product lifecycle.KEY WORDS: design of experiments, extractables, injection molding, leachables, process parameters, quality by design     

An Integrated Approach to Aggregate Control for Therapeutic Bispecific Antibodies Using an Improved Three Column Mab Platform-Like Purification Process

Single chain variable fragment-IgGs (scFv-IgG) are a class of bispecific antibodies consisting of two single chain variable fragments (scFv) that are fused to an intact IgG molecule. A common trend observed for expression of scFv-IgGs in mammalian cell culture is a higher level of aggregates (10%–30%) compared to mAbs, which results in lower purification yields in order to meet product quality targets. Furthermore, the high aggregate levels also pose robustness risks to a conventional mAb three column platform purification process which uses only the polishing steps (e.g., cation exchange chromatography [CEX]) for aggregate removal. Protein A chromatography with pH gradient elution, high performance tangential flow filtration (HP-TFF) and calcium phosphate precipitation were evaluated at the bench scale as means of introducing orthogonal aggregate removal capabilities into other aspects of the purification process. The two most promising process variants, namely Protein A pH gradient elution followed by calcium phosphate precipitation were evaluated at pilot scale, demonstrating comparable performance. Implementing Protein A chromatography with gradient elution and/or calcium phosphate precipitation removed a sufficient portion of the aggregate burden prior to the CEX polishing step, enabling CEX to be operated robustly under conditions favoring higher monomer yield. From starting aggregate levels ranging from 15% to 23% in the condition media, levels were reduced to between 2% and 3% at the end of the CEX step. The overall yield for the optimal process was 71%. Results of this work suggest an improved three-column mAb platform-like purification process for purification of high aggregate scFv-IgG bispecific antibodies is feasible. © 2018 The Authors. Biotechnology Progress published by Wiley Periodicals, Inc. on behalf of American Institute of Chemical Engineers. Biotechnol. Prog., 35: e2720, 2019     

Aminopeptidase Fingerprints, an Integrated Approach for Identification of Good Substrates and Optimal Inhibitors

Aminopeptidases process the N-terminal amino acids of target substrates by sequential cleavage of one residue at a time. They are found in all cell compartments of prokaryotes and eukaryotes, being implicated in the major proteolytic events of cell survival, defense, growth, and development. We present a new approach for the fast and reliable evaluation of the substrate specificity of individual aminopeptidases. Using solid phase chemistry with the 7-amino-4-carbamoylmethylcoumarin fluorophore, we have synthesized a library of 61 individual natural and unnatural amino acids substrates, chosen to cover a broad spectrum of the possible interactions in the S1 pocket of this type of protease. As proof of concept, we determined the substrate specificity of human, pig, and rat orthologs of aminopeptidase N (CD13), a highly conserved cell surface protease that inactivates enkephalins and other bioactive peptides. Our data reveal a large and hydrophobic character for the S1 pocket of aminopeptidase N that is conserved with aminopeptidase Ns. Our approach, which can be applied in principle to all aminopeptidases, yields useful information for the design of specific inhibitors, and more importantly, reveals a relationship between the kinetics of substrate hydrolysis and the kinetics of enzyme inhibition.     

A Statistical Approach to Biological Factors of Safety: Bending and Shearing inPsilotumAxes

A method is presented to calculate the factor of safety (basedon any criterion of failure) for populations of equivalent biologicalstructures that normally sustain mechanical loads (e.g. stemsof the same order of branching). The method uses the empiricallydetermined mean and standard deviation of the actual loads andthe load capabilities provided that both types of loads arenormally distributed in the population. The factor of safetySiscalculated from the formulaS