On Apples,Oranges and Camels

Abstract

Changing Schools through the Arts: How to Build on the Power of an Idea. New York: ACA Books, 1990, 284 pages. $14.95 paperback.     

Fold-Change Detection in a Whole-Pathway Model of Escherichia coli chemotaxis

There has been recent interest in sensory systems that are able to display a response which is proportional to a fold change in stimulus concentration, a feature referred to as fold-change detection (FCD). Here, we demonstrate FCD in a recent whole-pathway mathematical model of Escherichia coli chemotaxis. FCD is shown to hold for each protein in the signalling cascade and to be robust to kinetic rate and protein concentration variation. Using a sensitivity analysis, we find that only variations in the number of receptors within a signalling team lead to the model not exhibiting FCD. We also discuss the ability of a cell with multiple receptor types to display FCD and explain how a particular receptor configuration may be used to elucidate the two experimentally determined regimes of FCD behaviour. All findings are discussed in respect of the experimental literature.     

Logarithmic and Power Law Input-Output Relations in Sensory Systems with Fold-Change Detection

Two central biophysical laws describe sensory responses to input signals. One is a logarithmic relationship between input and output, and the other is a power law relationship. These laws are sometimes called the Weber-Fechner law and the Stevens power law, respectively. The two laws are found in a wide variety of human sensory systems including hearing, vision, taste, and weight perception; they also occur in the responses of cells to stimuli. However the mechanistic origin of these laws is not fully understood. To address this, we consider a class of biological circuits exhibiting a property called fold-change detection (FCD). In these circuits the response dynamics depend only on the relative change in input signal and not its absolute level, a property which applies to many physiological and cellular sensory systems. We show analytically that by changing a single parameter in the FCD circuits, both logarithmic and power-law relationships emerge; these laws are modified versions of the Weber-Fechner and Stevens laws. The parameter that determines which law is found is the steepness (effective Hill coefficient) of the effect of the internal variable on the output. This finding applies to major circuit architectures found in biological systems, including the incoherent feed-forward loop and nonlinear integral feedback loops. Therefore, if one measures the response to different fold changes in input signal and observes a logarithmic or power law, the present theory can be used to rule out certain FCD mechanisms, and to predict their cooperativity parameter. We demonstrate this approach using data from eukaryotic chemotaxis signaling.     

Fiber-Optic Microarray for Simultaneous Detection of Multiple Harmful Algal Bloom Species

Harmful algal blooms (HABs) are a serious threat to coastal resources, causing a variety of impacts on public health, regional economies, and ecosystems. Plankton analysis is a valuable component of many HAB monitoring and research programs, but the diversity of plankton poses a problem in discriminating toxic from nontoxic species using conventional detection methods. Here we describe a sensitive and specific sandwich hybridization assay that combines fiber-optic microarrays with oligonucleotide probes to detect and enumerate the HAB species Alexandrium fundyense, Alexandrium ostenfeldii, and Pseudo-nitzschia australis. Microarrays were prepared by loading oligonucleotide probe-coupled microspheres (diameter, 3 μm) onto the distal ends of chemically etched imaging fiber bundles. Hybridization of target rRNA from HAB cells to immobilized probes on the microspheres was visualized using Cy3-labeled secondary probes in a sandwich-type assay format. We applied these microarrays to the detection and enumeration of HAB cells in both cultured and field samples. Our study demonstrated a detection limit of approximately 5 cells for all three target organisms within 45 min, without a separate amplification step, in both sample types. We also developed a multiplexed microarray to detect the three HAB species simultaneously, which successfully detected the target organisms, alone and in combination, without cross-reactivity. Our study suggests that fiber-optic microarrays can be used for rapid and sensitive detection and potential enumeration of HAB species in the environment.     

Beyond Two-Cell Networks: Experimental Measurement of Neuronal Responses to Multiple Synaptic Inputs

Oscillations of large populations of neurons are thought to be important in the normal functioning of the brain. We have used phase response curve (PRC) methods to characterize the dynamics of single neurons and predict population dynamics. Our past experimental work was limited to special circumstances (e.g., 2-cell networks of periodically firing neurons). Here, we explore the feasibility of extending our methods to predict the synchronization properties of stellate cells (SCs) in the rat entorhinal cortex under broader conditions. In particular, we test the hypothesis that PRCs in SCs scale linearly with changes in synaptic amplitude, and measure how well responses to Poisson process-driven inputs can be predicted in terms of PRCs. Although we see nonlinear responses to excitatory and inhibitory inputs, we find that models based on weak coupling account for scaling and Poisson process-driven inputs reasonably accurately.     

Stomata Prioritize Their Responses to Multiple Biotic and Abiotic Signal Inputs

Stomata are microscopic pores in leaf epidermis that regulate gas exchange between plants and the environment. Being natural openings on the leaf surface, stomata also serve as ports for the invasion of foliar pathogenic bacteria. Each stomatal pore is enclosed by a pair of guard cells that are able to sense a wide spectrum of biotic and abiotic stresses and respond by precisely adjusting the pore width. However, it is not clear whether stomatal responses to simultaneously imposed biotic and abiotic signals are mutually dependent on each other. Here we show that a genetically engineered Escherichia coli strain DH5α could trigger stomatal closure in Vicia faba, an innate immune response that might depend on NADPH oxidase-mediated ROS burst. DH5α-induced stomatal closure could be abolished or disguised under certain environmental conditions like low [CO₂], darkness, and drought, etc. Foliar spraying of high concentrations of ABA could reduce stomatal aperture in high humidity-treated faba bean plants. Consistently, the aggressive multiplication of DH5α bacteria in Vicia faba leaves under high humidity could be alleviated by exogenous application of ABA. Our data suggest that a successful colonization of bacteria on the leaf surface is correlated with stomatal aperture regulation by a specific set of environmental factors.     

Resolving Cell Population Heterogeneity: Real-Time PCR for Simultaneous Multiplexed Gene Detection in Multiple Single-Cell Samples

Decoding the complexity of multicellular organisms requires analytical procedures to overcome the limitations of averaged measurements of cell populations, which obscure inherent cell-cell heterogeneity and restrict the ability to distinguish between the responses of individual cells within a sample. For example, defining the timing, magnitude and the coordination of cytokine responses in single cells is critical for understanding the development of effective immunity. While approaches to measure gene expression from single cells have been reported, the absolute performance of these techniques has been difficult to assess, which likely has limited their wider application. We describe a straightforward method for simultaneously measuring the expression of multiple genes in a multitude of single-cell samples using flow cytometry, parallel cDNA synthesis, and quantification by real-time PCR. We thoroughly assess the performance of the technique using mRNA and DNA standards and cell samples, and demonstrate a detection sensitivity of ∼30 mRNA molecules per cell, and a fractional error of 15%. Using this method, we expose unexpected heterogeneity in the expression of 5 immune-related genes in sets of single macrophages activated by different microbial stimuli. Further, our analyses reveal that the expression of one ‘pro-inflammatory’ cytokine is not predictive of the expression of another ‘pro-inflammatory’ cytokine within the same cell. These findings demonstrate that single-cell approaches are essential for studying coordinated gene expression in cell populations, and this generic and easy-to-use quantitative method is applicable in other areas in biology aimed at understanding the regulation of cellular responses.     

Anatomical Organization of Multiple Modulatory Inputs in a Rhythmic Motor System

In rhythmic motor systems, descending projection neuron inputs elicit distinct outputs from their target central pattern generator (CPG) circuits. Projection neuron activity is regulated by sensory inputs and inputs from other regions of the nervous system, relaying information about the current status of an organism. To gain insight into the organization of multiple inputs targeting a projection neuron, we used the identified neuron MCN1 in the stomatogastric nervous system of the crab, Cancer borealis. MCN1 originates in the commissural ganglion and projects to the stomatogastric ganglion (STG). MCN1 activity is differentially regulated by multiple inputs including neuroendocrine (POC) and proprioceptive (GPR) neurons, to elicit distinct outputs from CPG circuits in the STG. We asked whether these defined inputs are compact and spatially segregated or dispersed and overlapping relative to their target projection neuron. Immunocytochemical labeling, intracellular dye injection and three-dimensional (3D) confocal microscopy revealed overlap of MCN1 neurites and POC and GPR terminals. The POC neuron terminals form a defined neuroendocrine organ (anterior commissural organ: ACO) that utilizes peptidergic paracrine signaling to act on MCN1. The MCN1 arborization consistently coincided with the ACO structure, despite morphological variation between preparations. Contrary to a previous 2D study, our 3D analysis revealed that GPR axons did not terminate in a compact bundle, but arborized more extensively near MCN1, arguing against sparse connectivity of GPR onto MCN1. Consistent innervation patterns suggest that integration of the sensory GPR and peptidergic POC inputs occur through more distributed and more tightly constrained anatomical interactions with their common modulatory projection neuron target than anticipated.     

Comparing Simultaneous and Pointwise Confidence Intervals for Hydrological Processes

Distribution function estimation of the random variable of river flow is an important problem in hydrology. This issue is directly related to quantile estimation, and consequently to return level prediction. The estimation process can be complemented with the construction of confidence intervals (CIs) to perform a probabilistic assessment of the different variables and/or estimated functions. In this work, several methods for constructing CIs using bootstrap techniques, and parametric and nonparametric procedures in the estimation process are studied and compared. In the case that the target is the joint estimation of a vector of values, some new corrections to obtain joint coverage probabilities closer to the corresponding nominal values are also presented. A comprehensive simulation study compares the different approaches, and the application of the different procedures to real data sets from four rivers in the United States and one in Spain complete the paper.     

Semi-Mechanized Simultaneous Synthesis of Multiple Oligonucleotide Fragments

Abstract

A new strategy for the simultaneous synthesis of multiple oligonucleotides is described, which is based on asynchronous chain growth and use of a maximum number of support segments in each elongation. An approach to the mechanization of this strategy and applications to the preparation of gene fragments and sequentially variant probes are discussed.     

甘蔗一次多基因遗传转化及多重PCR检测

通过多基因遗传转化策略,水稻、玉米等农作物实现了一次多个性状的遗传改良,为探索甘蔗一次多基因遗传转化的可行性,用一个多基因植物表达载体,对甘蔗进行遗传转化。通过多重PCR法对4个外源基因在各个转基因株系内的整合与缺失进行检测。结果证明,甘蔗一次多基因遗传转化是可行的,但因插入的DNA片段较大,会发生基因丢失现象。因此,一次多基因遗传转化需要在转化的过程中获得相对较多的转化株系,才能从中筛选到有应用价值的株系。     

3种检测吡咯喹啉醌的方法比较

目的:研究和比较3种检测吡咯喹啉醌(PQQ)的方法,确定各种方法的特点和适用范围。方法:设计和改进了3种检测PQQ的方法,分别为活性电泳法、光谱法和NBT-Gly法,探讨其检测限和线性范围、精密度、样品检测和加样回收率。结果:活性电泳法专一性好,具有很高的灵敏度,可检测到12.6 ng/mL PQQ,可靠但准确性较差;NBT-Gly法操作简便,可用于大量样品的检测,但重复性不佳;光谱法精密度较好,但样品中存在吸光物质时对检测结果影响较大。结论:活性电泳法、光谱法和NBT-Gly法均可用于PQQ的检测,活性电泳法适于培养上清等复杂样品的粗略定量,NBT-Gly法适于大量样品的检测,光谱法适于纯度较高的PQQ的定量。     

Comparing Trials with Multiple Outcomes: The Multivariate One-Sided Hypothesis with Unknown Covariances

The paper deals with a problem arising for tests in clinical trials. The outcomes of a standard and a new treatment to be compared are multivariate normally distributed with common but unknown covariance matrix. Under the null hypothesis the means of the outcomes are equal, under the alternative the new treatment is assumed to be superior, i.e. the means are larger without further quantification. For known covariance matrix there is a variety of tests for this problem. Some of these procedures can be extended to the case of unknown covariances if one is willing to accept a bias. There is, however, also an efficient unbiased test. The paper contains some numerical comparisons of these different procedures and takes a look on the minimax properties of the unbiased test.     

三重RT-PCR同步检测马铃薯多种病毒影响因素

根据病毒外壳蛋白区序列设计PVX、PVS特异性引物对,根据P1基因区序列设计PVA特异性引物对,应用三重RT-PCR同步检测马铃薯X病毒,马铃薯A病毒及马铃薯S病毒,分别得到562bp、255bp、182bp大小的扩增片段。试验从反转录反应、PCR反应及循环条件3方面讨论了试剂和循环条件对三重RT-PCR同步检测3种病毒的影响。结果表明反转录反应中dNTPs浓度、3种病毒下游引物浓度比例对整个反应影响较大;其次是PCR反应中MgC12浓度和退火温度;反转录时间,循环条件对RT-PCR影响较小。     

Simultaneous PCR Detection of Multiple Classes of Integron Integrase Genes for Determining the Presence of Multidrug-Resistant Bacteria in Environmental Samples

Dissemination of multidrug-resistant bacteria, particularly in hospitals, has become a serious public health problem. Integrons impart antibiotic multidrug resistance in gram-negative and some gram-positive bacteria by capturing and then disseminating antibiotic resistance genes. This mechanism plays a major role in contributing to the alarmingly high prevalence of bacterial drug resistance. A universal polymerase chain reaction (PCR) primer set was attempted to design to more sensitively and specifically detect integrons in environmental samples. One set, designated intCiF3_a, intCiF3_b, intCiiiR3_a, and intCiiiR3_b, simultaneously amplifies the conserved region of the tyrosine recombinase gene family between box I and box II. This primer set generates PCR products derived from classes 1, 2, and 3 integron integrases from environmental samples such as wastewater. An unexpected finding of this study was the detection of new putative integron integrase gene sequences. This is the subject of ongoing research, which aims to provide a clear understanding of the risk to human health posed by these genetic elements.     

Fluctuating Asymmetry as a Bioindicator of Stress: Comparing Efficacy of Analyses Involving Multiple Traits

Researchers have suggested fluctuating asymmetry (FA) as an indicator of environmental stress and have usually tested this assertion by examining relations between FA of single traits and stress. Fluctuating asymmetry stress relations are real but are typically weak and difficult to detect. Researchers would like to maximize the probability of detecting FA-stress relations when they exist. We assert that analyses based on the FA of multiple traits may provide better methods for detecting stress. In this article, we used computer simulations to compare the ability of six analyses to detect differences in FA between stressed and unstressed populations. We show that the optimal analysis depends upon the underlying form of the FA distributions. We also show that two of the analyses had inflated Type I errors in some situations. Finally, we quantify the advantage of our preferred analysis over those of single-trait FA in detecting stress.