Crowdsourcing malaria parasite quantification: an online game for analyzing images of infected thick blood smears

Background: There are 600,000 new malaria cases daily worldwide. The gold standard for estimating the parasite burden and the corresponding severity of the disease consists in manually counting the number of parasites in blood smears through a microscope, a process that can take more than 20 minutes of an expert microscopist’s time. Objective: This research tests the feasibility of a crowdsourced approach to malaria image analysis. In particular, we investigated whether anonymous volunteers with no prior experience would be able to count malaria parasites in digitized images of thick blood smears by playing a Web-based game. Methods: The experimental system consisted of a Web-based game where online volunteers were tasked with detecting parasites in digitized blood sample images coupled with a decision algorithm that combined the analyses from several players to produce an improved collective detection outcome. Data were collected through the MalariaSpot website. Random images of thick blood films containing Plasmodium falciparum at medium to low parasitemias, acquired by conventional optical microscopy, were presented to players. In the game, players had to find and tag as many parasites as possible in 1 minute. In the event that players found all the parasites present in the image, they were presented with a new image. In order to combine the choices of different players into a single crowd decision, we implemented an image processing pipeline and a quorum algorithm that judged a parasite tagged when a group of players agreed on its position. Results: Over 1 month, anonymous players from 95 countries played more than 12,000 games and generated a database of more than 270,000 clicks on the test images. Results revealed that combining 22 games from nonexpert players achieved a parasite counting accuracy higher than 99%. This performance could be obtained also by combining 13 games from players trained for 1 minute. Exhaustive computations measured the parasite counting accuracy for all players as a function of the number of games considered and the experience of the players. In addition, we propose a mathematical equation that accurately models the collective parasite counting performance. Conclusions: This research validates the online gaming approach for crowdsourced counting of malaria parasites in images of thick blood films. The findings support the conclusion that nonexperts are able to rapidly learn how to identify the typical features of malaria parasites in digitized thick blood samples and that combining the analyses of several users provides similar parasite counting accuracy rates as those of expert microscopists. This experiment illustrates the potential of the crowdsourced gaming approach for performing routine malaria parasite quantification, and more generally for solving biomedical image analysis problems, with future potential for telediagnosis related to global health challenges.

A stochastic model updating method for parameter variability quantification based on response surface models and Monte Carlo simulation

Stochastic model updating must be considered for quantifying uncertainties inherently existing in real-world engineering structures. By this means the statistical properties,instead of deterministic values, of structural parameters can be sought indicating the parameter variability. However, the implementation of stochastic model updating is much more complicated than that of deterministic methods particularly in the aspects of theoretical complexity and low computational efficiency. This study attempts to propose a simple and cost-efficient method by decomposing a stochastic updating process into a series of deterministic ones with the aid of response surface models and Monte Carlo simulation. The response surface models are used as surrogates for original FE models in the interest of programming simplification, fast response computation and easy inverse optimization. Monte Carlo simulation is adopted for generating samples from the assumed or measured probability distributions of responses. Each sample corresponds to an individual deterministic inverse process predicting the deterministic values of parameters. Then the parameter means and variances can be statistically estimated based on all the parameter predictions by running all the samples. Meanwhile, the analysis of variance approach is employed for the evaluation of parameter variability significance. The proposed method has been demonstrated firstly on a numerical beam and then a set of nominally identical steel plates tested in the laboratory. It is found that compared with the existing stochastic model updating methods, the proposed method presents similar accuracy while its primary merits consist in its simple implementation and cost efficiency in response computation and inverse optimization.

Quantification of soluble fibre in feedstuffs for rabbits and evaluation of the interference between the determinations of soluble fibre and intestinal mucin.

This work compared the quantification of soluble fibre in feeds using different chemical and in vitro approaches, and studied the potential interference between soluble fibre and mucin determinations. Six ingredients: sugar beet pulp (SBP), SBP pectins, insoluble SBP, wheat straw, sunflower hulls and lignocellulose, and seven rabbit diets, differing in soluble fibre content, were evaluated. In experiment 1, ingredients and diets were analyzed for total dietary fibre (TDF), insoluble dietary fibre (IDF), soluble dietary fibre (SDF), aNDFom (corrected for protein, aNDFom-cp) and 2-step pepsin/pancreatin in vitro DM indigestibility (corrected for ash and protein, ivDMi2). Soluble fibre was estimated by difference using three procedures: TDF?IDF (SDFIDF), TDF?ivDMi2 (SDFivDMi2), and TDF?aNDFom-cp (SDFaNDFom-cp). Soluble fibre determined directly (SDF) or by difference as SDFivDMi2 were not different (109 g/kg DM, on average). However, when it was calculated as SDFaNDFom-cp the value was 40% higher (153 g/kg DM, P menor que 0.05), whereas SDFIDF (124 g/kg DM) did not differ from any of the other methods. The correlation between the four methods was high (r ? 0.96; P ? 0.001; n = 13), but it decreased or even disappeared when SBP pectins and SBP were excluded and a lower and more narrow range of variation of soluble fibre was used. In experiment 2, the ivDMi2 using crucibles (reference method) were compared to those made using individual or collective ankom bags in order to simplify the determination of SDFivDMi2. The ivDMi2 was not different when using crucibles or individual or collective ankom bags. In experiment 3, the potential interference between soluble fibre and intestinal mucin determinations was studied using rabbit intestinal raw mucus, digesta and SBP pectins, lignocelluloses and a rabbit diet. An interference was observed between the determinations of soluble fibre and crude mucin, as contents of TDF and apparent crude mucin were high in SBP pectins (994 and 709 g/kg DM) and rabbit intestinal raw mucus (571 and 739 g/kg DM). After a pectinase treatment, the coefficient of apparent mucin recovery of SBP pectins was close to zero, whereas that of rabbit mucus was not modified. An estimation of the crude mucin carbohydrates retained in digesta TDF is proposed to correct TDF and soluble fibre digestibility. In conclusion, the values of soluble fibre depend on the methodology used. The contamination of crude mucin with soluble fibre is avoided using pectinase.

Detection and quantification of peanut traces in wheat flour by near infrared hyperspectral imaging spectroscopy using principal-component analysis

The use of a common environment for processing different powder foods in the industry has increased the risk of finding peanut traces in powder foods. The analytical methods commonly used for detection of peanut such as enzyme-linked immunosorbent assay (ELISA) and real-time polymerase chain reaction (RT-PCR) represent high specificity and sensitivity but are destructive and time-consuming, and require highly skilled experimenters. The feasibility of NIR hyperspectral imaging (HSI) is studied for the detection of peanut traces down to 0.01% by weight. A principal-component analysis (PCA) was carried out on a dataset of peanut and flour spectra. The obtained loadings were applied to the HSI images of adulterated wheat flour samples with peanut traces. As a result, HSI images were reduced to score images with enhanced contrast between peanut and flour particles. Finally, a threshold was fixed in score images to obtain a binary classification image, and the percentage of peanut adulteration was compared with the percentage of pixels identified as peanut particles. This study allowed the detection of traces of peanut down to 0.01% and quantification of peanut adulteration from 10% to 0.1% with a coefficient of determination (r2) of 0.946. These results show the feasibility of using HSI systems for the detection of peanut traces in conjunction with chemical procedures, such as RT-PCR and ELISA to facilitate enhanced quality-control surveillance on food-product processing lines.

Modal analysis of buildings : quantification of uncertainties and model updating

Una estructura vibra con la suma de sus infinitos modos de vibración, definidos por sus parámetros modales (frecuencias naturales, formas modales y coeficientes de amortiguamiento). Estos parámetros se pueden identificar a través del Análisis Modal Operacional (OMA). Así, un equipo de investigación de la Universidad Politécnica de Madrid ha identificado las propiedades modales de un edificio de hormigón armado en Madrid con el método Identificación de los sub-espacios estocásticos (SSI). Para completar el estudio dinámico de este edificio, se ha desarrollado un modelo de elementos finitos (FE) de este edificio de 19 plantas. Este modelo se ha calibrado a partir de su comportamiento dinámico obtenido experimentalmente a través del OMA. Los objetivos de esta tesis son; (i) identificar la estructura con varios métodos de SSI y el uso de diferentes ventanas de tiempo de tal manera que se cuantifican incertidumbres de los parámetros modales debidos al proceso de estimación, (ii) desarrollar FEM de este edificio y calibrar este modelo a partir de su comportamiento dinámico, y (iii) valorar la bondad del modelo. Los parámetros modales utilizados en esta calibración han sido; espesor de las losas, densidades de los materiales, módulos de elasticidad, dimensiones de las columnas y las condiciones de contorno de la cimentación. Se ha visto que el modelo actualizado representa el comportamiento dinámico de la estructura con una buena precisión. Por lo tanto, este modelo puede utilizarse dentro de un sistema de monitorización estructural (SHM) y para la detección de daños. En el futuro, podrá estudiar la influencia de los agentes medioambientales, tales como la temperatura o el viento, en los parámetros modales. A structure vibrates according to the sum of its vibration modes, defined by their modal parameters (natural frequencies, damping ratios and modal shapes). These parameters can be identified through Operational Modal Analysis (OMA). Thus, a research team of the Technical University of Madrid has identified the modal properties of a reinforced-concrete-frame building in Madrid using the Stochastic Subspace Identification (SSI) method and a time domain technique for the OMA. To complete the dynamic study of this building, a finite element model (FE) of this 19-floor building has been developed throughout this thesis. This model has been updated from its dynamic behavior identified by the OMA. The objectives of this thesis are to; (i) identify the structure with several SSI methods and using different time blocks in such a way that uncertainties due to the modal parameter estimation are quantified, (ii) develop a FEM of this building and tune this model from its dynamic behavior, and (iii) Assess the quality of the model, the modal parameters used in this updating process have been; thickness of slabs, material densities, modulus of elasticity, column dimensions and foundation boundary conditions. It has been shown that the final updated model represents the structure with a very good accuracy. Thus, this model might be used within a structural health monitoring framework (SHM). The study of the influence of changing environmental factors (such as temperature or wind) on the model parameters might be considered as a future work.

Methods for the analysis of multi-scale cell dynamics from fluorescence microscopy images

La embriogénesis es el proceso mediante el cual una célula se convierte en un ser un vivo. A lo largo de diferentes etapas de desarrollo, la población de células va proliferando a la vez que el embrión va tomando forma y se configura. Esto es posible gracias a la acción de varios procesos genéticos, bioquímicos y mecánicos que interaccionan y se regulan entre ellos formando un sistema complejo que se organiza a diferentes escalas espaciales y temporales. Este proceso ocurre de manera robusta y reproducible, pero también con cierta variabilidad que permite la diversidad de individuos de una misma especie. La aparición de la microscopía de fluorescencia, posible gracias a proteínas fluorescentes que pueden ser adheridas a las cadenas de expresión de las células, y los avances en la física óptica de los microscopios han permitido observar este proceso de embriogénesis in-vivo y generar secuencias de imágenes tridimensionales de alta resolución espacio-temporal. Estas imágenes permiten el estudio de los procesos de desarrollo embrionario con técnicas de análisis de imagen y de datos, reconstruyendo dichos procesos para crear la representación de un embrión digital. Una de las más actuales problemáticas en este campo es entender los procesos mecánicos, de manera aislada y en interacción con otros factores como la expresión genética, para que el embrión se desarrolle. Debido a la complejidad de estos procesos, estos problemas se afrontan mediante diferentes técnicas y escalas específicas donde, a través de experimentos, pueden hacerse y confrontarse hipótesis, obteniendo conclusiones sobre el funcionamiento de los mecanismos estudiados. Esta tesis doctoral se ha enfocado sobre esta problemática intentando mejorar las metodologías del estado del arte y con un objetivo específico: estudiar patrones de deformación que emergen del movimiento organizado de las células durante diferentes estados del desarrollo del embrión, de manera global o en tejidos concretos. Estudios se han centrado en la mecánica en relación con procesos de señalización o interacciones a nivel celular o de tejido. En este trabajo, se propone un esquema para generalizar el estudio del movimiento y las interacciones mecánicas que se desprenden del mismo a diferentes escalas espaciales y temporales. Esto permitiría no sólo estudios locales, si no estudios sistemáticos de las escalas de interacción mecánica dentro de un embrión. Por tanto, el esquema propuesto obvia las causas de generación de movimiento (fuerzas) y se centra en la cuantificación de la cinemática (deformación y esfuerzos) a partir de imágenes de forma no invasiva. Hoy en día las dificultades experimentales y metodológicas y la complejidad de los sistemas biológicos impiden una descripción mecánica completa de manera sistemática. Sin embargo, patrones de deformación muestran el resultado de diferentes factores mecánicos en interacción con otros elementos dando lugar a una organización mecánica, necesaria para el desarrollo, que puede ser cuantificado a partir de la metodología propuesta en esta tesis. La metodología asume un medio continuo descrito de forma Lagrangiana (en función de las trayectorias de puntos materiales que se mueven en el sistema en lugar de puntos espaciales) de la dinámica del movimiento, estimado a partir de las imágenes mediante métodos de seguimiento de células o de técnicas de registro de imagen. Gracias a este esquema es posible describir la deformación instantánea y acumulada respecto a un estado inicial para cualquier dominio del embrión. La aplicación de esta metodología a imágenes 3D + t del pez zebra sirvió para desvelar estructuras mecánicas que tienden a estabilizarse a lo largo del tiempo en dicho embrión, y que se organizan a una escala semejante al del mapa de diferenciación celular y con indicios de correlación con patrones de expresión genética. También se aplicó la metodología al estudio del tejido amnioserosa de la Drosophila (mosca de la fruta) durante el cierre dorsal, obteniendo indicios de un acoplamiento entre escalas subcelulares, celulares y supracelulares, que genera patrones complejos en respuesta a la fuerza generada por los esqueletos de acto-myosina. En definitiva, esta tesis doctoral propone una estrategia novedosa de análisis de la dinámica celular multi-escala que permite cuantificar patrones de manera inmediata y que además ofrece una representación que reconstruye la evolución de los procesos como los ven las células, en lugar de como son observados desde el microscopio. Esta metodología por tanto permite nuevas formas de análisis y comparación de embriones y tejidos durante la embriogénesis a partir de imágenes in-vivo. ABSTRACT The embryogenesis is the process from which a single cell turns into a living organism. Through several stages of development, the cell population proliferates at the same time the embryo shapes and the organs develop gaining their functionality. This is possible through genetic, biochemical and mechanical factors that are involved in a complex interaction of processes organized in different levels and in different spatio-temporal scales. The embryogenesis, through this complexity, develops in a robust and reproducible way, but allowing variability that makes possible the diversity of living specimens. The advances in physics of microscopes and the appearance of fluorescent proteins that can be attached to expression chains, reporting about structural and functional elements of the cell, have enabled for the in-vivo observation of embryogenesis. The imaging process results in sequences of high spatio-temporal resolution 3D+time data of the embryogenesis as a digital representation of the embryos that can be further analyzed, provided new image processing and data analysis techniques are developed. One of the most relevant and challenging lines of research in the field is the quantification of the mechanical factors and processes involved in the shaping process of the embryo and their interactions with other embryogenesis factors such as genetics. Due to the complexity of the processes, studies have focused on specific problems and scales controlled in the experiments, posing and testing hypothesis to gain new biological insight. However, methodologies are often difficult to be exported to study other biological phenomena or specimens. This PhD Thesis is framed within this paradigm of research and tries to propose a systematic methodology to quantify the emergent deformation patterns from the motion estimated in in-vivo images of embryogenesis. Thanks to this strategy it would be possible to quantify not only local mechanisms, but to discover and characterize the scales of mechanical organization within the embryo. The framework focuses on the quantification of the motion kinematics (deformation and strains), neglecting the causes of the motion (forces), from images in a non-invasive way. Experimental and methodological challenges hamper the quantification of exerted forces and the mechanical properties of tissues. However, a descriptive framework of deformation patterns provides valuable insight about the organization and scales of the mechanical interactions, along the embryo development. Such a characterization would help to improve mechanical models and progressively understand the complexity of embryogenesis. This framework relies on a Lagrangian representation of the cell dynamics system based on the trajectories of points moving along the deformation. This approach of analysis enables the reconstruction of the mechanical patterning as experienced by the cells and tissues. Thus, we can build temporal profiles of deformation along stages of development, comprising both the instantaneous events and the cumulative deformation history. The application of this framework to 3D + time data of zebrafish embryogenesis allowed us to discover mechanical profiles that stabilized through time forming structures that organize in a scale comparable to the map of cell differentiation (fate map), and also suggesting correlation with genetic patterns. The framework was also applied to the analysis of the amnioserosa tissue in the drosophila’s dorsal closure, revealing that the oscillatory contraction triggered by the acto-myosin network organized complexly coupling different scales: local force generation foci, cellular morphology control mechanisms and tissue geometrical constraints. In summary, this PhD Thesis proposes a theoretical framework for the analysis of multi-scale cell dynamics that enables to quantify automatically mechanical patterns and also offers a new representation of the embryo dynamics as experienced by cells instead of how the microscope captures instantaneously the processes. Therefore, this framework enables for new strategies of quantitative analysis and comparison between embryos and tissues during embryogenesis from in-vivo images.