정확한 서열정렬기법과 인메모리 핵심 유전자 데이터베이스 기반의 향상된 메타유전체 분류법

학위논문 (박사) -- 서울대학교 대학원 : 자연과학대학 협동과정 생물정보학전공, 2020. 8. 천종식.샷건 메타지노믹스는 미생물과 숙주 또는 환경사이의 미치는 영향을 이해하는데 매우 중요한 역할을 하고 있다. 기술의 발달과 더불어 메타지노믹스를 통한 올바른 미생물 종의 동정과 각 종들의 분포는 마이크로바이옴 연구의 핵심 구성요소가 되었으며, 지난 10년간 샷건 메타지노믹스 분석을 위한 여러 알고리즘과 데이터베이스들이 개발되어져 왔다. 하지만 서로 다른 기준 데이터 혹은 알고리즘을 사용한 방법들은 서로 다른 분류 정보와 분석 파이프라인으로 인하여 편향된 결과를 나타내기도 하였는데, 이를 보완하고 보다 정확한 분류 동정을 위해 배양이 어려운 표준 균주와 같은 다양한 균주의 유전체 데이터를 포함하는 기준 데이터베이스의 중요성이 대두되고 있다. 샷건 메타지노믹스 분석에서 또 다른 중요한 요소는 분석에 소요되는 시간이라 할 수 있는데 대부분의 생물정보학적 프로그램들은 계산을 수행함에 있어 메모리와 알고리즘 최적화가 되어있지 않아 분석에 상당한 시간이 소요되는 문제점이 있다. 이러한 문제를 해결하기 위해, 본 연구에서는 exact match k-mer classification과 같은 방법을 사용하여 분석 속도를 향상시켰으며 Up-to-date Bacterial Core Gene (UBCG)를 기준 데이터베이스로 사용하여 보다 정확한 샷건 메타지노믹 분석을 수행할 수 있게 하였다. 분석의 효율성을 높이기 위해 두개의 기준 UBCG 데이터베이스가 만들어 졌으며 한 개는 박테리아의 분류체계에서 유효한 종명 (Valid names)만을 가지고 있는 데이터베이스와 다른 하나는 유효한 종명과 함께 EzBioCloud에 있는 genomospecies를 가지고 생성하였다. 검증을 위해 Streptococcus 종을 포함하는 (i) 합성된 메타지놈 샘플과 (ii) 만성 폐쇄성 폐질환(COPD) 환자의 임상 검체 (iii) 혈류 감염 환자의 임상 검체로 이루어진 세개의 데이터 셋을 이용하였으며 기존에 널리 알려진 샷것 파이프라인인 MetaPhlan2과 본 연구의 파이프라인을 비교 분석하였다. 위 검증 분석에서 UBCG를 기준 서열로 사용하기에 충분함을 검증하였으며, 빠르고 정확하게 기준 유전체에서 UBCG 서열을 뽑아 샷건 분석에 용이함을 증명하였다. 또한 genomospecies를 기준 데이터베이스에 추가함으로써, 보다 개선된 분류 정확도를 얻을 수 있음을 제시하였다. 마지막으로 비록 여러 파이프라인과 데이터베이스들이 존재하지만 보다 신뢰할 수 있는 분류결과를 얻기 위해선 기준 데이터베이스의 지속적인 업데이트와 분류 체계의 검증의 중요함을 강조하였다. 이후 본 연구에서 개발된 파이프라인을 이용하여 4,000개의 샷건 메타지놈 샘플에서 사람에 장내에 가장 많이 발견되는 Bacteroides 종에 대한 분석을 수행하였다. 많은 양의 데이터를 분석하여야 하기 때문에 기존에 많이 사용되는 MetaPhlAn2 과 같은 방법은 사용할 수 없었으며 분석 결과 Bacteroides는 도시화된 사람에게 많이 분포하는 반면 아프리카 혹은 남미지역에서 원시적 부족의 삶을 사는 사람에게서는 상대적으로 적게 분포함을 확인할 수 있었다. 또한 각 나라별 인구에서는 우점되는 Bacteroides 종이 다름을 확인할 수 있었는데 이는 각 연구의 샘플링 방법 혹은 위치에 따라 설명되어 질 수 있었다. 실험용 쥐의 결과에서는 가장 다양한 Bacteroides를 관찰할 수 있었으며 이는 많은 수의 기준 유전체가 생쥐에게서 나왔기 때문인 것으로 생각된다. 또한 고양이나 강아지 같은 반려동물의 샘플에서도 높은 상관관계를 발견할 수 있었는데 각 동물들의 생활양식과 먹이에 따른 결과인 것으로 보인다. 본 연구를 통해 보다 많은 메타지놈 데이터 분석의 필요성을 강조하고 있으며, 핵심 유전자들을 기준 데이터로 사용하는 방법의 실효성과 성능을 검증하였다. 이러한 핵심 유전자 기반의 기준 데이터베이스는 보다 정확하고 전체 미생물의 풍부도를 예측하는데 중요한 역할을 하는 것을 확인하였고 k-mer 방법을 통해 기존에 존재하던 다른 파이프라인 보다 더욱 빠른 결과를 도출할 수 있었다. 마지막으로 빠르게 기준 데이터베이스를 만들 수 있기 때문에 항상 최신의 데이터를 가지고 분석을 수행할 수 있으며 이는 궁극적으로 본 연구의 파이프라인을 실질적으로 연구나 진단 목적으로 이용하는 연구자들에게 큰 도움이 될 것이다.Shotgun metagenomics is of great importance to understand the microbial community composition of a sample and the impact it has on its host. The proper identification and quantification of bacterial species is a key component of any microbiome research that is based on metagenomic samples. In the last decade, several algorithms and databases have been developed, however the differences between references and the type of algorithm used for the classification makes the comparisons among themselves unfair and bias. The contents of the reference database, including genome sequences of type strains or reference genomes of uncultured species, have a great impact on the performance of the classification results of metagenomic samples. Another significant factor on shotgun metagenomics is the classification speed as most current bioinformatic tools lack computational and memory optimization. Here, I propose several enhancements to a well-known method, exact match k-mer classification in order to increase the overall speed of a metagenomic classification. This method was further improved by the use of Up-to-date Bacterial Core Gene (UBCG) sequences to provide better method for a faster and accurate shotgun metagenomic profiling classification. In order to prove the efficiency of our method, I built two UBCG-based reference databases: one containing UBCG sequences of valid named species, and the second one containing UBCG sequences of all valid named species and genomospecies in the EzBioCloud database. Three datasets containing Streptococcus species were used to evaluate the improved method against the MetaPhlan2 tool which is the most widely used open-source shotgun metagenomic classifier: (i) synthetic metagenomic samples, (ii) clinical sputum samples from patients with chronic obstructive pulmonary disease (COPD), and (iii) clinical samples of a blood stream infection. In this analysis, I demonstrated that UBCG sequences can be used as references for metagenomic classification, showing that they are easy to extract from genome sequences and accurate when predicting relative abundance. I also showed that the inclusion of genomospecies in the reference databases, significantly improves the classification accuracy of bacterial species within a metagenomic sample. Finally, I showed that while publicly available pipelines and databases are easily accessible, for accurate and reliable taxonomic classification, an updated database with proper taxonomic and genomic curation must be used. The method devised in this work is then applied to profile the Bacteroides species in over 4,000 shotgun metagenomic samples, which is one of most abundant members of the human gut microbiome. This task cannot be accomplished using conventional tools such as MetaPhlAn2 due to the high processing time they require. The results in this study showed that Bacteroides is high abundant in human samples from urban areas while being low abundant in humans from rural areas, particularly African and South American tribes. Countries showed dominance for a specific Bacteroides species, but this could also be explained by the type of study were the samples came from. Mice samples showed the most diversity of Bacteroides, this can be attributed by the number of bacterial references isolated from this organism. House cat and dog samples showed correlation between each other, this may be attributed to the similarities of their lifestyle and diet. This study shows the importance of having a great number of samples for any given metagenomic analysis, and even though, we have profiled thousands of samples, more might be needed in the future. The method proposed in this thesis demonstrates that core genes are reliable reference sequences for shotgun metagenomics. Their implementation as reference sequences in metagenomic databases improves the accuracy of the abundance prediction of any given sample. Additionally, with the use of a k-mer approach, this methods running time outperforms the most popular shotgun metagenomic tools. The work presented in this thesis aims to help microbial research by providing faster and accurate metagenomic taxonomic predictions. Finally, with the ability of updating a metagenomic database with ease, will help researchers to obtain the most up-to-date results to find potential diagnosis or treatments for diseases associated to human microbial communities.Chapter 1. General Introduction 1 1.1. Introduction to metagenomics 2 1.2. 16S rRNA sequencing 3 1.3. Shotgun metagenomic sequencing 5 1.3.1. History 5 1.3.2. Sample extraction 7 1.3.3. Library preparation 8 1.3.4. Sequencing 8 1.4. Shotgun metagenomic classification 9 1.4.1. Homology-based approaches 9 1.4.2. Exact match K-mer approaches 11 Chapter 2. An exact match k-mer algorithm 13 2.1. An exact match k-mer classification approach 14 2.1.1. Definition of the problem 14 2.1.2. Building a k-mer reference database 14 2.1.2.1. K-mer counting 14 2.1.2.2. K-mer mapping 16 2.1.3. Classification of a metagenomic read 16 2.1.3.1. K-mer search 19 2.1.3.2. Scoring a metagenomic read 20 2.1.4. Calculating the metagenome profile 20 2.1.4.1. Normalization for LCA-assigned reads 21 2.1.4.2. Normalization for cell count relative abundance 22 2.2. RAM memory usage 22 2.3. Quality Control 23 2.3.1. Read Trimming 23 2.3.2. Host read removal 24 Chapter 3. Revealing unrecognized species in the genus Streptococcus 28 3.1. A brief history of streptococcus in clinical metagenomics 29 3.2. Results and Discussion 32 3.2.1. Building a core gene reference database 32 3.2.2. Evaluation of Pipelines using Synthetic Metagenomes 36 3.2.3. Chronic obstructive pulmonary disease samples 44 3.2.3. Evaluating the value of genomospecies references in a metagenomic database 56 3.2.4. Identifying accurately a Streptococcal infection using clinical data 63 3.2.5. Effects of different ANI thresholds on the classification of genomospecies 69 3.3. Materials and Methods 76 3.3.1. Selecting the reference genomes 76 3.3.2. Average nucleotide identity and hierarchical clustering 76 3.3.3. Synthetic and Real metagenomic samples 77 3.3.4. Extracting the core genes 77 3.3.5. Taxonomic profiling 83 3.3.6. Biomarker discovery 84 3.4. Conclusions 85 Chapter 4. A large-scale shotgun metagenomic analysis on Bacteroides 86 4.1. Introduction 87 4.2. Bacteroides on the human gut 89 4.2.1. Collecting the samples 89 4.2.2. Methods 89 4.2.2.1. Reference Genomes 89 4.2.2.2. Metagenome profiling 90 4.2.3. Results 103 4.3. Bacteroides on Animal Species 128 4.3.1. Methods 128 4.3.2. Results 128 4.4. Discussion and conclusions 133 General Conclusion 135 References 139 Appendix I. A list of genomes from the genus Streptococcus used on Chapters 3 analysis. 146 국문초록 155Docto

Neurofuzzy controller based full vehicle nonlinear active suspension systems

To design a robust controller for active suspension systems is very important for guaranteeing the riding comfort for passengers and road handling quality for a vehicle. In this thesis, the mathematical model of full vehicle nonlinear active suspension systems with hydraulic actuators is derived to take into account all the motions of the vehicle and the nonlinearity behaviours of the active suspension system and hydraulic actuators. Four robust control types are designed and the comparisons among the robustness of those controllers against different disturbance types are investigated to select the best controller among them. The MATLAB SIMULINK toolboxes are used to simulate the proposed controllers with the controlled model and to display the responses of the controlled model under different types of disturbance. The results show that the neurofuzzy controller is more effective and robust than the other controller types. The implementation of the neurofuzzy controller using FPGA boards has been investigated in this work. The Xilinx ISE program is employed to synthesis the VHDL codes that describe the operation of the neurofuzzy controller and to generate the configuration file used to program the FPGA. The ModelSim program is used to simulate the operation of the VHDL codes and to obtain the expected output data of the FPGA boards. To confirm that FPGA the board used as the neurofuzzy controller system operated as expected, a MATLAB script file is used to compare the set of data obtained from the ModelSim program and the set of data obtained from the MATLAB SIMULINK model. The results show that the FPGA board is effective to be used as a neurofuzzy controller for full vehicle nonlinear active suspension systems. The active suspension system has a great performance for vibration isolation. However the main drawback of the active suspension is that it is high energy consumptive. Therefore, to use this suspension system in the proposed model, this drawback should be solved. Electromagnetic actuators are used to convert the vibration energy that arises from the rough road to useful electrical energy to reduce the energy consumption by the active suspension systems. The results show that the electromagnetic devices act as a power generator, i.e. the vibration energy excited by the rough road surface has been converted to a useful electrical energy supply for the actuators. Furthermore, when the nonlinear damper models are replaced by the electromagnetic actuators, riding comfort and the road handling quality are improved. As a result, two targets have been achieved by using hydraulic actuators with electromagnetic suspension systems: increasing fuel economy and improving the vehicle performance

Advanced Control Strategies for Mobile Hydraulic Applications

Mobile hydraulic machines are affected by numerous undesired dynamics, mainly discontinuous motion and vibrations. Over the years, many methods have been developed to limit the extent of those undesired dynamics and improve controllability and safety of operation of the machine. However, in most of the cases, today\u27s methods do not significantly differ from those developed in a time when electronic controllers were slower and less reliable than they are today. This dissertation addresses this aspect and presents a unique control method designed to be applicable to all mobile hydraulic machines controlled by proportional directional valves. In particular, the proposed control method is targeted to hydraulic machines such as those used in the field including construction (wheel loaders, excavators, and backhoes, etc.), load handling (cranes, reach-stackers, and aerial lift, etc.), agriculture (harvesters, etc.), forestry, and aerospace. For these applications the proposed control method is designed to achieve the following goals: A. Improvement of the machine dynamics by reducing mechanical vibrations of mechanical arms, load, as well as operator seat; B. Reduction of the energy dissipation introduced by current vibration damping methods; C. Reduction of system slowdowns introduced by current vibration damping methods. Goal A is generally intended for all machines; goal B refers to those applications in which the damping is introduced by means of energy losses on the main hydraulic transmission line; goal C is related to those applications in which the vibration attenuation is introduced by slowing down the main transmission line dynamics. Two case studies are discussed in this work: 1. Hydraulic crane: the focus is on the vibrations of the mechanical arms and load (goals A and B). 2. Wheel loader: the focus is on the vibrations of the driver\u27s seat and bucket (goals A and C). The controller structure is basically unvaried for different machines. However, what differs in each application are the controller parameters, whose adaptation and tuning method represent the main innovations of this work. The proposed controller structure is organized so that the control parameters are adapted with respect to the instantaneous operating point which is identified by means of feedback sensors. The Gain Scheduling technique is used to implement the controller whose set of parameters are function of the specific identified operating point. The optimal set of control parameters for each operating point is determined through the non-model-based controller tuning. The technique determines the optimal set of controller parameters through the optimization of the experimental machine dynamics. The optimization is based on an innovative application of the Extremum Seeking algorithm. The optimal controller parameters are then indexed into the Gain Scheduler. The proposed method does not require the modification of the standard valve controlled machine layout since it only needs for the addition of feedback sensors. The feedback signals are used by the control unit to modify the electric currents to the proportional directional valves and cancel the undesired dynamics of the machine by controlling the actuator motion. In order for the proposed method to be effective, the proportional valve bandwidth must be significantly higher than the frequency of the undesired dynamics. This condition, which is typically true for heavy machineries, is further investigated in the research. The research mostly focuses on the use of pressure feedback. In fact, although the use of position, velocity, or acceleration sensors on the vibrating bodies of the machine would provide a more straightforward measurement of the vibration, they are extremely rare on mobile hydraulic machines where mechanical and environmental stress harm their integrity. A comparison between pressure feedback and acceleration feedback alternatives of the proposed method is investigated with the aim to outline the conditions making one alternative preferable over the other one (for those applications were both alternatives are technically viable in terms of sensors and wiring reliability). A mid-sized hydraulic crane (case study 1) was instrumented at Maha Fluid Power Research Center to study the effectiveness of the proposed control method, its stability and its experimental validation. Up to 30% vibration damping and 40% energy savings were observed for a specific cycle over the standard vibration damping method for this application. The proposed control method was also applied to a wheel loader (case study 2), and up to 70% vibrations attenuation on the bucket and 30% on the driver\u27s cab were found in simulations. These results also served to demonstrate the applicability of the control method to different hydraulic machines. Improved system response and a straightforward controller parameters tuning methodology are the features which give to the proposed method the potential to become a widespread technology for fluid power machines. The proposed method also has potential for improving several current vibration damping methods in terms of energy efficiency as well as simplification of both the hydraulic system layout and tuning process

SPS phase control system performance via analytical simulation

A solar power satellite transmission system which incorporates automatic beam forming, steering, and phase control is discussed. The phase control concept centers around the notation of an active retrodirective phased array as a means of pointing the beam to the appropriate spot on Earth. The transmitting antenna (spacetenna) directs the high power beam so that it focuses on the ground-based receiving antenna (rectenna). A combination of analysis and computerized simulation was conducted to determine the far field performance of the reference distribution system, and the beam forming and microwave power generating systems

Skills Development for Inclusive and Sustainable Growth in Developing Asia-Pacific

Focusing on the Asia-Pacific region, which in recent years has been the engine of global economic growth , this volume surveys trends and prospects in technical and vocational education and training (TVET) with particular reference to achieving inclusive growth and the greening of economies. Underlying the increasing pressure for new models of TVET provision is the rapid pace of technological change, demand for a work force which is highly responsive to evolving needs and a transforming market place that calls for higher order skills and lifelong learning. The book proposes a re-engineered, modernized TVET system that fosters an innovative approach which enhances the employability of workers as well as the sustainability of their livelihoods. The book includes contributions from leading policy makers, researchers, and practitioners, including those in the private sector in analyzing and forecasting the most urgent priorities in skills development. The book argues for creative approaches to TVET design and delivery particularly with a view to improve job prospects , and meeting the goals of inclusion, sustainable development and social cohesion. Addressing issues such as the chronic mismatches between skills acquired and actual skills required in the work place, the volume proposes diversified approaches towards workforce development and partnerships with the private sector to improve the quality and relevance of skills development . The new imperatives created by ‘greening’ economies and responses required in skills development and training are addressed. Developing TVET is a high priority for governments in the Asia Pacific region as they seek to achieve long-term sustainable growth since the .continued success of their economic destinies depend on it. The volume also includes an emerging framework for skills development for inclusive and sustainable growth in the Asia and Pacific region

Analysis of high rate fracture tests of polymers

Imperial Users onl

Network-on-Chip

Addresses the Challenges Associated with System-on-Chip Integration Network-on-Chip: The Next Generation of System-on-Chip Integration examines the current issues restricting chip-on-chip communication efficiency, and explores Network-on-chip (NoC), a promising alternative that equips designers with the capability to produce a scalable, reusable, and high-performance communication backbone by allowing for the integration of a large number of cores on a single system-on-chip (SoC). This book provides a basic overview of topics associated with NoC-based design: communication infrastructure design, communication methodology, evaluation framework, and mapping of applications onto NoC. It details the design and evaluation of different proposed NoC structures, low-power techniques, signal integrity and reliability issues, application mapping, testing, and future trends. Utilizing examples of chips that have been implemented in industry and academia, this text presents the full architectural design of components verified through implementation in industrial CAD tools. It describes NoC research and developments, incorporates theoretical proofs strengthening the analysis procedures, and includes algorithms used in NoC design and synthesis. In addition, it considers other upcoming NoC issues, such as low-power NoC design, signal integrity issues, NoC testing, reconfiguration, synthesis, and 3-D NoC design. This text comprises 12 chapters and covers: The evolution of NoC from SoC—its research and developmental challenges NoC protocols, elaborating flow control, available network topologies, routing mechanisms, fault tolerance, quality-of-service support, and the design of network interfaces The router design strategies followed in NoCs The evaluation mechanism of NoC architectures The application mapping strategies followed in NoCs Low-power design techniques specifically followed in NoCs The signal integrity and reliability issues of NoC The details of NoC testing strategies reported so far The problem of synthesizing application-specific NoCs Reconfigurable NoC design issues Direction of future research and development in the field of NoC Network-on-Chip: The Next Generation of System-on-Chip Integration covers the basic topics, technology, and future trends relevant to NoC-based design, and can be used by engineers, students, and researchers and other industry professionals interested in computer architecture, embedded systems, and parallel/distributed systems