Real-Time PCR : A Molecular Approach to Investigate the Role of Intestinal Microbiota in the Pathophysiology of Irritable Bowel Syndrome

Irritable bowel syndrome (IBS) is a common multifactorial functional intestinal disorder, the pathogenesis of which is not completely understood. Increasing scientific evidence suggests that microbes are involved in the onset and maintenance of IBS symptoms. The microbiota of the human gastrointestinal (GI) tract constitutes a massive and complex ecosystem consisting mainly of obligate anaerobic microorganisms making the use of culture-based methods demanding and prone to misinterpretation. To overcome these drawbacks, an extensive panel of species- and group-specific assays for an accurate quantification of bacteria from fecal samples with real-time PCR was developed, optimized, and validated. As a result, the target bacteria were detectable at a minimum concentration range of approximately 10 000 bacterial genomes per gram of fecal sample, which corresponds to the sensitivity to detect 0.000001% subpopulations of the total fecal microbiota. The real-time PCR panel covering both commensal and pathogenic microorganisms was assessed to compare the intestinal microbiota of patients suffering from IBS with a healthy control group devoid of GI symptoms. Both the IBS and control groups showed considerable individual variation in gut microbiota composition. Sorting of the IBS patients according to the symptom subtypes (diarrhea, constipation, and alternating predominant type) revealed that lower amounts of Lactobacillus spp. were present in the samples of diarrhea predominant IBS patients, whereas constipation predominant IBS patients carried increased amounts of Veillonella spp. In the screening of intestinal pathogens, 17% of IBS samples tested positive for Staphylococcus aureus, whereas no positive cases were discovered among healthy controls. Furthermore, the methodology was applied to monitor the effects of a multispecies probiotic supplementation on GI microbiota of IBS sufferers. In the placebo-controlled double-blind probiotic intervention trial of IBS patients, each supplemented probiotic strain was detected in fecal samples. Intestinal microbiota remained stable during the trial, except for Bifidobacterium spp., which increased in the placebo group and decreased in the probiotic group. The combination of assays developed and applied in this thesis has an overall coverage of 300-400 known bacterial species, along with the number of yet unknown phylotypes. Hence, it provides good means for studying the intestinal microbiota, irrespective of the intestinal condition and health status. In particular, it allows screening and identification of microbes putatively associated with IBS. The alterations in the gut microbiota discovered here support the hypothesis that microbes are likely to contribute to the pathophysiology of IBS. The central question is whether the microbiota changes described represent the cause for, rather than the effect of, disturbed gut physiology. Therefore, more studies are needed to determine the role and importance of individual microbial species or groups in IBS. In addition, it is essential that the microbial alterations observed in this study will be confirmed using a larger set of IBS samples of different subtypes, preferably from various geographical locations.

Computation-Based Reliability Analysis in Real Time Applications

Reliability analysis for computing systems in aerospace applications must account for actual computations the system performs in the use environment. This paper introduces a theoretical nonhomogeneous Markov model for such applications.

A simple method for the measurement of stress in evaporated thin films by real-time holographic interferometry

The properties of thin films depend to a large extent upon their mechanical stability which in turn is dependent on the intrinsic stresses developed during evaporation. This paper describes a simple method for the measurement of stresses in thin films by the use of real-time holographic interferometry.

Secure Concurrency Control in Firm Real-Time Database Systems

Many real-time database applications arise in electronic financial services, safety-critical installations and military systems where enforcing security is crucial to the success of the enterprise. For real-time database systems supporting applications with firm deadlines, we investigate here the performance implications, in terms of killed transactions, of guaranteeing multilevel secrecy. In particular, we focus on the concurrency control (CC) aspects of this issue. Our main contributions are the following: First, we identify which among the previously proposed real-time CC protocols are capable of providing covert-channel-free security. Second, using a detailed simulation model, we profile the real-time performance of a representative set of these secure CC protocols for a variety of security-classified workloads and system configurations. Our experiments show that a prioritized optimistic CC protocol, OPT-WAIT, provides the best overall performance. Third, we propose and evaluate a novel "dual-CC" approach that allows the real-time database system to simultaneously use different CC mechanisms for guaranteeing security and for improving real-time performance. By appropriately choosing these different mechanisms, concurrency control protocols that provide even better performance than OPT-WAIT are designed. Finally, we propose and evaluate GUARD, an adaptive admission-control policy designed to provide fairness with respect to the distribution of killed transactions across security levels. Our experiments show that GUARD efficiently provides close to ideal fairness for real-time applications that can tolerate covert channel bandwidths of upto one bit per second.

A real transformation for modal analysis of flexible damped spacecraft

Modal approach is widely used for the analysis of dynamics of flexible structures. However, space analysts yet lack an intimate modal analysis of current spacecraft which are rich with flexibility and possess both structural and discrete damping. Mathematical modeling of such spacecraft incapacitates the existing real transformation procedure, for it cannot include discrete damping, demands uncomputable inversion of a modal matrix inaccessible due to its overwhelming size and does not permit truncation. On the other hand, complex transformation techniques entail more computational time and cannot handle structural damping. This paper presents a real transformation strategy which averts inversion of the associated real transformation matrix, allows truncation and accommodates both forms of damping simultaneously. This is accomplished by establishing a key relation between the real transformation matrix and its adjoint. The relation permits truncation of the matrices and leads to uncoupled pairs of coupled first order equations which contain a number of adjoint eigenvectors. Finally these pairs are solved to obtain a literal modal response of forced gyroscopic damped flexibile systems at arbitrary initial conditions.

Dual-trap optical tweezers with real-time force clamp control

Single molecule force clamp experiments are widely used to investigate how enzymes, molecular motors, and other molecular mechanisms work. We developed a dual-trap optical tweezers instrument with real-time (200 kHz update rate) force clamp control that can exert 0–100 pN forces on trapped beads. A model for force clamp experiments in the dumbbell-geometry is presented. We observe good agreement between predicted and observed power spectra of bead position and force fluctuations. The model can be used to predict and optimize the dynamics of real-time force clamp optical tweezers instruments. The results from a proof-of-principle experiment in which lambda exonuclease converts a double-stranded DNA tether, held at constant tension, into its single-stranded form, show that the developed instrument is suitable for experiments in single molecule biology.

Real-time multipurpose reservoir operation: a case study / Exploitation en temps reacuteel d'un reservoir agrave buts multiples: eacutetude d'un cas

A real-time operational methodology has been developed for multipurpose reservoir operation for irrigation and hydropower generation with application to the Bhadra reservoir system in the state of Karnataka, India. The methodology consists of three phases of computer modelling. In the first phase, the optimal release policy for a given initial storage and inflow is determined using a stochastic dynamic programming (SDP) model. Streamflow forecasting using an adaptive AutoRegressive Integrated Moving Average (ARIMA) model constitutes the second phase. A real-time simulation model is developed in the third phase using the forecast inflows of phase 2 and the operating policy of phase 1. A comparison of the optimal monthly real-time operation with the historical operation demonstrates the relevance, applicability and the relative advantage of the proposed methodology.

Specification of Real-Time Systems

Inspite of numerous research advancements made in recent years in the area of formal techniques, specification of real-time systems is still proving to be a very challenging and difficult problem. In this context, this paper critically examines state-of-the-art specification techniques for real-time systems and analyzes the emerging trends.