Gene regulatory networks: a coarse-grained, equation-free approach to multiscale computation

We present computer-assisted methods for analyzing stochastic models of gene regulatory networks. The main idea that underlies this equation-free analysis is the design and execution of appropriately-initialized short bursts of stochastic simulations; the results of these are processed to estimate coarse-grained quantities of interest, such as mesoscopic transport coefficients. In particular, using a simple model of a genetic toggle switch, we illustrate the computation of an effective free energy and of a state-dependent effective diffusion coefficient that characterize an unavailable effective Fokker-Planck equation. Additionally we illustrate the linking of equation-free techniques with continuation methods for performing a form of stochastic "bifurcation analysis"; estimation of mean switching times in the case of a bistable switch is also implemented in this equation-free context. The accuracy of our methods is tested by direct comparison with long-time stochastic simulations. This type of equation-free analysis appears to be a promising approach to computing features of the long-time, coarse-grained behavior of certain classes of complex stochastic models of gene regulatory networks, circumventing the need for long Monte Carlo simulations.Comment: 33 pages, submitted to The Journal of Chemical Physic

Under One Umbrella: Multiple Communities, One Church

It is my claim that the need for alternative communities within established church structures is not only growing, but is also paramount to the continuation of the gospel of Jesus Christ into the post-Christian, post-Modern world. These alternative communities often take the form of emerging worship services, Young Adult services, and churchwithin- a-church expressions of worship and community. They are seen in large, medium, and small church settings; as such, we see many of the same problems to varying degrees. The search for relevance has led us to these communities, and this author sees them as valid expressions of faith and community. I am further claiming that intrinsic within these community relationships is friction that often leads to challenges that, unless overcome, will not allow either community to see its full potential for the kingdom of heaven. I will explore different models of relationships, both for integration and separation, and suggest ways in which the conversations between communities can be facilitated in order to create preemptive understanding, which seeks in turn to allow each community a modicum of understanding and helps to create opportunities for all communities to thrive. I propose a book that will outline the challenges and look for opportunities therein. This book will address any church community that struggles with the growth of an alternative community within the more established church structure. The research will be broad enough to include churches of every size and can be seen cross-denominationally as a resource for easing the growing pains of alternative communities within already established church structures. The goal is to provide a framework for conversations between multiple communities housed under one ecclesial umbrella

Enhanced heterogeneity of rpoB in Mycobacterium tuberculosis found at low pH.

OBJECTIVES: The aim of this study was to gain an insight into the molecular mechanisms of the evolution of rifampicin resistance in response to controlled changes in the environment. METHODS: We determined the proportion of rpoB mutants in the chemostat culture and characterized the sequence of mutations found in the rifampicin resistance-determining region of rpoB in a steady-state chemostat at pH 7.0 and 6.2. RESULTS: The overall proportion of rpoB mutants of strain H37Rv remained constant for 37 days at pH 7.0, ranging between 3.6 x 10(-8) and 8.9 x 10(-8); however, the spectrum of mutations varied. The most commonly detected mutation, serine to leucine mutation at codon 531 (S531L), increased from 40% to 89%, while other mutations (S531W, H526Y, H526D, H526R, S522L and D516V) decreased over the 37 day sampling period. Changing the pH from 7.0 to 6.2 did not significantly alter the overall proportion of mutants, but resulted in a decrease in the percentage of strains harbouring S531L (from 89% to 50%) accompanied by an increase in the range of different mutations from 4 to 12. CONCLUSIONS: The data confirm that the fitness of strains with the S531L mutation is greater than that of strains containing other mutations. We also conclude that at low pH the environment is permissive for a wider spectrum of mutations, which may provide opportunities for a successful mutant to survive

Coastal fog detection using visual sensing

Use of visual sensing techniques to detect low visibility conditions may have a number of advantages when combined with other methods, such as satellite based remote sensing, as data can be collected and processed in real or near real time. Camera-enabled visual sensing can provide direct confirmation of modelling and forecasting results. Fog detection, modelling and prediction are a priority for maritime communities and coastal cities due to economic impacts of fog on aviation, marine, and land transportation. Canadian and Irish coasts are particularly vulnerable to dense fog under certain environmental conditions. Offshore oil and gas production on Grand Bank (off the Canadian East Coast) can be adversely affected by weather and sea state conditions. In particular, fog can disrupt the transfer of equipment and people to/from the production platforms by helicopter. Such disruptions create delays and the delays cost money. According to offshore oil and gas industry representatives at a recent workshop on metocean monitoring and forecasting for the NL offshore, there is a real need for improved forecasting of visibility (fog) out to 3 days. The ability to accurately forecast future fog conditions would improve the industry’s ability to adjust its schedule of operations accordingly. In addition, it was recognized by workshop participants that the physics of Grand Banks fog formation is not well understood, and that more and better data are needed

Conjugative Transposons and Their Cargo Genes Vary across Natural Populations of Rickettsia buchneri Infecting the Tick Ixodes scapularis

Rickettsia buchneri (formerly Rickettsia endosymbiont of Ixodes scapularis, or REIS) is an obligate intracellular endoparasite of the black-legged tick, the primary vector of Lyme disease in North America. It is noteworthy among the rickettsiae for its relatively large genome (1.8 Mb) and extraordinary proliferation of mobile genetic elements (MGEs), which comprise nearly 35% of its genome. Previous analysis of the R. buchneri genome identified several integrative conjugative elements named Rickettsiales amplified genomic elements (RAGEs); the composition of these RAGEs suggests that continued genomic invasions by MGEs facilitated the proliferation of rickettsial genes related to an intracellular lifestyle. In this study, we compare the genomic diversity at RAGE loci among sequenced rickettsiae that infect three related Ixodes spp., including two strains of R. buchneri and Rickettsia endosymbiont of Ixodes pacificus strain Humboldt, as well as a closely related species R. tamurae infecting Amblyomma testudinarium ticks. We further develop a novel multiplex droplet digital PCR assay and use it to quantify copy number ratios of chromosomal R. buchneri RAGE-A and RAGE-B to the single-copy gene gltA within natural populations of I. scapularis. Our results reveal substantial diversity among R. buchneri at these loci, both within individual ticks as well as in the I. scapularis population at large, demonstrating that genomic rearrangement of MGEs is an active process in these intracellular bacteria

Streamlining analysis methods for large acoustic surveys using automatic detectors with operator validation

1.  Passive acoustic surveys are becoming increasingly popular as a means of surveying for cetaceans and other marine species. These surveys yield large amounts of data, the analysis of which is time consuming and can account for a substantial proportion of the survey budget. Semi-automatic processes enable the bulk of processing to be conducted automatically while allowing analyst time to be reserved for validating and correcting detections and classifications. 2.  Existing modules within the Passive Acoustic Monitoring software PAMGuard were used to process a large (25.4 Terabyte) dataset collected during towed acoustic ship transits. The recently developed ‘Multi-Hypothesis Tracking Click Train Detector’ and the ‘Whistle and Moan Detector’ modules were used to identify occasions within the dataset at which vocalising toothed whales (odontocetes) were likely to be acoustically present. These putative detections were then reviewed by an analyst, with false positives being corrected. Target motion analysis provided a perpendicular distance to odontocete click events enabling the estimation of detection functions for both sperm whales and delphinids. Detected whistles were assigned to the lowest taxonomical level possible using the PAMGuard ‘Whistle Classifier’ module. 3.  After an initial tuning process, this semi-automatic method required 91 hr of an analyst's time to manually review both automatic click train and whistle detections from 1,696 hr of survey data. Use of the ‘Multi-Hypothesis Tracking Click Train Detector’ reduced the amount of data for the analyst to search by 74.5%, while the ‘Whistle and Moan Detector’ reduced data to search by 85.9%. In total, 443 odontocete groups were detected, of which 55 were from sperm whale groups, six were from beaked whales, two were from porpoise and the remaining 380 were identified to the level of delphinid group. An effective survey strip half width of 3,277 and 699 m was estimated for sperm whales and delphinids respectively. 4.  The semi-automatic workflow proved successful, reducing the amount of analyst time required to process the data, significantly reducing overall project costs. The workflow presented here makes use of existing modules within PAMGuard, a freely available and open-source software, readily accessible to acoustic analysts.Publisher PDFPeer reviewe

Four-month moxifloxacin-based regimens for drug-sensitive tuberculosis

Supported by the Global Alliance for TB Drug Development with support from the Bill and Melinda Gates Foundation, the European and Developing Countries Clinical Trials Partnership, U.S. Agency for International Development, U.K. Department for International Development, Directorate General for International Cooperation of the Netherlands, Irish Aid, Australia Department of Foreign Affairs and Trade, and National Institutes of Health, AIDS Clinical Trials Group and by grants from the National Institute of Allergy and Infectious Diseases (NIAID) (UM1AI068634, UM1 AI068636, and UM1AI106701) and by NIAID grants to the University of KwaZulu Natal, South Africa, AIDS Clinical Trials Group (ACTG) site 31422 (1U01AI069469); to the Perinatal HIV Research Unit, Chris Hani Baragwanath Hospital, South Africa, ACTG site 12301 (1U01AI069453); and to the Durban International Clinical Trials Unit, South Africa, ACTG site 11201 (1U01AI069426); Bayer Healthcare for the donation of moxifloxacin; and Sanofi for the donation of rifampin.Background: Early-phase and preclinical studies suggest that moxifloxacin-containing regimens could allow for effective 4-month treatment of uncomplicated, smear-positive pulmonary tuberculosis. Methods: We conducted a randomized, double-blind, placebo-controlled, phase 3 trial to test the noninferiority of two moxifloxacin-containing regimens as compared with a control regimen. One group of patients received isoniazid, rifampin, pyrazinamide, and ethambutol for 8 weeks, followed by 18 weeks of isoniazid and rifampin (control group). In the second group, we replaced ethambutol with moxifloxacin for 17 weeks, followed by 9 weeks of placebo (isoniazid group), and in the third group, we replaced isoniazid with moxifloxacin for 17 weeks, followed by 9 weeks of placebo (ethambutol group). The primary end point was treatment failure or relapse within 18 months after randomization. Results: Of the 1931 patients who underwent randomization, in the per-protocol analysis, a favorable outcome was reported in fewer patients in the isoniazid group (85%) and the ethambutol group (80%) than in the control group (92%), for a difference favoring the control group of 6.1 percentage points (97.5% confidence interval [CI], 1.7 to 10.5) versus the isoniazid group and 11.4 percentage points (97.5% CI, 6.7 to 16.1) versus the ethambutol group. Results were consistent in the modified intention-to-treat analysis and all sensitivity analyses. The hazard ratios for the time to culture negativity in both solid and liquid mediums for the isoniazid and ethambutol groups, as compared with the control group, ranged from 1.17 to 1.25, indicating a shorter duration, with the lower bounds of the 95% confidence intervals exceeding 1.00 in all cases. There was no significant difference in the incidence of grade 3 or 4 adverse events, with events reported in 127 patients (19%) in the isoniazid group, 111 (17%) in the ethambutol group, and 123 (19%) in the control group. Conclusions: The two moxifloxacin-containing regimens produced a more rapid initial decline in bacterial load, as compared with the control group. However, noninferiority for these regimens was not shown, which indicates that shortening treatment to 4 months was not effective in this setting. (Funded by the Global Alliance for TB Drug Development and others; REMoxTB ClinicalTrials.gov number, NCT00864383.)Publisher PDFPeer reviewe

Gamma Delta TCR and the WC1 Co-Receptor Interactions in Response to Leptospira Using Imaging Flow Cytometry and STORM

The WC1 cell surface family of molecules function as hybrid gamma delta (gamma delta) TCR co-receptors, augmenting cellular responses when cross-linked with the TCR, and as pattern recognition receptors, binding pathogens. It is known that following activation, key tyrosines are phosphorylated in the intracytoplasmic domains of WC1 molecules and that the cells fail to respond when WC1 is knocked down or, as shown here, when physically separated from the TCR. Based on these results we hypothesized that the colocalization of WC1 and TCR will occur following cellular activation thereby allowing signaling to ensue. We evaluated the spatio-temporal dynamics of their interaction using imaging flow cytometry and stochastic optical reconstruction microscopy. We found that in quiescent gamma delta T cells both WC1 and TCR existed in separate and spatially stable protein domains (protein islands) but after activation using Leptospira, our model system, that they concatenated. The association between WC1 and TCR was close enough for fluorescence resonance energy transfer. Prior to concatenating with the WC1 co-receptor, gamma delta T cells had clustering of TCR-CD3 complexes and exclusion of CD45. gamma delta T cells may individually express more than one variant of the WC1 family of molecules and we found that individual WC1 variants are clustered in separate protein islands in quiescent cells. However, the islands containing different variants merged following cell activation and before merging with the TCR islands. While WC1 was previously shown to bind Leptospira in solution, here we showed that Leptospira bound WC1 proteins on the surface of gamma delta T cells and that this could be blocked by anti-WC1 antibodies. In conclusion, gamma delta TCR, WC1 and Leptospira interact directly on the gamma delta T cell surface, further supporting the role of WC1 in gamma delta T cell pathogen recognition and cellular activation