Flux-sum analysis identifies metabolite targets for strain improvement.

BACKGROUND: Rational design of microbial strains for enhanced cellular physiology through in silico analysis has been reported in many metabolic engineering studies. Such in silico techniques typically involve the analysis of a metabolic model describing the metabolic and physiological states under various perturbed conditions, thereby identifying genetic targets to be manipulated for strain improvement. More often than not, the activation/inhibition of multiple reactions is necessary to produce a predicted change for improvement of cellular properties or states. However, as it is more computationally cumbersome to simulate all possible combinations of reaction perturbations, it is desirable to consider alternative techniques for identifying such metabolic engineering targets. RESULTS: In this study, we present the modified version of previously developed metabolite-centric approach, also known as flux-sum analysis (FSA), for identifying metabolic engineering targets. Utility of FSA was demonstrated by applying it to Escherichia coli, as case studies, for enhancing ethanol and succinate production, and reducing acetate formation. Interestingly, most of the identified metabolites correspond to gene targets that have been experimentally validated in previous works on E. coli strain improvement. A notable example is that pyruvate, the metabolite target for enhancing succinate production, was found to be associated with multiple reaction targets that were only identifiable through more computationally expensive means. In addition, detailed analysis of the flux-sum perturbed conditions also provided valuable insights into how previous metabolic engineering strategies have been successful in enhancing cellular physiology. CONCLUSIONS: The application of FSA under the flux balance framework can identify novel metabolic engineering targets from the metabolite-centric perspective. Therefore, the current approach opens up a new research avenue for rational design and engineering of industrial microbes in the field of systems metabolic engineering.

Software applications for flux balance analysis.

Flux balance analysis (FBA) is a widely used computational method for characterizing and engineering intrinsic cellular metabolism. The increasing number of its successful applications and growing popularity are possibly attributable to the availability of specific software tools for FBA. Each tool has its unique features and limitations with respect to operational environment, user-interface and supported analysis algorithms. Presented herein is an in-depth evaluation of currently available FBA applications, focusing mainly on usability, functionality, graphical representation and inter-operability. Overall, most of the applications are able to perform basic features of model creation and FBA simulation. COBRA toolbox, OptFlux and FASIMU are versatile to support advanced in silico algorithms to identify environmental and genetic targets for strain design. SurreyFBA, WEbcoli, Acorn, FAME, GEMSiRV and MetaFluxNet are the distinct tools which provide the user friendly interfaces in model handling. In terms of software architecture, FBA-SimVis and OptFlux have the flexible environments as they enable the plug-in/add-on feature to aid prospective functional extensions. Notably, an increasing trend towards the implementation of more tailored e-services such as central model repository and assistance to collaborative efforts was observed among the web-based applications with the help of advanced web-technologies. Furthermore, most recent applications such as the Model SEED, FAME, MetaFlux and MicrobesFlux have even included several routines to facilitate the reconstruction of genome-scale metabolic models. Finally, a brief discussion on the future directions of FBA applications was made for the benefit of potential tool developers.

Milwaukee Laboratory System Improvement Program (L-SIP).

The Laboratory System Improvement Program (L-SIP) of the Association of Public Health Laboratories aims to improve state public health laboratory (PHL) system performance through continuous quality improvement. We successfully applied this state assessment tool to a local PHL (LPHL) system by tailoring it to reflect local system needs and created an LPHL system definition explaining how a local system differs from, yet complements, a state system. On November 18, 2010, 75 stakeholders from 40 agencies assessed the Milwaukee, Wisconsin, PHL system, capturing themes, strengths and weaknesses of the system, and scores for each of the 10 Essential Public Health Services. A Laboratory Advisory Committee analyzed assessment results to identify a strategic focus of research and workforce development and define an action plan, which is now being carried out. Milwaukee's L-SIP process is effectively improving LPHL system research and workforce development while raising community awareness of the system.

Tuning the surface charge properties of epitaxial InN nanowires.

We have investigated the correlated surface electronic and optical properties of [0001]-oriented epitaxial InN nanowires grown directly on silicon. By dramatically improving the epitaxial growth process, we have achieved, for the first time, intrinsic InN both within the bulk and at nonpolar InN surfaces. The near-surface Fermi-level was measured to be ∼0.55 eV above the valence band maximum for undoped InN nanowires, suggesting the absence of surface electron accumulation and Fermi-level pinning. This result is in direct contrast to the problematic degenerate two-dimensional electron gas universally observed on grown surfaces of n-type degenerate InN. We have further demonstrated that the surface charge properties of InN nanowires, including the formation of two-dimensional electron gas and the optical emission characteristics can be precisely tuned through controlled n-type doping. At relatively high doping levels in this study, the near-surface Fermi-level was found to be pinned at ∼0.95-1.3 eV above the valence band maximum. Through these trends, well captured by the effective mass and ab initio materials modeling, we have unambiguously identified the definitive role of surface doping in tuning the surface charge properties of InN.

Mesh erosion following laparoscopic incisional hernia repair.

Laparascopic mesh repair is a safe and effective method of surgically treating incisional hernia. However, such an approach may lead to specific complications of both laparoscopy and mesh placement. The mesh may migrate, become infected or erode into adjacent structures. We describe the case of a woman who underwent laparoscopic incisional hernia repair with subsequent erosion of the mesh into the bladder.

NADPH-dependent pgi-gene knockout Escherichia coli metabolism producing shikimate on different carbon sources.

We explored the physiological and metabolic effects of different carbon sources (glucose, fructose, and glucose/fructose mixture) in phosphoglucose isomerase (pgi) knockout Escherichia coli mutant producing shikimic acid (SA). It was observed that the pgi(-) mutant grown on glucose exhibited significantly lower cell growth compared with the pgi(+) strain and its mixed glucose/fructose fermentation grew well. Interestingly, when fructose was used as a carbon source, the pgi(-) mutant showed the enhanced SA production compared with the pgi(+) strain. In silico analysis of a genome-scale E. coli model was then conducted to characterize the cellular metabolism and quantify NAPDH regeneration, which allowed us to understand such experimentally observed attenuated cell growth and enhanced SA production in glucose- and fructose-consuming pgi(-) mutant, respectively with respect to cofactor regeneration.

Ionicity in disordered GeSe2: a comparison of first-principles and atomistic potential models.

The structural properties of liquid GeSe(2), generated using two distinct computational methodologies, are compared. The results of molecular dynamics simulations, utilizing both first-principles density functional and a potential model which account for aspects of many-body interactions, are considered. The potential model favors ionic character in the bonding, resulting in a structure with very little chemical disorder and no homopolar bonds, in contrast to experimental observation. The use of a relatively simple potential model is shown to be useful in order to understand differences between the observed experimental structure and those obtained from the first-principles approach, the latter being affected by insufficient account of ionic character in the bonding. Both computational schemes are able to predict the appearance of the first sharp diffraction peak in the total neutron structure factor and in some of the partial structure factors as well as the concomitant presence of corner- and edge-sharing tetrahedral connections. For the potential model, this holds true provided the system temperatures are set to values high enough to allow for diffusion properties typical of a liquid. Structural properties obtained for the two sets of configurations are in closer agreement when the potential model is applied at very high temperatures.

The evolution of intermediate-range order in molten network-forming materials.

The atomistic origin of the intermediate-range order (IRO) is investigated for an archetypal network-forming liquid. A pairwise additive potential model is chosen which is augmented with a description of the (many-body) anion polarization. The anion polarizability and system temperature are both systematically varied in order to control the network topology. The change in the IRO is monitored via the construction of Bhatia-Thornton structure factors which highlight the effect of chemical composition and network topology. The atomistic origin of the first-sharp diffraction peak in the concentration-concentration function, S(CC)(k), is discussed in terms of the connectivity of the polyhedral network. The atomistic origin of the IRO is discussed by reference to previous analyses.

Stereotactic mammography imaging combined with 3D US imaging for image guided breast biopsy.

Stereotactic X-ray mammography (SM) and ultrasound (US) guidance are both commonly used for breast biopsy. While SM provides three-dimensional (3D) targeting information and US provides real-time guidance, both have limitations. SM is a long and uncomfortable procedure and the US guided procedure is inherently two dimensional (2D), requiring a skilled physician for both safety and accuracy. The authors developed a 3D US-guided biopsy system to be integrated with, and to supplement SM imaging. Their goal is to be able to biopsy a larger percentage of suspicious masses using US, by clarifying ambiguous structures with SM imaging. Features from SM and US guided biopsy were combined, including breast stabilization, a confined needle trajectory, and dual modality imaging. The 3D US guided biopsy system uses a 7.5 MHz breast probe and is mounted on an upright SM machine for preprocedural imaging. Intraprocedural targeting and guidance was achieved with real-time 2D and near real-time 3D US imaging. Postbiopsy 3D US imaging allowed for confirmation that the needle was penetrating the target. The authors evaluated 3D US-guided biopsy accuracy of their system using test phantoms. To use mammographic imaging information, they registered the SM and 3D US coordinate systems. The 3D positions of targets identified in the SM images were determined with a target localization error (TLE) of 0.49 mm. The z component (x-ray tube to image) of the TLE dominated with a TLEz of 0.47 mm. The SM system was then registered to 3D US, with a fiducial registration error (FRE) and target registration error (TRE) of 0.82 and 0.92 mm, respectively. Analysis of the FRE and TRE components showed that these errors were dominated by inaccuracies in the z component with a FREz of 0.76 mm and a TREz of 0.85 mm. A stereotactic mammography and 3D US guided breast biopsy system should include breast compression for stability and safety and dual modality imaging for target localization. The system will provide preprocedural x-ray mammography information in the form of SM imaging along with real-time US imaging for needle guidance to a target. 3D US imaging will also be available for targeting, guidance, and biopsy verification immediately postbiopsy.

Syndromic surveillance using regional emergency medicine internet.

STUDY OBJECTIVE: We demonstrate the feasibility and utility of emergency department (ED) syndromic surveillance using a regional emergency medicine Internet application to minimize impact on ED and public health staffing. METHODS: Regional (multi-ED) surveillance was established for 2 periods, one characterized by a high-profile national sports event and the other during an international disease outbreak. Counts of patient visits meeting syndrome criteria and total patient visits were reported daily on the secure regional emergency medicine Internet site and downloaded by public health staff. Trends were analyzed and displayed on the secure Web site. ED participants were surveyed about the acceptability and time cost of the project. RESULTS: In the first ("All Star Game") project, 8 departments reported daily counts for 4 weeks, covering more than 26,000 patient visits. In the second ("severe acute respiratory syndrome" [SARS]) project, an average of 11 departments in the same region reported daily data on febrile respiratory illnesses, travel, and contacts for 10 weeks. Experience with the first project allowed for rapid implementation of the second project during a 3-day period. In both instances, the surveillance efforts were undertaken without the need for extraordinary ED or public health staffing requirements. CONCLUSION: A regional emergency medicine Internet approach permitted rapid implementation of multisite syndromic surveillance without additional staff. Some problems were identified with the first project, related to clinician checklist completion and manual data tabulation and entry. The SARS project addressed these by simplifying data collection and restricting it to triage.

The public health dashboard: a surveillance model for bioterrorism preparedness.

The City of Milwaukee Health Department piloted a short-term, near real-time syndromic surveillance and communication tool by using an existing secure regional Internet infrastructure. Voluntary, active syndromic case reporting by hospital Emergency Departments was combined with other data streams, including clinical laboratory reports of communicable disease, hospital emergency room diversions, ambulance runs, medical examiner reports of unusual or suspicious deaths, poison control and nursing hotline call volumes, and pharmacy over-the-counter sales. These data were aggregated into a "Surveillance Dashboard" format that was used to communicate community syndromic health trends to hospitals, Emergency Departments, and other providers using a secure Internet technology. Emergency Departments at 8 area hospitals reported a total of 314 cases meeting syndromic criteria from 26,888 patient encounters. Participants were satisfied with data entry and communications. All participating Emergency Departments received e-mail and text pager alerts sent by the Milwaukee Health Department. No unexplained findings or suggestions of an early outbreak were reported through syndrome surveillance for the 4-week duration of the project. Similar surveillance and communications systems could provide multiple benefits to Emergency Department workflow and management, as well as to public health and emergency response.

Three Hundred Sixty Degree Feedback: program implementation in a local health department.

Three Hundred Sixty Degree Feedback systems, while popular in business, have been less commonly implemented in local public health agencies. At the same time, they are effective methods of improving employee morale, work performance, organizational culture, and attainment of desired organizational outcomes. These systems can be purchased "off-the-shelf," or custom applications can be developed for a better fit with unique organizational needs. We describe the City of Milwaukee Health Department's successful experience customizing and implementing a 360-degree feedback system in the context of its ongoing total quality improvement efforts.

Developmental language impairments with complex origins: learning from twins and multiple birth children.

Various factors that make language development vulnerable in twins and multiple birth children are discussed. Researchers have investigated single explanations for language impairment in twins. However, this paper argues that in individual cases the language impairment is more often the outcome of a complex interaction of factors. Some factors play a greater part than others in some cases. It is suggested that clinicians can develop a dynamic model of language acquisition from understanding the effect of multiple birth and other exceptional circumstances on language acquisition. This model is needed to underpin investigations into individual cases of language impairment. An approach to assessment is described that identifies factors operating in individual cases of multiple birth, providing an essential precursor to implementing appropriate case-based intervention.

Can phonological awareness training facilitate minimal pair therapy?

This study evaluated a new therapy approach. 18 children with phonological disorders resistant to conventional therapy had phonological awareness training followed by conventional speech therapy. The children's speech production improved as did their phonological awareness. In this heterogeneous group, the patterns revealed in their individual responses to therapy proved diagnostically significant.

Seeing and hearing rotated faces: influences of facial orientation on visual and audiovisual speech recognition.

It is well-known that facial orientation affects the processing of static facial information, but similar effects on the processing of visual speech have yet to be explored fully. Three experiments are reported in which the effects of facial orientation on visual speech processing were examined using a talking face presented at 8 orientations through 360 degrees. Auditory and visual forms of the syllables /ba/, /bi/, /ga/, /gi/, /ma/, /mi/, /ta/, and /ti/ were used to produce the following speech stimulus types: auditory, visual, congruent audiovisual, and incongruent audiovisual. Facial orientation did not affect identification of visual speed per se or the near-perfect accuracy of auditory speech report with congruent audiovisual speech stimuli. However, facial orientation did affect the accuracy of auditory speech report with incongruent audiovisual speech stimuli. Moreover, the nature of this effect depended on the type of incongruent visual speech used. Implications for the processing of visual and audiovisual speech are discussed.

From rodent glial precursor cell to human glial neoplasia in the oligodendrocyte-type-2 astrocyte lineage.

With only a few exceptions, the precursor cells representing the normal counterparts of human tumours are unknown. The comparative lack of information about the lineages involved in tissue development, and difficulties in growing many human tumors in a manner suitable for cellular biological analysis, together often make it difficult to study the differences between normal and tumor cells and to develop many of the model systems that would be useful in the study of human cancer. By applying techniques previously utilized to study glial progenitor cells, we have isolated a human glioblastoma multiforme (GBM)-derived population that expresses many properties otherwise uniquely expressed by oligodendrocyte-type-2 astrocyte (O-2A) progenitor cells. Hu-O-2A/Gb1 (for Human O-2A lineage Glioblastoma number 1) cells responded to similar mitogens and differentiation modulators as rodent O-2A progenitors, and generated cells with features of precursor cells, oligodendrocytes and astrocytes. Moreover, 1H-NMR analysis of amino acid composition demonstrated a striking conversation of types and quantities of free amino acids between the human tumour cells and the rodent primary cells. Hu-O-2A/Gb1 cells represent the first human glioma-derived population for which unambiguous lineage assignment has been possible, and our results indicate that the human O-2A lineage can contribute to one of the most malignant of glial tumours. In addition, the highly diagnostic 1H-NMR spectrum expressed by Hu-O-2A/Gb1 cells raises the possibility of eventual non-invasive identification of tumors of this lineage.

Assessing conversational disability.

In spite of current interest in conversational disability, and the growing number of children who are being diagnosed as exhibiting this disorder, there are few satisfactory procedures that can be used to assess their conversation. In the following paper, data from the naturally occurring conversations of two children, who were identified as having difficulties in participating in conversation, are presented. The data were scanned for the following conversational skills: securing the attention of the addressee before speaking, initiating new topics of conversation effectively, using the grammatical forms that are most frequently associated with particular linguistic functions, filling response slots effectively, maintaining cohesion within the discourse, and taking steps to repair mishearings or misunderstandings. This kind of assessment, based on the principles of conversational analysis, provided an objective appraisal of the conversational skills of the children, and also highlighted potential areas for therapeutic intervention. Many of the conversational behaviours exhibited by the two subjects corresponded with those that have been observed in very young children who are developing language normally. It is suggested that the subjects studied here presented with a severe delay in their conversational development rather than a disorder.

Word superiority over isolated letters: the neglected case of forward masking.

Previous research shows that when briefly presented alphabetic stimuli are followed by pattern masks, letters in words are reported more accurately than are isolated letters (the "Word-Letter Phenomenon," or WLP); however, when these masks are replaced by blank fields, the WLP disappears. These findings have led to the popular notion that the WLP reflects selective masking of ongoing stimulus processing and so critically depends on the use of poststimulus masks. Here we report three experiments which re-examine the role of masking in the WLP by contrasting the effects of postmasked displays with the effects of premasked displays in which words and isolated letters were preceded by a pattern mask and followed by a completely blank field. Despite the critical role generally assigned to poststimulus pattern masks, similar WLPs were obtained with pre- and postmasked displays. Implications for theories of word and letter recognition are discussed.

Synthesis and evaluation of 3-substituted 17-methylmorphinan analogs as potential anticonvulsant agents.

Dextromethorphan (1,(+)-3-methoxy-17-methylmorphinan) demonstrates anticonvulsant activity in a variety of in vitro and in vivo models of convulsive action. It is well known that 1 is metabolized to its phenolic derivative dextrorphan (2) and this metabolite is also a potent anticonvulsant. A series of (+)-3-substituted-17-methylmorphinans, which are structurally similar to 1 but are either not expected to be metabolized to 2 or might do so at a reduced rate, as compared to 1, were prepared. Three analogs, 5 ((+)-3-amino-17-methylmorphinan), 14 ((+)-3-ethoxy-17-methylmorphinan), and 15 ((+)-3-(2-propoxy)-17-methylmorphinan) were found to possess potent anticonvulsant activity with full efficacy (ED50 25, 5.6, and 3.9 mg/kg, sc, respectively) in the rat supramaximal electroshock (MES) test. Binding potencies of these compounds to receptor sites labeled with [3H]dextromethorphan ([3H]1), in rat brain and guinea pig brain subcellular fractions, and [3H]thienylcyclohexylpiperidine (TCP) and [3H]glycine in rat brain, were determined. Most of the analogs displaced [3H]1 from its binding sites, with compounds 14 (IC50 0.42 microM) and 15 (IC50 0.88 microM) having equivalent potencies to 1 (IC50 0.59 microM), in rat brain, and no appreciable activity at the [3H]TCP or [3H]glycine-labeled sites. Compound 5 did not bind with appreciable activity to the [3H]1 site, in rat brain, but did bind to the [3H]TCP site with lower potency than the parent 1 (IC50 7.8 and 2.0 microM, respectively). The mechanism of anticonvulsant action of these agents is not clear although it appears that interaction at the [3H]1 sites may be involved.

Retinoblastoma gene deletions in human glioblastomas.

The retinoblastoma susceptibility gene, RB, is the best characterised of the tumour suppressor genes, or 'anti-oncogenes'. Abnormal function of the RB protein is thought to result in loss of an inhibitory effect on cell growth, and thus contribute towards the development of certain human cancers. One group of human cancers of particular interest in relationship to retinoblastoma gene function are the gliomas, which are central nervous system tumours thought to originate from the neuroectoderm, the embryological tissue which also gives rise to retinoblastomas. We have therefore examined a group of benign and malignant gliomas for evidence of structural alterations of the RB gene. Four out of nine (44%) glioblastomas, the most malignant gliomas, showed loss of heterozygosity of a locus within this gene. In addition, one of these hemizygous tumours showed deletion of part of the RB protein-coding region, and this abnormality was also present in cells cultured from the tumour. These findings suggest that RB gene abnormalities may contribute to the development of glioblastomas.