The genome packaging machinery of dsDNA bacteriophage PRD1

Viral genomes are encapsidated within protective protein shells. This encapsidation can be achieved either by a co-condensation reaction of the nucleic acid and coat proteins, or by first forming empty viral particles which are subsequently packaged with nucleic acid, the latter mechanism being typical for many dsDNA bacteriophages. Bacteriophage PRD1 is an icosahedral, non-tailed dsDNA virus that has an internal lipid membrane, the hallmark of the Tectiviridae family. Although PRD1 has been known to assemble empty particles into which the genome is subsequently packaged, the mechanism for this has been unknown, and there has been no evidence for a separate packaging vertex, similar to the portal structures used for packaging in the tailed bacteriophages and herpesviruses. In this study, a unique DNA packaging vertex was identified for PRD1, containing the packaging ATPase P9, packaging factor P6 and two small membrane proteins, P20 and P22, extending the packaging vertex to the internal membrane. Lack of small membrane protein P20 was shown to totally abolish packaging, making it an essential part of the PRD1 packaging mechanism. The minor capsid proteins P6 was shown to be an important packaging factor, its absence leading to greatly reduced packaging efficiency. An in vitro DNA packaging mechanism consisting of recombinant packaging ATPase P9, empty procapsids and mutant PRD1 DNA with a LacZ-insert was developed for the analysis of PRD1 packaging, the first such system ever for a virus containing an internal membrane. A new tectiviral sequence, a linear plasmid called pBClin15, was identified in Bacillus cereus, providing material for sequence analysis of the tectiviruses. Analysis of PRD1 P9 and other putative tectiviral ATPase sequences revealed several conserved sequence motifs, among them a new tectiviral packaging ATPase motif. Mutagenesis studies on PRD1 P9 were used to confirm the significance of the motifs. P9-type putative ATPase sequences carrying a similar sequence motif were identified in several other membrane containing dsDNA viruses of bacterial, archaeal and eukaryotic hosts, suggesting that these viruses may have similar packaging mechanisms. Interestingly, almost the same set of viruses that were found to have similar putative packaging ATPases had earlier been found to share similar coat protein folds and capsid structures, and a common origin for these viruses had been suggested. The finding in this study of similar packaging proteins further supports the idea that these viruses are descendants of a common ancestor.

Herpes simplex virus DNA packaging sequences adopt novel structures that are specifically recognized by a component of the cleavage and packaging machinery

The product of the herpes simplex virus type 1 UL28 gene is essential for cleavage of concatemeric viral DNA into genome-length units and packaging of this DNA into viral procapsids. To address the role of UL28 in this process, purified UL28 protein was assayed for the ability to recognize conserved herpesvirus DNA packaging sequences. We report that DNA fragments containing the pac1 DNA packaging motif can be induced by heat treatment to adopt novel DNA conformations that migrate faster than the corresponding duplex in nondenaturing gels. Surprisingly, these novel DNA structures are high-affinity substrates for UL28 protein binding, whereas double-stranded DNA of identical sequence composition is not recognized by UL28 protein. We demonstrate that only one strand of the pac1 motif is responsible for the formation of novel DNA structures that are bound tightly and specifically by UL28 protein. To determine the relevance of the observed UL28 protein–pac1 interaction to the cleavage and packaging process, we have analyzed the binding affinity of UL28 protein for pac1 mutants previously shown to be deficient in cleavage and packaging in vivo. Each of the pac1 mutants exhibited a decrease in DNA binding by UL28 protein that correlated directly with the reported reduction in cleavage and packaging efficiency, thereby supporting a role for the UL28 protein–pac1 interaction in vivo. These data therefore suggest that the formation of novel DNA structures by the pac1 motif confers added specificity on recognition of DNA packaging sequences by the UL28-encoded component of the herpesvirus cleavage and packaging machinery.

Electro-magnetically Actuated Minute Polymer Pump Fabricated using Packaging Technology

Design, fabrication and preliminary testing of a flat pump with millimetre thickness are described in this paper. The pump is entirely made of polymer materials barring the magnet and copper coils used for electromagnetic actuation. The fabrication is carried out using widely available microelectronic packaging machinery and techniques. Therefore, the fabrication of the pump is straightforward and inexpensive. Two types of prototypes are designed and built. One consists of copper coils that are etched on an epoxy plate and the other has wound insulated wire of 90 mu m diameter to serve as a coil. The overall size of the first pump is 25 mm x 25 mm x 3.6 mm including the 3.1 mm-thick NdFeB magnet of diameter 12 mm. It consists of a pump chamber of 20 mm x 20 mm x 0.8 mm with copper coils etched from a copper-clad epoxy plate using dry-film lithography and milled using a CNC milling machine, two passive valves and the pump-diaphragm made of Kapton film of 0.089 mm thickness. The second pump has an overall size of 35 mm x 35 mm x 4.4 mm including the magnet and the windings. A breadboard circuit and DC power supply are used to test the pump by applying an alternating square-wave voltage pulse. A water slug in a tube attached to the inlet is used to observe and measure the air-flow induced by the pump against atmospheric pressure. The maximum flow rate was found to be 15 ml/min for a voltage of 2.5 V and a current of 19 mA at 68 Hz.

Automatització d’una màquina de tall rotatiu

Molt sovint, a l’indústria de l’alimentació o de la cosmètica, hi ha promocions on s’afegeixen productes de regal; aquests els embalen amb un sleever (tipus de plàstic força rígid) que després retractilen. Aquestes promocions solen ser esporàdiques i canviants i això fa que no sigui factible la modificació de la línia de producció general per a realitzar-les. Per aquest motiu, moltes empreses disposen d’una petita màquina de tall rotatiu per a poder tallar els sleevers ells mateixos, i així tallar a molta velocitat i precisió aquestes petites produccions. Així doncs, esdevé molt interessant l’adquisició d’aquest tipus de maquinària. L’objectiu del projecte és fer l’automatització d’una màquina que pugui tallar aquest tipus de material ràpidament per fer-lo rentable

Modulare Logistiksysteme in der Prozessindustrie unter Einbeziehung von Industrie 4.0

Zur Gewährleistung einer kundenindividuellen, flexiblen Produktion, welche auch kleine Losegrößen kostengünstig produzieren kann, werden zukünftig Fabriken der Prozessindustrie nicht nur modular sondern darüber hinaus mobil sein. Zur Erreichung dieses Zieles werden neben Produktionselemente, auch sämtliche Logistikkomponenten in Containern zu Modulen zusammengefasst. Hierfür ist es notwendig Lager, aber auch komplexe Verpackungsmaschinen, zu konzipieren welche sich zusammen mit Produktionsmodulen, nach dem Plug-and-Produce-Prinzip, zu kompletten Produktionslinien verknüpfen lassen. Die Steuerung und Organisation erfolgt nach den Gedanken von Industrie 4.0.

Convergent evolution of PICIs-mediated phage interference

Staphylococcal pathogenicity islands (SaPIs), the prototype members of the family of phage inducible chromosomal islands (PICIs), are extremely mobile phage satellites, which are transferred between bacterial hosts after their induction by a helper phage. The intimate relationship between SaPIs and their helper phages is one of the most studied examples of virus satellite interactions in prokaryotic cells. SaPIs encode and disseminate virulence and fitness factors, representing a driving force for bacterial adaptation and pathogenesis. Many SaPIs encode a conserved morphogenetic operon, including a core set of genes whose function allows them to parasitize and exploit the phage life cycle. One of the central mechanisms of this molecular piracy is the specific packaging of the SaPI genomes into reduced sized capsid structures derived from phage proteins. Pac phages were classically thought to be the only phages involved in the mobilisation of phage-mediated virulence genes, including the transfer of SaPIs within related and non-related bacteria. This study presents the involvement of S. aureus cos phages in the intra- and intergeneric transfer of cos SaPIs for the first time. A novel example of molecular parasitism is shown, by which this newly characterised group of cos SaPIs uses two distinct and complementary mechanisms to take over the helper phage packaging machinery for their own reproduction. SaPIbov5, the prototype of the cos SaPIs, does not encode the characteristic morphogenetic operon found in pac SaPIs. However, cos SaPIs features both pac and cos phage cleavage sequences in their genome, ensuring SaPI packaging in small- and full-sized phage particles, depending on the helper phage. Moreover, cos-site packaging in S. aureus was shown to require the activity of a phage HNH nuclease. The HNH protein functions together with the large terminase subunit, triggering cleavage and melting of the cos-site sequence. In addition, a novel piracy strategy, severely interfering with the helper phage reproduction, was identified in cos SaPIs and characterised. This mechanism of piracy depends on the cos SaPI-encoded ccm gene, which encodes a capsid protein involved in the formation of small phage particles, modifying the assembling process via a scaffolding mechanism. This strategy resembles the ones described for pac SaPIs and represents a remarkable example of convergent evolution. A further convergent mechanism of capsid size-reduction was identified and characterised for the Enterococcus faecalis EfCIV583 pathogenicity island, another member of the PICI family. In this case, the self-encoded CpmE conducts this molecular piracy through a putative scaffolding function. Similar to cos SaPIs, EfCIV583 carries the helper phage cleavage sequence in its genome enabling its mobilisation by the phage terminase complex. The results presented in this thesis show how two examples of non-related members of the PICI family follow the same evolutionary convergent strategy to interfere with their helper phage. These findings could indicate that the described strategies might be widespread among PICIs and implicate a significant impact of PICIs mediated-virulence gene transfer in bacterial evolution and the emergence of pathogenic bacteria.

Optimization and finite element analysis for reliable electronic packaging

This paper demonstrates a modeling and design approach that couples computational mechanics techniques with numerical optimisation and statistical models for virtual prototyping and testing in different application areas concerning reliability of eletronic packages. The integrated software modules provide a design engineer in the electronic manufacturing sector with fast design and process solutions by optimizing key parameters and taking into account complexity of certain operational conditions. The integrated modeling framework is obtained by coupling the multi-phsyics finite element framework - PHYSICA - with the numerical optimisation tool - VisualDOC into a fully automated design tool for solutions of electronic packaging problems. Response Surface Modeling Methodolgy and Design of Experiments statistical tools plus numerical optimisaiton techniques are demonstrated as a part of the modeling framework. Two different problems are discussed and solved using the integrated numerical FEM-Optimisation tool. First, an example of thermal management of an electronic package on a board is illustrated. Location of the device is optimized to ensure reduced junction temperature and stress in the die subject to certain cooling air profile and other heat dissipating active components. In the second example thermo-mechanical simulations of solder creep deformations are presented to predict flip-chip reliability and subsequently used to optimise the life-time of solder interconnects under thermal cycling.

Thermal, mechanical and optical modelling of VCSEL packaging

Hybrid OECB (Opto-Electrical Circuit Boards) are expected to make a significant impact in the telecomm switches arena within the next five years, creating optical backplanes with high speed point-to-point optical interconnects. OECB's incorporate short range optical interconnects, and are based on VCSEL (Vertical Cavity Surface Emitting Diode) and PD (Photo Diode) pairs, connected to each other via embedded waveguides in the OECB. The VCSEL device is flip-chip assembled onto an organic substrate with embedded optical waveguides. The performance of the VCSEL device is governed by the thermal, mechanical and optical characteristics of this assembly. During operation, the VCSEL device will heat up and the thermal change together with the CTE mismatch in the materials, will result in potential misalignment between the VCSEL apertures and the waveguide openings in the substrate. Any degree of misalignment will affect the optical performance of the package. This paper will present results from a highly coupled modelling analysis involving thermal, mechanical and optical models. The paper will also present results from an optimisation analysis based on Design of Experiments (DOE).

Optoelectronic packaging interfaces for submicron alignment (OPISA) and the dynamics of laser spot weld formation

The work presented in this paper is part of the OPISA project. This is a collaborative research project between the University of Greenwich and Bookham Technology. This report describes some of the initial work undertaken towards the goal of investigating optoelectronic packaging where alignment issues between optical sources and fibers can arise as part of the fabrication process. The focus of this study is on charting the dynamics of laser spot weld formation. This paper introduces some of the initial simulation work that has been undertaken and presents a model describing a transient heat source applied from a laser pulse to weld a stainless steel sleeve and ferrule and the resulting weld formation

Detection of faults in a high speed packaging machine using a multilayer perceptron (MLP)

In high speed manufacturing systems, continuous operation is desirable, with minimal disruption for repairs and service. An intelligent diagnostic monitoring system, designed to detect developing faults before catastrophic failure, or prior to undesirable reduction in output quality, is a good means of achieving this. Artificial neural networks have already been found to be of value in fault diagnosis of machinery. The aim here is to provide a system capable of detecting a number of faults, in order that maintenance can be scheduled in advance of sudden failure, and to reduce the necessity to replace parts at intervals based on mean time between failures. Instead, parts will need to be replaced only when necessary. Analysis of control information in the form of position error data from two servomotors is described.

Intelligent prognostics of machinery health utilising suspended condition monitoring data

The ability to forecast machinery failure is vital to reducing maintenance costs, operation downtime and safety hazards. Recent advances in condition monitoring technologies have given rise to a number of prognostic models for forecasting machinery health based on condition data. Although these models have aided the advancement of the discipline, they have made only a limited contribution to developing an effective machinery health prognostic system. The literature review indicates that there is not yet a prognostic model that directly models and fully utilises suspended condition histories (which are very common in practice since organisations rarely allow their assets to run to failure); that effectively integrates population characteristics into prognostics for longer-range prediction in a probabilistic sense; which deduces the non-linear relationship between measured condition data and actual asset health; and which involves minimal assumptions and requirements. This work presents a novel approach to addressing the above-mentioned challenges. The proposed model consists of a feed-forward neural network, the training targets of which are asset survival probabilities estimated using a variation of the Kaplan-Meier estimator and a degradation-based failure probability density estimator. The adapted Kaplan-Meier estimator is able to model the actual survival status of individual failed units and estimate the survival probability of individual suspended units. The degradation-based failure probability density estimator, on the other hand, extracts population characteristics and computes conditional reliability from available condition histories instead of from reliability data. The estimated survival probability and the relevant condition histories are respectively presented as “training target” and “training input” to the neural network. The trained network is capable of estimating the future survival curve of a unit when a series of condition indices are inputted. Although the concept proposed may be applied to the prognosis of various machine components, rolling element bearings were chosen as the research object because rolling element bearing failure is one of the foremost causes of machinery breakdowns. Computer simulated and industry case study data were used to compare the prognostic performance of the proposed model and four control models, namely: two feed-forward neural networks with the same training function and structure as the proposed model, but neglected suspended histories; a time series prediction recurrent neural network; and a traditional Weibull distribution model. The results support the assertion that the proposed model performs better than the other four models and that it produces adaptive prediction outputs with useful representation of survival probabilities. This work presents a compelling concept for non-parametric data-driven prognosis, and for utilising available asset condition information more fully and accurately. It demonstrates that machinery health can indeed be forecasted. The proposed prognostic technique, together with ongoing advances in sensors and data-fusion techniques, and increasingly comprehensive databases of asset condition data, holds the promise for increased asset availability, maintenance cost effectiveness, operational safety and – ultimately – organisation competitiveness.

Development of an online condition monitoring system for slow speed machinery

One of the main challenges of slow speed machinery condition monitoring is that the energy generated from an incipient defect is too weak to be detected by traditional vibration measurements due to its low impact energy. Acoustic emission (AE) measurement is an alternative for this as it has the ability to detect crack initiations or rubbing between moving surfaces. However, AE measurement requires high sampling frequency and consequently huge amount of data are obtained to be processed. It also requires expensive hardware to capture those data, storage and involves signal processing techniques to retrieve valuable information on the state of the machine. AE signal has been utilised for early detection of defects in bearings and gears. This paper presents an online condition monitoring (CM) system for slow speed machinery, which attempts to overcome those challenges. The system incorporates relevant signal processing techniques for slow speed CM which include noise removal techniques to enhance the signal-to-noise and peak-holding down sampling to reduce the burden of massive data handling. The analysis software works under Labview environment, which enables online remote control of data acquisition, real-time analysis, offline analysis and diagnostic trending. The system has been fully implemented on a site machine and contributing significantly to improve the maintenance efficiency and provide a safer and reliable operation.