bluuebunny/crossref_metadata_2025 huggingface dataset:
A geometric approach to robust performance of parametric uncertain systems
Authors: Bondia J. , Picó J.
Published: November 2003

A new approach for the robust performance problem for parametric uncertain systems is presented. Contrary to the classical approach, where specifications must be given in the frequency domain, this approach allows to deal with classical time specifications such as bounds on the overshoot, settling time and steady state error, which are matched to an uncertain reference model. Controller synthesis is then formulated as a set inclusion problem with a clear geometrical interpretation. Copyright © 2003 John Wiley & Sons, Ltd.
ℋ︁∞filtering for discrete‐time linear systems with Markovian jumping parameters†
Authors: de Souza Carlos E. , Fragoso Marcelo D.
Published: November 2003

This paper investigates the problem of ℋ︁∞filtering for discrete‐time linear systems with Markovian jumping parameters. It is assumed that the jumping parameter is available. This paper develops necessary and sufficient conditions for designing a discrete‐time Markovian jump linear filter which ensures a prescribed bound on the ℓ2‐induced gain from the noise signals to the estimation error. The proposed filter design is given in terms of linear matrix inequalities. Copyright © 2003 John Wiley & Sons, Ltd.
Robust inverse optimal control laws for nonlinear systems
Authors: El‐Farra Nael H. , Christofides Panagiotis D.
Published: November 2003

This work proposes a robust inverse optimal controller design for a class of nonlinear systems with bounded, time‐varying uncertain variables. The basic idea is that of re‐shaping the scalar nonlinear gain of anLgVcontroller, based on Sontag's formula, so as to guarantee certain uncertainty attenuation properties in the closed‐loop system. The proposed gain re‐shaping is shown to yield a control law that enforces global boundedness of the closed‐loop trajectories, robust asymptotic output tracking with an arbitrary degree of attenuation of the effect of uncertainty on the output, and inverse optimality with respect to a meaningful cost that penalizes the tracking error and the control action. The performance of the control law is illustrated through a simulation example and compared with other controller designs. Copyright © 2003 John Wiley & Sons, Ltd.
Microglial activation state and lysophospholipid acid receptor expression
Authors: Tham Chui‐Se , Lin Fen‐Fen , Rao Tadimeti S. , Yu Naichen , Webb Michael
Published: November 2003

We used a simple commercial magnetic immunobead method for the preparation of acutely isolated microglial cells from postnatal days 1–3 rat brain. With the exception of a 15 min enzyme incubation, all stages are carried out at 4 °C, minimizing the opportunity for changes in gene expression during the isolation to be reflected in changes in accumulated mRNA. The composition of the isolated cells was compared with that of microglial cultures prepared by conventional tissue culture methods, and the purity of microglia was comparable between the two preparations. RT‐PCR analysis of several genes related to inflammatory products indicated that the acutely prepared cells were in a less activated condition than the conventionally tissue cultured cells. We examined the pattern of expression of receptors for lysophosphatidic acid (lpa) and sphingosine‐1‐phosphate (S1P) using quantitative real‐time PCR (TaqMan PCR) techniques. mRNA for LPA1, S1P1, S1P2, S1P3 and S1P5 was detected in these preparations, but the levels of the different receptor mRNAs varied according to the state of activation of the cells. mRNA for LPA3 was only detected significantly in cultured cell after lipopolysaccharide (LPS) stimulation, being almost absent in cultured microglia and undetectable in the acutely isolated preparations. The levels of mRNA of LPA1 and S1P receptors was reduced by overnight exposure to S1P, while the same treatment significantly up‐regulated the level of LPA3 mRNA.
Effects of habitat degradation on avian guilds in East African papyrus Cyperus papyrus swamps
Authors: Maclean Ilya M. D. , Hassall Mark , Boar Rosalind , Nasirwa Oliver
Published: November 2003

The density and species richness of bird communities in disturbed and undisturbed stands of papyrus Cyperus papyrus were compared. Point counts, corrected for different probabilities of detection in different swamps, suggested that the species richness of bird communities in stands of papyrus disturbed by burning, grazing or pollution was higher than in nearby stands that were not disturbed. However, there were fewer species and individuals of highly specialized birds or species characteristic of papyrus, in disturbed stands than in undisturbed swamps. At < 1 ha-1, the density of Papyrus Gonolek Laniarius mufumbiri in Ugandan swamps was, for example, significantly lower in polluted and burnt sites than in undisturbed papyrus where up to 13 ha-1 were recorded in the centre of swamps. White-winged Swamp Warbler Bradypterus carpalis was only recorded in undisturbed papyrus. In the papyrus that fringes Lake Naivasha in Kenya, outside the geographical range of species restricted to papyrus, disturbance in the form of grazing selects against species classified as swamp-reliant. Species classified as generalist users of papyrus were much less sensitive to disturbance than specialists. The density of swamp specialists was also lower near the edge of swamps, where total species richness was higher. These results are discussed in relation to conservation management of papyrus swamps in East Africa.
Rating expressed emotion from initial family therapy sessions (a pilot study)
Authors: Berkowitz Ruth
Published: November 2003

A small pilot study is described in which an attempt was made to rate expressed emotion (EE) from an initial family interview. It was found that ratings could be made and that all but one of the families were high EE. Both empirical and clinical implications are discussed.
The family of the severely handicapped child: the sibling perspective
Authors: Shulman Shmuel
Published: November 2003

The presence of a severely handicapped child is a source of prolonged crisis and stress for the child's family. Parents may redefine their roles and develop coping strategies aimed at responding to the needs of the handicapped child but which may hamper the development of well siblings. Two cases of siblings of severely handicapped children who developed adjustment difficulties are presented and discussed from a family system perspective.
An introduction to some of the ideas of Humberto Maturana
Authors: Leyland Maureen L.
Published: November 2003

In recent years the work of Humberto Maturana, a research biologist and neurophysiologist from the University of Santiago, Chile, has come to the forefront of dialogue between several well‐known family therapists. This paper introduces the reader to some of his original ideas and shows its connection with family therapy through linkages between it and some of the theoretical underpinnings of the Milan School
Families' medical myths
Authors: Hardwick Peter J.
Published: November 2003

A medical myth is defined as a false or distorted belief regarding issues of health within a family. This empirical study considers phenomenology, function and origins of medical myths operating in some forty families encountered in the author's psychiatric practice. Therapeutic strategies found useful in re‐editing the myths are described. Implications for the family/medical professional relationship are discussed.
Annotation: culture and family therapy
Authors: Hodes Matthew
Published: November 2003

This annotation considers the importance of understanding culture for the theory and practice of family therapy. A number of theoretical approaches are described. Some are concerned with ubiquitous and universal family processes like the family life cycle, while others are concerned with the particularity of family life such as the content of myths and rituals. Some practical implications for the participants of the therapy, and the choice of therapeutic approach is discussed.
Anorexia nervosa: the treatment of a male case with combined behavioural and family therapy
Authors: Carr Alan , McDonnell Dermot , Afnan Shahin
Published: November 2003

The successful treatment of a case of anorexia nervosa in a 14‐year‐old boy is described in this paper. The treatment comprised an initial month‐long hospital‐based behavioural weight gain programme. Concurrently a family evaluation was conducted. Family therapy, involving the parents only, was conducted over a subsequent four month period. The management of a relapse four months after the termination of treatment is described and information obtained at 9 and 16 month follow up is presented.
‘Why should we talk to you? You'll only tell the Court!’ On being an informer and a family therapist
Authors: Crowther Catherine , Dare Christopher , Wilson Judy
Published: November 2003

The family therapy literature recognizes the constraints imposed on a therapist's freedom and impartiality in a statutory agency. This paper shows that clinical settings that are apparently independent are not free of such constraints. The wider professional welfare system has confused expectations of a clinical agency that it should provide simultaneously both therapy and a measure of social control. An account is given of one hospital‐based family therapy team's struggles to find manoeuvrability in child‐focused statutory cases, so as to be helpful both to families and referrers. One method of working is illustrated with a case example.
Should I give advice? A systemic view
Authors: Silver Evril
Published: November 2003

Giving advice is considered from i he viewpoint of Milan systemic family therapy, and the possibility of its always being a therapeutic error is discussed. The beliefs of both professionals and clients concerning advice are examined. What can systemic therapists do if families ask for advice, and when is direct advice useful? Effects on therapists and families of giving advice are discussed as well as ways of limiting possible damage to the process of therapy. Questioning techniques are considered as an alternative that allows families to discover their own solutions.
The end‐of‐session message in systemic family therapy: a descriptive study
Authors: Allman Peter , Blochf Sidney , Sharpe Michael
Published: November 2003

A retrospective case note review was conducted with the aim of describing the end‐of‐session messages conveyed during Milan‐style systemic family therapy. Fifty consecutive families treated in an adult family clinic were included. A classification of messages was developed; for each type of message the mean number per therapy session was calculated and the rates compared both within and between four systemic categories of family. ‘Supportive/engaging’ messages were given more frequently than ‘hypothesis‐related’ messages in each systemic category and this difference was most marked in families with grief as the central issue. More ‘acknowledgement’ messages were used in the ‘grief’ group than in the ‘separation/individuation’ group. ‘Hypothesis‐related’ messages were used more often in the ‘separation/individuation’ group than in the ‘grief’ group. We discuss possible reasons for the observed patterns and compare different ways in which the message can be conceived and implemented.
Engineering an online course: applying the ‘secrets’ of computer programming to course development
Authors: Ellis Timothy J. , Hafner William
Published: November 2003

Colleges and universities are increasingly migrating towards utilising the World Wide Web to convey at least part of, and in many cases, their entire curricular offering. Despite this trend there is little support for the professors responsible for translating courses refined over a career in the classroom for delivery via the Web. Teachers who are experts in their subject area and masters of their craft when in a classroom find themselves in the uncomfortable position of having to relearn how to teach in a new environment with little or no support. Development of an online course is, in many significant aspects, analogous to developing a computer product. The procedures and tools utilised in the software engineering field to manage computer software development, therefore, offer promise for developing online courses. This paper explores the potential of one process developed for the software engineering field—the System Development Lifecycle (SDL)—as a tool to effectively design and develop online college courses.
Boys of Class, Boys of Color: Negotiating the Academic and Social Geography of an Elite Independent School
Authors: Kuriloff Peter , Reichert Michael C.
Published: November 2003

How do boys from diverse backgrounds manage in an elite boys' school? Interviewing a representative sample of 27 boys, blocked for race, class, and academic performance, we found that they navigated the school's academic geography by mastering “a drill” that included hard work, unwavering commitment, a will to win, a cool style, and self knowledge as learners. Some developed a transformative love of learning. But many marginalized boys struggled with the school's social geography. African American boys managed most effectively as they developed intra‐group discourses of race and class enabling them to take up the school's offers of “hegemonic habitus” without “selling out.” We discuss the liberating implications of helping students in both independent and public schools develop similar critiques.
Complex Subjectivities: Class, Ethnicity, and Race in Women's Narratives of Upward Mobility
Authors: Jones Sandra J.
Published: November 2003

This article engages the intersection of class, race, and ethnicity and the ways in which class marginality informs self and academic practice by drawing on research with ten female professors from the working class. Two phenomenological interviews were conducted with each participant as a primary source of data. Secondary sources included classroom visits and relevant documents. Interview transcripts were analyzed using a combination of grounded theory and narrative analysis. The author presents a narrative analysis of two stories told by participants to illustrate two distinct conceptualizations of class and ethnicity. Following this, she draws on grounded theory analysis to illustrate the ways in which class marginality informs academic practice. Implications of this work for educational institutions are discussed.
Class Notes: Toward a Critical Psychology of Class and Schooling
Authors: Fine Michelle , Burns April
Published: November 2003

In this epilogue, we offer a theoretical mapping of notions that have emerged across the articles in this issue of the Journal of Social Issues specifically dedicated to questions of social class. Social class is often included within the “race, class, gender, and sexual orientation” mantra of feminist and critical race work in psychology, but rarely scrutinized with rigor or serious scholarship. Thus, for the purposes of this epilogue, we theorize the relationship between the material, social, psychological, and the political. We identify four theoretical venues through which these researchers have opened a conversation about class and schooling: ideology, institutions, contradictions and consciousness, and method. We conclude by crafting a research agenda for a critical psychology of class and schooling.
Fixed nucleotide differences on the Y chromosome indicate clear divergence between Equus przewalskii and Equus caballus
Authors: Wallner B. , Brem G. , Müller M. , Achmann R.
Published: November 2003

The phylogenetic relationship between Equus przewalskii and E. caballus is often a matter of debate. Although these taxa have different chromosome numbers, they do not form monophyletic clades in a phylogenetic tree based on mtDNA sequences. Here we report sequence variation from five newly identified Y chromosome regions of the horse. Two fixed nucleotide differences on the Y chromosome clearly display Przewalski's horse and domestic horse as sister taxa. At both positions the Przewalski's horse haplotype shows the ancestral state, in common with the members of the zebra/ass lineage. We discuss the factors that may have led to the differences in mtDNA and Y‐chromosomal observations.
Detection of QTL for immune response to sheep red blood cells in laying hens
Authors: Siwek M. , Cornelissen S. J. B. , Nieuwland M. G. B. , Buitenhuis A. J. , Bovenhuis H. , Crooijmans R. P. M. A. , Groenen M. A. M. , de Vries‐Reilingh G. , Parmentier H. K. , Van Der Poel J. J.
Published: November 2003

The aim of this study is to detect quantitative trait loci (QTL) involved in the regulation of the primary and the secondary immune response to sheep red blood cells (SRBC) in a resource population using microsatellite DNA markers. The F2 resource population originates from a cross of two divergently selected lines for either high (H line) or low (L line) primary antibody response to SRBC. The F2 population consisted of six half‐sib families, three families per each of reciprocal crosses. Total antibody titres to SRBC were determined by agglutination in serum from all birds. F2, F1 and F0 generations were genotyped for 170 microsatellite markers, using a whole‐genome scan approach. The half‐sib and the line‐cross analyses were performed to determine QTL regions associated with regulation of the immune response. In the half‐sib analysis, four QTL for SRBC primary response have been identified: on GGA3, GGA5, GGA16 and GGA23. No QTL was identified for SRBC secondary response under the half‐sib model. In the line‐cross analysis, three QTL were identified on GGA10, GGA16 and GGA27 for SRBC primary response and five QTL were identified on GGA6, GGA9, GGA15, GGA16 and GGA27 for SRBC secondary response. Subsequently, the family contribution of individual families to the QTL was analysed. The family with the largest contribution was genotyped with additional microsatellite markers in the QTL region on GGA5. The extended half‐sib analysis with additional genotype information results in narrowing down the QTL region on GGA5.
Protective effect of exogenous administration of α‐tocopherol in middle cerebral artery occlusion model of cerebral ischemia in rats
Authors: Chaudhary Geeta , Sinha Kusum , Gupta Y.K.
Published: November 2003

Oxidative stress due to increased free radical generation and depletion of endogenous antioxidants have been implicated in ischemia–reperfusion‐induced neuronal injury. The present study was carried out to study the effect of acute administration of α‐tocopherol in the middle cerebral artery (MCA) occlusion model of stroke in rats. Rats were anesthetized using chloral hydrate (400 mg/kg i.p.) and subjected to 2 h of transient MCA occlusion. α‐Tocopherol was administered at the dose of 125 and 250 mg/kg orally 1 h prior to the occlusion of MCA. Motor performance test (grip test, foot fault test, rotarod performance test, spontaneous locomotor activity), markers of oxidative stress and 2,3,5‐triphenyl tetrazolium chloride staining were carried out 24 h after MCA occlusion. A vehicle‐treated group was run parallel. It was observed that α‐tocopherol at the dose of 125 mg/kg neither improved neurologic deficit, nor decreased the raised level of oxidative stress markers in comparison with the MCA‐occluded rats. However, higher dose of α‐tocopherol (250 mg/kg p.o.) afforded significant protection as evident by increase in motor performance tests and a decrease in the volume of infarction. The raised levels of malondialdehyde after MCA occlusion were also significantly attenuated. The results demonstrate that exogenous administration of α‐tocopherol is able to reduce the neuronal damage caused during ischemia–reperfusion, which can be attributed to its antioxidant activity.
The effect of pretreatments on the drying rate and quality of dried bananas
Authors: Dandamrongrak Rak , Mason Richard , Young Gordon
Published: November 2003

The effects that four pretreatments (blanching, chilling, freezing, and combined blanching and freezing), used prior to drying, had on the drying rate and quality of bananas were investigated. An untreated sample was used as a control. The bananas were dried at 50 °C in a heat pump dehumidifier dryer, using an air velocity of 3.1 m s−1, until a final moisture content of approximately 25% dry weight basis was attained. While the initial drying rate was highest for the blanched treatment, the two pretreatments involving freezing resulted in the shortest drying times. The blanched sample was most preferred in terms of colour while the frozen samples exhibited extensive browning. The texture and flavour was significantly (P < 0.05) reduced in all samples that involved blanching and/or freezing.
Are Pets a Healthy Pleasure? The Influence of Pets on Blood Pressure
Authors: Allen Karen
Published: November 2003

Pet owners often describe their pets as important and cherished family members who offer solace in times of stress. This article considers evidence suggesting that pets influence human blood pressure. Studies on this topic extend current research testing the hypothesis that having other people around in stressful times can buffer the negative consequences of stress. The existing data suggest that people perceive pets as important, supportive parts of their lives and that the presence of a pet is associated with significant cardiovascular benefits, among both people with normal blood pressure and those with high blood pressure. Studies about pets and blood pressure have examined both naturally occurring and randomly assigned pet ownership but are limited by their focus on responses to short-term, acute stress. Future prospective studies should explore the influence of pets on people at risk for cardiovascular disease and also consider explanatory mechanisms for the pet effect.
Preliminary linkage map of the Turkey (Meleagris gallopavo) based on microsatellite markers
Authors: Burt D. W. , Morrice D. R. , Sewalem A. , Smith J. , Paton I. R. , Smith E. J. , Bentley J. , Hocking P. M.
Published: November 2003

The turkey is an agriculturally important species for which, until now, there is no published genetic linkage map based on microsatellite markers – still the markers most used in the chicken and other farm animals. In order to increase the number of markers on a turkey genetic linkage map we decided to map new microsatellite sequences obtained from a GT‐enriched turkey genomic library. In different chicken populations more than 35–55% of microsatellites are polymorphic. In the turkey populations tested here, 43% of all turkey primers tested were found to be polymorphic, in both commercial and wild type turkeys. Twenty linkage groups (including the Z chromosome) containing 74 markers have been established, along with 37 other unassigned markers. This map will lay the foundations for further genetic mapping and the identification of genes and quantitative trait loci in this economically important species. Genome comparisons, based on genetic maps, with related species such as the chicken would then also be possible. All primer information, polymerase chain reaction (PCR) conditions, allele sizes and genetic linkage maps can be viewed at http://roslin.thearkdb.org/. The DNA is also available on request through the Roslin Institute.
Mapping of ESTs derived from pre‐implantation stage cattle embryos to allocate 16 new additions to the ordered comparative map of cattle and human
Authors: Mamo S. , Wimmers K. , Gilles M. , Kata S. R. , Schellander K. , Ponsuksili S.
Published: November 2003

Twenty expressed sequence tags (ESTs) derived from cDNA libraries of different developmental stages of embryos were mapped using a whole genome bovine hamster radiation hybrid panel. These include 14 markers representing genes, most of which have not so far been mapped in cattle, with another three being novel in both cattle and human. The markers were placed on specific chromosomes with high LOD scores and except two all localizations fit the current human and cattle comparative map. The assignment of these genes further enriches the cattle genome map and also contributes to the international effort of generating comparative maps.
Effects of UVB‐irradiated algae on life history traits of Daphnia pulex
Authors: DE LANGE HENDRIKA , VAN DONK ELLEN
Published: November 2003

1. The impact of ultraviolet‐B (UVB)‐irradiated phytoplankton on the life history parameters of Daphnia was studied. Three species of Chlorophyceae (Chlamydomonas reinhardtii, Scenedesmus acutus and S. subspicatus) and one species of Cryptophyceae (Cryptomonas pyrenoidifera) were cultured with and without exposure to UVB radiation (biologically effective dose of 3 kJ m–2 day–1 DNA weighted) and fed to Daphnia pulex.

2. The daphnids feeding on UVB‐irradiated algae tended to be smaller in size, to produce less and smaller offspring, and to have a reduced population growth rate.

3. Fatty acid (FA) analysis of the phytoplankton showed a reduced total FA content under UVB radiation for all four species. Changes in the content of unsaturated fatty acids (UFA) were species specific.

4. The results indicate that the energy transfer between the primary producers’ and grazers’ level might be negatively influenced by UVB radiation. Direct effects on food quality are modest but detectable, changes in quantity or species composition may be more important in the energy transfer.
Effects of algal food quality on fecundity and population growth rates of Daphnia
Authors: KILHAM SUSAN , KREEGER DANIEL , GOULDEN CLYDE , LYNN SCOTT
Published: November 2003

1. Food quality was at least as important as food quantity for both fecundity and population growth responses of the cladoceran Daphnia pulicaria fed the green alga Ankistrodesmus falcatus grown under N limitation, P limitation, or non‐limited condition.

2. The fecundity of D. pulicaria was reduced under conditions of low food quality (low N or low P) compared with that for animals fed control non‐limited algae regardless of ration size. The reduced fecundity of D. pulicaria fed P‐limited food could be partially alleviated by increasing the ration (hence, compensation), but such was not the case for animals fed N‐limited food.

3. Population growth rates of D. pulicaria (rmax) were significantly reduced under conditions of low‐quality food for both N‐limited and P‐limited algae. Population growth rates were unaffected by ration size, indicating no compensation.
Effects of food type on the life history of Daphnia clones from lakes differing in trophic state. I. Daphnia galeata feeding on Scenedesmus and Oscillatoria*
Authors: REPKA SARI
Published: November 2003

1. Effects of the green alga Scenedesmus obliquus and the filamentous cyanobacterium Oscillatoria limnetica as food sources on the growth and reproduction of nine Daphnia galeata clones were studied. A high concentration of these two algae species was fed to Daphnia clones collected from four lakes with varying trophic conditions in a life history experiment.

2. Although Oscillatoria is regarded as a low quality food for Daphnia because of its filamentous morphology and suspected nutritional inadequacy, the cyanobacterium sustained both body growth and reproduction in D. galeata. Growth and reproduction were, however, lower and the onset of reproduction was delayed on a diet of Oscillatoria compared to a diet of Scenedesmus. The intrinsic growth rate, r, for each D. galeata clone was positive on a diet of Oscillatoria, ranging from 0.06 to 0.16 mm day–1, whereas the intrinsic growth rates on a diet of Scenedesmus ranged from 0.27 to 0.34 mm day–1.

3. The degree of growth depression on the Oscillatoria diet varied among clones. Clonal responses were correlated with body size; clones with larger body size decreased their growth and reproduction more on a diet of Oscillatoria. The responses of different clones from the same lake to the two diets resembled each other, irrespective of geography or trophic state of the lake.
Manipulation of dietary lipids, fatty acids and vitamins in zooplankton cultures
Authors: COUTTEAU PETER , SORGELOOS PATRICK
Published: November 2003

1. A wide range of species that are cultivated in commercial mariculture are planktonic during at least part of their life cycle; for example, the larval stages of shellfish (shrimp and molluscs) and the live feeds (rotifers, brine shrimp, copepods) used in the larviculture of marine fish and shellfish. Over the last decades various techniques have been developed to deliver nutrients to these zooplanktonic organisms either through artificial diets or by manipulating the composition of the live prey fed to the carnivorous stages. This paper reviews the methodology that has allowed aquaculturists to gain knowledge of nutritional requirements and may offer interesting opportunities for ecologists to verify the importance of key nutrients in the natural food chain of marine as well as freshwater ecosystems.

2. Live micro‐algae can be replaced partially or completely in the diet of filter‐feeders such as rotifers, Artemia, shrimp larvae and bivalves, by various types of preserved algae, micro‐encapsulated diets and yeast‐based diets, whereas lipid emulsions and liposomes may be utilized to supplement specific lipid‐and water‐soluble nutrients, respectively. Microbound and micro‐encapsulated diets have been designed to supplement live feed in the culture of micro‐predators such as fish and shrimp larvae.

3. Live prey organisms, in particular rotifers and Artemia, can be ‘bio‐encapsulated’ with a variety of enrichment diets to manipulate their content in certain nutrients, including ω3 highly unsaturated fatty acids (FA) and the vitamins C, A and E. Nevertheless, the enrichment techniques are not applicable for all nutrients and prey organisms. Phospholipid composition is difficult to manipulate through the diet of live feed and the enrichment of the essential FA docosahexaenoic acid (DHA) is hampered in most Artemia species due to the catabolism of this FA following enrichment.
Effect of physico‐chemical and molecular mobility parameters on Staphylococcus aureus growth
Authors: Vittadini Elena , Chinachoti Pavinee
Published: November 2003

The role played by water activity, ‘mobility’ and physico‐chemical properties of the media in modulating microbial response has been the object of large debate in the scientific community. In this study, Staphylococcus aureus growth parameters (lag phase and cell density at 24 h) in brain heart infusion (BHI) and BHI:NaCl (1:1) were analysed in their correlation with physico‐chemical/mobility parameters descriptive of the media [solid content, aw, kinematic viscosity, 17O NMR (R2, Pbw and )]. In these high moisture content, liquid and ‘homogeneous’ media S.aureus growth related to all the physico‐chemical and molecular mobility parameters analysed in a similar manner and it was found to be influenced more significantly by added NaCl than by the physico‐chemical and molecular mobility of the media. Staphylococcus aureus growth parameters correlated better with aw (relatively independent of NaCl concentration) than with any other parameter considered in this study.
Ssh10b, a conserved thermophilic archaeal protein, binds RNA in vivo
Authors: Guo Rong , Xue Hong , Huang Li
Published: November 2003

Proteins of the Sac10b family, which is highly conserved among hyperthermophilic archaea, have been regarded as DNA‐binding proteins. Based on their in vitro DNA‐binding properties, these proteins are thought to be involved in chromosomal organization or DNA repair/recombination. We show that Ssh10b, a member of the Sac10b family from Sulfolobus shibatae, bound with similar affinities to double‐stranded DNA, single‐stranded DNA and RNA in vitro. However, the protein was exclusively bound to RNA in S. shibatae cells, as revealed by in vivo UV cross‐linking and co‐immunoprecipitation. Ribosomal RNAs were among the RNA species co‐immunoprecipitated with Ssh10b. Consistent with this observation, Ssh10b was co‐purified with ribosomes under low salt conditions. Furthermore, we demonstrate by UV‐cross‐linking hybridization that, when the cells were irradiated with UV, Ssh10b became cross‐linked to 16S, 23S rRNAs and mRNAs. Our data indicate that RNA is the physiological binding target of the Sac10b family.
Constitutive expression of PcsB suppresses the requirement for the essential VicR (YycF) response regulator in Streptococcus pneumoniae R6§
Authors: Ng Wai‐Leung , Robertson Gregory T. , Kazmierczak Krystyna M. , Zhao Jingyong , Gilmour Raymond , Winkler Malcolm E.
Published: November 2003

We report several new findings about the function of the essential VicRK two‐component regulatory system (TCS) in the human pathogen Streptococcus pneumoniae. The vicR‐encoded response regulator, vicK‐encoded histidine kinase and the protein encoded by the downstream vicX gene are the homologues of the YycF, YycG and YycJ proteins, respectively, studied previously in Bacillus subtilis and Staphylococcus aureus. Using a regulatable promoter, we demonstrated that the VicK histidine kinase is conditionally required for growth of S. pneumoniae. Likewise, we found that the VicX protein is also conditionally required for growth and probably plays a role in the essential signal transduction pathway mediated by VicR and VicK. Recovery of limited substitutions in the conserved aspartate 52 residue (D52) of VicR was consistent with a requirement for phosphorylation of VicR for growth under some conditions. We applied microarrays to characterize the changes in transcription patterns in bacteria depleted for vicRKX operon expression. Our results suggest that the pcsB gene is a target of the VicRK TCS. We present evidence that downregulation of pcsB could account for many of the defects in cell growth, shape, size and morphology observed in bacteria depleted for vicRKX expression. Furthermore, constitutive expression of pcsB+ suppressed the essential requirement for the VicRK TCS and allowed the isolation of vicR null mutants.
Deletion analysis identifies key functional domains of the cytadherence‐associated protein HMW2 of Mycoplasma pneumoniae
Authors: Balish Mitchell F. , Ross Stephanie M. , Fisseha Makda , Krause Duncan C.
Published: November 2003

Mycoplasma pneumoniae attachment to host cells requires biogenesis of a functional attachment organelle, including proper localization of the adhesion protein P1 to this structure. Mutations in the hmw2 gene result in the inability to cytadhere, failure to localize P1 to the attachment organelle, altered cell morphology and accelerated turnover of the cytadherence‐associated proteins HMW1, HMW3 and P65. The hmw2 gene encodes HMW2 (190 kDa) and P28 (28 kDa), the latter apparently the product of internal translation initiation near the 3′ end of the hmw2 coding region. Transformation of hmw2 mutant I‐2 with recombinant wild‐type hmw2 restores a wild‐type phenotype. In the current study, a severely truncated hmw2 gene with an in frame internal deletion of 80% of the HMW2 coding region that leaves the P28‐encoding region intact restored cytadherence to mutant I‐2. Transformants produced the expected 38 kDa HMW2 derivative (HMW2Δmid) at levels comparable to that of HMW2 in wild‐type cells; like HMW2, HMW2Δmid exhibited marked Triton X‐100 insolubility. HMW3, P65 and P28 were fully restored, but not HMW1. These transformants were morphologically similar to wild‐type M. pneumoniae but failed to localize P1 to the attachment organelle. Finally, a C‐terminally truncated HMW2 derivative was partly Triton X‐100 soluble and incapable of restoring HMW1, HMW3 and P65 to wild‐type levels. These data are consistent with a model in which the C‐terminal domain of HMW2 imparts normal localization to the protein, and this localization itself is required for productive interactions with downstream cytadherence‐associated proteins. Furthermore, these results emphasize the association of HMW1 with P1 clustering.
Distribution of split DnaE inteins in cyanobacteria
Authors: Caspi Jonathan , Amitai Gil , Belenkiy Olga , Pietrokovski Shmuel
Published: November 2003

Inteins are genetic elements found inside the coding regions of different host proteins and are translated in frame with them. The intein‐encoded protein region is removed by an autocatalytic protein‐splicing reaction that ligates the host protein flanks with a peptide bond. This reaction can also occur in trans with the intein and host protein split in two. After translation of the two genes, the two intein parts ligate their flanking protein parts to each other, producing the mature protein. Naturally split inteins are only known in the DNA polymerase III alpha subunit (polC or dnaE gene) of a few cyanobacteria. Analysing the phylogenetic distribution and probable genetic propagation mode of these split inteins, we conclude that they are genetically fixed in several large cyanobacterial lineages. To test our hypothesis, we sequenced parts of the dnaE genes from five diverse cyanobacteria and found all species to have the same type of split intein. Our results suggest the occurrence of a genetic rearrangement in the ancestor of a large division of cyanobacteria. This event fixed the dnaE gene in a unique two‐genes one‐protein configuration in the progenitor of many cyanobacteria. Our hypothesis, findings and the cloning procedure that we established allow the identification and acquisition of many naturally split inteins. Having a large and diverse repertoire of these unique inteins will enable studies of their distinct activity and enhance their use in biotechnology.
Glutathione peroxidase genes in Arabidopsis are ubiquitous and regulated by abiotic stresses through diverse signaling pathways
Authors: Milla Miguel A. Rodriguez , Maurer Alberto , Huete Alicia Rodriguez , Gustafson J. Perry
Published: November 2003

Glutathione peroxidases (GPXs) are a group of enzymes that protect cells against oxidative damage generated by reactive oxygen species (ROS). The presence of GPXs in plants has been reported by several groups, but the roles of individual members of this family in a single plant species have not been studied. A family of seven related proteins named AtGPX1– AtGPX7 in Arabidopsis was identified, and the genomic organization of this family was reported. The putative subcellular localizations of the encoded proteins are the cytosol, chloroplast, mitochondria, and endoplasmic reticulum. Expressed sequence tags (ESTs) for all the genes except AtGPX7 were identified. Expression analysis of AtGPX genes in Arabidopsis tissues was performed, and different patterns were detected. Interestingly, several genes were up‐regulated coordinately in response to abiotic stresses. AtGPX6, like human phospholipid hydroperoxide GPX (PHGPX), possibly encodes mitochondrial and cytosolic isoforms by alternative initiation. In addition, this gene showed the strongest responses under most abiotic stresses tested. AtGPX6::GUS analysis in transgenic Arabidopsis showed that AtGPX6 is highly expressed throughout development in most tissues, thus supporting an important role for this gene in protection against oxidative damage. The different effects of salicylic acid (SA), jasmonic acid (JA), abscisic acid (ABA), and auxin on the expression of the genes indicate that the AtGPX family is regulated by multiple signaling pathways. Analysis of the upstream region of the AtGPX genes revealed the presence of multiple conserved motifs, and some of them resembled antioxidant‐responsive elements found in plant and human promoters. The potential regulatory role of specific sequences is discussed.
Temporal and spatial control of gene silencing in transgenic plants by inducible expression of double‐stranded RNA
Authors: Chen Shuai , Hofius Daniel , Sonnewald Uwe , Börnke Frederik
Published: November 2003

Downregulation of endogenous genes via post‐transcriptional gene silencing (PTGS) is a key to the characterization of gene function in plants. The recent discovery that double‐stranded RNA (dsRNA) is an extremely effective trigger of gene silencing greatly enhanced the predictability of this approach. However, strong constitutive silencing often leads to pleiotropic effects, which make it difficult to directly relate phenotype to gene function, or even interferes with the recovery of viable transgenic plants. Here, we show that strong genetic interference can be achieved in a chemically inducible fashion, allowing for temporal and spatial control of gene silencing in transgenic plants. To this end, transgenic tobacco plants were established expressing dsRNA in the form of intron‐spliced hairpin structures under the control of the ethanol‐inducible alc gene expression system. Targeting magnesium (Mg)‐chelatase subunit I (Chl I) and glutamate 1‐semialdehyde aminotransferase (GSA), both involved in chlorophyll (chl) biosynthesis, resulted in rapid and specific mRNA degradation upon induction with ethanol. Ethanol‐inducible silencing of the target genes caused strong but transient phenotypical alterations featured by a progressive loss of chl in young leaves, which persisted for about 7–9 days before newly growing leaves completely recovered. About 10–30% of the primary transformants showed phenotype development upon induction. Local silencing of Chl I could be achieved by confined ethanol treatment of a single leaf without affecting any other part of the plant. Inducible gene silencing using the alc system promises to obviate the problems associated with constitutive RNA silencing and enables to dissect primary and secondary effects of PTGS at temporal and spatial resolution.
A chloroplast‐localized PPR protein required for plastid ribosome accumulation
Authors: Williams Pascale M. , Barkan Alice
Published: November 2003

The pentatricopeptide repeat (PPR) is a degenerate 35‐amino acid repeating motif that is found in animal, fungal, and plant proteins. The PPR protein family is particularly large in plants, where the majority of family members are predicted to be targeted to mitochondria or chloroplasts. PPR proteins are believed to fall into the larger family of helical repeat proteins, which typically bind macromolecules through a surface formed by the stacking of consecutive helical repeating units. Prior findings implicate several PPR proteins in organellar RNA metabolism, but the biological functions of few PPR proteins have been explored and in no case has a direct substrate been definitively identified. We present a characterization of the maize nuclear gene ppr2, which encodes a chloroplast PPR protein. PPR2 is found in large, heterogeneous protein complexes in the chloroplast stroma, some of which may be associated with RNA. Null ppr2 mutants have albino leaves and lack plastid rRNA and translation products. Plastid rRNAs are absent in both dark‐ and light‐grown leaf tissues, indicating that their absence does not result from photo‐oxidative damage. The population of plastid transcripts in ppr2 mutants is similar to that in other maize mutants lacking plastid ribosomes, and no ppr2‐specific defects in plastid RNA metabolism have been detected. Taken together, the results suggest that ppr2 functions in the synthesis or assembly of one or more component of the plastid translation machinery.
Frequency of three BRCA1 gene founder mutations in breast/ovarian cancer families from the Pomerania–Kujawy region of Poland
Authors: Janiszewska H , Haus O , Lauda‐Świeciak A , Pasińska M , Laskowski R , Szymański W , Górski B , Lubiński J
Published: November 2003

A group of 63 families from the Pomerania–Kujawy region were analyzed for three BRCA1 gene Polish founder mutations, 5382insC, 300T>G, and 4153delA, because of breast (BrCa) and/or ovarian cancer (OvCa) history. The analysis was carried out by multiplex polymerase chain reaction method. BRCA1 mutation was revealed in nine (14%) families: in three (33%) of hereditary BrCa and OvCa families, in three (8%) of hereditary BrCa families, and in three (21%) of hereditary OvCa families. According to risk criteria, it was revealed in 45% of high‐risk families with more than three cancers, 13% of moderate‐risk families with two cancers, and 8% of families with sporadic OvCa. In six families, the mutation was found in a proband with BrCa or OvCa and in three families, the mutation was found in a healthy proband, first‐degree relative of a patient deceased of BrCa or OvCa. 5382insC frameshift mutation accounted for 67% and 300T>G missense mutation for 33% of all identified familial mutations. 4153delA frameshift mutation was not found in analyzed sample of families. 5382insC mutation was found in 9% and 300T>G in 5% of all investigated families, and in 27 and 18%, respectively, of high‐risk families. This underlines the importance of applying strict inclusion criteria to analyze mutation frequency in hereditary BrCa/OvCa families.
Novel isoforms of NADPH oxidase in vascular physiology and pathophysiology
Authors: Bengtsson Sara HM , Gulluyan Lerna M , Dusting Gregory J , Drummond Grant R
Published: November 2003

1. Vascular cells have evolved to use reactive oxygen species (ROS), such as superoxide and hydrogen peroxide, as signalling molecules. Under physiological conditions, ROS are important regulators of cell cycle, protein kinase activity and gene expression. However, in vascular disease states, such as hypertension and hypercholesterolaemia, excessive production of ROS may overwhelm the anti‐oxidant defence mechanisms of cells, resulting in ‘oxidative stress’, damage to the artery wall and, ultimately, development of atherosclerotic plaques.

2. The primary source of ROS in the vasculature is NADPH oxidase. There appear to be at least three isoforms of NADPH oxidase expressed in the vascular wall, each differing with respect to the flavin‐containing catalytic subunit it uses to transfer electrons from NADPH to molecular oxygen. Thus, although endothelial cells and adventitial fibroblasts express a gp91phox‐containing NADPH oxidase similar to that originally identified in phagocytes, vascular smooth muscle cells may rely on novel homologues of gp91phox, namely Nox1 and Nox4, to produce superoxide.

3. Controversy remains over which isoform(s) of NADPH oxidase is responsible for the oxidative stress associated with vascular diseases. We and others have shown that although gp91phox mRNA expression is upregulated during atherogenesis in human and animal models, expression of the Nox4 subunit remains unchanged. Nox1 expression is also likely to be increased in diseased arteries; however, its relative level of expression, at least at the mRNA level, appears to be markedly lower than that of the other gp91phox homologues, even after upregulation.

4. Whether these findings suggest that a gp91phox‐containing NADPH oxidase is more important than either Nox4 or Nox1 in vascular disease awaits studies examining relative protein expression and enzyme kinetics of each subunit, as well as the effects of targeted gene deletion of each of these gp91phox homologues on atherogenesis.