bluuebunny/crossref_metadata_2025 huggingface dataset:
Phenotypic plasticity of an invasive acacia versus two native Mediterranean species
Authors: Peperkorn Ralf , Werner Christiane , Beyschlag Wolfram
Published: October 2005

The phenotypic plasticity and the competitive ability of the invasive Acacia longifolia v. the indigenous Mediterranean dune species Halimium halimifolium and Pinus pinea were evaluated. In particular, we explored the hypothesis that phenotypic plasticity in response to biotic and abiotic factors explains the observed differences in competitiveness between invasive and native species. The seedlings’ ability to exploit different resource availabilities was examined in a two factorial experimental design of light and nutrient treatments by analysing 20 physiological and morphological traits. Competitiveness was tested using an additive experimental design in combination with 15N-labelling experiments. Light and nutrient availability had only minor effects on most physiological traits and differences between species were not significant. Plasticity in response to changes in resource availability occurred in morphological and allocation traits, revealing A. longifolia to be a species of intermediate responsiveness. The major competitive advantage of A. longifolia was its constitutively high shoot elongation rate at most resource treatments and its effective nutrient acquisition. Further, A. longifolia was found to be highly tolerant against competition from native species. In contrast to common expectations, the competition experiment indicated that A. longifolia expressed a constant allocation pattern and a phenotypic plasticity similar to that of the native species.
Galactosidases in opening, senescing and water-stressed Sandersonia aurantiaca flowers
Authors: O'Donoghue Erin M. , Eason Jocelyn R. , Somerfield Sheryl D. , Ryan Dacey A.
Published: October 2005

Three glycosyl hydrolase family 35 β-galactosidase-encoding cDNAs, SaGAL1 (full-length), SaGAL2 and SaGA3L (both partial), have been isolated from Sandersonia aurantiaca (Hook.) SaGAL1 protein was functionally expressed in E. coli and β-galactosidase identity confirmed by activity assay. All three clones are primarily expressed in tepal tissues of senescing sandersonia flowers. In order to identify relationships between tepal texture and galactose metabolism, cut sandersonia flowers were treated with sucrose, periods of dryness or PEG and parameters associated with galactose metabolism and firmness were monitored. Sucrose supplementation, known to increase tepal firmness, delayed expression of SaGAL1 and SaGAL3 in opening (stage 5) flowers, whereas the response to periods of dryness followed by rehydration depended on the maturity of the flower. These treatments also tended to hasten the onset of processes associated with programmed cell death, monitored by PRT5 (a senescence-associated protease) expression. Galactosidase activity and cell wall galactose content were also affected but in an inconsistent manner. PEG supplied to opening flowers for 1 d followed by water, induced a long period of wilt, and intensive PRT5 expression. However, β-galactosidase gene expression and activity was delayed in these flowers, and cell-wall galactose content changed apparently independently of galactosidase activity. We have not been able to demonstrate a causal connection between the change in petal texture and concurrent induction of galactose mobilisation in sandersonia during normal development and senescence.
The nucleotide sequence data reported appears in the EMBL, GenBank and DDBJ Nucleotide Sequence Databases under accession number AY280498 (SaGAL1), AY280499 (SaGAL2), AY280500 (SaGAL3).
Identification of microRNAs from rice
Authors: Lu Yu-Zhu , Yan Da-Wei , Lu Ying-Tang
Published: October 2005

MicroRNAs (miRNAs) are important regulators in the development of plants and animals. Several hundred have been identified from animals, and about a dozen have been cloned from plants, mainly Arabidopsis thaliana (L.) Heynh. We have identified nine miRNAs in Oryza sativa L., an important food crop that has been sequenced in recent years. The nine miRNAs include miRNA171 and miRNA167, which were also identified in Arabidopsis. These had the typical properties of miRNAs, including short length, an ability to form a stem–loop structure with a flanking genomic sequence and they could be identified by northern blot analyses. In addition, m-fold program and computational analyses indicted that the potential targets of six of the nine miRNAs are four known gene families and two unknown protein families, which comprise 16 unique genes.
Characterisation of a T-DNA-tagged gene of Arabidopsis thaliana that regulates gibberellin metabolism and flowering time
Authors: Svensson Maria , Lundh Dan , Bergman Per , Mandal Abul
Published: October 2005

A gene (At4g20010) involved in regulating flowering time in Arabidopsis thaliana (L.) Heynh. was identified by promoter trap T-DNA tagging. Plants containing a T-DNA insert in the 3′-UTR of At4g20010 flowered later under both long- and short-day conditions compared with control plants. Histochemical assays of the mutant plants showed that the promoterless gus gene is expressed predominantly in the shoot apex, but it is also expressed in root tips, stem nodes and in the abscission zone of developing siliques. Measurement of endogenous gibberellin (GA) showed that bioactive GA4 levels in mutant plants were reduced compared with wild type (WT) plants. Like other known mutants defective in GA biosynthesis, the late-flowering phenotype observed in our T-DNA-tagged line could be largely repressed by application of exogenous GA3. The T-DNA-tagged gene At4g20010 encodes a previously uncharacterised protein belonging to the DUF731 family. Sequence analysis showed similarity to a single-stranded binding domain and to an RNA-binding protein of Chlamydomonas reinhardtii. Considering the above results (sequence similarity, mutant phenotype and level of endogenous GA), we propose that At4g20010 is an RNA-binding protein involved in regulating GA biosynthesis, possibly at the post-transcriptional level.
Ca2+ pulsation in BY-2 cells and evidence for control of mechanosensory Ca2+-selective channels by the plasmalemmal reticulum
Authors: Pickard Barbara G. , Fujiki Masaaki
Published: October 2005

A previously unknown cytoskeletal structure, now named the plasmalemmal reticulum (Gens et al. 2000, Protoplasma 212, 115–134), was found in cultured BY-2 tobacco cells during a search for a force-focusing mechanism that might enhance signal transduction by the cells’ mechanosensory Ca2+-selective cation channels (MCaCs). This polyhedral structure, which links cell wall, plasma membrane, and internal cytoplasm, prominently contains arabinogalactan protein (AGP). To check for reticulum-promoted Ca2+ elevation, the AGP-binding reagent (β-d-glucosyl)3 Yariv phenylglycoside has been applied to BY-2 cells expressing a free cameleon Ca2+ reporter. Ca2+ elevation was substantial and prolonged. Moreover it occurred in the nucleus as well as the cytoplasm. Cells treated with non-binding mannosyl Yariv reagent could not be discriminated from untreated controls or those treated with carrier solution alone. Supply of the MCaC inhibiter Gd3+ just before treatment with Yariv reagent prevented Ca2+ rise. These data strongly support the hypothesis that the plasmalemmal reticulum controls MCaC activity. The massive inward spread of Ca2+ suggested that entry of the ion through the channels initiated a wave of release from the ER, and YCX in the ER showed Ca2+ levels consistent with this premise. Cytosolic and nuclear Ca2+ often pulsed in control cells in near synchrony and at rates ranging from zero to five cycles per ∼20-min recording. (Pulsation was over-ridden by the applied amounts of glucosyl Yariv compound.) Suggestively but very crudely, oscillation rate was assessed as possibly correlating with stage of cell cycle. Because cell Ca2+ was lowered and pulsation was eliminated by Gd3+, MCaCs appear to participate in these endogenous fluctuations. The extent to which pulsing plays regulatory roles in relatively undifferentiated types of cells should be evaluated.
Potential yield and water-use efficiency benefits in sorghum from limited maximum transpiration rate
Authors: Sinclair Thomas R. , Hammer Graeme L. , van Oosterom Erik J.
Published: October 2005

Limitations on maximum transpiration rates, which are commonly observed as midday stomatal closure, have been observed even under well-watered conditions. Such limitations may be caused by restricted hydraulic conductance in the plant or by limited supply of water to the plant from uptake by the roots. This behaviour would have the consequences of limiting photosynthetic rate, increasing transpiration efficiency, and conserving soil water. A key question is whether the conservation of water will be rewarded by sustained growth during seed fill and increased grain yield. This simulation analysis was undertaken to examine consequences on sorghum yield over several years when maximum transpiration rate was imposed in a model. Yields were simulated at four locations in the sorghum-growing area of Australia for 115 seasons at each location. Mean yield was increased slightly (5–7%) by setting maximum transpiration rate at 0.4 mm h–1. However, the yield increase was mainly in the dry, low-yielding years in which growers may be more economically vulnerable. In years with yield less than ∼450 g m–2, the maximum transpiration rate trait resulted in yield increases of 9–13%. At higher yield levels, decreased yields were simulated. The yield responses to restricted maximum transpiration rate were associated with an increase in efficiency of water use. This arose because transpiration was reduced at times of the day when atmospheric demand was greatest. Depending on the risk attitude of growers, incorporation of a maximum transpiration rate trait in sorghum cultivars could be desirable to increase yields in dry years and improve water use efficiency and crop yield stability.
Measuring the transmission dynamics of a sexually transmitted disease
Authors: Ryder Jonathan J. , Webberley K. Mary , Boots Michael , Knell Robert J.
Published: October 2005

Sexually transmitted diseases (STDs) occur throughout the animal kingdom and are generally thought to affect host population dynamics and evolution very differently from other directly transmitted infectious diseases. In particular, STDs are not thought to have threshold densities for persistence or to be able to regulate host population density independently; they may also have the potential to cause host extinction. However, these expectations follow from a theory that assumes that the rate of STD spread depends on the proportion (rather than the density) of individuals infected in a population. We show here that this key assumption (“frequency dependence”), which has not previously been tested in an animal STD system, is invalid in a simple and general experimental model. Transmission of an STD in the two-spot ladybird depended more on the density of infected individuals in the study population than on their frequency. We argue that, in this system, and in many other animal STDs in which population density affects sexual contact rate, population dynamics may exhibit some characteristics that are normally reserved for diseases with density-dependent transmission.
Heat-shock protein 60 is required for blastema formation and maintenance during regeneration
Authors: Makino Shinji , Whitehead Geoffrey G. , Lien Ching-Ling , Kim Soo , Jhawar Payal , Kono Akane , Kawata Yasushi , Keating Mark T.
Published: October 2005

Zebrafish fin regeneration requires the formation and maintenance of blastema cells. Blastema cells are not derived from stem cells but behave as such, because they are slow-cycling and are thought to provide rapidly proliferating daughter cells that drive regenerative outgrowth. The molecular basis of blastema formation is not understood. Here, we show that heat-shock protein 60 (
hsp60
) is required for blastema formation and maintenance. We used a chemical mutagenesis screen to identify no blastema (
nbl
), a zebrafish mutant with an early fin regeneration defect. Fin regeneration failed in
nbl
due to defective blastema formation.
nbl
also failed to regenerate hearts. Positional cloning and mutational analyses revealed that
nbl
results from a V324E missense mutation in
hsp60
. This mutation reduced
hsp60
function in binding and refolding denatured proteins.
hsp60
expression is increased during formation of blastema cells, and dysfunction leads to mitochondrial defects and apoptosis in these cells. These data indicate that
hsp60
is required for the formation and maintenance of regenerating tissue.
Sex differences in estrogenic regulation of neuronal activity in neonatal cultures of ventromedial nucleus of the hypothalamus
Authors: Zhou Jin , Pfaff Donald W. , Chen Gong
Published: October 2005

Estrogenic effects have been implicated in sexual differentiation of brain and behavior, in part by affecting neuronal activity in the ventromedial nucleus of the hypothalamus (VMN). We report here a remarkable sex difference in estrogenic regulation of neuronal activity in male vs. female neural networks. Spontaneous synaptic currents originating from a population of neurons were recorded in primary VMN cultures using the whole-cell patch-clamp technique. Treatment with 17β-estradiol (E2, 10 nM) for 24 h induced opposite effects in the two sexes: the frequency of spontaneous synaptic events decreased significantly in neurons derived from males but increased in those from females. Interestingly, the 24-hour E2 effect was partially reversed by an acute application (5 min) of a second dose of E2 (10 nM), suggesting an interaction between extended (24-hr) and acute (5-min) effects of E2 in VMN neurons. To understand the underlying mechanism of this sexually dimorphic action of E2, we analyzed the E2 effect on GABAergic neurotransmission by recording miniature inhibitory postsynaptic currents. After 24-hour E2 treatment, both the amplitude and frequency of miniature inhibitory postsynaptic currents increased in neurons derived from males but decreased in those from females. These results suggest that E2-induced changes in GABAergic inhibition could at least partially explain E2 effects on neuronal activity. We conclude that E2 may have sexually dimorphic effects on the synaptic output of VMN neurons by modulating GABAergic neurotransmission.
CREDO: a web-based tool for computational detection of conserved sequence motifs in noncoding sequences
Authors: Hindemitt Tobias , Mayer Klaus F. X.
Published: October 2005

Summary: CREDO is a user-friendly, web-based tool that integrates the analysis and results of different algorithms widely used for the computational detection of conserved sequence motifs in noncoding sequences. It enables easy comparison of the individual results. CREDO offers intuitive interfaces for easy and rapid configuration of the applied algorithms and convenient views on the results in graphical and tabular formats.

Availability:  

Contact:  kmayer@gsf.de

Supplementary information: A detailed help file for CREDO is available on . Further supplementary material is available on Bioinformatics online.
Discrimination of outer membrane proteins using support vector machines
Authors: Park Keun-Joon , Gromiha M. Michael , Horton Paul , Suwa Makiko
Published: October 2005

Motivation: Discriminating outer membrane proteins from other folding types of globular and membrane proteins is an important task both for dissecting outer membrane proteins (OMPs) from genomic sequences and for the successful prediction of their secondary and tertiary structures.

Results: We have developed a method based on support vector machines using amino acid composition and residue pair information. Our approach with amino acid composition has correctly predicted the OMPs with a cross-validated accuracy of 94% in a set of 208 proteins. Further, this method has successfully excluded 633 of 673 globular proteins and 191 of 206 α-helical membrane proteins. We obtained an overall accuracy of 92% for correctly picking up the OMPs from a dataset of 1087 proteins belonging to all different types of globular and membrane proteins. Furthermore, residue pair information improved the accuracy from 92 to 94%. This accuracy of discriminating OMPs is higher than that of other methods in the literature, which could be used for dissecting OMPs from genomic sequences.

Availability: Discrimination results are available at

Contact:  michael-gromiha@aist.go.jp
Persister cells mediate tolerance to metal oxyanions in Escherichia coli
Authors: Harrison Joe J. , Ceri Howard , Roper Nicole J. , Badry Erin A. , Sproule Kimberley M. , Turner Raymond J.
Published: October 2005

Bacterial cultures produce subpopulations of cells termed ‘persisters’, reputedly known for high tolerance to killing by antibiotics. Ecologically, antibiotics produced by competing microflora are only one potential stress encountered by bacteria. Another pressure in the environment is toxic metals that are distributed ubiquitously by human pollution, volcanic activity and the weathering of minerals. This study evaluated the time- and concentration-dependent killing of Escherichia coli planktonic and biofilm cultures by the water-soluble metal(loid) oxyanions chromate (), arsenate (), arsenite (), selenite (), tellurate () and tellurite (). Correlative to previous reports in the literature, control antibiotic assays indicated that a small proportion of E. coli biofilm populations remained recalcitrant to killing by antibiotics (even with 24 h exposure). In contrast, metal oxyanions presented a slow, bactericidal action that eradicated biofilms. When exposed for 2 h, biofilms were up to 310 times more tolerant to killing by metal oxyanions than corresponding planktonic cultures. However, by 24 h, planktonic cells and biofilms were eradicated at approximately the same concentration in all instances. Coloured complexes of metals and chelators could not be generated in biofilms exposed to or , suggesting that the extracellular polymeric matrix of E. coli may have a low binding affinity for metal oxyanions. Viable cell counts at 2 and 24 h exposure revealed that, at high concentrations, all of the metal oxyanions had killed 99 % (or a greater proportion) of the bacterial cells in biofilm populations. It is suggested here that the short-term survival of <1 % of the bacterial population corresponds well with the hypothesis that a small population of persister cells may be responsible for the time-dependent tolerance of E. coli biofilms to high concentrations of metal oxyanions.
σ B contributes to Listeria monocytogenes invasion by controlling expression of inlA and inlB
Authors: Kim Heesun , Marquis Hélène , Boor Kathryn J.
Published: October 2005

The ability ofListeria monocytogenesto invade non-phagocytic cells is important for development of a systemic listeriosis infection. The authors previously reported that aL. monocytogenesΔsigBstrain is defective in invasion into human intestinal epithelial cells, in part, due to decreased expression of a major invasion gene,inlA. To characterize additional invasion mechanisms under the control ofσB, mutants were generated carrying combinations of in-frame deletions ininlA,inlBandsigB. Quantitative assessment of bacterial invasion into the human enterocyte Caco-2 and hepatocyte HepG-2 cell lines demonstrated thatσBcontributes to both InlA and InlB-mediated invasion ofL. monocytogenes. Previous identification of theσB-dependent P2prfApromoter upstream of the major virulence gene regulator, positive regulatory factor A (PrfA), suggested that the contributions ofσBto expression of various virulence genes, includinginlA, could be at least partially mediated through PrfA. To test this hypothesis, relative invasion capabilities of ΔsigBand ΔprfAstrains were compared. Exponential-phase cells of the ΔsigBand ΔprfAstrains were similarly defective at invasion; however, stationary-phase ΔsigBcells were significantly less invasive than stationary-phase ΔprfAcells, suggesting that the contributions ofσBto invasion extend beyond those mediated through PrfA in stationary-phaseL. monocytogenes. TaqMan quantitative reverse-transcriptase PCRs further demonstrated that expression ofinlAandinlBwas greatly increased in aσB-dependent manner in stationary-phaseL. monocytogenes. Together, results from this study provide strong biological evidence of a critical role forσBinL. monocytogenesinvasion into non-phagocytic cells, primarily mediated through control ofinlAandinlBexpression.
Functional analysis of EspB from enterohaemorrhagic Escherichia coli
Authors: Chiu Hao-Jie , Syu Wan-Jr
Published: October 2005

In enterohaemorrhagic Escherichia coli (EHEC), the type III secretion protein EspB is translocated into the host cells and plays an important role in adherence, pore formation and effector translocation during infection. The secretion domain of EspB has been mapped previously. To define the other functional determinants of EspB, several plasmids encoding different fragments of EspB were created and analysed to see which of them lost the functions of the full-length molecule. One finding was that residues 118–190 of EspB were required for both efficient translocation of EspB and interaction of EspB with EspA. Additionally, the segment consisting of residues 217–312 was necessary for bacterial adherence. Furthermore, a predicted transmembrane domain (residues 99–118) was found to be critical for EHEC to cause red blood cell haemolysis, presumably by forming pores in the cell membrane. The same segment was also important for actin accumulation induced beneath the bacterial-attachment site. Taken together, these data indicate that the EspB protein (312 residues in total) has functions associated with its different regions. These regions may interact with each other or with other components of the type III system to orchestrate the intricate actions of EHEC during infection.
Colony sectorization of Metarhizium anisopliae is a sign of ageing
Authors: Wang Chengshu , Butt Tariq M. , Leger Raymond J. St
Published: October 2005

Spontaneous phenotypic degeneration resulting in sterile sectors is frequently observed when culturing filamentous fungi on artificial medium. Sterile sectors from two different strains of the insect pathogenic fungusMetarhizium anisopliaewere investigated and found to contain reduced levels of cAMP and destruxins (insecticidal peptides). Microarray analysis using slides printed with 1730 clones showed that compared to wild-type, sterile sectors down-regulated 759 genes and upregulated 27 genes during growth in Sabouraud glucose broth or on insect cuticle. The differentially expressed genes are largely involved in cell metabolism (18·8 %), cell structure and function (13·6 %) and protein metabolism (8·8 %). Strong oxidative stress was demonstrated in sectorial cultures using the nitro blue tetrazolium assay and these cultures show other syndromes associated with ageing, including mitochondrial DNA alterations. However, genes involved in deoxidation and self-protection (e.g. heat-shock proteins, HSPs) were also upregulated. Further evidence of physiological adaptation by the degenerative sectorial cultures included cell-structure reorganization and the employment of additional signalling pathways. In spite of their very similar appearance, microarray analysis identified 181 genes differentially expressed between the two sectors, and the addition of exogenous cAMP only restored conidiation in one of them. Most of the differentially expressed genes were involved in catabolic or anabolic pathways, but the latter included genes for sporulation. Compared to the mammalian ageing process, sectorization inM. anisopliaeshowed many similarities, including similar patterns of cAMP production, oxidative stress responses and the involvement of HSPs. Thus, a common molecular machinery for ageing may exist throughout the eukaryotes.
Multilocus sequence analysis and comparative evolution of virulence-associated genes and housekeeping genes of Clostridium difficile
Authors: Lemée Ludovic , Bourgeois Ingrid , Ruffin Elodie , Collignon Anne , Lemeland Jean-François , Pons Jean-Louis
Published: October 2005

A multilocus sequence analysis of ten virulence-associated genes was performed to study the genetic relationships between 29 Clostridium difficile isolates of various origins, hosts and clinical presentations, and selected from the main lineages previously defined by multilocus sequence typing (MLST) of housekeeping genes. Colonization-factor-encoding genes (cwp66, cwp84, fbp68, fliC, fliD, groEL and slpA), toxin A and B genes (tcdA and tcdB), and the toxin A and B positive regulator gene (tcdD) were investigated. Binary toxin genes (cdtA and cdtB) were also detected, and internal fragments were sequenced for positive isolates. Virulence-associated genes exhibited a moderate polymorphism, comparable to the polymorphism of housekeeping genes, whereas cwp66 and slpA genes appeared highly polymorphic. Isolates recovered from human pseudomembranous colitis cases did not define a specific lineage. The presence of binary toxin genes, detected in five of the 29 isolates (17 %), was also not linked to clinical presentation. Conversely, toxigenic A−B+ isolates defined a very homogeneous lineage, which is distantly related to other isolates. By clustering analysis, animal isolates were intermixed with human isolates. Multilocus sequence analysis of virulence-associated genes is consistent with a clonal population structure for C. difficile and with the lack of host specificity. The data suggest a co-evolution of several of the virulence-associated genes studied (including toxins A and B and the binary toxin genes) with housekeeping genes, reflecting the genetic background of C. difficile, whereas flagellin, cwp66 and slpA genes may undergo recombination events and/or environmental selective pressure.
Effect of methanobactin on the activity and electron paramagnetic resonance spectra of the membrane-associated methane monooxygenase in Methylococcus capsulatus Bath
Authors: Choi Dong W. , Antholine William E. , Do Young S. , Semrau Jeremy D. , Kisting Clint J. , Kunz Ryan C. , Campbell Damon , Rao Vinay , Hartsel Scott C. , DiSpirito Alan A.
Published: October 2005

Improvements in the purification of methanobactin (mb) from eitherMethylosinus trichosporiumOB3bTorMethylococcus capsulatusBath resulted in preparations that stimulated methane-oxidation activity in both whole-cell and cell-free fractions ofMethylococcus capsulatusBath expressing the membrane-associated methane monooxygenase (pMMO). By using washed membrane factions with pMMO activities in the 290 nmol propylene oxidized min−1(mg protein)−1range, activities approaching 400 nmol propylene oxidized min−1(mg protein)−1were commonly observed following addition of copper-containing mb (Cu–mb), which represented 50–75 % of the total whole-cell activity. The stimulation of methane-oxidation activity by Cu–mb was similar to or greater than that observed with equimolar concentrations of Cu(II), without the inhibitory effects observed with high copper concentrations. Stimulation of pMMO activity was not observed with copper-free mb, nor was it observed when the copper-to-mb ratio was <0·5 Cu atoms per mb. The electron paramagnetic resonance (EPR) spectra of mb differed depending on the copper-to-mb ratio. At copper-to-mb ratios of <0·4 Cu(II) per mb, Cu(II) addition to mb showed an initial coordination by both sulfur and nitrogen, followed by reduction to Cu(I) in <2 min. At Cu(II)-to-mb ratios between 0·4 and 0·9 Cu(II) per mb, the intensity of the Cu(II) signal in EPR spectra was more representative of the Cu(II) added and indicated more nitrogen coordination. The EPR spectral properties of mb and pMMO were also examined in the washed membrane fraction following the addition of Cu(II), mb and Cu–mb in the presence or absence of reductants (NADH or duroquinol) and substrates (CH4and/or O2). The results indicated that Cu–mb increased electron flow to the pMMO, increased the free radical formed following the addition of O2and decreased the residual free radical following the addition of O2plus CH4. The increase in pMMO activity and EPR spectral changes to the pMMO following Cu–mb addition represent the first positive evidence of interactions between the pMMO and Cu–mb.
The role of Dam methylation in phase variation of Haemophilus influenzae genes involved in defence against phage infection
Authors: Zaleski Piotr , Wojciechowski Marek , Piekarowicz Andrzej
Published: October 2005

Haemophilus influenzae uses phase variation (PV) to modulate the activity of its defence systems against phage infection. The PV of the restriction–modification (R-M) system HindI, the main defence system against phage infection and incoming chromosomal and phage DNA in H. influenzae Rd, is driven by changes of the pentanucleotide repeat tract within the coding sequence of the hsdM gene and is influenced by lack of Dam methylation. Phase-variable resistance/sensitivity to phage infection correlates with changes in lipooligosaccharide (LOS) structure and occurs by slippage of tetranucleotide repeats within the gene lic2A, coding for a step in the biosynthesis of LOS. The lack of Dam activity destabilizes the tetranuclotide (5′-CAAT) repeat tract and increases the frequency of switching from sensitivity to resistance to phage infection more than in the opposite direction. The PV of the lgtC gene does not influence resistance or sensitivity to phage infection. Insertional inactivation of lic2A, but not lgtC or lgtF, leads to resistance to phage infection and to the same structure of the LOS as observed among phase-variable phage-resistant variants. This indicates that in the H. influenzae Rd LOS only the first two sugars (Glc-Gal) extending from the third heptose are part of bacterial phage receptors.
Organization of the biosynthetic gene cluster for the macrolide concanamycin A in Streptomyces neyagawaensis ATCC 27449
Authors: Haydock Stephen F. , Appleyard Anthony N. , Mironenko Tatiana , Lester John , Scott Natasha , Leadlay Peter F.
Published: October 2005

The macrolide antibiotic concanamycin A has been identified as an exceptionally potent inhibitor of the vacuolar (V-type) ATPase. Such compounds have been mooted as the basis of a potential drug treatment for osteoporosis, since the V-ATPase is involved in the osteoclast-mediated bone resorption that underlies this common condition. To enable combinatorial engineering of altered concanamycins, the biosynthetic gene cluster governing the biosynthesis of concanamycin A has been cloned fromStreptomyces neyagawaensisand shown to span a region of over 100 kbp of contiguous DNA. An efficient transformation system has been developed forS. neyagawaensisand used to demonstrate the role of the cloned locus in the formation of concanamycin A. Sequence analysis of the 28 ORFs in the region has revealed key features of the biosynthetic pathway, in particular the biosynthetic origin of portions of the backbone, which arise from the unusual polyketide building blocks ethylmalonyl-CoA and methoxymalonyl-ACP, and the origin of the pendant deoxysugar moiety 4′-O-carbamoyl-2′-deoxyrhamnose, as well as the presence of a modular polyketide synthase (PKS) encoded by six giant ORFs. Examination of the methoxymalonyl-specific acyltransferase (AT) domains has led to recognition of an amino acid sequence motif which can be used to distinguish methylmalonyl-CoA- from methoxymalonyl-ACP-specific AT domains in natural PKSs.
Two new major subunits in the cellulosome of Clostridium thermocellum: xyloglucanase Xgh74A and endoxylanase Xyn10D
Authors: Zverlov Vladimir V. , Schantz Nicolaus , Schmitt-Kopplin Philippe , Schwarz Wolfgang H.
Published: October 2005

The structure and enzymic activity of xyloglucanase Xgh74A and endoxylanase Xyn10D, components in the cellulosomes of cellulose-grown Clostridium thermocellum, were determined. Xyn10D is a thermostable endo-1,4-β-xylanase with a module composition identical to Xyn10C (CBM22-GH10-Doc). It hydrolyses xylan and mixed-linkage 1,3-1,4-β-glucan with a temperature optimum of 80 °C. Xyloglucanase Xgh74A contains a catalytic module of GHF74 in addition to a C-terminal dockerin module. It hydrolyses every fourth β-1,4-glucan bond in the xyloglucan backbone, thus producing decorated cellotetraose units. Its low activity on CMC and lack of activity on amorphous cellulose indicates recognition of the xylosidic side chains present in xyloglucan, which is readily hydrolysed (295 U mg−1). The pattern of the hydrolysis products from tamarind xyloglucan resembles that of other GHF74 xyloglucan endoglucanases. The data indicate that Xgh74A and Xyn10D contribute to the in vivo degradation of the hemicelluloses xyloglucan and xylan by the cellulosome of C. thermocellum. Xgh74A is the first xyloglucanase identified in C. thermocellum and the only enzyme in the cellulosome that hydrolyses tamarind xyloglucan.
Dimeric Brucella abortus Irr protein controls its own expression and binds haem
Authors: Martínez Marcela , Ugalde Rodolfo A. , Almirón Marta
Published: October 2005

Brucella abortus needs to synthesize haem in order to replicate intracellularly and to produce virulence in mice. Thus, to gain insight into the pathogenesis of the bacterium, regulatory proteins of the haem biosynthetic pathway were sought. An iron response regulator (Irr) from Bradyrhizobium japonicum, which is a close relative of Brucella, was previously described as being involved in the coordination of haem biosynthesis and iron availability. The Bru. abortus genome was searched for an irr orthologue gene, and the Bru. abortus irr gene was cloned, sequenced and disrupted. A null mutant was constructed that accumulated protoporphyrin IX under conditions of iron deprivation. This phenotype was overcome by a complementing plasmid carrying the wild-type irr. Purified recombinant Bru. abortus Irr behaved as a stable dimer and bound haem. Interestingly, in vivo, Irr was only detected in cells obtained from iron-limited cultures and the protein downregulated its own transcription. Through lacZ fusion, it was demonstrated that iron did not regulate irr transcription. The data reported show that Bru. abortus Irr is a homodimeric protein that is accumulated in iron-limited cells, controls its own transcription and downregulates the biosynthesis of haem precursors.
Evolution and diversification of a sexually dimorphic luminescent system in ponyfishes (Teleostei: Leiognathidae), including diagnoses for two new genera
Authors: Sparks John S. , Dunlap Paul V. , Smith W. Leo
Published: October 2005

A phylogeny was generated for Leiognathidae, an assemblage of bioluminescent, Indo‐Pacific schooling fishes, using 6175 characters derived from seven mitochondrial genes (16S,COI,ND4,ND5,tRNA‐His,tRNA‐Ser,tRNA‐Leu), two nuclear genes (28S,histone H3), and 15 morphological transformations corresponding to features of the fishes' sexually dimorphic light‐organ system (LOS; e.g., circumesophageal light organ, lateral lining of the gas bladder, transparent flank and opercular patches). Leiognathidae comprises three genera,Gazza,Leiognathus, andSecutor. Our results demonstrate that Leiognathidae,Gazza, andSecutorare monophyletic, whereasLeiognathusis not. The recovered pattern of relationships reveals that a structurally complex, strongly sexually dimorphic and highly variable species‐specific light organ is derived from a comparatively simple non‐dimorphic structure, and that evolution of other sexually dimorphic internal and external features of the male LOS are closely linked with these light‐organ modifications. Our results demonstrate the utility of LOS features, both for recovering phylogeny and resolving taxonomic issues in a clade whose members otherwise exhibit little morphological variation. We diagnose two new leiognathid genera,PhotopectoralisandPhotoplagios, on the basis of these apomorphic LOS features and also present derived features of the LOS to diagnose several additional leiognathid clades, includingGazzaandSecutor. Furthermore, we show that five distinct and highly specialized morphologies for male‐specific lateral luminescence signaling, which exhibit species‐specific variation in structure, have evolved in these otherwise outwardly conservative fishes. Leiognathids inhabit turbid coastal waters with poor visibility and are often captured in mixed assemblages of several species. We hypothesize that the species‐specific, sexually dimorphic internal and external modifications of the leiognathid LOS provide compelling evidence for an assortative mating scheme in which males use species‐specific patterns of lateral luminescence signaling to attract mates, and that this system functions to maintain reproductive isolation in these turbid coastal environments.

© The Willi Hennig Society 2005.
Generating implied alignments under direct optimization using POY
Authors: Giribet Gonzalo
Published: October 2005

The concept of implied alignments, as used in modern systematics literature, is discussed. Basic notions about how to generate implied alignments are given together with explicit examples. The objective of this article is to guide researchers on how to produce implied alignments for validating purposes in an efficient manner using POY.

© The Willi Hennig Society 2005.
Targeting optimal introns for phylogenetic analyses in non‐model taxa: experimental results in Asian pitvipers
Authors: Creer Simon , Malhotra Anita , Thorpe Roger S. , Pook Catharine E.
Published: October 2005

Nuclear introns are increasingly used as phylogenetic markers. Here, we present a multidisciplinary approach towards optimal locus selection and amplification using Asian pitvipers as an example of a non‐model taxon, and raise the profile of length variant heterozygotes (LVHs) in intron loci. Taxon‐specific primers were identified using a bioinformatic approach, and also designed from existing exon primed, intron crossing (EPIC) primer amplifications. Eleven further universal EPIC primer pairs were assayed using a range of PCR optimization strategies. Taxon‐specific primers yielded the most consistent amplifications, but assaying a large number of universal EPIC primers yielded another appropriate locus for phylogenetic purposes. Modified Taq DNA polymerases such as JumpStart™Taq either significantly improved the specificity and yield of EPIC PCR amplifications (of low copy number nuclear targets), or resulted in amplifications that were not significantly worse than those derived from a generic Taq DNA polymerase. Finally, LVHs were detected in all loci that were sequenced suggesting that they are relatively common in introns. This study provides an efficient and cost effective template for the successful identification of intron markers for molecular systematics which is universally applicable to other non‐model taxon groups.

© The Willi Hennig Society 2005.
Survey of paediatric case management practices in Australia for children and young people with acquired brain injury (ABI)
Authors: Scheinberg A. M. , Gibson W. , Hughes D. , Miles A. , Murphy P. , Noronha J.
Published: October 2005

Aim  To investigate the characteristics of paediatric acquired brain injury (ABI) case managers and their organizations in Australia, and to determine the ways in which case managers deliver their service, maintain professional development and evaluate outcomes.

Methods  Postal survey of 57 known paediatric ABI case managers working at 24 separate organizations in Australia.

Results  Out of 40 questionnaires returned (70%), the majority of respondents had been providing case management for over 5 years (55%). Co‐ordination of services was ranked the most important component of case management, with services most frequently delivered by telephone. Evaluation of case management outcomes was reported by only 52% of the case managers. Whilst the majority of case managers had received some form of training (83%), this varied widely from informal in‐house training, to workshops run by insurance companies and government agencies.

Discussion  This survey provides information about current case management practices for children with ABI in Australia. Detailing what case managers do is a first step towards developing controlled research designs which are required to demonstrate whether case management is effective in terms of clinical outcomes and cost–benefit.
Audit of local performance compared with standards recommended by the national guidelines for aetiologic investigation of permanent childhood hearing impairment
Authors: Yoong S. , Spencer N.
Published: October 2005

Background  National guidelines for aetiologic investigation of childhood deafness were developed as the Newborn Hearing Screening Program (NHSP) was being implemented in the United Kingdom. This guidance document was expected to be incorporated into the operational procedure of the NHSP.

Method  This criterion‐based audit compared local care set against developed guidelines that can be used to assess the appropriateness of specific investigations, services and outcomes. Data on children diagnosed to have sensorineural deafness from March 2002–2004 were extracted from an established computerized database for analysis.

Results  Forty‐seven children were included; 17 have bilateral severe to profound hearing loss, 25 have bilateral mild to moderate loss and 5 with unilateral loss. A high proportion of Pakistani children were from consanguineous marriages with a family history of deafness. Total 29.8% of children were diagnosed through newborn screening and 70.2% detected through hearing surveillance programmes. For children with bilateral severe to profound deafness, 53.0% accepted, 5.9% declined and 41.2% were not offered imaging of their inner ears. A total of 47.1% accepted and 52.9% declined electrocardiograph (ECG) evaluation. Total 70.6% accepted and 29.4% declined connexin mutations testing. Parental requests were required for those with lesser degree of hearing loss. Total 24% accepted, 28% declined and 48% were not offered connexin testing. None were offered ECG and imaging. Testing for congenital infections was inappropriate for children over 1 year old. Ten subjects accepted and five declined this investigation. In the total group, 63.8% accepted, 17.0% declined and 19.1% were not offered referral to the ophthalmic service. Total 46.8% accepted, 44.7% declined and 8.5% were not offered referral to genetics service. Investigations resulted in two connexin‐positive children with moderate loss.

Conclusion  Our study identified key areas where guidelines were not followed. These were related to lack of funding and parental choice. This sample has a higher connexin ‘hit’ rate for lesser degree deafness.
Parental reported bullying among Nordic children: a population‐based study
Authors: Nordhagen R. , Nielsen A. , Stigum H. , Köhler L.
Published: October 2005

Background  Bullying has been shown to be a serious problem amongst school children, but few studies have been population‐based and included pre‐school children.

Methods  The study is part of a cross‐sectional comparative study in 1984 and 1996, focusing on children's and their families’ health and welfare in the Nordic countries. At each point of time parents of 3000 randomly selected children aged 2–17 years in each of five Nordic countries received a postal questionnaire. Altogether approximately 20 000 questionnaires were completed. The prevalence of bullying, risk factors for bullying and possible effect factors were analysed.

Results  Parents reported bullying of their child in 15.1% of the cases. Bullying varied from 7.2% in Sweden to about 20% in Denmark and Finland. There was a small increase in bullying from 13.7% in 1984 to 16.4% in 1996. Bullying was most frequent in boys (OR: 1.4) and in children 2–6 and 7–12 years old (OR: 2.0 and 2.2 compared with older children). Children of single parents and of parents with low education had increased risks (OR: 1.4 and 1.4). Children with chronic conditions had higher risks for being bullied (OR: 2.3). In 1996 children with psychiatric/nervous problems and hyperactivity had high risks for being bullied (OR: 8.8 and 10.5) and for bullying others (3.9 and 3.5). Being bullied was associated with poor thriving and psychosomatic and psychological problems. No countries had national interventions before 1984, but Sweden had early focused on the problem and implemented a strong national policy before 1996. After 1996 national anti‐bullying policies were strengthened in the Nordic countries, most in Sweden and Norway.

Conclusion  Bullying is common among Nordic children, including pre‐school children. Bullying is a threat to children's health, and augments problems in children with chronic conditions. The low prevalence of bullying in Sweden may be a result from sustained, strong anti‐bullying policies. There is still a need for continuing interventions.
Symptoms of inattention and hyperactivity in children with habitual snoring: evidence from a community‐based study in Istanbul
Authors: Arman A. R. , Ersu R. , Save D. , Karadag B. , Karaman G. , Karabekiroglu K. , Karakoc F. , Dagli E. , Berkem M.
Published: October 2005

Background  Neurobehavioural symptoms of inattention and hyperactivity are common in children with sleep‐disordered breathing (SDB). Prevalence rates of habitual snoring and attention deficit hyperactivity disorder (ADHD) are very similar and both have a substantial negative effect on children's behavioural health.

Objective  We examined the differences for subjective attentional and hyperactivity measures reported by parents and teachers among primary school children with habitual snoring and age‐ and sex‐matched controls in a community‐based case‐control study in Istanbul.

Methods  In 2002, a survey was carried out to determine the prevalence of snoring in 2147 primary school children. After one year, in 2003, 151 children with habitual snoring and 302 controls from this survey were studied with parental SDB questionnaire, Conners’ Parent (Conners‐P) and Teacher Scales, and an inattention hyperactivity scale (IHS). Exclusion criteria included history of ADHD diagnosis, controls who started to snore and habitual snorers (HS) who no longer snored in this follow‐up study.

Results  Ninety‐six HS and 190 control subjects (mean age: 9.4 ± 1.3) were evaluated. HS had significantly more symptoms of hyperactivity (Conners‐ADHD index) (P: 0.033), attentional (P: 0.019), and conduct and oppositional defiant in subscales (P: 0.001) of Conners‐P and IHS‐Parents. A pooled score of Conners‐P ADHD Index > 60 and IHS‐Parent score > 1.25 showed considerable difference in HS when compared with controls (5.1% vs. 1.4%) (P < 0.0001). Daytime hyperactivity and excessive daytime sleepiness reported by parents correlated with scores of Conners‐P and IHS‐P (P < 0.01). Teachers’ observations showed significant correlations with learning disability and the level of academic performance in HS (P < 0.01). Other behavioural parameters related to SDB were not significantly correlated with teachers’ ADHD ratings in HS.

Conclusion  Increased rates of moderate hyperactivity as well as conduct and oppositional defiant symptoms in HS reported by the parents might reflect a negative impact on overall neurobehavioural health. The teachers’ scores yielded no significant results among HS and controls. This may be caused by the limitation due to shared method variance. The negative effect of crowded classes on teachers’ evaluations must be also taken into consideration. After exclusion of a diagnosis of ADHD in children presenting with hyperactivity and inattention, children with habitual snoring with prominent scores of behavioural measures should be considered as candidates for further assessment by a sleep specialist.