bluuebunny/crossref_metadata_2025 huggingface dataset:
Modelling late Quaternary changes in plant distribution, vegetation and climate using pollen data from Georgia, Caucasus
Authors: Connor Simon E. , Kvavadze Eliso V.
Published: November 2008

Aim  To use pollen data, numerical analysis and modelling to reconstruct late Quaternary vegetation and climate in a complex, mountainous environment.

Location  Georgia (Caucasus region).

Methods  Pollen data were assembled from various sources and used to map: (1) changing frequencies of individual taxa; (2) vegetation changes; and (3) reconstructed climatic parameters for the past 14,000 years. Numerical analyses were performed using two‐way indicator species analysis (twinspan), detrended correspondence analysis (DCA), the modern analogue technique (MAT) and weighted averaging (WA).

Results  Mapping of pollen taxa showed that Chenopodiaceae, Artemisia and Ephedra were most abundant in the study area during the late‐glacial. Betula and Corylus expanded during the early Holocene, yielding to Abies, Carpinus, Fagus, Quercus and Castanea. Picea, Pinus, Juglans and Ostrya‐type expanded during the late Holocene. Mixed forests grew in the moist, Black Sea refugium throughout the late Quaternary. Elsewhere in Georgia, the Pleistocene–Holocene transition is recorded as a shift from desert‐steppes to oak‐xerophyte communities and mixed forests. This kind of vegetation remained relatively stable until the mid–late Holocene, when coniferous forests and mountain grasslands advanced. DCA showed that rainfall was most strongly correlated with pollen composition in the study area (r2 = 0.55). No temperature signal was detected. A weighted‐averaging transfer function linking pollen percentages to annual precipitation was selected over a MAT model as it performed better when applied to a validation data set. Rainfall reconstructions indicate widespread aridity at the terminal Pleistocene, followed by a gradual increase in precipitation, peaking during the mid Holocene (7000–4000 cal. yr bp) and generally decreasing thereafter.

Main conclusions  On a regional scale, the results confirm those from previous studies of palaeovegetation and palaeoclimate in Western Asia. On a local scale, reconstructions from individual sites often diverge from the regional trend because of edaphic changes, ecological succession, human impacts and other disturbances. Some of these factors are probably responsible for the increasing heterogeneity of Georgia’s vegetation in the latter half of the Holocene.
Microrefugia
Authors: Rull Valentí
Published: November 2008

The concept of microrefugia is widely accepted to explain the post‐glacial colonization of continents, which led to the present‐day biotic configuration of the Earth. However, so far, microrefugia are no more than a ‘theoretical necessity’ without an appropriate biogeographical and ecological characterization. Here, a tentative definition and a classification of microrefugia, based on the existing literature, are proposed for discussion. A deeper biogeographical analysis of the concept is suggested using novel ecological models and methods, in order to develop new hypotheses to be tested with palaeoecological and molecular phylogenetic tools.
Marine, intertidal, freshwater and terrestrial biodiversity of an isolated polar archipelago
Authors: Barnes David K. A. , Kaiser Stefanie , Griffiths Huw J. , Linse Katrin
Published: November 2008

Aim  We ask how biodiverse is a polar archipelago; how this faunal richness is spread across marine, intertidal, freshwater, terrestrial and parasitic realms; and how fast species are accumulated with increased sampling effort.

Location  The South Orkney Islands (SOI), Scotia Arc, Southern Ocean.

Methods  We sampled mega‐ and macro‐benthos at the SOI using scuba in the shallows (0–10 m), a rough bottom otter trawl at 150–250 m and an Agassiz trawl and epibenthic sledge, both at depths of 200, 500, 1000 and 1500 m. We also collated species occurrence at the SOI in each realm from a century of literature and modern databases to investigate patterns in species accumulation, endemism, faunistic affinities and bathymetric ranges in three model taxa.

Results  Our 11 benthic samples showed that point biodiversity at the SOI is high, yielding 19 classes and 158 species. Nearly a third were new to the area, whilst five species and one genus were new to science. The shallowest samples were richest but had fewest new records of species. Known richness at the SOI is dominated by marine species (1026), of which 821 (83.3%) were benthic. Across all realms, 1224 species (50 classes, 24 phyla) were recorded, of which 43 were intertidal, 64 freshwater, 100 terrestrial, 60 parasitic and 40 birds. Species accumulation curves for model taxa showed new sampling yields about 0.75% per sample of known benthic richness, so by Antarctic standards we know the SOI quite well. Most species are Southern Ocean endemics, but very few occur only at the SOI.

Main conclusions  This first estimate of faunal biodiversity of a polar locality demonstrates both high richness and high levels of knowledge at the SOI. As suspected but never quantified, the benthos dominates polar biodiversity, at least at the SOI. Marine species there constitute 20% of those recently listed for the entire Southern Ocean, whilst > 60% of terrestrial species are known from Antarctica. The SOI, being one of the better‐studied polar locations, of known age and with a discrete shelf, represent an important source of comparison for biodiversity studies. Our data clearly show that richness and our knowledge of the polar fauna differ across environments.
Phylogeography: retrospect and prospect
Authors: Avise John C.
Published: November 2008

Phylogeography has grown explosively in the two decades since the word was coined and the discipline was outlined in 1987. Here I summarize the many achievements and novel perspectives that phylogeography has brought to population genetics, phylogenetic biology and biogeography. I also address future directions for the field. From the introduction of mitochondrial DNA assays in the late 1970s, to the key distinction between gene trees and species phylogenies, to the ongoing era of multi‐locus coalescent theory, phylogeographic perspectives have consistently challenged conventional genetic and evolutionary paradigms, and they have forged empirical and conceptual bridges between the formerly separate disciplines of population genetics (microevolutionary analysis) and phylogenetic biology (in macroevolution).
Local–regional diversity relationship varies with spatial scale in lotic diatoms
Authors: Soininen Janne , Heino Jani , Kokocinski Mikolaj , Muotka Timo
Published: November 2008

Aim  Ecologists have shown increasing interest in the relative roles of local and regional factors in structuring biotic communities. One approach to studying this is to examine the relationship between local species richness (LSR) and regional species richness (RSR). We examined the LSR–RSR relationship in stream diatoms, using two data sets that varied in spatial extent. At broad spatial extent ranging across drainage systems, we expected climatic and dispersal‐related factors to constrain LSR, thus resulting in a linear LSR–RSR relationship. However, at small spatial scales dispersal across sites should be unconstrained, resulting in strong local interactions and a weak or asymptotic LSR–RSR relationship.

Location  Boreal streams in Finland.

Methods  For data set 1, we sampled 15 stream riffles (localities) in each of eight drainage systems (regions), with the latitudinal gradient between the southernmost and northernmost sites being almost 1100 km. For data set 2, a locality for estimating LSR was a single stone, and each riffle represented a region for estimating RSR. We sampled 20 stones in each of eight riffles. We used linear regressions to examine the relationship between LSR and RSR across regions. We used both observed richness values, as well as values estimated with the Chao1 estimator.

Results  We found a relatively strong linear relationship between the Chao1‐estimated mean LSR and RSR (R2 = 0.654, P = 0.015) across drainage systems. The slope of the regression was 0.643 and it did not differ from 1.0, thus indicating linearity. At the riffle scale, however, LSR and RSR were not linearly related, and the slope of the regression (0.039) differed significantly from 1.0, indicating curvilinearity.

Main conclusions  These results suggest that the relationship between mean LSR and RSR varies across spatial scales in diatoms – from significantly linear at large scales to curvilinear at small scales. These plots imply strong regional enrichment in stream diatoms across drainage systems. Their diversity is thus determined largely by the composition of the regional species pool, as also in many macroorganisms. In contrast, at small spatial scales the LSR–RSR relationship implied a hard limit to local diversity, reflecting the primacy of local processes.
Possible impact of inflammatory status on C‐telopeptide pyridinoline cross‐links of Type I collagen and osteocalcin levels around oral implants with peri‐implantitis: a controlled clinical trial
Authors: TÜMER C. , AKSOY Y. , GÜNCÜ G. N. , NOHUTCU R. M. , KILINC K. , TÖZÜM T. F.
Published: November 2008

Summary  Detection of progression level of peri‐implantitis may help in the prevention of oral implant failure. C‐telopeptide pyridinoline crosslinks of Type I collagen (ICTP) and osteocalcin (OC) are specific markers of bone turnover and bone degradation. Determination of the ICTP and OC levels in the peri‐implant sulcus fluid (PISF) may predict the metabolic and/or inflammatory changes in the peri‐implant bone. The aim of this clinical study was to evaluate ICTP and OC levels in the PISF for oral implants with and without peri‐implant bone destruction and correlate these levels with the traditional clinical peri‐implant parameters (probing depth, plaque index, gingival index and gingival bleeding time index) and radiographic bone level measurements. Fifteen patients with 30 peri‐implant sites with bone destruction (radiographic bone loss) and health were included. Clinical parameters were measured and PISF was collected from the sites. Peri‐implant sulcus fluid ICTP and OC levels were detected by radioimmunoassay technique from PISF samples. All clinical parameters demonstrated a significant increase in peri‐implantitis sites compared with healthy sites. The PISF volume of the peri‐implantitis sites was also significantly higher than of the healthy peri‐implant sites. Although not statistically significant, a trend of increase was demonstrated in ICTP PISF samples sampled from peri‐implantitis sites compared with healthy sites. A significant increase was noticed for OC PISF level in peri‐implantitis sites compared with healthy ones. As well as peri‐implant clinical measurements, volumetric changes at PISF may be counted as an important clinical parameter to distinguish the bone destruction sites from healthy sites around oral implants.
Effects of tooth loss and denture wear on tongue‐tip motion in elderly dentulous and edentulous people
Authors: YOSHIKAWA M. , YOSHIDA M. , NAGASAKI T. , TANIMOTO K. , TSUGA K. , AKAGAWA Y.
Published: November 2008

Summary  The purpose of this study was to clarify quantitatively the differences in tongue‐tip motion among the dentulous elderly people and also among the elderly edentulous, both with and without their dentures and, to identify the influence of tooth loss and denture wear on tongue‐tip motion. Fourteen young dentulous people, 12 elderly dentulous people and 13 elderly edentulous people participated in this study. Subjects were asked to swallow a 10 mL barium sulfate solution three times. The elderly edentulous people were asked to swallow the solution while wearing dentures and with dentures removed. Functional swallowing was recorded on cine‐film with a digital subtraction angiography system. Lateral cinefluorography images were obtained from seated subjects. Using a cine‐projector, the movements of the tongue surface were traced as dots and lines frame by frame on a single tracing sheet within a definite period of time from the beginning of the oral phase to the end of the pharyngeal phase. With counting the number of ‘trajectories’ of tongue‐tip motion, tongue movements were classified as ‘stable’ and ‘hyperactive’ types. The results was that significantly more ‘hyperactive’ type subjects were found among the elderly edentulous who were not wearing dentures (12 of 13) compared with the dentulous young (1 of 14), the elderly dentulous (1 of 13) or the elderly edentulous wearing dentures (1 of 13) (P < 0.001). The tongue‐tip motion for the ‘hyperactive’ type was very complex and the tongue‐tip anchoring against the palate was always instable.
Cytotoxicity of one‐step dentin‐bonding agents toward dental pulp and odontoblast‐like cells
Authors: YASUDA Y. , INUYAMA H. , MAEDA H. , AKAMINE A. , NÖR J. E. , SAITO T.
Published: November 2008

Summary  The purpose of this study was to compare the cytotoxicity of five one‐step dentin‐bonding agents on human dental pulp and odontoblast‐like cells (MDPC‐23). Photopolymerized and unpolymerized samples of these dentin‐bonding agents were prepared and incubated with dental pulp or MDPC‐23 cells. After 24 or 72 h of incubation, the number of unstained cells with trypan blue was counted. The staining of cells with trypan blue stands for a cytotoxicity. The pulp cell and MDPC‐23 cytotoxicity of polymerized sample treatment increased in the order of AQ Bond Plus (AQ)<Clearfil Tri‐S Bond (TS)=G‐bond (GB)<Absolute (AB)<Adper Prompt (AP) for 24 and 72 h. The pulp cell cytotoxicity of unpolymerized sample treatment for 24 h increased in the order of AQ<GB = AB<TS<AP. The MDPC‐23 cytotoxicity of unpolymerized sample treatment for 24 h increased in the order of AQ<GB<TS = AB<AP. Whether polymerized or unpolymerized, AQ was the least cytotoxic agent, while AP was the strongest. All polymerized dentin‐bonding agents exhibited lower cytotoxicity by 2–65% than their unpolymerized counterparts. The appearance of the cytotoxicity of dentin‐bonding agents was time‐dependent, and cell viability was lower at 72 h by 2–46% than at 24 h. The cytotoxicity to MDPC‐23 cells was about 5–24% higher than that to pulp cells. These results indicate that one‐step dentin‐bonding agents differ markedly in their cytotoxicity. Differential cytotoxic effects of one‐step dentin‐bonding agents should be considered during clinical application of operative restoration.
In vitro evaluation of the implant abutment connection sealing capability of different implant systems
Authors: COELHO P. G. , SUDACK P. , SUZUKI M. , KURTZ K. S. , ROMANOS G. E. , SILVA N. R. F. A.
Published: November 2008

Summary  This study sought to evaluate the sealing capability of the implant abutment connection of different dental implant systems. Five Nobel Replace select, Straumann and Intra‐lock implants of approximately 4·5 mm diameter with their respective abutments were provided by the manufacturers. A calibration curve was determined by placing toluidine blue (TB) increments of 0·1 μL into 1·5 mL of distilled water and recording its absorbance in a spectrophotometer until reaching 0·7 μL. Then, 0·7 μL of TB was placed in the deepest portion of each implant’s internal screw, the abutments were adapted to the implant according to the manufacturer’s instructions and the specimens were placed in vials with 1·5 mL of distilled water. Spectrophotometric analysis was performed at 1, 3, 6, 24, 48, 72, 96 and 144 h. Statistical analysis was performed by One‐way anova at 95% level of significance. The calibration curve was linear with respect to the TB amount in 1·5 μL distilled water (R2 = 0·9961). All implant abutment systems presented an increase in absorbance as a function of time. As time elapsed in vitro, significantly higher amounts of TB was released from the Straumann and Nobel Replace Select connection systems (P < 0·0001). Leakage was significant between the groups. Despite controlled torquing, the seal between the implant body and the abutment could not be maintained in all three of the systems tested.
Nonsteroidal anti‐inflammatory agents and musculoskeletal injuries in Thoroughbred racehorses in Kentucky
Authors: DIRIKOLU L. , WOODS W. E. , BOYLES J. , LEHNER A. F. , HARKINS J. D. , FISHER M. , SCHAEFFER D. J. , TOBIN T.
Published: November 2008

Injuries sustained by horses during racing have been considered as an unavoidable part of horse racing. Many factors may be associated with the musculoskeletal injuries of Thoroughbred race horses. This study surveyed the amounts of nonsteroidal anti‐inflammatory agents (NSAIDs) in injured horse’s biological system (plasma) at Kentucky racetracks from January 1, 1995 through December 31, 1996. During that period, there were 84 catastrophic cases (euthanized horses) and 126 noncatastrophic cases. Plasma concentrations of NSAIDs were determined by High Performance Liquid Chromatography in injured and control horses. The possible role of anti‐inflammatory agents in musculoskeletal injuries of Thoroughbred race horses was investigated by comparing the apparent concentrations of NSAIDs in injured horses to concentrations in control horses. The plasma concentrations of phenylbutazone and flunixin were higher in injured horses than in control horses. Most injured and control horses did not have a detectable level of naproxen in their plasma samples. Further studies must be carried out to determine whether horses with higher plasma concentrations of NSAIDs have an altered risk of musculoskeletal injuries compared with other horses.
Pharmacokinetics of intravenous ceftiofur sodium and concentration in body fluids of foals
Authors: MEYER S. , GIGUÈRE S. , RODRIGUEZ R. , ZIELINSKI R. J. , GROVER G. S. , BROWN S. A.
Published: November 2008

The objectives of this study were to determine pharmacokinetics of intravenous (i.v.) ceftiofur in foals, to compare ultra‐high performance liquid chromatography tandem mass spectometry (UPLC‐MS/MS) and microbiologic assay for the measurement of ceftiofur concentrations, and to determine the minimum inhibitory concentration (MIC) of ceftiofur against common equine bacterial pathogens. In a cross‐over design, ceftiofur sodium was administered i.v. to six foals (1–2 days‐of‐age and 4–5 weeks‐of‐age) at dosages of 5 and 10 mg/kg. Subsequently, five doses of ceftiofur were administered i.v. to six additional foals between 1 and 5 days of age at a dose of 5 mg/kg q 12 h. Concentrations of desfuroylceftiofur acetamide (DCA), the acetamide derivative of ceftiofur and desfuroylceftiofur‐related metabolites were measured in plasma, synovial fluid, urine, and CSF by use of UPLC‐MS/MS. A microbiologic assay was used to measure ceftiofur activity for a subset of plasma samples. Following i.v. administration of ceftiofur at a dose of 5 mg/kg to 1–2 day‐old foals, DCA had at½of 7.8 ± 0.1 h, a body clearance of 74.4 ± 8.4 mL/h/kg, and an apparent volume of distribution of 0.83 ± 0.09 L/kg. After multiple i.v. doses at 5 mg/kg, DCA concentrations in CSF were significantly lower than concurrent plasma concentrations. Ceftiofur activity using a microbiologic assay significantly underestimated plasma concentrations of DCA. TheMICof ceftiofur required to inhibit growth of 90% of isolates ofEscherichia coli,Pasteurellaspp,Klebsiellaspp, and β‐hemolytic streptococci was <0.5 μg/mL. Intravenous administration of ceftiofur sodium at the rate of 5 mg/kg every 12 h would provide sufficient coverage for the treatment of susceptible bacterial isolates.
A matter of life and death: cell wall homeostasis and the WalKR (YycGF) essential signal transduction pathway
Authors: Dubrac Sarah , Bisicchia Paola , Devine Kevin M. , Msadek Tarek
Published: November 2008

The WalK/WalR (aka YycG/YycF) two‐component system (TCS), originally identified in Bacillus subtilis, is very highly conserved and specific to low G+C Gram‐positive bacteria, including a number of important pathogens. An unusual feature is that this system is essential for viability in most of these bacteria. Recent studies have revealed conserved functions for this system, defining this signal transduction pathway as a crucial regulatory system for cell wall metabolism, that we have accordingly renamed WalK/WalR. Here we review the cellular role of the WalK/WalR TCS in different bacterial species, focusing on the function of genes in its regulon, as well as variations in walRK operon structure and the composition of its regulon. We also discuss the nature of its essentiality and the potential type of signal being sensed. The WalK histidine kinase of B. subtilis has been shown to localize to the divisome and we suggest that the WalKR system acts as an information conduit between extracytoplasmic cellular structures and intracellular processes required for their synthesis, playing a vital role in effectively co‐ordinating peptidoglycan plasticity with the cell division process.
The Pta–AckA pathway controlling acetyl phosphate levels and the phosphorylation state of the DegU orphan response regulator both play a role in regulating Listeria monocytogenes motility and chemotaxis
Authors: Gueriri Ibtissem , Bay Sylvie , Dubrac Sarah , Cyncynatus Camille , Msadek Tarek
Published: November 2008

DegU is considered to be an orphan response regulator in Listeria monocytogenes since the gene encoding the cognate histidine kinase DegS is absent from the genome. We have previously shown that DegU is involved in motility, chemotaxis and biofilm formation and contributes to L. monocytogenes virulence. Here, we have investigated the role of DegU phosphorylation in Listeria and shown that DegS of Bacillus subtilis can phosphorylate DegU of L. monocytogenes in vitro. We introduced the B. subtilis degS gene into L. monocytogenes, and showed that this leads to highly increased expression of motility and chemotaxis genes, in a DegU‐dependent fashion. We inactivated the predicted phosphorylation site of DegU by replacing aspartate residue 55 with asparagine and showed that this modified protein (DegUD55N) is no longer phosphorylated by DegS in vitro. We show that although the unphosphorylated form of DegU retains much of its activity in vivo, expression of motility and chemotaxis genes is lowered in the degUD55N mutant. We also show that the small‐molecular‐weight metabolite acetyl phosphate is an efficient phosphodonor for DegU in vitro and our evidence suggests this is also true in vivo. Indeed, a L. monocytogenesΔptaΔackA mutant that can no longer synthesize acetyl phosphate was found to be strongly affected in chemotaxis and motility gene expression and biofilm formation. Our findings suggest that phosphorylation by acetyl phosphate could play an important role in modulating DegU activity in vivo, linking its phosphorylation state to the metabolic status of L.  monocytogenes.
Spa32 interaction with the inner‐membrane Spa40 component of the type III secretion system of Shigella flexneri is required for the control of the needle length by a molecular tape measure mechanism
Authors: Botteaux Anne , Sani Musa , Kayath Christian A. , Boekema Egbert J. , Allaoui Abdelmounaaïm
Published: November 2008

The effectors of enterocyte invasion by Shigella are dependent on a type III secretion system that contains a needle whose length average does not exceed 50 nm. Previously, we reported that Spa32 is required for needle length control as well as to switch substrate specificity from MxiH to Ipa proteins secretion. To identify functional domains of Spa32, 11 truncated variants were constructed and analysed for their capacity (i) to control the needle's length; (ii) to secrete the Ipa proteins; and (iii) to invade HeLa cells. Deletion at either the N‐terminus or C‐terminus affect Spa32 function in all cases, but Spa32 variants lacking internal residues 37–94 or 130–159 retained full Spa32 function. Similarly, a Spa32 variant obtained by inserting of the YscP's ruler domain retained Spa32 function although it programmed slightly elongated needles. Using the GST pull‐down assay, we show that residues 206–246 are required for Spa32 binding to the C‐terminus of Spa40, an inner membrane protein required for Ipa proteins secretion. Our data clearly demonstrate that shortening Spa32 affects the length of the needle in a comparable manner to the spa32 mutant, indicating that the control of needle length does not require a molecular ruler mechanism.
Ubiquitin‐dependent and ‐independent mitochondrial protein quality controls: implications in ageing and neurodegenerative diseases
Authors: Germain Doris
Published: November 2008

The ubiquitin‐independent protein quality control of matrix proteins of the mitochondrion is well characterized and until recently the mitochondrion was considered a ‘ubiquitination‐free’ organelle. However, a number of studies now indicate multiple roles of the ubiquitin–proteasome pathway in the regulation and maintenance of mitochondrial integrity. Of particular interest to this review is the finding of a mitochondrial ubiquitin‐dependent protein quality control and that this pathway may share similarity to the endoplasmic reticulum‐associated degradation (ERAD) pathway that acts to eliminate misfolded proteins from the lumen of the endoplasmic reticulum. The potential cross‐talk between the ubiquitin‐dependent and ‐independent protein quality controls and their implications in ageing and neurodegenerative diseases, notably in Parkinson's disease, are discussed.
Post‐transcriptional control of Crp–cAMP by RNase LS in Escherichia coli
Authors: Iwamoto Akira , Lemire Sébastien , Yonesaki Tetsuro
Published: November 2008

Escherichia coli ribonuclease LS was first characterized as a potential antagonist of bacteriophage T4; the E. coli rnlA gene is required for this activity. When rnlA mutant cells were grown on Luria–Bertani agar containing a high concentration of NaCl, their growth was substantially impaired, and introduction of a mutation into crp or cyaA alleviated the NaCl sensitivity. A mutation in rnlA caused fivefold overexpression of Crp. At the same time, the expression of σ38 was lower by two‐ to threefold in an rnlA mutant than in the wild type, which probably accounts for the susceptibility to high NaCl concentration. The overproduction of Crp was eliminated by deletion of the Crp‐site II, to which Crp binds to enhance its own transcription in the presence of abnormally high concentration of cAMP. Consistently, introduction of a mutation into cyaA also eliminated the overproduction of Crp. In fact, all of CyaA, cAMP and cyaA transcripts accumulated to high levels and, after induction, cyaA transcripts were markedly stabilized in an rnlA mutant compared with the wild type. We conclude that RNase LS regulates Crp–cAMP concentration by degrading the cyaA transcripts.
Transcription regulation of the Saccharomyces cerevisiae PIS1 gene by inositol and the pleiotropic regulator, Ume6p
Authors: Jani Niketa M. , Lopes John M.
Published: November 2008

In Saccharomyces cerevisiae, transcription of most of the phospholipid biosynthetic genes (e.g. INO1, CHO1, CHO2 and OPI3) is repressed by growth in the presence of inositol and choline and derepressed in their absence. This regulation requires the Ino2p and Ino4p activators and the Opi1p repressor. The PIS1 structural gene is required for the synthesis of the essential lipid phosphatidylinositol. Previous reports show that PIS1 expression is uncoupled from inositol/choline regulation, but is regulated by carbon source, hypoxia and zinc. However, in this study we found that the expression of PIS1 is induced twofold by inositol. This regulation did not require Ino2p and Ino4p, although Ino4p was required for full expression. Ino4p is a basic helix‐loop‐helix protein that requires a binding partner. Curiously, none of the other basic helix‐loop‐helix proteins affected PIS1 expression. Inositol induction did require another general regulator of phospholipid biosynthesis, Ume6p. Ume6p was found to be a positive regulator of PIS1 gene expression. Ume6p, and several associated factors, were required for inositol‐mediated induction and chromatin immunoprecipitation analysis showed that Ume6p directly regulates PIS1 expression. Thus, we demonstrate novel regulation of the PIS1 gene by Ume6p.
Structure and non‐essential function of glycerol kinase in Plasmodium falciparum blood stages
Authors: Schnick Claudia , Polley Spencer D. , Fivelman Quinton L. , Ranford‐Cartwright Lisa C. , Wilkinson Shane R. , Brannigan James A. , Wilkinson Anthony J. , Baker David A.
Published: November 2008

Malaria pathology is caused by multiplication of asexual parasites within erythrocytes, whereas mosquito transmission of malaria is mediated by sexual precursor cells (gametocytes). Microarray analysis identified glycerol kinase (GK) as the second most highly upregulated gene in Plasmodium falciparum gametocytes with no expression detectable in asexual blood stage parasites. Phosphorylation of glycerol by GK is the rate‐limiting step in glycerol utilization. Deletion of this gene from P. falciparum had no effect on asexual parasite growth, but surprisingly also had no effect on gametocyte development or exflagellation, suggesting that these life cycle stages do not utilize host‐derived glycerol as a carbon source. Kinetic studies of purified PfGK showed that the enzyme is not regulated by fructose 1,6 bisphosphate. The high‐resolution crystal structure of P. falciparum GK, the first of a eukaryotic GK, reveals two domains embracing a capacious ligand‐binding groove. In the complexes of PfGK with glycerol and ADP, we observed closed and open forms of the active site respectively. The 27° domain opening is larger than in orthologous systems and exposes an extensive surface with potential for exploitation in selective inhibitor design should the enzyme prove to be essential in vivo either in the human or in the mosquito.
Spontaneous mutagenesis is elevated in protease‐defective cells
Authors: Al Mamun Abu Amar M. , Humayun M. Zafri
Published: November 2008

As a first step towards describing the role of proteolysis in maintaining genomic integrity, we have determined the effect of the loss of ClpXP, a major energy‐dependent cytoplasmic protease that degrades truncated proteins as well as a number of regulatory proteins, on spontaneous mutagenesis. In a rifampicin‐sensitive to rifampicin‐resistance assay that detects base substitution mutations in the essential rpoB gene, there is a modest, but appreciable increase in mutagenesis in Δ(clpP‐clpX) cells relative to wild‐type cells. A colony papillation analysis using a set of lacZ strains revealed that genetic −1 frameshift mutations are strongly elevated in Clp‐defective cells. A quantitative analysis using a valine‐sensitive to valine‐resistance assay that detects frameshift mutations showed that mutagenesis is elevated 50‐fold in Clp‐defective cells. Elevated frameshift mutagenesis observed in Clp‐deficient cells is essentially abolished in lexA1[Ind‐] (SOS‐uninducible) cells, and in cells deleted for the SOS gene dinB, which codes for DNA polymerase IV. In contrast, mutagenesis is unaffected or stimulated in cells deleted for umuC or umuD, which code for critical components of DNA polymerase V. Loss of rpoS, which codes for a stress‐response sigma factor known to upregulate dinB expression in stationary phase, does not affect mutagenesis. We propose that elevated DinB expression, as well as stabilization of UmuD/UmuD′ heterodimers in Δ(clpP‐clpX) cells, contributes to elevated mutagenesis. These findings suggest that in normal cells, Clp‐mediated proteolysis plays an important role in preventing gratuitous mutagenesis.
Protein unfolding is an essential requirement for transport across the parasitophorous vacuolar membrane of Plasmodium falciparum
Authors: Gehde Nina , Hinrichs Corinna , Montilla Irine , Charpian Stefan , Lingelbach Klaus , Przyborski Jude M.
Published: November 2008

Plasmodium falciparum traffics a large number of proteins to its host cell, the mature human erythrocyte. How exactly these proteins gain access to the red blood cell is poorly understood. Here we have investigated the effect of protein folding on the transport of model substrate proteins to the host cell. We find that proteins must pass into the erythrocyte cytoplasm in an unfolded state. Our data strongly support the presence of a protein‐conducing channel in the parasitophorous vacoular membrane, and additionally imply an important role for molecular chaperones in keeping parasite proteins in a ‘translocation competent’ state prior to membrane passage.
The histidine kinase inhibitor Sda binds near the site of autophosphorylation and may sterically hinder autophosphorylation and phosphotransfer to Spo0F
Authors: Cunningham Katherine A. , Burkholder William F.
Published: November 2008

Histidine kinases are widely used by bacteria, fungi and plants to sense and respond to changing environmental conditions. Signals in addition to those directly sensed by the kinase are often integrated by proteins that fine‐tune the biological response by modulating the activity of the kinase or its targets. The Bacillus subtilis histidine kinase KinA promotes the initiation of sporulation when nutrients are limiting, but sporulation can be delayed by two inhibitors of KinA, Sda (when DNA replication is perturbed) or KipI (under unknown conditions). We have identified residues in the dimerization/histidine‐phosphotransfer (DHp) domain of KinA that are functionally important for inhibition by Sda and KipI and overlapping surface‐exposed residues that lie close to or comprise the Sda binding site. Sda inhibits the intermolecular transfer of phosphate from the catalytic ATP‐binding (CA) domain of KinA to the autophosphorylation site in the DHp domain when the domains are split into separate polypeptides, either by steric hindrance or by altering the conformation of the DHp domain. Sda also slows the rate of phosphotransfer from KinA∼P to its target, Spo0F, consistent with our finding that a KinA residue important for Sda function overlaps with the predicted Spo0F binding site on KinA.
SpoT governs Legionella pneumophila differentiation in host macrophages
Authors: Dalebroux Zachary D. , Edwards Rachel L. , Swanson Michele S.
Published: November 2008

During its life cycle, Legionella pneumophila alternates between a replicative and a transmissive state. To determine their contributions to L. pneumophila differentiation, the two ppGpp synthetases, RelA and SpoT, were disrupted. Synthesis of ppGpp was required for transmission, as relA spoT mutants were killed during entry to and exit from macrophages. RelA, which senses amino acid starvation induced by serine hydroxamate, is dispensable in macrophages, as relA mutants spread efficiently. SpoT monitors fatty acid biosynthesis (FAB), since following cerulenin treatment, wild‐type and relA strains expressed the flaA transmissive gene, but relA spoT mutants did not. As in Escherichia coli, the SpoT response to FAB perturbation likely required an interaction with acyl‐carrier protein (ACP), as judged by the failure of the spoT‐A413E allele to rescue transmissive trait expression of relA spoT bacteria. Furthermore, SpoT was essential for transmission between macrophages, since secondary infections by relA spoT mutants were restored by induction of spoT, but not relA. To resume replication, ppGpp must be degraded, as mutants lacking spoT hydrolase activity failed to convert from the transmissive to the replicative phase in either bacteriological medium or macrophages. Thus, L. pneumophila requires SpoT to monitor FAB and to alternate between replication and transmission in macrophages.
Alcohol oxidase (AOX1) from Pichia pastoris is a novel inhibitor of prion propagation and a potential ATPase
Authors: Zhang Hong , Loovers Harriët M. , Xu Li‐Qiong , Wang Mingzhu , Rowling Pamela J. E. , Itzhaki Laura S. , Gong Weimin , Zhou Jun‐Mei , Jones Gary W. , Perrett Sarah
Published: November 2008

Previous results suggest that methylotrophic yeasts may contain factors that modulate prion stability. Alcohol oxidase (AOX), a key enzyme in methanol metabolism, is an abundant protein that is specific to methylotrophic yeasts. We examined the effect of Pichia pastoris AOX1 on prion phenotypes in Saccharomyces cerevisiae. The S. cerevisiae prion states [PSI+] and [URE3] arise from aggregation of the proteins Sup35p and Ure2p respectively, and correlate with the ability of Sup35p and Ure2p to form amyloid‐like fibrils in vitro. We found that expression of P. pastoris AOX1 in S. cerevisiae had no effect on propagation of the [PSI+] prion, but inhibited propagation of [URE3]. Addition of AOX1 early in the time‐course of fibril formation inhibits Ure2p fibril formation in vitro. AOX1 has not previously been identified as an ATPase. However, we discovered that in addition to its flavin adenine dinucleotide‐dependent AOX activity, AOX1 possesses ATPase activity. This study identifies AOX1 as a novel prion inhibitory factor and a potential ATPase.
Age‐related changes in the development of oocytes and testicular follicles after emergence in Stavsolus japonicus (Plecoptera)
Authors: YOSHIMURA MAYUMI
Published: November 2008

Abstract Female stoneflies oviposit several times during the adult stage of their life cycle. The number of eggs within the deposited egg masses decreases at successive ovipositions. To clarify the reason for this decrease and to determine the conditions of testicular follicles, the patterns of development of oocytes and testicular follicles on different days after emergence are investigated in the systellognathan species Stavsolus japonicus (Okamoto) (Perlodidae). The size of the mature oocytes in the ovariole peaks a few weeks after emergence but decreases to the lowest level by 35 days after emergence. Several maturing oocytes can be observed in the ovarioles of individuals a few weeks after emergence but only one mature oocyte is observed at 35 days after emergence. The decreased number of eggs laid per mass by older individuals may therefore be due to the lower maturation of all the ovarioles.
Respiratory strategies of tenebrionid beetles in arid Australia: does physiology beget nocturnality?
Authors: DUNCAN FRANCES D. , DICKMAN CHRISTOPHER R.
Published: November 2008

Abstract The respiratory patterns and role of mesothoracic and abdominal spiracles in gas exchange are examined using flow‐through respirometry in three species of Heleini beetles from the Simpson Desert, central Australia. Two species, Helea (Heleus) waitei and Helea sp., show a form of continuous respiration with 70% and 75% of the CO2 being emitted from the mesothoracic spiracles, respectively. Their mass specific metabolic rates are similar and similar to other nocturnally active desert‐dwelling tenebrionid beetles. Brises blairi also shows a continuous form of respiration, with 66% of CO2 being emitted from the mesothoracic spiracle but has a significantly higher mass specific metabolic rate. Unusually for arid‐dwelling tenebrionid beetles, all three of the study species are confirmed to be exclusively nocturnal. There is no evidence that this activity pattern is driven by current ecological factors, such as competition or predation, and it is proposed instead that nocturnality arises from physiological constraint. Heleini probably evolved under mesic conditions and lack some of the key physiological adaptations to reduce water loss that characterize day‐active tenebrionid beetles in arid environments elsewhere. In consequence, they are able to exploit arid conditions only by restricting their activity to the most benign phase of the 24‐h cycle.
Impact of Chemokine ReceptorCCR2andCCR5Gene Polymorphism on Allograft Outcome in North Indian Renal Transplant Recipients
Authors: Singh R. , Kapoor R. , Srivastava A. , Mittal R. D.
Published: November 2008

The CC chemokine receptors, CCR2 and CCR5, contribute to the trafficking of leucocytes into the sites of the immune response in transplanted kidney. Therefore, the inter‐individual differences inCCR2andCCR5gene expression, due to single‐nucleotide polymorphism, have the potential to influence various immune responses within the graft, eventually deciding the allograft outcome. In this study, we genotyped 296 North Indian renal transplant recipients and 277 healthy controls forCCR2V64IandCCR5‐Δ32polymorphisms by sequence‐specific primers and restriction fragment length polymorphism and examined their association with allograft outcome. The frequency ofCCR2+/64I (heterozygous) andCCR2‐64I/64I (homozygous mutant) genotype were comparatively higher in non‐rejecters when compared with transplant recipients experiencing one or more than one rejection episode (20.4% versus 8.2%), thereby resulting in a significantly reduced risk of allograft rejection (OR = 0.331,P = 0.026). The Kaplan–Meier curve also suggested higher mean time for the first rejection episode inCCR2‐64I allele carriers (32.83 ± 1.36 months) when compared withCCR2+/+ recipients (28.09 ± 0.93 months, log P = 0.027). TheCCR5‐Δ32variant had no profound effect on allograft outcome. In conclusion, our study confirmedCCR2‐64I allele to be associated with reduced risk for allograft rejection in North Indian transplant recipients influencing allograft outcome.
Silencing a prohibitin alters plant development and senescence
Authors: Chen Jen‐Chih , Jiang Cai‐Zhong , Reid Michael S.
Published: November 2008

Prohibitins, highly conserved mitochondrial proteins, have been shown to play important roles in cell cycling and senescence in animals and yeast. Sequences with high similarity to prohibitins have been identified in a number of plant species, but their function has not yet been demonstrated. The deduced amino acid sequences of PhPHB1 and PhPHB2, sequences that we identified in a petunia floral expressed sequence tag (EST) database, show high similarity to those of prohibitin‐1 and prohibitin‐2 proteins, respectively, reported from yeast, animals and plants. Southern analysis suggested that these genes were members of small gene families with at least two prohibitin‐1 homologs and four prohibitin‐2 homologs. When we downregulated expression of prohibitin‐1 using a Tobacco rattle virus‐based (TRV), virus‐induced gene silencing system (VIGS), we observed plants with smaller and distorted leaves and flowers. Cells in silenced flowers were larger than in control flowers, indicating a substantial reduction in the number of cell divisions that took place during corolla development. The life of silenced flowers was shorter than that of controls, whether on the plant or detached. The respiration of silenced flowers was higher than that of controls, and we observed a marked increase in the abundance of transcripts of a catalase and a small heat‐shock protein in the silenced flowers. Our data indicate that prohibitins play a key role in plant development and senescence.
National survey of the health and nutrition of schoolchildren in Ethiopia
Authors: Hall Andrew , Kassa Tamiru , Demissie Tsegaye , Degefie Tedbabe , Lee Seung
Published: November 2008

Objective  To estimate the prevalence of chronic health and nutritional conditions of schoolchildren in Ethiopia.

Methods  Cross‐sectional survey in schools in each region randomly selected in proportion to size, then a random sample of 50–68 children in grades 3 and 4 in each school. Children were examined for signs of micronutrient deficiencies and chronic infections; weighed and measured; provided a faecal sample to diagnose intestinal parasitic infections; and were interviewed about their recent diet and hygiene practices.

Results  A total of 7572 children were studied in 142 schools in all 11 regions of Ethiopia. Nearly 17% of children were orphans. The prevalence of stunting was 22.3% and 23.1% of children were thin for age, but these may be underestimates as there was evidence that age may have been estimated based on stature when children enrol in school. Just under 10% of children were anaemic when altitude was accounted for. The overall prevalence of trachoma was 13% and children who washed their face before school were at lower risk of trachoma than children who had not. Children who reported that they had eaten fruits or vegetables the day before also had a lower risk of xerophthalmia than children who had not. Only 30% of children were infected with intestinal worms.

Conclusions  Trachoma is a problem but anaemia and intestinal worms are relatively uncommon in Ethiopian schoolchildren. These data provide a basis for developing a school health policy and programmes.
Antiretroviral treatment and the health workforce in South Africa: how have ART workers been affected by scaling up?
Authors: Tobi Patrick , George Gavin , Schmidt Elena , Renton Adrian
Published: November 2008

Objective  To investigate the effect of scaling up antiretroviral treatment (ART) on the working environment and motivation of health workers in South Africa; and to suggest strategies to minimize negative effects and maximise positive effects.

Methods  Exploratory interviews with health managers and senior clinical staff were used to identify locally relevant work environment indicators. A self ‐reported Likert scale questionnaire was administered to a randomly selected cohort of 269 health professionals at health facilities in KwaZulu Natal and Western Cape provinces of South Africa that included ART delivery sites. The cohort was disaggregated into ART and non‐ART groups and differences between the two compared with Fisher’s exact test and the non‐parametric Mann–Whitney U‐test.

Results  The ART sub‐cohort reported: (i) a lighter workload (P = 0.013), (ii) higher level of staffing (P = 0.010), (iii) lower sickness absence (P = 0.032), (iv) higher overall job satisfaction (P = 0.010), (v) poorer physical state of their work premises (P = 0.003), and (vi) higher staff turnover (P = 0.036).

Conclusion  Scale‐up affects the work environment in ways that influence workers’ motivation both positively and negatively. A net negative balance is likely to drive staff out‐migration, undermine the quality of care and compromise the capacity of the programme to achieve significant scale. As health workers are the most important element of the health system, a comprehensive and systematic understanding of scale‐up impacts on their working conditions and motivation needs to be an integral part of any delivery strategy.
User fee exemptions are not enough: out‐of‐pocket payments for ‘free’ delivery services in rural Tanzania
Authors: Kruk Margaret E. , Mbaruku Godfrey , Rockers Peter C. , Galea Sandro
Published: November 2008

Objective  To identify the main drivers of costs of facility delivery and the financial consequences for households among rural women in Tanzania, a country with a policy of delivery fee exemptions.

Methods  We selected a representative sample of households in a rural district in western Tanzania. Women who given birth within 5 years were asked about payments for doctor’s/nurse’s fees, drugs, non‐medical supplies, medical tests, maternity waiting home, transport and other expenses. Wealth was assessed using a household asset index. We estimated the proportion of women who cut down on spending or borrowed money/sold household items to pay for delivery in each wealth group.

Results  In all, 73.3% of women with facility delivery reported having made out‐of‐pocket payments for delivery‐related costs. The average cost was 6272 Tanzanian shillings (TZS), [95% Confidence Interval (CI): 4916, 7628] or 5.0 United States dollars. Transport costs (53.6%) and provider fees (26.6%) were the largest cost components in government facilities. Deliveries in mission facilities were twice as expensive as those in government facilities. Nearly half (48.3%) of women reported cutting down on spending or borrowing money/selling household assets to pay for delivery, with the poor reporting this most frequently.

Conclusion  Out‐of‐pocket payments for facility delivery were substantial and were driven by high transport costs, unofficial provider payments, and preference for mission facilities, which levy user charges. Novel approaches to financing maternal health services, such as subsidies for transport and care from private providers, are required to reduce the cost barriers to attended delivery.