bluuebunny/crossref_metadata_2025 huggingface dataset:
Optimising microwave vacuum puffing for blue honeysuckle snacks
Authors: Liu Chenghai , Zheng Xianzhe , Shi John , Xue Jun , Lan Yubin , Jia Shuhua
Published: February 2010

Fresh blue honeysuckle fruit slices were puffed in a microwave vacuum dryer up to a final moisture content about 5% (w.b.). The effect of initial moisture content (IMC) (25–45%), vacuum pressure (VP) (70–90 kPa) and microwave intensity (MI) (10–30 W g−1) on quality attributes, in terms of expansion ratio (ER), hardness (HD), crispness (CR) and colour of the products, were analysed by response surface methodology. Besides the effect of MI on chroma (CH), the high IMC and low VP had a significantly positive impact on the quality attributes of blue honeysuckle snacks. The optimum product qualities, which were ER (1.62 times), HD (5836.31 g), CR (4.48), and CH (28.7) were obtained at an IMC of 38.42%, VP of 82.02 kPa, and MI of 22.42 W g−1. The microwave vacuum method has obvious advantages when puffing the blue honeysuckle snacks.
Dough and crumb grain changes during mixing and fermentation and their relation with extension properties and bread quality of yeasted sweet dough
Authors: Tlapale‐Valdivia Alma D. , Chanona‐Pérez Jorge , Mora‐Escobedo Rosalva , Farrera‐Rebollo Reynold R. , Gutiérrez‐López Gustavo F. , Calderón‐Domínguez Georgina
Published: February 2010

Dough and crumb grain evolution of yeasted sweet dough, as affected by the combined effect of mixing (dough development) and fermentation times, and their relation to rheological properties and bread quality was studied. Structural changes of under or optimum developed dough, without fermentation, were related mainly to increments in extensibility (P > 95%), without a noticeable change in pH. At all mixing stages, fermentation duration increased Rm, whereas extensibility and pH decreased. When plotting Rm with bread specific volume, cell area, total cell area fraction or yellow crumb colour parameter (b*) simple mathematical models, with determination coefficients larger than 0.85 were obtained. Bread crumb image analysis showed that crumb cell size was not affected by mixing time, while fermentation duration had a direct influence on this parameter, increasing the proportion of the smallest and the largest crumb cells, without changing cell density, possibly as a result of Ostwald ripening, coalescence and separation‐redistribution mechanisms.
Insight into an island radiation: the Tarentola geckos of the Cape Verde archipelago
Authors: Vasconcelos Raquel , Carranza Salvador , James Harris D.
Published: February 2010

Aim  To reassess the relationships between Tarentola geckos from the Cape Verde Islands by including specimens from all islands in the range. To determine the variation within forms by sequencing over 400 specimens, thereby allowing the discovery of cryptic forms and resolving some of the issues raised previously. This extensive sampling was also used to shed light on distributions and to explain genetic diversity by comparing the ages and ecological and geological features of the islands (size, elevation and habitat diversity).

Location  The Cape Verde Islands: an oceanic archipelago belonging to the Macaronesian biogeographic region, located around 500 km off Senegal.

Methods  A total of 405 new specimens of Tarentola geckos were collected from nine islands with very different geological histories, topography, climate and habitats. Mitochondrial cytochrome b (cyt b) gene and 12S rRNA partial sequences were obtained and analysed using phylogenetic methods and networks to determine molecular diversity, demographic features and phylogeographic patterns.

Results  The phylogenetic relationships between all known forms of Cape Verdean Tarentola specimens were estimated for the first time, the relationships between new forms were assessed and previously hypothesized relationships were re‐examined. Despite the large sample size, low intraspecific diversity was found using a 303‐bp cyt b fragment. Star‐like haplotype networks and statistical tests suggest the past occurrence of a rapid demographic and geographical expansion over most of the islands. Genetic variability is positively correlated with size, elevation and habitat diversity of the islands, but is not linearly related to the age of the islands. Biogeographical patterns have, in general, high concordance with phylogenetic breaks and with the three eco‐geographical island groups. Volcanism and habitat diversity, both tightly linked with island ontogeny, as postulated by the general dynamic model of oceanic island biogeography, as well as present and historical size of the islands appear to be the main factors explaining the genetic diversity of this group.

Main conclusions  The Tarentola radiation was clarified and is clearly associated with the geological and ecological features of the islands. Two factors may account for the low intraspecific variation: (1) recent volcanic activity and high ecological stress, and (2) poor habitat diversity within some islands. More studies are needed to align taxonomy with phylogenetic relationships, whereas GIS modelling may help to predict precise species distributions.
Climatic water deficit, tree species ranges, and climate change in Yosemite National Park
Authors: Lutz James A. , van Wagtendonk Jan W. , Franklin Jerry F.
Published: February 2010

Aim  (1) To calculate annual potential evapotranspiration (PET), actual evapotranspiration (AET) and climatic water deficit (Deficit) with high spatial resolution; (2) to describe distributions for 17 tree species over a 2300‐m elevation gradient in a 3000‐km2 landscape relative to AET and Deficit; (3) to examine changes in AET and Deficit between past (c. 1700), present (1971–2000) and future (2020–49) climatological means derived from proxies, observations and projections; and (4) to infer how the magnitude of changing Deficit may contribute to changes in forest structure and composition.

Location  Yosemite National Park, California, USA.

Methods  We calculated the water balance within Yosemite National Park using a modified Thornthwaite‐type method and correlated AET and Deficit with tree species distribution. We used input data sets with different spatial resolutions parameterized for variation in latitude, precipitation, temperature, soil water‐holding capacity, slope and aspect. We used climate proxies and climate projections to model AET and Deficit for past and future climate. We compared the modelled future water balance in Yosemite with current species water‐balance ranges in North America.

Results  We calculated species climatic envelopes over broad ranges of environmental gradients – a range of 310 mm for soil water‐holding capacity, 48.3°C for mean monthly temperature (January minima to July maxima), and 918 mm yr−1 for annual precipitation. Tree species means were differentiated by AET and Deficit, and at higher levels of Deficit, species means were increasingly differentiated. Modelled Deficit for all species increased by a mean of 5% between past (c. 1700) and present (1971–2000). Projected increases in Deficit between present and future (2020–49) were 23% across all plots.

Main conclusions  Modelled changes in Deficit between past, present and future climate scenarios suggest that recent past changes in forest structure and composition may accelerate in the future, with species responding individualistically to further declines in water availability. Declining water availability may disproportionately affect Pinus monticola and Tsuga mertensiana. Fine‐scale heterogeneity in soil water‐holding capacity, aspect and slope implies that plant water balance may vary considerably within the grid cells of kilometre‐scale climate models. Sub‐grid‐cell soil and topographical data can partially compensate for the lack of spatial heterogeneity in gridded climate data, potentially improving vegetation‐change projections in mountainous landscapes with heterogeneous topography.
Chronic pain in people with an intellectual disability: under‐recognised and under‐treated?
Authors: McGuire B. E. , Daly P. , Smyth F.
Published: February 2010

Aim  To examine the nature, prevalence and impact of chronic pain in adults with an intellectual disability (ID) based on carer report.

Methods  Postal questionnaires were sent to 250 care‐givers and 157 responses were received (63%).

Results  Chronic pain was reported in 13% of the sample (n = 21), 6.3% had pain in two sites and 2% had pain in three or more sites. Of those with chronic pain, 19 experienced mild chronic pain, while severe pain was reported for two service users. Pain problems were more prevalent in those with a Mild ID than in those with more severe disability, perhaps reflecting the ability of the Mild group to communicate about their pain. Non‐prescription medication was the most common form of treatment and there was a notable absence of involvement of specialist pain services.

Conclusions  Given their increased risk for chronic pain, we concluded that pain in the ID population may be under‐recognised and under‐treated, especially in those with impaired capacity to communicate about their pain.
Effects of an MHC‐DRB genotype and allele number on the load of gut parasites in the bank vole Myodes glareolus
Authors: KLOCH AGNIESZKA , BABIK WIESŁAW , BAJER ANNA , SIŃSKI EDWARD , RADWAN JACEK
Published: February 2010

The major histocompatibility complex (MHC) genes code for the proteins responsible for pathogen recognition. The MHC class II DRB gene is multiplicated in the bank vole, Myodes glareolus, with different numbers of loci found in different individuals. Possessing large numbers of loci should increase the probability of pathogen recognition, but according to the optimality hypothesis, there is a cost of possessing too many MHC alleles. Using 454 technology, we determined the individual DRB allelic diversity and related it to the load of intestinal parasites in voles collected from three sites separated by a distance of 12 to 27 km. The analysis of six microsatellite loci revealed significant population structure (FST = 0.07). The sites differed significantly in the prevalence and abundance of nematode species as well. We found two significant associations between MHC alleles and the intensity of the infection with the most prevalent nematode, Aspiculuris tetraptera. One of these associations was population‐specific. This result suggests that the directions of selection can differ between populations connected by a low level of gene flow, which may contribute to the maintenance of high DRB allele diversity. In accordance with the optimality hypothesis, individuals with an intermediate number of alleles carried the lowest number of nematode species and had the lowest prevalence of A. tetraptera. However, the intensity of infection with A. tetraptera was linearly and negatively associated with the number of alleles.
Mining transcriptome sequences towards identifying adaptive single nucleotide polymorphisms in lake whitefish species pairs (Coregonus spp. Salmonidae)
Authors: RENAUT SÉBASTIEN , NOLTE ARNE W. , BERNATCHEZ LOUIS
Published: February 2010

Next‐generation sequencing allows the discovery of large numbers of single nucleotide polymorphisms (SNPs) in species where little genomic information was previously available. Here, we assembled, de novo, over 130 mb of non‐normalized cDNA using 454 pyrosequencing data from dwarf and normal lake whitefish and backcross hybrids. Our main goals were to gather a large data set of SNP markers, document their distribution within coding regions, evaluate the effect of species divergence on allele frequencies and combine results with previous genomic studies to identify candidate genes underlying the adaptive divergence of lake whitefish. We identified 6094 putative SNPs in 2674 contigs (mean size: 576 bp, range: 101–6116) and 1540 synonymous and 1734 non‐synonymous mutations for a genome‐wide non‐synonymous to synonymous substitution rate ratio (pN/pS) of 0.37. As expected based on the young age (<15 000 years) of whitefish species pair, the overall level of divergence between them was relatively weak. Yet, 89 SNPs showed pronounced allele frequency differences between sympatric normal and dwarf whitefish. Among these, SNPs in genes annotated to energy metabolic functions were the most abundant and this, in addition to previous experimental data at the gene expression and phenotypic level, brings compelling evidence that genes involved in energy metabolism are prime candidates explaining the adaptive divergence of lake whitefish species pairs. Finally, we unexpectedly identified 44 contigs annotated to transposable elements and these were predominantly composed of backcross hybrids sequences. This indicates an elevated activity of transposable elements, which could potentially contribute to the reduced fitness of hybrids previously documented.
Diversity in a hidden world: potential and limitation of next‐generation sequencing for surveys of molecular diversity of eukaryotic microorganisms
Authors: MEDINGER RALPH , NOLTE VIOLA , PANDEY RAM VINAY , JOST STEFFEN , OTTENWÄLDER BIRGIT , SCHLÖTTERER CHRISTIAN , BOENIGK JENS
Published: February 2010

With the delivery of millions of sequence reads in a single experiment, next‐generation sequencing (NGS) is currently revolutionizing surveys of microorganism diversity. In particular, when applied to Eukaryotes, we are still lacking a rigorous comparison of morphological and NGS‐based diversity estimates. In this report, we studied the diversity and the seasonal community turnover of alveolates (Ciliophora and Dinophyceae) in an oligotrophic freshwater lake by SSU amplicon sequencing with NGS as well as by classical morphological analysis. We complemented the morphological analysis by single‐cell PCR followed by Sanger sequencing to provide an unambiguous link to the NGS data. We show that NGS and morphological analyses generally capture frequency shifts of abundant taxa over our seasonal samples. The observed incongruencies are probably largely due to rDNA copy number variation among taxa and heterogeneity in the efficiency of cell lysis. Overall, NGS‐based amplicon sequencing was superior in detecting rare species. We propose that in the absence of other nuclear markers less susceptible to copy number variation, rDNA‐based diversity studies need to be adjusted for confounding effects of copy number variation.
Multiple marker parallel tag environmental DNA sequencing reveals a highly complex eukaryotic community in marine anoxic water
Authors: STOECK THORSTEN , BASS DAVID , NEBEL MARKUS , CHRISTEN RICHARD , JONES MEREDITH D. M. , BREINER HANS‐WERNER , RICHARDS THOMAS A.
Published: February 2010

Sequencing of ribosomal DNA clone libraries amplified from environmental DNA has revolutionized our understanding of microbial eukaryote diversity and ecology. The results of these analyses have shown that protist groups are far more genetically heterogeneous than their morphological diversity suggests. However, the clone library approach is labour‐intensive, relatively expensive, and methodologically biased. Therefore, even the most intensive rDNA library analyses have recovered only small samples of much larger assemblages, indicating that global environments harbour a vast array of unexplored biodiversity. High‐throughput parallel tag 454 sequencing offers an unprecedented scale of sampling for molecular detection of microbial diversity. Here, we report a 454 protocol for sampling and characterizing assemblages of eukaryote microbes. We use this approach to sequence two SSU rDNA diversity markers—the variable V4 and V9 regions—from 10 L of anoxic Norwegian fjord water. We identified 38 116 V4 and 15 156 V9 unique sequences. Both markers detect a wide range of taxonomic groups but in both cases the diversity detected was dominated by dinoflagellates and close relatives. Long‐tailed rank abundance curves suggest that the 454 sequencing approach provides improved access to rare genotypes. Most tags detected represent genotypes not currently in GenBank, although many are similar to database sequences. We suggest that current understanding of the ecological complexity of protist communities, genetic diversity, and global species richness are severely limited by the sequence data hitherto available, and we discuss the biological significance of this high amplicon diversity.
mlRho – a program for estimating the population mutation and recombination rates from shotgun‐sequenced diploid genomes
Authors: HAUBOLD BERNHARD , PFAFFELHUBER PETER , LYNCH MICHAEL
Published: February 2010

Improvements in sequencing technology over the past 5 years are leading to routine application of shotgun sequencing in the fields of ecology and evolution. However, the theory to estimate evolutionary parameters from these data is still being worked out. Here we present an extension and implementation of part of this theory, mlRho. This program can efficiently compute the following three maximum likelihood estimators based on shotgun sequence data obtained from single diploid individuals: the population mutation rate (4Neμ), the sequencing error rate, and the population recombination rate (4Nec). We demonstrate the accuracy of mlRho by applying it to simulated data sets. In addition, we analyse the genomes of the sea squirt Ciona intestinalis and the water flea Daphnia pulex. Ciona intestinalis is an obligate outcrosser, while D. pulex is a cyclic parthenogen, and we discuss how these contrasting life histories are reflected in our parameter estimates. The program mlRho is freely available from http://guanine.evolbio.mpg.de/mlRho.
Assembly free comparative genomics of short‐read sequence data discovers the needles in the haystack
Authors: CANNON CHARLES H. , KUA CHAI‐SHIAN , ZHANG D. , HARTING J.R.
Published: February 2010

Most comparative genomic analyses of short‐read sequence (SRS) data rely upon the prior assembly of a reference sequence. Here, we present an assembly free analysis of SRS data that discovers sequence variants among focal genomes by tabulating the presence and frequency of ‘complex’ fragments in the data. Using data from nine tree species, we compare genomic diversity from populations to families. As a control, we simulated SRS data for three known plant genomes. The results provide insight into the quality and distributional bias of the sequencing reaction. Three main types of informative complexmers were identified, each possessing unique statistical properties. Type I complexmers are unique to a genome but suffer from a high false positive rate, being highly dependent on read coverage and distribution. Type II complexmers are shared between two genomes and can highlight potential copy‐number differences. Type III complexmers are exclusive to a subset of genomes and can be useful for associating genetic differences with phenotypic or geographic variation. At the population level in an endangered timber species, numerous markers were identified that could potentially determine geographic origin of individuals and regulate international trade. We observed that the genomic data for the four fig species were more divergent than for stone oak species, possibly due to their complex pollination syndrome and high rates of gene flow. Our approach greatly enhances the application of SRS technology to the study of non‐model organisms and directly identifies the most informative genetic elements for more detailed study and assembly.
Genome‐wide SNP detection in the great titParus majorusing high throughput sequencing
Authors: BERS NIKKIE E. M. VAN , OERS KEES VAN , KERSTENS HINDRIK H. D. , DIBBITS BERT W. , CROOIJMANS RICHARD P. M. A. , VISSER MARCEL E. , GROENEN MARTIEN A. M.
Published: February 2010

Identifying genes that underlie ecological traits will open exiting possibilities to study gene–environment interactions in shaping phenotypes and in measuring natural selection on genes. Evolutionary ecology has been pursuing these objectives for decades, but they come into reach now that next generation sequencing technologies have dramatically lowered the costs to obtain the genomic sequence information that is currently lacking for most ecologically important species. Here we describe how we generated over 2 billion basepairs of novel sequence information for an ecological model species, the great titParus major.We used over 16 million short sequence reads for thede novoassembly of a reference sequence consisting of 550 000 contigs, covering 2.5% of the genome of the great tit. This reference sequence was used as the scaffold for mapping of the sequence reads, which allowed for the detection of over 20 000 novel single nucleotide polymorphisms. Contigs harbouring 4272 of the single nucleotide polymorphisms could be mapped to a unique location on the recently sequenced zebra finch genome. Of all the great tit contigs, significantly more were mapped to the microchromosomes than to the intermediate and the macrochromosomes of the zebra finch, indicating a higher overall level of sequence conservation on the microchromosomes than on the other types of chromosomes. The large number of great tit contigs that can be aligned to the zebra finch genome shows that this genome provides a valuable framework for large scale genetics, e.g. QTL mapping or whole genome association studies, in passerines.
Comparative genomics based on massive parallel transcriptome sequencing reveals patterns of substitution and selection across 10 bird species
Authors: KÜNSTNER AXEL , WOLF JOCHEN B. W. , BACKSTRÖM NICLAS , WHITNEY OSCEOLA , BALAKRISHNAN CHRISTOPHER N. , DAY LAINY , EDWARDS SCOTT V. , JANES DANIEL E. , SCHLINGER BARNEY A. , WILSON RICHARD K. , JARVIS ERICH D. , WARREN WESLEY C. , ELLEGREN HANS
Published: February 2010

Next‐generation sequencing technology provides an attractive means to obtain large‐scale sequence data necessary for comparative genomic analysis. To analyse the patterns of mutation rate variation and selection intensity across the avian genome, we performed brain transcriptome sequencing using Roche 454 technology of 10 different non‐model avian species. Contigs from de novo assemblies were aligned to the two available avian reference genomes, chicken and zebra finch. In total, we identified 6499 different genes across all 10 species, with ∼1000 genes found in each full run per species. We found evidence for a higher mutation rate of the Z chromosome than of autosomes (male‐biased mutation) and a negative correlation between the neutral substitution rate (dS) and chromosome size. Analyses of the mean dN/dS ratio (ω) of genes across chromosomes supported the Hill–Robertson effect (the effect of selection at linked loci) and point at stochastic problems with ω as an independent measure of selection. Overall, this study demonstrates the usefulness of next‐generation sequencing for obtaining genomic resources for comparative genomic analysis of non‐model organisms.
Rapid evolution and selection inferred from the transcriptomes of sympatric crater lake cichlid fishes
Authors: ELMER K. R. , FAN S. , GUNTER H. M. , JONES J. C. , BOEKHOFF S. , KURAKU S. , MEYER A.
Published: February 2010

Crater lakes provide a natural laboratory to study speciation of cichlid fishes by ecological divergence. Up to now, there has been a dearth of transcriptomic and genomic information that would aid in understanding the molecular basis of the phenotypic differentiation between young species. We used next‐generation sequencing (Roche 454 massively parallel pyrosequencing) to characterize the diversity of expressed sequence tags between ecologically divergent, endemic and sympatric species of cichlid fishes from crater lake Apoyo, Nicaragua: benthicAmphilophus astorquiiand limneticAmphilophus zaliosus. We obtained 24 174A. astorquiiand 21 382A. zaliosushigh‐quality expressed sequence tag contigs, of which 13 106 pairs are orthologous between species. Based on the ratio of nonsynonymous to synonymous substitutions, we identified six sequences exhibiting signals of strong diversifying selection (Ka/Ks > 1). These included genes involved in biosynthesis, metabolic processes and development. This transcriptome sequence variation may be reflective of natural selection acting on the genomes of these young, sympatric sister species. Based onKs ratios and p‐distances between 3′‐untranslated regions (UTRs) calibrated to previously published species divergence times, we estimated a neutral transcriptome‐wide substitutional mutation rate of ∼1.25 × 10−6per site per year. We conclude that next‐generation sequencing technologies allow us to infer natural selection acting to diversify the genomes of young species, such as crater lake cichlids, with much greater scope than previously possible.
Genome‐wide analysis of alternative splicing evolution among Mus subspecies
Authors: HARR BETTINA , TURNER LESLIE M.
Published: February 2010

Alternative splicing, the combination of different exons to produce a variety of transcripts from a single gene, contributes enormously to transcriptome diversity in mammals, and the majority of genes encode alternatively spliced products. Previous research comparing mouse, rat and human has shown that a significant proportion of splice forms are not conserved across species, suggesting that alternative transcripts are an important source of evolutionary novelty. Here, we studied the evolution of alternative splicing in the early stages of species divergence in the house mouse. We sequenced the testis transcriptomes of three Mus musculus subspecies and Mus spretus using Illumina technology. On the basis of a genome‐wide analysis of read coverage differences among subspecies, we identified several hundred candidate alternatively spliced regions. We conservatively estimate that 6.5% of testis‐expressed genes show alternative splice differences between at least one pair of M. musculus subspecies, a proportion slightly higher than the proportion of genes differentially expressed among subspecies. These results suggest that differences in both the structure and abundance of transcripts contribute to early transcriptome divergence.
On the use of large marker panels to estimate inbreeding and relatedness: empirical and simulation studies of a pedigreed zebra finch population typed at 771 SNPs
Authors: SANTURE ANNA W. , STAPLEY JESSICA , BALL ALEXANDER D. , BIRKHEAD TIM R. , BURKE TERRY , SLATE JON
Published: February 2010

In recent years there has been a dramatic increase in the availability of high density genetic marker data for both model and non‐model organisms. A potential application of these data is to infer relatedness in the absence of a complete pedigree. Using a marker panel of 771 SNPs genotyped in three generations of an extensive zebra finch pedigree, correlations between pedigree relatedness and seven marker‐based estimates of relatedness were examined, as was the relationship between heterozygosity and inbreeding. Although marker‐based and pedigree relatedness were highly correlated, the variance in estimated relatedness was high. Further, the correlation between heterozygosity and inbreeding was weak, even though mean inbreeding coefficient is typical of that seen in wild vertebrate pedigrees; the weak relationship was in part due to the small variance in inbreeding in the pedigree. Our data suggest that using marker information to reconstruct the pedigree, and then calculating relatedness from the pedigree, is likely to give more accurate relatedness estimates than using marker‐based estimators directly.
The earliest stages of adaptation in an experimental plant population: strong selection on QTLS for seed dormancy
Authors: HUANG XUEQING , SCHMITT JOHANNA , DORN LISA , GRIFFITH CONVERSE , EFFGEN SIGI , TAKAO SHAUN , KOORNNEEF MAARTEN , DONOHUE KATHLEEN
Published: February 2010

Colonizing species may often encounter strong selection during the initial stages of adaptation to novel environments. Such selection is particularly likely to act on traits expressed early in development since early survival is necessary for the expression of adaptive phenotypes later in life. Genetic studies of fitness under field conditions, however, seldom include the earliest developmental stages. Using a new set of recombinant inbred lines, we present a study of the genetic basis of fitness variation in Arabidopsis thaliana in which genotypes, environments, and geographic location were manipulated to study total lifetime fitness, beginning with the seed stage. Large‐effect quantitative trait loci (QTLs) for fitness changed allele frequency and closely approached 90% in some treatments within a single generation. These QTLs colocated with QTLs for germination phenology when seeds were dispersed following a schedule of a typical winter annual, and they were detected in two geographic locations at different latitudes. Epistatically interacting loci affected both fitness and germination in many cases. QTLs for field germination phenology colocated with known QTLs for primary dormancy induction as assessed in laboratory tests, including the candidate genes DOG1 and DOG6. Therefore fitness, germination phenology, and primary dormancy are genetically associated at the level of specific chromosomal regions and candidate loci. Genes associated with the ability to arrest development at early life stages and assess environmental conditions are thereby likely targets of intense natural selection early in the colonization process.
Distinct subcellular localization in the cytosol and apicoplast, unexpected dimerization and inhibition of Plasmodium falciparum glyoxalases
Authors: Urscher Miriam , Przyborski Jude M. , Imoto Masaya , Deponte Marcel
Published: February 2010

The ubiquitous glyoxalase system removes methylglyoxal as a harmful by‐product of glycolysis. Because malaria parasites have drastically increased glycolytic fluxes, they could be highly susceptible to the inhibition of this detoxification pathway. Here we analysed the intracellular localization, oligomerization and inhibition of the glyoxalases from Plasmodium falciparum. Glyoxalase I (GloI) and one of the two glyoxalases II (cGloII) were located in the cytosol of the blood stages. The second glyoxalase II (tGloII) was detected in the apicoplast pointing to alternative metabolic pathways. Using a variety of methods, cGloII was found to exist in a monomer–dimer equilibrium that might have been overlooked for homologues from other organisms and that could be of physiological importance. The compounds methyl‐gerfelin and curcumin, which were previously shown to inhibit mammalian GloI, also inhibited P. falciparum GloI. Inhibition patterns were predominantly competitive but were complicated because of the two different active sites of the enzyme. This effect was neglected in previous inhibition studies of monomeric glyoxalases I, with consequences for the interpretation of inhibition constants. In summary, the present work reveals novel general glyoxalase properties that future research can build on and provides a significant advance in characterizing the glyoxalase system from P. falciparum.
Estimating Z‐ring radius and contraction in dividing Escherichia coli
Authors: Strömqvist Johan , Skoog Karl , Daley Daniel O. , Widengren Jerker , Von Heijne Gunnar
Published: February 2010

We present a fluorescence recovery after photobleaching‐based method for monitoring the progression of septal Z‐ring contraction in dividing Escherichia coli cells. In a large number of cells undergoing division, we irreversibly bleached cytosolically expressed Enhanced Green Fluorescent Protein on one side of the septal invagination and followed the fluorescence relaxation on both sides of the septum. Since the relaxation time depends on the cross‐sectional area of the septum, it can be used to determine the septal radius r. Assuming that the fraction of the observed cells with r‐values in a given interval reflects the duration of that interval in the division process we could derive an approximate time‐course for the contraction event, as a population average. By applying the method repeatedly on individual cells, the contraction process was also followed in real time. On a population average level, our data are best described by a linear contraction process in time. However, on the single cell level the contraction processes display a complex behaviour, with varying levels of activity. The proposed approach provides a simple yet versatile method for studying Z‐ring contraction in vivo, and will help to elucidate its underlying mechanisms.
Caulobacter PopZ forms a polar subdomain dictating sequential changes in pole composition and function
Authors: Bowman Grant R. , Comolli Luis R. , Gaietta Guido M. , Fero Michael , Hong Sun‐Hae , Jones Ying , Lee Julie H. , Downing Kenneth H. , Ellisman Mark H. , McAdams Harley H. , Shapiro Lucy
Published: February 2010

The bacterium Caulobacter crescentus has morphologically and functionally distinct cell poles that undergo sequential changes during the cell cycle. We show that the PopZ oligomeric network forms polar ribosome exclusion zones that change function during cell cycle progression. The parS/ParB chromosomal centromere is tethered to PopZ at one pole prior to the initiation of DNA replication. During polar maturation, the PopZ‐centromere tether is broken, and the PopZ zone at that pole then switches function to act as a recruitment factor for the ordered addition of multiple proteins that promote the transformation of the flagellated pole into a stalked pole. Stalked pole assembly, in turn, triggers the initiation of chromosome replication, which signals the formation of a new PopZ zone at the opposite cell pole, where it functions to anchor the newly duplicated centromere that has traversed the long axis of the cell. We propose that pole‐specific control of PopZ function co‐ordinates polar development and cell cycle progression by enabling independent assembly and tethering activities at the two cell poles.
AglP is a S‐adenosyl‐L‐methionine‐dependent methyltransferase that participates in the N‐glycosylation pathway of Haloferax volcanii
Authors: Magidovich Hilla , Yurist‐Doutsch Sophie , Konrad Zvia , Ventura Valeria V. , Dell Anne , Hitchen Paul G. , Eichler Jerry
Published: February 2010

While pathways for N‐glycosylation in Eukarya and Bacteria have been solved, considerably less is known of this post‐translational modification in Archaea. In the halophilic archaeon Haloferax volcanii, proteins encoded by the agl genes are involved in the assembly and attachment of a pentasaccharide to select asparagine residues of the S‐layer glycoprotein. AglP, originally identified based on the proximity of its encoding gene to other agl genes whose products were shown to participate in N‐glycosylation, was proposed, based on sequence homology, to serve as a methyltransferase. In the present report, gene deletion and mass spectrometry were employed to reveal that AglP is responsible for adding a 14 Da moiety to a hexuronic acid found at position four of the pentasaccharide decorating the Hfx. volcanii S‐layer glycoprotein. Subsequent purification of a tagged version of AglP and development of an in vitro assay to test the function of the protein confirmed that AglP is a S‐adenosyl‐L‐methionine‐dependent methyltransferase.
Spatiotemporal patterns of oxidative burst and micronecrosis in resistance of wheat to brown rust infection
Authors: Orczyk W. , Dmochowska‐Boguta M. , Czembor H. J. , Nadolska‐Orczyk A.
Published: February 2010

The accumulation of H2O2 (oxidative burst) and the progress of pathogen development were studied in compatible and incompatible wheat‐brown rust interactions. The accumulation of H2O2 was detected in 98·7% of guard cells with appressoria 8 h post inoculation (hpi). The reaction in both susceptible and resistant plants declined 2–3 days post inoculation (dpi). The second phase of the oxidative burst was observed in the mesophyll and/or epidermis. In susceptible plants it began 4–5 dpi and was detected only in the epidermis. In resistant plants the response was observed in the mesophyll. In moderately resistant plants it was induced 1–3 dpi, and the percentage of infection units reached 80–90% 8 dpi. This corresponded with severe necrotic symptoms. In highly resistant plants, the oxidative burst was short and transient. The percentage of infection units with H2O2 accumulation reached its highest level (60–70%) 2 dpi, and decreased thereafter. Four days later, the low percentage and weak DAB staining indicated very low H2O2 accumulation. The localization and the time‐course changes of the oxidative burst correlated with the profiles of the micronecrotic response, haustorium mother cell formation and pathogen development termination. An early and localized induction of oxidative burst followed by its rapid quenching correlated with high resistance and almost no disease symptoms. The possible correlation of the oxidative burst and pathogen development patterns with the level and durability of resistance conferred by Lr genes are discussed.
Development of Melaleuca oils as effective natural‐based personal insect repellents
Authors: Greive Kerryn A , Staton John A , Miller Peter F , Peters Bryce A , Oppenheim V M Jane
Published: February 2010

For many decades effective insect repellents have relied on synthetic actives such as N,N‐diethyl‐meta‐toluamide. Increasingly, consumers are seeking natural‐based alternatives to many everyday products including insect repellents. While many studies have been published detailing the potential of essential oils to act as insect repellents, few oils have been identified as viable alternatives to synthetic actives. This study details the process involved in the selection of Australian essential oils effective as repellents and the subsequent testing of natural‐based insect repellents using the selected oils. Using a combination of laboratory‐based and field‐based testing, oil from Melaleuca ericifolia was identified as being an effective insect repellent. When formulated into three different bases: an alcohol‐based spray, an emulsion and a gel, these Melaleuca‐based repellents were shown to be as effective at repelling mosquitoes Aedes vigilax (Skuse) (Diptera: Culicidae) and Verrallina carmenti (Edwards) (Diptera: Culicidae), the bush fly Musca vetustissima (Walker) (Diptera: Muscidae), and biting midges Culicoides ornatus (Taylor) (Diptera: Ceratopogonidae) and Culicoides immaculatus (Lee & Reye) (Diptera: Ceratopogonidae) as a synthetic‐based commercial repellent. This study has shown that effective insect repellents based on natural active ingredients can deliver repellency on par with synthetic actives in the field. Three Melaleuca‐based formulations have been registered as repellents and are now commercially available.
Life history traits and foraging behaviour of Cotesia nonagriae (Olliff) (Hymenoptera: Braconidae), a newly recognised member of the Cotesia flavipes complex of stemborer parasitoids
Authors: Muirhead Kate A , Sallam Nader , Austin Andrew D
Published: February 2010

Cotesia nonagriae (Olliff) from Australia, a parasitoid of the incidental native pest of sugarcane, Bathytricha truncata (Walker) (Lepidoptera: Noctuidae), was previously thought to be a synonym of Cotesia flavipes Cameron. However, recent studies using DNA sequences, morphology and preliminary biological information show that this parasitoid is clearly a different species than C. flavipes and other members of the species complex. Here we further examine differences in the biology of these species by undertaking a detailed study of the life history traits of C. nonagriae, including adult longevity and the potential and realised fecundity of females. In addition, the influence of learning on microhabitat location and foraging behaviour were investigated. Duration of the larval stages and adult longevity of C. nonagriae were longer than previously recorded for other members of the species complex. The potential fecundity of females was similar to C. flavipes (∼200 eggs); however, C. nonagriae oviposited a average of over 100 eggs into each host, almost three times more than for other species in the C. flavipes complex (30–40). The propensity of C. nonagriae to allocate a large number of eggs to each host may be an evolutionary strategy due to the high mortality rate (50–57%) of ovipositing adult wasps. During microhabitat location, both naïve and experienced females demonstrated a strong response towards the plant host complex, with experienced wasps benefiting by having a more rapid response time to host‐induced volatiles and cues.
Predicting the probability of significant prostate cancer in Japanese men with serum prostate‐specific antigen less than 10 ng/mL: Development of a novel pre‐biopsy nomogram
Authors: Ohigashi Takashi , Kanao Kent , Mizuno Ryuichi , Kikuchi Eiji , Nakashima June , Oya Mototsugu
Published: February 2010

Objectives:  To develop and assess a new nomogram incorporating pre‐biopsy clinical data to predict significant prostate cancer in Japanese men with a serum prostate‐specific antigen (PSA) level of less than 10 ng/mL.

Methods:  We collected pre‐biopsy data from 620 men with a serum total PSA of less than 10 ng/mL. They included 491 men with a negative biopsy and 129 men who were confirmed to have histological prostate cancer and subsequently underwent radical prostatectomy. Clinically significant tumors were defined as those with a tumor volume larger than 0.5 mL and/or a Gleason score of 7 or more.

Results:  One hundred and seven prostatectomy patients had clinically significant cancers. Stepwise multivariate logistic regression analysis showed that digital rectal examination findings, PSA adjusted for transition zone volume and free‐to‐total PSA ratio were the significant independent predictors of significant cancers (P < 0.0001). Using these pre‐biopsy independent factors, a nomogram was developed to predict significant cancers. According to a receiver operating characteristics analysis, the nomogram showed an area under the curve of 0.831.

Conclusion:  This represents the first nomogram to predict the probability of clinically significant cancers before biopsy. This tool is most likely to be useful in the management of patients with moderate to elevated PSA.
Metatranscriptomic array analysis of ‘Candidatus Accumulibacter phosphatis’‐enriched enhanced biological phosphorus removal sludge
Authors: He Shaomei , Kunin Victor , Haynes Matthew , Martin Hector Garcia , Ivanova Natalia , Rohwer Forest , Hugenholtz Philip , McMahon Katherine D.
Published: February 2010

Here we report the first metatranscriptomic analysis of gene expression and regulation of ‘Candidatus Accumulibacter’‐enriched lab‐scale sludge during enhanced biological phosphorus removal (EBPR). Medium density oligonucleotide microarrays were generated with probes targeting most predicted genes hypothesized to be important for the EBPR phenotype. RNA samples were collected at the early stage of anaerobic and aerobic phases (15 min after acetate addition and switching to aeration respectively). We detected the expression of a number of genes involved in the carbon and phosphate metabolisms, as proposed by EBPR models (e.g. polyhydroxyalkanoate synthesis, a split TCA cycle through methylmalonyl‐CoA pathway, and polyphosphate formation), as well as novel genes discovered through metagenomic analysis. The comparison between the early stage anaerobic and aerobic gene expression profiles showed that expression levels of most genes were not significantly different between the two stages. The majority of upregulated genes in the aerobic sample are predicted to encode functions such as transcription, translation and protein translocation, reflecting the rapid growth phase of Accumulibacter shortly after being switched to aerobic conditions. Components of the TCA cycle and machinery involved in ATP synthesis were also upregulated during the early aerobic phase. These findings support the predictions of EBPR metabolic models that the oxidation of intracellularly stored carbon polymers through the TCA cycle provides ATP for cell growth when oxygen becomes available. Nitrous oxide reductase was among the very few Accumulibacter genes upregulated in the anaerobic sample, suggesting that its expression is likely induced by the deprivation of oxygen.
The autoinducer synthase LqsA and putative sensor kinase LqsS regulate phagocyte interactions, extracellular filaments and a genomic island of Legionella pneumophila
Authors: Tiaden André , Spirig Thomas , Sahr Tobias , Wälti Martin A. , Boucke Karin , Buchrieser Carmen , Hilbi Hubert
Published: February 2010

The amoebae‐resistant opportunistic pathogen Legionella pneumophila employs a biphasic life cycle to replicate in host cells and spread to new niches. Upon entering the stationary growth phase, the bacteria switch to a transmissive (virulent) state, which involves a complex regulatory network including the lqs gene cluster (lqsA‐lqsR‐hdeD‐lqsS). LqsR is a putative response regulator that promotes host–pathogen interactions and represses replication. The autoinducer synthase LqsA catalyses the production of the diffusible signalling molecule 3‐hydroxypentadecan‐4‐one (LAI‐1) that is presumably recognized by the sensor kinase LqsS. Here, we analysed L. pneumophila strains lacking lqsA or lqsS. Compared with wild‐type L. pneumophila, the ΔlqsS strain was more salt‐resistant and impaired for the Icm/Dot type IV secretion system‐dependent uptake by phagocytes. Legionella pneumophila strains lacking lqsS, lqsR or the alternative sigma factor rpoS sedimented more slowly and produced extracellular filaments. Deletion of lqsA moderately reduced the uptake of L. pneumophila by phagocytes, and the defect was complemented by expressing lqsA in trans. Unexpectedly, the overexpression of lqsA also restored the virulence defect and reduced filament production of L. pneumophila mutant strains lacking lqsS or lqsR, but not the phenotypes of strains lacking rpoS or icmT. These results suggest that LqsA products also signal through sensors not encoded by the lqs gene cluster. A transcriptome analysis of the ΔlqsA and ΔlqsS mutant strains revealed that under the conditions tested, lqsA regulated only few genes, whereas lqsS upregulated the expression of 93 genes at least twofold. These include 52 genes clustered in a 133 kb high plasticity genomic island, which is flanked by putative DNA‐mobilizing genes and encodes multiple metal ion efflux pumps. Upon overexpression of lqsA, a cluster of 19 genes in the genomic island was also upregulated, suggesting that LqsA and LqsS participate in the same regulatory circuit.