bluuebunny/crossref_metadata_2025 huggingface dataset:
Micro, macro mineral and proximate composition of Atlantic bonito and horse mackerel: a monthly differentiation
Authors: Özden Özkan
Published: February 2010

Atlantic Bonito and horse mackerel were investigated for proximate and elemental composition throughout the year. Two fish species were collected from the Istanbul local fish market from December (2006) to November (2007) during the fishing season. Highest protein values were found in the June for Atlantic Bonito, in July for horse mackerel. Highest fat value of Atlantic Bonito and horse mackerel were determined in November. Trace and potential toxic minerals (mercury, cadmium, lead, copper and zinc) exceeded the legislative limits in some months of the year. The highest concentration among the elements was obtained for sodium, potassium, calcium, magnesium, manganese, iron, zinc, iodine and selenium throughout the year.
The effects of chickpea on the functional properties of white and whole wheat bread
Authors: Yamsaengsung Ram , Schoenlechner Regine , Berghofer Emmerich
Published: February 2010

The effects of the addition of chickpea (Cicer arietinum), emulsifier and altered amount of water on the functional properties (bread volume, colour of crust, crumb texture and crumb porosity) of white and whole wheat bread were investigated applying a screening three factorial design. Addition of chickpea increased crumb firmness and slightly decreased bread volume in both bread types. Addition of emulsifier increased bread volume and decreased bread firmness. Altering the amount of water addition had no significant effects on white bread, but decreased crumb firmness and increased bread volume in whole wheat bread. None of the investigated parameter had an effect of whole wheat bread colour, while in white bread chickpea addition increased darkness and yellowness of the bread. Optimum addition for breads of highest quality would be 56.9–58% water, 1.0% emulsifier and 10.0–16.0% chickpea in white bread and 58% water, 1.0% emulsifier and 11.8% chickpea in whole wheat bread.
An alternative strategy for corn drying (Zea mays) resulted in both energy savings and reduction of fumonisins B1 and B2 contamination
Authors: Yılmaz Neşe , Tuncel Necati Barış
Published: February 2010

The objectives of this study were to investigate the effect of harvest date on fumonisins contamination and to offer an alternative drying procedure with low energy costs. Corn kernel samples above 13% moisture content were dried with hot air (HA), the technique more usually utilised by farmers and compared with infrared and infrared–hot air (IR–HA) combination techniques. The effect of harvest date on fumonisin contamination was found to be significant while drying had no influence. Due to the fact that the corn remains in the field longer than necessary for economic reasons, energy expenses of the drying techniques were calculated in terms of specific energy consumption for unit of evaporated water. It was found that the energy expenses to reduce the moisture level from 15 to 13% with HA drying were higher than the expenses to reduce the moisture level from 29 to 13% with IR–HA drying combination.
Acrylamide content in potato chips on the Italian market determined by liquid chromatography tandem mass spectrometry
Authors: Tateo Fernando , Bononi Monica , Gallone Franco
Published: February 2010

An easy‐to‐use and simplified liquid chromatography‐tandem mass spectrometry (LC‐ESI (+)‐MS/MS) method was adopted to determine the amount of acrylamide in thirty‐two samples of potato chips purchased on the South Italian market in 2009. Extract produced from matrices was directly analysed without derivatisation, the use of isotopically labelled acrylamide or the use of clean up cartridges. Quality analytical data were reported. The limit of detection and limit of quantification were 6 μg kg−1 and 18 μg kg−1, respectively and recovery values ranged from 90.7–96.3%. The relative standard deviation (RSD) ranged between 2.1% and 5.8%. The values ranged between 27 and 1400 μg kg−1 and the arithmetic mean acrylamide content resulted 363 μg kg−1. According to Foot et al. [Food Additives and Contaminants, 24 (2007)(S1), 37] and considering 500 μg kg−1 as the minimum level possible with the actual available mitigation tools, the number of samples showing an acrylamide level higher than 500 μg kg−1 resulted to be 22%. This paper gives a contribution to data collection in response to the monitoring activity of EFSA and following the Commission Recommendation 2007/331/EC: really, the number of Italian samples of potato chips and potato products for which acrylamide values was reported in the ‘results on the monitoring of acrylamide’ of EFSA (http://www.efsa.europa.eu/cs/BlobServer/Report/datex_report_acrylamide_en,0.pdf?ssbinary=true, 2009) issued on 30 April 2009 is very limited.
Thermo‐physical properties of composite bread dough with maize and cassava flours
Authors: Oladunmoye Olufunmilola Olaitan , Ojo Ademola , Akinoso Rahman , Akanbi Charles Taiwo
Published: February 2010

Composite wheat–cassava and wheat–maize flours were produced in ratio 100:0. 60:40, 50:50, 40:60 and 0:100 respectively. Thermo‐physical properties of bread dough were determined. For wheat –cassava composite bread dough, moisture content ranged between 44.02 ± 2.04 to 51.31 ± 2.99% dry basis (db), density (1035.2 ± 20.4 to 975.6 ± 12.6 kg m−3), specific heat capacity (2.51 ± 0.61 to 3.01 ± 0.42 kJ kg−1 K) and thermal conductivity (0.362 ± 0.13 to 0.473 ± 0.12 W mK−1). While wheat–maize mixture gave 44.14 ± 1.94 to 45.09 ± 1.26%(db) of moisture content, 981.4 ± 16.3–960.4 ± 22.5 kg m−3 density, 1.77 ± 0.17–2.61 ± 0.63 kJ kg−1 K specific heat capacity and 0.36 ± 0.07–0.39 ± 0.02 W mK−1 thermal conductivity. Effects of substitutions was significant on moisture content and thermal conductivity of dough while non significant influence was recorded on density and specific heat capacity at P < 0.05.
Probiotic lactic acid bacteria from Kung‐Som: isolation, screening, inhibition of pathogenic bacteria
Authors: Hwanhlem Noraphat , Watthanasakphuban Nisit , Riebroy Siriporn , Benjakul Soottawat , H‐Kittikun Aran , Maneerat Suppasil
Published: February 2010

Lactic acid bacteria (LAB) were isolated from Kung‐Som at various fermentation periods. Only ten strains, named D2SM22, D6SM3, D6SM24, D6SM26, D8SM21, D10SM5, D10SM11, D10SM16, D10SM20 and D16SM26 showed a survival rate of more than 50% under the simulated gastric juice. After being subjected to simulated gastric juice, four strains (D6SM3, D8SM21, D10SM16 and D10SM20) showed a survival rate of more than 50% in simulated small intestinal juices. Growth of strain D6SM3, D8SM21 and D10SM16 under micro‐aerobic and anaerobic conditions was not different. Tested pathogenic strains (Escherichia coli, Staphylococcus aureus, Bacillus cereus, Vibrio parahaemolyticus and Salmonella sp.) were inhibited by probiotic LAB. However, none of strains could produce bacteriocins. All strains were identified as Lactobacillus plantarum. No differences in pH, acidity, LAB count and liking scores between Kung‐Som produced with starter culture and conventional method were observed (P > 0.01).
Determination of simple sugars, malic acid and total phenolic compounds in apple pomace by infrared spectroscopy and PLSR
Authors: Queji Mary D. , Wosiacki Gilvan , Cordeiro Gilcélia A. , Peralta‐Zamora Patricio G. , Nagata Noemi
Published: February 2010

The aim of this work was to develop a fast, versatile, inexpensive and environmentally safe analytical method to quantify simple sugars, malic acid and total phenolic compounds in apple pomace, considering its potential use as a raw material with value instead of as an industrial waste. Diffuse reflectance infrared spectroscopy (DRIFTS) measurements of twenty‐six samples of apple pomace were analysed by partial least squares regression (PLSR), using several signal pre‐processing methods. Multivariate models developed with four to five latent variables (LVs) and based in the MIR (mid‐infrared) region had good prediction for the determination of sucrose, fructose, malic acid and total phenolic compounds, with average errors between 3.9% and 6.6%. By contrast, glucose was better determined by models developed in the NIR (near‐infrared) region and using six LVs, yielding an average error lower than 7.4%. These results confirmed the feasibility of the multivariate spectroscopic approach as an alternative for expensive and time‐consuming conventional chemical methods.
Differential effects of experimental increases in sociality on ectoparasites of free‐ranging raccoons
Authors: Monello Ryan J. , Gompper Matthew E.
Published: February 2010

1.Parasite transmission depends on the rate at which hosts come into contact with one another or the infectious stages of parasites. However, host contact rates and their influence on parasite transmission are difficult to quantify in natural settings and can fluctuate with host behaviour and the ecological constraints of parasites.

2.We investigated how experimental increases in rates of contact and social aggregation affected ectoparasite prevalence and intensity of free‐ranging raccoons (Procyon lotor). Twelve independent raccoon populations were subjected to differential resource provisions for 2 years: a clumped food distribution to aggregate hosts (n = 5 aggregated populations), a dispersed food distribution to control for the effects of food without aggregating hosts (n = 3) and a no food treatment (n = 4).

3.Remote cameras indicated that aggregation sizes and rates of contact were three to four times greater in aggregated compared with that in non‐aggregated populations. The number of ticks (adultDermacentor variabilis) on raccoons in aggregated populations was 1·5–2·5 times greater from May to July, the primary time of tick seasonal occurrence. Conversely, louse (Trichodectes octomaculatus) populations werec.40% sparser on male raccoons in aggregated (compared with that in non‐aggregated) populations because of greater overdispersion of lice and a larger number of male hosts harbouring fewer parasites. No treatment‐related differences were found among fleas (Orchopeas howardi).

4.These results were not consistent with our current understanding of parasite transmission; greater rates of host sociality led to increases in a parasite that does not rely on host contact for transmission (ticks) and declines in a parasite that depends on host contact for transmission (lice). We concluded thatD. variabilisincreased in aggregated sites because they can detect and seek out hosts and were more likely to drop off after obtaining a blood meal and re‐attach to raccoons in these locations. Several factors may have contributed to sparser louse populations on male hosts, including a dilution effect that lowered per capita infestation levels.

5.These results indicate that ectoparasites can interact in unique ways with their hosts that are not consistent with other types of parasite species or models of their transmission.
Population dynamics of an Arctiid caterpillar–tachinid parasitoid system using state‐space models
Authors: Karban Richard , De Valpine Perry
Published: February 2010

1. Population dynamics of insect host–parasitoid systems are important in many natural and managed ecosystems and have inspired much ecological theory. However, ecologists have a limited knowledge about the relative strengths of species interactions, abiotic effects and density dependence in natural host–parasitoid dynamics. Statistical time‐series analyses would be more informative by incorporating multiple factors, measurement error and noisy dynamics.

2. We use a novel maximum likelihood and model‐selection analysis of a state‐space model for host–parasitoid dynamics to examine 21 years of annual census data for woolly bear caterpillars (Platyprepia virginalis) and their locally host‐specific tachinid parasitoids (Thelaira americana).

3. Caterpillar densities varied by three orders of magnitude and were driven by density dependence and precipitation from the previous March but not detectably by parasitoids, despite variable and sometimes high (>50%) parasitism.

4. Fly fluctuations, as estimated from per cent parasitism, were affected by density dependence and precipitation from the previous July. There was marginal evidence that host abundance drives fly fluctuations as a generic linear effect but no evidence for classical Nicholson–Bailey coupling.

5. The state‐space model analysis includes new methods for likelihood calculation and allows a balanced consideration of effect magnitude and statistical significance in a nonlinear model with multiple alternative explanatory variables.
Plio‐Pleistocene diversification and genetic population structure of an endangered lizard (the Blue Mountains water skink, Eulamprus leuraensis) in south‐eastern Australia
Authors: Dubey Sylvain , Shine Richard
Published: February 2010

Aim  Although climatic fluctuations occurred world‐wide during the Pleistocene, the severity of glacial and drought events – and hence their influence on animal and plant biogeography – differed among regions. Many Holarctic species were forced to warmer‐climate refugia during glacial periods, leaving the genetic signature of recent expansion and gene flow among modern‐day populations. Montane south‐eastern Australia experienced less extreme glaciation, but the effects of drier and colder climatic conditions over this period on biotic distributions, and hence on the present‐day genetic structure of animal and plant populations, are poorly known.

Location  South‐eastern Australia.

Methods  The endangered Blue Mountains water skink (Eulamprus leuraensis) is a viviparous lizard known from fewer than 40 isolated small swamps at 560–1060 m elevation in south‐eastern Australia. We conducted molecular phylogenetic, dating and population genetics analyses using the mitochondrial NADH dehydrogenase 4 (ND4) of 224 individuals of E. leuraensis sampled across the species’ distribution.

Results  Ancient divergences in haplotype groups between lizards from the Blue Mountains and the Newnes Plateau, and strong genetic differences, even between swamps separated by only a few kilometres, suggest that the species has persisted as a series of relatively isolated populations within its current distribution for about a million years. Presumably, habitat patches similar to current‐day swamps persisted throughout glacial–interglacial cycles in this region, allowing the development of high levels of genetic structuring within and among present‐day populations.

Main conclusions  Our results suggest that less extreme glacial conditions occurred in the Southern Hemisphere compared with the Northern Hemisphere, allowing cold‐adapted species (such as E. leuraensis) to persist in montane areas. However, additional studies are needed before we can assemble a comprehensive view of the impact of Pleistocene climatic variation on the phylogeography of Southern Hemisphere taxa.
To kill or be killed: how viruses interact with the cell death machinery
Authors: Kaminskyy V. , Zhivotovsky B.
Published: February 2010

Abstract.  Kaminskyy V, Zhivotovsky B (Karolinska Institutet, Stockholm, Sweden). To kill or be killed: how viruses interact with the cell death machinery (Symposium). J Intern Med 2010; 267: 473–482.

A virus (from the Latin virus meaning toxin or poison) is a small infectious agent that can only replicate inside the cells of another organism. Viruses are found wherever there is life and have probably existed since living cells first evolved. Viruses do not have their own metabolism and require a host cell to make new products. The range of structural and biochemical (i.e., cytopathic) effects that viruses have on the host cell is extensive. Most viral infections eventually result in the death of the host cell. The causes of death include cell lysis, alterations to the cell’s surface membrane and various modes of programmed cell death. Some viruses cause no apparent changes to the infected cell. Cells in which the virus is latent and inactive show few signs of infection and often function normally. This causes persistent infection and the virus is often dormant for many months or years. Some viruses can cause cells to proliferate without causing malignancy, whereas others are established causes of cancer. Human organisms use a genetically controlled cell death programme that prevents the spreading of viral infection and kills the virus. Between 19 and 21 November 2009, with sponsorship from the Journal of Internal Medicine, the Swedish Research Foundation and the Swedish Cancer Society hosted a conference in Stockholm entitled: ‘To kill or to be killed. Viral evasion strategies and interference with cell death machinery’. Four comprehensive reviews from this conference are presented in this issue of the Journal of Internal Medicine. These reviews include descriptions of: the modulation of host innate and adaptive immune defenses by cytomegalovirus; the impact of gamma‐chain family cytokines on T cell homoeostasis in HIV‐1 infection and the therapeutic implications; approaches to killing tumours by depriving them of the mechanisms for detoxification; and viral strategies for the evasion of immunogenic cell death.
Viral strategies for the evasion of immunogenic cell death
Authors: Galluzzi L. , Kepp O. , Morselli E. , Vitale I. , Senovilla L. , Pinti M. , Zitvogel L. , Kroemer G.
Published: February 2010

Abstract.  Galluzzi L, Kepp O, Morselli E, Vitale I, Senovilla L, Pinti M, Zitvogel L, Kroemer G (INSERM, U848; Institut Gustave Roussy; and Université Paris Sud‐XI, Paris, France; Università degli studi di Modena e Reggio Emilia, Modena, Italy; INSERM, U805, Paris, France). Viral strategies for the evasion of immunogenic cell death (Symposium). J Intern Med 2010; 267: 526–542.

Driven by co‐evolutionary forces, viruses have refined a wide arsenal of strategies to interfere with the host defences. On one hand, viruses can block/retard programmed cell death in infected cells, thereby suppressing one of the most ancient mechanisms against viral dissemination. On the other hand, multiple viral factors can efficiently trigger the death of infected cells and uninfected cells from the immune system, which favours viral spreading and prevents/limits an active antiviral response, respectively. Moreover, several viruses are able to inhibit the molecular machinery that drives the translocation of calreticulin to the surface of dying cells. Thereby, viruses block the exposure of an engulfment signal that is required for the efficient uptake of dying cells by dendritic cells and for the induction of the immune response. In this review, we discuss a variety of mechanisms by which viruses interfere with the cell death machinery and, in particular, by which they subvert immunogenic cell death.
Characterization of duplicate gene evolution in the recent natural allopolyploid Tragopogon miscellus by next‐generation sequencing and Sequenom iPLEX MassARRAY genotyping
Authors: BUGGS RICHARD J. A. , CHAMALA SRIKAR , WU WEI , GAO LU , MAY GREGORY D. , SCHNABLE PATRICK S. , SOLTIS DOUGLAS E. , SOLTIS PAMELA S. , BARBAZUK W. BRAD
Published: February 2010

Tragopogon miscellus (Asteraceae) is an evolutionary model for the study of natural allopolyploidy, but until now has been under‐resourced as a genetic model. Using 454 and Illumina expressed sequence tag sequencing of the parental diploid species of T. miscellus, we identified 7782 single nucleotide polymorphisms that differ between the two progenitor genomes present in this allotetraploid. Validation of a sample of 98 of these SNPs in genomic DNA using Sequenom MassARRAY iPlex genotyping confirmed 92 SNP markers at the genomic level that were diagnostic for the two parental genomes. In a transcriptome profile of 2989 SNPs in a single T. miscellus leaf, using Illumina sequencing, 69% of SNPs showed approximately equal expression of both homeologs (duplicate homologous genes derived from different parents), 22% showed apparent differential expression and 8.5% showed apparent silencing of one homeolog in T. miscellus. The majority of cases of homeolog silencing involved the T. dubius SNP homeolog (164/254; 65%) rather than the T. pratensis homeolog (90/254). Sequenom analysis of genomic DNA showed that in a sample of 27 of the homeologs showing apparent silencing, 23 (85%) were because of genomic homeolog loss. These methods could be applied to any organism, allowing efficient and cost‐effective generation of genetic markers.
Ecogenomics: using massively parallel pyrosequencing to understand virus ecology
Authors: ROOSSINCK MARILYN J. , SAHA PRASENJIT , WILEY GRAHAM B. , QUAN JIAXI , WHITE JAMES D. , LAI HONGSHING , CHAVARRÍA FELIPE , SHEN GUOAN , ROE BRUCE A.
Published: February 2010

Environmental samples have been analysed for viruses in metagenomic studies, but these studies have not linked individual viruses to their hosts. We designed a strategy to isolate double‐stranded RNA, a hallmark of RNA virus infection, from individual plants and convert this to cDNA with a unique four nucleotide Tag at each end. Using 96 different Tags allowed us to pool samples and still retain the link to the original sample. We then analysed the sequence of pooled samples using massively parallel sequencing with Roche 454 pyrosequencing such that 384 samples could be assessed per picotiter plate. Using this method we have been able to analyse thousands of plants, and we have discovered several thousand new plant viruses, all linked to their specific plant hosts. Here we describe the method in detail, including the results and analysis for eight pools of samples. This technology will be extremely useful in understanding the full scope of plant virus biodiversity.
Nucleotide divergence vs. gene expression differentiation: comparative transcriptome sequencing in natural isolates from the carrion crow and its hybrid zone with the hooded crow
Authors: WOLF JOCHEN B. W. , BAYER TILL , HAUBOLD BERNHARD , SCHILHABEL MARKUS , ROSENSTIEL PHILIP , TAUTZ DIETHARD
Published: February 2010

Recent advances in sequencing technology promise to provide new strategies for studying population differentiation and speciation phenomena in their earliest phases. We focus here on the black carrion crow (Corvus [corone] corone), which forms a zone of hybridization and overlap with the grey coated hooded crow (Corvus [corone] cornix). However, although these semispecies are taxonomically distinct, previous analyses based on several types of genetic markers did not reveal significant molecular differentiation between them. We here corroborate this result with sequence data obtained from a set of 25 nuclear intronic loci. Thus, the system represents a case of a very early phase of species divergence that requires new molecular approaches for its description. We have therefore generated RNAseq expression profiles using barcoded massively parallel pyrosequencing of brain mRNA from six individuals of the carrion crow and five individuals from a hybrid zone with the hooded crow. We obtained 856 675 reads from two runs, with average read length of 270 nt and coverage of 8.44. Reads were assembled de novo into 19 552 contigs, 70% of which could be assigned to annotated genes in chicken and zebra finch. This resulted in a total of 7637 orthologous genes and a core set of 1301 genes that could be compared across all individuals. We find a clear clustering of expression profiles for the pure carrion crow animals and disperse profiles for the animals from the hybrid zone. These results suggest that gene expression differences may indeed be a sensitive indicator of initial species divergence.
Ultrasequencing of the meiofaunal biosphere: practice, pitfalls and promises
Authors: CREER S. , FONSECA V. G. , PORAZINSKA D. L. , GIBLIN‐DAVIS R. M. , SUNG W. , POWER D. M. , PACKER M. , CARVALHO G. R. , BLAXTER M. L. , LAMBSHEAD P. J. D. , THOMAS W. K.
Published: February 2010

Biodiversity assessment is the key to understanding the relationship between biodiversity and ecosystem functioning, but there is a well‐acknowledged biodiversity identification gap related to eukaryotic meiofaunal organisms. Meiofaunal identification is confounded by the small size of taxa, morphological convergence and intraspecific variation. However, the most important restricting factor in meiofaunal ecological research is the mismatch between diversity and the number of taxonomists that are able to simultaneously identify and catalogue meiofaunal diversity. Accordingly, a molecular operational taxonomic unit (MOTU)‐based approach has been advocated for en mass meiofaunal biodiversity assessment, but it has been restricted by the lack of throughput afforded by chain termination sequencing. Contemporary pyrosequencing offers a solution to this problem in the form of environmental metagenetic analyses, but this represents a novel field of biodiversity assessment. Here, we provide an overview of meiofaunal metagenetic analyses, ranging from sample preservation and DNA extraction to PCR, sequencing and the bioinformatic interrogation of multiple, independent samples using 454 Roche sequencing platforms. We report two examples of environmental metagenetic nuclear small subunit 18S (nSSU) analyses of marine and tropical rainforest habitats and provide critical appraisals of the level of putative recombinant DNA molecules (chimeras) in metagenetic data sets. Following stringent quality control measures, environmental metagenetic analyses achieve MOTU formation across the eukaryote domain of life at a fraction of the time and cost of traditional approaches. The effectiveness of Roche 454 sequencing brings substantial advantages to studies aiming to elucidate the molecular genetic richness of not only meiofaunal, but also all complex eukaryotic communities.
Finding a (pine) needle in a haystack: chloroplast genome sequence divergence in rare and widespread pines
Authors: WHITTALL J. B. , SYRING J. , PARKS M. , BUENROSTRO J. , DICK C. , LISTON A. , CRONN R.
Published: February 2010

Critical to conservation efforts and other investigations at low taxonomic levels, DNA sequence data offer important insights into the distinctiveness, biogeographic partitioning and evolutionary histories of species. The resolving power of DNA sequences is often limited by insufficient variability at the intraspecific level. This is particularly true of studies involving plant organelles, as the conservative mutation rate of chloroplasts and mitochondria makes it difficult to detect polymorphisms necessary to track genealogical relationships among individuals, populations and closely related taxa, through space and time. Massively parallel sequencing (MPS) makes it possible to acquire entire organelle genome sequences to identify cryptic variation that would be difficult to detect otherwise. We are using MPS to evaluate intraspecific chloroplast‐level divergence across biogeographic boundaries in narrowly endemic and widespread species of Pinus. We focus on one of the world’s rarest pines – Torrey pine (Pinus torreyana) – due to its conservation interest and because it provides a marked contrast to more widespread pine species. Detailed analysis of nearly 90% (∼105 000 bp each) of these chloroplast genomes shows that mainland and island populations of Torrey pine differ at five sites in their plastome, with the differences fixed between populations. This is an exceptionally low level of divergence (1 polymorphism/∼21 kb), yet it is comparable to intraspecific divergence present in widespread pine species and species complexes. Population‐level organelle genome sequencing offers new vistas into the timing and magnitude of divergence within species, and is certain to provide greater insight into pollen dispersal, migration patterns and evolutionary dynamics in plants.
Characterization of a hotspot for mimicry: assembly of a butterfly wing transcriptome to genomic sequence at the HmYb/Sb locus
Authors: FERGUSON LAURA , LEE SIU FAI , CHAMBERLAIN NICOLA , NADEAU NICOLA , JORON MATHIEU , BAXTER SIMON , WILKINSON PAUL , PAPANICOLAOU ALEXIE , KUMAR SUJAI , KEE THUAN‐JIN , CLARK RICHARD , DAVIDSON CLAIRE , GLITHERO REBECCA , BEASLEY HELEN , VOGEL HEIKO , FFRENCH‐CONSTANT RICHARD , JIGGINS CHRIS
Published: February 2010

The mimetic wing patterns of Heliconius butterflies are an excellent example of both adaptive radiation and convergent evolution. Alleles at the HmYb and HmSb loci control the presence/absence of hindwing bar and hindwing margin phenotypes respectively between divergent races of Heliconius melpomene, and also between sister species. Here, we used fine‐scale linkage mapping to identify and sequence a BAC tilepath across the HmYb/Sb loci. We also generated transcriptome sequence data for two wing pattern forms of H. melpomene that differed in HmYb/Sb alleles using 454 sequencing technology. Custom scripts were used to process the sequence traces and generate transcriptome assemblies. Genomic sequence for the HmYb/Sb candidate region was annotated both using the MAKER pipeline and manually using transcriptome sequence reads. In total, 28 genes were identified in the HmYb/Sb candidate region, six of which have alternative splice forms. None of these are orthologues of genes previously identified as being expressed in butterfly wing pattern development, implying previously undescribed molecular mechanisms of pattern determination on Heliconius wings. The use of next‐generation sequencing has therefore facilitated DNA annotation of a poorly characterized genome, and generated hypotheses regarding the identity of wing pattern at the HmYb/Sb loci.
Bacterial diversity in a glacier foreland of the high Arctic
Authors: SCHÜTTE URSEL M.E. , ABDO ZAID , FOSTER JAMES , RAVEL JACQUES , BUNGE JOHN , SOLHEIM BJØRN , FORNEY LARRY J.
Published: February 2010

Over the past 100 years, Arctic temperatures have increased at almost twice the global average rate. One consequence is the acceleration of glacier retreat, exposing new habitats that are colonized by microorganisms whose diversity and function are unknown. Here, we characterized bacterial diversity along two approximately parallel chronosequences in an Arctic glacier forefield that span six time points following glacier retreat. We assessed changes in phylotype richness, evenness and turnover rate through the analysis of 16S rRNA gene sequences recovered from 52 samples taken from surface layers along the chronosequences. An average of 4500 sequences was obtained from each sample by 454 pyrosequencing. Using parametric methods, it was estimated that bacterial phylotype richness was high, and that it increased significantly from an average of 4000 (at a threshold of 97% sequence similarity) at locations exposed for 5 years to an average of 7050 phylotypes per 0.5 g of soil at sites that had been exposed for 150 years. Phylotype evenness also increased over time, with an evenness of 0.74 for 150 years since glacier retreat reflecting large proportions of rare phylotypes. The bacterial species turnover rate was especially high between sites exposed for 5 and 19 years. The level of bacterial diversity present in this High Arctic glacier foreland was comparable with that found in temperate and tropical soils, raising the question whether global patterns of bacterial species diversity parallel that of plants and animals, which have been found to form a latitudinal gradient and be lower in polar regions compared with the tropics.
A genetic basis for the phenotypic differentiation between siscowet and lean lake trout (Salvelinus namaycush)
Authors: GOETZ FREDERICK , ROSAUER DANIEL , SITAR SHAWN , GOETZ GILES , SIMCHICK CRYSTAL , ROBERTS STEVEN , JOHNSON RONALD , MURPHY CHERYL , BRONTE CHARLES R. , MACKENZIE SIMON
Published: February 2010

In Lake Superior there are three principal forms of lake trout (Salvelinus namaycush): lean, siscowet and humper. Wild lean and siscowet differ in the shape and relative size of the head, size of the fins, location and size of the eyes, caudal peduncle shape and lipid content of the musculature. To investigate the basis for these phenotypic differences, lean and siscowet lake trout, derived from gametes of wild populations in Lake Superior, were reared communally under identical environmental conditions for 2.5 years. Fish were analysed for growth, morphometry and lipid content, and differences in liver transcriptomics were investigated using Roche 454 GS‐FLX pyrosequencing. The results demonstrate that key phenotypic differences between wild lean and siscowet lake trout such as condition factor, morphometry and lipid levels, persist in these two forms when reared in the laboratory under identical environmental conditions. This strongly suggests that these differences are genetic and not a result of environmental plasticity. Transcriptomic analysis involving the comparison of hepatic gene frequencies (RNA‐seq) and expression (quantitative reverse transcription–polymerase chain reaction (qPCR)) between the two lake trout forms, indicated two primary gene groups that were differentially expressed; those involving lipid synthesis, metabolism and transport (acyl‐CoA desaturase, acyl‐CoA binding protein, peroxisome proliferator‐activated receptor gamma, and apolipoproteins), and those involved with immunity (complement component C3, proteasome, FK506 binding protein 5 and C1q proteins). The results demonstrate that RNA‐seq can be used to identify differentially expressed genes; however, some discrepancies between RNA‐seq analysis and qPCR indicate that methods for deep sequencing may need to be refined and/or different RNA‐seq platforms utilized.
Massively parallel 454‐sequencing of fungal communities in Quercus spp. ectomycorrhizas indicates seasonal dynamics in urban and rural sites
Authors: JUMPPONEN ARI , JONES KENNETH L. , DAVID MATTOX J. , YAEGE CHULEE
Published: February 2010

We analysed two sites within and outside an urban development in a rural background to estimate the fungal richness, diversity and community composition in Quercus spp. ectomycorrhizas using massively parallel 454‐sequencing in combination with DNA‐tagging. Our analyses indicated that shallow sequencing (∼150 sequences) of a large number of samples (192 in total) provided data that allowed identification of seasonal trends within the fungal communities: putative root‐associated antagonists and saprobes that were abundant early in the growing season were replaced by common ectomycorrhizal fungi in the course of the growing season. Ordination analyses identified a number of factors that were correlated with the observed communities including host species as well as soil organic matter, nutrient and heavy metal enrichment. Overall, our application of the high throughput 454 sequencing provided an expedient means for characterization of fungal communities.
Microbial ecological response of the intestinal flora of Peromyscus maniculatus and P. leucopus to heavy metal contamination
Authors: COOLON JOSEPH D. , JONES KENNETH L. , NARAYANAN SANJEEV , WISELY SAMANTHA M.
Published: February 2010

Heavy metal contamination negatively affects natural systems including plants, birds, fish and bacteria by reducing biodiversity at contaminated sites. At the Tri‐State Mining District, efforts have been made to remediate sites to mitigate the detrimental effects that contamination has caused on human health. While the remediation effort has returned the site to within federal safety standards, it is unclear if this effort is sufficient to restore floral and faunal communities. Intrinsic to ecosystem and organism health is the biodiversity and composition of microbial communities. We have taken advantage of recent advances in sequencing technology and surveyed the bacterial community of remediated and reference soils as well as the intestinal microbial community of two ubiquitous rodent species to provide insight on the impacts of residual heavy metal contamination on the ecosystem. Rodents found on the remediated site had reduced body mass, smaller body size and lower body fat than animals on reference sites. Using bar‐coded, massively parallel sequencing, we found that bacterial communities in both the soil and Peromyscus spp. gastrointestinal tracts had no difference in diversity between reference and remediated sites but assemblages differed in response to contamination. These results suggest that niche voids left by microbial taxa that were unable to deal with the remnant levels of heavy metals on remediated sites were replaced by taxa that could persist in this environment. Whether this replacement provided similar ecosystem services as ancestral bacterial communities is unknown.
Key considerations for measuring allelic expression on a genomic scale using high‐throughput sequencing
Authors: FONTANILLAS PIERRE , LANDRY CHRISTIAN R. , WITTKOPP PATRICIA J. , RUSS CARSTEN , GRUBER JONATHAN D. , NUSBAUM CHAD , HARTL DANIEL L.
Published: February 2010

Differences in gene expression are thought to be an important source of phenotypic diversity, so dissecting the genetic components of natural variation in gene expression is important for understanding the evolutionary mechanisms that lead to adaptation. Gene expression is a complex trait that, in diploid organisms, results from transcription of both maternal and paternal alleles. Directly measuring allelic expression rather than total gene expression offers greater insight into regulatory variation. The recent emergence of high‐throughput sequencing offers an unprecedented opportunity to study allelic transcription at a genomic scale for virtually any species. By sequencing transcript pools derived from heterozygous individuals, estimates of allelic expression can be directly obtained. The statistical power of this approach is influenced by the number of transcripts sequenced and the ability to unambiguously assign individual sequence fragments to specific alleles on the basis of transcribed nucleotide polymorphisms. Here, using mathematical modelling and computer simulations, we determine the minimum sequencing depth required to accurately measure relative allelic expression and detect allelic imbalance via high‐throughput sequencing under a variety of conditions. We conclude that, within a species, a minimum of 500–1000 sequencing reads per gene are needed to test for allelic imbalance, and consequently, at least five to 10 millions reads are required for studying a genome expressing 10 000 genes. Finally, using 454 sequencing, we illustrate an application of allelic expression by testing for cis‐regulatory divergence between closely related Drosophila species.
From signal perception to signal transduction: ligand‐induced dimeric switch of DctB sensory domain in solution
Authors: Nan Beiyan , Liu Xin , Zhou Yanfeng , Liu Jiwei , Zhang Le , Wen Jin , Zhang Xiaodong , Su Xiao‐Dong , Wang Yi‐Ping
Published: February 2010

Sinorhizobium meliloti DctB is a typical transmembrane sensory histidine kinase, which senses C4‐dicarboxylic acids (DCA) and regulates the expression of DctA, the DCA transporter. We previously reported the crystal structures of its periplasmic sensory domain (DctBp) in apo and succinate‐bound states, and these structures showed dramatic conformational changes at dimeric level. Here we show a ligand‐induced dimeric switch in solution and a strong correlation between DctBp's dimerization states and the in vivo activities of DctB. Using site‐directed mutagenesis, we identify important determinants for signal perception and transduction. Specifically, we show that the ligand‐binding pocket is essential for DCA‐induced ‘on’ activity of DctB. Mutations at different sections of DctBp's dimerization interface can lock full‐length DctB at either ‘on’ or ‘off’ state, independent of ligand binding. Taken together, these results suggest that DctBp's signal perception and transduction occur through a ‘ligand‐induced dimeric switch’, in which the changes in the dimeric conformations upon ligand binding are responsible for the signal transduction in DctB.
A high‐affinity putrescine‐cadaverine transporter from Trypanosoma cruzi
Authors: Hasne Marie‐Pierre , Coppens Isabelle , Soysa Radika , Ullman Buddy
Published: February 2010

Whereas mammalian cells and most other organisms can synthesize polyamines from basic amino acids, the protozoan parasite Trypanosoma cruzi is incapable of polyamine biosynthesis de novo and therefore obligatorily relies upon putrescine acquisition from the host to meet its nutritional requirements. The cell surface proteins that mediate polyamine transport into T. cruzi, as well as most eukaryotes, however, have by‐in‐large eluded discovery at the molecular level. Here we report the identification and functional characterization of two polyamine transporters, TcPOT1.1 and TcPOT1.2, encoded by alleles from two T. cruzi haplotypes. Overexpression of the TcPOT1.1 and TcPOT1.2 genes in T. cruzi epimastigotes revealed that TcPOT1.1 and TcPOT1.2 were high‐affinity transporters that recognized both putrescine and cadaverine but not spermidine or spermine. Furthermore, the activities and subcellular locations of both TcPOT1.1 and TcPOT1.2 in intact parasites were profoundly influenced by extracellular putrescine availability. These results establish TcPOT1.1 and TcPOT1.2 as key components of the T. cruzi polyamine transport pathway, an indispensable nutritional function for the parasite that may be amenable to therapeutic manipulation.
Discovery of an operon that participates in agmatine metabolism and regulates biofilm formation in Pseudomonas aeruginosa
Authors: Williams Bryan J. , Du Rui‐Hong , Calcutt M. Wade , Abdolrasulnia Rasul , Christman Brian W. , Blackwell Timothy S.
Published: February 2010

Agmatine is the decarboxylation product of arginine and a number of bacteria have devoted enzymatic pathways for its metabolism. Pseudomonas aeruginosa harbours the aguBA operon that metabolizes agmatine to putrescine, which can be subsequently converted into other polyamines or shunted into the TCA cycle for energy production. We discovered an alternate agmatine operon in the P. aeruginosa strain PA14 named agu2ABCA′ that contains two genes for agmatine deiminases (agu2A and agu2A′). This operon was found to be present in 25% of clinical P. aeruginosa isolates. Agu2A′ contains a twin‐arginine translocation signal at its N‐terminus and site‐directed mutagenesis and cell fractionation experiments confirmed this protein is secreted to the periplasm. Analysis of the agu2ABCA′ promoter demonstrates that agmatine induces expression of the operon during the stationary phase of growth and during biofilm growth and agu2ABCA′ provides only weak complementation of aguBA, which is induced during log phase. Biofilm assays of mutants of all three agmatine deiminase genes in PA14 revealed that deletion of agu2ABCA′, specifically its secreted product Agu2A′, reduces biofilm production of PA14 following addition of exogenous agmatine. Together, these findings reveal a novel role for the agu2ABCA′ operon in the biofilm development of P. aeruginosa.
Identification of the gate regions in the primary structure of the secretin pIV
Authors: Spagnuolo Julian , Opalka Natacha , Wen Wesley X. , Gagic Dragana , Chabaud Elodie , Bellini Pierdomenico , Bennett Matthew D. , Norris Gillian E. , Darst Seth A. , Russel Marjorie , Rakonjac Jasna
Published: February 2010

Secretins are a family of large bacterial outer membrane channels that serve as exit ports for folded proteins, filamentous phage and surface structures. Despite the large size of their substrates, secretins do not compromise the barrier function of the outer membrane, implying a gating mechanism. The region in the primary structure that forms the putative gate has not previously been determined for any secretin. To identify residues involved in gating the pIV secretin of filamentous bacteriophage f1, we used random mutagenesis of the gene followed by positive selection for mutants with compromised barrier function (‘leaky’ mutants). We identified mutations in 34 residues, 30 of which were clustered into two regions located in the centre of the conserved C‐terminal secretin family domain: GATE1 (that spanned 39 residues) and GATE2 (that spanned 14 residues). An internal deletion constructed in the GATE2 region resulted in a severely leaky phenotype. Three of the four remaining mutations are located in the region that encodes the N‐terminal, periplasmic portion of pIV and could be involved in triggering gate opening. Two missense mutations in the 24‐residue region that separates GATE1 and GATE2 were also constructed. These mutant proteins were unstable, defective in multimerization and non‐functional.
Update on Legionnaires’ disease: pathogenesis, epidemiology, detection and control
Authors: Hilbi Hubert , Jarraud Sophie , Hartland Elizabeth , Buchrieser Carmen
Published: February 2010

Legionellosis or Legionnaires’ disease is an emerging and often‐fatal form of pneumonia that is most severe in elderly and immunocompromised people, an ever‐increasing risk group for infection. In recent years, the genomics of Legionella spp. has significantly increased our knowledge of the pathogenesis of this disease by providing new insights into the evolution and genetic and physiological basis of Legionella–host interactions. The seventh international conference on Legionella, Legionella 2009, illustrated many recent conceptual advances in epidemiology, pathogenesis and ecology. Experts in different fields presented new findings on basic mechanisms of pathogen–host interactions and bacterial evolution, as well as the clinical management and environmental prevalence and persistence of Legionella. The presentations revealed remarkable facts about the genetic and metabolic basis of the intracellular lifestyle of Legionella and reported on its striking ability to manipulate host cell processes by molecular mimicry. Together, these investigations will lead to new approaches for the treatment and prevention of Legionnaires’ disease.
Effects of resistance of Eremocitrus glauca and Microcitrus australis to viroid infection: replication, accumulation and long‐distance movement of six citrus viroids
Authors: Bani Hashemian S. M. , Barbosa C. J. , Serra P. , Duran‐Vila N.
Published: February 2010

In studies to identify genotypes resistant to infection with citrus viroids, Eremocitrus glauca and Microcitrus australis were selected because their evolution in their habitat in Australia and New Guinea may have led to the selection of unusual traits. The movement and accumulation of Citrus exocortis viroid (CEVd), Hop stunt viroid, Citrus bent leaf viroid, Citrus dwarfing viroid, Citrus bark cracking viroid and Citrus viroid V (CVd‐V) in self‐rooted as well as in graft‐ propagated E. glauca and M. australis plants was assessed by northern hybridization, RT‐PCR and by topworking to the sensitive selection 861‐S1 of Etrog citron. In both plant species the inoculated viroids were undetectable unless these plants were grafted to a susceptible Citrus partner, the rough lemon rootstock and/or the topworked Etrog citron, which acted as viroid sources. The results obtained indicate that M. australis and in particular E. glauca are poor viroid hosts in which viroid replication/accumulation does not occur or is extremely inefficient. However, viroid downward and upward movement to grafted Citrus partners in which viroid replication and accumulation occurs efficiently was not impaired. Eremocitrus glauca and M. australis showed differences regarding their properties as viroid hosts, but for both species CEVd seemed to have the lowest affinity among the viroid species tested and CVd‐V the highest. Even though E. glauca and M. australis do not appear to be truly resistant to viroid infection, they are interesting genotypes for further characterization of the mechanisms involved in viroid infection.