bluuebunny/crossref_metadata_2025 huggingface dataset:
Swimming in the deep end of the gene pool: global population structure of an oceanic giant
Authors: BRADSHAW COREY J. A.
Published: October 2007

Despite the impression held by some that few biological mysteries remain, even evocative species such as humpback whales (Megaptera novaeangliae), white sharks (Carcharodon carcharias) and green turtles (Chelonia mydas) have poorly documented movement patterns, reproductive strategies and population dynamics despite years of dedicated research. This is largely due to the difficulty of observing wide‐ranging marine species over the majority of their life cycle. The advent of powerful tracking devices has certainly improved our understanding, but it is usually only with molecular tools that the nature of population structure becomes apparent. In this issue of Molecular Ecology, Castro and colleagues have provided the first global‐scale assessment of population structure for the largest fish — whale sharks (Rhincodon typus). Whale sharks can reach lengths > 12 m and are a popular tourist attraction at places where they aggregate, yet for most of their life cycle, we know little indeed of where they go and how they interact with other populations. Previous tracking studies imply a high dispersal capacity, but only now have Castro and colleagues demonstrated high gene flow and haplotype diversity among the major ocean basins where they are found.
Genetic structure and evolved malaria resistance in Hawaiian honeycreepers
Authors: FOSTER JEFFREY T. , WOODWORTH BETHANY L. , EGGERT LORI E. , HART PATRICK J. , PALMER DANIELLE , DUFFY DAVID C. , FLEISCHER ROBERT C.
Published: October 2007

Infectious diseases now threaten wildlife populations worldwide but population recovery following local extinction has rarely been observed. In such a case, do resistant individuals recolonize from a central remnant population, or do they spread from small, perhaps overlooked, populations of resistant individuals? Introduced avian malaria (Plasmodium relictum) has devastated low‐elevation populations of native birds in Hawaii, but at least one species (Hawaii amakihi,Hemignathus virens) that was greatly reduced at elevations below about 1000 m tolerates malaria and has initiated a remarkable and rapid recovery. We assessed mitochondrial and nuclear DNA markers from amakihi and two other Hawaiian honeycreepers, apapane (Himatione sanguinea) and iiwi (Vestiaria coccinea), at nine primary study sites from 2001 to 2003 to determine the source of re‐establishing birds. In addition, we obtained sequences from tissue from amakihi museum study skins (1898 and 1948–49) to assess temporal changes in allele distributions. We found that amakihi in lowland areas are, and have historically been, differentiated from birds at high elevations and had unique alleles retained through time; that is, their genetic signature was not a subset of the genetic variation at higher elevations. We suggest that high disease pressure rapidly selected for resistance to malaria at low elevation, leaving small pockets of resistant birds, and this resistance spread outward from the scattered remnant populations. Low‐elevation amakihi are currently isolated from higher elevations (> 1000 m) where disease emergence and transmission rates appear to vary seasonally and annually. In contrast to results from amakihi, no genetic differentiation between elevations was found in apapane and iiwi, indicating that slight variation in genetic or life‐history attributes can determine disease resistance and population recovery. Determining the conditions that allow for the development of resistance to disease is essential to understanding how species evolve resistance across a landscape of varying disease pressures.
Phylogeny, divergence times and species limits of spiny lizards (Sceloporus magisterspecies group) in western North American deserts and Baja California
Authors: LEACHÉ ADAM D. , MULCAHY DANIEL G.
Published: October 2007

The broad distribution of theSceloporus magisterspecies group (squamata: phrynosomatidae) throughout western North America provides an appropriate model for testing biogeographical hypotheses explaining the timing and origins of diversity across mainland deserts and the Baja California Peninsula. We inferred concordant phylogenetic trees describing the higher‐level relationships within themagistergroup using 1.6 kb of mitochondrial DNA (mtDNA) and 1.7 kb of nuclear DNA data. These data provide strong support for the parallel divergence of lineages endemic to the Baja California Peninsula (S. zosteromusand theorcutticomplex) in the form of two sequential divergence events at the base of themagistergroup phylogeny. A relaxed phylogenetic analysis of the mtDNA data using one fossil and one biogeographical constraint provides a chronology of these divergence events and evidence that further diversification within the Baja California clades occurred simultaneously, although patterns of geographical variation and speciation between clades differ. We resolved four major phylogeographical clades withinS. magisterthat (i) provide a novel phylogenetic placement of the Chihuahuan Desert populations sister to the Mojave Desert; (ii) illustrate a mixed history for the Colorado Plateau that includes Mojave and Sonoran Desert components; and (iii) identify an area of overlap between the Mojave and Sonoran Desert clades near Yuma, Arizona. Estimates of bidirectional migration rates among populations ofS. magisterusing four nuclear loci support strong asymmetries in gene flow among the major mtDNA clades. Based on the nonexclusivity of mtDNA haplotypes, nuclear gene flow among populations and wide zones of phenotypic intergradation,S. magisterappears to represent a single geographically variable and widespread species.
Fundamental links between genes and elements: evolutionary implications of ecological stoichiometry
Authors: JEYASINGH PUNIDAN D. , WEIDER LAWRENCE J.
Published: October 2007

Organisms require elements to live and reproduce. We already know that availability of certain elements [e.g. phosphorus (P)] is highly variable spatiotemporally. In addition, there is variability in demand for various elements ontogenetically, as well as phylogenetically. Nonetheless, we know little about the underlying causes for such variation. In this study, we surveyed the literature to identify genes involved in the homeostasis of one biogenic element, P. Evidence from the literature suggests that variation in the environmental supply of P affects expression of highly conserved genes (e.g. the phosphate transporter system). In addition, we found evidence for genetic variation in the acquisition, assimilation, and allocation of P. Such effects of P supply should impinge on fitness, and drive evolutionary change. Further understanding of the inter‐ and intraspecific fitness consequences to imbalances in the availability of P in relation to other biogenic elements may be useful to disentangle primary mechanisms driving diversifications. We argue that studying the interactions between stoichiometric constraints and underlying genes is relevant to the majority of organisms, and conclude by furnishing information on designing and interpreting experiments using this approach.
Phenotype matching and inbreeding avoidance in African elephants
Authors: MOORE JIM
Published: October 2007

Perhaps the most important ‘decision’ made by any animal (or plant) is whether to disperse — leave kith and kin, or remain with the familiar and related. The benefits of staying at home are obvious, so dispersal requires an explanation — and the most popular is that dispersal functions to avoid inbreeding depression. Strong support comes from the observation that dispersal is so often sex biased. Simply put, all else being equal members of both sexes should prefer to remain philopatric, but this would lead to inbreeding depression so members of one sex have to disperse. In principle, this link between inbreeding depression and sex‐biased dispersal could be broken if individuals recognize close kin and avoid mating with them. Archie et al. (2007) provide one of the most compelling analyses to date of the interaction among inbreeding avoidance, kin recognition and mating strategies in any mammal, clearly showing that elephants recognize even close paternal kin and avoid mating with them. Their important results illuminate the subtleties of elephant inbreeding avoidance as well as illustrate the difficulty of arriving at definitive answers to questions about the evolution of dispersal behaviour.
Microsatellite variation points to local landscape plantings as sources of invasive pampas grass (Cortaderia selloana) in California
Authors: OKADA MIKI , AHMAD RIAZ , JASIENIUK MARIE
Published: October 2007

International trade in horticultural plants is a major pathway of introduction of invasive species. Pampas grass (Cortaderia selloana) is an invasive species of horticultural origin that is native to South America but cultivated as an ornamental in regions with Mediterranean climates worldwide. To gain insight into the introduction history of invasive populations in California, we analysed microsatellite marker variation in cultivated and invasive C. selloana. We sampled 275 cultivated plants from diverse sources and 698 invasive plants from 33 populations in four geographical regions of California. A model‐based Bayesian clustering analysis identified seven distinct gene pools in cultivated C. selloana. Probabilities of assignment of invasive individuals to cultivated gene pools indicated that two gene pools accounted for the genomic origin of 78% of the invasive C. selloana sampled. Extensive admixture between cultivated source gene pools was detected within invasive individuals. Sources of admixed invasive individuals are probably landscape plantings. Consistent with the Bayesian assignment results indicating that multiple cultivated gene pools and landscape plantings are probable sources of invasive populations, FST and neighbour‐joining clustering analyses indicated multiple escapes from shared sources in each geographical region. No isolation by distance or geographical trend in reduction of genetic diversity was evident. Furthermore, a generally random and discontinuous distribution of proportional assignments of invasive populations to cultivated gene pools suggests that introductions occurred recurrently within each geographical region. Our results strongly suggest that dispersal through local landscape plantings has contributed to the range expansion of invasive C. selloana in California.
Role of the Trypanosoma brucei natural cysteine peptidase inhibitor ICP in differentiation and virulence
Authors: Santos Camila C. , Coombs Graham H. , Lima Ana Paula C. A. , Mottram Jeremy C.
Published: October 2007

ICP is a chagasin‐family natural tight binding inhibitor of Clan CA, family C1 cysteine peptidases (CPs). We investigated the role of ICP in Trypanosoma brucei by generating bloodstream form ICP‐deficient mutants (Δicp). A threefold increase in CP activity was detected in lysates of Δicp, which was restored to the levels in wild type parasites by re‐expression of the gene in the null mutant. Δicp displayed slower growth in culture and increased resistance to a trypanocidal synthetic CP inhibitor. More efficient exchange of the variant surface glycoprotein (VSG) to procyclin during differentiation from bloodstream to procyclic form was observed in Δicp, a phenotype that was reversed in the presence of synthetic CP inhibitors. Furthermore, we showed that degradation of anti‐VSG IgG is abolished when parasites are pretreated with synthetic CP inhibitors, and that parasites lacking ICP degrade IgG more efficiently than wild type. In addition, Δicp reached higher parasitemia than wild type parasites in infected mice, suggesting that ICP modulates parasite infectivity. Taken together, these data suggest that CPs of T. brucei bloodstream form play a role in surface coat exchange during differentiation, in the degradation of internalized IgG and in parasite infectivity, and that their function is regulated by ICP.
Selective chromosome amplification in Vibrio cholerae
Authors: Srivastava Preeti , Chattoraj Dhruba K.
Published: October 2007

Most bacteria have one chromosome but some have more than one, as is common in eukaryotes. How multiple chromosomes are maintained in bacteria remains largely obscure. Here we have examined the behaviour of the two Vibrio cholerae chromosomes as a function of growth rate. At slow growth rates, both chromosomes were maintained at copy numbers of one to two per cell. Increasing the growth rate by nutritional shift‐up amplified the origin‐proximal DNA of the larger chromosome (chrI) to four copies per cell, but not that of the smaller chrII. The latter was amplified when its specific initiator was supplied in excess or a specific negative regulator was deleted. The growth rate‐insensitive behaviour of chrII, whose origin is similar to origins of members of a major class of plasmids, was shared by some but not all of several representative plasmids tested in V. cholerae. Also, unlike plasmid replication, chrII replication is known to be initiated at a specific stage of the cell cycle. Raising chrII copy number decreased growth rate, suggesting that this chromosome might serve as a repository for necessary but potentially deleterious genes.
P2Y1receptors mediate inhibitory neuromuscular transmission and enteric neuronal activation in small intestine
Authors: Gallego D. , Vanden Berghe P. , Farré R. , Tack J. , Jiménez M.
Published: October 2007

Abstract There is increasing evidence that adenosine 5′‐triphosphate or a related purine plays a crucial role in smooth muscle relaxation and enteric synaptic neurotransmission. Accordingly, the aim of the present work is to investigate the role P2Y1receptors in purinergic inhibitory neurotransmission (pig ileum) and enteric neuronal activation in the small intestine (guinea‐pig ileum). Using contractility measurements, micro‐electrode recordings and Ca2+imaging we found that (i) adenosine 5′‐Ο‐2‐thiodiphosphate (ADPβS) (10 μmol L−1) caused smooth muscle relaxation and hyperpolarization that was antagonized by MRS2179 (10 μmol L−1) a P2Y1receptor antagonist and apamin (1 μmol L−1); (ii) electrical field stimulation (EFS) caused a non‐nitrergic inhibitory junction potential (IJP) and relaxation that was antagonized by MRS2179 (10 μmol L−1); (iii) P2Y1receptors were immunolocalized in smooth muscle cells and enteric neurons; (iv) superfusion of ADPβS (1 μmol L−1) induced Ca2+transients in myenteric neurons that were inhibited by MRS2179 (1 μmol L−1), but not by tetrodotoxin (1 μmol L−1); and (v) EFS induced calcium transients were partially inhibited by MRS2179 (1 μmol L−1). We conclude that in the small intestine purinergic neuromuscular transmission responsible for the IJP and non‐nitrergic relaxation is mediated by P2Y1receptors located in smooth muscle cells. Functional P2Y1receptors are also present in guinea‐pig myenteric neurons. Therefore, P2Y1receptors might be an important pharmacological target to modulate gastrointestinal functions.
Inhibitors of advanced glycation end‐products prevent loss of enteric neuronal nitric oxide synthase in diabetic rats
Authors: Jeyabal P. V. S. , Kumar R. , Gangula P. R. R. , Micci M.‐a. , Pasricha P. J.
Published: October 2007

Abstract  Gastrointestinal dysfunction is common in diabetes, and several studies indicate that loss of neuronal nitrergic inhibition may play an important role in its pathogenesis. However, the mechanisms responsible for this effect remain largely unknown. We have previously shown that advanced glycation end‐products (AGEs) formed by non‐enzymatic glycation dependent processes, can inhibit the expression of intestinal neuronal nitric oxide synthase (nNOS) in vitro acting via their receptor, receptor for AGEs. We now hypothesized that this effect may also be important in experimental diabetes in vivo. We aimed to evaluate the role of AGEs on duodenal nNOS expression and the effects of aminoguanidine (a drug that prevents AGE formation) and ALT‐711 (AGE cross‐link breaker) in experimental diabetes. Streptozotocin induced diabetic rats were randomized to no treatment, treatment with aminoguanidine (1 g L−1 daily through drinking water) at the induction of diabetes, or treatment with ALT‐711 (3 mg kg−1 intraperitoneally), beginning at week 6. A fourth group was used as healthy controls. We performed real time polymerase chain reaction, Western blotting and immunohistochemistry to detect nNOS expression. AGE levels were analysed using sandwich ELISA. Diabetes enhanced accumulation of AGEs in serum, an effect that was prevented by treatment with aminoguanidine and ALT‐711. Further, diabetic rats showed a significant reduction in duodenal nNOS expression by mRNA, protein and immunocytochemistry, an effect that was prevented by aminoguanidine. ALT‐711 had similar effects on nNOS protein and immunohistochemistry (but not on mRNA levels). The generation of AGEs in diabetes results in loss of intestinal nNOS expression and may be responsible for enteric dysfunction in this condition. This study suggests that treatment directed against AGEs may be useful for the treatment of gastrointestinal complications of diabetes.
An exploratory study of the association of adrenergic and serotonergic genotype and gastrointestinal motor functions
Authors: Grudell A. B. M. , Camilleri M. , Carlson P. , Gorman H. , Ryks M. , Burton D. , Baxter K. , Zinsmeister A. R.
Published: October 2007

Abstract  Adrenergic and serotonergic mechanisms alter human gut motor functions. Genotype variation influences phenotype. Our aim was to test the hypothesis that variation in genes that control these functions is associated with gastrointestinal (GI) motor functions in humans with functional GI disorders (FGID). A database of 251 people was assembled by combining genotype data with measurements of gut transit and gastric volumes. Genetic variations evaluated were: α2A adrenergic (C‐1291G), α2C (Del 332‐325), 5‐HT transporter (SLC6A4) and GNβ3 (C825T). We sought associations between motor function or disease groups and genotypes, adjusting for age, gender and body mass index. Among 251 participants, 82 were healthy, 20 with irritable bowel syndrome (IBS) with mixed bowel habit, 49 with constipation‐predominant IBS, 67 with diarrhoea‐predominant IBS and 33 with functional dyspepsia. For all candidate genes, there was no significant association between motor function and wildtype vs non‐wildtype gene status. There were significant interactions between genotype and motility phenotype, specifically GNβ3 and α2A and gastric emptying at 4 h. Borderline associations were noted for SCL6A4 and α2A and postprandial gastric volume, and for α2C and gastric emptying at 2 h. We conclude that genotype variation may affect gastric motor functions in different FGID phenotypes. However, these candidate genes account for only a limited amount of the variance in gastric function of patients with FGID.
Cross‐protection between attenuated Plasmodium berghei and P. yoelii sporozoites
Authors: SEDEGAH M. , WEISS W. W. , HOFFMAN S. L.
Published: October 2007

An attenuated Plasmodium falciparum sporozoite (PfSPZ) vaccine is under development, in part, based on studies in mice with P. berghei. We used P. berghei and P. yoelii to study vaccine‐induced protection against challenge with a species of parasite different from the immunizing parasite in BALB/c mice. One‐hundred percent of mice were protected against homologous challenge. Seventy‐nine percent immunized with attenuated P. berghei sporozoite (PbSPZ) (six experiments) were protected against challenge with P. yoelii sporozoite (PySPZ), and 63% immunized with attenuated PySPZ (three experiments) were protected against challenge with PbSPZ. Antibodies in sera of immunized mice only recognized homologous sporozoites and could not have mediated protection against heterologous challenge. Immunization with attenuated PySPZ or PbSPZ induced CD8+ T cell‐dependent protection against heterologous challenge. Immunization with attenuated PySPZ induced CD8+ T cell‐dependent protection against homologous challenge. However, homologous protection induced by attenuated PbSPZ was not dependent on CD8+ or CD4+ T cells, and depletion of both populations only reduced protection by 36%. Immunization of C57BL/10 mice with PbSPZ induced CD8+ T cell‐dependent protection against P. berghei, but no protection against P. yoelii. The cross‐protection data in BALB/c mice support testing a human vaccine based on attenuated PfSPZ for its efficacy against P. vivax.
The role of TNF‐α in Trichuris muris infection II: global enhancement of ongoing Th1 or Th2 responses
Authors: HAYES K. S. , BANCROFT A. J. , GRENCIS R. K.
Published: October 2007

Host resistance to Trichuris muris is driven by Th2 responses. However, TNF‐α has also been shown to play a role in protection. As TNF‐α has a variety of actions, the exact role of TNF‐α in immunity to T. muris is yet to be established. Here we demonstrate that although blocking TNF‐α has been shown to abrogate resistance, rTNF‐α treatment does not promote resistance. Further, we show that TNF‐α functions to enhance the ongoing immune response. AKR animals that typically respond to infection with a polarized Th1 response produce greater levels of Th1 cytokines when treated with TNF‐α and BALB/c animals that normally respond with a polarized Th2 response produce higher levels of Th2 cytokines. Crucially, blocking TNF‐α in the strong Th2 responder strain BALB/c does not prevent expulsion of T. muris, thus supporting its role as a biological enhancer. TNF‐α does increase transcription of both IFN‐γ and IL‐13 in vitro but can also act synergistically with IL‐13 in vitro to promote production of RELMβ, which has also been shown to play a role in resistance to T. muris. Thus, this data demonstrates that TNF‐α acts to enhance an ongoing immune response but is not necessary for a strong protective Th2 response.
The overexpression of an alternative oxidase gene triggers ozone sensitivity in tobacco plants
Authors: PASQUALINI STEFANIA , PAOLOCCI FRANCESCO , BORGOGNI ANDREA , MORETTINI ROBERTA , EDERLI LUISA
Published: October 2007

The alternative oxidase (AOX) of plant mitochondria transfers electrons from the ubiquinione pool to oxygen without energy conservation and prevents the formation of reactive oxygen species (ROS) when the ubiquinone pool is over‐reduced. Thus, AOX may be involved in plant acclimation to a number of oxidative stresses. To test this hypothesis, we exposed wild‐type (WT) Xanthi tobacco plants as well as Xanthi plants transformed with the Bright Yellow tobacco AOX1a cDNA with enhanced (SN21 and SN29), and decreased (SN10) AOX capacity to an acute ozone (O3) fumigation. As a result of 5 h of O3 exposition (250 nL L−1), SN21 and SN29 plants surprisingly showed localized leaf damage, whereas SN10, similarly to WT plants, was undamaged. In keeping with this observation, WT and SN21 plants differed in their response to O3 for the expression profiles of catalase 1 (CAT1), catalase 2 (CAT2), glutathione peroxidase (GPX) and ascorbate peroxidase (APX) genes, and for the activity of these antioxidant enzymes, which were induced in WT. Concomitantly, although ozone induced H2O2 accumulation in WT and in all transgenic lines, only in transgenics with high AOX capacity the H2O2 level in the post‐fumigation period was high. The alternative pathway of WT plants was strongly stimulated by O3, whereas in SN21 plants, the respiratory capacity was always high across the treatment. The present results show that, far from exerting a protective role, the overexpression of AOX triggers an increased O3 sensitivity in tobacco plants. We hypothesize that the AOX overexpression results in a decrease of mitochondrial ROS level that in turn alters the defensive mitochondrial to nucleus signalling pathway that activates ROS scavenging systems.
Cord Blood Cytokine Profile Detection in Neonates with T1D Parents – Monitoring of Cellular Auto‐reactivity Using Protein Microarray
Authors: Bohmova K. , Hladikova Z. , Cerny M. , Flajsmanova K. , Vrabelova Z. , Skramlikova T. , Spalova I. , Cerna M. , Chudoba D. , Pithova P. , Stadlerova G. , Bartaskova D. , Faresjo M. , Stechova K.
Published: October 2007

Type 1 diabetes (T1D) is a great medical challenge and its incidence rises rapidly. T lymphocytes and their cytokine production are supposed to play a major role in T1D development. So far, there is no potent tool to recognize the early signs of cellular auto‐reactivity which leads to β‐cell damage. The naïve immune system of the newborn (not yet influenced by external factors) can be used as an important model for T1D pathogenesis studies. Cord blood samples of 22 healthy neonates born at term to a diabetic parent (T1DR) and 15 newborns with no family history of any autoimmune disease (controls) were collected. Determination of 23 cytokines was performed before and after the stimulation with diabetogenic autoantigens using protein microarray. We observed lower basal production of all detected cytokines in the T1DR group – granulocyte/macrophage colony‐stimulating factor (GM‐CSF) (P = 0.025), growth regulated protein (GRO) (P = 0.002), GRO‐α (P = 0.027), interleukin (IL)‐1‐α (P = 0.051), IL‐3 (P = 0.008), IL‐7 (P = 0.027), IL‐8 (P = 0.042), monocyte chemoattractant proteins (MCP)‐3 (P = 0.022), monokine‐induced by IFN‐γ (MIG) (P = 0.034) and regulated upon activation normal T‐cell express sequence (RANTES) (P = 0.004). Exclusively lower post‐stimulative levels of G‐CSF (P = 0.030) and GRO‐α (P = 0.04) were observed in controls in comparison with the basal levels. A significant post‐stimulative decrease in G‐CSF (P = 0.030) and MCP‐2 (P = 0.009) levels was observed in controls in comparison with T1DR neonates. We also observed the interesting impact of the risky genotype on the protein microarray results. Protein microarray seems to be a useful tool to characterize a risk pattern of the immune response for T1D also in newborns.
Human Soluble CD80 is Generated by Alternative Splicing, and Recombinant Soluble CD80 Binds to CD28 and CD152 Influencing T‐cell Activation
Authors: Kakoulidou M. , Giscombe R. , Zhao X. , Lefvert A. K. , Wang X.
Published: October 2007

CD80 is a costimulatory factor mainly expressed on the surface of activated monocytes, B cells and dendritic cells. In this study, we demonstrate that 24% of healthy individuals have soluble forms of CD80, sCD80, in their serum. The concentration of sCD80 ranged from 0 to 1 mg/l. At the mRNA level, we detected a spliced form s1CD80 (771 bp), in unstimulated monocytes and B cells, while another form named s2CD80 (489 bp) was expressed in activated T cells as well as in freshly isolated and activated monocytes. s1CD80 lacks the transmembrane domain, and the IgC‐like domain plus the transmembrane domain are spliced out of s2CD80. We also present data demonstrating that recombinant s1CD80 binds to recombinant CD152‐Ig and CD28‐Ig. It can also bind to T cells, preferentially to activated T cells. Recombinant sCD80 had immunomodulatory effects shown by its inhibition of the mixed lymphocyte reaction and inhibition of T‐cell proliferation. sCD80 in human serum adds a new member to the family of soluble receptors, implying a network of soluble costimulatory factors with functional relevance. The inhibitory effect of the recombinant protein on T‐cell activation makes it a possible candidate for treatment of diseases associated with hyperactivated T cells.
The Effect of the Cholinergic Anti‐Inflammatory Pathway on Experimental Colitis
Authors: Bai A. , Guo Y. , Lu N.
Published: October 2007

Inflammatory bowel diseases (IBD) are characterized by proinflammatory cytokines, tissue damage and loss of neuron in inflamed mucosa, which implies the cholinergic anti‐inflammatory pathway may be destroyed during the process of inflammatory response. In the study, we identified the effect of cholinergic agonist as anabaseine (AN) and nicotinic receptor antagonist as chlorisondamine diiodide (CHD) on trinitrobenzene sulfonic acid (TNBS)‐induced colitis, to investigate the potential therapeutic effect of the cholinergic anti‐inflammatory pathway on IBD. Experimental colitis was induced by TNBS at day 1, 10 μg AN or 1.5 μg CHD was injected i.p. to mouse right after the induction of colitis, and repeated on interval day till the mice were sacrificed at day 8. Colonic inflammation was examined by histological analysis, myeloperoxidase (MPO) activity, and the production of tumour necrosis factor (TNF)‐α in tissue. Lamina propria mononuclear cells (LPMC) were isolated, and NF‐κB activation was detected by western blot. The mice with colitis treated by AN showed less tissue damage, less MPO activity, less TNF‐α production in colon, and inhibited NF‐κB activation in LPMC, compared with those mice with colitis untreated, whereas the mice with colitis treated by CHD showed the worst tissue damage, the highest MPO activity, the highest TNF‐α level, and enlarged NF‐κB activation in LPMC. Agonist of the cholinergic anti‐inflammatory pathway inhibits colonic inflammatory response by downregulating the production of TNF‐α, and inhibiting NF‐κB activation, which suggests that modulating the cholinergic anti‐inflammatory pathway may be a new potential management for IBD.
Long‐term Protection in Hamsters against Human Parainfluenza Virus Type 3 Following Mucosal or Combinations of Mucosal and Systemic Immunizations with Chimeric Alphavirus‐based Replicon Particles
Authors: Greer C. E. , Zhou F. , Goodsell A. , Legg H. S. , Tang Z. , Zur Megede J. , Uematsu Y. , Polo J. M. , Vajdy M.
Published: October 2007

No licensed vaccines are available to protect against parainfluenza virus type 3 (PIV3), a significant health risk for infants. In search of a safe vaccine, we used an alphavirus‐based chimeric vector, consisting of Sindbis virus (SIN) structural proteins and Venezuelan equine encephalitis virus (VEE) replicon RNA, expressing the PIV3 hemagglutinin‐neuraminidase (HN) glycoprotein (VEE/SIN‐HN). We compared different routes of intramuscular (IM), intranasal (IN), or combined IN and IM immunizations with VEE/SIN‐HN in hamsters. Six months after the final immunization, all hamsters were protected against live PIV3 IN challenge in nasal turbinates and lungs. This protection appeared to correlate with antibodies in serum, nasal turbinates and lungs. This is the first report demonstrating mucosal protection against PIV3 for an extended time following immunizations with an RNA replicon delivery system.
Comparison of five xylan synthesis mutants reveals new insight into the mechanisms of xylan synthesis
Authors: Brown David M. , Goubet Florence , Wong Vicky W. , Goodacre Royston , Stephens Elaine , Dupree Paul , Turner Simon R.
Published: October 2007

Previous studies using co‐expression analysis have identified a large number of genes likely to be involved in secondary cell‐wall formation. However, the function of very few of these genes is known. We have studied the cell‐wall phenotype of irx7, irx8 and irx9, three previously described irregular xylem (irx) mutants, and irx14 and parvus‐3, which we now show also to be secondary cell‐wall mutants. All five mutants, which have mutations in genes encoding putative glycosyltransferases, exhibited large decreases in xylan. In addition, all five mutants were found to have the same specific defect in xylan structure, retaining MeGlcUA but lacking GlcUA side branches. Polysaccharide analysis by carbohydrate gel electrophoresis (PACE) was used to determine the xylan structure in Arabidopsis, and revealed that side branches are added to approximately one in every eight xylose residues. Interestingly, this ratio is constant in all the lines analysed despite the wide variation in xylan content and the absence of GlcUA branches. Xylanase digestion of xylan from wild‐type plants released a short oligosaccharide sequence at the reducing end of the xylan chain. MALDI‐TOF MS analysis indicated that this sequence of sugars was absent in xylan from irx7, irx8 and parvus‐3 mutants, but was present in irx9 and irx14. This is consistent with previous NMR analysis of xylan from irx7, irx8 and irx9, and suggests that PARVUS may be involved in the synthesis of a xylan primer whereas IRX14 may be required to synthesize the xylan backbone. This hypothesis is supported by assays showing that irx9 and irx14 are both defective in incorporation of radiolabel from UDP 14C‐xylose. This study has important implications for both our understanding of xylan biosynthesis and the functional analysis of cell‐wall biosynthesis genes.
Effect of an inhaled adenosine A2A agonist on the allergen‐induced late asthmatic response
Authors: Luijk B. , Van Den Berge M. , Kerstjens H. A. M. , Postma D. S. , Cass L. , Sabin A. , Lammers J.‐W. J.
Published: October 2007

Background:  Adenosine receptor activation is suggested to play a role in asthmatic airway inflammation. Inhibition of adenosine receptors may have an effect on the late asthmatic response (LAR) after allergen inhalation and this mechanism could offer a potential new treatment in asthma.

Methods:  We evaluated the effect of an inhaled adenosine‐2A (A2A)‐receptor agonist (GW328267X), 25 μg, in 15 nonsmoking atopic asthmatics who underwent an inhaled allergen challenge following twice daily treatment for 1 week in a double‐blind, placebo‐ and fluticasone propionate (250 μg) controlled study.

Results:  In contrast to fluticasone, treatment with the A2A‐receptor agonist neither significantly protect against the allergen‐induced early and late asthmatic reaction, nor the accompanying inflammatory response as measured by sputum total cell counts, number of EG2+ cells, and the concentration of interleukin‐8 and eosinophil cationic protein.

Conclusion:  The inhaled A2A‐receptor agonist, GW328267X, 25 μg does not affect the allergen‐induced LAR or the associated inflammatory response in asthma.
Galectin‐10 mRNA is overexpressed in peripheral blood of aspirin‐induced asthma
Authors: Devouassoux G. , Pachot A. , Laforest L. , Diasparra J. , Freymond N. , Van Ganse E. , Mougin B. , Pacheco Y.
Published: October 2007

Background:  In sensitive patients, aspirin is associated with nasal and bronchial inflammation, eliciting local symptoms. Although the disease is clinically well characterized, its physiopathology is incompletely understood and noninvasive procedures, allowing an effective distinction between aspirin‐induced asthma (AIA) and aspirin‐tolerant asthma (ATA) are missing.

Objectives:  The aims of the study were to compare AIA and ATA cohorts for clinical characteristics and to screen peripheral blood for differential mRNA expression.

Methods:  Patients experiencing symptoms following aspirin ingestion were considered as aspirin sensitive. Peripheral blood was collected to quantify mRNA expression, using microarray technology and quantitative RT‐PCR.

Results:  Data indicated that AIA and ATA share large number of similarities for clinical phenotype. Screening of mRNA expression using microarray showed an overexpression of galectin‐10 mRNA in AIA (AIA/ATA ratio = 1.9, P < 0.05). Results were confirmed using qRT‐PCR. A positive correlation was established between microarray and qRT‐PCR results for galectin‐10 mRNA expression (r = 0.92, P < 0.0001). Finally, qRT‐PCR results were validated on a subset of asthmatics and controls, showing an increased expression of galectin‐10 mRNA in AIA vs ATA (P < 0.001) and vs controls (P < 0.01).

Conclusions:  Our results demonstrate that AIA and ATA remain difficult to distinguish using clinical criteria. Employing two molecular biological methods, we demonstrate that galectin‐10 mRNA is overexpressed in AIA, suggesting a novel candidate gene and a potentially innovative pathway for mucosal inflammation in aspirin intolerance.
Rapid detection of the cariogenic pathogens Streptococcus mutans and Streptococcus sobrinus using loop‐mediated isothermal amplification
Authors: Nagashima S. , Yoshida A. , Ansai T. , Watari H. , Notomi T. , Maki K. , Takehara T.
Published: October 2007

Introduction:  Streptococcus mutans and Streptococcus sobrinus are associated with the development of dental caries in humans. In this study, we developed a rapid, sensitive method for detecting these major cariogenic pathogens using loop‐mediated isothermal amplification (LAMP). The assay procedure is quite simple: the amplification is carried out in a single tube under isothermal conditions at 63°C, and the result can be obtained in less than 1 h.

Methods:  Initially, a set of six primers was designed by targeting S. mutans‐specific and S. sobrinus‐specific regions, identified using the genomic subtractive hybridization technique. We evaluated the specificities and sensitivities of these assays. Furthermore, we detected and quantified these bacteria in saliva and carious dentin from eight children.

Results:  The sensitivities of the S. mutans‐specific and S. sobrinus‐specific LAMP methods, examined using agarose gel electrophoresis, were each one cell for a 30‐min reaction. The detection limits using real‐time turbidimetry analysis were 1 to 107 cells (3.28 × 101 to 3.28 × 108 fg S. mutans template DNA) per reaction tube and 1 to 105 cells (2.72 × 103 to 2.72 × 108 fg S. sobrinus template DNA) per reaction tube. Using these assays, we detected and quantified these cariogenic bacteria for evaluation of the LAMP assay for clinical diagnosis.

Conclusions:  Our results suggest that the LAMP‐based assay in combination with subtractive hybridization is valuable for preparing species‐specific primers for closely related species. Furthermore, the LAMP‐based assay will be a useful tool for the rapid and sensitive prediction of dental caries.
Nitrite reductase in Streptoccocus mutans plays a critical role in the survival of this pathogen in oral cavity
Authors: Choudhury T. , Sato E. F. , Inoue M.
Published: October 2007

Background/aims:  The mechanisms of nitric oxide (NO) production by bacteria in the oral cavity are still not clearly defined but salivary streptococci have been reported to generate NO. The aim of this study was to clarify the mechanism of nitrite metabolism and generation of NO by Streptococcus mutans, a major pathogen of dental caries.

Methods:  We searched the genomic database of oral pathogens for nitrite reductase and used a polymerase chain reaction (PCR) to clone the nirJ gene from S. mutans GS5. His‐tagged recombinant NirJ protein was expressed in Escherichia coli BL21 and characterized. We constructed a nirJ gene‐disrupted mutant strain of S. mutans (ΔnirJ) to analyze the physiological significance of nirJ.

Results:  S. mutans generates NO from nitrite, probably as a result of the possession of nitrite reductase. We cloned the nirJ gene from S. mutans GS5 by PCR. The recombinant NirJ protein catalyzed the reduction of nitrite with a Km value of 3.37 μm and a specific activity of 2.5 μmol/min/mg of protein at 37°C. Biochemical analysis revealed that the nitrite‐reducing activity of the mutant (ΔnirJ) strain was significantly lower than that of the wild‐type strain. The growth of the mutant strain, but not of the wild‐type strain, was strongly suppressed by the presence of physiological levels of nitrite (∼0.2 mm) in saliva.

Conclusion:  These observations suggest that the elimination of nitrite and/or the generation of NO are important for the survival of S. mutans in the oral cavity.
Quantitative messenger RNA expression of Toll‐like receptors and interferon‐α1 in gingivitis and periodontitis
Authors: Kajita K. , Honda T. , Amanuma R. , Domon H. , Okui T. , Ito H. , Yoshie H. , Tabeta K. , Nakajima T. , Yamazaki K.
Published: October 2007

Introduction:  In addition to bacteria, viruses have been reportedly implicated in periodontitis. However, the available data are confined to Toll‐like receptor 2 (TLR2) and TLR4, which recognize bacterial products in periodontitis. In the present study, we investigated the expression levels of TLR5, ‐7, and ‐9 messenger RNAs (mRNAs) in addition to those of TLR2 and ‐4, and compared gingivitis and periodontitis. Interferon‐α1 (IFN‐α1), which is important for the antiviral response, was also compared.

Methods:  Gene expression was analyzed using quantitative real‐time polymerase chain reaction for 59 periodontitis and 27 gingivitis tissue samples together with viral serology in some patients. The presence of plasmacytoid dendritic cells (pDCs), a robust producer of IFN‐α, was immunohistochemically analyzed in an additional seven periodontitis and two gingivitis specimens.

Results:  The expression levels of TLR2, ‐4, ‐7, and ‐9 were significantly higher in periodontitis lesions than gingivitis lesions. The expression level of TLR5 was comparable to levels of TLR2 and ‐4; however, no significant difference was found between gingivitis and periodontitis. Although the expression of IFN‐α1 mRNA was higher in periodontitis lesions compared with gingivitis lesions, the level was quite low. Only a few pDCs were found in some periodontitis specimens. No difference was found for antibody‐positivity between gingivitis and periodontitis.

Conclusion:  This is the first study to show that a variety of TLRs are up‐regulated in periodontitis lesions compared with gingivitis lesions, suggesting that diverse microbial and possibly viral antigens are involved in the pathogenic mechanisms for periodontal diseases. However, the ligands recognized by the various TLRs in periodontal lesions remain to be determined.
Bacterial examination of endodontic infections by clonal analysis in concert with denaturing high‐performance liquid chromatography
Authors: Jacinto R. C. , Gomes B. P. F. A. , Desai M. , Rajendram D. , Shah H. N.
Published: October 2007

Background/aims:  The aim of this study was to examine the diversity of bacterial species in the infected root canals of teeth associated with endodontic abscesses by cloning and sequencing techniques in concert with denaturing high‐performance liquid chromatography.

Methods:  Samples collected from five infected root canals were subjected to polymerase chain reaction (PCR) with universal 16S ribosomal DNA primers. Products of these PCRs were cloned and sequenced. Denaturing high‐performance liquid chromatography (DHPLC) was used as a screening method to reduce the number of clones necessary for DNA sequencing.

Results:  All samples were positive for the presence of bacteria and a range of 7–13 different bacteria were found per root canal sample. In total, 48 different oral clones were detected among the five root canal samples. Olsenella profusa was the only species present in all samples. Porphyromonas gingivalis, Dialister pneumosintes, Dialister invisus, Lachnospiraceae oral clone, Staphylococcus aureus, Pseudoramibacter alactolyticus, Peptostreptococcus micros and Enterococcus faecalis were found in two of the five samples. The majority of the taxa were present in only one sample, for example Tannerella forsythia, Shuttleworthia satelles and Filifactor alocis. Some facultative anaerobes that are frequently isolated from endodontic infections such as E. faecalis, Streptococcus anginosus and Lactobacillus spp. were also found in this study.

Conclusion:  Clonal analysis of the microflora associated with endodontic infections revealed a wide diversity of oral species.
Growth‐phase‐dependent gene expression profiling of poplar (Populus alba × Populus tremula var. glandulosa) suspension cells
Authors: Lee Hyoshin , Bae Eun‐Kyung , Park So‐Young , Sjödin Andreas , Lee Jae‐Soon , Noh Eun‐Woon , Jansson Stefan
Published: October 2007

Complex sequences of morphological and biochemical changes occur during the developmental course of a batch plant cell culture. However, little information is available about the changes in gene expression that could explain these changes, because of the difficulties involved in isolating specific cellular events or developmental phases in the overlapping phases of cell growth. In an attempt to obtain such information we have examined the global growth phase‐dependent gene expression of poplar cells in suspension cultures by cDNA microarray analysis. Our results reveal that significant changes occur in the expression of genes with functions related to protein synthesis, cell cycling, hormonal responses and cell wall biosynthesis, as cultures progress from initiation to senescence, that are highly correlated with observed developmental and physiological changes in the cells. Genes encoding protein kinases, calmodulin and proteins involved in both ascorbate metabolism and water‐limited stress responses also showed strong stage‐specific expression patterns. Our report provides fundamental information on molecular mechanisms that control cellular changes throughout the developmental course of poplar cell cultures.
A case of subcutaneous Mycoleptodiscus indicus infection in a liver transplant recipient successfully treated with antifungal therapy
Authors: Garrison A.P. , Procop G.W. , Vincek V. , Moon J. , Morris M.I. , Doblecki‐Lewis S. , Cleary T.J. , Brust D. , Rosa‐Cunha I.
Published: October 2007

Abstract: Mycoleptodiscus indicus, a dematiaceous mold, occurs on the leaves of a number of different host plants and has been only recently described as a cause of human infection. Immunosuppressed individuals are at risk for developing infections with opportunistic fungal pathogens, which are a major cause of morbidity and mortality in this population. In addition, the treatment of infections caused by these fungi is frequently challenging. We report a case of M. indicus subcutaneous infection in a 51‐year‐old man with human immunodeficiency virus and hepatitis C co‐infection, who had a liver transplant. He developed skin nodules with a sporotrichoid lymphangitic distribution. Histopathology demonstrated unusual fungal elements with angioinvasion. Mycology cultures isolated a dematiaceous mold with the characteristic curved hyaline conidia of M. indicus. Initial treatment involved a combination of amphotericin B lipid complex and voriconazole, followed by monotherapy with voriconazole. The subcutaneous lesions resolved completely after 4 months of antifungal therapy.