bluuebunny/crossref_metadata_2025 huggingface dataset:
Safety, immunogenicity and efficacy of peptide‐pulsed cellular immunotherapy in macaques
Authors: De Rose Robert , Mason Rosemarie D. , Loh Liyen , Peut Viv , Smith Miranda Z. , Fernandez Caroline S. , Alcantara Sheilajen , Amarasena Thakshila , Reece Jeanette , Seddiki Nabila , Kelleher Anthony D. , Zaunders John , Kent Stephen J.
Published: January 2009

Background  Simple and effective delivery methods for cellular immunotherapies are needed. We recently published on the effectiveness of using ex vivo pulsing of overlapping SIV Gag 15mer peptides onto fresh peripheral blood cells in 32 SIVmac251‐infected pigtail macaques.

Methods  We now report on the safety of this approach, analysis of a novel assay for immunogenicity, the effect of an MHC allele, Mane‐A*10, on CD8 T cell escape occurring and disease outcome.

Results  The vaccine strategy was safe, with no perturbations in weight or hematological profiles in comparison to controls. The high levels of SIV‐specific T cell immunogenicity of this approach was confirmed using a novel assay measuring upregulation of surface CD134 of CD4 T cells. A substantial effect of the Mane‐A*10 allele in reducing SIV viral load of pigtail macaques was observed in both vaccinees and controls; the virologic efficacy of the immunotherapy in comparison to controls was greatest in Mane‐A*10− animals. Escape mutations at several new CD8 T cell epitopes throughout the SIV proteome were observed, primarily in animals with poorer virologic control.

Conclusions  In summary, we provide further information that peptide‐pulsed PBMC are a safe, immunogenic and effective immunotherapy. The observed influence of MHC alleles and immune escape allows us to design more insightful future immunotherapy studies.
Epistaxis during Treatment with Paclitaxel
Authors: Yarom Nirit , Cyjon Arnoldo , Kovel Svetlana , Yachnin Adelya , Sella Avishay , Evron Ella
Published: January 2009

Abstract:  The purpose of this study was to determine the incidence and severity of epistaxis in patients treated with paclitaxel. Patients who were treated with paclitaxel filled a questionnaire regarding their general health, medications and incidents of epistaxis. Relevant clinical information was obtained from the patients’ charts. Forty‐seven consecutive patients were recruited to the study. Twenty‐four (51%) of the patients reported epistaxis during paclitaxel therapy. Twenty‐three of 39 (59%) patients who received weekly paclitaxel had epistaxis at least once during treatment, compared with one out of eight patients who were treated every 3 weeks (P = 0.045). All episodes of epistaxis were mild, occurred with normal platelets counts and did not require blood product transfusions or treatment modification. The majority of the patients experienced the first episode of epistaxis on the third week of weekly paclitaxel treatment and then repeatedly throughout therapy. It is concluded that epistaxis is a common mild side‐effect of weekly paclitaxel that has not been reported previously. In this trial, epistaxis did not have any major clinical consequences. However, when paclitaxel is combined with other drugs that may cause bleeding, such as bevacizumab, physicians should be alerted to the potential risk of this phenomenon.
Testosterone Replacement Therapy Following Radical Prostatectomy
Authors: Khera Mohit , Grober Ethan D. , Najari Bobby , Colen John S. , Mohamed Osama , Lamb Dolores J. , Lipshultz Larry I.
Published: January 2009

Introduction

Controversy exists regarding testosterone replacement therapy (TRT) in men following radical prostatectomy (RP). Many clinicians are hesitant to offer patients TRT after an RP, out of concern that the increased androgen levels may promote tumor progression or recurrence from residual tumor. Recently, several small studies have demonstrated the use of TRT in men following an RP and have shown an improvement in serum testosterone levels with no increase in prostate-specific antigen (PSA) values.

Aims

The aim of this article is to assess changes in PSA and testosterone values in hypogonadal patients on TRT after RP and also to evaluate the impact of pathologic Gleason grade on ultimate PSA values.

Methods

All hypogonadal men who were treated with TRT by members of our department following RP were retrospectively reviewed. PSA values before RP, after RP, and after TRT were evaluated. Serum testosterone levels before and after TRT were also examined. Only patients with undetectable PSA values and negative surgical margins on pathologic specimen were offered TRT and included in the study.

Main Outcome Measures

Main outcome measures were changes in PSA and testosterone values after initiation of TRT.

Results

Fifty-seven men, ages 53–83 years (mean 64), were identified as having initiated TRT following RP. Men received TRT for an average of 36 months following RP (range 1–136 months). Patients were followed an average of 13 months after initiation of TRT (range 1–99 months). The mean testosterone values rose from 255 ng/dL before TRT to 459 ng/dL after TRT (P < 0.001). There was no increase in PSA values after initiation of TRT and thus no patient had a biochemical PSA recurrence.

Conclusion

TRT is effective in improving testosterone levels, without increasing PSA values, in hypogonadal men who have undergone RP.
On Three‐Dimensional Holographic Vector of Atomic Interaction Field Analysis for Influenza Neuraminidase Inhibitors
Authors: Li Zhiliang S. , Sun Jiaying Y. , Liang Guizhao Z. , Lu Fenglin L. , Zhu Wanping P. , Zhang Mengjun J. , Zhang Yonghong , Yang Shanbin B. , Shu Mao , Chen Guohua H. , Lu Tingting T.
Published: January 2009

Influenza is a major respiratory infection associated with significant morbidity in the general population and mortality in elderly and high‐risk patients. It is an RNA virus that contains two major surface glycoproteins, neuraminidase and hemagglutinin. These proteins are essential for infection. Neuraminidase has been found to be a potential target to control influenza virus. Here, we have developed three‐dimensional holographic vector of atomic interaction field analysis as a new method of quantitative structure–activity relationships for different sets of compounds to understand chemical–biological interactions governing their activities toward influenza neuraminidase. Good results, R = 0.885, SD = 0.848, RCV = 0.858 (the maximum) and SDCV = 0.934 (the minimum), showed that holographic vector of atomic interaction field analysis can be applicable to molecular structural characterization and biological activity prediction and quantitative structure–activity relationship model had favorable stability and prediction capability.
Identification of Allosteric PIF‐Pocket Ligands for PDK1 using NMR‐Based Fragment Screening and 1H‐15N TROSY Experiments
Authors: Stockman Brian J. , Kothe Michael , Kohls Darcy , Weibley Laura , Connolly Brendan J. , Sheils Alissa L. , Cao Qing , Cheng Alan C. , Yang Lily , Kamath Ajith V. , Ding Yuan‐Hua , Charlton Maura E.
Published: January 2009

Aberrant activation of the phosphoinositide 3‐kinase pathway because of genetic mutations of essential signalling proteins has been associated with human diseases including cancer and diabetes. The pivotal role of 3‐phosphoinositide‐dependent kinase‐1 in the PI3K signalling cascade has made it an attractive target for therapeutic intervention. The N‐terminal lobe of the 3‐phosphoinositide‐dependent kinase‐1 catalytic domain contains a docking site which recognizes the non‐catalytic C‐terminal hydrophobic motifs of certain substrate kinases. The binding of substrate in this so‐called PDK1 Interacting Fragment pocket allows interaction with 3‐phosphoinositide‐dependent kinase‐1 and enhanced phosphorylation of downstream kinases. NMR spectroscopy was used to a screen 3‐phosphoinositide‐dependent kinase‐1 domain construct against a library of chemically diverse fragments in order to identify small, ligand‐efficient fragments that might interact at either the ATP site or the allosteric PDK1 Interacting Fragment pocket. While majority of the fragment hits were determined to be ATP‐site binders, several fragments appeared to interact with the PDK1 Interacting Fragment pocket. Ligand‐induced changes in 1H‐15N TROSY spectra acquired using uniformly 15N‐enriched PDK1 provided evidence to distinguish ATP‐site from PDK1 Interacting Fragment‐site binding. Caliper assay data and 19F NMR assay data on the PDK1 Interacting Fragment pocket fragments and structurally related compounds identified them as potential allosteric activators of PDK1 function.
Synthesis, Evaluation of Pharmacological Activities and Quantitative Structure–Activity Relationship Studies of a Novel Group of bis(4‐Nitroaryl‐1,4‐dihyropyridine)
Authors: Miri Ramin , Javidnia Katayoun , Hemmateenejad Bahram , Tabarzad Maryam , Jafarpour Mehrnaz
Published: January 2009

Voltage‐dependent calcium channels are crucial targets for a wide range of clinically active pharmacological agents. From these agents, 1,4‐dihydropyridines constitute a group of small organic compounds are based on a core pyridine structure which can both block and enhance calcium currents. They are considered specific for L‐Type calcium channels; however, other channel types, and in particular certain T‐Type channels, may show sensitivity to dihydropyridine compounds. In this study, we synthesized a novel group of bis‐1,4‐dihydropyridines using the procedure reported by Dagnino that involved the condensation of n‐alkyl diacetoacetate (n = 2–7) with methyl‐3‐aminocrotonate and nitrophenylaldehyde. The synthesis was run under two conditions: (i) reflux and (ii) microwave. Calcium channels antagonist activity were determined in vitro using guinea‐pig ileum longitudinal smooth muscle assay. Synthesis of these compounds was confirmed with 1H‐NMR, IR and mass spectrometry. Then IC50 of them are calculated and compared with Nifedipine. Finally, the result of this pharmacological assay was used in quantitative structure–activity relationship studies utilizing multiple linear regression analysis. Most of these compounds are less active compared with Nifedipine. Decrease in activity is the result of increase in steric hindrance. The quantitative structure–activity relationship study indicates that the activity is related to the electrostatic and topological parameters and the distance between two C5‐esteric groups of 1,4‐dihydropyridine rings.
Avian influenza surveillance in wild birds in the European Union in 2006
Authors: Hesterberg Uta , Harris Kate , Stroud David , Guberti Vittorio , Busani Luca , Pittman Maria , Piazza Valentina , Cook Alasdair , Brown Ian
Published: January 2009

Background  Infections of wild birds with highly pathogenic avian influenza (AI) subtype H5N1 virus were reported for the first time in the European Union in 2006.

Objectives  To capture epidemiological information on H5N1 HPAI in wild bird populations through large‐scale surveillance and extensive data collection.

Methods  Records were analysed at bird level to explore the epidemiology of AI with regard to species of wild birds involved, timing and location of infections as well as the applicability of different surveillance types for the detection of infections.

Results  In total, 120,706 records of birds were sent to the Community Reference Laboratory for analysis. Incidents of H5N1 HPAI in wild birds were detected in 14 EU Member States during 2006. All of these incidents occurred between February and May, with the exception of two single cases during the summer months in Germany and Spain.

Conclusions  For the detection of H5N1 HPAI virus, passive surveillance of dead or diseased birds appeared the most effective approach, whilst active surveillance offered better detection of low pathogenic avian influenza (LPAI) viruses. No carrier species for H5N1 HPAI virus could be identified and almost all birds infected with H5N1 HPAI virus were either dead or showed clinical signs. A very large number of Mallards (Anas platyrhynchos) were tested in 2006 and while a high proportion of LPAI infections were found in this species, H5N1 HPAI virus was rarely identified in these birds. Orders of species that appeared to be very clinically susceptible to H5N1 HPAI virus were swans, diving ducks, mergansers and grebes, supporting experimental evidence. Surveillance results indicate that H5N1 HPAI virus did not establish itself successfully in the EU wild bird population in 2006.
Pre‐weaning morbidity and mortality of llamas and alpacas
Authors: Sharpe MS , Lord LK , Wittum TE , Anderson DE
Published: January 2009

Objectives To describe the morbidity and mortality patterns and identify factors associated with morbidity in pre‐weaning llamas and alpacas.

Design Cross‐sectional observational study of 287 crias born on four farms in Ohio, USA

Procedure Historical data representing all crias born over a 6‐year period were obtained from two llama farms and two alpaca farms in Ohio. Multivariable generalised linear mixed effects regression models were used to identify factors associated with morbidity outcomes.

Results In total, 105 (37%) of the llamas and alpacas had some reported morbidity during the pre‐weaning period, and mortality rate was 2.1%. In addition, 51 (18%) of llamas and alpacas experienced morbidity because of infectious disease, and 47 (16%) experienced morbidity during the neonatal period. The three most commonly reported morbidity events were undifferentiated diarrhoea (23%), umbilical hernia (16%) and unspecified infectious disease (15%). Difficult birth was an important risk factor for pre‐weaning morbidity in this population.

Conclusions Camelid veterinarians and their clients can expect that pre‐weaning health events are common among crias, although mortality is low. Crias experiencing difficult births may require additional monitoring for health events during the pre‐weaning period.
Unidentified microsporidian in western pygmy perch Edelia vittata (Nannopercidae) from Australia
Authors: Stephens FJ
Published: January 2009

Specimens of western pygmy perch, Edelia vittata Castelnau 1873, were submitted for examination because of the presence of black masses in the body wall in the abdominal area. The owner reported that approximately 10% of mature fish in one dam were affected. The masses had no apparent effect on the health of affected fish, but they were unsuitable for sale for aesthetic reasons. After necropsy and histological examination, the masses were found to be microsporidial xenomas within the abdominal cavity. Transmission electron microscopy results suggest that this microsporidian is a previously unreported species, because the melanised xenoma wall and internal structure of the spores differ from previously described species. The epidemiology of the parasite remains largely unknown and highlights the need for further surveys of the disease status and parasites of Australian native fish, especially those of interest to aquaculturists.
Effect of the non‐steroidal anti‐inflammatory drug, carprofen, on weaned sheep following non‐surgical mulesing by intradermal injection of cetrimide
Authors: Colditz IG , Lloyd JB , Paull DR , Lee C , Giraudo A , Pizzato C , Fisher AD
Published: January 2009

Objective   To assess in weaned lambs the palliative effects of the non‐steroidal anti‐inflammatory drug, carprofen, following intradermal injection of cetrimide to induce non‐surgical mulesing.

Procedures   We allocated 40 weaned lambs (20–22 weeks old) to four groups of 10 animals: (1) control, 2) conventional surgical mules, (3) intradermal treatment and (4) intradermal treatment + carprofen. Non‐surgical mulesing was induced by intradermal injection of 4% (w/w) cetrimide + 3% (w/w) polyvinylpyrrolidone in water. In group 4, carprofen (4 mg/kg, SC) was administered 1 h before intradermal treatment. Five weaners, including an animal from each treatment, were run in each pen. Neutrophil to lymphocyte ratio, cortisol, beta‐endorphin and haptoglobin levels and rectal temperature were monitored at least daily for the first 7 days after treatment, then weekly until day 28. Body weight was measured weekly and behaviour was measured every 15 min for 12 h on the day of treatment, then on days 1, 2, 4, 6, 12, 21 and 28 following treatment.

Results   The intradermal treatment resulted in high fever and elevated blood cortisol by 12 h. Rectal temperatures were significantly elevated until 5 days after treatment, cortisol was elevated until 3 days after treatment, haptoglobin for at least 7 days after treatment and the neutrophil to lymphocyte ratio until 4 days after treatment. Average daily gain was depressed in the week following treatment. Abnormal behaviours (hunched standing, stiff walking, pawing, lateral lying and lying intention) were increased on the day of treatment and for 6 days post treatment. Carprofen reduced the time spent in abnormal behaviours by approximately two‐thirds but did not ameliorate the physiological responses to the intradermal treatment.

Conclusions   In weaner sheep, carprofen ameliorated the behavioural responses, but was unable to provide relief from the intense and sustained physiological responses to non‐surgical mulesing by intradermal injection of cetrimide. Systemic side‐effects may be unavoidable with formulations based on quaternary ammonium compounds that are designed to reduce the risk of fly strike in sheep by remodelling breech tissue through induction of tissue necrosis.
Sulfur‐associated polioencephalomalacia in cattle grazing plants in the Family Brassicaceae
Authors: McKenzie RA , Carmichael AM , Schibrowski ML , Duigan SA , Gibson JA , Taylor JD
Published: January 2009

Polioencephalomalacia was diagnosed histologically in cattle from two herds on the Darling Downs, Queensland, during July–August 2007. In the first incident, 8 of 20 18‐month‐old Aberdeen Angus steers died while grazing pastures comprising 60%Sisymbrium irio(London rocket) and 40%Capsella bursapastoris(shepherd's purse). In the second incident, 2 of 150 mixed‐breed adult cattle died, and another was successfully treated with thiamine, while grazing a pasture comprising almost 100%Raphanus raphanistrum(wild radish). Affected cattle were either found dead or comatose or were seen apparently blind and head‐pressing in some cases. For both incidents, plant and water assays were used to calculate the total dietary sulfur content in dry matter as 0.62% and 1.01% respectively, both exceeding the recommended 0.5% for cattle eating more than 40% forage. Blood and tissue assays for lead were negative in both cases. No access to thiaminase, concentrated sodium ion or extrinsic hydrogen sulfide sources were identified in either incident. Below‐median late summer and autumn rainfall followed by above‐median unseasonal winter rainfall promoted weed growth at the expense of wholesome pasture species before these incidents.
Cross‐species amplification tests and diversity analysis using 56 PCR markers in Dactylis glomerata and Lolium perenne
Authors: LITRICO I. , BECH N. , FLAJOULOT S. , CADIER D. , TALON C. , GIBELIN C. , BARRE P.
Published: January 2009

We report results of cross‐species amplification in Dactylis glomerata and Lolium perenne of 12 simple sequence repeats (SSRs) isolated from Lolium multiflorum×Festuca glaucescens, 42 SSRs from Festuca arundinacea and two sequence tagged sites from Oryza sativa. We compared the transferability and diversity between D. glomerata and L. perenne, which are important forage crops. While Nei's gene diversity values were equivalent in both species (from 0.14 to 0.92), the mean number of allele per locus was more important in D. glomerata than in L. perenne (5.45 vs. 4.50). These markers will be used for analysing population structure in grassland populations under agronomic practices.
Polymorphic microsatellite loci from Sprague's pipit (Anthus spragueii), a grassland endemic passerine bird
Authors: CRAWFORD NICHOLAS G. , PETERS MAUREEN B. , HAGEN CRIS , GLENN TRAVIS C. , DAVIS STEPHEN K. , SOMERS CHRISTOPHER M.
Published: January 2009

We developed 11 novel microsatellite loci for Sprague's pipit (Anthus spragueii), a songbird endemic to the Northern Great Plains of central North America. These loci were screened in 21 individuals from southern Saskatchewan, Canada. All 11 loci were polymorphic, with the number of alleles per locus ranging from five to 23, polymorphic information content from 0.312 to 0.947, and observed heterozygosity from 0.476 to 0.900. These loci will be suitable for a wide variety of studies on pipit natural history, ecology, and conservation.
Anonymous nuclear markers for Malagasy plated lizards (Zonosaurus)
Authors: NOONAN BRICE P. , YODER ANNE D.
Published: January 2009

We report the development of 18, single‐copy, anonymous nuclear loci from the Malagasy plated lizard Zonosaurus madagascariensis. More than 140 clones from a genomic library were examined and 38 potential loci tested across both closely and distantly related lizards. Of the 18 loci reported here, more than half (10) work in closely related zonosaurines although only one successfully amplified a homologous fragment in the distantly related iguanid (Oplurus). Sequences of these loci revealed a high frequency of single nucleotide polymorphisms, supporting previous reports of high levels of intraspecific variation in lizards.
Microsatellite DNA markers for population‐genetic studies of blue mackerel (Scomber australasicus) and cross‐specific amplification in S. japonicus
Authors: TANG C. Y. , TZENG C. H. , CHEN C. S. , CHIU T. S.
Published: January 2009

Blue mackerel (Scomber australasicus) is targeted by large‐scale purse‐seiners in the western North Pacific, and its stock structure is still contentious. Herein, we described 10 polymorphic microsatellite loci for blue mackerel. The number of alleles among 32 individuals surveyed ranged from five to 27 (average of 16.2 alleles per locus). Departures from Hardy–Weinberg expectation were observed at two loci. Cross‐specific amplification in the congener, S. japonicus, was successful, except for one locus, revealed to be diagnostic for these congeners. These microsatellite loci will be useful tools to address queries in population genetic structure, fishery management unit and taxonomic species status in the genus Scomber.
Heteroduplex molecules cause sexing errors in a standard molecular protocol for avian sexing
Authors: CASEY ASHLEY E. , JONES KENNETH L. , SANDERCOCK BRETT K. , WISELY SAMANTHA M.
Published: January 2009

Molecular methods are a necessary tool for sexing monomorphic birds. These molecular approaches are usually reliable, but sexing protocols should be evaluated carefully because biochemical interactions may lead to errors. We optimized laboratory protocols for genetic sexing of a monomorphic shorebird, the upland sandpiper (Bartramia longicauda), using two independent sets of primers, P2/P8 and 2550F/2718R, to amplify regions of the sex‐linked CHD‐Z and CHD‐W genes. We discovered polymorphisms in the region of the CHD‐Z intron amplified by the primers P2/P8 which caused four males to be misidentified as females (n = 90 mated pairs). We cloned and sequenced one CHD‐W allele (370 bp) and three CHD‐Z alleles in our population: Z° (335 bp), Z′ (331 bp) and Z″ (330 bp). Normal (Z°Z°) males showed one band in agarose gel analysis and were easily differentiated from females (Z°W), which showed two bands. However, males heterozygous for CHD‐Z alleles (Z′Z″) unexpectedly showed two bands in a pattern similar to females. While the Z′ and Z″ fragments contained only short deletions, they annealed together during the polymerase chain reaction (PCR) process and formed heteroduplex molecules that were similar in size to the W fragment. Errors previously reported for molecular sex‐assignment have usually been due to allelic dropout, causing females to be misidentified as males. Here, we report evidence that events in PCRs can lead to the opposite error, with males misidentified as females. We recommend use of multiple primer sets and large samples of known‐sex birds for validation when designing protocols for molecular sex analysis.
Isolation and characterization of microsatellite markers in the lowland leopard frog (Rana yavapaiensis) and the relict leopard frog (R. onca), two declining frogs of the North American desert southwest
Authors: SAVAGE ANNA E , JAEGER JEF R
Published: January 2009

We characterized 15 microsatellite loci for the lowland leopard frog (Rana yavapaiensis) and the relict leopard frog (R. onca) for future studies of population genetic structure and relatedness. Analysis of 20 individuals from single populations of each species showed that all markers were polymorphic in at least one species. Observed and expected heterozygosities ranged from 0 to 0.94 and from 0.11 to 0.85, respectively, and there were three to 11 alleles per locus. No loci were in linkage disequilibrium, but six loci deviated significantly from Hardy–Weinberg equilibrium, and the presence of a null allele was detected in two of these loci.
Ten polymorphic microsatellite loci isolated from the alpine caddisfly Allogamus uncatus Brauer (Trichoptera: Limnephilidae)
Authors: SHAMA LISA N. S , KUBOW KAREN B , ROBINSON CHRISTOPHER T
Published: January 2009

Here we report the development of 10 microsatellite loci for the alpine caddisfly, Allogamus uncatus. Polymorphism as detected in 24 individuals ranged from three to 17 alleles per locus, and observed heterozygosity ranged from 0.087 to 0.864. These primers will enable research on the genetic population structure of this species, the extent of gene flow among alpine permanent and temporary streams, and the genetic consequences of extinction/recolonization events.
Isolation and characterization of 13 polymorphic nuclear microsatellite primers for the widespread Indo‐Pacific three‐spot damselfish, Dascyllus trimaculatus, and closely related D. auripinnis
Authors: LERAY MATTHIEU , BELDADE RICARDO , HOLBROOK SALLY J. , SCHMITT RUSSELL J. , PLANES SERGE , BERNARDI GIACOMO
Published: January 2009

A set of 13 simple sequence repeat markers was developed from D. trimaculatus genomic DNA, tested for D. auripinnis and characterized using 40 individuals per species. All the loci were polymorphic with a number of alleles ranging from three to 30. Observed heterozygosities varied from 0.23 to 0.89 for D. trimaculatus and from 0.11 to 0.85 for D. auripinnis. Early results show that these will be powerful markers for the study of ecological and evolutionary mechanism in this coral reef fish species complex.
Characterization of microsatellite loci in the myco‐heterotrophic plant Monotropa hypopitys (Ericaceae) and amplification in related taxa
Authors: KLOOSTER M. R. , HOENLE A. W. , CULLEY T. M.
Published: January 2009

The myco‐heterotroph Monotropa hypopitys is a perennial, circumboreally distributed herb of significant importance in studies of nonphotosynthetic plant biology. To address a deficiency in our knowledge of myco‐heterotroph population genetics, 11 microsatellite markers were developed using a cost‐effective, nonradioactive protocol. Multiplex reactions revealed polymorphism in the red and yellow colour forms of M. hypopitys with an average of 2.69 alleles per primer. Many primers additionally amplified in the congener Monotropa uniflora and five other closely related genera. This is the first report of microsatellite primer development and amplification in the Monotropoideae (Ericaceae).
Two‐step multiplex polymerase chain reaction improves the speed and accuracy of genotyping using DNA from noninvasive and museum samples
Authors: ARANDJELOVIC M. , GUSCHANSKI K. , SCHUBERT G. , HARRIS T. R. , THALMANN O. , SIEDEL H. , VIGILANT L.
Published: January 2009

Many studies in molecular ecology rely upon the genotyping of large numbers of low‐quantity DNA extracts derived from noninvasive or museum specimens. To overcome low amplification success rates and avoid genotyping errors such as allelic dropout and false alleles, multiple polymerase chain reaction (PCR) replicates for each sample are typically used. Recently, two‐step multiplex procedures have been introduced which drastically increase the success rate and efficiency of genotyping. However, controversy still exists concerning the amount of replication needed for suitable control of error. Here we describe the use of a two‐step multiplex PCR procedure that allows rapid genotyping using at least 19 different microsatellite loci. We applied this approach to quantified amounts of noninvasive DNAs from western chimpanzee, western gorilla, mountain gorilla and black and white colobus faecal samples, as well as to DNA from ~100‐year‐old gorilla teeth from museums. Analysis of over 45 000 PCRs revealed average success rates of > 90% using faecal DNAs and 74% using museum specimen DNAs. Average allelic dropout rates were substantially reduced compared to those obtained using conventional singleplex PCR protocols, and reliable genotyping using low (< 25 pg) amounts of template DNA was possible. However, four to five replicates of apparently homozygous results are needed to avoid allelic dropout when using the lowest concentration DNAs (< 50 pg/reaction), suggesting that use of protocols allowing routine acceptance of homozygous genotypes after as few as three replicates may lead to unanticipated errors when applied to low‐concentration DNAs.
Development of microsatellite markers in the Australasian snake‐necked turtle Chelodina rugusa and cross‐species amplification
Authors: ALACS E. A , HILLYER M. J. , GEORGES A. , FITZSIMMONS N. N. , HUGHES J. M.
Published: January 2009

Seventeen microsatellite loci were developed for the snake‐necked turtle, Chelodina rugosa (Ogilby, 1890). Sixteen of the loci were polymorphic but three of these loci had null alleles. One locus displayed linkage disequilibrium. These 17 markers were tested for amplification in eight congeneric species with varying success; 98% amplification in Chelodina burrungandjii, 72% in C. canni, 38% in C. expansa, 58% in C. longicollis, 67% in C. mccordi, 73% in C. oblonga, 81% in C. parkeri, and 68% in C. pritchardi. These microsatellite markers will be useful for population assignment, gene flow, mating systems and hybridization studies in the genus Chelodina.
Expressed sequence tag‐derived polymorphic SSR markers for Fucus serratus and amplification in other species of Fucus
Authors: COYER J. A. , HOARAU G. , BESZTERI B. , PEARSON G. , OLSEN J. L.
Published: January 2009

The seaweed genus Fucus is a dominant component of intertidal shores throughout the North Atlantic and North Pacific and has been the focus of considerable developmental, ecological, and evolutionary research for the past century. Here, we present details of 21 expressed sequence tag‐derived simple sequence repeat markers (microsatellites). All 21 were polymorphic for F. serratus, which also display considerable cross‐reactivity with the sister species F. distichus (18) and the more distantly related F. vesiculosus (13), and F. spiralis (5).
Subspecies‐informative SNP assays for evaluating introgression between native golden trout and introduced rainbow trout
Authors: STEPHENS MOLLY R. , CLIPPERTON NEIL W. , MAY BERNIE
Published: January 2009

We characterize 20 single nucleotide polymorphism assays for evaluating hybridization between native golden trout subspecies (Oncorhynchus mykiss aguabonita and O. m. whitei) and introduced rainbow trout strains. These assays utilize the 5′‐nuclease reaction, facilitating high‐throughput genotyping of many individuals and making them useful in quantifying and monitoring introgression and potentially applicable to studies of other O. mykiss groups. Minor allele frequency differentials (δq) among native and introduced rainbow groups ranged from 0 to 1, with an average differential of 0.75 for both subspecies aguabonita and whitei relative to the hatchery rainbow trout strain.
Development of nine polymorphic microsatellite markers for the phytoparasitic nematode Xiphinema index, the vector of the grapevine fanleaf virus
Authors: VILLATE L. , ESMENJAUD D. , COEDEL S. , PLANTARD O.
Published: January 2009

We report isolation, characterization and cross‐species amplification of nine microsatellite loci from the phytoparasitic nematode Xiphinema index, the vector of grapevine fanleaf virus. Levels of polymorphism were evaluated in 62 individuals from two X. index populations. The number of alleles varies between two and 10 depending on locus and population. Observed heterozygosity on loci across both populations varied from 0.32 to 0.857 (mean 0.545). The primers were tested for cross‐species amplification in three other species of phytoparasitic nematodes of the Xiphinema genus. These nine microsatellite loci constitute valuable markers for population genetics and phylogeographical studies of X. index.
Isolation of microsatellite loci from expressed sequence tag library of Puccinia striiformis f. sp. tritici
Authors: CHEN C. Q. , ZHENG W. M. , BUCHENAUER H. , HUANG L. L. , LU N. H. , KANG Z. S.
Published: January 2009

We described twenty polymorphic microsatellite loci derived from the expressed sequence tags of Puccinia striiformis f. sp. tritici, which causes yellow rust disease on wheat. The numbers of alleles range from two to six and eight microsatellite loci show significant similarities to known genes. Observed and expected heterozygosities ranged from 0.12 to 0.78 and from 0.24 to 0.87, respectively.
Isolation and characterization of 19 polymorphic microsatellite DNA markers in the Japanese brown frog (Rana japonica)
Authors: KOIZUMI NORIYUKI , WATABE KEIJI , MORI ATSUSHI , TAKEMURA TAKESHI
Published: January 2009

We report the isolation and characterization of 19 polymorphic microsatellite DNA markers in the Japanese brown frog (Rana japonica). These markers were tested in 24 individuals each collected from three distinct populations in Ichikai‐machi, Tochigi Prefecture. The number of observed alleles per locus ranged from 3 to 24 across all populations, and the values of observed and expected heterozygosities ranged from 0.130 to 1 and from 0.125 to 0.941, respectively. These microsatellite loci will be useful for investigating the intraspecific genetic variation and population structure of this species.
Isolation of highly polymorphic microsatellite loci for a species with a large genome size: sharp‐ribbed salamander (Pleurodeles waltl)
Authors: VAN DE VLIET MIRJAM S. , DIEKMANN ONNO E. , SERRÃO ESTER A. , BEJA PEDRO
Published: January 2009

Fourteen highly polymorphic microsatellite markers were developed and characterized for the sharp‐ribbed salamander, Pleurodeles waltl. Isolating microsatellites with more than 12 single repeat type units was only successful for a tetranucleotide repeat (ATAG). Compared to microsatellite libraries constructed simultaneously for two anuran amphibian species, a greater number of primer pairs designed for P. waltl had to be discarded, due to consistent amplification problems. Low amplification success rate for P. waltl may be due to its larger genome size. Consequently, to avoid nonspecific binding and to increase amplification success, polymerase chain reaction programmes with touchdown cycles were used. For 14 microsatellite markers, amplification was successful and consistent with number of alleles and expected heterozygosity ranging from seven to 22 and from 0.79 to 0.94, respectively. All 14 microsatellite markers will be extremely useful for metapopulation studies of this unique amphibian species.
Characterization of polymorphic microsatellite markers for the green turtle (Chelonia mydas)
Authors: DUTTON PETER H. , FREY AMY
Published: January 2009

We describe primers and polymerase chain reaction conditions to amplify 12 microsatellite loci from the green turtle (Chelonia mydas), including one dinucleotide, four trinucleotide and seven tetranucleotide loci. The primers were tested on 78 individuals from a Pacific population nesting in the Hawaiian Islands. The primer pairs developed in this study yielded an average of 8.33 alleles per locus (range of 3–15 alleles), an average observed heterozygosity of 0.668 (range 0.309–0.910), and an average polymorphic information content of 0.647 (range 0.287–0.894).
Characterization of polymorphic microsatellite loci in the endangered Miho spine loach (Iksookimia choii) and cross‐species amplification within the Cobitidae family
Authors: BANG IN‐CHUL , KIM WOO‐JIN , LEE IL‐RO
Published: January 2009

The Miho spine loach (Iksookimia choii) is an endangered freshwater fish endemic to Korea. To aid a conservation programme for this species, 20 polymorphic microsatellite loci were isolated and characterized in 67 individuals from three populations. The number of alleles per locus ranged from 8 to 33. Within populations, the observed and expected heterozygosities ranged from 0.59 to 1.00 and from 0.56 to 0.96, respectively, indicating its usefulness in population genetics studies. Cross‐species amplification of the loci was tested in six species of the family Cobitidae, and 18 loci were amplified in two or more related species.
Development and characterization of 16 polymorphic microsatellite loci in the Tibet endemic fish Glyptosternum maculatum
Authors: GUO B. Y. , XIE C. X. , SHAN L. , ZHANG Y. , WANG H. L. , XIONG D. M. , LIU H. P. , JI Q.
Published: January 2009

Sixteen novel polymorphic microsatellite loci were developed and characterized in Glyptosternum maculatum, an endemic fish inhabiting the Yarlung Zangbo River valley, Tibet, China. These loci were developed after the enrichment of microsatellite‐containing genomic DNA fragments with a subtractive hybridization protocol. The polymorphism of these loci was analysed with 36 individuals representing a geographical population. The number of alleles found at these loci ranged from 3 to 10. The observed heterozygosity and expected heterozygosity ranged from 0 to 1 and from 0.31 to 0.75, respectively. These microsatellite loci will certainly facilitate the determination of the genetic structure of G. maculatum and the conservation of its genetic resource.
Polymorphic microsatellite markers for the striped skunk, Mephitis mephitis, and other mephitids
Authors: DRAGOO JERRY W. , COAN KATHRYN E. , MOORE KRISTIN A. , HENKE SCOTT E. , FLEISCHER ROBERT C. , WISELY SAMANTHA M.
Published: January 2009

We report 10 polymorphic microsatellite loci primers developed for striped skunks (Mephitis mephitis), a widespread mesocarnivore in North America. Numbers of alleles in these loci ranged from seven to 14 and the observed heterozygosity ranged from 0.76 to 1.0. These primers will be useful for studying population dynamics of skunks where rabies is endemic and will be useful to estimate genetic relatedness among females sharing winter dens. Most of these primers amplify across species within the Mephitidae.
Molecular methods for arthropod bloodmeal identification and applications to ecological and vector‐borne disease studies
Authors: KENT REBEKAH J.
Published: January 2009

DNA‐based methods have greatly enhanced the sensitivity and specificity of hematophagous arthropod bloodmeal identification. A variety of methods have been applied to study the blood‐feeding behaviour of mosquitoes, ticks, black flies and other blood‐feeding arthropods as it relates to host–parasite interactions and pathogen transmission. Overviews of the molecular techniques used for bloodmeal identification, their advantages, disadvantages and applications are presented for DNA sequencing, group‐specific polymerase chain reaction primers, restriction fragment length polymorphism, real‐time polymerase chain reaction, heteroduplex analysis, reverse line‐blot hybridization and DNA profiling. Technical challenges to bloodmeal identification including digestion and analysis of mixed bloodmeals are discussed. Analysis of bloodmeal identification results remains a challenge to the field, particularly with regard to incorporation of vertebrate census and ecology data. Future research directions for molecular analysis of arthropod bloodmeals are proposed.
Isolation and characterization of microsatellites in the harlequin ladybird, Harmonia axyridis (Coleoptera, Coccinellidae), and cross‐species amplification within the family Coccinellidae
Authors: LOISEAU ANNE , MALAUSA THIBAUT , LOMBAERT ERIC , MARTIN JEAN‐FRANÇOIS , ESTOUP ARNAUD
Published: January 2009

A total of 18 microsatellite DNA loci were isolated and characterized from the harlequin ladybird, Harmonia axyridis (Coleoptera: Coccinellidae). We optimized a multiplex panel consisting of two polymerase chain reactions, allowing the genotyping of all loci. The number of alleles and heterozygosity observed at each locus ranged from 1 to 12 and from 0 to 100%, respectively. After Bonferroni correction for multiple tests, none of the loci deviated significantly from Hardy–Weinberg equilibrium and there was no indication of significant linkage disequilibrium among pairs of loci. Successful cross‐species amplification was obtained for only three of the seven tested species of Coccinellidae.
Sphingolipid‐mediated restoration of Mitf expression and repigmentation in vivo in a mouse model of hair graying
Authors: Saha Bidisha , Singh Suman Kumar , Mallick Shampa , Bera Rabindranath , Datta Pijush K. , Mandal Mriganka , Roy Syamal , Bhadra Ranjan
Published: January 2009

Recent advances in the identification and characterisation of stem cell populations has led to substantial interest in understanding the precise triggers that would operate to induce activation of quiescent stem cells. Melanocyte stem cells (MSCs) reside in the bulge region of the hair follicles and are characterised by reduced expression of the microphthalmia‐associated transcription factor (Mitf) and its target genes implicated in differentiation. Vitiligo is characterised by progressive destruction of differentiated melanocytes. However, therapies using UV irradiation therapy can induce a degree of repigmentation, suggesting that MSCs may be activated. As Mitf is implicated in control of proliferation, we have explored the possibility that inducing Mitf expression via lipid‐mediated activation of the p38 stress‐signalling pathway may represent a re‐pigmentation strategy. Here we have isolated from placental extract a C18:0 sphingolipid able to induce Mitf and tyrosinase expression via activation of the p38 stress‐signalling pathway. Strikingly, in age‐onset gray‐haired C57BL/6J mice that exhibit decaying Mitf expression, topical application of placental sphingolipid leads to increased Mitf in follicular melanocytes and fresh dense black hair growth. The results raise the possibility that lipid‐mediated activation of the p38 pathway may represent a novel approach to an effective vitiligo therapy.
Canadian Narcotics Legislation, 1908–1923: A Conflict Model Interpretation
Authors: Cook Shirley J.
Published: January 2009

La legislation canadienne sur les narcotiques s'est developpee en tant que processus politique et elle traduisit les differenciations d'alors dans l'exercice du pouvoir. L'analyse met en relief les positions de subordination et de superordination definies par la loi et les interets incompatibles des legislateurs et des deviants. Les donnees provenant du Hansard et d'autres sources complementaires durant les annees 1908–1923, mettent en lumiere comment les positions de classe, les croyances culturelles et I'hostilite interethnique ont accentue le contenu punitif de la legislation. Le processus politique, dans une culture differente, aurait peut‐etre produit une loi moins coercitive et moins repressive. De la meme maniere, la loi n'aurait peut‐etre pas ete mise en force avec tant de zele et d'energie dans un climat moral et politique different.

Canadian narcotics legislation developed as a political process and reflected current power differentials. The analysis emphasizes the superordinate‐subordinate relationship defined by the law and the incompatible interests of the rule‐makers and deviants. The data from Hansard and elsewhere during the years 1908–1923 show how the stratification order, cultural beliefs and interracial hostility increased the punitive content of the legislation. The political process in a different culture and social structure might have produced a less coercive and repressive law. In a different political and moral climate the law might not have been enforced so energetically.
Effect of Benazepril on Survival and Cardiac Events in Dogs with Asymptomatic Mitral Valve Disease: A Retrospective Study of 141 Cases
Authors: Pouchelon J.‐L. , Jamet N. , Gouni V. , Tissier R. , Serres F. , Carlos Sampedrano C. , Castaignet M. , Lefebvre H. P. , Chetboul V.
Published: January 2009

Background:Angiotensin‐converting enzyme inhibitors (ACEIs) improve quality of life and extend the life span of dogs with naturally acquired ISACHC class II‐III congestive heart failure (CHF). However, their effects on asymptomatic heart disease remain controversial.

Hypothesis:Benazepril (BNZ), an ACEI, could have beneficial effects at the asymptomatic stage of degenerative mitral valve disease (MVD).

Animals:Dogs with ISACHC class Ia MVD and moderate‐to‐severe mitral regurgitation (MR) assessed by the color Doppler mapping technique at entry (Day 0) were retrospectively included.

Methods:Dogs were assigned to the treated group (BNZ group) if they received BNZ (and no other cardiac medication) from Day 0 or to the untreated group (UT group) if they did not receive any cardioactive treatment until occurrence of CHF.

Results:A total of 141 dogs were included in the study, 66 in the BNZ group (dosage: 0.30 ± 0.13 mg/kg) and 75 in the UT group. In the population (n = 93) including all breeds except Cavalier (CKC) and King Charles Spaniels (KC), median survival time to all causes of death in the BNZ group (n = 34, 3.3 years) was significantly longer than in the UT group (n = 59, 1.9 years) as was time to cardiac event (P< .05). Conversely, no effect of the BNZ treatment was observed in the CKC and KC population.

Conclusions and Clinical Relevance:BNZ had beneficial effects in asymptomatic dogs other than CKC and KC affected by MVD with moderate‐to‐severe MR. Breed distribution should be taken into account for interpretation of clinical trials performed in dogs with cardiac disease.
Association between Cough and Cytology of Bronchoalveolar Lavage Fluid and Pulmonary Function in Horses Diagnosed with Inflammatory Airway Disease
Authors: Bedenice D. , Mazan M.R. , Hoffman A.M.
Published: January 2009

Background:The objective of this study was to examine the relationship between chronic cough, pulmonary mechanical function, bronchial hyper‐responsiveness, and peripheral airway inflammation in adult performance horses with nonseptic inflammatory airway disease (IAD).

Hypothesis:We hypothesized that horses with a presenting complaint of cough have a higher percentage of inflammatory cells on bronchoalveolar lavage (BAL), greater airway obstruction, and airway hyper‐responsiveness (AHR) than do horses without cough.

Animals/Sample Population:Adult performance horses (n = 137) referred for evaluation of signs of IAD including cough or exercise intolerance (university hospital patients) and BAL samples (n = 142) taken in first opinion practice.

Methods:A descriptive, retrospective cohort analysis was performed to evaluate the association between chronic cough and pulmonary mechanical function, AHR, BAL cytology, patient signalment, and comorbid features (multivariable logistic regression).

Results:Cough was significantly more prevalent in horses >7 years, and best characterized by a high BAL neutrophil count (>5%) and nasal discharge. Lung function mechanics, abnormal thoracic auscultation, and exercise intolerance did not retain statistical significance in the logistical regression analysis of cough. Although AHR was not related to neutrophilic airway inflammation (BAL neutrophils >5%), it was significantly associated with BAL mast cells >2%.

Conclusions and Clinical Importance:Our data support that neutrophilic airway inflammation may potentiate cough without further changing respiratory mechanical function in IAD. In contrast, mast cell release increased AHR without affecting the incidence of cough. Cough may be used as an indicator of neutrophilic airway inflammation in the presence of low‐grade nonseptic respiratory disease.