Flotation—A New Method to Circumvent PCR Inhibitors in the Diagnosis of Lawsonia intracellularis

The obligate intracellular bacterium Lawsonia intracellularis causes enteritis and poor growth in weaned pigs. Cultivation is difficult and diagnosis ante mortem is mainly based on techniques such as polymerase chain reaction. However, false negative results caused by the presence of PCR-inhibitory factors constitute a problem. This study aimed to develop and evaluate a new technique, flotation, to separate L. intracellularis from inhibitors in faeces prior to PCR. The technique was evaluated by comparison to two previously evaluated and commonly used methods, preparation by boiling lysate combined with nested PCR and preparation by a commercial kit combined with conventional PCR. Continuous density centrifugation of faecal samples containing L. intracellularis suggested the buoyant density of the microbe to be between 1.064 and 1.077 g/mL. Several flotation setups were tested to achieve optimal separation of the microbe from inhibitors and faecal particles. The finally selected setup floated whole L. intracellularis from the application site at the bottom to the upper part of the gradient while inhibitory components mainly remained in the bottom. PCR was performed directly on material recovered from the upper interphase. The method was evaluated on 116 clinical samples. As compared to sample preparation by boiling combined with nested PCR, fewer samples were inhibited but also fewer positives were identified. In comparison to preparation by a commercial kit combined with conventional PCR, presently used for routine diagnosis, similar results were obtained. However, the new method was comparably faster to perform. The new method, based on flotation of Lawsonia intracellularis combined with conventional PCR, was well suited for routine diagnosis

Genetic Risk Factors in Lupus Nephritis and IgA Nephropathy - No Support of an Overlap

Background: IgA nephropathy (IgAN) and nephritis in Systemic Lupus Erythematosus (SLE) are two common forms of glomerulonephritis in which genetic findings are of importance for disease development. We have recently reported an association of IgAN with variants of TGFB1. In several autoimmune diseases, particularly in SLE, IRF5, STAT4 genes and TRAF1-C5 locus have been shown to be important candidate genes. The aim of this study was to compare genetic variants from the TGFB1, IRF5, STAT4 genes and TRAF1-C5 locus with susceptibility to IgAN and lupus nephritis in two Swedish cohorts. Patients and Methods: We genotyped 13 single nucleotide polymorphisms (SNPs) in four genetic loci in 1252 DNA samples from patients with biopsy proven IgAN or with SLE (with and without nephritis) and healthy age-and sex-matched controls from the same population in Sweden. Results: Genotype and allelic frequencies for SNPs from selected genes did not differ significantly between lupus nephritis patients and SLE patients without nephritis. In addition, haplotype analysis for seven selected SNPs did not reveal a difference for the SLE patient groups with and without nephritis. Moreover, none of these SPNs showed a significant difference between IgAN patients and healthy controls. IRF5 and STAT4 variants remained significantly different between SLE cases and healthy controls. In addition, the data did not show an association of TRAF1-C5 polymorphism with susceptibility to SLE in this Swedish population. Conclusion: Our data do not support an overlap in genetic susceptibility between patients with IgAN or SLE and reveal no specific importance of SLE associated SNPs for the presence of lupus nephritis.Original Publication: Mai Tuyet Vuong, Iva Gunnarsson, Sigrid Lundberg, Elisabet Svenungsson, Lars Wramner, Anders Fernström, Ann-Christine Syvanen, Lieu Thi Do, Stefan H. Jacobson and Leonid Padyukov, Genetic Risk Factors in Lupus Nephritis and IgA Nephropathy - No Support of an Overlap, 2010, PLOS ONE, (5), 5. http://dx.doi.org/10.1371/journal.pone.0010559 Licensee: Public Library of Science (PLoS) http://www.plos.org/</p

Thermohaline instability and rotation-induced mixing. I - Low- and intermediate-mass solar metallicity stars up to the end of the AGB

(abridged) Numerous spectroscopic observations provide compelling evidence for non-canonical processes that modify the surface abundances of low- and intermediate-mass stars beyond the predictions of standard stellar theory. We study the effects of thermohaline instability and rotation-induced mixing in the 1-4 Msun range at solar metallicity. We present evolutionary models by considering both thermohaline and rotation-induced mixing in stellar interior. We discuss the effects of these processes on the chemical properties of stars from the zero age main sequence up to the end of the second dredge-up on the early-AGB for intermediate-mass stars and up to the AGB tip for low-mass stars. Model predictions are compared to observational data for lithium,12C/13C,[N/C],[Na/Fe],16O/17O, and 16O/18O in Galactic open clusters and in field stars with well-defined evolutionary status,as well as in planetary nebulae. Thermohaline mixing simultaneously accounts for the observed behaviour of 12C/13C,[N/C], and lithium in low-mass stars that are more luminous than the RGB bump, and its efficiency is increasing with decreasing initial stellar mass. On the TP-AGB,thermohaline mixing leads to lithium production, although the 7Li yields remain negative. Although the 3He stellar yields are much reduced thanks to this process, we find that solar-metallicity, low-mass stars remain net 3He producers. Rotation-induced mixing is found to change the stellar structure so that in the mass range between \sim 1.5 and 2.2 Msun the thermohaline instability occurs earlier on the red giant branch than in non-rotating models. Finally rotation accounts for the observed star-to-star abundance variations at a given evolutionary status, and is necessary to explain the features of CN-processed material in intermediate-mass stars.Comment: 18 pages, 22 figures, accepted for publication in A&

Genetic variation in the transforming growth factor-β1 gene is associated with susceptibility to IgA nephropathy

Background. There is growing evidence of genetic risk for susceptibility to IgA nephropathy. Among several candidate genes related to immunological regulation in renal tissue, TGFB1 is known to be a contributor to proliferation and the development of fibrosis

Tillstånd och trender för arter och deras livsmiljöer

2015 års upplaga av den svenska rödlistan är den fjärde i ordningen. Den är baserad på IUCN:s rödlistningskriterier och revideras vart femte år. I rödlistan bedöms risken som enskilda arter av djur, växter och svampar löper att försvinna från Sverige. Bedömningen utförs av ArtDatabankens medarbetare i samverkan med över 100 externa experter, indelade i 14 expertkommittéer för olika organismgrupper. Under arbetet med 2015 års rödlista har tillstånd och trender bedömts för 21 600 arter och 1 318 lägre taxa (apomiktiska arter, underarter och varieteter), sammanlagt ca 22 900 taxa. Av de bedömda arterna klassificerades 2 029 som hotade (kategorierna CR, EN och VU) och 4 273 som rödlistade (inkluderar även kategorierna NT, RE och DD). Förhållandet mellan antalet rödlistade och antalet bedömda arter ar 19,8 %, vilket är ungefär samma värde som 2010 och 2005. I denna rapport jämförs antalet och andelen rödlistade arter mellan olika organismgrupper, biotoper, substrat och påverkansfaktorer. Texten ar indelad i en allmän del och åtta kapitel inriktade på olika landskapstyper. Landskapstyperna utgör en grov indelning av landets miljöer enligt följande kategorier: Skog, Jordbrukslandskap, Urbana miljöer, Fjäll, Våtmarker, Sötvatten, Havsstränder och Havsmiljöer. Skogen och jordbrukslandskapet är de artrikaste landskapstyperna med 1 800 respektive 1 400 arter som har en stark anknytning dit, och ytterligare flera hundra arter som förekommer där mer sporadiskt. De faktorer som påverkar flest rödlistade arter i Sverige är skogsavverkning och igenväxning, som båda utgör ett hot mot vardera ca 30 % av de rödlistade arterna. Avverkning minskar arealen av skog där naturliga strukturer och naturlig dynamik upprätthålls, och den orsakar därmed förlust av livsmiljöer. Igenväxning orsakas av ett antal faktorer, bland annat upphörande hävd (bete och slåtter), gödsling, trädplantering och brist på naturliga störningsregimer som t.ex. regelbundna översvämningar kring vattendrag och sjöar. Andra viktiga påverkansfaktorer är fiske, torrläggning av våtmarker, tillbakagång hos värdarter (främst alm och ask som drabbats av invasiva svampsjukdomar), klimatförändringar och konkurrens från invasiva arter. IUCN:s rödlisteindex beräknas för ett urval av de bedömda organismgrupperna. Rödlisteindex visar att skillnaderna mellan rödlistorna från 2000, 2005, 2010 och 2015 är små. Ett par undantag finns dock. Groddjur och stora däggdjur har fått en något förbättrad situation sedan 2000. Totalt förefaller det ändå som att trycket mot Sveriges artstock har förblivit relativt konstant under de senaste 15 åren

Open clusters towards the Galactic center: chemistry and dynamics. A VLT spectroscopic study of NGC6192, NGC6404, NGC6583

In the framework of the study of the Galactic metallicity gradient and its time evolution, we present new high-resolution spectroscopic observations obtained with FLAMES and the fiber link to UVES at VLT of three open clusters (OCs) located within $\sim$7~kpc from the Galactic Center (GC): NGC~6192, NGC~6404, NGC~6583. We also present new orbit determination for all OCs with Galactocentric distances (R$_{\rm{GC}}) \leq$8~kpc and metallicity from high-resolution spectroscopy. We aim to investigate the slope of the inner disk metallicity gradient as traced by OCs and at discussing its implication on the chemical evolution of our Galaxy. We have derived memberships of a group of evolved stars for each clusters, obtaining a sample of 4, 4, and 2 member stars in NGC~6192, NGC~6404, and NGC~6583, respectively. Using standard LTE analysis we derived stellar parameters and abundance ratios for the iron-peak elements Fe, Ni, Cr, and for the $\alpha$-elements Al, Mg, Si, Ti, Ca. We calculated the orbits of the OCs currently located within 8~kpc from the GC, and discuss their implication on the present-time radial location. {The average metallicities of the three clusters are all oversolar: [Fe/H]= $+0.12\pm0.04$ (NGC~6192), $+0.11\pm0.04$ (NGC 6404), $+0.37\pm0.03$ (NGC 6583). They are in qualitative agreement with their Galactocentric distances, being all internal OCs, and thus expected to be metal richer than the solar neighborhood. The abundance ratios of the other elements over iron [X/Fe] are consistent with solar values. The clusters we have analysed, together with other OC and Cepheid data, confirm a steep gradient in the inner disk, a signature of an evolutionary rate different than in the outer disk.Comment: 17 pages, 13 figures, A&A accepted for publicatio

Genetic, serological and biochemical characterization of Leishmania tropica from foci in northern Palestine and discovery of zymodeme MON-307

Background Many cases of cutaneous leishmaniasis (CL) have been recorded in the Jenin District based on their clinical appearance. Here, their parasites have been characterized in depth. Methods Leishmanial parasites isolated from 12 human cases of CL from the Jenin District were cultured as promastigotes, whose DNA was extracted. The ITS1 sequence and the 7SL RNA gene were analysed as was the kinetoplast minicircle DNA (kDNA) sequence. Excreted factor (EF) serotyping and multilocus enzyme electrophoresis (MLEE) were also applied. Results This extensive characterization identified the strains as Leishmania tropica of two very distinct sub-types that parallel the two sub-groups discerned by multilocus microsatellite typing (MLMT) done previously. A high degree of congruity was displayed among the results generated by the different analytical methods that had examined various cellular components and exposed intra-specific heterogeneity among the 12 strains. Three of the ten strains subjected to MLEE constituted a new zymodeme, zymodeme MON-307, and seven belonged to the known zymodeme MON-137. Ten of the 15 enzymes in the profile of zymodeme MON-307 displayed different electrophoretic mobilities compared with the enzyme profile of the zymodeme MON-137. The closest profile to that of zymodeme MON-307 was that of the zymodeme MON-76 known from Syria. Strains of the zymodeme MON-307 were EF sub-serotype A2 and those of the zymodeme MON-137 were either A9 or A9B4. The sub-serotype B4 component appears, so far, to be unique to some strains of L. tropica of zymodeme MON-137. Strains of the zymodeme MON-137 displayed a distinctive fragment of 417 bp that was absent in those of zymodeme MON-307 when their kDNA was digested with the endonuclease RsaI. kDNA-RFLP after digestion with the endonuclease MboI facilitated a further level of differentiation that partially coincided with the geographical distribution of the human cases from which the strains came. Conclusions The Palestinian strains that were assigned to different genetic groups differed in their MLEE profiles and their EF types. A new zymodeme, zymodeme MON-307 was discovered that seems to be unique to the northern part of the Palestinian West Bank. What seemed to be a straight forward classical situation of L. tropica causing anthroponotic CL in the Jenin District might be a more complex situation, owing to the presence of two separate sub-types of L. tropica that, possibly, indicates two separate transmission cycles involving two separate types of phlebotomine sand fly vector

Positive Inotropic Effects by Uridine Triphosphate (UTP) and Uridine Diphosphate (UDP) via P2Y2 and P2Y6 Receptors on Cardiomyocytes and Release of UTP in Man During Myocardial Infarction

The aim of this study was to examine a possible role for extracellular pyrimidines as inotropic factors for the heart. First, nucleotide plasma levels were measured to evaluate whether UTP is released in patients with coronary heart disease. Then, inotropic effects of pyrimidines were examined in isolated mouse cardiomyocytes. Finally, expression of pyrimidine-selective receptors (a subgroup of the P2 receptors) was studied in human and mouse heart, using real time polymerase chain reaction, Western blot, and immunohistochemistry. Venous plasma levels of UTP were increased (57%) in patients with myocardial infarction. In electrically stimulated cardiomyocytes the stable P2Y2/4 agonist UTPγS increased contraction by 52%, similar to β1-adrenergic stimulation with isoproterenol (65%). The P2Y6-agonist UDPγS also increased cardiomyocyte contraction (35%), an effect abolished by the P2Y6-blocker MRS2578. The phospholipase C inhibitor {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 inhibited both the UDPγS and the UTPγS-induced inotropic effect, indicating an IP3-mediated effect via P2Y6 receptors. The P2Y14 agonist UDP-glucose was without effect. Quantification of mRNA with real time polymerase chain reaction revealed P2Y2 as the most abundant pyrimidine receptor expressed in cardiomyocytes from man. Presence of P2Y6 receptor mRNA was detected in both species and confirmed at protein level with Western blot and immunohistochemistry in man. In conclusion, UTP levels are increased in humans during myocardial infarction, giving the first evidence for UTP release in man. UTP is a cardiac inotropic factor most likely by activation of P2Y2 receptors in man. For the first time we demonstrate inotropic effects of UDP, mediated by P2Y6 receptors via an IP3-dependent pathway. Thus, the extracellular pyrimidines (UTP and UDP) could be important inotropic factors involved in the development of cardiac disease