Molecular Phylogeny of Microsporidians with Particular Reference to Species that Infect the Muscles of Fish

Ribosomal DNA from eight species of microsporidians infecting fish have been sequenced. Seven of these species infect the skeletal muscle of fish ( Pleistophora spp.) and one species infects migratory mesenchyma cells ( Glugea anomala ). These sequences, in addition to other available microsporidian rDNA sequences from a broad range of host taxa, have been used in phylogenetic analysis. This analysis revealed that muscle-infecting microsporidians from fish are a polyphyletic group, indicating that characters supposed to be important in the classification of the genus Pleistophora have to be re-evaluated. One character that probably has a polyphyletic origin is the amorphous coat, which has been extensively used in the definition of this genus. Furthermore, our results showed that the insect parasitizing Pleistophora spp. are not related to the true pleistophorans parasitic in skeletal muscle of fish. Phylogenetic analysis of small subunit rDNA sequences revealed disagreements between the molecular phylogeny and classifications based upon ultrastruclure. Many of the morphological characters claimed to be important in microsporidian classifications appeared to have arisen several times during evolution: for example, the diplokaryon and sporophorous vesicles.     

Pre-Analytical Sample Quality: Metabolite Ratios as an Intrinsic Marker for Prolonged Room Temperature Exposure of Serum Samples

Advances in the “omics” field bring about the need for a high number of good quality samples. Many omics studies take advantage of biobanked samples to meet this need. Most of the laboratory errors occur in the pre-analytical phase. Therefore evidence-based standard operating procedures for the pre-analytical phase as well as markers to distinguish between ‘good’ and ‘bad’ quality samples taking into account the desired downstream analysis are urgently needed. We studied concentration changes of metabolites in serum samples due to pre-storage handling conditions as well as due to repeated freeze-thaw cycles. We collected fasting serum samples and subjected aliquots to up to four freeze-thaw cycles and to pre-storage handling delays of 12, 24 and 36 hours at room temperature (RT) and on wet and dry ice. For each treated aliquot, we quantified 127 metabolites through a targeted metabolomics approach. We found a clear signature of degradation in samples kept at RT. Storage on wet ice led to less pronounced concentration changes. 24 metabolites showed significant concentration changes at RT. In 22 of these, changes were already visible after only 12 hours of storage delay. Especially pronounced were increases in lysophosphatidylcholines and decreases in phosphatidylcholines. We showed that the ratio between the concentrations of these molecule classes could serve as a measure to distinguish between ‘good’ and ‘bad’ quality samples in our study. In contrast, we found quite stable metabolite concentrations during up to four freeze-thaw cycles. We concluded that pre-analytical RT handling of serum samples should be strictly avoided and serum samples should always be handled on wet ice or in cooling devices after centrifugation. Moreover, serum samples should be frozen at or below -80°C as soon as possible after centrifugation.     

Selectivity is species-dependent: Characterization of standard agonists and antagonists at human,rat, and mouse adenosine receptors

Adenosine receptors (ARs) have emerged as new drug targets. The majority of data on affinity/potency and selectivity of AR ligands described in the literature has been obtained for the human species. However, preclinical studies are mostly performed in mouse or rat, and standard AR agonists and antagonists are frequently used for studies in rodents without knowing their selectivity in the investigated species. In the present study, we selected a set of frequently used standard AR ligands, 8 agonists and 16 antagonists, and investigated them in radioligand binding studies at all four AR subtypes, A₁, A_2A, A_2B, and A₃, of three species, human, rat, and mouse. Recommended, selective agonists include CCPA (for A₁AR of rat and mouse), CGS-21680 (for A_2A AR of rat), and Cl-IB-MECA (for A₃AR of all three species). The functionally selective partial A_2B agonist BAY60-6583 was found to additionally bind to A₁ and A₃AR and act as an antagonist at both receptor subtypes. The antagonists PSB-36 (A₁), preladenant (A_2A), and PSB-603 (A_2B) displayed high selectivity in all three investigated species. MRS-1523 acts as a selective A₃AR antagonist in human and rat, but is only moderately selective in mouse. The comprehensive data presented herein provide a solid basis for selecting suitable AR ligands for biological studies.

Electronic supplementary material

The online version of this article (doi:10.1007/s11302-015-9460-9) contains supplementary material, which is available to authorized users.     

Do Parents Meet Adolescents’ Monitoring Standards? Examination of the Impact on Teen Risk Disclosure and Behaviors if They Don’t

In this study, we examined how adolescents compare monitoring efforts by their parents to those of a "good parent" standard and assessed the impact of these comparisons on adolescent self-disclosure and risk behavior and their perceptions of their parents'' monitoring knowledge. Survey responses from 519 adolescents (12–17 years) at baseline of a larger, longitudinal study examining parental monitoring and adolescent risk were examined. Adolescents’ “good parent comparisons” differed greatly by monitoring areas (e.g., telephone use, health, money); however, between 5.5% and 25.8% of adolescents believed their parents needed to monitor their activities more than they currently were monitoring. Alternatively, between 8.5% and 23.8% of adolescents believed their parents needed to monitor their activities less often. These perceptions significantly distinguished adolescents in terms of their level of disclosure, perceived monitoring knowledge, and risk involvement. Adolescents who viewed their parents as needing to monitor more were less likely to disclose information to their parents (p<.001), less likely to perceive their parents as having greater monitoring knowledge (p<.001), and more likely to be involved in a risk behaviors (p<.001) than adolescents who perceived their parents needed no change. Adolescent disclosure to a parent is a powerful predictor of adolescent risk and poor health outcomes. These findings demonstrate that adolescents'' comparisons of their parents'' monitoring efforts can predict differences in adolescent disclosure and future risk. Obtaining adolescent "good parent" comparisons may successfully identify intervention opportunities with the adolescent and parent by noting the areas of need and direction of monitoring improvement.     

The European General Practice Research Network Presents the Translations of Its Comprehensive Definition of Multimorbidity in Family Medicine in Ten European Languages

Background

Multimorbidity, according to the World Health Organization, exists when there are two or more chronic conditions in one patient. This definition seems inaccurate for the holistic approach to Family Medicine (FM) and long-term care. To avoid this pitfall the European General Practitioners Research Network (EGPRN) designed a comprehensive definition of multimorbidity using a systematic literature review.

Objective

To translate that English definition into European languages and to validate the semantic, conceptual and cultural homogeneity of the translations for further research.

Method

Forward translation of the EGPRN’s definition of multimorbidity followed by a Delphi consensus procedure assessment, a backward translation and a cultural check with all teams to ensure the homogeneity of the translations in their national context. Consensus was defined as 70% of the scores being higher than 6. Delphi rounds were repeated in each country until a consensus was reached

Results

229 European medical expert FPs participated in the study. Ten consensual translations of the EGPRN comprehensive definition of multimorbidity were achieved.

Conclusion

A comprehensive definition of multimorbidity is now available in English and ten European languages for further collaborative research in FM and long-term care.     

Serum Cardiac Troponin-I is Superior to Troponin-T as a Marker for Left Ventricular Dysfunction in Clinically Stable Patients with End-Stage Renal Disease

Background

Serum troponin assays, widely used to detect acute cardiac ischemia, might be useful biomarkers to detect chronic cardiovascular disease (CVD). Cardiac-specific troponin-I (cTnI) and troponin-T (cTnT) generally detect myocardial necrosis equally well. In dialysis patients however, serum cTnT levels are often elevated, unlike cTnI levels. The present study aims to elucidate the associations of cTnI and cTnT with CVD in clinically stable dialysis patients.

Methods

Troponin levels were measured using 5^th generation hs-cTnT assays (Roche) and STAT hs-cTnI assays (Abbott) in a cohort of dialysis patients. Serum troponin levels were divided into tertiles with the lowest tertile as a reference value. Serum troponins were associated with indicators of CVD such as left ventricular mass index (LVMI), left ventricular ejection fraction (LVEF) and the presence of coronary artery disease (CAD). Associations were explored using regression analysis.

Results

We included 154 consecutive patients, 68±7 years old, 77% male, 70% hemodialysis. Median serum cTnT was 51ng/L (exceeding the 99^th percentile of the healthy population in 98%) and median serum cTnI was 13ng/L (elevated in 20%). A high cTnI (T3) was significantly associated with a higher LVMI (Beta 31.60; p=0.001) and LVEF (Beta -4.78; p=0.005) after adjusting for confounders whereas a high serum cTnT was not. CAD was significantly associated with a high cTnT (OR 4.70 p=0.02) but not with a high cTnI. Unlike cTnI, cTnT was associated with residual renal function (Beta:-0.09; p=0.006).

Conclusion

In the present cohort, serum cTnI levels showed a stronger association with LVMI and LVEF than cTnT. However, cTnT was significantly associated with CAD and residual renal function, unlike cTnI. Therefore, cTnI seems to be superior to cTnT as a marker of left ventricular dysfunction in asymptomatic dialysis patients, while cTnT might be better suited to detect CAD in these patients.     

Effects of the new adiponectin paralogous protein CTRP-3 and of LPS on cytokine release from monocytes of patients with type 2 diabetes mellitus

Aims/Hypothesis: It was the aim to investigate the hypothesis that the new C1q/TNF-family member CTRP-3 (C1q/TNF-related protein-3) acts anti-inflammatory in human monocytes from healthy controls and patients with type 2 diabetes mellitus (T2D). Methods: Monocytes were isolated from 20 healthy controls and 30 patients with T2D. IL-6 and TNF concentrations were measured by ELISA. CTRP-3 was expressed in insect cells and used for stimulation experiments. Results: Basal IL-6 and TNF were not different in control and in T2D monocytes. LPS-stimulation (1 μg/ml) significantly (p < 0.001) increased IL-6 and TNF in the supernatants of control and in T2D monocytes to a similar extent. CTRP-3 (1 μg/ml) significantly (p = 0.03) inhibited LPS-induced IL-6 in control monocytes but not in T2D monocytes. TNF upon co-stimulation with LPS and CTRP-3 was significantly (p = 0.012) lower in control than in T2D monocytes. LPS-induced TNF concentration was significantly and positively correlated with serum total cholesterol and LDL cholesterol in T2D patients. Conclusions: CTRP-3 inhibits LPS-induced IL-6 and TNF release. This anti-inflammatory effect is lost in T2D. Serum cholesterol concentration affects the pro-inflammatory potential of LPS to induce TNF release from T2D monocytes in the presence or absence of CTRP-3. CTRP-3 might partly account for the pro-inflammatory state in T2D.     

Actin-binding Protein-1 Interacts with WASp-interacting Protein to Regulate Growth Factor-induced Dorsal Ruffle Formation

Growth factor stimulation induces the formation of dynamic actin structures known as dorsal ruffles. Mammalian actin-binding protein-1 (mAbp1) is an actin-binding protein that has been implicated in regulating clathrin-mediated endocytosis; however, a role for mAbp1 in regulating the dynamics of growth factor–induced actin-based structures has not been defined. Here we show that mAbp1 localizes to dorsal ruffles and is necessary for platelet-derived growth factor (PDGF)-mediated dorsal ruffle formation. Despite their structural similarity, we find that mAbp1 and cortactin have nonredundant functions in the regulation of dorsal ruffle formation. mAbp1, like cortactin, is a calpain 2 substrate and the preferred cleavage site occurs between the actin-binding domain and the proline-rich region, generating a C-terminal mAbp1 fragment that inhibits dorsal ruffle formation. Furthermore, mAbp1 directly interacts with the actin regulatory protein WASp-interacting protein (WIP) through its SH3 domain. Finally, we demonstrate that the interaction between mAbp1 and WIP is important in regulating dorsal ruffle formation and that WIP-mediated effects on dorsal ruffle formation require mAbp1. Taken together, these findings identify a novel role for mAbp1 in growth factor–induced dorsal ruffle formation through its interaction with WIP.     

Time Trends of Helicobacter pylori Resistance to Antibiotics in Children Living in Vienna,Austria

Background: Increase of antibiotic resistance is a worldwide problem. Within the 4 years before the turn of the millennium Helicobacter pylori strains isolated in children living in Vienna, Austria, showed a primary clarithromycin and metronidazole resistance of 20% and 16%, respectively. The aim of this retrospective follow‐up survey was to assess the further development and current antimicrobial resistance status. Methods: Children having undergone upper endoscopy between March 2002 and March 2008 at the same two co‐operating pediatric gastroenterology units which had also been collaborating on the prior assessment were included. H. pylori infection was diagnosed by rapid urease test, histology, and culture. If the latter was positive, susceptibility testing to amoxicillin, clarithromycin and metronidazole by E‐test followed. From March 2004 onwards, susceptibility to levofloxacin, tetracycline and rifampin was additionally assessed. Results: Out of 897 children, 153 had a proven infection with H. pylori and no history of prior eradication treatment. Their median age was 11.5 years (range 0.5–20.9 years). Primary resistance to clarithromycin and metronidazole were 34% and 22.9%, respectively; dual resistance was found in 9.8% of the strains; 0.9% was resistant to tetracycline and rifampin, respectively. No case of amoxicillin resistance was detected. The only independent risk factor for clarithromycin resistance turned out to be the origin of a child from Austrian parents. Conclusions: In the last decade, the rate of primary resistance of H. pylori to clarithromycin continued to rise. No significant change was found regarding primary resistance to metronidazole or dual resistance to metronidazole and clarithromycin, respectively.     

Spatial dynamics of Norwegian tetraonid populations

Different species in a given site or population of a given species in different sites may fluctuate in synchrony if they are affected similarly by factors such as spatially autocorrelated climate, predation, or by dispersal between populations of one species. We used county wise time series of hunting bag records of four Norwegian tetraonid species covering 24 years to examine patterns of interspecific and intraspecific synchrony. We estimated synchrony at three spatial scales; national, regional (consisting of counties with similar climate), and county level. Ecologically related species with overlapping distributions exhibited strong synchrony across Norway, but there was much variation between the different regions and counties. Regions with a long coastline to both the North Sea and the Norwegian Ocean exhibited an overall stronger synchrony than those consisting of more continental areas. Intraspecific synchrony was generally low across all counties, but stronger synchrony between counties within regions defined by climatic conditions. Synchrony was negatively related to distance between populations in three of four species. Only the synchrony in willow ptarmigan showed a clear negative relationship with distance, while the other species had both strong positive and negative correlations at short distances. Strong interspecific synchrony between some species pairs within regions and weak intraspecific synchrony across counties within regions suggest a stronger synchronizing effect from environmental factors such as weather or predation and less effect from dispersal. Our results suggest that the complete tetraonid community is structured by environmental factors affecting the different species similarly and causes widespread interspecific synchrony. Local factors affecting the population dynamics nevertheless frequently forces neighbouring populations out of phase.