Development of a heptaplex PCR assay for identification of Staphylococcus aureus and CoNS with simultaneous detection of virulence and antibiotic resistance genes

Background Staphylococcal toxicity and antibiotic resistance (STAAR) have been menacing public health. Although vancomycin-resistant Staphylococcus aureus (VRSA) is currently not as widespread as methicillin-resistant S. aureus (MRSA), genome evolution of MRSA into VRSA, including strains engineered within the same patient under anti-staphylococcal therapy, may build up to future public health concern. To further complicate diagnosis, infection control and anti-microbial chemotherapy, non-sterile sites such as the nares and the skin could contain both S. aureus and coagulase-negative staphylococci (CoNS), either of which could harbour mecA the gene driving staphylococcal methicillin-resistance and required for MRSA-VRSA evolution. Results A new heptaplex PCR assay has been developed which simultaneously detects seven markers for: i) eubacteria (16S rRNA), ii) Staphylococcus genus (tuf), iii) Staphylococcus aureus (spa), iv) CoNS (cns), v) Panton-Valentine leukocidin (pvl), vi) methicillin resistance (mecA), and vii) vancomycin resistance (vanA). Following successful validation using 255 reference bacterial strains, applicability to analyse clinical samples was evaluated by direct amplification in spiked blood cultures (n = 89) which returned 100 % specificity, negative and positive predictive values. The new assay has LoD of 1.0x103 CFU/mL for the 16S rRNA marker and 1.0x104 CFU/mL for six other markers and completes cycling in less than one hour. Conclusion The speed, sensitivity (100 %), NPV (100 %) and PPV (100 %) suggest the new heptaplex PCR assay could be easily integrated into a routine diagnostic microbiology workflow. Detection of the cns marker allows for unique identification of CoNS in mono-microbial and in poly-microbial samples containing mixtures of CoNS and S. aureus without recourse to the conventional elimination approach which is ambiguous. In addition to the SA-CoNS differential diagnostic essence of the new assay, inclusion of vanA primers will allow microbiology laboratories to stay ahead of the emerging MRSA-VRSA evolution. To the best of our knowledge, the new heptaplex PCR assay is the most multiplexed among similar PCR-based assays for simultaneous detection of STAAR

Calibration : the Achilles heel of predictive analytics

Acknowledgements This work was developed as part of the international STRengthening Analytical Thinking for Observational Studies (STRATOS) initiative. The objective of STRATOS is to provide accessible and accurate guidance in the design and analysis of observational studies (http://stratos-initiative.org/). Members of the STRATOS Topic Group ‘Evaluating diagnostic tests and prediction models’ are (alphabetically) Patrick Bossuyt, Gary S. Collins, Petra Macaskill, David J. McLernon, Karel G.M. Moons, Ewout W. Steyerberg, Ben Van Calster, Maarten van Smeden, and Andrew Vickers. Funding This work was funded by the Research Foundation – Flanders (FWO; grant G0B4716N) and Internal Funds KU Leuven (grant C24/15/037). The funders had no role in study design, data collection, data analysis, interpretation of results, or writing of the manuscript. Contributions All authors conceived of the study. BVC drafted the manuscript. All authors reviewed and edited the manuscript and approved the final version.Peer reviewedPublisher PD

Examining the generalizability of research findings from archival data

This initiative examined systematically the extent to which a large set of archival research findings generalizes across contexts. We repeated the key analyses for 29 original strategic management effects in the same context (direct reproduction) as well as in 52 novel time periods and geographies; 45% of the reproductions returned results matching the original reports together with 55% of tests in different spans of years and 40% of tests in novel geographies. Some original findings were associated with multiple new tests. Reproducibility was the best predictor of generalizability-for the findings that proved directly reproducible, 84% emerged in other available time periods and 57% emerged in other geographies. Overall, only limited empirical evidence emerged for context sensitivity. In a forecasting survey, independent scientists were able to anticipate which effects would find support in tests in new samples.info:eu-repo/semantics/publishedVersio

Patients treated for hyperthyroidism are at increased risk of becoming obese: findings from a large prospective secondary care cohort

YesBackground: The most commonly reported symptom of hyperthyroidism is weight loss; successful treatment increases weight. Weight gain faced by patients with hyperthyroidism is widely considered a simple reaccumulation of premorbid weight, whereas many patients feel there is a significant weight “overshoot” attributable to the treatment. We aimed to establish if weight gain seen following treatment for hyperthyroidism represents replenishment of premorbid weight or “overshoot” beyond expected regain and, if there is excessive weight gain, whether this is associated with the applied treatment modality. Methods: We calculated the risk of becoming obese (body mass index [BMI] >30 kg/m2) following treatment for hyperthyroidism by comparing BMI of 1373 patients with overt hyperthyroidism seen in a secondary care setting with the age- and sex-matched background population (Health Survey for England, 2007–2009). Next, we investigated the effect of treatment with an antithyroid drug (ATD) alone in regard to ATD with radioactive iodine (131I) therapy. We modeled the longitudinal weight data in relation to the treatment pathway to thyroid function and the need for long-term thyroxine replacement. Results: During treatment of hyperthyroidism, men gained 8.0 kg (standard deviation ±7.5) and women 5.5 kg (±6.8). At discharge, there was a significantly increased risk of obesity in male (odds ratio = 1.7 [95% confidence interval 1.3–2.2], p 10 mIU/L; 0.5 kg, 0.3–0.7, p < 0.001) or free thyroxine (fT4) was reduced (fT4 ≤ 10 pmol/L (0.8 ng/dL); 0.3 kg, 0.1–0.4, p < 0.001) during follow-up. Initiation of levothyroxine was associated with further weight gain (0.4 kg, 0.2–0.6, p < 0.001) and the predicted excess weight gain in 131I-induced hypothyroidism was 1.8 kg. Conclusions: Treatment for hyperthyroidism is associated with significant risks of becoming obese. 131I treatment and subsequent development of hypothyroidism were associated with small but significant amounts of excess weight gain compared with ATD alone. We advocate that the discussion over the weight “overshoot” risk forms part of the individualized treatment decision-making process

Examining the generalizability of research findings from archival data

This initiative examined systematically the extent to which a large set of archival research findings generalizes across contexts. We repeated the key analyses for 29 original strategic management effects in the same context (direct reproduction) as well as in 52 novel time periods and geographies; 45% of the reproductions returned results matching the original reports together with 55% of tests in different spans of years and 40% of tests in novel geographies. Some original findings were associated with multiple new tests. Reproducibility was the best predictor of generalizability—for the findings that proved directly reproducible, 84% emerged in other available time periods and 57% emerged in other geographies. Overall, only limited empirical evidence emerged for context sensitivity. In a forecasting survey, independent scientists were able to anticipate which effects would find support in tests in new samples.</p

Masitinib (AB1010), a Potent and Selective Tyrosine Kinase Inhibitor Targeting KIT

International audienceBackground: The stem cell factor receptor, KIT, is a target for the treatment of cancer, mastocytosis, and inflammatory diseases. Here, we characterise the in vitro and in vivo profiles of masitinib (AB1010), a novel phenylaminothiazole-type tyrosine kinase inhibitor that targets KIT. Methodology/Principal Findings: In vitro, masitinib had greater activity and selectivity against KIT than imatinib, inhibiting recombinant human wild-type KIT with an half inhibitory concentration (IC50) of 200 ± 40 nM and blocking stem cell factor-induced proliferation and KIT tyrosine phosphorylation with an IC50 of 150 ± 80 nM in Ba/F3 cells expressing human or mouse wild-type KIT. Masitinib also potently inhibited recombinant PDGFR and the intracellular kinase Lyn, and to a lesser extent, fibroblast growth factor receptor 3. In contrast, masitinib demonstrated weak inhibition of ABL and c-Fms and was inactive against a variety of other tyrosine and serine/threonine kinases. This highly selective nature of masitinib suggests that it will exhibit a better safety profile than other tyrosine kinase inhibitors; indeed, masitinib-induced cardiotoxicity or genotoxicity has not been observed in animal studies. Molecular modelling and kinetic analysis suggest a different mode of binding than imatinib, and masitinib more strongly inhibited degranulation, cytokine production, and bone marrow mast cell migration than imatinib. Furthermore, masitinib potently inhibited human and murine KIT with activating mutations in the juxtamembrane domain. In vivo, masitinib blocked tumour growth in mice with subcutaneous grafts of Ba/F3 cells expressing a juxtamembrane KIT mutant. Conclusions: Masitinib is a potent and selective tyrosine kinase inhibitor targeting KIT that is active, orally bioavailable in vivo, and has low toxicit

Revisiting the Dexamethasone Suppression Test in unipolar major depression: an exploratory study

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