Design and evaluation of a unique RT-qPCR assay for diagnostic quality control assessment that is applicable to pathogen detection in three species of salmonid fish

BACKGROUND: The detection of pathogens at early stages of infection is a key point for disease control in aquaculture. Therefore, accurate diagnostic procedures are a must. Real-time PCR has been a mainstay in diagnostics over the years due to its speed, specificity, sensitivity, reproducibility and throughput; as such, real-time PCR is a target for improvement. Nevertheless, to validate a novel diagnostic tool, correct setup of the assay, including proper endogenous controls to evaluate the quantity and quality of the samples and to detect possible sample degradation, is compulsory. This work aims to design a unique RT-qPCR assay for pathogen detection in the three salmonid species reared in Chile. The assay uses elongation factor 1 alpha as the single endogenous control, thus avoiding the need for multiple endogenous controls, as well as multiple validations and non-comparable quality control parameters. RESULTS: The in vivo and in vitro analyses of samples from Salmo salar, Oncorhynchus mykiss and Oncorhynchus kisutch showed that when primers were accurately selected to target conserved regions of the elongation factor 1 alpha (ELF1α) gene, a single novel RT-qPCR assay yielding similar and reproducible Ct values between the three species could be designed. The opposite occurred when an assay originally designed for Salmo salar was tested in samples from the two species of the genus Oncorhynchus. CONCLUSIONS: Here, we report the design and evaluation of an accurate trans-species RT-qPCR assay that uses the elongation factor 1 alpha (ELF1α) gene as an endogenous control and is applicable for diagnostic purposes in samples obtained from the three salmonid species reared in Chile

Investment case for two-year post university speciality training in family medicine in Tajikistan: how much is needed for continuing and scaling up the improved education of family doctors?

Background: A new two-year Post University Specialty Training (PUST) programme in family medicine was introduced to improve the quality of postgraduate speciality medical education in Tajikistan. Postgraduate education of family doctors (FDs) needs to be urgently scaled up, as 38% of FD positions in Tajikistan remained unfilled in 2018. Moreover, the international financial support for the PUST programme is ending. This investment case assesses the minimum funding needed for the continuation and scale-up of PUST and establishes the rationale for the investment in the light of a recent evaluation. Methods: The costs of the programme were calculated for 2018 and a scale-up forecast made for the period 2019-2023. The impact of the scale-up on the shortage of FDs was assessed. An evaluation using a Multiple Choice Questionnaire and Objective Structured Clinical Examination (OSCE) assessed and compared theoretical knowledge, clinical skills and competencies of PUST trained and conventionally trained FDs. Results: The annual costs of the programme were US$228,000 in 2018. The total investment needed for scaling up PUST from 31 new FDs in 2018 to 100 FD graduates each year by 2023 was US$ 802,000.However, when the retirement of FDs and population growth are considered, the scale-up will result only in maintaining the current level of FDs working and not solve the country's FD shortage. The PUST FDs demonstrated significantly better clinical skills than the conventionally trained interns, scoring 60 and 45% of OSCE points, respectively. Theoretical knowledge showed a similar trend; PUST FDs answered 44% and interns 38% of the questions correctly. Conclusions: The two-year PUST programme has clearly demonstrated it produces better skilled family doctors than the conventional one-year internship, albeit some enduring quality concerns do still prevail. The discontinuation of international support for PUST would be a major setback and risks potentially losing the benefits of the programme for family medicine and also other specialities. To guarantee the supply of adequately trained FDs and address the FD shortage, the PUST should be continued and scaled up. Therefore, it is essential that international support is extended and a gradual transition to sustainable national financing gets underway

Universality of Symmetry and Mixed-symmetry Collective Nuclear States

The global correlation in the observed variation with mass number of the $E2$ and summed $M1$ transition strengths is examined for rare earth nuclei. It is shown that a theory of correlated $S$ and $D$ fermion pairs with a simple pairing plus quadrupole interaction leads naturally to this universality. Thus a unified and quantitative description emerges for low-lying quadrupole and dipole strengths.Comment: In press, Phys. Rev. Lett. 199

BCG directly induces cell cycle arrest in human transitional carcinoma cell lines as a consequence of integrin cross-linking

BACKGROUND: Current models of the mechanism by which intravesical BCG induces an anti-tumor effect in urothelial carcinoma propose a secondary cellular immune response as principally responsible. Our group has demonstrated that BCG mediated cross-linking of α5 [Image: see text] 1 integrin receptors present on the tumor surface elicits a complex biologic response involving AP1 and NF-κB signaling as well as the transactivation of immediate early genes. This study evaluated the direct biologic effect of cross-linking α5β1 integrin on cell cycle progression and apoptosis in two human urothelial carcinoma cell lines. METHODS: Two independent assays (MTT and Colony forming ability) were employed to measure the effect of α5β1 cross-linking (antibody mediated or BCG) on cellular proliferation. Flow cytometry was employed to measure effect of BCG and α5β1 antibody mediated cross-linking on cell cycle progression. Apoptosis was measured using assays for both DNA laddering and Caspase 3 activation. RESULTS: Results demonstrate that integrin cross-linking by BCG, or antibody mediated crosslinking of α5β1 resulted in a decrease in proliferating cell number. BCG treatment or α5β1 cross-linking increased the percentage of cells in G0/G1, in both 253J and T24 cell lines. Peptide mediated blockade of integrin binding site using RGDS reversed the effect BCG on both proliferation and cell cycle arrest. Apoptosis in response to BCG was not identified by either DNA laddering or Caspase 3 activation. CONCLUSION: These findings show that BCG exerts a direct cytostatic effect on human urothelial carcinoma cell lines. Cell cycle arrest at the G1/S interface is a mechanism by which BCG inhibits cellular proliferation. This effect is duplicated by antibody mediated cross-linking of α5β1 and likely occurs as a consequence of crosslink-initiated signal transduction to cell cycle regulatory genes

Switchable Coupling of Vibrations to Two-Electron Carbon-Nanotube Quantum Dot States

We report transport measurements on a quantum dot in a partly suspended carbon nanotube. Electrostatic tuning allows us to modify and even switch 'on' and 'off' the coupling to the quantized stretching vibration across several charge states. The magnetic-field dependence indicates that only the two-electron spin-triplet excited state couples to the mechanical motion, indicating mechanical coupling to both the valley degree of freedom and the exchange interaction, in contrast to standard models