Performance of the neoBona test: a new paired-end massively parallel shotgun sequencing approach for cell-free DNA-based aneuploidy screening.

OBJECTIVE: To assess the performance of screening for fetal trisomies 21, 18 and 13 by cell-free (cf) DNA analysis of maternal blood using a new method based on paired-end massively parallel shotgun sequencing (MPSS). METHODS: This was a blinded study of plasma samples (1mL) obtained from 1000 women undergoing screening for fetal trisomies 21, 18 and 13 at 11-13 weeks' gestation. The study included 50 cases with confirmed fetal trisomy 21, 30 with trisomy 18, 10 with trisomy 13 and 910 unaffected pregnancies. Paired-end MPSS with the neoBona® test allowed simultaneous assessment of fetal fraction, cfDNA fragment size distribution and chromosome counting, which were integrated into a new analysis algorithm to calculate trisomy likelihood ratios (t-score) for each chromosome of interest. Each sample was classified as trisomic or unaffected using chromosome-specific cut-offs set at t-score values of 1.5 for trisomy 21 and 3.0 for trisomies 18 and 13. RESULTS: Valid results were provided for 988 (98.8%) cases; 12 (1.2%) samples, from nine euploid and three trisomy 21 pregnancies, did not pass quality-control criteria and were excluded from further analysis. All 47 cases of trisomy 21, all 10 of trisomy 13, 29 of 30 with trisomy 18 and all 901 unaffected cases were classified correctly. Median fetal fraction was 10.5% (range, 0.3-33.8%) and trisomic and unaffected cases with low fetal fractions of < 1% were identified correctly. CONCLUSIONS: This novel method for cfDNA analysis of maternal plasma, which utilizes paired-end MPSS, can provide accurate prediction of fetal trisomies. Use of a new multicomponent t-score removes the need to reject samples with fetal fraction < 4%, which potentially extends the benefits of non-invasive prenatal cfDNA analysis to a larger proportion of pregnancies. © 2016 Authors. Ultrasound in Obstetrics & Gynecology published by John Wiley & Sons Ltd on behalf of International Society of Ultrasound in Obstetrics and Gynecology.

Performance of screening for aneuploidies by cell-free DNA analysis of maternal blood in twin pregnancies.

OBJECTIVES: To report clinical implementation of cell-free DNA (cfDNA) analysis of maternal blood in screening for trisomies 21, 18 and 13 in twin pregnancies and examine variables that could influence the failure rate of the test. METHODS: cfDNA testing was performed in 515 twin pregnancies at 10-28 weeks' gestation. The failure rate of the test to provide results was compared with that in 1847 singleton pregnancies, and logistic regression analysis was used to determine which factors among maternal and pregnancy characteristics were significant predictors of test failure. RESULTS: Failure rate of the cfDNA test at first sampling was 1.7% in singletons and 5.6% in twins. Of those with a test result, the median fetal fraction in twins was 8.7% (range, 4.1-30.0%), which was lower than that in singletons (11.7% (range, 4.0-38.9%)). Multivariable regression analysis demonstrated that twin pregnancy, higher maternal weight and conception by in-vitro fertilization provided significant independent prediction of test failure. Follow-up was available in 351 (68.2%) of the twin pregnancies and comprised 334 with euploid fetuses, 12 discordant for trisomy 21 and five discordant for trisomy 18. In all 323 euploid cases with a result, the risk score for each trisomy was < 1:10 000. In 11 of the 12 cases with trisomy 21 and in the five with trisomy 18, the cfDNA test gave a high-risk result, but in one case of trisomy 21, the score was < 1:10 000. CONCLUSION: In twin pregnancies screening by cfDNA testing is feasible, but the failure rate is higher and detection rate may be lower than in singletons.

Uso de las técnicas psicodramáticas en terapia de pareja / No disponible

Se expone en el siguiente trabajo el empleo de técnicas psicodramáticas en terapia de pareja, siguiendo el encuadre de sociodrama de diadas. El fin del mismo es mostrar cómo a través de la metodología del psicodrama se pueden estudiar los vínculos y sus características, cómo influyen en la relaciones de pareja y cómo es posible hacer consciente de los mismos a los protagonistas, promoviendo un cambio en aquellos casos que es necesario (AU)

This study presents the use of psychodrama techniques in couples’s therapy, following the framing of dyads skit. The purpose is to show how through the methodology of psychodrama may be studied the links and their characteristics, how they influence over the couple relationships and how it may make them aware of the protagonists and promote a change in necessary cases (AU)

Clinical application of midtrimester non-invasive fetal RHD genotyping and identification of RHD variants in a mixed-ethnic population.

OBJECTIVE: This study aims to assess the suitability of non-invasive prenatal RHD genotyping in non-immunized midtrimester pregnant women from a mixed ethnic population, to prevent unnecessary anti-D immunoglobulin prophylaxis and to identify RHD variants METHODS: Rhesus D-negative pregnant women were offered fetal RHD genotyping at 24 gestational weeks. A total of 284 samples were tested for RHD status using multiplex rt-PCR amplification of exons 5 and 7 of the RHD gene and exons 6 and 10 in selected cases. Women carrying RHD-negative fetuses were counseled about their option to avoid routine antenatal anti-D immunoglobulin administration. Diagnostic accuracy of RHD genotyping was compared with postnatal Rhesus D serotyping. RESULTS: A total of 184 positives (65%), 91 negatives (32%) and 7 cases (2.5%) compatibles with RHD variants were detected by RHD genotyping. No false negative results were found, and a single false positive was observed in a twin pregnancy. Genotyping was accepted when offered by 94% of women (284/302), and anti-D immunoglobulin was avoided in 95% (90/95) of RHD-negative fetuses. CONCLUSIONS: Non-invasive routine antenatal RHD genotyping at 24 weeks of pregnancy is a highly accurate method, resulting in the avoidance of 95% of unnecessary administrations of anti-D immunoglobulin, with no false negative results.

Chemometric-assisted method development in reversed-phase liquid chromatography.

The computer-assisted development of separation procedures in liquid chromatography (LC), which has involved the adaptation of diverse chemometric techniques and the incorporation of successive advances in informatics and computing over a period of more than four decades, has provided chromatographers with commercial and academic tools of great utility. These tools enable the development, optimization (including robustness studies) and transfer of procedures that constitute one of the analytical activities most commonly used at present in laboratories and industries worldwide. Departing from a typical operational scheme of manually developed chromatographic procedures, this review describes some of the different tools and options that chemometrics provides to the chromatographer for the computer-assisted approach, and outlines the present status of this approach and possible reasons why it is still not considered fully satisfactory.

Seaweed as a source of novel nutraceuticals: sulfated polysaccharides and peptides.

Seaweeds and seaweed-derived products are underexploited marine bioresources and a source of natural ingredients for functional foods. Nutritional studies on seaweeds indicate that brown and red seaweeds possess a good nutritional quality and could be used as an alternative source of dietary fiber, protein, and minerals. Moreover, bioactive sulfated polysaccharides are the main components of soluble fiber in seaweeds and also bioactive peptides can be prepared from seaweed protein. This chapter gives an overview of the main biological properties of sulfated polysaccharides and peptides from brown and red seaweeds. Recent studies have provided evidence that sulfated polysaccharides from seaweeds can play a vital role in human health and nutrition. Besides, peptides derived from algal protein are most promising as antihypertensive agents. Further research work, especially in vivo studies, are needed in order to gain a better knowledge of the relation structure-function by which bioactive compounds from seaweeds exert their bioactivity.

Muerte encefálica: criterios diagnósticos clínicos. Pruebas diagnósticas instrumentales / Brain death: diagnostic clinical criteria: diagnostic instrumental tests

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Retention of arsenate using genetically modified coryneform bacteria and determination of arsenic in solid samples by ICP-MS.

A novel method for the retention of arsenate [As(V)] combining time-controlled solid-phase extraction with living bacterial biomass is presented. As(V) retention was carried out by exposing the extractant, consisting of a living double-mutant of Corynebacterium glutamicum strain ArsC1-C2, to the sample for a retention time of 1-7min, before the arsenic distribution equilibrium between the sample solution and the extractant was established. The amount of As(V) retained in the biomass was measured by inductively coupled plasma-mass spectrometry (ICP-MS) after the sample had been treated with nitric acid. A theoretical model of the retention process was developed to describe the experimental retention-time profiles obtained with the bacterial cells. This relationship provided a feasible quantification of the retention process before steady-state was reached, providing that the agitation conditions and the retention time had been controlled. An analytical procedure for the retention/quantification of As(V) was then developed; the detection limit was 0.1 ng As(V)mL(-1) and the relative standard deviation 2.4-3.0%. The maximum effective retention capacity for As(V) was about 12.5mgAs(g biomass)(-1). The developed procedure was applied to the determination of total arsenic in coal fly ash, using a sample that had undergone oxidative pre-treatment.

Evidence that some imidazoline derivatives inhibit peripherally the vasopressor sympathetic outflow in pithed rats.

Imidazoline derivatives (e.g. clonidine and moxonidine) and alpha(2)-adrenoceptor agonists (e.g. B-HT 933) have been shown to inhibit sympathetically-induced [(3)H]noradrenaline release in several isolated blood vessels. The present study has compared the potential capability of agonists at imidazoline I(1/2) receptors and/or alpha(1/2)-adrenoceptors to inhibit the sympathetically-induced vasopressor responses in pithed rats. For this purpose, male Wistar rats were pithed and prepared for measurement of diastolic blood pressure and heart rate. Then, the vasopressor responses induced by either selective electrical stimulation (2 ms, 60 V; 0.03, 0.1, 0.3, 1 and 3 Hz) of the vascular sympathetic outflow (T(7)-T(9)) or i.v. bolus injections of exogenous noradrenaline (0.03, 0.1, 0.3, 1 and 3 microg/kg) were determined before and during i.v. continuous infusions of the agonists B-HT 933 (alpha(2)), clonidine (alpha(2), I(1)), moxonidine (alpha(2), I(1)), cirazoline (alpha(1), I(2)), agmatine (putative endogenous ligand of imidazoline receptors) and methoxamine (alpha(1)), or equivalent volumes of physiological saline. Electrical sympathetic stimulation elicited frequency-dependent vasopressor responses which were significantly inhibited during the continuous infusions of B-HT 933, clonidine, moxonidine, cirazoline and agmatine, but not of physiological saline. Interestingly, the vasopressor responses to exogenous noradrenaline, which remained unaffected during the infusions of physiological saline, B-HT 933, moxonidine, cirazoline and agmatine, were significantly blocked during the infusions of clonidine or methoxamine. These results suggest that B-HT 933, moxonidine, cirazoline and agmatine induced a prejunctional inhibition of the vasopressor sympathetic outflow in pithed rats, whilst clonidine inhibited the vasopressor sympathetic outflow by both prejunctional and postjunctional mechanisms.

Retention of inorganic arsenic by coryneform mutant strains.

The natural resistance mechanisms of corynebacteria to respond to the environments containing high levels of arsenic were successfully adopted to develop inexpensive and selective extractants for submicrogram amounts of arsenic. Kinetic and equilibrium characteristics were evaluated, and a preliminary exploration of the capability of these strains to be used for arsenic speciation was also made in this work. Three kinetics models were used to fit the experimental data. It was found that the pseudo-first-order kinetics model was not quite adequate to describe the retention process, while the intraparticle diffusion and the pseudo-second-order kinetics models provide the best fits. The equilibrium isotherm showed that the retention of arsenic was consistent with the Langmuir equation and that the Freundlich and Dubinin-Radushkevich models provided poorer fits to the experimental data. The maximum effective retention capacity for arsenic was about 15.4 ng As/mg biomass. The amount of arsenic retained was directly measured in the biomass by forward planning a slurry electrothermal atomic absorption spectrometric procedure.

Identification of the emerging skin pathogen Corynebacterium amycolatum using PCR-amplification of the essential divIVA gene as a target.

The actinomycete Corynebacterium amycolatum is a saprophytic bacterium usually associated with the human skin, but it is at present considered an emergent pathogen as it is isolated from nosocomial settings from samples of immunosuppressed patients. The conventional method to distinguish C. amycolatum from closely related species is mainly based on phenotypic or chemotaxonomic studies. We developed a molecular method to identify rapidly C. amycolatum based on the use of different primers for amplification of the cell division divIVA gene using conventional or real-time PCR. This technique was used for the first time to distinguish C. amycolatum from the closely related Corynebacterium striatum, Corynebacterium minutissimum and Corynebacterium xerosis, without the requirement of further molecular analysis. The suitability of the identification method was tested on 51 clinical isolates belonging to the nonlipophilic fermentative group of corynebacteria (cluster C. striatum/C. amycolatum), which were accurately characterized by sequencing a 0.8 kb fragment of the 16S rRNA gene.

Evaluación de la PCR cuantitativa fluorescente para el diagnóstico prenatal rápido de anomalías cromosómicas / Tittle

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Rapid prenatal diagnosis of common chromosome aneuploidies by QF-PCR. Assessment on 18,000 consecutive clinical samples.

The quantitative fluorescent PCR (QF-PCR) assay, introduced during the last few years, allows prenatal diagnoses of common chromosome aneuploidies in a few hours after sampling. We report the first assessment of QF-PCR performed on a large cohort of 18,000 consecutive clinical specimens analysed in two different Centres. All samples were analysed by QF-PCR using several selected STR markers together with amelogenin and, occasionally, SRY for fetal sexing. Results were compared with those obtained by conventional cytogenetic analysis. In 17,129 tests, normal fetuses were detected by QF-PCR. No false positives were observed. All 732 cases of trisomy 21, 18, 13, triploidies, double trisomies as well as all but one fetuses with X and Y aneuploidies were correctly diagnosed. Chromosome mosaicism could also be suspected in several samples. In some cases of in vitro culture failures, QF-PCR was the only evidence of fetal X, Y, 21, 18 and 13 chromosome complement. QF-PCR proved to be efficient and reliable in detecting major numerical chromosome disorders. The main advantages of the molecular assay are its very low cost, speed and automation enabling a single operator to perform up to 40 assays per day. QF-PCR relieves anxiety of most parents within 24 h from sampling and accelerates therapeutic interventions in the case of an abnormal result. In countries where large scale conventional cytogenetics is hampered by its high cost and lack of technical expertise, QF-PCR may be used as the only prenatal diagnostic test.

Inadvertent ureteral catheterizaion with a microtip catheter at cystometry.

Two patients in whom the right ureter was inadvertently catheterized at water cystometry are described. Accidental ureteral catheterization and filling was followed by colicky pain in the right flank and by an abrupt increase in the recorded pressure, up to 148 cm H2O. The pain disappeared and the intravesical pressure returned to baseline after the microtip catheter was withdrawn.

[Role of adenosine triphosphate (ATP) in head-up tilt-induced syncope]. / Papel del adenosín trifosfato (ATP) en la inducción de síncope mediante test de basculación.

INTRODUCTION AND OBJECTIVES: Recent studies have demonstrated that adenosine is an endogenous modulator of the cardiac excitatory afferent nerves, and could provoke a vasovagal response during head-up tilt test. Isoproterenol has been the drug of choice to increase the sensitivity of this testing. The aim of the present study was to analyze the role of adenosine in head-up tilt-induced syncope in susceptible patients, and to compare the relative sensitivities of adenosine and isoproterenol. METHODS: Thirty patients with unexplained syncope (16 female and 14 male, mean age 37.1 +/- 18 years), no heart disease and negative baseline head-up tilt test were studied. After the baseline test, patients were randomized to receive adenosine triphosphate (bolus injections of 3, 6 and 9 mg/ 5 min) or isoproterenol (bolus injections of 2, 4 and 6 micrograms/5 min) and underwent a second tilt test. After 15 min at rest, patients received the alternative drug and a third test was performed. Eleven normal control subjects were tested with adenosine in the upright position to determine its effects. RESULTS: A vasovagal response was induced in 7 patients (23.3%) after ATP administration. Nine patients (30%) showed a positive response with isoproterenol. Only 2 patients (6.6%) showed a positive response with both drugs. Of the control subjects, one (9%) had a vasovagal response after ATP administration. CONCLUSIONS: We conclude that adenosine triphosphate seems to be a useful tool to provoke vasovagal reaction in susceptible patients during head-up tilt test.

[Acute respiratory infections in pediatric intensive care units. A multicenter prospective study]. / Infecciones respiratorias agudas en unidades de cuidados intensivos pediátricos. Estudio prospectivo multicéntrico.

OBJECTIVE: The purpose of this study was to know the etiology, clinical background, treatment an evolution of severe infectious diseases in children admitted to Pediatric Intensive Care Units (PICUs). PATIENTS AND METHODS: A multicenter prospective study was carried out. Children with respiratory infections admitted to 10 PICUs throughout Spain between May 1994 and April 1995 were included in a long term survey. The nosocomial infections were not included. Student's t and Wilcoxon tests were used for quantitative variables and Chi square with Yates correction and Fisher's test for the qualitative variables. RESULTS: One hundred twenty-two patients with acute respiratory infections were studied. The mean value on Downes score at admittance was 5.2 +/- 2.3. Diagnosis were allocated as follows: 47 bronchopneumonia (38.5%), 40 bronchiolitis (33%), 15 epiglotitis (12%), 14 laryngitis (11.5%) and "others" 6 (5%). Etiologic agents were identified in 69 cases (56.5%), with respiratory syncytial virus being the most frequently isolated agent (35 cases, 51%), followed by Hemophilus influenzae in 13 cases (19%). The mean PICU stay was 5.8 +/- 7.9 days (1-67 days). Of these cases, 112 (92%) recovered completely and 9 (7%) died (8 with bronchopneumonia and 1 with epiglotitis). A significant association could be seen between the increase in mortality and the variables Downes' score and diagnosis of bronchopneumonia. CONCLUSIONS: The most frequent respiratory infections in the PICU were pneumonia and bronchopneumonia. Viral etiology, with a frequency of 54%, was the main cause of respiratory infection. Bacterial etiology represented 46% of the total cases, with Hemophilus influenzae as the most frequent etiologic agent.

Role of endogenous opioids in syncope induced by head-up tilt test and its relationship with isoproterenol-dependent and isoproterenol-independent neurally-mediated syncope.

This study was designed to evaluate the role of endogenous opioids in neurally-mediated syncope. Head-up tilt test was performed on 35 patients with syncope of unknown origin. Plasma beta-endorphin was measured (1) at baseline, (2) at the end of tilt test or at time of syncope, (3) 15 min before isoproterenol-test, (4) at the end of the isoproterenol-test or at time of syncope. Subjects with a positive tilt testing showed a larger rise in plasma beta-endorphin concentrations at time of syncope (baseline 13.7+/-8.0 vs. syncope 41.4+/-26.4 pmol l(-1); P<0.01). On the contrary, patients with a positive isoproterenol-test showed no rise in plasma beta-endorphin levels (baseline 7.9+/-3.6 vs. syncope 7.4+/-2.7 pmol l(-1); P=ns). Patients with a passive negative tilt test (baseline 6.7+/-2.8 vs. end of test 7.0+/-3.3 pmol l(-1); P=ns) and negative isoproterenol tilt test (baseline 7.4+/-3.8 vs. end of test 8.1+/-3.4 pmol l(-1); P=ns) showed no changes in beta-endorphin concentrations. To further examine the efficacy of i.v. naloxone to prevent syncope, 10 patients were randomized to naloxone (0.02 mg/kg) or placebo. Second head-up tilt testing was negative in 1/5 patients with naloxone and in 2/5 patients with placebo. We conclude that, (1) endogenous opioids seem to be involved in vasovagal syncope induced by baseline head-up tilt test, (2) changes in plasma beta-endorphin concentrations show significant differences between patients who have isoproterenol-dependent and isoproterenol-independent syncope, this finding might occur in the setting of different pathophysiologic mechanisms, and (3) intravenous naloxone at a dose of 0.02 mg/kg was not superior to placebo in order to prevent positive responses to baseline tilt test.

[Epidemiological changes in infectious endocarditis. A prospective study, 1992-96]. / Cambios epidemiológicos en la endocarditis infecciosa. Estudio prospectivo 1992-96.

BACKGROUND: To know the present epidemiological situation of the infective endocarditis in our environment and its evolution in the last few years. RESULTS: The incidence of infective endocarditis was 0.85 per thousand patients admitted to hospital, with a mean age of 43 years. The predisposed factors more frequently found were: drug addiction (32%) and cardiac prosthetic valves (23%). In the greatest number of our patients the cardiac valves involved were: tricuspid (28%), mitral (27%) and prosthetic valves (23%). The causative organism were: S. aureus (19 cases), Streptococcus (15 cases) and S. epidermidis (11 cases). The echocardiography study resulted diagnostic in 90% of the patients, valve replacements were performed in 22% of the cases. The overall mortality rate was 10%. CONCLUSIONS: The current profile of infective endocarditis is characterized by a high incidence of parenterally drug addict patients or prosthetic valves carriers. Increase of the infections of S aureus and a decrease of Streptococcus infections, as well as a less overall mortality.