Comparison of different PCR amplification targets for molecular diagnosis of Strongyloides stercoralis.

Molecular techniques are an alternative for the diagnosis of strongyloidiasis, produced by Strongyloides stercoralis. However, it is necessary to determine the best amplification target for the populations of this parasite present in a geographical area and standardize a polymerase chain reaction (PCR) protocol for its detection. The objectives of this work were the comparison of different PCR targets for molecular detection of S. stercoralis and the standardization of a PCR protocol for the selected target with the best diagnostic results. DNA extraction was performed from parasite larvae by saline precipitation. Three amplification targets of the genes encoding ribosomal RNA 18S (18S rDNA) and 5.8S (5.8S rDNA) and cytochrome oxidase 1 (COX1) of S. stercoralis were compared, and the PCR reaction conditions for the best target were standardized (concentration of reagents and template DNA, hybridization temperature, and number of cycles). The analytical sensitivity and specificity of the technique were determined. DNA extraction by saline precipitation made it possible to obtain DNA of high purity and integrity. The ideal target was the 5.8S rDNA, since the 18S rDNA yielded non-reproducible results and COX1 never amplified under any condition tested. The optimal conditions for the 5.8S rDNA-PCR were: 1.5 mM MgCl2, 100 µM dNTPs, 0.4 µM primers, and 0.75 U DNA polymerase, using 35 cycles and a hybridization temperature of 60 °C. The analytical sensitivity of the PCR was 1 attogram of DNA, and the specificity was 100%. Consequently, the 5.8S rDNA was shown to be highly sensitive and specific for the detection of S. stercoralis DNA.

A hybrid artificial intelligence model leverages multi-centric clinical data to improve fetal heart rate pregnancy prediction across time-lapse systems.

STUDY QUESTION: Can artificial intelligence (AI) algorithms developed to assist embryologists in evaluating embryo morphokinetics be enriched with multi-centric clinical data to better predict clinical pregnancy outcome? SUMMARY ANSWER: Training algorithms on multi-centric clinical data significantly increased AUC compared to algorithms that only analyzed the time-lapse system (TLS) videos. WHAT IS KNOWN ALREADY: Several AI-based algorithms have been developed to predict pregnancy, most of them based only on analysis of the time-lapse recording of embryo development. It remains unclear, however, whether considering numerous clinical features can improve the predictive performances of time-lapse based embryo evaluation. STUDY DESIGN, SIZE, DURATION: A dataset of 9986 embryos (95.60% known clinical pregnancy outcome, 32.47% frozen transfers) from 5226 patients from 14 European fertility centers (in two countries) recorded with three different TLS was used to train and validate the algorithms. A total of 31 clinical factors were collected. A separate test set (447 videos) was used to compare performances between embryologists and the algorithm. PARTICIPANTS/MATERIALS, SETTING, METHODS: Clinical pregnancy (defined as a pregnancy leading to a fetal heartbeat) outcome was first predicted using a 3D convolutional neural network that analyzed videos of the embryonic development up to 2 or 3 days of development (33% of the database) or up to 5 or 6 days of development (67% of the database). The output video score was then fed as input alongside clinical features to a gradient boosting algorithm that generated a second score corresponding to the hybrid model. AUC was computed across 7-fold of the validation dataset for both models. These predictions were compared to those of 13 senior embryologists made on the test dataset. MAIN RESULTS AND THE ROLE OF CHANCE: The average AUC of the hybrid model across all 7-fold was significantly higher than that of the video model (0.727 versus 0.684, respectively, P = 0.015; Wilcoxon test). A SHapley Additive exPlanations (SHAP) analysis of the hybrid model showed that the six first most important features to predict pregnancy were morphokinetics of the embryo (video score), oocyte age, total gonadotrophin dose intake, number of embryos generated, number of oocytes retrieved, and endometrium thickness. The hybrid model was shown to be superior to embryologists with respect to different metrics, including the balanced accuracy (P ≤ 0.003; Wilcoxon test). The likelihood of pregnancy was linearly linked to the hybrid score, with increasing odds ratio (maximum P-value = 0.001), demonstrating the ranking capacity of the model. Training individual hybrid models did not improve predictive performance. A clinic hold-out experiment was conducted and resulted in AUCs ranging between 0.63 and 0.73. Performance of the hybrid model did not vary between TLS or between subgroups of embryos transferred at different days of embryonic development. The hybrid model did fare better for patients older than 35 years (P < 0.001; Mann-Whitney test), and for fresh transfers (P < 0.001; Mann-Whitney test). LIMITATIONS, REASONS FOR CAUTION: Participant centers were located in two countries, thus limiting the generalization of our conclusion to wider subpopulations of patients. Not all clinical features were available for all embryos, thus limiting the performances of the hybrid model in some instances. WIDER IMPLICATIONS OF THE FINDINGS: Our study suggests that considering clinical data improves pregnancy predictive performances and that there is no need to retrain algorithms at the clinic level unless they follow strikingly different practices. This study characterizes a versatile AI algorithm with similar performance on different time-lapse microscopes and on embryos transferred at different development stages. It can also help with patients of different ages and protocols used but with varying performances, presumably because the task of predicting fetal heartbeat becomes more or less hard depending on the clinical context. This AI model can be made widely available and can help embryologists in a wide range of clinical scenarios to standardize their practices. STUDY FUNDING/COMPETING INTEREST(S): Funding for the study was provided by ImVitro with grant funding received in part from BPIFrance (Bourse French Tech Emergence (DOS0106572/00), Paris Innovation Amorçage (DOS0132841/00), and Aide au Développement DeepTech (DOS0152872/00)). A.B.-C. is a co-owner of, and holds stocks in, ImVitro SAS. A.B.-C. and F.D.M. hold a patent for 'Devices and processes for machine learning prediction of in vitro fertilization' (EP20305914.2). A.D., N.D., M.M.F., and F.D.M. are or have been employees of ImVitro and have been granted stock options. X.P.-V. has been paid as a consultant to ImVitro and has been granted stocks options of ImVitro. L.C.-D. and C.G.-S. have undertaken paid consultancy for ImVitro SAS. The remaining authors have no conflicts to declare. TRIAL REGISTRATION NUMBER: N/A.

Recalculating the staff required to run a modern assisted reproductive technology laboratory.

STUDY QUESTION: What number of staff is sufficient to perform increasingly complicated processes in today's modern ART laboratories? SUMMARY ANSWER: The adequate number of personnel required for the efficient and safe operation of modern ART laboratories needs to be calculated. WHAT IS KNOWN ALREADY: In today's modern ART laboratories, the amount of time required to perform increasingly complicated processes has more than doubled, with a downward trend in the amount of work an embryologist can do. Different workload unit values have been used to evaluate each workload task and efficiency in a particular ART laboratory, as well as to occasionally compare one laboratory with another. STUDY DESIGN, SIZE, DURATION: Seven senior embryologists working at different IVF centers, three public and four private centers, participated in this multicenter study conducted between 2019 and 2020. We prepared a survey to create a calculator for staff using the average (of three attempts) time spent in every laboratory by each embryologist of the center to perform any ART process. PARTICIPANTS/MATERIALS, SETTING, METHODS: Different laboratory processes and activities related to quality control, time spent and conventional human double witnessing were included in the survey. To calculate the number of processes that each embryologist can perform per year, an embryologist was considered to be having a full-time contract and working 7 or 8 h/day. The times included in the calculation of each task were those corresponding to the 95th percentile. For the calculations, Microsoft® Office Excel® Professional Plus 2019 was used. MAIN RESULTS AND THE ROLE OF CHANCE: The survey showed that the time needed per embryologist to perform the different processes necessary for a classic IVF cycle without time lapse (TL) was 8.11 h, and with TL, it was 10.27 h. The calculated time also considered the time spent in documentation handling, cycle preparation, database management and conventional human double witnessing verification. An ICSI without TL needed 8.55 h, and with TL, it needed 10.71 h. An ICSI-PGT without a TL cycle needed 11.75 h, and with TL, it needed 13.91 h. Furthermore, 1.81 h should be added for every vitrification support needed. The time needed to control more than 200 critical steps, including equipment control and culture parameters, was 30 min per day plus 3.9 min per device to control.The time spent in semen analysis (including documentation handling, cycle preparation and database management) or intrauterine insemination with a partner sperm was 2.7 h. For donor sperm, an additional hour was required for the management involved. The time required to perform a testicular biopsy and cryopreserve the sample was 4 h. Similarly, the time required to perform seminal cryopreservation was 3.7 h. LIMITATIONS, REASONS FOR CAUTION: The study was conducted considering a full-time contract embryologist working 7 or 8 h/day, 5 days a week, with days off according to the Spanish regulations. However, our findings can be adapted to foreign regulations using the developed online calculation platform. WIDER IMPLICATIONS OF THE FINDINGS: A new advanced staff calculator allows any IVF laboratory to estimate the minimum number of embryologists necessary without compromising the security or success of the results. Nevertheless, we recommend a minimum of two qualified embryologists in every laboratory, regardless of the workload. STUDY FUNDING/COMPETING INTEREST(S): This work was funded by the Asociación para el Estudio de la Biología de la Reproducción (ASEBIR). None of the authors has any conflict of interest to declare. TRIAL REGISTRATION NUMBER: N/A.

Mycotoxins in raw materials, beverages and supplements of botanicals: A review of occurrence, risk assessment and analytical methodologies.

Over recent years, consumer interest in natural products, such as botanicals has increased considerably. One of the factors affecting their quality is the presence of mycotoxins. This review focuses on exploring the mycotoxin occurrence in botanicals (raw material and ready-to-eat forms such as infusions or tablets) and the risk assessment due to their ingestion. Aflatoxins, Ochratoxin A, and Fumonisins are the most commonly studied mycotoxins and data in the literature report levels ranging from traces to 1000 µg/kg in raw materials. In general, the highest contents observed in raw materials decreased to unconcerning levels after the preparation of the infusions, reaching values that generally do not exceed 100 µg/L. Regarding botanical dietary supplements, the levels observed were lower than those reported for other matrices, although higher levels (of up to 1000 µg/kg) have been reported in some cases. Risk assessment studies in botanicals revealed a higher risk when they are consumed as tablets compared to infusions. Analytical methodologies implied in mycotoxin determination have also been contemplated. In this sense, liquid chromatography coupled to fluorescence detection has been the most frequently employed analytical technique, although in recent years tandem mass spectrometry has been widely used.

Domiciliation and sympatry of Triatoma maculata and Rhodnius prolixus, risk of Trypanosoma cruzi transmission in villages of Anzoátegui, Venezuela.

The domiciliation of Triatoma maculata and Rhodnius prolixus and the entomological risk indicators for the transmission of Trypanosoma cruzi, an etiological agent of Chagas Disease-CD, were studied in rural villages of Anzoátegui state, Venezuela. Nightly home visits were made for 4 months/year, for 2 years, to search for and capture triatomines in human settlements. For six of the evaluated villages, 16.4% (11/67) of houses were found with triatomine infestation; obtaining 151 triatomines in all their ontogenetic stages, of which 54.3% (82/151) corresponded to T. maculata and 45.7% (69/151) to R. prolixus. In 7.5% of the evaluated houses, both species were presented in sympatry. Entomological indicators of transmission risk were higher for T. maculata in relation to R. prolixus. Inoculation of fecal flagellates of triatomines produced 2.92 × 105 flagellates/mL of blood in mean and 100% mortality in the murine model. Molecular tests (satellite DNA, kDNA and DTUs studies) demonstrated the presence of T. cruzi, all compatible with TcI. The food source determined by IESPA, revealed that R. prolixus showed less eclecticism in relation to T. maculata in the use of blood sources. This could be an indicator of an older domiciliation with low dispersion between ecotopes. The sympatry of T. maculata and R. prolixus had been recorded in natural niches, but for the first time it is recorded inside the houses in rural villages of the Anzoátegui state. Human dwellings can constitute an adequate niche, with available food sources for both triatomines species and with the risk of establishing AT/CD as zoonosis or zooanthroponosis.

Seroepidemiological evidence for Taenia solium taeniasis/cysticercosis in three Venezuelan rural communities.

Taenia solium is the most common parasite infection of the brain, causing neurocysticercosis and typically found in rural communities with free-ranging pigs. Identification of transmission in rural areas is essential for its control. Risk factors and transmission of the parasite were evaluated in three rural Venezuelan communities (Valle del Rio and Potrero Largo, Cojedes state; and Palmarito, Portuguesa state) by a questionnaire (112 households) and coprological (492 samples) and serological (433 human and 230 porcine sera) analysis, respectively. Typical risk factors were found in all three communities: free-foraging pig husbandry, deficient sanitary conditions, high open defecation and ignorance of the parasite life cycle. Coprological examinations revealed a high level of soil-transmitted parasites. Importantly, two T. solium adult worm carriers were identified in each of the three communities. Anti-metacestode antibodies and the HP10 secreted metacestode glycoprotein were detected at significant levels in human and porcine sera in Valle del Rio, Potrero Largo and Palmarito. In conclusion, these communities may be considered to be endemic for taeniasis/cysticercosis, and the instigation of an appropriate control programme is recommended.

Molecular characterization of Trypanosoma cruzi and Leishmania spp. coinfection in mammals of Venezuelan coendemic areas.

BACKGROUND & OBJECTIVES: Trypanosoma cruzi and Leishmania spp. are protozoans that cause American trypanosomiasis and leishmaniasis, respectively. In endemic foci where both diseases coincide, coinfection can occur. The objective of this work was the characterization of the parasites involved in coinfection in several endemic areas of Venezuela. METHODS: Molecular characterization was done in 30 samples of several species of mammals (Didelphis marsupialis, Equus mulus, Rattus rattus, Canis familiaris, Felis catus, and Sciurus granatensis) from the states of Anzoategui, Cojedes and Capital District diagnosed with T. cruzi and Leishmania spp. coinfections. For the typing of T. cruzi DTUs, the markers of miniexon, 24Sa rDNA, 18Sa rDNA, and hsp60-PCR-RFLP (EcoRV) were used. Infection by Leishmania spp. was characterized by miniexon multiplex PCR for complexes of Leishmania and ITS1-PCR-RFLP (HaeIII, HhaI, and RsaI) for the identification of the species. RESULTS: The T. cruzi TcI was present in 100% of the coinfected mammals, which included 76.7% of triple infection by T. cruzi TcI-complex-L. (L) mexicana-L. infantum/chagasi, 13.3% of double infection by T. cruzi TcI-L. mexicana and 10% of double infection by T. cruzi Tcl-L. infantum/chagasi. INTERPRETATION & CONCLUSION: These results suggest that the double or triple infection is a phenomenon existing in almost all the coendemics areas and mammals studied, which might influence the mechanisms of adaptation and pathogenicity of these parasites.

[Hypertensive urgencies: Description of clinical characteristics, management, variables associated with re-entry and short-term prognosis]. / Urgencias hipertensivas: descripción de características clínicas, manejo, variables asociadas a reconsulta y pronóstico a corto plazo.

OBJECTIVES: To describe the characteristics of hypertensive urgencies at the emergency department, as well as the variables associated with early re-admission (<7 days) and re-admission at one month (<30 days). METHODS: We conducted a descriptive, retrospective study of all patients who were admitted to the emergency department of a third level hospital during 2013. Subsequently, a case-control analysis was performed to analyze the group of patients with readmission. RESULTS: A total of 398 hypertensive urgencies were collected (32.4% men, mean age 67.75 years), which led to an incidence of 3.28/1000 visits. Eighty point nine percent had a previous history of hypertension, and the mean Charlson Index was 2.23. Headache was the most frequent symptom (49.1%), followed by dizziness/instability (29.5%) and nausea/vomiting (17.1%). Eighty point seven percent of the patients were prescribed pharmacological treatment. The rates of cardiovascular events or mortality at one month were low (2.26% and 0.25% respectively). Despite this, 7.53% and 11.31% of patients were readmitted in under 7 days or 30 days, respectively. The variables associated with readmission in the multivariate analysis were elevated systolic blood pressure in the first determination, previous hypertension and the presence of palpitations. CONCLUSIONS: Hypertensive emergencies are high-incidence conditions in the Emergency Department. In our study, patients with a prior diagnosis of hypertension and elevated systolic blood pressure at the first determination had a higher risk of re-entry and would be candidates for closer follow-up on discharge.

Presence of mycotoxins in ready-to-eat food and subsequent risk assessment.

A study on a set of ready-to-eat meals (n = 328) based on cereals, legumes, vegetables, fish and meat was carried out to determine the natural presence of twenty-seven mycotoxins by both liquid chromatography and gas chromatography coupled mass spectrometry in tandem (MS/MS) after QuEChERS extraction. The occurrence of mycotoxins was headed by cereal samples with 35% of samples contaminated by at least one mycotoxin followed by vegetables (32%), legumes (15%) and lastly, 9% of fish and meat samples were contaminated. DON was the most detected mycotoxin in vegetables, meat, fish and cereals with an incidence of 13% 18% 19% and 60%, respectively, and the highest mean levels were found in fish (1.19 µg/kg) and vegetable (1.53 µg/kg), respectively. The highest levels means were for HT-2 toxin ranging from 4.03 to 7.79 µg/kg, in cereal and legume samples respectively. In this last, HT-2 toxin was also the most prevalent (54%). In meat samples, OTA resulted with highest value with 8.09 µg/kg. Likewise, PCA analysis revealed a high correlation between the mycotoxins and the food groups analyzed. The findings indicate that there is no toxicological concern associated with exposure to mycotoxins for consumers as all levels were in accordance with the legislation.

[Hydrocephalus due to hyperplasia of the choroid plexuses in a patient with trisomy 9 mosaicism. A real diagnostic and therapeutic challenge]. / Hidrocefalia por hiperplasia de plexos coroideos en un paciente con mosaicismo de trisomia 9. Un verdadero reto diagnostico y terapeutico.

INTRODUCTION: Trisomy 9 is an unusual chromosome abnormality in live-born patients, which is frequently accompanied by functional and structural anomalies of the central nervous system. Among many other alterations, several studies have been published in the English-speaking literature that show an association between chromosome 9 abnormality and pathologies affecting the choroid plexuses. CASE REPORT: We report the case of a 4-month-old male with trisomy 9 mosaicism associated to hydrocephalus secondary to choroid plexus hyperplasia, who was referred due to a clinical picture of intracranial hypertension. The cerebrospinal fluid (CSF) drainage procedure that was initially chosen caused massive ascites due to an excessive production of CSF, and led to a cascade of multiple surgical interventions, which included endoscopic and drainage procedures. CONCLUSIONS: This is another example of an association between choroid plexus pathologies and chromosome 9 abnormality. Due to its scarce incidence, diagnosis of hydrocephalus secondary to plexus hyperplasia is difficult, as is selecting its most suitable treatment. In this type of hydrocephalus there is a double pathophysiological mechanism, which involves an increase in CSF production and a decrease in its reabsorption. Despite taking these considerations into account, the treatment of hydrocephalus secondary to plexus hyperplasia is a real challenge that usually leads to multiple surgical interventions ranging from plexectomy or coagulation of the choroid plexuses to the implantation of CSF drainage devices.

TITLE: Hidrocefalia por hiperplasia de plexos coroideos en un paciente con mosaicismo de trisomia 9. Un verdadero reto diagnostico y terapeutico.Introduccion. La trisomia 9 es una cromosomopatia inusual en pacientes nacidos vivos, que frecuentemente se acompaña de anomalias funcionales y estructurales del sistema nervioso central. Entre otras muchas alteraciones, varios trabajos en la bibliografia anglosajona demuestran una asociacion entre cromosomopatia 9 y patologia de los plexos coroideos. Caso clinico. Varon de 4 meses de vida con mosaicismo de trisomia 9 asociado a hidrocefalia secundaria a hiperplasia de los plexos coroideos, que fue remitido por clinica de hipertension intracraneal. El procedimiento derivativo de liquido cefalorraquideo por el que optamos inicialmente provoco una ascitis masiva debida a la produccion desmesurada de liquido cefalorraquideo, y desemboco en una cascada de multiples intervenciones quirurgicas, entre las que se incluyeron procedimientos endoscopicos y derivativos. Conclusiones. Se trata de un ejemplo mas de asociacion entre patologia de los plexos coroideos y cromosomopatia 9. Debido a su escasa incidencia, es dificil establecer el diagnostico de hidrocefalia secundaria a hiperplasia de los plexos y, por tanto, el tratamiento mas adecuado. En este tipo de hidrocefalia existe un doble mecanismo fisiopatologico, que implica un aumento de produccion de liquido cefalorraquideo y una disminucion de su reabsorcion. A pesar de tener en cuenta dicha consideracion, el tratamiento de la hidrocefalia secundaria a hiperplasia de los plexos supone un verdadero reto que habitualmente pasa por multiples procedimientos quirurgicos, desde la plexectomia o coagulacion de los plexos coroideos hasta la implantacion de dispositivos de derivacion de liquido cefalorraquideo.

Risk factors for diabetic macular oedema in type 2 diabetes: A case-control study in a United Kingdom primary care setting.

AIM: To identify risk factors associated with the development of DMO among patients diagnosed with type 2 diabetes managed in a primary care setting in the UK. METHODS: A case-control study nested in a cohort of incident Type 2 diabetes identified in The Health Improvement Network database from 2000-2007. Cases were people with DMO (N=211) and controls were a DMO-free sample (N=2194). No age restrictions were applied. Adjusted odds ratios and 95%CIs were estimated (OR; 95%CI). RESULTS: DMO increased with high alcohol use (2.88; 1.49-5.55), cataracts (4.10; 2.73-6.15), HbA1c ≥7% (1.58; 1.08-2.32), systolic blood pressure ≥160mm Hg (2.03; 1.17-3.53), total cholesterol ≥5mmol/L (1.66; 1.15-2.39), LDL ≥3mmol/L (1.73; 1.14-2.61), and microalbuminuria (1.78; 1.16-2.73). Diuretic drugs were associated with a reduced risk of DMO (0.68; 0.47-0.99), as did smoking (0.47; 0.28-0.77), overweight (0.53; 0.30-0.96) or obesity (0.52; 0.30-0.91) at diabetes diagnosis, and high triglyceride levels (0.51; 0.35-0.74). Patients treated with anti-diabetic drugs showed higher risk of DMO than non-treated patients, particularly those with sulphonylureas (3.40; 2.42-4.78), insulin (3.21; 1.92-5.36) or glitazones (1.88; 1.17-3.04). CONCLUSION: In patients with type 2 diabetes managed in primary care, multiple factors associated with DMO were identified, such as cataracts, microalbuminuria and high levels of HbA1c, systolic BP, total cholesterol, and LDL. Diuretic drugs were associated with a reduced risk of DMO. Treated diabetes, particularly with sulphonylureas, insulin or glitazones showed highest risk of DMO. The inverse association between smoking, obesity, and triglycerides and DMO deserves further research.

Mitigation of enniatins in edible fish tissues by thermal processes and identification of degradation products.

Emerging mycotoxins, such as enniatins and beauvericin, are common contaminants in vegetal matrices, but recently, the occurrence of mycotoxins in foodstuffs from animal origin has been also reported as they can be present in edible tissues of animals fed with contaminated feedstuffs. Sea bass, sea bream, Atlantic salmon and rainbow trout from aquaculture analyzed in the present survey showed contamination by emerging Fusarium mycotoxins enniatins (ENs). ENs were extracted from raw and cooked fish with acetonitrile and analyzed by Liquid Chromatography coupled to Mass Spectrometry. In this study, the stability of ENs was evaluated during food processing by the application of different cooking methods (broiling, boiling, microwaving and baking treatments). All treated samples showed a reduction in mycotoxin levels with different percentages depending on the type of EN and the fish species. Thus, the reduction obtained ranged from 30 to 100%. The thermal treatments have shown to be a good strategy to mitigate ENs content in edible fish tissues. On the other hand, some ENs degradation products originated during the application of thermal treatments were identified.

Elvitegravir concentrations in seminal plasma in HIV-1-infected men.

OBJECTIVES: The aim of the study was to quantify elvitegravir (EVG) concentrations in the semen of HIV-1-infected men receiving antiretroviral therapy (ART) consisting of an elvitegravir/cobicistat/emtricitabine/tenofovir (EVG/COBI/FTC/TDF) single-tablet regimen. METHODS: A phase IV, cross-sectional study was carried out including HIV-1-infected male adults with suppressed plasma HIV-1 RNA who switched ART to EVG/COBI/FTC/TDF. Total EVG concentrations at the end of the dosing interval (C24 h ) and HIV-1 RNA were measured in paired seminal plasma (SP) and blood plasma (BP) samples 4 weeks after switching to EVG/COBI/FTC/TDF. Validated liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to quantify EVG concentrations, and HIV-1 RNA was determined by real-time polymerase chain reaction (PCR). RESULTS: Ten men were included. Their median age was 40 years (range 24-47 years), the median time on ART was 50 months (range 10-186 months), the median time with plasma HIV-1 RNA < 40 copies/mL was 37 months (range 7-113 months), and the median CD4 count was 737 cells/µL (range 190-1122 cells/µL). Four weeks after switching to EVG/COBI/FTC/TDF, all subjects had HIV-1 RNA < 40 copies/mL in both BP and SP. Median EVG C24 h was 277 ng/mL (range 64.8-1790 ng/mL) in BP and 169 ng/mL (range 12.8-792 ng/mL) in SP. A significant correlation was observed between BP and SP EVG concentrations (Spearman rho 0.952; P < 0.001). The median SP:BP EVG concentration ratio was 0.39 (range 0.20-0.92). EVG C24 h in SP was at least 23-fold the in vitro protein-unbound 50% effective response (EC50 ) of HIV-1 clinical isolates (0.04-0.55 ng/mL). In all but one individual, EVG C24 h in SP was also higher than the blood plasma protein binding-adjusted 95% inhibitory concentration (IC95 ) of wild-type HIV-1 (45 ng/mL). CONCLUSIONS: Seminal EVG concentrations in HIV-infected men treated with EVG/COBI/FTC/TDF sufficed to contribute to maintaining HIV-1 RNA suppression in this compartment.

Broad autism spectrum and obsessive-compulsive symptoms in adults with the fragile X premutation.

OBJECTIVE: Clinical observations and a limited number of research studies provide evidence that the fragile X premutation may confer risk for autism, executive dysfunction, and psychopathology. The link to autism spectrum symptoms and social cognition deficits with the premutation remains uncertain, and thus was the focus of the present investigation. METHOD: Our sample included 131 individuals, 42 men/22 women with the FMR1 premutation (mean age = 31.83 ± 8.59 years) with a normal neurological exam, and 48 men/19 women healthy age-matched controls (mean age = 29.48 ± 7.29 years). Individuals completed a comprehensive neuropsychological battery with additional assessments for social cognition, broad autism spectrum, and obsessive-compulsive (OC) symptoms. RESULTS: Premutation carriers self-reported higher rates of autism-related symptoms (Autism Quotient; p = .001). Among males only, premutation carriers showed more atypical social interaction (p < .001) and stereotyped behavior (p = .014) during standardized clinical examination on the Autism Diagnostic Observation Schedule (ADOS) relative to controls. Female premutation carriers reported significantly higher rates of OC symptoms compared to control females (p = .012). Molecular measures defining the expanded premutation (FMR1 CGG repeat length and/or mRNA) were significantly associated with a measure of theory of mind (Reading the Mind in the Eyes Task). CONCLUSIONS: The results of this study indicate a higher rate of broad autism spectrum symptoms in some males with the premutation and provide evidence for an obsessive-compulsive subtype in female premutation carriers.

The influence of landscape configuration and environment on population genetic structure in a sedentary passerine: insights from loci located in different genomic regions.

The study of the factors structuring genetic variation can help to infer the neutral and adaptive processes shaping the demographic and evolutionary trajectories of natural populations. Here, we analyse the role of isolation by distance (IBD), isolation by resistance (IBR, defined by landscape composition) and isolation by environment (IBE, estimated as habitat and elevation dissimilarity) in structuring genetic variation in 25 blue tit (Cyanistes caeruleus) populations. We typed 1385 individuals at 26 microsatellite loci classified into two groups by considering whether they are located into genomic regions that are actively (TL; 12 loci) or not (NTL; 14 loci) transcribed to RNA. Population genetic differentiation was mostly detected using the panel of NTL. Landscape genetic analyses showed a pattern of IBD for all loci and the panel of NTL, but genetic differentiation estimated at TL was only explained by IBR models considering high resistance for natural vegetation and low resistance for agricultural lands. Finally, the absence for IBE suggests a lack of divergent selection pressures associated with differences in habitat and elevation. Overall, our study shows that markers located in different genomic regions can yield contrasting inferences on landscape-level patterns of realized gene flow in natural populations.

Multimycotoxin analysis in water and fish plasma by liquid chromatography-tandem mass spectrometry.

High performance liquid chromatography-mass spectrometry was used for the determination of 15 mycotoxins in water and fish plasma samples, including aflatoxins, fumonisins, ochratoxin A, sterigmatocistin, fusarenon-X and emerging Fusarium mycotoxins. In this work, dispersive liquid-liquid microextraction (DLLME) was assessed as a sample treatment for the simultaneous extraction of mycotoxins. Results showed differences in recovery assays when different extraction solvents were employed. Ethyl acetate showed better recoveries for the major part of mycotoxins analyzed, except for aflatoxins B2, G1 and G2, which showed better recoveries when employing chloroform as extractant solvent. Fumonisins and beauvericin exhibited low recoveries in both water and plasma. This method was validated according to guidelines established by European Commission and has shown to be suitable to be applied in dietary and/or toxicokinetic studies in fish where is necessary to check mycotoxin contents in rearing water and fish plasma.

Analysis of mycotoxins in coffee and risk assessment in Spanish adolescents and adults.

Mycotoxins are toxic compounds produced by fungal secondary metabolism that cause toxicological effects. Coffee is a highly popular beverage that is susceptible to contamination by mycotoxigenic fungi. The aim of the present study was to determine the presence of the following 21 mycotoxins in coffee using liquid chromatography tandem mass spectrometry (LC-MS/MS-IT): aflatoxin B1, B2, G1 and G2; ochratoxin A; nivalenol; deoxynivalenol; 3-acetyldeoxynivalenol; 15-acetyldeoxynivalenol; diacetoxyscirpenol; neosolaniol; T-2 and HT-2 toxin; sterigmatocystin; enniatin A, A1, B, and B1; beauvericin; and fumonisin B1 and B2. We aimed to determine differences by coffee process (coffee maker, electrical machine, soluble and traditional Turkish process) and to calculate the estimated daily intake (EDI) and risk assessment of mycotoxins from coffee consumption using deterministic approach at various scenarios of food consumption in Spanish adolescents and adults. The results demonstrate that all studied mycotoxins were detected in samples with mean concentrations ranging from 0.69 µg/kg to 282.89 µg/kg. Eleven percent of samples did not show contamination with legislated mycotoxins. Only 15-acetyldeoxynivalenol, deoxynivalenol, neosolaniol, fumonisin B1, and ochratoxin A exhibited significant differences between methods of coffee brewing. The results show that coffee intake does not represent a potential risk for consumers with respect to individual mycotoxin contamination.

Interactions of a standardized flavonoid fraction from Tilia americana with Serotoninergic drugs in elevated plus maze.

ETHNOPHARMACOLOGICAL RELEVANCE: Tilia americana var. mexicana (Schltdl) Hardin (Tiliaceae) aerial parts (bracts and flowers) are used in the traditional Mexican medicine to treat nervous disorders, as sedative and to treat insomnia. A fraction of this species called FC1 (organic fraction from this plant) was proposed, described as anxiolytic and characterized by the presence of flavonoids. In the present work, this fraction was standardized, and its interaction with different serotonergic drugs was tested. We used the elevated plus maze model as anxiety test and the open field test so as to observe a possible effect on mice׳s motor behavior. MATERIAL AND METHODOLOGY: HPLC technique was used to quantify the flavonoids contained in a fraction called F1C. Different doses of F1C were administered to ICR mice (12.5, 25, 37.5 and 50mg/kg, oral pathway) then they were exposed to elevated plus maze or open field test. After, each dose of F1C fraction was co-administered with different drugs, in order to evaluate the animal׳s behavior: DOI agonist (2.0mg/kg) and KET antagonist (0.03mg/kg) of 5-HT2A receptors; 8-OH-DPAT (0.1mg/kg) selective agonist and WAY100635 (0.5mg/kg) antagonist of 5HT1 receptors. RESULTS: The HPLC quantitative analysis revealed the F1C composition (mg/g of extract): tiliroside (28.56), glucoside of quercetin (16.25), quercitrin (7.96), rutin (3.93), Kaempferol (2.83). The Emax for F1C curve was 80.6% for time to open arms with an ED50 of 15.09 mg/kg. The combination of F1C with DOI gives a significant increase of the F1C anxiolytic effect (Emax=111% and ED50=13.51 mg/kg), while KET blocks it completely (Emax=12.25% and ED50=2.4 mg/kg). The administration of F1C with 8-OH-DPAT does not generate significant changes on the time to open arms, although it does induce a decrement in F1C potency (Emax=83.3% and ED50=33.3mg/kg). When F1C and WAY-100365 are combined, the anxiolytic activity of the fraction decreases (Emax=33.3% and ED50=102.10mg/kg). CONCLUSIONS: The medicinal use attributed to Tilia americana for their effect on central nervous system, could be in part in the flavonoid fraction (F1C) with anxiolytic activity which is dose dependent, and has the ability to interact with the serotonergic system. It is necessary to advance in the study of the mechanism of action, using other techniques such in vitro analysis.