Selective pressure acting on influenza virus neuraminidase protein and relation with development of resistance to antiviral drugs

Neuraminidase (NA) protein of influenza viruses has the particularity of being under antibody and antiviral drug selective pressure, as it is one of the main surface antigens and the target of neuraminidase inhibitors(NAIs). The aim of this study is to investigate the selective pressure(SP) acting on the NA of seasonal and pandemic influenza viruses. It comprises two objectives: (a)to evaluate the contribution of positive SP for the emergence of NAIs resistant viruses; and (b)to determine the impact of NAIs introduction into clinic and its wide use during pandemic on the SP acting on NA. For the 1st objective it will be analysed the SP acting on the sites associated with NAIs resistance or reduction in susceptibility. The 2nd objective implies a differential evolutionary pressure analysis according to time, with 3 sub-datasets of NA sequences being considered: (1)before worldwide introduction of NAIs into clinic(1999); (2)before wide use of oseltamivir during A(H1N1)2009 pandemic(2009); and (3)from 2009 to date. A large dataset of full-length NA coding sequences will be used for each (sub)type/variant, comprising sequences obtained at national level(since 2000/2001) and sequences available at GISAID and NCBI. A(H1N1)seasonal dataset was already created, including a total of 1523 sequences, from which 94 belong to 1st sub-dataset, 1094 to 2nd and 335 to 3rd. All SP analysis will be performed using the expertise acquired with this workshop. This study may contribute for understanding the role of antiviral drug selective pressure in NAIs resistance, patterns of emergency of resistant viruses and NA evoluti

Evaluation and Characterization of Influenza Antiviral Drug Resistance in Portugal: Major Results and Achievements of a 5-Year Study

In 2007 started to be carried out for the first time in Portugal a study focused on influenza antiviral drug resistance. Three main objectives were established:(1)to determine the antiviral profile of influenza viruses to oseltamivir, zanamivir and amantadine;(2)to determine and monitor the baseline level of susceptibility along winter seasons and for each influenza sub(type);(3)to analyse and characterize the whole genome of viruses that showed phenotypic levels of inhibition to neuraminidase inhibitors(NAIs). NAIs profile was determined phenotypically, using a fluorescence MUNUNA assay, and genotypically by NA and HA sequencing. A total of 340 seasonal viruses(117 A(H3N2),93 A(H1N1),130 B) were tested for oseltamivir and of 297(112 A(H3N2),68 A(H1N1),117 B) for zanamivir. Additionally, 142 A(H1N1)pdm09 viruses were evaluated for both NAIs. Whole genome sequencing was performed in 27 of the A(H1N1)pdm09 viruses. Amantadine profile was determined through M2 pyrosequencing or conventional sequencing in a total of 205 seasonal A viruses(138 A(H3N2),84 A(H1N1)) and of 117 A(H1N1)pdm09 viruses. Main results are: -Resistance to oseltamivir in 27 A(H1N1) seasonal viruses(29%,N=93) from 2007/2008 and 2008/2009 and in one A(H1N1)pdm09 virus(0.7%,N=142) from 2010/2011. These viruses exhibited a highly reduced level of inhibition to oseltamivir by phenotypic analysis (170-650 IC50 fold-change) and NA H275Y mutation; -One suspected case of clinical resistance to oseltamivir with a mixed population of H275Y viruses(73,8%H275,26.2%Y275); -No resistance to zanamivir; -Dual reduced susceptibility to oseltamivir and zanamivir in one B virus(0,85%,N=117) and in two A(H1N1)pdm09 viruses(1,41%,N=142). These viruses exhibited a 2-4 IC50 fold-change level of inhibition to both NAIs. A mixed population of D197N viruses was found in the B virus(56%D197,44%N197) and the two A(H1N1)pdm09 viruses shared NA I223V and PB2 V480I mutations; -Resistance to amantadine in 49 A(H3N2) viruses(35,5%,N=138) from 2005/2006 to 2008/2009(46 S31N,3 S31N+V27A), and in all A(H1N1)pdm09 viruses(S31N). This 5-year study allowed to establish a technical platform for influenza antiviral drug resistance evaluation, to timely detect the emergence of resistant viruses, to acquire know-how on the natural variation of virus susceptibility, and to contribute for the management of cases suspected of clinical resistance. Additionally, it allowed the gathering of a large amount of data that will be used in more advanced studies, focused on evolutionary analysis and on detailed characterization of specific mutations

Estresse Salino Na Qualidade Fisiológica De Sementes De Cryptostegia Madagascariensis Bojer Ex Decne

The Cryptostegia madagascariensis Bojer ex Decne. species was introduced in Brazil with ornamental purposes, but currently, it is invasive in the Caatinga biome. Despite its high level of occupancy of altered areas of Caatinga, there is lack of information about this species, particularly in relation to the capacity of its seeds to germinate in adverse conditions. The objective of this study was to evaluate seed germination and the initial seedling growth of Cryptostegia madagascariensis in function of salinity at different temperatures. The experiment was conducted following a completely randomized design, with treatments arranged in a 6 × 3 factorial design with six salinity levels promoted by sodium chloride (NaCl): 0.0 (control); 2.0; 4.0; 6.0; 8.0 and 10.0 dS m-1 and three temperatures: 25 and 30°C constant and alternating 20-35°C. The physiological quality of seeds was evaluated using the following variables: germination, first count and the speed of germination index, length and dry weight of roots and hypocotyl. The results indicate that there is high probability of seed germination of Cryptostegia madagascariensis subjected to salt stress at constant temperature of 30°C and alternating 20-35°C, which enhances its invasive character. © 2016, Universidade Federal de Santa Maria. All rights reserved.2641189119

Caloric restriction induces energy-sparing alterations in skeletal muscle contraction, fiber composition and local thyroid hormone metabolism that persist during catch-up fat upon refeeding.

Weight regain after caloric restriction results in accelerated fat storage in adipose tissue. This catch-up fat phenomenon is postulated to result partly from suppressed skeletal muscle thermogenesis, but the underlying mechanisms are elusive. We investigated whether the reduced rate of skeletal muscle contraction-relaxation cycle that occurs after caloric restriction persists during weight recovery and could contribute to catch-up fat. Using a rat model of semistarvation-refeeding, in which fat recovery is driven by suppressed thermogenesis, we show that contraction and relaxation of leg muscles are slower after both semistarvation and refeeding. These effects are associated with (i) higher expression of muscle deiodinase type 3 (DIO3), which inactivates tri-iodothyronine (T3), and lower expression of T3-activating enzyme, deiodinase type 2 (DIO2), (ii) slower net formation of T3 from its T4 precursor in muscles, and (iii) accumulation of slow fibers at the expense of fast fibers. These semistarvation-induced changes persisted during recovery and correlated with impaired expression of transcription factors involved in slow-twitch muscle development. We conclude that diminished muscle thermogenesis following caloric restriction results from reduced muscle T3 levels, alteration in muscle-specific transcription factors, and fast-to-slow fiber shift causing slower contractility. These energy-sparing effects persist during weight recovery and contribute to catch-up fat

A new measure of health motivation influencing food choices and its association with food intakes and nutritional biomarkers in European adolescents

Objective:To develop a scale to assess health motivation influencing food choices and to explore its performance in the associations with food intakes and nutritional biomarkers. Design:Psychometric study using cross-sectional self-report questionnaires and nutritional biomarkers. Setting:Multi-centre investigation conducted in ten European cities. Participants:2954 adolescents who were included in the HELENA study and completed the Food Choices and Preferences (FCP) questionnaire. Results:Nineteen out of 124 items of the FCP questionnaire were in the same dimension. Sixteen presented adequate parameters for the Scale of evaluatiOn of Food choIcEs (SOFIE). The scores were positively associated with the intakes of cereals, dairy products, meats and eggs, and fish, as well as with blood concentrations of vitamin C, ß-carotene, n-3 fatty acids, cobalamin, holo-transcobalamin and folate; scores were negatively associated with the intake of alcohol. Conclusions:SOFIE can improve the assessment of motivation influencing food choices based on items with the best performance and is proposed as a new measure to health-related studies

Measuring nutritional knowledge using Item Response Theory and its validity in European adolescents

Objective: To analyse the Nutritional Knowledge Test (NKT) using Item Response Theory (ITR) analysis and to assess the construct validity of the Nutritional Knowledge Scale (NKTS) and its associations with adolescent food group consumption and nutritional biomarkers. DesignCross-sectional study: SettingMulticentre investigation conducted in ten European cities.ParticipantsAdolescents aged 12·5-17·5 years (n 3215) who completed over 75 % of the NKT. Results: Factor analysis indicated that the NKT can be analysed with a one-dimensional model. Eleven out of twenty-three items from the NKT presented adequate parameters and were selected to be included in the NKTS. Nutrition knowledge was positively associated with consumption of fruits, cereals, dairy products, pulses, meat and eggs, and fish, as well as with blood concentrations of vitamin C, ß-carotene, n-3 fatty acids, holo-transcobalamin, cobalamin and folate; nutrition knowledge was negatively associated with intake of olives and avocado, alcohol and savoury snacks. Conclusions: The NKTS assessed nutritional knowledge adequately and it is proposed as a new tool to investigate this subject in future studies

Standard addition method based on four-way PARAFAC decomposition to solve the matrix interferences in the determination of carbamate pesticides in lettuce using excitation–emission fluorescence data

The simultaneous determination of two carbamate pesticides (carbaryl and carbendazim) and of the degradation product of carbaryl (1-naphthol) in iceberg lettuce was achieved by means of PARAFAC decomposition and excitation–emission fluorescence matrices. A standard addition method for a calibration based on four-way data was applied using different dilutions of the extract from iceberg lettuce as a fourth way that provided the enough variation of the matrix to carry out the four-way analysis. A high fluorescent overlapping existed between the three analytes and the fluorophores of the matrix. The identification of two fluorescent matrix constituents through the four-way model enabled to know the matrix contribution in each dilution of the extract. This contribution was subtracted from the previous signals and a subsequent three-way analysis was carried out with the tensors corresponding to each dilution. The PARAFAC decomposition of these resulting tensors showed a CORCONDIA index equal to 99%. For the identification of the analytes, the correlation between the PARAFAC spectral loadings and the reference spectra has been used. The trueness of the method, in the concentration range studied, was guaranteed because there was neither constant nor proportional bias according to the appropriate hypothesis tests. The best recovery percentages were obtained with the data from the most diluted extract, being the results: 127.6% for carbaryl, 125.55% for carbendazim and 87.6% for 1-naphthol. When the solvent calibration was performed, the decision limit (CCα) and the capability of detection (CCβ) values, in x0=0, were 2.21 and 4.38 μg L−1 for carbaryl, 4.87 and 9.64 μg L−1 for carbendazim; and 3.22 and 6.38 μg L−1 for 1-naphthol, respectively, for probabilities of false positive and false negative fixed at 0.05. However, these values were 5.30 and 10.49 μg L−1 for carbaryl, 18.05 and 35.73 μg L−1 for carbendazim; and 1.92 and 3.79 μg L−1 for 1-naphthol, respectively, when the matrix-matched calibration using the most diluted extract was carried out in the recovery study.Ministerio de Economía y Competitividad(CTQ2011-26022) and JuntadeCastillayLeón(BU108A11-2)