Application of a Rapid and Sensitive Method for Hormonal and Vitamin E Profiling Reveals Crucial Regulatory Mechanisms in Flower Senescence and Fruit Ripening

Knowledge of ripeness and regulation of postharvest processes is an important tool to prevent loss of commercial value in both fruit and cut flower markets. The joint analysis of hormones and vitamin E levels can reveal complex interactions between hormones and oxidative stress as key regulators of postharvest processes. Profiling of both groups of metabolic compounds was performed during the ripening of non-climacteric fruits (red raspberry, Rubus idaeus L.) and senescence of ethylene-insensitive flowers (Dutch Iris, Iris x hollandica L.). After an initial extraction of the sample, without further purification steps, the hormonal profile was analyzed by UPLC-MS/MS and vitamin E levels were measured by HPLC. This methodological approach was very fast and had enough sensitivity for the analysis of small samples. Raspberry fruit maturation was characterized by a decline of cytokinin levels [zeatin, zeatin riboside, 2-isopentenyl adenine, and isopentenyl adenosine (Z, ZR, 2-iP, and IPA, respectively)] and gibberellins (GA₁ in particular). Exogenous application of ABA prevented δ-tocopherol loss during fruit ripening. Iris floral senescence was also under strict hormonal control, also mediated by cytokinins and gibberellins. Z, ZR, 2-iP, GA₉, and GA₂₄ levels decreased in inner tepals, whereas the level of IPA decreased in style-merged-to-stigma tissues, thus suggesting tissue-specific roles for different hormones. α-Tocopherol levels decreased during senescence of inner tepals, hence suggesting enhanced oxidative stress. In conclusion, the rapid and sensitive hormonal and vitamin E profiling presented here can help in understanding the key physiological processes underlying fruit ripening and floral senescence.     

Hybridisation between oilseed rape (Brassica napus) and tetraploid Brassica rapa under field conditions

Cross-compatible relatives of crop species contribute to the uncertainty regarding the potential risk of transgene escape from genetically modified varieties. The most successful crossing partner of oilseed rape (Brassica napus L.) is diploid Brassica rapa L. Variation of ploidy level among B. rapa cultivars has, until recently, been neglected in the context of gene flow and hybridisation with oilseed rape. We estimated the extent of hybridisation between autotetraploid B. rapa varieties (female) and B. napus (pollen donor) under experimental field conditions. Morphology, variation of relative DNA amount, and microsatellite markers were used to distinguish between intraspecific offspring of tetraploid B. rapa and interspecific hybrids with B. napus. Of 517 seed progenies of tetraploid B. rapa, 45 juvenile plants showed species specific morphological traits of oilseed rape. The detection of putative hybrids based on variation in relative DNA amounts was problematic due to the occurrence of aneuploidy. In total, 84 offspring showed relative DNA amounts deviating from tetraploid B. rapa, four of which were hexaploids. Of the 205 offspring analysed at three microsatellite loci, 67 had oilseed rape alleles. Based on molecular evidence a minimum hybridisation rate of 13.0% was estimated. A few mother plants accounted for the majority of hybrids. The mean pollen viability of hybrids between B. napus and tetraploid B. rapa (80.6%) was high in comparison with mean pollen viability of triploid hybrids between B. napus and diploid B. rapa. Therefore, the occurrence of tetraploid B. rapa should be taken into consideration when estimating the likelihood of gene flow from oilseed rape to close relatives at the landscape level. Tetraploid B. rapa is a common component of several seed mixtures and establishes feral populations in northwest Germany. Assuming a similar abundance of diploid and tetraploid B. rapa, gene flow from B. napus to tetraploid may be more likely than gene flow to diploid B. rapa.     

A Comparative pO2 Probe and [18F]-Fluoro-Azomycinarabino-Furanoside ([18F]FAZA) PET Study Reveals Anesthesia-Induced Impairment of Oxygenation and Perfusion in Tumor and Muscle

Tumor hypoxia can be identified by [¹⁸F]FAZA positron emission tomography, or invasively using oxygen probes. The impact of anesthetics on tumor hypoxia remains controversial. The aim of this comprehensive study was to investigate the impact of isoflurane and ketamine/xylazine anesthesia on [¹⁸F]FAZA uptake and partial oxygen pressure (pO₂) in carcinoma and muscle tissue of air- and oxygen-breathing mice.

Methods

CT26 colon carcinoma-bearing mice were anesthetized with isoflurane (IF) or ketamine/xylazine (KX) while breathing air or oxygen (O₂). We performed 10 min static PET scans 1 h, 2 h and 3 h after [¹⁸F]FAZA injection and calculated the [¹⁸F]FAZA-uptake and tumor-to-muscle ratios (T/M). In another experimental group, we placed a pO₂ probe in the tumor as well as in the gastrocnemius muscle to measure the pO₂ and perfusion.

Results

Ketamine/xylazine-anesthetized mice yielded up to 3.5-fold higher T/M-ratios compared to their isoflurane-anesthetized littermates 1 h, 2 h and 3 h after [¹⁸F]FAZA injection regardless of whether the mice breathed air or oxygen (3 h, KX-air: 7.1 vs. IF-air: 1.8, p = 0.0001, KX-O₂: 4.4 vs. IF-O₂: 1.4, p < 0.0001). The enhanced T/M-ratios in ketamine/xylazine-anesthetized mice were mainly caused by an increased [¹⁸F]FAZA uptake in the carcinomas. Invasive pO₂ probe measurements yielded enhanced intra-tumoral pO₂ values in air- and oxygen-breathing ketamine/xylazine-anesthetized mice compared to isoflurane-anesthetized mice (KX-air: 1.01 mmHg, IF-air: 0.45 mmHg; KX-O₂ 9.73 mmHg, IF-O₂: 6.25 mmHg). Muscle oxygenation was significantly higher in air-breathing isoflurane-anesthetized (56.9 mmHg) than in ketamine/xylazine-anesthetized mice (33.8 mmHg, p = 0.0003).

Conclusion

[¹⁸F]FAZA tumor uptake was highest in ketamine/xylazine-anesthetized mice regardless of whether the mice breathed air or oxygen. The generally lower [¹⁸F]FAZA whole-body uptake in isoflurane-anesthetized mice could be due to the higher muscle pO₂-values in these mice compared to ketamine/xylazine-anesthetized mice. When performing preclinical in vivo hypoxia PET studies, oxygen should be avoided, and ketamine/xylazine-anesthesia might alleviate the identification of tumor hypoxia areals.     

Evaluation of the Becton Dickinson Rapid Influenza Diagnostic Tests in Outpatients in Germany during Seven Influenza Seasons

BackgroundAn extensive retrospective study spanning several seasons was undertaken to evaluate the diagnostic performance of the BD rapid influenza diagnostic test (RIDT) in comparison with the RT-PCR assay.MethodsA total of 2,179 respiratory samples were tested in parallel by in-house RT-PCR and the RIDT. During the 2003-2004, 2006-2007, 2007-2008, and 2008-2009 (n=1671) seasons, the BD Directigen Flu A+B test was used, and during the 2010-2011, 2011-2012 and 2012-2013 (n=508) seasons, the BD Directigen EZ Flu A+B test b was used.ResultsThe sensitivity, specificity, PPV and NPV for the BD Directigen Flu A+B test calculated for types A and B together were 39%, 99%, 98%, and 56%, respectively. For the BD Directigen EZ Flu A+B test, these values were 47%, 100%, 100%, 55%, respectively. The sensitivity of the BD Directigen Flu A+B test did not differ significantly from season to season or between types A (44%) and B (37%). The sensitivity of the BD Directigen EZ Flu A+B test calculated for type A only was 59%, which was considerably higher than the sensitivity of this test for type B (23%). The sensitivity of the RIDT was approximately 40-50% in children and teenagers, but it was only 18.% in adults aged 20 years and older. The specificity of both RIDTs was very high (>99%) during all seasons.ConclusionsDue to their rapid turnaround time, RIDTs can help guide decisions about the clinical management of influenza. Because of the high specificity, a positive result can be interpreted as a true positive, and antiviral therapy as well as appropriate measures to prevent the transmission of influenza can be initiated. The best sensitivity of the RIDT is achieved in children. However, even in this group, the RIDT will only recognize influenza infection in approximately half of the cases, and influenza should still be considered in patients with negative results; negative RIDT results must be confirmed by PCR.     

Orally Disintegrating Films and Mini-Tablets—Innovative Dosage Forms of Choice for Pediatric Use

Oral drug delivery is a non-invasive and therefore a very convenient route of administration. Orally disintegrating dosage forms, like soluble films and (mini-)tablets, appear promising for use in the pediatric population. New guidance for the development of pediatric medicines has been published, which provides considerations on how pediatric products should be designed. However, most of the considerations leave a lot of room for interpretations. Bearing in mind the different aspects discussed in the latest guideline, the use of orally disintegrating films and tablets, in particular, small-sized tablets, is discussed and reflected upon by providing evidence from the scientific literature. The available dosage forms for children are various and examples of currently licensed products for use in the pediatric population were compiled. Aspects such as the appropriateness for pediatrics, the choice of excipients, the opportunities for modified drug release preparations or fixed-dose combinations, the acceptability and palatability, and also limitations were discussed with respect to the new dosage forms of orally disintegrating films and mini-tablets. This paper points out that innovation in pediatric medicines are planned and should be encouraged; however, supported by the regulatory guidance, only general considerations are provided. Nevertheless, the guideline summarizes multiple points to consider during the development of medicines for pediatric use. Considering the scientific evidence and the regulatory guidance, orally disintegrating dosage forms, like soluble films and (mini-)tablets, offer an innovative solution for pediatric drug delivery.KEY WORDS: children, orally disintegrating dosage forms, pediatric drug delivery, pediatrics, orodispersible films and tablets     

Informed Choice in the German Mammography Screening Program by Education and Migrant Status: Survey among First-Time Invitees

Breast cancer is the most prevalent cancer among women and mammography screening programs are seen as a key strategy to reduce breast cancer mortality. In Germany, women are invited to the population-based mammography screening program between ages 50 to 69. It is still discussed whether the benefits of mammography screening outweigh its harms. Therefore, the concept of informed choice comprising knowledge, attitude and intention has gained importance. The objective of this observational study was to assess the proportion of informed choices among women invited to the German mammography screening program for the first time. A representative sample of 17,349 women aged 50 years from a sub-region of North Rhine Westphalia was invited to participate in a postal survey. Turkish immigrant women were oversampled. The effects of education level and migration status on informed choice and its components were assessed. 5,847 (33.7%) women responded to the postal questionnaire of which 4,113 were used for analyses. 31.5% of the women had sufficient knowledge. The proportion of sufficient knowledge was lower among immigrants and among women with low education levels. The proportion of women making informed choices was low (27.1%), with similar associations with education level and migration status. Women of low (OR 2.75; 95% CI 2.18–3.46) and medium education level (OR 1.49; 95% CI 1.27–1.75) were more likely to make an uninformed choice than women of high education level. Turkish immigrant women had the greatest odds for making an uninformed choice (OR 5.30, 95% CI 1.92–14.66) compared to non-immigrant women. Other immigrant women only had slightly greater odds for making an uninformed choice than non-immigrant women. As immigrant populations and women with low education level have been shown to have poor knowledge, they need special attention in measures to increase knowledge and thus informed choices.     

Regulation of neuronal excitability by release of proteins from glial cells

Effects of glial cells on electrical isolation and shaping of synaptic transmission between neurons have been extensively studied. Here we present evidence that the release of proteins from astrocytes as well as microglia may regulate voltage-activated Na⁺ currents in neurons, thereby increasing excitability and speed of transmission in neurons kept at distance from each other by specialized glial cells. As a first example, we show that basic fibroblast growth factor and neurotrophin-3, which are released from astrocytes by exposure to thyroid hormone, influence each other to enhance Na⁺ current density in cultured hippocampal neurons. As a second example, we show that the presence of microglia in hippocampal cultures can upregulate Na⁺ current density. The effect can be boosted by lipopolysaccharides, bacterial membrane-derived stimulators of microglial activation. Comparable effects are induced by the exposure of neuron-enriched hippocampal cultures to tumour necrosis factor-α, which is released from stimulated microglia. Taken together, our findings suggest that release of proteins from various types of glial cells can alter neuronal excitability over a time course of several days. This explains changes in neuronal excitability occurring in states of thyroid hormone imbalance and possibly also in seizures triggered by infectious diseases.     

Whose trait is it anyways? Coevolution of joint phenotypes and genetic architecture in mutualisms

Evolutionary biologists typically envision a trait’s genetic basis and fitness effects occurring within a single species. However, traits can be determined by and have fitness consequences for interacting species, thus evolving in multiple genomes. This is especially likely in mutualisms, where species exchange fitness benefits and can associate over long periods of time. Partners may experience evolutionary conflict over the value of a multi-genomic trait, but such conflicts may be ameliorated by mutualism’s positive fitness feedbacks. Here, we develop a simulation model of a host–microbe mutualism to explore the evolution of a multi-genomic trait. Coevolutionary outcomes depend on whether hosts and microbes have similar or different optimal trait values, strengths of selection and fitness feedbacks. We show that genome-wide association studies can map joint traits to loci in multiple genomes and describe how fitness conflict and fitness feedback generate different multi-genomic architectures with distinct signals around segregating loci. Partner fitnesses can be positively correlated even when partners are in conflict over the value of a multi-genomic trait, and conflict can generate strong mutualistic dependency. While fitness alignment facilitates rapid adaptation to a new optimum, conflict maintains genetic variation and evolvability, with implications for applied microbiome science.