An optimized FAIRE procedure for low cell numbers in yeast

We report an optimized low‐input FAIRE‐seq (Formaldehyde‐Assisted Isolation of Regulatory Elements‐sequencing) procedure to assay chromatin accessibility from limited amounts of yeast cells. We demonstrate that the method performs well on as little as 4 mg of cells scraped directly from a few colonies. Sensitivity, specificity and reproducibility of the scaled‐down method are comparable with those of regular, higher input amounts, and allow the use of 100‐fold fewer cells than existing procedures. The method enables epigenetic analysis of chromatin structure without the need for cell multiplication of exponentially growing cells in liquid culture, thus opening the possibility of studying colony cell subpopulations, or those that can be isolated directly from environmental samples.     

The effectiveness of ozone and acidulant treatments in extending the refrigerated shelf life of fresh-cut potatoes

Abstract: The objective of the study was to determine the effectiveness of acidulant dip treatments (with or without aqueous ozone) to reduce enzymatic browning and to extend the shelf life of fresh‐cut potato slices during refrigerated storage (4 °C) for 28 d. Potato slices subjected to aqueous ozone (2 ppm) had significantly (P≤ 0.05) higher L‐values and lower a‐values, but ozone did not appear to have any effect on aerobic plate counts (APCs) or polyphenol oxidase (PPO) activity. NatureSeal (NS) and sodium acid sulfate (SAS) were the most effective acidulant treatments in reducing browning (significantly [P≤ 0.05] higher L‐values, lower a‐values, and browning index values) regardless of ozone treatment. NS and SAS also had lower PPO activity compared to other treatments on days 0 and 28, and significantly (P≤ 0.05) lower APCs (≤2.00 log CFU/g) over refrigerated storage. Therefore, the SAS treatment was comparable to NS, a commercially available product, and showed promise as an effective antibrowning dip to reduce browning and spoilage in fresh‐cut potato products. Practical Application: A 1% SAS dip treatment which included 1% citric and 1% ascorbic acid was found to be an effective antibrowning dip for fresh‐cut potatoes along with NatureSeal®'s PS‐10, compared to other treatments. They were both effective in maintaining low microbial counts over refrigerated storage. Additionally, aqueous ozone washes (2 ppm) showed significant benefits to reduce browning; however, ozone did not affect microbial counts or PPO enzyme activity. Therefore, the SAS treatment could have potential use in the fruit and vegetable industry to reduce browning and spoilage in fresh‐cut potato products.     

Randomised trial investigating effect of a novel nicotine delivery device (Eclipse) and a nicotine oral inhaler on smoking behaviour, nicotine and carbon monoxide exposure, and motivation to quit

OBJECTIVE: To monitor the effect of a novel nicotine delivery device that may produce fewer carcinogens (Eclipse) on cigarette smoking, carbon monoxide and nicotine concentrations, and motivation to give up smoking. The smoker's own brand of cigarette and a nicotine replacement product (Nicotrol inhaler) were used as comparisons. DESIGN: After baseline data were recorded, smokers were randomised to either Eclipse or inhaler for two weeks and then switched to the other product for another two weeks. Thereafter a second baseline was obtained. SETTING AND PARTICIPANTS: Fifty smokers were included and data are reported for the 40 with complete data sets. The smokers were not trying to quit but were interested in trying a new product to reduce their risk. They visited a smoking clinic 10 times during the six week period of the trial. INTERVENTION: No counselling to aid reduction by Eclipse or inhaler was given. MAIN OUTCOME MEASURES: At each visit smoking status and carbon monoxide concentrations were recorded. In half of the visits withdrawal symptoms, attitudes towards smoking, heart rate, and blood nicotine concentrations were also recorded. RESULTS: Eclipse use decreased the number of cigarettes smoked per day (cpd) from 19.1 cpd at baseline to 2.1 cpd (p < 0.001), but increased carbon monoxide concentrations in parts per million (ppm) from 21.0 ppm to 33.0 ppm (p < 0.001). A similar decrease in cigarettes smoked per day was seen with the Nicotrol inhaler, from 19.1 cpd to 4.8 cpd (p < 0.001), but carbon monoxide decreased from 21.0 ppm to 12.7 ppm (p < 0.001). The blood nicotine concentration remained fairly stable with Eclipse, increasing slightly from 16.8 ng/ml to 18.0 ng/ml, while for the inhaler a significant drop was noted, from 16.8 ng/ml to 12. 2 ng/ml (p < 0.002). Craving and withdrawal did not increase with Eclipse. Few significant adverse events occurred with Eclipse. CONCLUSIONS: Eclipse can dramatically decrease cigarette consumption without causing withdrawal symptoms or decreases in nicotine concentrations or motivation to quit altogether. Unlike the inhaler, Eclipse produces an increase in carbon monoxide concentration. Thus Eclipse may not be a safer cigarette.     

The relationship between sensory quality and volatile compounds in raw juice processed from elderberries (Sambucus nigra L.)

Elderberry juice was processed by pressing of 101 samples of elderberries (Sambucus nigra L.) picked in Austria, England and Denmark at optimum maturity. Taste panels trained in determination of elderberry flavour evaluated the juices and the results from the sensory evaluation were correlated to the content of volatile compounds. The volatile compounds were isolated from elderberry juices by dynamic headspace technique and analysed by GC-FID and GC-MS. A total of 59 volatile compounds were identified, 9 of which have not previously been detected in elderberry products. A total of 53 compounds occurred in quantifiable amounts and significant differences among the investigated elderberry samples were observed for 50 compounds. The importance of the isolated volatile compounds to elderberry flavour was determined by classical factor analysis using the results from the sensory evaluation and information on the odour of the individual volatile compounds. Classical factor analysis were used including determination of eigenvalues, factor loadings and factor scores by varimax rotation of standardised variables for extraction of four non correlated factors (principal components) explaining approximately 73% of the variation in elderberry flavour. Twenty-eight volatile compounds and sensory quality were included in the four factors. Using the compounds with highest numerical values (loadings) the four factors were named: fresh-elderberry-elder flower (35.2%), fresh-fruity-sweet (17.3%), fresh-fruity (13.7%) and green-floral-fruity (6.7%). Three compounds with characteristic elderberry odour (-damascenone, nonanal, dihydroedulan) were closely correlated with the sensory quality of elderberry together with compounds with fruity, floral, green and/or miscellaneous odours.     

Effects of breed and a concentrate or grass silage diet on beef quality in cattle of 3 ages. I: Animal performance, carcass quality and muscle fatty acid composition

An increase in the intake of the n-3 series polyunsaturated fatty acids (PUFA) is recommended by nutritionists for the human diet and beef is a significant source of these fatty acids. Enhancing the n-3 PUFA content of beef is important in view of the generally saturated nature of fatty acids in ruminant meats and the potentially negative effect this can have on human health. This study examined the effects of breed and diet on the fatty acid composition of beef M. longissimus. Ninety-six steers were used, 48 Aberdeen Angus cross (AA) and 48 Holstein-Friesian (HF). At 6months of age, 3 groups were identified, to be slaughtered at 14, 19 and 24months, respectively. Each group consisted of eight steers of each breed fed on a concentrate or a grass silage diet, rich in n-6 and n-3 PUFA, respectively. The intake of the concentrate diet was restricted so that steers of each breed grew at a similar rate on each diet. The early maturing AA produced heavier, fatter carcasses with better conformation. Animals fed grass silage had higher carcass fatness and conformation scores and higher levels of neutral lipid and total lipid in muscle than those fed concentrate. When all animals were pooled, a decline in PUFA% as total muscle lipid increased was evident. Feeding a grass silage diet rich in α-linolenic acid (18:3n-3) increased levels of this fatty acid in muscle neutral lipid by a factor of about 3.0 compared with the concentrate diet, as well as enhancing the synthesis of the n-3 series long-chain C20-22 PUFA in the phospholipid fraction, including docosahexaenoic acid (DHA, 22:6n-3). In contrast, both levels and proportions of linoleic acid (18:2n-6) and the n-6 series C20-22 PUFA were higher in animals fed the concentrate diet. The proportions of 18:1trans and conjugated linoleic acid (CLA) in muscle neutral lipid were higher in animals fed concentrate compared with silage in all 3 groups. This was partly due to increased consumption of 18:2n-6. The ratio of PUFA to saturated fatty acids (P:S) in muscle was reduced by feeding grass silage, partly as the result of increased fat deposition. However, the increase in levels of n-3 series fatty acids with silage-feeding resulted in beneficially low n-6:n-3 ratios in muscle in all age groups (approximately 1.2 compared with 12.0 in the concentrate diet). Subtle breed differences in PUFA amounts and proportions were noted. Holstein-Friesians had higher proportions of PUFA and higher P:S ratios compared with AA, partly due to a higher proportion of phospholipid in total lipid. In phospholipid itself, HF in the 19 and 24months groups had higher proportions of most n-3 PUFA. In all age groups the ratio of DHA to its precursor, 18:3n-3 was higher in HF.     

The Influence of Ethanol on the Foaming Properties of Beer Protein Fractions: A Comparison of Rudin and Microconductivity Methods of Foam Assessment

The effect of ethanol on the foaming properties of beer protein fractions was studied using a microconductivity method and nitrogen gas to generate the foam. Increasing the ethanol concentration resulted in a decrease in foam stability. Interfacial studies including thin film drainage and dilational elasticity measurements indicated that ethanol reduced the rigidity of the adsorbed protein layer resulting in accelerated drainage from the foam lamellae and increased probability of film rupture. These results conflict with data from the Rudin method (using nitrogen gas to generate the foam) which indicate that, at low concentration, ethanol improves foam stability. These apparently conflicting results may be explained by the foam positive effects of a decline in bubble size and increase in bulk viscosity observed for the Rudin method, contrasted with the negative influence of a reduction in surface viscosity observed for the microconductivity foam assessment method.     

Persistent thrombin generation in humans during specific thrombin inhibition with hirudin

BACKGROUND: The degree to which antithrombotic drugs suppress thrombin generation is unknown. Because hirudin, unlike antithrombin III, binds intravascular thrombin rapidly and selectively to yield a circulating inactive complex of 3- to 4-hour half-life, we used intravenous hirudin in humans to investigate the course of thrombin generation during and early after anticoagulation with this potent, direct antithrombin. METHODS AND RESULTS: Intravascular thrombin was measured with an ELISA for the thrombin-hirudin complex formed during and for 18 hours after stopping a 6-hour infusion of hirudin at 0.1, 0.2, and 0.3 mg.kg-1.h-1 in three groups of six patients each. With free hirudin in 20- to 10,000-fold molar excess of thrombin and peak activated partial thromboplastin times of 2.3 to 3.0 times baseline, mean plasma thrombin-hirudin complex increased from 794 +/- 85 pg/mL (mean +/- SEM) 15 minutes after the start of the infusion to 1617 +/- 151 pg/mL at 6 hours of infusion to 2667 +/- 654 pg/mL at 24 hours. During the 24-hour observation period, plasma concentration of fragment 1.2 (the peptide released during conversion of prothrombin to thrombin) never fell below baseline but rather increased transiently during the hirudin infusion. Plasma concentrations of thrombin-antithrombin III complex (in ng/mL) decreased from 4.34 +/- 0.40 at baseline to 1.64 +/- 0.13 at 6 hours (P < .001) and gradually increased after stopping the infusion to 5.7 +/- 0.87 at 24 hours (nonsignificant compared with baseline). CONCLUSIONS: Measurement of thrombin-hirudin complex may be used as a marker of thrombin generation in humans. Persistent accumulation of thrombin-hirudin complex and generation of fragment 1.2 during and after completion of potent anticoagulation with hirudin suggest thrombin generation is not blocked by high-affinity thrombin inhibition. The persistent formation of thrombin during declining plasma levels of hirudin may contribute to the pathogenesis of rethrombosis early after antithrombin therapy or during inadequate anticoagulation.     

The perinatal mortality rate as an indicator of quality of care in international comparisons

Priming and recollection are expressions of human memory mediated by different brain events. These brain events were monitored while people discriminated words from nonwords. Mean response latencies were shorter for words that appeared in an earlier study phase than for new words. This priming effect was reduced when the letters of words in study-phase presentations were presented individually in succession as opposed to together as complete words. Based on this outcome, visual word-form priming was linked to a brain potential recorded from the scalp over the occipital lobe about 450 ms after word onset. This potential differed from another potential previously associated with recollection, suggesting that distinct operations associated with these two types of memory can be monitored at the precise time that they occur in the human brain.     

Growth of a human intestinal Desulfovibrio desulfuricans in continuous cultures containing defined populations of saccharolytic and amino acid fermenting bacteria

Ecological and physiological effects of the sulphate-reducing bacterium (SRB) Desulfovibrio desulfuricans on other intestinal organisms were investigated in anaerobic chemostats (dilution rate approximately 0.2 h-1). Reproducible defined bacterial communities were used in these experiments, comprising 14 different saccharolytic and amino acid fermenting species: Bifidobacterium longum, Bif. adolescentis, Bif. pseudolongum, Bif. infantis, Bacteroides thetaiotaomicron, Bact. vulgatus, Lactobacillus acidophilus, Enterococcus faecalis, Ent. faecium, Escherichia coli, Clostridium perfringens, Cl. butyricum, Cl. innocuum, Cl. bifermentans. Lactobacillus and Cl. bifermentans populations never rose above minimum detection limits (log10 2.0 and 4.0, respectively) under the experimental conditions employed in these studies. Inclusion of Des. desulfuricans in bacterial cultures (c. log10 8.4 viable cells ml-1) resulted in marked reductions (i.e. greater than 1 log) in planktonic cell population densities of several species, particularly Bif. longum, Cl. perfringens and Bif. pseudolongum. The two bacteroides species were unaffected by Des. desulfuricans, while numbers of Cl. butyricum increased. Extensive wall growth developed in the SRB culture, consisting mainly of Des. desulfuricans (log10 9.2 viable cells ml-1), Bact. thetaiotaomicron and Bact. vulgatus, with lesser numbers of facultative anaerobes, Cl. perfringens and Bif. longum. Wall growth was associated with a reduction in planktonic cell mass and increased acid production by the cultures. Chemotaxonomic study of chemostat microbiotas, on the basis of cellular fatty acid methyl ester (FAME) analyses, showed the existence of characteristic bacteroides (C15) and bifidobacterial (C18) markers, but desulfovibrio markers (i-C15:0, C16:0, i-C17:1) could be identified. The metabolic activities of saccharolytic organisms were altered in the SRB chemostat, including synthesis of a number of hydrolytic enzymes involved in carbohydrate breakdown, such as alpha-galactosidase, alpha-glucosidase and beta-galactosidase, together with several mucinolytic enzymes. High concentrations of sulphide (8.2 mmol 1-1) were detected in the SRB chemostat, suggesting that this metabolite may have been inhibitory to some species. Saccharolytic organisms growing in the SRB fermenter utilized more starch, but less galactose-containing polymers, which correlated with the observed glycosidase activities. Profound differences were also recorded with respect to fermentation product formation in the chemostats, where a major switch to acetate production occurred in the SRB culture, with concomitant reductions in propionate, butyrate and lactate, which is an important electron donor for desulfovibrios.