Food additives: Assessing the impact of exposure to permitted emulsifiers on bowel and metabolic health - introducing the FADiets study.

Emulsifiers are common components of processed foods consumed as part of a Western diet. Emerging in vitro cell-line culture, mouse model and human intestinal tissue explant studies have all suggested that very low concentrations of the food emulsifier polysorbate 80 may cause bacterial translocation across the intestinal epithelium, intestinal inflammation and metabolic syndrome. This raises the possibility that dietary emulsifiers might be factors in conditions such as coronary artery disease, type 2 diabetes and Crohn's disease. The potential mechanism behind the observed effects of this emulsifier is uncertain but may be mediated via changes in the gut microbiota or by increased bacterial translocation, or both. It is also unknown whether these effects are generalisable across all emulsifiers and detergents, including perhaps the natural emulsifier lecithin or even conjugated bile acids, particularly if the latter escape reabsorption and pass through to the distal ileum or colon. A major objective of the Medical Research Council (MRC)-funded Mechanistic Nutrition in Health (MECNUT) Emulsifier project is therefore to investigate the underlying mechanisms and effects of a range of synthetic and natural emulsifiers and detergents in vitro and in vivo, and to determine the effects of a commonly consumed emulsifier (soya lecithin) on gut and metabolic health through a controlled dietary intervention study in healthy human volunteers - the FADiets study. This report provides an overview of the relevant literature, discussing the impact of emulsifiers and other additives on intestinal and metabolic health, and gives an overview of the studies being undertaken as part of the MECNUT Emulsifier project.

Variable alterations of the microbiota, without metabolic or immunological change, following faecal microbiota transplantation in patients with chronic pouchitis.

Faecal microbiota transplantation (FMT) is effective in the treatment of Clostridium difficile infection, where efficacy correlates with changes in microbiota diversity and composition. The effects of FMT on recipient microbiota in inflammatory bowel diseases (IBD) remain unclear. We assessed the effects of FMT on microbiota composition and function, mucosal immune response, and clinical outcome in patients with chronic pouchitis. Eight patients with chronic pouchitis (current PDAI ≥7) were treated with FMT via nasogastric administration. Clinical activity was assessed before and four weeks following FMT. Faecal coliform antibiotic sensitivities were analysed, and changes in pouch faecal and mucosal microbiota assessed by 16S rRNA gene pyrosequencing and (1)H NMR spectroscopy. Lamina propria dendritic cell phenotype and cytokine profiles were assessed by flow cytometric analysis and multiplex assay. Following FMT, there were variable shifts in faecal and mucosal microbiota composition and, in some patients, changes in proportional abundance of species suggestive of a "healthier" pouch microbiota. However, there were no significant FMT-induced metabolic or immunological changes, or beneficial clinical response. Given the lack of clinical response following FMT via a single nasogastric administration our results suggest that FMT/bacteriotherapy for pouchitis patients requires further optimisation.

Relaxin-3 receptor (Rxfp3) gene deletion reduces operant sucrose- but not alcohol-responding in mice.

The pervasive use of refined sugars in highly accessible, palatable foods and persistent exposure to reinforcing food-associated cues has contributed to overconsumption of sugar-rich diets and the current obesity epidemic in Western society. We have shown previously that brain relaxin-3 mRNA levels positively correlate with sucrose and alcohol intake, and that central antagonism of relaxin-3 receptors (RXFP3) attenuates alcohol self-administration and alcohol-seeking in rats, but food-seeking behaviour and palatable food consumption in mice. To further examine the relationship between motivated appetitive behaviours and relaxin-3/RXFP3 signalling, we investigated the effect of Rxfp3 gene deletion in C57BL/6J mice on sucrose and alcohol self-administration and cue-induced reinstatement (RNST) of sucrose- and alcohol-seeking. Acquisition and maintenance of sucrose and alcohol self-administration was assessed in male wild-type (WT) and Rxfp3 knockout (KO) (C57BL/6J(RXFP3TM1) (/) (DGen) ) littermate mice using fixed ratio (FR) schedules of reinforcement. Mice were subsequently challenged with a progressive ratio (PR) test to measure motivation and, following extinction training, re-exposed to reward-associated cues to evaluate RNST of active lever-responding. Wild-type and Rxfp3 KO mice displayed similar acquisition of FR1 sucrose self-administration, but Rxfp3 KO mice responded less when the instrumental requirement was increased to FR3. These mice also showed a lower breakpoint for sucrose and attenuated cue-induced RNST of sucrose-seeking. Notably, no marked genotype differences in alcohol-responding were observed. In mice, endogenous relaxin-3/RXFP3 signalling promotes self-administration of sucrose under high response requirements and cue-induced RNST of sucrose-seeking, but does not apparently regulate motivation to consume alcohol or alcohol-seeking behaviour.

Review article: faecal transplantation therapy for gastrointestinal disease.

BACKGROUND: Evidence is emerging regarding the relationship between a dysbiosis of the human gut microbiota and a number of gastrointestinal diseases as well as diseases beyond the gut. Probiotics have been investigated in many gastrointestinal disease states, with variable and often modest outcomes. Faecal transplantation is an alternative approach to manipulate the gut microbiota. AIM: To review the use of faecal transplantation therapy for the management of gastrointestinal disorders. METHODS: Available articles on faecal transplantation in the management of gastrointestinal disorders were identified using a Pubmed search and bibliographies of review articles on the subject were collated. RESULTS: A total of 239 patients who had undergone faecal transplantation were reported. Seventeen of 22 studies of faecal transplantation were in fulminant or refractory Clostridium difficile. Studies of faecal transplantation are heterogeneous regarding the patients, donors, screening, methods of administration and definition of response. Faecal transplantation for C. difficile has been demonstrated to be effective in 145/166 (87%) patients. Small numbers of patients are reported to have undergone successful faecal transplantation for irritable bowel syndrome and inflammatory bowel disease. CONCLUSIONS: Faecal transplantation has been reported with good outcomes for fulminant and refractory C. difficile. No adverse effects of faecal transplantation have been reported. However, there are no level 1 data of faecal transplantation and reports to date may suffer from reporting bias of positive outcomes and under-reporting of adverse effects. This therapy holds great promise, where a dysbiosis of the gut microbiota is responsible for disease and further studies are necessary to explore this potential.

Evaluation of the effects of cage height and stocking density on the behaviour of laying hens in furnished cages.

1. Limited information is available on how changes in horizontal and vertical space within enriched or furnished layer cages (as defined by Directive 1999/74/EC) influence hen behaviour. This study evaluated the effects of varying minimum cage heights and space allowances on the behaviour of laying hens housed in furnished cages. It was conducted on two flocks of medium brown hybrid hens housed in furnished cages with access to perches and nest boxes on a semi-commercial scale at ADAS Gleadthorpe. 2. Flock 1 consisted of two layer strains (ISA Brown and Babcock 380), housed at two minimum cage heights (38 and 45 cm) and 5 stocking densities between 609 and 870 cm2/bird, with 12 replicates of each of the 20 strain/cage height/stocking density treatment combinations. Stocking density was varied by varying the number of birds per cage from 10 to 7 in standard full-width cages or housing 7 hens in a narrower cage. As a consequence stocking density, group size and trough width per bird co-varied for 4 out of 5 stocking density treatments. 3. Behaviour of flock 1 was sampled at 33 to 36, 46 and 68 weeks of age. At each age one top-tier, one middle-tier and one bottom-tier cage was sampled for each treatment. 4. Few behavioural differences due to cage treatments were detected. Hens at 870 cm2 had shorter feeding bouts than hens at 609 and 762 cm2. Yawning was more common in the cages with greater cage height. 5. Video recordings of flock 1 examined cage height effects on hens' use of vertical space and provided additional data on stretching and self-maintenance activities. No differences in behaviour between 38 and 45 cm cages were found except that scratching head was more common in cages with greater cage height. 6. Flock 2 consisted of two layer strains (Shaver Brown and Hy-Line Brown), housed at 38 and 45 cm and 609, 762 and 1016 cm2/bird, with 18 replicates of each of the 12 strain/cage height/stocking density treatment combinations. Stocking density was varied by housing 10, 8 or 6 hens in standard full-width cages. Behaviour of flock 2 was sampled at 30, 48, 60 and 67 weeks from video recordings. Three cages per treatment from middle-tiers only were sampled at each age. 7. Hens housed at 609 cm2/hen had the longest mean feeding bout, greater than for hens at 762 cm2/hen but not hens at 1016 cm2/hen. More unsuccessful attempts to reach the feeder and sideways and backwards displacements from the feeder occurred at 762 and 609 cm2/hen than at 1016 cm2/hen. A maximum of 8 hens were observed feeding synchronously. 8. These results suggest that changes in horizontal and vertical space over the ranges we studied had little effect on behaviour other than feeding behaviour. Specifying a minimum useable trough space per hen, rather than calculating feeder space from total length of feeder per cage, irrespective of accessibility, might help avoid crowding at the feeder and associated disturbance of feeding bouts.

A constructed microbial consortium for biodegradation of the organophosphorus insecticide parathion.

A consortium comprised of two engineered microorganisms was assembled for biodegradation of the organophosphate insecticide parathion. Escherichia coli SD2 harbored two plasmids, one encoding a gene for parathion hydrolase and a second carrying a green fluorescent protein marker. Pseudomonas putida KT2440 pSB337 contained a p-nitrophenol-inducible plasmid-borne operon encoding the genes for p-nitrophenol mineralization. The co-culture effectively hydrolyzed 500 microM parathion (146 mg l(-1)) and prevented the accumulation of p-nitrophenol in suspended culture. Kinetic analyses were conducted to characterize the growth and substrate utilization of the consortium members. Parathion hydrolysis by E. coli SD2 followed Michaelis-Menten kinetics. p-Nitrophenol mineralization by P. putida KT2440 pSB337 exhibited substrate-inhibition kinetics. The growth of both strains was inhibited by increasing concentrations of p-nitrophenol, with E. coli SD2 completely inhibited by 600 microM p-nitrophenol (83 mg l(-1)) and P. putida KT2440 pSB337 inhibited by 1,000 microM p-nitrophenol (139 mg l(-1)). Cultivation of the consortium as a biofilm indicated that the two species could cohabit as a population of attached cells. Analysis by confocal microscopy showed that the biofilm was predominantly comprised of P. putida KT2440 pSB337 and that the distribution of E. coli SD2 within the biofilm was heterogeneous. The use of biofilms for the construction of degradative consortia may prove beneficial.

Development of furnished cages for laying hens.

1. A 3-year trial was carried out of cages for laying hens, occupying a full laying house. The main cage designs used were 5000 cm2 in area, 50 cm high at the rear and furnished with nests and perches. F cages had a front rollaway nest at the side, lined with artificial turf. FD cages also had a dust bath containing sand over the nest. H cages had two nest hollows at the side, one in front of the other. They were compared with conventional cages 2500 cm2 in area and 38 cm high at the rear. 2. Cages were stocked with from 4 to 8 ISA Brown hens per cage, resulting in varied allowances of area, feeder and perch per bird. No birds were beak trimmed. In F and FD cages two further treatments were applied: nests and dust baths were sometimes fitted with gates to exclude birds from dust baths in the morning and from both at night; elevated food troughs, with a lip 33 cm above the cage floor, were compared with standard troughs. 3. Management of the house was generally highly successful, with temperature control achieved by ventilation. Egg production was above breeders' standards and not significantly affected by cage design. More eggs per bird were collected when there were fewer birds per cage but food consumption also then tended to be higher. 4. The number of downgraded eggs was variable, with some tendency for more in furnished cages. Eggs laid in dust baths were often downgraded. Those laid at the back of the cage were frequently dirty because of accumulation of droppings. H nests were unsuccessful, with less than 50% of eggs laid in the nest hollows. However, up to 93% of eggs were laid in front rollaways, and few of these were downgraded. 5. Feather and foot damage were generally less in furnished than in conventional cages, greater where there were more birds per cage. With an elevated food trough there was less feather damage but more overgrowth of claws. In year 2, mortality was greater in cages with more birds. 6. Pre-laying behaviour was mostly settled in front rollaway nests. Dust baths were used more for pecking and scratching than for dust bathing. Comfort behaviour was more frequent in furnished cages than conventional, although still not frequent. Locomotion was strongly affected by number of birds per cage or by space per bird, being reduced by crowding. Most birds perched at night except in one treatment providing only 10.7 cm perch per bird. 7. Behaviour was more unrestricted and varied, and physical condition was better, in furnished than in conventional cages. However, egg production will cost more in furnished cages, partly because more eggs are downgraded. Dust baths must be fitted with gates that the birds cannot open from outside, but gates for nest boxes were found unnecessary. If a low perch is fitted it must be far enough from the back of the cage for birds to walk there. 8. Where there was less space per bird (more birds per cage) than the requirements in the 1999 European Commission Directive on laying hens, there were: fewer eggs per hen, but still above the breeders' target; lower food consumption; more feather and foot damage, but less than in conventional cages; higher mortality in one trial out of three; less freedom of movement. However, the results were still very good even with 8 birds per cage, and support the principle that furnished cages provide an acceptable way of protecting the welfare of laying hens.

Spatial distribution and behaviour of laying hens housed in an alternative system.

The spatial distribution and behaviour of perchery housed laying hens were compared at a constant stocking density (18.5 birds/m(2)) in eight pens with colonies of five different sizes (323 birds (N=1), 374 birds (N=2), 431 birds (N=2), 572 birds (N=1) and 912 birds (N=2)). The birds were placed in the perchery when they were 12 weeks old. Observations began when they were 26 weeks old and continued at 8 week intervals until 61 weeks of age. Colony size did not appear to affect the spatial distribution of birds, but more standing behaviour and less feeding behaviour were observed in the smallest and largest colony sizes. Older birds spent more time on the floor areas and less time on perches. Young birds (26-28 weeks) spent more time feeding, foraging, drinking and preening, and less time standing idle than older birds. In the afternoons, there were fewer birds on the perches and more on the floor levels, corresponding with less time spent resting and more time spent performing active behaviours. Birds did not distribute themselves evenly throughout their pens: within specific areas of pens densities varied between 9 and 41 birds/m(2). This variation, which reflects the flux of birds from one part of the pen to another, was greatest for the larger colony sizes, and may have adverse implications for welfare in terms of crowding and hysteria.

Laying hens in large flocks in a perchery system: influence of stocking density on location, use of resources and behaviour.

1. ISA Brown laying hens (3000) were housed in a perchery in 10 pens, each with 300 birds. The pens varied in size to produce 4 different stocking densities: 9.9 birds/m2 (n = 3), 13.5/m2 (n = 2), 16.0/m2 (n = 2) and 19.0/m2 (n = 3). Observations began at 20 weeks of age and continued until 69 weeks to establish the spatial distribution of the birds, usage of the different resources and the expression of behaviour. 2. Overall, birds spent most time on the perch frame (47%), litter area (23%), slatted floor (17%) and nestbox area (9%). 3. There was no effect of density on the proportion of birds observed on the slatted floor or on the elevated perches but as density increased the proportion on the littered area decreased. 4. Space usage was determined vertically, horizontally and longitudinally. Individual birds were seen to use about 80% of the pen volume available to them. This value was similar for all densities and showed that individuals did not have separate home ranges. 5. Fewer vertical movements were made within the main perch frame at the upper than at the lower levels but movements between the perches of the main frame and the nestbox rails were relatively frequent. This may help birds move up and down through the main frame. 6. Behaviours which decreased in incidence with crowding included moving, foraging and dust-bathing. Behaviours which increased with crowding included standing. Behaviours which were unaffected included resting, preening, prelaying behaviour, comfort behaviour and the minor behaviours. 7. The proportion of birds engaged in feeding and drinking was unaffected by density, except each time the chain feeders (which operated intermittently) ran more hens were seen feeding at the lower densities. This suggests that food delivery stimulated feeding behaviour; there may have been some restriction at the higher densities on birds feeding when and where they wanted. 8. Stocking density had no effect on the frequency of agonistic interactions: threats, lunges, comb/head pecks, chases and fights. 9. The incidence of damaging pecking was low and not density dependent. 10. Increasing density within the range investigated inhibited the expression of a number of behaviours and limited the use of specific resources: bird welfare at 19 birds/m2 may have been very slightly impaired.

Egg shell colour is affected by laying cage design.

1. When laying hens are stressed some retain their eggs in the shell gland beyond the normal time of laying and this can result in the deposition of extra-cuticular calcium which makes brown eggs appear paler. 2. Three different types of enriched modified cage were compared: the location where eggs were laid was recorded and shell colour was measured using a reflectometer. 3. In 2 types of cage with enclosed nest boxes more eggs (80%) were laid in the nests than in a design with nest hollows in the open part of the cage (41%). 4. The eggs from the cages with enclosed nests were darker (had less extraneous calcium) than those with open nest hollows. This implies that in the designs with nest boxes fewer eggs had been retained and the hens may have been less stressed. 5. The results support previous evidence that to reduce stress and improve welfare it is desirable to provide enclosed nest sites for caged laying hens.

Measurement of aluminium in clinical samples.

Measurement of aluminium in clinical samples is important for the protection of patients with chronic renal failure, and for investigations of dementia. The types of specimen that need to be analysed include body fluids and tissues, dialysis fluids, water and foods and the concentrations that may be found can range from around 1 microgram/L to several mg/L. Although techniques must afford exceptional sensitivity they will also be susceptible to the effects of external contamination and it is imperative that those involved with the analysis, from sample collection through to the determination, ensure that precautions are taken to prevent undue addition of the metal. Of the methods that may be used ETAAS is clearly the most appropriate. Great care is required to achieve reliable performance but with experienced personnel very good results can be obtained. Regular reviews of recent analytical developments are presented in the Atomic Spectrometry Updates series and these are recommended to those who may wish to look for further information.

Diagnostic significance of octadeca-9,11-dienoic acid in cervical neoplasia.

The percentage molar ratio (%MR) of the 9,11 and 9,12 isomers of octadecadienoic acid was determined in cervical exfoliated cells from 148 subjects, of whom 27 had cytologically proven intraepithelial neoplasia and in cervical biopsy specimens from 43 subjects, of whom 24 had histologically diagnosed cervical intraepithelial neoplasia. The %MR in both cervical biopsy specimens and exfoliated cells did not significantly differ in subjects with or without cervical intraepithelial neoplasia. The measurement of the %MR of 9,11:9,12 octadecadienoic acid has no role in the detection of cervical intraepithelial neoplasia.

Intestinal alkaline phosphatase activity in relation to age in humans.

Early studies of a few subjects suggested that intestinal alkaline phosphatase (IAP) activities in children 5 years of age or less were higher than in older individuals. To further investigate this finding, the IAP and disaccharidase activities of 298 subjects (133 were 5 years of age or less) with normal intestinal histology were assayed. Ninety-five of the children had serum alkaline phosphatase determined. The youngest individual with a low lactase activity was 5 years of age, which supported the earlier findings. When the whole population was tested, there was no correlation between the intestinal and serum alkaline phosphatase values. The mean IAP activity of subjects 1 year old and less was greater than in older individuals, but there was greater statistical dispersion and the data were not normally distributed. When studying the natural logarithm of the data, a wide dispersion of values about the mean in the 0 to 3 year-old age group was observed. This qualitative behavior is characteristic of functions involving the base of the natural logarithm and of processes that "age" in a simple way.

Determination of aluminium in serum: findings of an external quality assessment scheme.

The analytical performance of laboratories participating in the serum aluminium programme of the Guildford Trace Element Quality Assessment Scheme has been evaluated. The considerable disparity between reported results was not attributable to the nature of the samples distributed, nor to the standard aluminium solutions used in different laboratories. All laboratories used electrothermal atomic absorption spectrometry but sample preparation procedures and temperature programmes varied enormously. While some laboratories performed better than others in the scheme, and the standard of analysis appears to be improving with time, no laboratory showed consistent acceptable performance.

Enzymic determination of plasma cholesterol on discrete automatic analysers.

Enzymic procedures for the determination of plasma cholesterol, using cholesterol esterase and cholesterol oxidase, have been adapted to the Vickers D-300, Vickers M,-300, and Vitatron AKES discrete analysers. The results obtained by these methods have been compared to those obtained by manual and continuous flow Liebermann-Burchard methods. The enzymic methods were found to be accurate, precise and of adequate sensitivity.