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Declines in species diversity carry profound implications for ecosystem functioning. Communities of primary producers and consumers interact on evolutionary as well as ecological time scales, shaping complex relationships between biodiversity and ecosystem functioning. In subsidized ecosystems, resource inputs are independent of consumer actions, offering a simplified view of the relationship between species diversity and function for higher trophic levels. With food webs supported by substantial but variable inputs of detritus from adjacent marine ecosystems, sandy beaches are classic examples of subsidized ecosystems. We investigated effects of consumer species diversity and identity on a key ecological function, consumption of kelp wrack from nearshore giant kelp (Macrocystis pyrifera) forests. We assessed effects of species richness on kelp consumption by experimentally manipulating richness of six common species of invertebrate detritivores in laboratory mesocosms and conducting field assays of kelp consumption on beaches. Consumer richness had no effect on kelp consumption in the field and a slight negative effect in laboratory experiments. Kelp consumption was most strongly affected by the species composition of the detritivore community. Species identity and body size of intertidal detritivores drove variation in kelp consumption rates in both experiments and field assays. Our results provide further evidence that species traits, rather than richness per se, influence ecosystem function most, particularly in detrital-based food webs with high functional redundancy across species. On sandy beaches, where biodiversity is threatened by rising sea levels and expanding development, our findings suggest that loss of large-bodied consumer species could disproportionally impact ecosystem function.

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Acute-phase proteins (APP) are secreted from the liver as a result of inflammation or infection and are measurable in serum and plasma. To determine whether the constitutive APP serum amyloid A (SAA), alpha-1-acid glycoprotein (AGP), ceruloplasmin (Cp), and ovotransferrin (Ovt) have changed as a result of selection for improved production and growth characteristics over the last 40 yr two historical broilers lines were compared to a modern line of the same lineage. Serum was harvested from blood samples taken from the 3 broiler lines on days 10, 17, and 20, and the APP concentrations were determined using immunoassay methods. Most of the significant changes observed were age related, with SAA and Cp having significantly lower concentrations at day 20 than days 10 and 17 in all lines. The only significant difference between lines was observed at day 20 on which both Cp (P = 0.01) and AGP (P = 0.03) were significantly higher in the modern line than the 90s line, though no significant differences were noted between the modern and 70s line. When evaluating the difference in APP concentrations between males (Cx) and females (Px) across all 3 lines, females had a higher SAA at day 17 and lower SAA at day 20, P = 0.0078 and 0.0327 respectively, and males had a significantly higher Ovt on days 17 and 20 (P = 0.0002 and P = 0.003 respectively). These results reveal that APP concentrations fluctuate over this early period of growth and that the changes in APP serum concentration appear uniform between 3 lines with very contrasting selection history, suggesting the improvements made in meat production efficiency since the 1970s have not affected the circulating concentrations of these constitutively expressed APP.

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Habitat complexity can influence predation rates (e.g. by providing refuge) but other ecosystem processes and species interactions might also be modulated by the properties of habitat structure. Here, we focussed on how complexity of artificial habitat (plastic plants), in microcosms, influenced short-term processes driven by three aquatic detritivores. The effects of habitat complexity on leaf decomposition, production of fine organic matter and pH levels were explored by measuring complexity in three ways: 1. as the presence vs. absence of habitat structure; 2. as the amount of structure (3 or 4.5 g of plastic plants); and 3. as the spatial configuration of structures (measured as fractal dimension). The experiment also addressed potential interactions among the consumers by running all possible species combinations. In the experimental microcosms, habitat complexity influenced how species performed, especially when comparing structure present vs. structure absent. Treatments with structure showed higher fine particulate matter production and lower pH compared to treatments without structures and this was probably due to higher digestion and respiration when structures were present. When we explored the effects of the different complexity levels, we found that the amount of structure added explained more than the fractal dimension of the structures. We give a detailed overview of the experimental design, statistical models and R codes, because our statistical analysis can be applied to other study systems (and disciplines such as restoration ecology). We further make suggestions of how to optimise statistical power when artificially assembling, and analysing, 'habitat complexity' by not confounding complexity with the amount of structure added. In summary, this study highlights the importance of habitat complexity for energy flow and the maintenance of ecosystem processes in aquatic ecosystems.

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Quantitative polymerase chain reaction (qPCR) is a powerful technique for quantification of gene expression, especially genes involved in immune responses. Although qPCR is a very efficient and sensitive tool, variations in the enzymatic efficiency, quality of RNA and the presence of inhibitors can lead to errors. Therefore, qPCR needs to be normalised to obtain reliable results and allow comparison. The most common approach is to use reference genes as internal controls in qPCR analyses. In this study, expression of seven genes, including ß-actin (ACTB), ß-2-microglobulin (B2M), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), ß-glucuronidase (GUSB), TATA box binding protein (TBP), α-tubulin (TUBAT) and 28S ribosomal RNA (r28S), was determined in cells isolated from chicken lymphoid tissues and stimulated with three different mitogens. The stability of the genes was measured using geNorm, NormFinder and BestKeeper software. The results from both geNorm and NormFinder were that the three most stably expressed genes in this panel were TBP, GAPDH and r28S. BestKeeper did not generate clear answers because of the highly heterogeneous sample set. Based on these data we will include TBP in future qPCR normalisation. The study shows the importance of appropriate reference gene normalisation in other tissues before qPCR analysis.

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OBJECTIVE: To evaluate changes in glycemic control following the initial canagliflozin pharmacy claim in a real-world population. RESEARCH DESIGN AND METHODS: A retrospective cohort analysis of adult patients with type 2 diabetes mellitus (T2DM) was conducted using 2013 medical, pharmacy and laboratory claims from the Inovalon MORE 2 Registry. Patients with T2DM aged ≥18 years with ≥60 days of canagliflozin supply and HbA1c test results within 120 days before and ≥60 days after initial canagliflozin claim (defined as index date) were included. The differences between HbA1c levels pre- and post-index were assessed. Changes pre- and post-index in Healthcare Effectiveness Data and Information Set (HEDIS) glycemic control criteria of HbA1c <7% and <8% and poor control of HbA1c >9% were evaluated. Subgroup analyses of patients with HbA1c >7% at baseline and patients aged ≥65 were also conducted. RESULTS: Among the 268 patients meeting the study criteria, mean HbA1c pre-index was 8.3% and post-index was 7.6%; the mean reduction in HbA1c pre-post index was 0.7% (95% CI: 0.6%, 0.9%). The proportions of patients meeting the HEDIS glycemic control measures (HbA1c <7%, <8% and poor control of >9%) improved and was significantly different pre- and post-index (all p < 0.001). Of the patients with an HbA1c >7% prior to index (81% of the cohort; mean pre-index HbA1c = 8.8%), HbA1c was reduced by 0.9% (95% CI: 0.8%, 1.1%). The aged ≥65 subgroup consisted of 15% of the cohort, with a pre-index HbA1c of 8.3%. The mean reduction in HbA1c test results pre- and post-canagliflozin index was 0.6% (95% CI: 0.4%, 0.9%). This analysis did not adjust for changes in antihyperglycemic agents during the study period. CONCLUSION: Patients with T2DM were observed to have improved glycemic control following initial canagliflozin pharmacy claim as measured by HbA1c change and attainment of specific glycemic control criteria.

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Biodiversity loss is occurring rapidly worldwide, yet it is uncertain whether few or many species are required to sustain ecosystem functioning in the face of environmental change. The importance of biodiversity might be enhanced when multiple ecosystem processes (termed multifunctionality) and environmental contexts are considered, yet no studies have quantified this explicitly to date. We measured five key processes and their combined multifunctionality at three temperatures (5, 10 and 15 °C) in freshwater aquaria containing different animal assemblages (1-4 benthic macroinvertebrate species). For single processes, biodiversity effects were weak and were best predicted by additive-based models, i.e. polyculture performances represented the sum of their monoculture parts. There were, however, significant effects of biodiversity on multifunctionality at the low and the high (but not the intermediate) temperature. Variation in the contribution of species to processes across temperatures meant that greater biodiversity was required to sustain multifunctionality across different temperatures than was the case for single processes. This suggests that previous studies might have underestimated the importance of biodiversity in sustaining ecosystem functioning in a changing environment.

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1. Numerous studies have revealed (usually positive) relationships between biodiversity and ecosystem functioning (B-EF), but the underpinning drivers are rarely addressed explicitly, hindering the development of a more predictive understanding. 2. We developed a suite of statistical models (where we combined existing models with novel ones) to test for richness and evenness effects on detrital processing in freshwater microcosms. Instead of using consumer species as biodiversity units, we used two size classes within three species (six types). This allowed us to test for diversity effects and also to focus on the role of body size and biomass. 3. Our statistical models tested for (i) whether performance in polyculture was more than the sum of its parts (non-additive effects), (ii) the effects of specific type combinations (assemblage identity effects) and (iii) whether types behaved differently when their absolute or relative abundances were altered (e.g. because type abundance in polyculture was lower compared with monoculture). The latter point meant we did not need additional density treatments. 4. Process rates were independent of richness and evenness and all types performed in an additive fashion. The performance of a type was mainly driven by the consumers' metabolic requirements (connected to body size). On an assemblage level, biomass explained a large proportion of detrital processing rates. 5. We conclude that B-EF studies would benefit from widening their statistical approaches. Further, they need to consider biomass of species assemblages and whether biomass is comprised of small or large individuals, because even if all species are present in the same biomass, small species (or individuals) will perform better.

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In a dose-escalation trial for a new drug, each successive dose is tested on a new cohort of volunteer subjects, so that if any dose produces severe adverse reactions then higher doses are not tested. However, if there are other differences between the cohorts, such as differences in environmental health factors, type of person or experimental procedure, then these differences may obscure the differences between doses. Therefore, cohorts should be fitted in the analysis, as either fixed or random effects. I suggest that, if this is done, then there are three simple principles that reduce variance (i) allocating no more than half the subjects in any cohort to any single dose; (ii) subject to safety constraints, using as many different doses as possible in each cohort; (iii) using one more cohort than the number of doses, without increasing the total number of subjects. Using these principles, I propose some new designs that conform to the safety rules of traditional dose-escalation trials while reducing the variance of the estimators of differences between the doses by a factor of two or more, for the same number of subjects.

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AIMS: To determine the prevalence and number of Salmonella and Campylobacter in sausages and to evaluate their destruction during cooking. METHODS AND RESULTS: One hundred and sixty-two packs of uncooked economy or catering sausages, comprising 53 packs of frozen and 109 of chilled sausages, were purchased in Devon between March and July 2000. All were tested for the presence of Salmonella and 51 packs of chilled sausages were also examined for the presence of Campylobacter spp. To investigate the heat tolerance of Salmonella enterica serovar Typhimurium DT104 in sausage-meat, chilled, handmade and frozen sausages were inoculated with approx. 1.5 x 10(4) bacterial cells per sausage (approximately 300 cfu g(-1)) and then cooked by frying, grilling or barbecuing. The levels of creatinine kinase, lactate dehydrogenase and alkaline phosphatase in uncooked and cooked sausages were measured to evaluate their potential as indicators of adequate cooking and, therefore, pathogen elimination. Salmonella were detected in 7.5% of frozen and 9.1% of the chilled sausages (8.6% overall) but Campylobacter spp. were not isolated. After cooking, a visual assessment suggested that all of the sausages were thoroughly cooked. Despite this, barbecuing and frying sometimes allowed Salmonella cells to survive and the temperature profiles during cooking indicated that the lethal range was sometimes not reached. The enzyme levels tested were not reliable indicators of the inactivation of bacterial pathogens because Salmonella were sometimes isolated from sausages with low values of all three enzymes. CONCLUSIONS: Salmonella spp. are present in a significant proportion of sausages and are not always killed during the cooking process. SIGNIFICANCE AND IMPACT OF THE STUDY: These findings have clear implications for public health.

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Detailed mapping of glucose and lactate metabolism along the radius of the hepatic lobule was performed in situ in rat livers perfused with 1.5 mM lactate before and during the addition of 5 mM fructose. The majority of fructose uptake occurred in the periportal region; 45% of fructose taken up in the periportal half of the lobular volume being converted into glucose. Periportal lactate uptake was markedly decreased by addition of fructose. Basal perivenous lactate output, which was derived from glucose synthesized periportally, was increased in the presence of fructose. During fructose infusion there was a small decrease in cell pH periportally, but acidification of up to 0.5 pH units perivenously. The evidence suggests that in situ the apparent direct conversion of fructose into lactate represents, to a substantial extent, the result of periportal conversion of fructose into glucose and the subsequent uptake and glycolysis to lactate in the perivenous zone of some of that glucose. (31)P NMR spectroscopy showed that the cellular concentration of phosphomonoesters changes very little periportally during fructose infusion, but there was an approximate twofold increase perivenously, presumably due to the accumulation of fructose 1-phosphate. It may be inferred that fructokinase activity is expressed throughout the hepatic lobule.

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The intralobular distribution of metabolism was examined in the livers from rats with severe diabetic ketoacidosis (DKA), perfused at pH 6.8, and compared with that in livers from normal starved animals perfused at either pH 7.4 or 6.8. With lactate and palmitate as substrates, the perivenous uptake of periportally synthesized glucose seen in normal livers at pH 7.4 was abolished during DKA; indeed, gluconeogenesis was most active in the perivenous region. Whereas in normal livers perfused at pH 7.4 the periportal region showed a markedly elevated intracellular pH (pH(i)) compared with the perivenous zone, this distribution of pH(i) was reversed in DKA, with an intermediate distribution in normal livers perfused at pH 6. 8. 3-Hydroxybutyrate was generated throughout the lobule. Some acetoacetate generated periportally was converted to 3-hydroxybutyrate more perivenously. A steep gradient of oxygen uptake along the radius of the lobule was apparent in all three groups; oxygen uptake was greatly decreased perivenously despite adequate oxygen supply. These findings are consistent with our previous observations of the lobular co-location of high pH(i) and gluconeogenesis, and might offer an explanation of how high gluconeogenic rates can continue in spite of severe systemic acidosis in DKA. The findings provide direct evidence for a marked redistribution of intralobular metabolism in DKA.

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Ramu stunt disease of sugarcane (ScRS) was responsible for large yield losses in commercial sugarcane varieties (interspecific hybrids of Saccharum spp.) in the Ramu Valley in northeast Papua New Guinea during the late 1980s. Losses were total in the cultivar Ragnar; Q90 and Yasawa were also affected but Cadmus and Q107 were resistant. Since that time, replanting with resistant cultivars has kept the disease under control. The disease spreads rapidly in susceptible cultivars, where it results in severe stunting of the cane and a yellow mottled striping of the leaves. Although several attempts have been made to detect a viral pathogen, no evidence for viral etiology exists and the causal agent remains unknown. With a nested polymerase chain reaction (PCR) of general phytoplasma primers from the 16S rDNA (1), phytoplasma-specific products were consistently amplified from the leaves of field-grown sugarcane, from sugarcane with ScRS symptoms grown in the glasshouse at IACR-Rothamsted, UK, and from samples of the putative vector collected at Ramu, the delphacid plant hopper Eumetopina flavipes Muir, which had been found to transmit symptoms of Ramu stunt in pot trials (2). Digestion of the amplimers with restriction enzymes RsaI and HaeIII gave profiles that matched those of members of the sugarcane white leaf (SCWL) phytoplasma group. The DNA sequence of the intergenic spacer region of the phytoplasma associated with ScRS showed a 95.98% homology with that of SCWL, suggesting that this newly discovered phytoplasma can provisionally be placed in this group. The 16S-23S intergenic spacer sequence has been submitted to GenBank (accession no. AF 106061). References: (1) C. P. R. Cronjé et al. Ann. Appl. Biol. 133:177, 1998; (2) L. S. Kuniata et al. J. Aust. Entomol. Soc. 33:185, 1994.

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Mutations in GJB2 encoding the gap junction protein connexin-26 (Cx26) have been established as the basis of autosomal recessive non-syndromic hearing loss. The involvement of GJB2 in autosomal dominant deafness has also been proposed, although the putative mutation identified in one family with both deafness and palmoplantar keratoderma has recently been suggested to be merely a non-disease associated polymorphism. We have observed a similar phenotype in an Egyptian family that segregated with a heterozygous missense mutation of GJB2, leading to a non-conservative amino acid substitution (R75W). The deleterious dominant-negative effect of R75W on gap channel function was subsequently demonstrated in the paired oocyte expression system. Not only was R75W alone incapable of inducing electrical conductance between adjacent cells, but it almost completely suppressed the activity of co-expressed wildtype protein. The Cx26 mutant W77R, which has been implicated in autosomal recessive deafness, also failed to form functional gap channels by itself but did not significantly interfere with the function of wildtype Cx26. These data provide compelling evidence for the serious functional consequences of Cx26 mutations in dominant and recessive deafness.

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Erythrokeratodermia variabilis (EKV, OMIM 133200) is an autosomal dominant genodermatosis with considerable intra- and interfamilial variability. It has a disfiguring phenotype characterized by the independent occurrence of two morphologic features: transient figurate red patches and localized or generalized hyperkeratosis. Both features can be triggered by external factors such as trauma to the skin. After initial linkage to the RH locus on 1p, EKV was mapped to an interval of 2.6 cM on 1p34-p35, and a candidate gene (GJA4) encoding the gap junction protein alpha-4 (connexin 31, Cx31) was excluded by sequence analysis. Evidence in mouse suggesting that the EKV region harbours a cluster of epidermally expressed connexin genes led us to characterize the human homologues of GJB3 (encoding Cx31) and GJB5 (encoding Cx31.1). GJB3, GJB5 and GJA4 were localized to a 1.1-Mb YAC in the candidate interval. We detected heterozygous missense mutations in GJB3 in four EKV families leading to substitution of a conserved glycine by charged residues (G12R and G12D), or change of a cysteine (C86S). These mutations are predicted to interfere with normal Cx31 structure and function, possibly due to a dominant inhibitory effect. Our results implicate Cx31 in the pathogenesis of EKV, and provide evidence that intercellular communication mediated by Cx31 is crucial for epidermal differentiation and response to external factors.

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Maternal protein restriction is a model of fetal programming of adult glucose intolerance. Perfused livers of 48-h- starved adult offspring of rat dams fed 8% protein diets during pregnancy and lactation produced more glucose from 6 mM lactate than did control livers from rats whose dams were fed 20% protein. In control livers, a mean of 24% of the glucose formed from lactate in the periportal region of the lobule was taken up by the most distal perivenous cells; this distal perivenous uptake was greatly diminished in maternal low protein (MLP) livers, accounting for a major fraction of the increased glucose output of MLP livers. In control livers, the distal perivenous cells contained 40% of the total glucokinase of the liver; this perivenous concentration of glucokinase was greatly reduced in MLP livers. Intralobular distribution of phosphenolpyruvate carboxykinase was unaltered, though overall increased activity could have contributed to the elevated glucose output. Hepatic lobular volume in MLP livers was twice that in control livers, indicating that MLP livers had half the normal number of lobules. Fetal programming of adult glucose metabolism may operate partly through structural alterations and changes in glucokinase expression in the immediate perivenous region.

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The results of increasing blood flow capability in a modified system for plasma exchange with a rotating filter are reported. There were 742 treatments performed with the authors' original system (OS), limited to blood flows of 100 ml/ min, and 327 treatments performed with the updated system (US), allowing for blood flows of 150 ml/min. Blood flows for OS were 98 +/- 5 ml/min (mean +/- SD) vs 145 +/- 12 ml/min for US (p < 0.001). Plasma flows were 65 +/- 7 ml/min for OS vs 98 +/- 12 ml/min for US (p < 0.001). Plasma removal rate was 42 +/- 8 ml/min for OS vs 61 +/- 14 ml/min for US (p < 0.001). Mean treatment time was reduced from 76 +/- 23 min for OS to 52 +/- 17 min for US (p < 0.001) in spite of providing a similar amount of plasma removed per treatment (3,113 +/- 577 ml/Rx for OS vs 3078 +/- 797 ml/Rx for US; p = 0.48). Despite statistical significance, there were only small differences in filtration fractions (65 +/- 12% for OS vs 62 +/- 11% for US; p < 0.001) and patient hematocrits (34 +/- 6% for OS vs 33 +/- 6% for US; p < 0.001). In conclusion, modification of the OS to allow for increased blood flow has resulted in a substantial improvement in procedure efficiency and a clinically useful decrease in treatment time.

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