What is wrong with non-respondents? Alcohol-, drug- and smoking related mortality and morbidity in a 12-year follow up study of respondents and non-respondents in the Danish Health and Morbidity Survey

Aim: Response rates in health surveys have diminished over the last two decades, making it difficult to obtain reliable information on health and health-related risk factors in different population groups. This study compared cause-specific mortality and morbidity among survey respondents and different types of non-respondents to estimate alcohol-, drug- and smoking related mortality and morbidity among non-respondents. Design: Prospective follow-up study of respondents and non-respondents in two cross-sectional health surveys. Setting: Denmark. Participants: A total sample of 39,540 Danish citizens aged 16 or older. Measurements: Register-based information on cause-specific mortality and morbidity at the individual level was obtained for respondents (n=28,072) and different types of non-respondents (refusals n=8,954; illness/disabled n=731, uncontactable n=1,593). Cox proportional hazards models were used to examine differences in alcohol-, drug- and smoking-related mortality and morbidity, respectively, in a 12 year follow-up period. Findings: Overall, non-response was associated with a significantly increased hazard ratio of 1.56 (95% CI: 1.36–1.78) for alcohol-related morbidity, 1.88 (95% CI: 1.38-2.57) for alcohol-related mortality, 1.55 (95% CI: 1.27–1.88) for drug-related morbidity, 3.04 (95% CI: 1.57–5.89) for drug-related mortality and 1.15 (95% CI: 1.03–1.29) for smoking-related morbidity. The hazard ratio for smoking-related mortality also tended to be higher among non-respondents compared with respondents although no significant association was evident (HR: 1.14; 95% CI: 0.95-1.36). Uncontactable and ill/disabled non-respondents generally had a higher hazard ratio of alcohol-, drug- and smoking related mortality and morbidity compared with refusal non-respondents. Conclusion: Health survey non-respondents in Denmark have an increased hazard ratio of alcohol-, drug-, and smoking-related mortality and morbidity compared with respondents, which may indicate more unfavourable health behaviours among non-respondents

Akin House Curriculum Development and Living History Programming

This unit plan is comprised of a variety of inquiry-based lessons that explore the culture and way of life of the Native Americans who occupied New England. After studying the Akin house documents, materials, and narratives, I chose to focus my unit on the land and the people who came before the Akin family so that students will learn the long-view of our rich New England history

Senior Voice Recital: Charlotte Derwin, Mezzo-Soprano; Harold Gray, Piano; March 14, 1971

Centennial Recital HallMarch 14, 19713:00 p.m

Gating NO Release from Nitric Oxide Synthase

We have investigated the kinetics of NO escape from Geobacillus stearothermophilus nitric oxide synthase (gsNOS). Previous work indicated that NO release was gated at position 223 in mammalian enzymes; our kinetics experiments include mutants at that position along with measurements on the wild type enzyme. Employing stopped-flow UV–vis methods, reactions were triggered by mixing a reduced enzyme/N-hydroxy-l-arginine complex with an aerated buffer solution. NO release kinetics were obtained for wt NOS and three mutants (H134S, I223V, H134S/I223V). We have confirmed that wt gsNOS has the lowest NO release rate of known NOS enzymes, whether bacterial or mammalian. We also have found that steric clashes at positions 223 and 134 hinder NO escape, as judged by enhanced rates in the single mutants. The empirical rate of NO release from the gsNOS double mutant (H134/I223V) is nearly as rapid as that of the fastest mammalian enzymes, demonstrating that both positions 223 and 134 function as gates for escape of the product diatomic molecule

Beta-Lactamase Resistance Harboured by Escherichia coli isolated from a Dairy Farm

CTX-M beta-lactamases have become one of the most prevalent extended spectrum beta-lactamases (ESBL) globally. Their association with mobile elements such as ISEcp1, has allowed for their capture and mobilisation within both human and animal environments. The EVAL farms research programme generated a collection of 1,000 Escherichia coli bovine faecal samples isolated from the environment of a dairy farm on selective media and characterised phenotypically for antibiotic resistance via the disc diffusion method. This current study further characterised the antibiotic resistances of 86 E. coli from the EVAL farms collection, chosen for their beta-lactamase type resistance phenotype. These 86 isolates were grouped according to their resistance phenotypes, which included those suspected of encoding blaCTX-M (reduced susceptibility to ampicillin (AMP), cefotaxime (CTX) and aztreonam (ATM) but susceptibility to amoxicillin-clavulanic acid (AMC) and cefoxitin (FOX)) and those suspected of overexpressing ampC (reduced susceptibility to AMP, AMC, FOX, CTX and ATM. Confirmation of the presence of blaCTX-M by PCR resulted in 39 blaCTX strains being identified and the remaining 47 isolates categorised as putative ampC strains. The presence of an ISEcp1 element was also shown in all blaCTX PCR-positive isolates. The 86 isolates were further characterised phenotypically using minimum inhibitory concentration (MIC) assays via the agar dilution method, with an extended panel of 25 antibiotics which included cefquinome (CFQ). The extended panel included any classes of antibiotics that could cover different resistance mechanisms such as carbapenemases, ESBLs as well as aminoglycoside, tetracycline and colistin resistance. High levels of resistance were seen to AMP, CTX, cefpodoxime (CPD) and CFQ along with resistance to ceftazidime (CAZ), ATM and tetracycline (TET) in the blaCTX isolates. Genotypic characterisation via whole genome sequencing (WGS) was conducted via both short read Illumina and long read MinION Oxford Nanopore Technologies (ONT), with hybrid assembly on all isolates. This WGS was able to show that all the blaCTX-M were of blaCTX-M-15 type, were chromosomally encoded and in association with ISEcp1. WGS also revealed the ISEcp1 element additionally encoded qnrS1 in all isolates and tetAR in 34 of the isolates, with 4 found to contain no tetAR genes and one that had the tetAR genes located separately from the ISEcp1 element, in a different region of the genome. Subsequent in silico multi-locus sequence typing (MLST) using the MLST finder from the Centre for Genomic Epidemiology (CGE), showed all within the blaCTX group to be ST2325. Comparison of the 39 isolates in the blaCTX group by single nucleotide polymorphism (SNP) analysis with 105 ST2325 isolates from the Enterobase database, produced a maximum likelihood tree, showing that the 39 blaCTX EVAL farms isolates were part of their own clonal branch of the tree and within 1-5 SNPs of each other, demonstrating that spread of blaCTX-M-15 on this dairy farm was likely as a result of clonal expansion. Other ST2325 isolates from Enterobase were found not to be closely related to the 39 blaCTX isolates, with the closest an E. coli bovine isolate from Spain which was within 36-45 SNPs of the 39 blaCTX isolates. It was also noted that ST2325 isolates from other published studies analysed from Enterobase appeared to form separate clonal study-associated clusters, with clonal groups of ST2325 within 0-6 SNPs of each other. The use of WGS allowed resistance genes to be identified and compared to the phenotypic data, and plasmids and other mobile elements including the ISEcp1 elements to be characterised. The potential for ISEcp1 to mobilise a chromosomally-encoded blaCTX-M-15 to a resident plasmid and transpose to another strain and whether sub-lethal levels of antibiotics used in dairy farming (including AMP, cloxacillin (CLOX) and CAZ) might enhance this transposition of ISEcp1, was addressed through transposition experiments with four isolates from the blaCTX group. The sub-lethal levels of these antibiotics used, looked to mimic the concentrations that might be encountered by bacteria of treated animals or within the environment of the dairy farm. Transposition of the ISEcp1 element in association with blaCTX-M-15, was successful with all concentrations of AMP, CLOX and CAZ and enhanced transposition with an increased rate of transfer (when compared to the baseline rate of transfer in non-selective media) was successful with some concentrations of AMP, CLOX and CAZ. Levels of enhancement varied from 1.07 fold to 45 fold the baseline rate. The characterisation of subsequent transconjugants encoding ISEcp1 elements using WGS via both Illumina short and MinION (ONT) long read with hybrid assembly showed that the ISEcp1 elements could either lose or gain downstream genes, through the recognition of a new imperfect IRR site. This revealed a possible mechanism for the loss or gain of a phenotype within the dairy farm E. coli isolates. Isolates of the second group of E. coli displaying a beta-lactamase type phenotype, the ampC group, when further characterised through MIC assays showed that many phenotypic resistances indicated by the disc assay were lost and only four of the 47 isolates in the ampC group were resistant to streptomycin (STREP), six were resistant to TET and only one isolate was resistant to trimethoprim/sulfamethoxazole combination (SXT). Only 22 isolates were showing likely overexpression of ampC according to the MIC assay results, with high level resistance to AMP along with resistance to CAZ, CPD and ATM with intermediate resistance to CTX. Through a combination of both PCR and Sanger sequencing, and WGS, 22 isolates were confirmed as overexpressing ampC by locating mutations in the promoter regions of ampC. WGS of the 47 isolates in the ampC group identified additional resistance genes including beta-lactamase type resistance genes blaTEM-1 and blaOXA-1 in one isolate each respectively, mobile genetic elements (MGEs) and a small number of virulence genes. MLST typing showed, 21 isolates were ST1308 with 20 of these overexpressing ampC. These 21 ST1308 were subject to SNP analysis via snippy and a maximum likelihood tree was constructed. From these data it appeared the earliest sampled isolate, which was not overexpressing ampC, was within 431-561 SNPs of the remaining 20 ST1308 isolates and therefore was not closely related but did appear to share a common ancestor them. Within the remaining 20, 19 were within 5-1 SNPs of each other and one was within 8-16 SNPs of those 19. Therefore, it appeared the 19 were likely clonal and the 1 within 8-16 SNPs appeared to be very closely related to those 19. This suggested that in this particular dairy farm environment, there had been a small clonal expansion of isolates of the same ST, with the majority also encoding overexpression of ampC. The results of this study showed a potential mechanism for mobility of blaCTX-M-15 within the environment of a dairy farm, demonstrated there had been spread of blaCTX-M-15 as a result of the clonal expansion of ST2325 and also revealed several different mechanisms in place for beta-lactamase type resistance including both blaCTX-M-15 and overexpression of ampC. The study also showed the benefits of utilising both phenotypic and genotypic methods together for the identification of resistance mechanisms within E. coli

SSCC TD: a serial and simultaneous configural-cue compound stimuli representation for temporal difference learning

This paper presents a novel representational framework for the Temporal Difference (TD) model of learning, which allows the computation of configural stimuli – cumulative compounds of stimuli that generate perceptual emergents known as configural cues. This Simultaneous and Serial Configural-cue Compound Stimuli Temporal Difference model (SSCC TD) can model both simultaneous and serial stimulus compounds, as well as compounds including the experimental context. This modification significantly broadens the range of phenomena which the TD paradigm can explain, and allows it to predict phenomena which traditional TD solutions cannot, particularly effects that depend on compound stimuli functioning as a whole, such as pattern learning and serial structural discriminations, and context-related effects

Beta-Lactamase Resistance Harboured by Escherichia coli isolated from a Dairy Farm

CTX-M beta-lactamases have become one of the most prevalent extended spectrum beta-lactamases (ESBL) globally. Their association with mobile elements such as ISEcp1, has allowed for their capture and mobilisation within both human and animal environments. The EVAL farms research programme generated a collection of 1,000 Escherichia coli bovine faecal samples isolated from the environment of a dairy farm on selective media and characterised phenotypically for antibiotic resistance via the disc diffusion method. This current study further characterised the antibiotic resistances of 86 E. coli from the EVAL farms collection, chosen for their beta-lactamase type resistance phenotype. These 86 isolates were grouped according to their resistance phenotypes, which included those suspected of encoding blaCTX-M (reduced susceptibility to ampicillin (AMP), cefotaxime (CTX) and aztreonam (ATM) but susceptibility to amoxicillin-clavulanic acid (AMC) and cefoxitin (FOX)) and those suspected of overexpressing ampC (reduced susceptibility to AMP, AMC, FOX, CTX and ATM. Confirmation of the presence of blaCTX-M by PCR resulted in 39 blaCTX strains being identified and the remaining 47 isolates categorised as putative ampC strains. The presence of an ISEcp1 element was also shown in all blaCTX PCR-positive isolates. The 86 isolates were further characterised phenotypically using minimum inhibitory concentration (MIC) assays via the agar dilution method, with an extended panel of 25 antibiotics which included cefquinome (CFQ). The extended panel included any classes of antibiotics that could cover different resistance mechanisms such as carbapenemases, ESBLs as well as aminoglycoside, tetracycline and colistin resistance. High levels of resistance were seen to AMP, CTX, cefpodoxime (CPD) and CFQ along with resistance to ceftazidime (CAZ), ATM and tetracycline (TET) in the blaCTX isolates. Genotypic characterisation via whole genome sequencing (WGS) was conducted via both short read Illumina and long read MinION Oxford Nanopore Technologies (ONT), with hybrid assembly on all isolates. This WGS was able to show that all the blaCTX-M were of blaCTX-M-15 type, were chromosomally encoded and in association with ISEcp1. WGS also revealed the ISEcp1 element additionally encoded qnrS1 in all isolates and tetAR in 34 of the isolates, with 4 found to contain no tetAR genes and one that had the tetAR genes located separately from the ISEcp1 element, in a different region of the genome. Subsequent in silico multi-locus sequence typing (MLST) using the MLST finder from the Centre for Genomic Epidemiology (CGE), showed all within the blaCTX group to be ST2325. Comparison of the 39 isolates in the blaCTX group by single nucleotide polymorphism (SNP) analysis with 105 ST2325 isolates from the Enterobase database, produced a maximum likelihood tree, showing that the 39 blaCTX EVAL farms isolates were part of their own clonal branch of the tree and within 1-5 SNPs of each other, demonstrating that spread of blaCTX-M-15 on this dairy farm was likely as a result of clonal expansion. Other ST2325 isolates from Enterobase were found not to be closely related to the 39 blaCTX isolates, with the closest an E. coli bovine isolate from Spain which was within 36-45 SNPs of the 39 blaCTX isolates. It was also noted that ST2325 isolates from other published studies analysed from Enterobase appeared to form separate clonal study-associated clusters, with clonal groups of ST2325 within 0-6 SNPs of each other. The use of WGS allowed resistance genes to be identified and compared to the phenotypic data, and plasmids and other mobile elements including the ISEcp1 elements to be characterised. The potential for ISEcp1 to mobilise a chromosomally-encoded blaCTX-M-15 to a resident plasmid and transpose to another strain and whether sub-lethal levels of antibiotics used in dairy farming (including AMP, cloxacillin (CLOX) and CAZ) might enhance this transposition of ISEcp1, was addressed through transposition experiments with four isolates from the blaCTX group. The sub-lethal levels of these antibiotics used, looked to mimic the concentrations that might be encountered by bacteria of treated animals or within the environment of the dairy farm. Transposition of the ISEcp1 element in association with blaCTX-M-15, was successful with all concentrations of AMP, CLOX and CAZ and enhanced transposition with an increased rate of transfer (when compared to the baseline rate of transfer in non-selective media) was successful with some concentrations of AMP, CLOX and CAZ. Levels of enhancement varied from 1.07 fold to 45 fold the baseline rate. The characterisation of subsequent transconjugants encoding ISEcp1 elements using WGS via both Illumina short and MinION (ONT) long read with hybrid assembly showed that the ISEcp1 elements could either lose or gain downstream genes, through the recognition of a new imperfect IRR site. This revealed a possible mechanism for the loss or gain of a phenotype within the dairy farm E. coli isolates. Isolates of the second group of E. coli displaying a beta-lactamase type phenotype, the ampC group, when further characterised through MIC assays showed that many phenotypic resistances indicated by the disc assay were lost and only four of the 47 isolates in the ampC group were resistant to streptomycin (STREP), six were resistant to TET and only one isolate was resistant to trimethoprim/sulfamethoxazole combination (SXT). Only 22 isolates were showing likely overexpression of ampC according to the MIC assay results, with high level resistance to AMP along with resistance to CAZ, CPD and ATM with intermediate resistance to CTX. Through a combination of both PCR and Sanger sequencing, and WGS, 22 isolates were confirmed as overexpressing ampC by locating mutations in the promoter regions of ampC. WGS of the 47 isolates in the ampC group identified additional resistance genes including beta-lactamase type resistance genes blaTEM-1 and blaOXA-1 in one isolate each respectively, mobile genetic elements (MGEs) and a small number of virulence genes. MLST typing showed, 21 isolates were ST1308 with 20 of these overexpressing ampC. These 21 ST1308 were subject to SNP analysis via snippy and a maximum likelihood tree was constructed. From these data it appeared the earliest sampled isolate, which was not overexpressing ampC, was within 431-561 SNPs of the remaining 20 ST1308 isolates and therefore was not closely related but did appear to share a common ancestor them. Within the remaining 20, 19 were within 5-1 SNPs of each other and one was within 8-16 SNPs of those 19. Therefore, it appeared the 19 were likely clonal and the 1 within 8-16 SNPs appeared to be very closely related to those 19. This suggested that in this particular dairy farm environment, there had been a small clonal expansion of isolates of the same ST, with the majority also encoding overexpression of ampC. The results of this study showed a potential mechanism for mobility of blaCTX-M-15 within the environment of a dairy farm, demonstrated there had been spread of blaCTX-M-15 as a result of the clonal expansion of ST2325 and also revealed several different mechanisms in place for beta-lactamase type resistance including both blaCTX-M-15 and overexpression of ampC. The study also showed the benefits of utilising both phenotypic and genotypic methods together for the identification of resistance mechanisms within E. coli

Phylogenomics and analysis of shared genes suggest a single transition to mutualism in Wolbachia of nematodes

Wolbachia, endosymbiotic bacteria of the order Rickettsiales, are widespread in arthropods but also present in nematodes. In arthropods, A and B supergroup Wolbachia are generally associated with distortion of host reproduction. In filarial nematodes, including some human parasites, multiple lines of experimental evidence indicate that C and D supergroup Wolbachia are essential for the survival of the host, and here the symbiotic relationship is considered mutualistic. The origin of this mutualistic endosymbiosis is of interest for both basic and applied reasons: How does a parasite become a mutualist? Could intervention in the mutualism aid in treatment of human disease? Correct rooting and high-quality resolution of Wolbachia relationships are required to resolve this question. However, because of the large genetic distance between Wolbachia and the nearest outgroups, and the limited number of genomes so far available for large-scale analyses, current phylogenies do not provide robust answers. We therefore sequenced the genome of the D supergroup Wolbachia endosymbiont of Litomosoides sigmodontis, revisited the selection of loci for phylogenomic analyses, and performed a phylogenomic analysis including available complete genomes (from isolates in supergroups A, B, C, and D). Using 90 orthologous genes with reliable phylogenetic signals, we obtained a robust phylogenetic reconstruction, including a highly supported root to the Wolbachia phylogeny between a (A + B) clade and a (C + D) clade. Although we currently lack data from several Wolbachia supergroups, notably F, our analysis supports a model wherein the putatively mutualist endosymbiotic relationship between Wolbachia and nematodes originated from a single transition event

Maternal fructose and/or salt intake and reproductive outcome in the rat: effects on growth, fertility, sex ratio, and birth order

Maternal diet can significantly skew the secondary sex ratio away from the expected value of 0.5 (proportion males), but the details of how diet may do this are unclear. Here, we altered dietary levels of salt (4% salt in the feed) and/or fructose (10% in the drinking water) of pregnant rats to model potential effects that consumption of a "Western diet" might have on maternofetal growth, development, and sex ratio. We demonstrate that excess fructose consumption before and during pregnancy lead to a marked skew in the secondary sex ratio (proportion of males, 0.60; P < 0.006). The effect was not mediated by selective developmental arrest of female embryos or influenced by fetal position in the uterine horn or sex-specific effects on sperm motility, suggesting a direct effect of glycolyzable monosaccharide on the maternal ovary and/or ovulated oocyte. Furthermore, combined excess maternal consumption of salt and fructose-sweetened beverage significantly reduced fertility, reflected as a 50% reduction in preimplantation and term litter size. In addition, we also noted birth order effects in the rat, with sequential implantation sites tending to be occupied by the same sex