Pedigree-free animal models : the relatedness matrix reloaded

Animal models typically require a known genetic pedigree to estimate quantitative genetic parameters. Here we test whether animal models can alternatively be based on estimates of relatedness derived entirely from molecular marker data. Our case study is the morphology of a wild bird population, for which we report estimates of the genetic variance-covariance matrices (G) of six morphological traits using three methods: the traditional animal model; a molecular marker-based approach to estimate heritability based on Ritland's pairwise regression method; and a new approach using a molecular genealogy arranged in a relatedness matrix (R) to replace the pedigree in an animal model. Using the traditional animal model, we found significant genetic variance for all six traits and positive genetic covariance among traits. The pairwise regression method did not return reliable estimates of quantitative genetic parameters in this population, with estimates of genetic variance and covariance typically being very small or negative. In contrast, we found mixed evidence for the use of the pedigree-free animal model. Similar to the pairwise regression method, the pedigree-free approach performed poorly when the full-rank R matrix based on the molecular genealogy was employed. However, performance improved substantially when we reduced the dimensionality of the R matrix in order to maximize the signal to noise ratio. Using reduced-rank R matrices generated estimates of genetic variance that were much closer to those from the traditional model. Nevertheless, this method was less reliable at estimating covariances, which were often estimated to be negative. Taken together, these results suggest that pedigree-free animal models can recover quantitative genetic information, although the signal remains relatively weak. It remains to be determined whether this problem can be overcome by the use of a more powerful battery of molecular markers and improved methods for reconstructing genealogies.

Large body size in an island-dwelling bird : a microevolutionary analysis

Island races of passerine birds display repeated evolution towards larger body size compared with their continental ancestors. The Capricorn silvereye (Zosterops lateralis chlorocephalus) has become up to six phenotypic standard deviations bigger in several morphological measures since colonization of an island approximately 4000 years ago. We estimated the genetic variance-covariance (G) matrix using full-sib and 'animal model' analyses, and selection gradients, for six morphological traits under field conditions in three consecutive cohorts of nestlings. Significant levels of genetic variance were found for all traits. Significant directional selection was detected for wing and tail lengths in one year and quadratic selection on culmen depth in another year. Although selection gradients on many traits were negative, the predicted evolutionary response to selection of these traits for all cohorts was uniformly positive. These results indicate that the G matrix and predicted evolutionary responses are consistent with those of a population evolving in the manner observed in the island passerine trend, that is, towards larger body size.

Isolation of microsatellite loci in the Capricorn silvereye, Zosterops lateralis chlorocephalus (Aves: Zosteropidae)

The Capricorn silvereye (Zosterops lateralis chlorocephalus) is ideally suited to investigating the genetic basis of body size evolution. We have isolated and characterized a set of microsatellite markers for this species. Seven out of 11 loci were polymorphic. The number of alleles detected ranged from two to five and observed heterozygosities between 0.12 and 0.67. One locus, ZL49, was found to be sex-linked. This moderate level of diversity is consistent with that expected in an isolated, island population.

Molecular cocrystals of 5-(4-bromophenyl)-1,3,4-thiadiazol-2-amine: hydrogen bonding in the structures of the 1:1 adduct with 2-(naphthalen-2-yloxy)acetic acid and the salt with 3,5-dinitrobenzoic acid

The structures of the anhydrous products from the interaction of 2-amino-5-(4-bromophenyl)-1,3,4-thiadiazole with (2-naphthoxy)acetic acid, the 1:1 adduct C8H6BrN3S . C12H10O3 (I) and 3,5-dinitrobenzoic acid, the salt C8H7BrN3S+ C7H3N2O6- (II) have been determined. In the adduct (I), a heterodimer is formed through a cyclic hydrogen-bonding motif [graph set R2/2(8)], involving carboxylic acid O-H...N(hetero)and amine N-H...O(carboxyl) interactions. The heterodimers are essentially planar with a thiadiazole to naphthyl ring dihedral angle of 15.9(2)deg. and the intramolecular thiadiazole to phenyl ring angle of 4.7(2)deg. An amine N-H...N(hetero) hydrogen bond between the heterodimers generates a one-dimensional chain structure extending down [001]. Also present are weak benzene-benzene and naphthalene-naphthalene pi-pi stacking interactions down the b axis [minimum ring centroid separation, 3.936(3) Ang.]. With the salt (II), the cation-anion association is also through a cyclic R2/2(8) motif but involving duplex N-H...O(carboxyl) hydrogen bonds, giving a heterodimer which is close to planar [dihedral angles between the thiadiazole ring and the two benzene rings, 5.00(16)deg. (intra) and 7.23(15)deg. (inter)]. A secondary centrosymmetric cyclic N-H...O(carboxyl) hydrogen-bonding association involving the second amino H-atom generates a heterotetramer. Also present in the crystal are weak pi-pi i-\p interactions between thiadiazolium rings [minimum ring centroid separation, 3.936(3)Ang.], as well as a short Br...O(nitro) interaction [3.314(4)Ang.]. The two structures reported here now provide a total of three crystallographically characterized examples of co-crystalline products from the interaction of 2-amino-5-(4-bromophenyl)-1,3,4-thiadiazole with carboxylic acids, of which only one involves proton-transfer.

Variation in the vitamin D receptor gene is associated with multiple sclerosis in an Australian population

Multiple Sclerosis (MS) is a chronic inflammatory demyelinating disease of the central nervous system (CNS) resulting in accumulating neurological disability. The disorder is more prevalent at higher latitudes. To investigate VDR gene variation using three intragenic restriction fragment length polymorphisms (Apa I, Taq I and Fok I) in an Australian MS case-control population. One hundred and four Australian MS patients were studied with patients classified clinically as Relapsing Remitting MS (RR-MS), Secondary Progressive MS (SP-MS) or Primary Progressive MS (PP-MS). Also, 104 age-, sex-, and ethnicity-matched controls were investigated as a comparative group. Our results show a significant difference of genotype distribution frequency between the case and control groups for the functional exon 9 VDR marker Taq I (p(Gen) = 0.016) and interestingly, a stronger difference for the allelic frequency (p(All) = 0.0072). The Apa I alleles were also found to be associated with MS (p(All) = 0.04) but genotype frequencies were not significantly different from controls (p(Gen) = 0.1). The Taq and Apa variants are in very strong and significant linkage disequilibrium (D' = 0.96, P < 0.0001). The genotypic associations are strongest for the progressive forms of MS (SP-MS and PP-MS). Our results support a role for the VDR gene increasing the risk of developing multiple sclerosis, particularly the progressive clinical subtypes of MS.

Isolation and characterization of tumor cells from the ascites of ovarian cancer patients : molecular phenotype of chemoresistant ovarian tumors

Tumor cells in ascites are a major source of disease recurrence in ovarian cancer patients. In an attempt to identify and profile the population of ascites cells obtained from ovarian cancer patients, a novel method was developed to separate adherent (AD) and non-adherent (NAD) cells in culture. Twenty-five patients were recruited to this study; 11 chemonaive (CN) and 14 chemoresistant (CR). AD cells from both CN and CR patients exhibited mesenchymal morphology with an antigen profile of mesenchymal stem cells and fibroblasts. Conversely, NAD cells had an epithelial morphology with enhanced expression of cancer antigen 125 (CA125), epithelial cell adhesion molecule (EpCAM) and cytokeratin 7. NAD cells developed infiltrating tumors and ascites within 12-14 weeks after intraperitoneal (i.p.) injections into nude mice, whereas AD cells remained non-tumorigenic for up to 20 weeks. Subsequent comparison of selective epithelial, mesenchymal and cancer stem cell (CSC) markers between AD and NAD populations of CN and CR patients demonstrated an enhanced trend in mRNA expression of E-cadherin, EpCAM, STAT3 and Oct4 in the NAD population of CR patients. A similar trend of enhanced mRNA expression of CD44, MMP9 and Oct4 was observed in the AD population of CR patients. Hence, using a novel purification method we demonstrate for the first time a distinct separation of ascites cells into epithelial tumorigenic and mesenchymal non-tumorigenic populations. We also demonstrate that cells from the ascites of CR patients are predominantly epithelial and show a trend towards increased mRNA expression of genes associated with CSCs, compared to cells isolated from the ascites of CN patients. As the tumor cells in the ascites of ovarian cancer patients play a dominant role in disease recurrence, a thorough understanding of the biology of the ascites microenvironment from CR and CN patients is essential for effective therapeutic interventions.

Weight-related differences in glucose metabolism and free fatty acid production in two South African population groups

OBJECTIVE The effects of free fatty acids (FFA), leptin, tumour necrosis factor (TNF) alpha and body fat distribution on in vivo oxidation of a glucose load were studied in two South African ethnic groups. DESIGN AND MEASUREMENTS Anthropometric and various metabolic indices were measured at fasting and during a 7h oral glucose tolerance test (OGTT). Body composition was measured using bioelectrical impedance analysis and subcutaneous and visceral fat mass was assessed using a five- and two-level CT-scan respectively. Glucose oxidation was evaluated by measuring the ratio of (13)CO(2) to (12)CO(2) in breath following ingestion of 1-(13)C-labelled glucose. SUBJECTS Ten lean black women (LBW), ten obese black women (OBW), nine lean white women (LWW) and nine obese white women (OWW) were investigated after an overnight fast. RESULTS Visceral fat levels were significantly higher (P < 0.01) in obese white than black women, despite similar body mass indexes (BMIs). There were no ethnic differences in glucose oxidation however; in the lean subjects of both ethnic groups the area under the curve (AUC) was higher than in obese subjects (P < 0.05 for both) and was found to correlate negatively with weight (r = -0.69, P < 0.01) after correcting for age. Basal TNF alpha concentrations were similar in all groups. Percentage suppression of FFAs at 30 min of the OCTT was 24 +/- 12% in OWW and - 38 +/- 23% (P < 0.05) in OBW, ie the 30 min FFA level was higher than the fasting level in the latter group. AUC for FFAs during the late postprandial period (120 - 420 min) was significantly higher in OWW than OBW (P < 0.01) and LWW (P < 0.01) and correlated positively with visceral fat mass independent of age (r = 0.78, P < 0.05) in the OWW only. Leptin levels were higher (P < 0.01) both at fasting and during the course of the OCTT in obese women from both ethnic groups compared to the lean women. CONCLUSIONS Glucose oxidation is reduced in obese subjects of both ethnic groups; inter- and intra-ethnic differences were observed in visceral fat mass and FFA production and it is possible that such differences may play a role in the differing prevalences of obesity-related disorders that have been reported in these two populations.

Ethnic differences in lipid metabolism in two groups of obese South African women

There is a higher prevalence of ischemic heart disease (IHD) in South African white than black women. The objective of this study was to determine biochemical explanations for this prevalence. The study group contained 15 obese black women (OBW) and 14 obese white women (OWW), ah premenopausal, who were examined after an overnight fast. Anthropometric measurements and blood concentrations of glucose, non-esterified fatty acids (NEFAs), catecholamines, plasminogen activator inhibitor-1, C-peptide, proinsulin, lipograms, cortisol, growth hormone, and post-heparin Lipoprotein Lipase activity were measured during an oral glucose tolerance test (OGTT), Body composition was measured using bioelectrical impedance analysis, and subcutaneous and visceral fat mass were assessed with CT-scans. Visceral fat area was higher in OWW (139.7 +/- 10.7 cm(2)) than in OBW (72.3 +/- 3.9 cm(2)) (P < 0.01), as were fasting and 3 h triglyceride concentrations (P < 0.05 for all). OWW also had higher NEFA levels than OBW at 3 and 4 h compared, with OBW (P < 0.05 for both). Fasting cortisol (266 +/- 24 vs. 197 +/- 19 nmol/l; P < 0.05) was higher in OWW than in OBW. These data demonstrate that OWW have higher visceral fat mass than OBW, which may lead to a more atherogenic fasting and postprandial Lipid profile. The higher cortisol levels of the OWW may promote visceral fat deposition. - Punyadeera, C., M-T. van der Merwe, N.J. Crowther, M. Toman, C. P. Schlaphoff, and I. P. Gray. Ethnic differences in lipid metabolism in two groups of obese South African women.

A two-arm cluster randomized control trial to determine the effectiveness of a pressure ulcer prevention bundle for critically ill patients

Purpose This study tested the effectiveness of a pressure ulcer (PU) prevention bundle in reducing the incidence of PUs in critically ill patients in two Saudi intensive care units (ICUs). Design A two-arm cluster randomized experimental control trial. Methods Participants in the intervention group received the PU prevention bundle, while the control group received standard skin care as per the local ICU policies. Data collected included demographic variables (age, diagnosis, comorbidities, admission trajectory, length of stay) and clinical variables (Braden Scale score, severity of organ function score, mechanical ventilation, PU presence, and staging). All patients were followed every two days from admission through to discharge, death, or up to a maximum of 28 days. Data were analyzed with descriptive correlation statistics, Kaplan-Meier survival analysis, and Poisson regression. Findings The total number of participants recruited was 140: 70 control participants (with a total of 728 days of observation) and 70 intervention participants (784 days of observation). PU cumulative incidence was significantly lower in the intervention group (7.14%) compared to the control group (32.86%). Poisson regression revealed the likelihood of PU development was 70% lower in the intervention group. The intervention group had significantly less Stage I (p = 002) and Stage II PU development (p = 026). Conclusions Significant improvements were observed in PU-related outcomes with the implementation of the PU prevention bundle in the ICU; PU incidence, severity, and total number of PUs per patient were reduced. Clinical Relevance Utilizing a bundle approach and standardized nursing language through skin assessment and translation of the knowledge to practice has the potential to impact positively on the quality of care and patient outcome.

Integration over song classification replicates: song variant analysis in the hihi

Human expert analyses are commonly used in bioacoustic studies and can potentially limit the reproducibility of these results. In this paper, a machine learning method is presented to statistically classify avian vocalizations. Automated approaches were applied to isolate bird songs from long field recordings, assess song similarities, and classify songs into distinct variants. Because no positive controls were available to assess the true classification of variants, multiple replicates of automatic classification of song variants were analyzed to investigate clustering uncertainty. The automatic classifications were more similar to the expert classifications than expected by chance. Application of these methods demonstrated the presence of discrete song variants in an island population of the New Zealand hihi (Notiomystis cincta). The geographic patterns of song variation were then revealed by integrating over classification replicates. Because this automated approach considers variation in song variant classification, it reduces potential human bias and facilitates the reproducibility of the results.

Charge transport studies in donor-acceptor block copolymer PDPP-TNT and PC71BM based inverted organic photovoltaic devices processed in room conditions

Diketopyrrolopyrole-naphthalene polymer (PDPP-TNT), a donor-acceptor co-polymer, has shown versatile behavior demonstrating high performances in organic field-effect transistors (OFETs) and organic photovoltaic (OPV) devices. In this paper we report investigation of charge carrier dynamics in PDPP-TNT, and [6,6]-phenyl C71 butyric acid methyl ester (PC71BM) bulk-heterojunction based inverted OPV devices using current density-voltage (J-V) characteristics, space charge limited current (SCLC) measurements, capacitance-voltage (C-V) characteristics, and impedance spectroscopy (IS). OPV devices in inverted architecture, ITO/ZnO/PDPP-TNT:PC71BM/MoO3/Ag, are processed and characterized at room conditions. The power conversion efficiency (PCE) of these devices are measured ∼3.8%, with reasonably good fill-factor 54.6%. The analysis of impedance spectra exhibits electron’s mobility ∼2 × 10−3 cm2V−1s−1, and lifetime in the range of 0.03-0.23 ms. SCLC measurements give hole mobility of 1.12 × 10−5 cm2V−1s−1, and electron mobility of 8.7 × 10−4 cm2V−1s−1.

Stochastic linear multistep methods for the simulation of chemical kinetics

In this paper, we introduce the Stochastic Adams-Bashforth (SAB) and Stochastic Adams-Moulton (SAM) methods as an extension of the tau-leaping framework to past information. Using the theta-trapezoidal tau-leap method of weak order two as a starting procedure, we show that the k-step SAB method with k >= 3 is order three in the mean and correlation, while a predictor-corrector implementation of the SAM method is weak order three in the mean but only order one in the correlation. These convergence results have been derived analytically for linear problems and successfully tested numerically for both linear and non-linear systems. A series of additional examples have been implemented in order to demonstrate the efficacy of this approach.

Cardiometabolic effects of genetic upregulation of the interleukin 1 receptor antagonist: A Mendelian randomisation analysis

Background To investigate potential cardiovascular and other effects of long-term pharmacological interleukin 1 (IL-1) inhibition, we studied genetic variants that produce inhibition of IL-1, a master regulator of inflammation. Methods We created a genetic score combining the effects of alleles of two common variants (rs6743376 and rs1542176) that are located upstream of IL1RN, the gene encoding the IL-1 receptor antagonist (IL-1Ra; an endogenous inhibitor of both IL-1α and IL-1β); both alleles increase soluble IL-1Ra protein concentration. We compared effects on inflammation biomarkers of this genetic score with those of anakinra, the recombinant form of IL-1Ra, which has previously been studied in randomised trials of rheumatoid arthritis and other inflammatory disorders. In primary analyses, we investigated the score in relation to rheumatoid arthritis and four cardiometabolic diseases (type 2 diabetes, coronary heart disease, ischaemic stroke, and abdominal aortic aneurysm; 453 411 total participants). In exploratory analyses, we studied the relation of the score to many disease traits and to 24 other disorders of proposed relevance to IL-1 signalling (746 171 total participants). Findings For each IL1RN minor allele inherited, serum concentrations of IL-1Ra increased by 0·22 SD (95% CI 0·18–0·25; 12·5%; p=9·3 × 10−33), concentrations of interleukin 6 decreased by 0·02 SD (−0·04 to −0·01; −1·7%; p=3·5 × 10−3), and concentrations of C-reactive protein decreased by 0·03 SD (−0·04 to −0·02; −3·4%; p=7·7 × 10−14). We noted the effects of the genetic score on these inflammation biomarkers to be directionally concordant with those of anakinra. The allele count of the genetic score had roughly log-linear, dose-dependent associations with both IL-1Ra concentration and risk of coronary heart disease. For people who carried four IL-1Ra-raising alleles, the odds ratio for coronary heart disease was 1·15 (1·08–1·22; p=1·8 × 10−6) compared with people who carried no IL-1Ra-raising alleles; the per-allele odds ratio for coronary heart disease was 1·03 (1·02–1·04; p=3·9 × 10−10). Per-allele odds ratios were 0·97 (0·95–0·99; p=9·9 × 10−4) for rheumatoid arthritis, 0·99 (0·97–1·01; p=0·47) for type 2 diabetes, 1·00 (0·98–1·02; p=0·92) for ischaemic stroke, and 1·08 (1·04–1·12; p=1·8 × 10−5) for abdominal aortic aneurysm. In exploratory analyses, we observed per-allele increases in concentrations of proatherogenic lipids, including LDL-cholesterol, but no clear evidence of association for blood pressure, glycaemic traits, or any of the 24 other disorders studied. Modelling suggested that the observed increase in LDL-cholesterol could account for about a third of the association observed between the genetic score and increased coronary risk. Interpretation Human genetic data suggest that long-term dual IL-1α/β inhibition could increase cardiovascular risk and, conversely, reduce the risk of development of rheumatoid arthritis. The cardiovascular risk might, in part, be mediated through an increase in proatherogenic lipid concentrations. Funding UK Medical Research Council, British Heart Foundation, UK National Institute for Health Research, National Institute for Health Research Cambridge Biomedical Research Centre, European Research Council, and European Commission Framework Programme 7.

Genome-wide association study to identify genetic determinants of severe asthma

Background: The genetic basis for developing asthma has been extensively studied. However, association studies to date have mostly focused on mild to moderate disease and genetic risk factors for severe asthma remain unclear. Objective: To identify common genetic variants affecting susceptibility to severe asthma. Methods: A genome-wide association study was undertaken in 933 European ancestry individuals with severe asthma based on Global Initiative for Asthma (GINA) criteria 3 or above and 3346 clean controls. After standard quality control measures, the association of 480 889 genotyped single nucleotide polymorphisms (SNPs) was tested. To improve the resolution of the association signals identified, non-genotyped SNPs were imputed in these regions using a dense reference panel of SNP genotypes from the 1000 Genomes Project. Then replication of SNPs of interest was undertaken in a further 231 cases and 1345 controls and a meta-analysis was performed to combine the results across studies. Results: An association was confirmed in subjects with severe asthma of loci previously identified for association with mild to moderate asthma. The strongest evidence was seen for the ORMDL3/GSDMB locus on chromosome 17q12-21 (rs4794820, p=1.03×10 (-8)following meta-analysis) meeting genome-wide significance. Strong evidence was also found for the IL1RL1/IL18R1 locus on 2q12 (rs9807989, p=5.59×10 (-8) following meta-analysis) just below this threshold. No novel loci for susceptibility to severe asthma met strict criteria for genome-wide significance. Conclusions: The largest genome-wide association study of severe asthma to date was carried out and strong evidence found for the association of two previously identified asthma susceptibility loci in patients with severe disease. A number of novel regions with suggestive evidence were also identified warranting further study.

Label-free isolation of a prostate cancer cell among blood cells and the single-cell measurement of drug accumulation using an integrated microfluidic chip

Circulating tumor cells (CTCs) are found in the blood of patients with cancer. Although these cells are rare, they can provide useful information for chemotherapy. However, isolation of these rare cells from blood is technically challenging because they are small in numbers. An integrated microfluidic chip, dubbed as CTC chip, was designed and fabricated for conducting tumor cell isolation. As CTCs usually show multidrug resistance (MDR), the effect of MDR inhibitors on chemotherapeutic drug accumulation in the isolated single tumor cell is measured. As a model of CTC isolation, human prostate tumor cells were mixed with mouse blood cells and the labelfree isolation of the tumor cells was conducted based on cell size difference. The major advantages of the CTC chip are the ability for fast cell isolation, followed by multiple rounds of single-cell measurements, suggesting a potential assay for detecting the drug responses based on the liquid biopsy of cancer patients.