Importing genetically altered animals: ensuring quality.

The reproducibility of research using laboratory animals requires reliable management of their quality, in particular of their genetics, health and environment, all of which contribute to their phenotypes. The point at which these biological materials are transferred between researchers is particularly sensitive, as it may result in a loss of integrity of the animals and/or their documentation. Here, we describe the various aspects of laboratory animal quality that should be confirmed when sharing rodent research models. We also discuss how repositories of biological materials support the scientific community to ensure the continuity of the quality of laboratory animals. Both the concept of quality and the role of repositories themselves extend to all exchanges of biological materials and all networks that support the sharing of these reagents.

Community based research network: opportunities for coordination of care, public health surveillance, and farmworker research.

INTRODUCTION: The lack of aggregated longitudinal health data on farmworkers has severely limited opportunities to conduct research to improve their health status. To correct this problem, we have created the infrastructure necessary to develop and maintain a national Research Data Repository of migrant and seasonal farmworker patients and other community members receiving medical care from Community and Migrant Health Centers (C/MHCs). Project specific research databases can be easily extracted from this repository. METHODS: The Community Based Research Network (CBRN) has securely imported and merged electronic health records (EHRs) data from five geographically dispersed C/MHCs. To demonstrate the effectiveness of our data aggregation methodologies, we also conducted a small pilot study using clinical, laboratory and demographic data from the CBRN Data Repository from two initial C/MHCs to evaluate HbA1c management. RESULTS: Overall, there were 67,878 total patients (2,858 farmworkers) that were seen by two C/MHCs from January to August 2013. A total of 94,189 encounters were captured and all could be linked to a unique patient. HbA1c values decreased as the number of tests or intensity of testing increased. CONCLUSION: This project will inform the foundation for an expanding collection of C/MHC data for use by clinicians for medical care coordination, by clinics to assess quality of care, by public health agencies for surveillance, and by researchers under Institutional Review Board (IRB) oversight to advance understanding of the needs and capacity of the migrant and seasonal farmworker population and the health centers that serve them. Approved researchers can request data that constitute a Limited Data Set from the CBRN Data Repository to establish a specific research database for their project.

The Flannotator--a gene and protein expression annotation tool for Drosophila melanogaster.

SUMMARY: Understanding developmental processes and building towards integrative systems biology require detailed knowledge of the spatio-temporal expression of genes and proteins. We have developed a software package for collecting, storing and searching the annotation of protein or gene expression patterns in Drosophila melanogaster. Using standard Drosophila anatomy and Gene Ontologies, the system can readily capture expression patterns at any stage of development and in all recognized tissue types as well as details of sub-cellular localization. The web-based system allows multiple groups to work in collaboration and share images and annotation. AVAILABILITY: http://www.flannotator.org.uk/.

Chemosensory control of surface antigen switching in the nematode Caenorhabditis elegans.

Nematodes change their surface compositions in response to environmental signals, which may allow them to survive attacks from microbial pathogens or host immune systems. In the free-living species Caenorhabditis elegans, wild-type worms are induced to display an L1 (first larval stage) surface epitope at later larval stages when grown on an extract of spent culture medium (Inducible Larval Display or ILD). Before this study, it was not known whether ILD was regulated by the well-characterized, neurologically based chemical senses of C. elegans, which mediate other behavioural and developmental responses to environmental signals such as chemotaxis and formation of the facultatively arrested dauer larva stage. We show here that ILD requires the activities of three genes that are essential for the function of the C. elegans chemosensory neurons. ILD was abolished in chemotaxis-defective che-3, osm-3 and tax-4 mutants. In contrast, chemotaxis-defective mutants altered in a different gene, srf-6, show constitutive display of the L1 epitope on all four larval stages. The ILD-defective che-3, osm-3 and tax-4 mutations blocked the constitutive larval display of an srf-6 mutant. Combining srf-6 and certain dauer-constitutive mutations in double mutants enhanced constitutive dauer formation, consistent with the idea that srf-6 acts in parallel with specific components of the dauer formation pathway. These results taken together are consistent with the hypothesis that ILD is triggered by environmental signals detected by the nematode's chemosensory neurons.

Canine RPGRIP1 mutation establishes cone-rod dystrophy in miniature longhaired dachshunds as a homologue of human Leber congenital amaurosis.

Cone-rod dystrophy 1 (cord1) is a recessive condition that occurs naturally in miniature longhaired dachshunds (MLHDs). We mapped the cord1 locus to a region of canine chromosome CFA15 that is syntenic with a region of human chromosome 14 (HSA14q11.2) containing the retinitis pigmentosa GTPase regulator-interacting protein 1 (RPGRIP1) gene. Mutations in RPGRIP1 have been shown to cause Leber congenital amaurosis, a group of retinal dystrophies that represent the most common genetic causes of congenital visual impairment in infants and children. Using the newly available canine genome sequence we sequenced RPGRIP1 in affected and carrier MLHDs and identified a 44-nucleotide insertion in exon 2 that alters the reading frame and introduces a premature stop codon. All affected and carrier dogs within an extended inbred pedigree were homozygous and heterozygous, respectively, for the mutation. We conclude the mutation is responsible for cord1 and demonstrate that this canine disease is a valuable model for exploring disease mechanisms and potential therapies for human Leber congenital amaurosis.

MAMMOT--a set of tools for the design, management and visualization of genomic tiling arrays.

UNLABELLED: The MAMMOT software suite is a collection of Perl and PHP scripts for designing, annotating and visualizing genome tiling arrays to, for example, facilitate studies into the epigenetics of gene regulation. The web design allows rapid experimental data entry from multiple users, and results can easily be shared between groups and individuals. AVAILABILITY: http://www.mammot.org.uk/ CONTACT: e.ryder@gen.cam.ac.uk.

Plasma fibrinogen level and the risk of major cardiovascular diseases and nonvascular mortality: an individual participant meta-analysis.

CONTEXT: Plasma fibrinogen levels may be associated with the risk of coronary heart disease (CHD) and stroke. OBJECTIVE: To assess the relationships of fibrinogen levels with risk of major vascular and with risk of nonvascular outcomes based on individual participant data. DATA SOURCES: Relevant studies were identified by computer-assisted searches, hand searches of reference lists, and personal communication with relevant investigators. STUDY SELECTION: All identified prospective studies were included with information available on baseline fibrinogen levels and details of subsequent major vascular morbidity and/or cause-specific mortality during at least 1 year of follow-up. Studies were excluded if they recruited participants on the basis of having had a previous history of cardiovascular disease; participants with known preexisting CHD or stroke were excluded. DATA EXTRACTION: Individual records were provided on each of 154,211 participants in 31 prospective studies. During 1.38 million person-years of follow-up, there were 6944 first nonfatal myocardial infarctions or stroke events and 13,210 deaths. Cause-specific mortality was generally available. Analyses involved proportional hazards modeling with adjustment for confounding by known cardiovascular risk factors and for regression dilution bias. DATA SYNTHESIS: Within each age group considered (40-59, 60-69, and > or =70 years), there was an approximately log-linear association with usual fibrinogen level for the risk of any CHD, any stroke, other vascular (eg, non-CHD, nonstroke) mortality, and nonvascular mortality. There was no evidence of a threshold within the range of usual fibrinogen level studied at any age. The age- and sex- adjusted hazard ratio per 1-g/L increase in usual fibrinogen level for CHD was 2.42 (95% confidence interval [CI], 2.24-2.60); stroke, 2.06 (95% CI, 1.83-2.33); other vascular mortality, 2.76 (95% CI, 2.28-3.35); and nonvascular mortality, 2.03 (95% CI, 1.90-2.18). The hazard ratios for CHD and stroke were reduced to about 1.8 after further adjustment for measured values of several established vascular risk factors. In a subset of 7011 participants with available C-reactive protein values, the findings for CHD were essentially unchanged following additional adjustment for C-reactive protein. The associations of fibrinogen level with CHD or stroke did not differ substantially according to sex, smoking, blood pressure, blood lipid levels, or several features of study design. CONCLUSIONS: In this large individual participant meta-analysis, moderately strong associations were found between usual plasma fibrinogen level and the risks of CHD, stroke, other vascular mortality, and nonvascular mortality in a wide range of circumstances in healthy middle-aged adults. Assessment of any causal relevance of elevated fibrinogen levels to disease requires additional research.

New skills for a new age: leading the introduction of public health concepts in healthcare curricula.

Health policy in the UK is going through significant changes. At the heart of the transformation is a dedicated focus on public health. The new primary-care-based health system will not only be premised on a specialist public health workforce, but also on broader based public-health-oriented health professionals. Within primary care, widening the foundation of health professionals with public health competencies suggests that higher education bodies will need to adapt their curricula to an approach that highlights population-based health principles, preventive philosophy, and public health concepts and methods. The first part of this paper describes the mapping of the public health content of healthcare curricula at one university in England, based on the 10 public health standard areas of competencies of the Faculty of Public Health Medicine. The second part examines, through the findings of a strengths, weaknesses, opportunities, threats (SWOT) analysis, the factors that advocates for a public-health-oriented educational strategy must examine before embarking on the instigation and development of public health concepts in the healthcare curricula. The aspects that necessitate consideration include strengths such as the prevailing policy, market forces, commitment, and motivation to the effort, and the availability of resources, information and external contacts. Features such as political drive and advocacy, interest in the education debate, collaborative links through joint working and partnerships, and ongoing internal reforms and restructuring could all act as opportunities. However, resistance and anxiety are to be expected, the operationalization of the effort and empowerment of those leading it need to be thought about, and issues of control and interests are critical. The presence of conflicting priorities and competition or the lack of vision and directives, or uncertainty about change, could act as threats and barriers to the effort. If shifting the 'traditional' healthcare curricula to a more 'innovative' public-health-oriented one is to be a success, administrators of educational change will need to take into account a 'melange' of factors and stakeholders involved in a gradual and incremental process.

Lean adolescents with increased risk for metabolic syndrome

The aim of the present study was to determine in adolescents the relationship between insulin levels and body mass index (BMI), body fat distribution, diet, life style and lipid profile. We studied 167 adolescents (68 boys and 99 girls) whose ages ranged from 14 to 17 years. A detailed medical (including pubertal stage) and nutritional record was obtained from each subject. Biochemical measurements included fasting serum insulin, glucose, total cholesterol (TC), triglycerides (Tg), HDL-C, LDL-C and VLDL-C. HOMA insulin resistance (IR) and HOMA beta-cell function (beta-cell) were calculated. Insulin levels were over 84 pmol/L (cut off normal value in our lab) in 56 per cent of the boys and 43 per cent of the girls. Thirty-seven percent of lean adolescents whose BMI was 21.5 +/- 1.9 kg/m2 presented higher fasting insulin levels. HOMA IR, Tg, systolic (SBP) and diastolic blood pressure (DBP) values when compared to a lean normoinsulinemic group. Insulin levels were correlated (p < 0.01) with body mass index. Both boys and girls in the highest BMI quartile (BMI > 24 kg/m2) had significantly higher serum insulin, HOMA beta-cell, and Tg levels, and the lowest HDL-C levels. A high-energy intake rich in saturated fat and low physical activity were found in this lean but metabolically altered adolescents. We conclude that even with a BMI as low as 21 kg/m2 an inappropriate diet and low physical activity might be responsible for the high insulin levels and dislipidemias in adolescents.

Chromosome-specific single-locus FISH probes allow anchorage of an 1800-marker integrated radiation-hybrid/linkage map of the domestic dog genome to all chromosomes.

We present here the first fully integrated, comprehensive map of the canine genome, incorporating detailed cytogenetic, radiation hybrid (RH), and meiotic information. We have mapped a collection of 266 chromosome-specific cosmid clones, each containing a microsatellite marker, to all 38 canine autosomes by fluorescence in situ hybridization (FISH). A 1500-marker RH map, comprising 1078 microsatellites, 320 dog gene markers, and 102 chromosome-specific markers, has been constructed using the RHDF5000-2 whole-genome radiation hybrid panel. Meiotic linkage analysis was performed, with at least one microsatellite marker from each dog autosome on a panel of reference families, allowing one meiotic linkage group to be anchored to all 38 dog autosomes. We present a karyotype in which each chromosome is identified by one meiotic linkage group and one or more RH groups. This updated integrated map, containing a total of 1800 markers, covers >90% of the dog genome. Positional selection of anchor clones enabled us, for the first time, to orientate nearly all of the integrated groups on each chromosome and to evaluate the extent of individual chromosome coverage in the integrated genome map. Finally, the inclusion of 320 dog genes into this integrated map enhances existing comparative mapping data between human and dog, and the 1000 mapped microsatellite markers constitute an invaluable tool with which to perform genome scanning studies on pedigrees of interest.

Statistics: detecting differences among groups.

Some of the statistical tests most commonly used (and misused) in biological research are presented here. The tests discussed are those used for comparisons among groups (e.g., t test and ANOVA). The purpose of this appendix is to enable the investigator to determine rapidly the most appropriate way to analyze data, and to point out some of the most common errors to avoid.

Statistics for the molecular biologist: group comparisons.

In this appendix, some of the statistical tests most commonly used (and misused) in biological research are discussed. These tests are used for comparisons among groups (e.g., t test and ANOVA). A number of other important areas (e.g., linear regression, correlation, and goodness-of-fit testing) are not covered. The purpose is to enable the reader to determine rapidly the most appropriate way to analyze data, and to point out some of the most common errors to avoid.

Characterization of Distinct Tombusviruses that Cause Diseases of Lettuce and Tomato in the Western United States.

A soilborne disease of lettuce, associated with necrosis and dieback, has been found with increasing frequency in California and Arizona over the last 10 years. An isometric virus, serologically related to Tomato bushy stunt virus (TBSV), was consistently isolated from lettuce plants with these disease symptoms. Back-inoculation to healthy lettuce plants and subsequent reisolation of the virus from symptomatic lettuce leaves suggested that this virus was the causal agent of this disease. A tombusvirus was also associated with a necrosis disease of greenhouse-grown tomatoes in Colorado and New Mexico. Complementary DNA representing the 3' end of viral genomic RNAs recovered from diseased lettuce and tomato plants had identical nucleotide sequences. However, these sequences were divergent (12.2 to 17.1%) from sequences of the previously described strains of TBSV, Petunia asteroid mosaic virus (PAMV), Artichoke mottled crinkle virus, and Carnation Italian ringspot virus. Additional tombusvirus isolates were recovered from diseased lettuce and tomato plants and these were most closely related to the TBSV-cherry strain (synonymous with PAMV) and to Cucumber necrosis virus based on comparison of 3'-end sequences (0.1 to 0.6% and 4.8 to 5.1% divergence, respectively). Western blot analysis revealed that the new tombusvirus isolated from diseased lettuce and tomato plants in the western United States is serologically distinct from previously described tombusvirus species and strains. Based on genomic and serological properties, we propose to classify this virus as a new tombusvirus species and name it Lettuce necrotic stunt virus.

[Glucose/insulin response to a glucose overload in subjects at risk for type 2 diabetes]. / Respuesta de la Glucosa/insulina a una sobrecarga glucosada en sujetos con riesgo a diabetes tipo 2.

With the purpose of determining how certain risk factors for type 2 diabetes such as family history of diabetes, obesity and dyslipidemia, affect the glucose-insulin response to a glucose challenge, 135 individuals (77 women and 58) men were studied. Their ages ranged from 20-68 years, their basal glycemic values were less than 110 mg/dL but they were considered at risk for diabetes due to the presence of one or more of those factors. We found that the presence of those risk factors did not affect the glycemic response in any case. However, the basal insulin levels as well as the post-challenge values were increased significantly (p < 0.0001) by the presence of obesity in men as well as in women. Dyslipidemia increased the basal and post challenge glucose insulin values only in men (p < 0.002). The coexistence of obesity and family history of diabetes provoked a decrease in the basal insulin levels as well as in the insulin response to glucose. We conclude that, without alteration of the glycemic response, the presence of risk factors as obesity, dyslipidemia or family history of diabetes leads to basal hyperinsulinemia, as well as glucose stimulated hyperinsulinemia, however the coexistence of obesity and family history of diabetes, is responsible for a deficit in the insulin secretion by the pancreas.

Computational methods for single-point and multipoint analysis of genetic variants associated with a simulated complex disorder in a general population.

Several techniques for association analysis have been applied to simulated genetic data for a general population. We describe and compare the performance of three single-point methods and two multipoint approaches rooted in machine learning and data mining.

Needs of older people in the community: issues for PCGs and PCTs.

National policy (Department of Health, 1997, 2000) is directing care provision to being appropriate to meet people's needs; to be effective; evidence-based; efficient and economic. Primary care groups (PCGs) and trusts (PCTs) have been identified as being the organizations to deliver such care. Since the introduction of the NHS and Community Care Act 1990, certain issues have been highlighted as having implications for the provision of an 'ideal' service for older people on the community. These issues focus around three main areas: assessment of need; working in partnership and quality of service provision. This article discusses these issues and their implications for PCGs/PCTs in relation to supporting older people in the community.