Chromosome-level genome sequence of the Genetically Improved Farmed Tilapia (GIFT, Oreochromis niloticus) highlights regions of introgression with O. mossambicus.

BACKGROUND: The Nile tilapia (Oreochromis niloticus) is the third most important freshwater fish for aquaculture. Its success is directly linked to continuous breeding efforts focusing on production traits such as growth rate and weight. Among those elite strains, the Genetically Improved Farmed Tilapia (GIFT) programme initiated by WorldFish is now distributed worldwide. To accelerate the development of the GIFT strain through genomic selection, a high-quality reference genome is necessary. RESULTS: Using a combination of short (10X Genomics) and long read (PacBio HiFi, PacBio CLR) sequencing and a genetic map for the GIFT strain, we generated a chromosome level genome assembly for the GIFT. Using genomes of two closely related species (O. mossambicus, O. aureus), we characterised the extent of introgression between these species and O. niloticus that has occurred during the breeding process. Over 11 Mb of O. mossambicus genomic material could be identified within the GIFT genome, including genes associated with immunity but also with traits of interest such as growth rate. CONCLUSION: Because of the breeding history of elite strains, current reference genomes might not be the most suitable to support further studies into the GIFT strain. We generated a chromosome level assembly of the GIFT strain, characterising its mixed origins, and the potential contributions of introgressed regions to selected traits.

Management of posterior malleolus fractures: A multicentre cohort study in the United Kingdom.

BACKGROUND: There is a need to assess clinical practice in light of increasing literature recommending fixation of posterior malleolus (PM) fractures. This multicentre study examines treatment approaches, within both Major Trauma Centres and District General Hospitals and role of radiographs and CT scanning. METHODS: Trauma lists and databases were used to identify patients and data collected from electronic and paper medical records and imaging systems between August 2017-18. Analysis of treatment and outcomes was then conducted. RESULTS: One-hundred-and-sixty ankle fractures were included in the study, only 68 ankle fractures underwent CT scanning following initial radiographs and of these, 65 were managed operatively, with 32 undergoing PM fixation. Syndesmotic stabilisation was performed in 9.6% where the PM was fixed. CONCLUSION: CT is still under-utilised, PM fractures that appear to be anything other than small avulsion-type injuries should undergo CT scanning. Syndesmotic stabilisation is statistically less likely to be performed with fixation of the PM.

Large-scale expansion of no-take closures within the Great Barrier Reef has not enhanced fishery production.

A rare opportunity to test hypotheses about potential fishery benefits of large-scale closures was initiated in July 2004 when an additional 28.4% of the 348 000 km2 Great Barrier Reef (GBR) region of Queensland, Australia was closed to all fishing. Advice to the Australian and Queensland governments that supported this initiative predicted these additional closures would generate minimal (10%) initial reductions in both catch and landed value within the GBR area, with recovery of catches becoming apparent after three years. To test these predictions, commercial fisheries data from the GBR area and from the two adjacent (non-GBR) areas of Queensland were compared for the periods immediately before and after the closures were implemented. The observed means for total annual catch and value within the GBR declined from preclosure (2000-2003) levels of 12780 Mg and Australian $160 million, to initial post-closure (2005-2008) levels of 8143 Mg and $102 million; decreases of 35% and 36% respectively. Because the reference areas in the non-GBR had minimal changes in catch and value, the beyond-BACI (before, after, control, impact) analyses estimated initial net reductions within the GBR of 35% for both total catch and value. There was no evidence of recovery in total catch levels or any comparative improvement in catch rates within the GBR nine years after implementation. These results are not consistent with the advice to governments that the closures would have minimal initial impacts and rapidly generate benefits to fisheries in the GBR through increased juvenile recruitment and adult spillovers. Instead, the absence of evidence of recovery in catches to date currently supports an alternative hypothesis that where there is already effective fisheries management, the closing of areas to all fishing will generate reductions in overall catches similar to the percentage of the fished area that is closed.

Condom promotion in Belize: self-efficacy of Belizean nurses.

BACKGROUND: Outside of abstinence, correct and consistent condom use is the single most effective tool to prevent the transmission human immunodeficiency virus (HIV). This is particularly true in countries such as Belize where incidence rates remain high. Women are physiologically at higher risk for HIV, and many feel powerless to insist on condom use. Although nurses are in a position to promote condom use, variables that influence this decision are not clearly understood. In this study, we examined variables that influence a nurses' self-efficacy to promote and teach condom use to women specifically to reduce their HIV risk. METHODS: Data related to self-efficacy, vicarious experience related to condom use promotion and a nurse's sexual relationship power were collected from nurses practising in Belize (n = 60). These data were cross-sectional and collected at the annual nurses' conference. RESULTS: Both years of nursing education and positive vicarious experience promoting and teaching condom use to women were positively correlated to their self-efficacy to do so. Vicarious experience was significantly correlated to self-efficacy in a subgroup of nurses with lower sexual relationship power but not in those with higher sexual relationship power. CONCLUSIONS: When designing HIV continuing education programmes for nurses in Belize, it is important to consider level of nursing education and access to vicarious experience such as mentoring and role modelling. An additional factor to consider is the influence that a nurse's power in her own primary sexual relationship may play in the formation of her self-efficacy.

Genetic mapping in mammals: chromosome map of domestic cat.

A genetic map of 31 biochemical loci located on 17 feline syntenic (linkage) groups has been derived by somatic cell genetic analysis of cat-rodent hybrids. Most of these syntenic groups have been assigned to one of the 19 feline chromosomes. Comparative linkage analysis of the feline biochemical loci and homologous human loci revealed considerable conservation of linkage associations between the primates and the Felidae (order Carnivora). Many of these same linkage groups have not been conserved in the murine genome. The genetic and evolutionary implications of comparative mapping analysis among mammalian species are discussed.

The human homologs of the raf (mil) oncogene are located on human chromosomes 3 and 4.

Two human genes that are homologous to both the murine transforming gene (oncogene) v-raf and the chicken transforming gene v-mil have been mapped by means of human-rodent somatic cell hybrids to human chromosomes previously devoid of known oncogenes. One gene, c-raf-2, which appears to be a processed pseudogene, is located on chromosome 4. The other gene, c-raf-1, which appears to be the active gene, is located on chromosome 3 and has been regionally mapped by chromosomal in situ hybridization to 3p25. This assignment correlates with specific chromosomal abnormalities associated with certain human malignancies.

Gene for T-cell growth factor: location on human chromosome 4q and feline chromosome B1.

T-cell growth factor (TCGF) or interleukin-2 (IL-2), an immunoregulatory lymphokine, is produced by lectin- or antigen-activated mature T lymphocytes and in a constitutive manner by certain T-cell lymphoma cell lines. By means of a molecular clone of human TCGF and DNA extracted from a panel of somatic cell hybrids (rodent cells X normal human lymphocytes), the TCGF structural gene was identified on human chromosome 4. In situ hybridization of the TCGF clone to human chromosomes resulted in significant labeling of the midportion of the long arm of chromosome 4, indicating that the TCGF gene was located at band q26-28. Genomic DNA from a panel of hybrids prepared with HUT-102 B2 cells was examined with the same molecular clone. In this clone of cells, which produces human T-cell leukemia virus, the TCGF gene was also located on chromosome 4 and was apparently not rearranged. The homologous TCGF locus in the domestic cat was assigned to chromosome B1 by using a somatic cell hybrid panel that segregates cat chromosomes. Linkage studies as well as high-resolution G-trypsin banding indicate that this feline chromosome is partially homologous to human chromosome 4.

The promise of comparative genomics in mammals.

Dense genetic maps of human, mouse, and rat genomes that are based on coding genes and on microsatellite and single-nucleotide polymorphism markers have been complemented by precise gene homolog alignment with moderate-resolution maps of livestock, companion animals, and additional mammal species. Comparative genetic assessment expands the utility of these maps in gene discovery, in functional genomics, and in tracking the evolutionary forces that sculpted the genome organization of modern mammalian species.

A Maladaptive Combination of Traits Contributes to the Maintenance of a Drosophila Hybrid Zone.

Drosophila teissieri and D. yakuba diverged approximately 3 mya and are thought to share a large, ancestral, African range [1-3]. These species now co-occur in parts of continental Africa and in west Africa on the island of Bioko [1, 4]. While D. yakuba is a human commensal, D. teissieri seems to be associated with Parinari fruits, restricting its range to forests [4-6]. Genome data indicate introgression, despite no evidence of contemporary hybridization. Here we report the discovery of D. yakuba-D. teissieri hybrids at the interface of secondary forests and disturbed, open habitats on Bioko. We demonstrate that hybrids are the F1 progeny of D. yakuba females and D. teissieri males. At high temperatures like those found on Bioko, D. teissieri females are generally less receptive to mating, and in combination with temperature effects on egg lay and egg-to-adult viability, this decreases the potential for gene flow between female D. teissieri and male D. yakuba relative to the reciprocal cross. Field and laboratory experiments demonstrate that F1 hybrids have a maladaptive combination of D. yakuba behavior and D. teissieri physiology, generating additional barriers to gene flow. Nevertheless, analysis of introgressed and non-introgressed regions of the genome indicate that, while rare, gene flow is relatively recent. Our observations identify precise intrinsic and extrinsic factors that, along with hybrid male sterility, limit gene flow and maintain these species. These data contribute to a growing body of literature that suggests the Gulf of Guinea may be a hotspot for hybridization.

Myocardial energy production and consumption remain balanced during positive inotropic stimulation when coronary flow is restricted to basal rates in rabbit heart.

The effect on myocardial energy balance of increasing oxygen demand without altering basal myocardial perfusion rate was assessed in isolated, isovolumic, retrograde blood perfused rabbit hearts. Myocardial energy requirements were increased with paired stimulation. The capacity of rapid paired stimulation to increase mechanical energy consumption was demonstrated in the presence of increased perfusion with the rate X pressure product and oxygen consumption increasing 86 and 148%, respectively, compared with control values. In contrast, rapid paired stimulation under constant, basal flow conditions did not alter the rate X pressure product, while oxygen extraction and consumption increased only 40% relative to control. Myocardial ATP, creatine-phosphate, and lactate content were identical under control and constant flow-paired stimulation conditions. The results of this study indicate that no detectable energy imbalance was produced by rapid paired stimulation with flow held constant at basal rates. These results suggest that the myocardium does not increase mechanical energy expenditure in response to inotropic or rate stimulation in the presence of restricted flow reserve and are inconsistent with the concept of "demand-induced" or "relative" myocardial ischemia.

Genetic characterization of human c-rel sequences.

We isolated and sequenced a human genomic-DNA segment that is homologous to a portion of v-rel, the transforming gene of reticuloendotheliosis virus (strain T). We also localized the human rel sequences to human chromosome 2 by screening a panel of rodent X human somatic-cell hybrids with the newly described human rel segment.

Coronal Knee Alignment 40 Years after Total Meniscectomy in Adolescents: A Prospective Cohort Study.

INTRODUCTION: Meniscectomies result in altered knee biomechanics and increase contact forces on the operated knee joint. METHODS: We assessed coronal knee alignment in relation to radiological osteoarthritis grading, clinical range of movement and patient reported outcome measures 40 years after total open meniscectomies in adolescence. Thirty eight knees (30 patients) that underwent total open meniscectomy were assessed on standardised weight-bearing anteroposterior radiographs for deviation from 'physiological valgus angle' in either direction (magnitude of malalignment). These values were analysed as per site of meniscectomy for correlations with radiographic scoring systems, range of motion and patient reported outcome measures. RESULTS: Tibiofemoral angle was significantly more varus, and the magnitude of malalignment was significantly higher for the medial meniscectomy patients. The range of flexion was lower for those patients who underwent medial and lateral meniscectomies of the same knee. The patients who underwent meniscectomies of both knees had worse scores for IKDC and KOOS quality of life. Tibiofemoral angle, magnitude of malalignment and range of flexion strongly correlated with Ahlback, and Kellgren and Laurence scores, but patient reported that outcome measures did not correlate. CONCLUSION: Meniscectomy induced malalignment corresponds to the site of meniscectomy and the radiographic degree of osteoarthritis. While malalignment and reduced range of movement correlate well with worsening radiographic signs of arthritis, patient reported outcome measures do not correlate.

Surgical management of the forefoot in patients with rheumatoid arthritis - a review article.

Foot and ankle pathologies cause a significant disease burden on rheumatoid patients. Forefoot pathologies causes pain, callosities and possibly ulceration, and can cause problems with footwear. Forefoot correction in rheumatoid patients has historically comprised of excision of diseased joints. While satisfaction was high with this procedure, complications, changing expectations and improvement in medical therapy have raised expectation of patients, physicians and surgeons alike. This review assesses the role of joint preserving osteotomies and arthrodesis, as well as associated complications. It also describes the role of the multidisciplinary team in the management of these patients.

Insulin, unlike food intake, does not suppress ghrelin in human subjects.

UNLABELLED: Food intake suppresses plasma levels of the gastric peptide ghrelin in humans. We hypothesize that the food intake- suppression of ghrelin could be secondary to the plasma glucose and insulin changes after a meal. The aim of this study was to compare the effect of the administration of a combined pulse of glucose and insulin to the effect of one meal on plasma ghrelin in human subjects. A secondary aim was to study the effect of an oral glucose load on ghrelin levels. METHODS: Experiment 1 (n = 10) studied plasma glucose, insulin, leptin and ghrelin for 6 hours after a 790 kcal liquid meal. In Experiment 2 (n = 7), a subcutaneous pulse of insulin (Humalog 0.03U/kg) and an I.V. infusion of glucose were administered in order to mimic the plasma changes of glucose and insulin after a meal, and plasma ghrelin levels were monitored for 9 h. The OGTT data was used to study the effect of oral glucose on ghrelin. RESULTS: A mixed liquid meal decreased basal serum ghrelin by 26% at 40 minutes (p = 0.009). A 75 gr oral glucose load suppresses ghrelin by 28% at 30 minutes. Contrary to the meal effect, the parenteral administration of insulin and glucose did not suppress serum ghrelin. CONCLUSION: Unlike food intake, the administration of insulin and glucose does not suppress ghrelin levels. These data suggest that the suppressive effect of food intake or oral glucose on serum ghrelin is unlikely mediated by the changes of plasma insulin and glucose observed after the ingestion.

Tyramide signal amplification (TSA)-FISH applied to mapping PCR-labeled probes less than 1 kb in size.

Tyramide signal amplification (TSA)-FISH was used to map one mouse and two human DNA probes of less than 1 kb in size. The two human probes were 319 and 608 bp, and the mouse probe was 855 bp. Probes, made from PCR products, were labeled by incorporating biotin-11-dUTP (human) and biotin-16-dUTP (mouse) during PCR amplification. Signals were readily observed in both interphase and metaphase cells following TSA-FISH for all three genes, whereas conventional FISH experiments produced no signals. The two human ATP-binding cassette (ABC) genes, EST883227 (GenBank Accession No. AA243820) and EST990006 (GenBank Accession No. AA348546), mapped to human chromosomes 7p21 and 17q25. The mouse gene, cmyc (exon 2) mapped to band D2 of mouse chromosome 15. These findings demonstrate the ability of this technique to map small probes (PCR products and expressed sequence tags) of less than 1 kb through highly increased signal amplification.

Assessment of the [18F]fluorodeoxyglucose kinetic model in calculations of myocardial glucose metabolism during ischemia.

The lumped constant--a term in the operational equation of the Sokoloff tracer kinetic model for deoxyglucose that accounts for the difference in transport and phosphorylation between glucose and its analog, deoxyglucose--could potentially vary from normal to ischemic conditions in the heart. To test the stability of the lumped constant during ischemia, we evaluated the ratio of the extraction fraction for (F-18)-fluorodeoxyglucose (FDG) to that for glucose (a measure of the lumped constant if there is no significant dephosphorylation of FDG-6-PO4) and the rate constant for dephosphorylation of FDG-6-PO4 (k4*) in the isolated, arterially perfused interventricular septum of the rabbit during moderate and severe demand-induced and reduced-flow ischemias. The lumped constant and k4* in each of the four ischemic experimental conditions were found not to be significantly different from the value obtained from the nonischemic controls.

Physiological 3 beta-hydroxy-5-ene steroid substrates bind to 3 beta-hydroxysteroid dehydrogenase without the prior binding of cofactor.

3 beta-Hydroxy-delta 5-steroid dehydrogenase (3 beta-HSD)/steroid delta 5-4-isomerase catalyses the conversion of 3 beta-hydroxy-5-ene steroids (e.g. pregnenolone) to 3-oxo-4-ene-steroids (progesterone) in human placenta. Isotope exchange at equilibrium using NAD+/NADH and the 5 alpha-reduced steroids, 5 alpha-androstane-3 beta, 17 beta-diol and 5 alpha-androstan-17 beta-ol-3-one, determined a cofactor-first order of binding for these 3 beta-HSD substrates [1]. Exchange at equilibrium cannot be performed with 3 beta-hydroxy-5-ene steroids because 3 beta-HSD is not reversible with the 5-ene substrates. To compare their cofactor requirements for binding, 3 beta-hydroxy-5-ene and 3 beta-hydroxy-5 alpha-reduced steroids were tested as protectors against the inactivation of purified human placental 3 beta-HSD by 2 alpha-bromoacetoxyprogesterone (2 alpha-BAP) in the presence or absence of cofactor. In incubations without cofactor, pregnenolone or dehydroepiandrosterone dramatically slowed (protected) the rate of 3 beta-HSD inactivation by 2 alpha-BAP, an affinity alkylator that binds specifically at the 3 beta-HSD substrate site. In contrast, 5 alpha-androstan-3 alpha-ol-17-one, 5 alpha-androstane-3 beta, 17 beta-diol, or 11 alpha-acetoxy-5 alpha-pregnan-3,20-dione protected 3 beta-HSD from inactivation by 2 alpha-BAP only in the presence of NADH (0.3 microM) or NAD+ (10 microM). At these low concentrations, neither NADH nor NAD+ slowed the inactivation of 3 beta-HSD by 2 alpha-BAP in the absence of protector-steroid. Further, the 3-oxo-5 alpha-reduced alkylator, 11 alpha-bromoacetoxy-5 alpha-pregnan-3,20-dione (11 alpha-BA-5 alpha-P), did not inactivate 3 beta-HSD in a specific manner. After pre-incubation with NAD+ (10 microM), 11 alpha-BA-5 alpha-P inactivated 3 beta-HSD rapidly and specifically (t1/2 = 3.7 min). 11 alpha-Bromoacetoxyprogesterone inactivated 3 beta-HSD at the same rate (t1/2 = 5.0 min) in the presence or absence of NAD+. These affinity labelling studies confirm the cofactor-first binding order for 3 beta-hydroxy-5 alpha-reduced steroids, and conclusively show that the more important, physiological 3 beta-hydroxy-5-ene substrates bind to 3 beta-HSD without a cofactor requirement.

Site-directed mutagenesis identifies amino acid residues associated with the dehydrogenase and isomerase activities of human type I (placental) 3beta-hydroxysteroid dehydrogenase/isomerase.

3beta-hydroxysteroid dehydrogenase/steroid delta5-->4-isomerase (3beta-HSD/isomerase) was expressed by baculovirus in Spodoptera fungiperda (Sf9) insect cells from cDNA sequences encoding human wild-type I (placental) and the human type I mutants - H261R, Y253F and Y253,254F. Western blots of SDS-polyacrylamide gels showed that the baculovirus-infected Sf9 cells expressed the immunoreactive wild-type, H261R, Y253F or Y253,254F protein that co-migrated with purified placental 3beta-HSD/isomerase (monomeric Mr=42,000 Da). The wild-type, H261R and Y253F enzymes were each purified as a single, homogeneous protein from a suspension of the Sf9 cells (5.01). In kinetic studies with purified enzyme, the H261R mutant enzyme had no 3beta-HSD activity, whereas the Km and Vmax values of the isomerase substrate were similar to the values obtained with the wild-type and native enzymes. The Vmax (88 nmol/min/mg) for the conversion of 5-androstene-3,17-dione to androstenedione by the Y253F isomerase activity was 7.0-fold less than the mean Vmax (620 nmol/min/mg) measured for the isomerase activity of the wild-type and native placental enzymes. In microsomal preparations, isomerase activity was completely abolished in the Y253,254F mutant enzyme, but Y253,254F had 45% of the 3beta-HSD activity of the wild-type enzyme. In contrast, the purified Y253F, wild-type and native enzymes had similar Vmax values for substrate oxidation by the 3beta-HSD activity. The 3beta-HSD activities of the Y253F, Y253,254F and wild-type enzymes reduced NAD+ with similar kinetic values. Although NADH activated the isomerase activities of the H261R and wild-type enzymes with similar kinetics, the activation of the isomerase activity of H261R by NAD+ was dramatically decreased. Based on these kinetic measurements, His261 appears to be a critical amino acid residue for the 3beta-HSD activity, and Tyr253 or Tyr254 participates in the isomerase activity of human type I (placental) enzyme.

Over-expression of human type I (placental) 3 beta-hydroxy-5-ene-steroid dehydrogenase/isomerase in insect cells infected with recombinant baculovirus.

Human type I placental 3 beta-hydroxy-5-ene-steroid dehydrogenase/steroid 5-->4-ene-isomerase (3 beta-HSD/isomerase) synthesizes androstenedione from fetal dehydroepiandrosterone and progesterone from pregnenolone. The full length cDNA that encodes type I 3 beta-HSD/isomerase was inserted into the baculovirus, Autographa californica multiple nucleocapsid polyhedrosis virus, and expressed in Spodoptera fungiperda (Sf-9) insect cells. Western blots showed that the baculovirus-infected Sf-9 cells produced an immunoreactive protein that co-migrated with purified placental 3 beta-HSD/isomerase. Ultracentrifugation localized the expressed enzyme activities in all the membrane-associated organelles of the Sf-9 cell (nuclear, mitochondrial and microsomal). Kinetic studies showed that the expressed enzyme has 3 beta-HSD and isomerase activities. The Michaelis-Menton constant is very similar for the 3 beta-HSD substrate, 5 alpha-androstan-3 beta- ol-17-one, in the Sf-9 cell homogenate (Km = 17.9 microM) and placental microsomes (Km = 16.7 microM). The 3 beta-HSD activity (Vmax = 14.5 nmol/min/mg) is 1.6-fold higher in the Sf-9 cell homogenate compared to placental microsomes (Vmax = 9.1 nmol/min/mg). The Km values are almost identical for the isomerase substrate, 5-androstene-3,17-dione, in the Sf-9 cell homogenate (Km = 14.7 microM) and placental microsomes (Km = 14.4 microM). The specific isomerase activity is 1.5-fold higher in the Sf-9 cells (Vmax = 25.7 nmol/min/mg) relative to placenta (Vmax = 17.2 nmol/min/mg). These studies show that our recombinant baculovirus system over-expresses fully active enzyme that is kinetically identical to native 3 beta-HSD/isomerase in human placenta.