The utility of multiple genomic technologies for interpretation of modern next generation sequencing: A novel case of three FANCA gene variants resulting in autosomal recessive Fanconi anaemia.

Fanconi anaemia (FA) is a rare autosomal recessive condition resulting in changes in the FANC gene family. This report describes a case of Fanconi anaemia in a family with complex biallelic variants. The patient is a 32-year-old female diagnosed with FA on cascade testing during childhood with chromosome breakage studies. On examination she had a fixed deformity of the right thumb and the proximal interphalangeal joint was immobile. Her brother shared this radial abnormality and had FA, requiring a bone marrow transplant. She presented in adulthood seeking further BRCA advice and had next generation sequencing that showed three variants in the FANCA gene. One allele a known pathogenic change, the other had two sequence variants in tandem that have been reported as variants of uncertain significance. There is one other unrelated case of these two variants occurring together in cis, resulting in Fanconi anaemia. This case is an interesting example of three variants in the FANCA gene, one allele with a pathogenic deletion and the other with a single complex allele made up of two missense variants of uncertain significance, likely manifesting with FA. It highlights the utility of different genetic technologies in the interpretation of next generation sequencing.

Whole angiosperms Wolffia columbiana disperse by gut passage through wildfowl in South America.

For the first time to our knowledge, we demonstrate that whole angiosperm individuals can survive gut passage through birds, and that this occurs in the field. Floating plants of the genus Wolffia are the smallest of all flowering plants. Fresh droppings of white-faced whistling duck Dendrocygna viduata ( n = 49) and coscoroba swan Coscoroba coscoroba ( n = 22) were collected from Brazilian wetlands. Intact Wolffia columbiana were recovered from 16% of D. viduata and 32% of Coscoroba samples (total = 164 plantlets). The viability of plants was tested, and asexual reproduction was confirmed. Wolffia columbiana is an expanding alien in Europe. Avian endozoochory of asexual angiosperm propagules may be an important, overlooked dispersal means for aquatic plants, and may contribute to the invasive character of alien species.

ABCG2 loss-of-function polymorphism predicts poor response to allopurinol in patients with gout.

Many patients fail to achieve the recommended serum urate (SU) target (<6 mgdl-1) with allopurinol. The aim of our study was to examine the association of ABCG2 with SU target in response to standard doses of allopurinol using a cohort with confirmed adherence. Good response was defined as SU<6 mgdl-1 on allopurinol ⩽300 mgd-1 and poor response as SU⩾6 mgdl-1 despite allopurinol >300 mgd-1. Adherence was confirmed by oxypurinol concentrations. ABCG2 genotyping was performed using pre-designed single nucleotide polymorphism (SNP) TaqMan assays. Of 264 patients, 120 were good responders, 68 were poor responders and 76 were either non-adherent or could not be classified. The minor allele of ABCG2 SNP rs2231142 conferred a significantly increased risk of poor response to allopurinol (odds ratio=2.71 (1.70-4.48), P=6.0 × 10-5). This association remained significant after adjustment for age, sex, body mass index, ethnicity, estimated glomerular filtration rate, diuretic use and SU off urate-lowering therapy. ABCG2 rs2231142 predicts poor response to allopurinol, as defined by SU⩾6 mgdl-1 despite allopurinol >300 mgd-1.

Low-Mach-number turbulence in interstellar gas revealed by radio polarization gradients.

The interstellar medium of the Milky Way is multiphase, magnetized and turbulent. Turbulence in the interstellar medium produces a global cascade of random gas motions, spanning scales ranging from 100 parsecs to 1,000 kilometres (ref. 4). Fundamental parameters of interstellar turbulence such as the sonic Mach number (the speed of sound) have been difficult to determine, because observations have lacked the sensitivity and resolution to image the small-scale structure associated with turbulent motion. Observations of linear polarization and Faraday rotation in radio emission from the Milky Way have identified unusual polarized structures that often have no counterparts in the total radiation intensity or at other wavelengths, and whose physical significance has been unclear. Here we report that the gradient of the Stokes vector (Q, U), where Q and U are parameters describing the polarization state of radiation, provides an image of magnetized turbulence in diffuse, ionized gas, manifested as a complex filamentary web of discontinuities in gas density and magnetic field. Through comparison with simulations, we demonstrate that turbulence in the warm, ionized medium has a relatively low sonic Mach number, M(s) ≲ 2. The development of statistical tools for the analysis of polarization gradients will allow accurate determinations of the Mach number, Reynolds number and magnetic field strength in interstellar turbulence over a wide range of conditions.

Twenty-eight loci that influence serum urate levels: analysis of association with gout.

OBJECTIVES: Twenty-eight genetic loci are associated with serum urate levels in Europeans. Evidence for association with gout at most loci is absent, equivocal or not replicated. Our aim was to test the loci for association with gout meeting the American College of Rheumatology gout classification criteria in New Zealand European and Polynesian case-control sample sets. METHODS: 648 European cases and 1550 controls, and 888 Polynesian (Ma¯ori and Pacific) cases and 1095 controls were genotyped. Association with gout was tested by logistic regression adjusting for age and sex. Power was adequate (>0.7) to detect effects of OR>1.3. RESULTS: We focused on 24 loci without previous consistent evidence for association with gout. In Europeans, we detected association at seven loci, one of which was the first report of association with gout (IGF1R). In Polynesian, association was detected at three loci. Meta-analysis revealed association at eight loci-two had not previously been associated with gout (PDZK1 and MAF). In participants with higher Polynesian ancestry, there was association in an opposing direction to Europeans at PRKAG2 and HLF (HLF is the first report of association with gout). There was obvious inconsistency of gout association at four loci (GCKR, INHBC, SLC22A11, SLC16A9) that display very similar effects on urate levels. CONCLUSIONS: We provide the first evidence for association with gout at four loci (IGF1R, PDZK1, MAF, HLF). Understanding why there is lack of correlation between urate and gout effect sizes will be important in understanding the aetiology of gout.

MUTYH-Associated Polyposis: The Irish Experience>.

MUTYH is involved in DNA damage repair. Bi-allelic MUTYH mutations predispose to polyposis and gastrointestinal malignancies, distinct genetically from autosomal dominant familial adenomatous polyposis coli. Two common European MUTYH mutations account for 90% of MUTYH-associated polyposis (MAP). We aimed to examine the incidence of MAP in Ireland. A retrospective cohort study was undertaken. Patients undergoing MUTYH testing from 2003-2016 were identified by searching electronic databases using terms "MUTYH" and "MYH". Phenotypic and genotypic details were obtained by chart review. Bi-allelic mutations were confirmed in 26 individuals (17 families), of whom 16 (62%) developed colorectal malignancies, and 22(85%) polyposis. Eleven families had bi-allelic status for one/both common European mutations. Regional variation was noted, with over-representation of bi-allelic mutation carriers in the South-west of Ireland. MAP is under-diagnosed in Ireland. Increased awareness is required to facilitate appropriate identification and surveillance of bi-allelic mutation carriers for colorectal pathology.

Quality control for retinal OCT in multiple sclerosis: validation of the OSCAR-IB criteria.

BACKGROUND: Retinal optical coherence tomography (OCT) permits quantification of retinal layer atrophy relevant to assessment of neurodegeneration in multiple sclerosis (MS). Measurement artefacts may limit the use of OCT to MS research. OBJECTIVE: An expert task force convened with the aim to provide guidance on the use of validated quality control (QC) criteria for the use of OCT in MS research and clinical trials. METHODS: A prospective multi-centre (n = 13) study. Peripapillary ring scan QC rating of an OCT training set (n = 50) was followed by a test set (n = 50). Inter-rater agreement was calculated using kappa statistics. Results were discussed at a round table after the assessment had taken place. RESULTS: The inter-rater QC agreement was substantial (kappa = 0.7). Disagreement was found highest for judging signal strength (kappa = 0.40). Future steps to resolve these issues were discussed. CONCLUSION: Substantial agreement for QC assessment was achieved with aid of the OSCAR-IB criteria. The task force has developed a website for free online training and QC certification. The criteria may prove useful for future research and trials in MS using OCT as a secondary outcome measure in a multi-centre setting.

Retinal axonal loss in very early stages of multiple sclerosis.

BACKGROUND AND PURPOSE: The lack of surrogates of clinical progression has limited the design of neuroprotection trials in multiple sclerosis (MS). Our aim was to study the association between time-domain optical coherence tomography measures and clinical and magnetic resonance imaging outcomes in early MS. METHODS: Forty-three relapsing-remitting MS patients within 1 year of onset were followed for up to 3 years. RESULTS: The peripapillary retinal nerve fiber layer (RNFL) decreased annually by 2 µm (95% confidence interval -3.89, -0.11; P = 0.038). The RNFL tended to be associated with normalized normal appearing white matter volume in cross-sectional (P = 0.08) and longitudinal analyses (P = 0.06). CONCLUSIONS: There is substantial RNFL loss even in very early MS. Our data suggest that retinal axonal atrophy is associated with atrophy in global white matter volume in early MS.

The antibody-based targeted delivery of TNF in combination with doxorubicin eradicates sarcomas in mice and confers protective immunity.

BACKGROUND: Soft-tissue sarcomas are a group of malignancies of mesenchymal origin, which typically have a dismal prognosis if they reach the metastatic stage. The observation of rare spontaneous remissions in patients suffering from concomitant bacterial infections had triggered the clinical investigation of the use of heat-killed bacteria as therapeutic agents (Coley's toxin), which induced complete responses in patients in the pre-chemotherapy era and is now known to mediate substantial elevations in serum TNF levels. METHODS: We designed and developed a novel immunocytokine based on murine TNF sequentially fused to the antibody fragment F8 (specific to extra-domain A of fibronectin). The antitumor activity was studied in two syngeneic murine sarcoma models. RESULTS: The L19 antibody (specific to extra-domain B of fibronectin) has shown by SPECT imaging procedures to selectively localise on sarcoma in a patient with a peripheral nerve sheath tumour, and immunohistochemical analysis of human soft-tissue sarcoma samples showed comparable antigen expression of EDA and EDB. The antibody-based pharmacodelivery of TNF by the fusion protein 'F8-TNF' to oncofetal fibronectin in sarcoma-bearing mice leads to complete and long-lasting tumour eradications when administered in combination with doxorubicin, the first-line drug for the treatment of sarcomas in humans. Doxorubicin alone did not display any therapeutic effect in both tested models of this study. The cured mice had acquired protective immunity against the tumour, as they rejected subsequent challenges with sarcoma cells. CONCLUSION: The findings of this study provide a rationale for the clinical study of the fully human immunocytokine L19-TNF in combination with doxorubicin in patients with soft-tissue sarcoma.

Loss of heterozygosity on chromosome 16p13.3 in hamartomas from tuberous sclerosis patients.

Tuberous sclerosis (TSC) is an autosomal dominant condition with characteristic skin lesions, mental handicap, seizures and the development of hamartomas in the brain, heart, kidneys and other organs. Linkage studies have shown locus heterogeneity with a TSC gene mapped to chromosome 9q34 and a second, recently identified on 16p13.3. We have analysed DNA markers in eight hamartomas and one tumour from TSC patients and found allele loss on 16p13.3 in three angiomyolipomas, one cardiac rhabdomyoma, one cortical tuber and one giant cell astrocytoma. We suggest that the TSC gene on 16p13.3 functions like a tumour suppressor gene, in accordance with Knudsen's hypothesis.

Structural and electrochemical insights into novel Wadsley Roth Nb7Ti1.5Mo1.5O25 and Ta7Ti1.5Mo1.5O25 anodes for Li-ion battery application.

Niobium based anodes are gaining increasing popularity for application in high-power lithium-ion batteries, due to their high theoretical capacities, inherent safety at high current densities, and long-term stability. Here, we report the discovery and characterisation of a new Wadsley Roth niobate system, Nb7Ti1.5Mo1.5O25, showing that it is isostructural with known systems: Nb9PO25 and Nb9VO25. To evaluate the material's electrochemical performance, including performance at high current densities (for potential high power applications), and long-term stability, Li half-coin cells were prepared. The material showed an initial capacity of 268(9) mA h g-1 at 0.01 A g-1 (voltage range of 2.5-1.0 V). However, in subsequent cycles, some of this initial capacity is lost, which is attributed to Li trapping associated with the presence of reducible MoO4 units, similar to the situation observed for isostructural Nb9VO25. After this initial irreversible capacity loss, the material showed good performance at high current density rates, such that at 2 A g-1 and 4 A g-1 respective capacities of 132(10) mA h g-1 and 115(14) mA g-1 were delivered. Moreover, the material showed respectable capacity retention (97%) after being cycled for 100 cycles at 0.2 A g-1. In order to identify the different Nb, Ti, Mo redox couples involved in this system, a Ta analogue was also synthesized (Ta7Ti1.5Mo1.5O25) and the electrochemical performance for this phase is also reported. This phase shows a lower initial capacity at 0.01 A g-1 (140(3) mA h g-1) than the Nb analogue in the same voltage range, which can be increased (225 mA h g-1) if a lower cutoff voltage (0.5 V) is applied. The capacity retention for this Ta system after 100 cycles at 0.2 A g-1 is similar to the Nb analogue (97%). Further work has explored whether the Nb-Ti-Mo contents could be varied, and these results showed that single phase Nb10-2xTixMoxO25 samples could be prepared for 1.5 ≤ x ≤ 1.75, and electrochemical testing results for the x = 1.75 endmember are also reported. Overall, this research highlights the synthesis and electrochemical characterisation of two new Wadsley Roth phases, and further highlights the challenges associated with the presence of reducible cations in tetrahedral sites in such structures with respect to minimising initial irreversible capacity loss.

Rate and risk predictors for development of self-reported type-2 diabetes mellitus over a 5-year period: the SHIELD study.

AIMS: This investigation determined the proportion of adults newly diagnosed as having type-2 diabetes mellitus (T2DM), and ascertained risk predictors for development of self-reported T2DM. METHODS: The US Study to Help Improve Early evaluation and management of risk factors Leading to Diabetes (SHIELD) survey was a 5-year longitudinal study of adults with and without diabetes mellitus. Adults completed a baseline health questionnaire in 2004 and ≥1 annual follow-up survey through 2009. Respondents with no self-reported diagnosis of diabetes at baseline were followed to measure rate of and assess risk factors for development of T2DM over 5 years. RESULTS: Among 8582 respondents without diabetes at baseline, 622 (7.2%) reported a diagnosis of T2DM over the subsequent 5 years. Increasing age, family history of T2DM, body mass index ≥30 kg/m(2), abdominal obesity, excessive thirst, asthma, gestational diabetes and 'high blood sugar without diabetes' significantly increased the risk of developing T2DM (p < 0.05 for each). Good to excellent health status and self-reported circulatory problems decreased the risk (p < 0.05 for each). CONCLUSIONS: Among this representative US adult population, the rate of developing T2DM was 7.2% over 5 years. Predictors of T2DM diagnosis identified in this analysis were readily obtainable via self-report.

Changes in water level, land use, and hydrological budget in a semi-permanent playa lake, southwest Spain.

Medina playa lake, a Ramsar site in western Andalusia, is a brackish lowland lake of 120 ha with an average depth of 1 m. Water flows into Medina from its 1,748-ha watershed, but the hydrology of the lake has not previously been studied. This paper describes the application of a water budget model on a monthly scale over a 6-year period, based on a conceptual hydrological model, and considers different future scenarios after calibration to improve the understanding of the lake's hydrological functioning. Climatic variables from a nearby weather station and observational data (water-level evolution) were used to develop the model. Comparison of measured and predicted values demonstrated that each model component provided a reasonable output with a realistic interaction among the components. The model was then used to explore the potential consequences of land-use changes. Irrigation of olive groves would significantly reduce both the hydroperiod (becoming dry 15% of the time) and the average depth of the lake (water level <0.5 m 40% of the time). On the other hand, removal of an artificial overflow would double the average flooded surface area during high-water periods. The simulated water balance demonstrates that the catchment outputs are dominated by lake evaporation and surface outflow from the lake system to a creek. Discrepancies between predicted and observed water levels identify key areas of uncertainty for future empirical research. The study provides an improved basis for future hydrological management of the catchment and demonstrates the wider utility of this methodology in simulating this kind of system. This methodology provides a realistic appraisal of potential land-use management practices on a catchment-wide scale and allows predictions of the consequences of climate change.

Clinical evidence of efficient tumor targeting based on single-chain Fv antibody selected from a combinatorial library.

We present a system for cancer targeting based on single-chain Fv (scFv) antibodies selected from combinatorial libraries, produced in bacteria and purified by using an engineered tag. Combinatorial libraries of scFv genes contain great diversity, and scFv antibodies with characteristics optimized for a particular task can be selected from them using filamentous bacteriophage. We illustrate the benefits of this system by imaging patients with carcinoembryonic antigen (CEA)-producing cancers using an iodine-123 labeled scFv anti-CEA selected for high affinity. All known tumor deposits were located, and advantages over current imaging technology are illustrated. ScFvs are produced in a cloned form and can be readily engineered to have localizing and therapeutic functions that will be applicable in cancer and other diseases.

Helping the general physician to improve outcomes after PEG insertion: how we changed our practice.

During their careers, most general physicians are involved in the decision-making process for patients that potentially require percutaneous endoscopic gastrostomy (PEG) insertion. However, poor patient selection and less than favourable outcomes are frequently observed in this group. With the aim of identifying and addressing the underlying issues, the PEG service at University Hospital Llandough was radically changed over an eight-year period. The development of a nurse-led pre-assessment service and design of a specific referral form was successful in reducing the number of PEG referrals and consequently the 30-day mortality rate. Furthermore, the educational and training needs of general physicians of all grades regarding the issues surrounding PEG placement were identified and addressed at formal teaching sessions. A combination of these factors has positively impacted on our service, with more appropriate patient selection and a reduced 30-day mortality rate.

Invertebrate dispersal by waterbird species in neotropical wetlands.

Endozoochory by waterbirds is particularly relevant to the dispersal of non-flying aquatic invertebrates. This ecological function exercised by birds has been demonstrated in different biogeographical regions, but there are no studies for the neotropical region. In this work, we identified propagules of invertebrates in faeces of 14 syntopic South American waterbird species representing six families, and hatched additional invertebrates from cultured faeces. We tested whether propagule abundance, species richness and composition varied among bird species, and between the cold and warm seasons. We found 164 invertebrate propagules in faecal samples from seven different waterbirds species, including eggs of the Temnocephalida and Notonectidae, statoblasts of bryozoans (Plumatella sp.) and ephippia of Cladocera. Ciliates (including Paramecium sp. and Litostomatea), nematodes and rotifers (Adineta sp. and Nottomatidae) hatched from cultured samples. Potential for endozoochory was confirmed for 12 of 14 waterbird species. Our statistical models suggest that richness and abundance of propagules are associated with bird species and not affected by seasonality. Dispersal by endozoochory is potentially important to a broad variety of invertebrates, being promoted by waterbirds with different ecological and morphological traits, which are likely to drive the dispersal of invertebrates in neotropical wetlands.

Urate-induced epigenetic modifications in myeloid cells.

OBJECTIVES: Hyperuricemia is a metabolic condition central to gout pathogenesis. Urate exposure primes human monocytes towards a higher capacity to produce and release IL-1ß. In this study, we assessed the epigenetic processes associated to urate-mediated hyper-responsiveness. METHODS: Freshly isolated human peripheral blood mononuclear cells or enriched monocytes were pre-treated with solubilized urate and stimulated with LPS with or without monosodium urate (MSU) crystals. Cytokine production was determined by ELISA. Histone epigenetic marks were assessed by sequencing immunoprecipitated chromatin. Mice were injected intraarticularly with MSU crystals and palmitate after inhibition of uricase and urate administration in the presence or absence of methylthioadenosine. DNA methylation was assessed by methylation array in whole blood of 76 participants with normouricemia or hyperuricemia. RESULTS: High concentrations of urate enhanced the inflammatory response in vitro in human cells and in vivo in mice, and broad-spectrum methylation inhibitors reversed this effect. Assessment of histone 3 lysine 4 trimethylation (H3K4me3) and histone 3 lysine 27 acetylation (H3K27ac) revealed differences in urate-primed monocytes compared to controls. Differentially methylated regions (e.g. HLA-G, IFITM3, PRKAB2) were found in people with hyperuricemia compared to normouricemia in genes relevant for inflammatory cytokine signaling. CONCLUSION: Urate alters the epigenetic landscape in selected human monocytes or whole blood of people with hyperuricemia compared to normouricemia. Both histone modifications and DNA methylation show differences depending on urate exposure. Subject to replication and validation, epigenetic changes in myeloid cells may be a therapeutic target in gout.

Evidence of interaction of CARD8 rs2043211 with NALP3 rs35829419 in Crohn's disease.

The location of CARD8 within an inflammatory bowel disease (IBD) locus and its role in the NALP3 inflammasome and as a nuclear factor (NF)kappaB inhibitor make it an attractive candidate risk gene for IBD. However, studies testing for the association of the CARD8 loss-of-function single-nucleotide polymorphism (SNP) rs2043211 with IBD have yielded mixed results. A recent study provided evidence that this discordance may result from an interaction of rs2043211 with loss-of-function variants in nucleotide-binding oligomerization domain protein 2 (NOD2) and a gain-of-function SNP (rs35829419) in NALP3. To confirm this interaction, we conducted a replication in an independent IBD sample set (n=1009 patients, n=517 controls). We found that the presence of the minor allele of rs2043211 with the major allele of rs35829419 conferred a protective effect against Crohn's disease (and vice versa), which intensified in the absence of NOD2 mutations (P(1,2/1,1)=0.009, odds ratio (OR)=0.66, 95% confidence interval (CI) (0.48-0.90); P(1,1/1,2)=0.015, OR=0.35, 95% CI (0.15-0.82)). We propose that these genotype combinations protect against gut inflammation by preventing the NALP3 inflammasome from producing excessive interleukin-1beta.