Optimizing DDRADseq in non-model species: A Case Study in Eucalyptus dunnii Maiden

Restriction site-associated DNA sequencing (RADseq) and its derived protocols, such as double digest RADseq (ddRADseq), offer a flexible and highly cost-effective strategy for efficient plant genome sampling. This has become one of the most popular genotyping approaches for breeding, conservation, and evolution studies in model and non-model plant species. However, universal protocols do not always adapt well to non-model species. Herein, this study reports the development of an optimized and detailed ddRADseq protocol in Eucalyptus dunnii, a non-model species, which combines different aspects of published methodologies. The initial protocol was established using only two samples by selecting the best combination of enzymes and through optimal size selection and simplifying lab procedures. Both single nucleotide polymorphisms (SNPs) and simple sequence repeats (SSRs) were determined with high accuracy after applying stringent bioinformatics settings and quality filters, with and without a reference genome. To scale it up to 24 samples, we added barcoded adapters. We also applied automatic size selection, and therefore obtained an optimal number of loci, the expected SNP locus density, and genome-wide distribution. Reliability and cross-sequencing platform compatibility were verified through dissimilarity coefficients of 0.05 between replicates. To our knowledge, this optimized ddRADseq protocol will allow users to go from the DNA sample to genotyping data in a highly accessible and reproducible way.Instituto de BiotecnologíaFil: Aguirre, Natalia Cristina. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Filippi, Carla Valeria. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Zaina, Giusi. University of Udine. Department of Agricultural, Food, Environmental and Animal Sciences; ItaliaFil: Rivas, Juan Gabriel. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Acuña, Cintia Vanesa. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Villalba, Pamela Victoria. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Garcia, Martin Nahuel. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Gonzalez, Sergio Alberto. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Rivarola, Maximo Lisandro. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Martinez, Maria Carolina. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Puebla, Andrea Fabiana. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Morgante, Michele. University of Udine. Department of Agricultural, Food, Environmental and Animal Sciences; ItaliaFil: Hopp, Horacio Esteban. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Paniego, Norma Beatriz. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Marcucci Poltri, Susana Noemi. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Intravascular Large B-Cell Lymphoma Genomic Profile Is Characterized by Alterations in Genes Regulating NF-κB and Immune Checkpoints.

Intravascular large B-cell lymphoma (IVLBCL) is an uncommon lymphoma with an aggressive clinical course characterized by selective growth of tumor cells within the vessels. Its pathogenesis is still uncertain and there is little information on the underlying genomic alterations. In this study, we performed a clinicopathologic and next-generation sequencing analysis of 15 cases of IVLBCL using a custom panel for the detection of alterations in 68 recurrently mutated genes in B-cell lymphomagenesis. Six patients had evidence of hemophagocytic syndrome. Four patients presented concomitantly a solid malignancy. Tumor cells outside the vessels were observed in 7 cases, 2 with an overt diffuse large B-cell cell lymphoma. In 4 samples, tumor cells infiltrated lymphatic vessel in addition to blood capillaries. Programmed death-ligand 1 (PD-L1) was positive in tumor cells in 4 of 11 evaluable samples and in macrophages intermingled with tumor cells in 8. PD-L1 copy number gains were identified in a higher proportion of cases expressing PD-L1 than in negative tumors. The most frequently mutated gene was PIM1 (9/15, 60%), followed by MYD88L265P and CD79B (8/15, 53% each). In 6 cases, MYD88L265P and CD79B mutations were detected concomitantly. We also identified recurrent mutations in IRF4 , TMEM30A , BTG2 , and ETV6 loci (4/15, 27% each) and novel driver mutations in NOTCH2 , CCND3 , and GNA13 , and an IRF4 translocation in 1 case each. The mutational profile was similar in patients with and without evidence of hemophagocytic syndrome and in cases with or without dissemination of tumor cells outside the vessels. Our results confirm the relevance of mutations in B-cell receptor/nuclear factor-κB signaling and immune escape pathways in IVLBCL and identify novel driver alterations. The similar mutational profile in tumors with extravascular dissemination suggests that these cases may also be considered in the spectrum of IVLBCL

Unlocking the Mysteries of Diastolic Function Deciphering the Rosetta Stone 10 Years Later

It has now been a quarter of a century since the first description by Kitabatake and his associates of the use of echo-Doppler to characterize the transmitral flow velocity curves in various disease states. A decade ago we described the role of echocardiography in the “Evaluation of Diastolic Filling of Left Ventricle in Health and Disease: Doppler Echocardiography Is the Clinician’s Rosetta Stone.” Over the ensuing decade, advances in echo-Doppler have helped to further decipher the morphologic and physiological expression of cardiovascular disease and unlock additional mysteries of diastology. The purpose of this review is to highlight the developments in echo-Doppler and refinements in our knowledge that have occurred over the past decade that enhance our understanding of diastology

Removal of Hepatitis B virus surface HBsAg and core HBcAg antigens using microbial fuel cells producing electricity from human urine

© 2019, The Author(s). Microbial electrochemical technology is emerging as an alternative way of treating waste and converting this directly to electricity. Intensive research on these systems is ongoing but it currently lacks the evaluation of possible environmental transmission of enteric viruses originating from the waste stream. In this study, for the first time we investigated this aspect by assessing the removal efficiency of hepatitis B core and surface antigens in cascades of continuous flow microbial fuel cells. The log-reduction (LR) of surface antigen (HBsAg) reached a maximum value of 1.86 ± 0.20 (98.6% reduction), which was similar to the open circuit control and degraded regardless of the recorded current. Core antigen (HBcAg) was much more resistant to treatment and the maximal LR was equal to 0.229 ± 0.028 (41.0% reduction). The highest LR rate observed for HBsAg was 4.66 ± 0.19 h−1 and for HBcAg 0.10 ± 0.01 h−1. Regression analysis revealed correlation between hydraulic retention time, power and redox potential on inactivation efficiency, also indicating electroactive behaviour of biofilm in open circuit control through the snorkel-effect. The results indicate that microbial electrochemical technologies may be successfully applied to reduce the risk of environmental transmission of hepatitis B virus but also open up the possibility of testing other viruses for wider implementation

Aggregating behaviour in invasive Caribbean lionfish is driven by habitat complexity

Caribbean lionfish (Pterois spp.) are considered the most heavily impacting invasive marine vertebrate ever recorded. However, current management is largely inadequate, relying on opportunistic culling by recreational SCUBA divers. Culling efficiency could be greatly improved by exploiting natural aggregations, but to date this behaviour has only been recorded anecdotally, and the drivers are unknown. We found aggregations to be common in situ, but detected no conspecific attraction through visual or olfactory cues in laboratory experiments. Aggregating individuals were on average larger, but showed no further differences in morphology or life history. However, using visual assessments and 3D modelling we show lionfish prefer broad-scale, but avoid fine-scale, habitat complexity. We therefore suggest that lionfish aggregations are coincidental based on individuals’ mutual attraction to similar reef structure to maximise hunting efficiency. Using this knowledge, artificial aggregation devices might be developed to concentrate lionfish densities and thus improve culling efficiency

Assessing road effects on bats: the role of landscape, road features, and bat activity on road-kills

Recent studies suggest that roads can significantly impact bat populations. Though bats are one of the most threatened groups of European vertebrates, studies aiming to quantify bat mortality and determine the main factors driving it remain scarce. Between March 16 and October 31 of 2009, we surveyed road-killed bats daily along a 51-km-long transect that incorporates different types of roads in southern Portugal. We found 154 road-killed bats of 11 species. The two most common species in the study area, Pipistrellus kuhlii and P. pygmaeus, were also the most commonly identified road-kill, representing 72 % of the total specimens collected. About two-thirds of the total mortality occurred between mid July and late September, peaking in the second half of August. We also recorded casualties of threatened and rare species, including Miniopterus schreibersii, Rhinolophus ferrumequinum, R. hipposideros, Barbastella barbastellus, and Nyctalus leisleri. These species were found mostly in early autumn, corresponding to the mating and swarming periods. Landscape features were the most important variable subset for explaining bat casualties. Road stretches crossing or in the vicinity of high-quality habitats for bats—including dense Mediterranean woodland (‘‘montado’’) areas, water courses with riparian gallery, and water reservoirs—yielded a significantly higher number of casualties. Additionally, more roadkilled bats were recorded on high-traffic road stretches with viaducts, in areas of higher bat activity and near known roosts

Acidithiobacillus ferrooxidans metabolism: from genome sequence to industrial applications

<p>Abstract</p> <p>Background</p> <p><it>Acidithiobacillus ferrooxidans </it>is a major participant in consortia of microorganisms used for the industrial recovery of copper (bioleaching or biomining). It is a chemolithoautrophic, γ-proteobacterium using energy from the oxidation of iron- and sulfur-containing minerals for growth. It thrives at extremely low pH (pH 1–2) and fixes both carbon and nitrogen from the atmosphere. It solubilizes copper and other metals from rocks and plays an important role in nutrient and metal biogeochemical cycling in acid environments. The lack of a well-developed system for genetic manipulation has prevented thorough exploration of its physiology. Also, confusion has been caused by prior metabolic models constructed based upon the examination of multiple, and sometimes distantly related, strains of the microorganism.</p> <p>Results</p> <p>The genome of the type strain <it>A. ferrooxidans </it>ATCC 23270 was sequenced and annotated to identify general features and provide a framework for <it>in silico </it>metabolic reconstruction. Earlier models of iron and sulfur oxidation, biofilm formation, quorum sensing, inorganic ion uptake, and amino acid metabolism are confirmed and extended. Initial models are presented for central carbon metabolism, anaerobic metabolism (including sulfur reduction, hydrogen metabolism and nitrogen fixation), stress responses, DNA repair, and metal and toxic compound fluxes.</p> <p>Conclusion</p> <p>Bioinformatics analysis provides a valuable platform for gene discovery and functional prediction that helps explain the activity of <it>A. ferrooxidans </it>in industrial bioleaching and its role as a primary producer in acidic environments. An analysis of the genome of the type strain provides a coherent view of its gene content and metabolic potential.</p

Co-limitation towards lower latitudes shapes global forest diversity gradients

The latitudinal diversity gradient (LDG) is one of the most recognized global patterns of species richness exhibited across a wide range of taxa. Numerous hypotheses have been proposed in the past two centuries to explain LDG, but rigorous tests of the drivers of LDGs have been limited by a lack of high-quality global species richness data. Here we produce a high-resolution (0.025° × 0.025°) map of local tree species richness using a global forest inventory database with individual tree information and local biophysical characteristics from ~1.3 million sample plots. We then quantify drivers of local tree species richness patterns across latitudes. Generally, annual mean temperature was a dominant predictor of tree species richness, which is most consistent with the metabolic theory of biodiversity (MTB). However, MTB underestimated LDG in the tropics, where high species richness was also moderated by topographic, soil and anthropogenic factors operating at local scales. Given that local landscape variables operate synergistically with bioclimatic factors in shaping the global LDG pattern, we suggest that MTB be extended to account for co-limitation by subordinate drivers

The positioning system of the ANTARES Neutrino Telescope

The ANTARES neutrino telescope, located 40km off the coast of Toulon in the Mediterranean Sea at a mooring depth of about 2475m, consists of twelve detection lines equipped typically with 25 storeys. Every storey carries three optical modules that detect Cherenkov light induced by charged secondary particles (typically muons) coming from neutrino interactions. As these lines are flexible structures fixed to the sea bed and held taut by a buoy, sea currents cause the lines to move and the storeys to rotate. The knowledge of the position of the optical modules with a precision better than 10cm is essential for a good reconstruction of particle tracks. In this paper the ANTARES positioning system is described. It consists of an acoustic positioning system, for distance triangulation, and a compass-tiltmeter system, for the measurement of the orientation and inclination of the storeys. Necessary corrections are discussed and the results of the detector alignment procedure are described