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The incorporation of [methyl-3H]thymidine into three macromolecular fractions, designated as DNA, RNA, and protein, by bacteria from Hartbeespoort Dam, South Africa, was measured over 1 year by acid-base hydrolysis procedures. Samples were collected at 10 m, which was at least 5 m beneath the euphotic zone. On four occasions, samples were concurrently collected at the surface. Approximately 80% of the label was incorporated into bacterial DNA in surface samples. At 10 m, total incorporation of label into bacterial macromolecules was correlated to bacterial utilization of glucose (r = 0.913, n = 13, P < 0.001). The labeling of DNA, which ranged between 0 and 78% of total macromolecule incorporation, was inversely related to glucose uptake (r = -0.823), total thymidine incorporation (r = -0.737), and euphotic zone algal production (r = -0.732, n = 13, P < 0.005). With decreased DNA labeling, increasing proportions of label were found in the RNA fraction and proteins. Enzymatic digestion followed by chromatographic separation of macromolecule fragments indicated that DNA and proteins were labeled while RNA was not. The RNA fraction may represent labeled lipids or other macromolecules or both. The data demonstrated a close coupling between phytoplankton production and heterotrophic bacterial activity in this hypertrophic lake but also confirmed the need for the routine extraction and purification of DNA during [methyl-3H]thymidine studies of aquatic bacterial production.
Spatial and Temporal Variations in Bacterial Macromolecule Labeling with [methyl-3H]Thymidine in a Hypertrophic Lake †
Richard D. Robarts,1,* Richard J. Wicks,2 and Lynne M. Sephton1
In inherited primary arrhythmia syndromes (PAS) and cardiomyopathies (CMP), the yield of genetic testing varies between 20 and 75% in different diseases according to studies performed in the pre next-generation sequencing (NGS) era. It is unknown whether retesting historical negative samples with NGS techniques is worthwhile. Therefore, we assessed the value of NGS-based panel testing in previously genotype negative-phenotype positive probands. We selected 107 patients (47 PAS and 60 CMP) with a clear phenotype who remained genotype negative after genetic analysis of the main genes implicated in their specific phenotype. Targeted sequencing of the coding regions of 71 PAS- and CMP-related genes was performed. Variant interpretation and classification was done according to a cardiology-specific scoring algorithm (‘Amsterdam criteria’) and the ACMG-AMP criteria. Co-segregation analysis was performed when DNA and clinical data of family members were available. Finally, a genetic diagnosis could be established in 21 patients (20%), 5 PAS (11%) and 16 CMP (27%) patients, respectively. The increased detection rate was due to sequencing of novel genes in 52% of the cases and due to technical failures with the historical analysis in 48%. A total of 118 individuals were informed about their carrier state and either reassured or scheduled for proper follow-up. To conclude, genetic retesting in clinically overt PAS and CMP cases, who were genotype negative with older techniques, resulted in an additional genetic diagnosis in up to 20% of the cases. This clearly supports a policy for genetic retesting with NGS-based panels.
Repeat genetic testing with targeted capture sequencing in primary arrhythmia syndrome and cardiomyopathy
Tomas Robyns,corresponding author1 Cuno Kuiperi,2 Jeroen Breckpot,2 Koenraad Devriendt,2 Erika Souche,2 Johan Van Cleemput,1 Rik Willems,1 Dieter Nuyens,3 Gert Matthijs,2 and Anniek Corveleyn2
The Loeys-Dietz syndrome (LDS) is a connective tissue disorder affecting the cardiovascular, skeletal, and ocular system. Most typically, LDS patients present with aortic aneurysms and arterial tortuosity, hypertelorism, and bifid/broad uvula or cleft palate. Initially, mutations in transforming growth factor?β (TGF?β) receptors (TGFBR1 and TGFBR2) were described to cause LDS, hereby leading to impaired TGF?β signaling. More recently, TGF?β ligands, TGFB2 and TGFB3, as well as intracellular downstream effectors of the TGF?β pathway, SMAD2 and SMAD3, were shown to be involved in LDS. This emphasizes the role of disturbed TGF?β signaling in LDS pathogenesis. Since most literature so far has focused on TGFBR1/2, we provide a comprehensive review on the known and some novel TGFB2/3 and SMAD2/3 mutations. For TGFB2 and SMAD3, the clinical manifestations, both of the patients previously described in the literature and our newly reported patients, are summarized in detail. This clearly indicates that LDS concerns a disorder with a broad phenotypical spectrum that is still emerging as more patients will be identified. All mutations described here are present in the corresponding Leiden Open Variant Database.
aneurysm, Loeys-Dietz syndrome, SMAD2, SMAD3, TGFB2, TGFB3
A mutation update on the LDS?associated genes TGFB2/3 and SMAD2/3
Dorien Schepers, 1 , † Giada Tortora, 2 , 3 , † Hiroko Morisaki, 4 , 5 , 6 Gretchen MacCarrick, 7 Mark Lindsay, 8 David Liang, 9 Sarju G. Mehta, 10 Jennifer Hague, 10 Judith Verhagen, 11 Ingrid van de Laar, 11 Marja Wessels, 11 Yvonne Detisch, 12 Mieke van Haelst, 13 , 14 Annette Baas, 13 Klaske Lichtenbelt, 13 Kees Braun, 15 Denise van der Linde, 16 Jolien Roos?Hesselink, 16 George McGillivray, 17 Josephina Meester, 1 Isabelle Maystadt, 18 Paul Coucke, 19 Elie El?Khoury, 20 Sandhya Parkash, 21 Birgitte Diness, 22 Lotte Risom, 22 Ingrid Scurr, 23 Yvonne Hilhorst?Hofstee, 24 Takayuki Morisaki, 4 , 5 Julie Richer, 25 Julie Desir, 26 Marlies Kempers, 27 Andrea L. Rideout, 28 Gabrielle Horne, 29 Chris Bennett, 30 Elisa Rahikkala, 31 Geert Vandeweyer, 1 Maaike Alaerts, 1 Aline Verstraeten, 1 Hal Dietz, 7 Lut Van Laer, 1 and Bart Loeyscorresponding author 1 , 27