Birmingham Environment for Academic Research : Case studies volume 3

This collection of case studies was brought together to showcase the extent and diversity of research that is supported by the University of Birmingham’s Environment for Academic Research (BEAR). BEAR is a collection of contemporary IT resources designed to help research. The following case studies demonstrate how BEAR services such as the Research Data Store (RDS), BEAR software and the University supercomputer BlueBEAR are integral to the progression of important research across disciplines. BlueBEAR is a key component of BEAR, providing compute power and specialist applications free to enable staff and students to delve deeper into their research. Upgraded in 2023, the cluster includes many large memory nodes and a GPU service alongside standard compute nodes. Alongside BlueBEAR, the RDS is a popular choice amongst researchers to securely store their working research data. As of publication, more than 5000 researchers across all five colleges were actively using BlueBEAR and/or the RDS. In this volume, we showcase case studies representing diverse research from every college. From estimating snow coverage to modelling second language acquisition, we show how BEAR services are enabling exciting and important research across the university

Robust estimation of bacterial cell count from optical density

Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data

Report on the 15th international symposium on geo-disaster reduction, 25–30 august 2017, Oki Islands - Matsue - Kyoto, Japan

Abstract On 25–30 August 2017, the 15th International Symposium on Geo-disaster Reduction has been held in Oki Islands, Matsue and Kyoto, Japan, focusing on the theme of “Global Strategy for Geo-disaster Reduction”. Through Field Excursion, High Level Forum, Keynote Lectures, Invited Lectures, Scientific Session, Youth Forum and Poster Session, this symposium has successfully provided a platform for scientists, industrial professionals and students to share their research and created a forum to exchange ideas for promoting geo-disaster reduction

The DNA Methylome and Association of Differentially Methylated Regions with Differential Gene Expression during Heat Stress in Brassica rapa

Cytosine DNA methylation is a critical epigenetic mechanism in the silencing of transposable elements, imprinting and regulating gene expression. However, little is known about the potential role of mC in response to heat stress. To determine and explore the functions of the dynamic DNA methylome during heat stress, we characterized single-base resolution methylome maps of Brassica rapa and assessed the dynamic changes of mC under heat stress using whole genome bisulfite sequencing. On average, the DNA methylation levels of CG, CHG and CHH are 39.3%, 15.38% and 5.24% in non-heading Chinese cabbage (NHCC), respectively. We found that the patterns of methylation are similar to other eudicot plants, but with higher CHH methylation levels. Further comparative analysis revealed varying patterns for three sequence contexts (mCG, mCHG and mCHH) under heat stress indicating context- and position-dependent methylation regulation. DNA methylation near the TSS and TES may be closely associated with methylation-dependent transcriptional silencing. Association analysis of differential methylation and differential gene expression revealed a different set of methDEGs involved at early and late stages under heat stress. The systemic characterization of the dynamic DNA methylome during heat stress will improve our understanding of the mechanism of epigenetic regulation under heat stress

Molecular cloning, characterization and RNA interference assay of two toll-like receptors in giant freshwater prawn, Macrobrachium rosenbergii

Toll-like receptors are a family of conserved membrane receptors which are involved in the regulation of a series of innate immune responses in invertebrates and vertebrates. Two different types of Toll-like receptors from giant freshwater prawn (Macrobrachium rosenbergii), which were designated as MrToll1 and MrToll2, were cloned in the present study. The full-length cDNA of MrToll1 is 4234 bp in length, which includes a 3015-bp open reading frame (ORF) encoding a deduced protein of 1004 amino acids. The sequence of MrToll2 cDNA is 4383 bp in length containing a 2739-bp ORF which encodes 912 amino acid residues. The results from the multiple alignments of amino acid sequences and phylogenetic analysis indicated the two MrTolls identified in the present study should be classified as crustacean type-1 and type-2 Toll-like receptor, respectively. MrToll1 and MrToll2 were constitutively expressed in all the examined tissues, including gill, stomach, intestine, eyestalk, muscle, heart, ganglion, brain, hemocytes and hepatopancreas. RNA interference assay demonstrated that knockdown of MrToll1 with specific-siRNA significantly suppressed the expression of myeloid differentiation factor 88 (MyD88), anti-lipopolysaccharide factor (ALF), crustin and prophenoloxidase (proPO) and enhanced the expression level of MrToll2 in M. rosenbergii. Additionally, an obvious down-regulation of MyD88 and crustin expression was observed in MrToll2-silenced prawns. These findings suggest that MrToll1 and MrToll2 might play essential roles in the innate immunity of M. rosenbergii through regulating the expression of immune-related genes

Identification and characterization of TOR in Macrobrachium rosenbergii and its role in muscle protein and lipid production

Abstract The recent scarcity of fishmeal and other resources means that studies on the intrinsic mechanisms of nutrients in the growth and development of aquatic animals at the molecular level have received widespread attention. The target of rapamycin (TOR) pathway has been reported to receive signals from nutrients and environmental stresses, and regulates cellular anabolism and catabolism to achieve precise regulation of cell growth and physiological activities. In this study, we cloned and characterized the full-length cDNA sequence of the TOR gene of Macrobrachium rosenbergii (MrTOR). MrTOR was expressed in all tissues, with higher expression in heart and muscle tissues. In situ hybridization also indicated that MrTOR was expressed in muscle, mainly around the nucleus. RNA interference decreased the expression levels of MrTOR and downstream protein synthesis-related genes (S6K, eIF4E, and eIF4B) (P < 0.05) and the expression and enzyme activity of the lipid synthesis-related enzyme, fatty acid synthase (FAS), and increased enzyme activity of the lipolysis-related enzyme, lipase (LPS). In addition, amino acid injection significantly increased the transcript levels of MrTOR and downstream related genes (S6K, eIF4E, eIF4B, and FAS), as well as triglyceride and total cholesterol tissue levels and FAS activity. Starvation significantly increased transcript levels and enzyme activities of adenylate-activated protein kinase and LPS and decreased transcript levels and enzyme activities of FAS, as well as transcript levels of MrTOR and its downstream genes (P < 0.05), whereas amino acid injection alleviated the starvation-induced decreases in transcript levels of these genes. These results suggested that arginine and leucine activated the TOR signaling pathway, promoted protein and lipid syntheses, and alleviated the pathway changes induced by starvation

Enhancement of β-Lactam-Mediated Killing of Gram-Negative Bacteria by Lysine Hydrochloride

ABSTRACT Widespread bacterial resistance among Gram-negative bacteria is rapidly depleting our antimicrobial arsenal. Adjuvants that enhance the bactericidal activity of existing antibiotics provide a way to alleviate the resistance crisis, as new antimicrobials are becoming increasingly difficult to develop. The present work with Escherichia coli revealed that neutralized lysine (lysine hydrochloride) enhances the bactericidal activity of β-lactams in addition to increasing bacteriostatic activity. When combined, lysine hydrochloride and β-lactam increased expression of genes involved in the tricarboxylic acid (TCA) cycle and raised reactive oxygen species (ROS) levels; as expected, agents known to mitigate bactericidal effects of ROS reduced lethality from the combination treatment. Lysine hydrochloride had no enhancing effect on the lethal action of fluoroquinolones or aminoglycosides. Characterization of a tolerant mutant indicated involvement of the FtsH/HflkC membrane-embedded protease complex in lethality enhancement. The tolerant mutant, which carried a V86F substitution in FtsH, exhibited decreased lipopolysaccharide levels, reduced expression of TCA cycle genes, and reduced levels of ROS. Lethality enhancement by lysine hydrochloride was abolished by treating cultures with Ca2+ or Mg2+, cations known to stabilize the outer membrane. These data, plus damage observed by scanning electron microscopy, indicate that lysine stimulates β-lactam lethality by disrupting the outer membrane. Lethality enhancement of β-lactams by lysine hydrochloride was also observed with Acinetobacter baumannii and Pseudomonas aeruginosa, thereby suggesting that the phenomenon is common among Gram-negative bacteria. Arginine hydrochloride behaved in a similar way. Overall, the combination of lysine or arginine hydrochloride and β-lactam offers a new way to increase β-lactam lethality with Gram-negative pathogens. IMPORTANCE Antibiotic resistance among Gram-negative pathogens is a serious medical problem. The present work describes a new study in which a nontoxic nutrient increases the lethal action of clinically important β-lactams. Elevated lethality is expected to reduce the emergence of resistant mutants. The effects were observed with significant pathogens (Escherichia coli, Acinetobacter baumannii, and Pseudomonas aeruginosa), indicating widespread applicability. Examination of tolerant mutants and biochemical measurements revealed involvement of endogenous reactive oxygen species in response to outer membrane perturbation. These lysine hydrochloride–β-lactam data support the hypothesis that lethal stressors can stimulate the accumulation of ROS. Genetic and biochemical work also revealed how an alteration in a membrane protease, FtsH, abolishes lysine stimulation of β-lactam lethality. Overall, the work presents a method for antimicrobial enhancement that should be safe, easy to administer, and likely to apply to other nutrients, such as arginine