Identification and characterization of a spore-like morphotype in chronically starved Mycobacterium avium subsp. paratuberculosis cultures

Mycobacteria are able to enter into a state of non-replication or dormancy, which may result in their chronic persistence in soil, aquatic environments, and permissive hosts. Stresses such as nutrient deprivation and hypoxia provide environmental cues to enter a persistent state; however, a clear definition of the mechanism that mycobacteria employ to achieve this remains elusive. While the concept of sporulation in mycobacteria is not novel, it continues to spark controversy and challenges our perceptions of a non-replication. We investigated the potential role of sporulation in one-year old broth cultures of Mycobacterium subsp. paratuberculosis (MAP). We show that dormant cultures of MAP contain a mix of vegetative cells and a previously unknown morphotype resembling a spore. These spore-like structures can be enriched for using sporulating media. Furthermore, purified MAP spore forms survive exposure to heat, lysozyme and proteinase K. Heat-treated spores are positive for MAP 16SrRNA and IS900. MAP spores display enhanced infectivity as well as maintain acid-fast characteristics upon germination in a well-established bovine macrophage model. This is the first study to demonstrate a new MAP morphotype possessing spore-like qualities. Data suggest that sporulation may be a viable mechanism by which MAP accomplishes persistence in the host and/or environment. Thus, our current understanding of mycobacterial persistence, pathogenesis, epidemiology and rational drug and vaccine design may need to be reevaluated.     

Chromatographic retention time prediction for posttranslationally modified peptides

Retention time prediction of peptides in liquid chromatography has proven to be a valuable tool for mass spectrometry-based proteomics, especially in designing more efficient procedures for state-of-the-art targeted workflows. Additionally, accurate retention time predictions can also be used to increase confidence in identifications in shotgun experiments. Despite these obvious benefits, the use of such methods has so far not been extended to (posttranslationally) modified peptides due to the absence of efficient predictors for such peptides. We here therefore describe a new retention time predictor for modified peptides, built on the foundations of our existing Elude algorithm. We evaluated our software by applying it on five types of commonly encountered modifications. Our results show that Elude now yields equally good prediction performances for modified and unmodified peptides, with correlation coefficients between predicted and observed retention times ranging from 0.93 to 0.98 for all the investigated datasets. Furthermore, we show that our predictor handles peptides carrying multiple modifications as well. This latest version of Elude is fully portable to new chromatographic conditions and can readily be applied to other types of posttranslational modifications. Elude is available under the permissive Apache2 open source License at http://per-colator.com or can be run via a web-interface at http://elude.sbc.su.se.     

Nanoflake‐Assembled Hierarchical Na3V2(PO4)3/C Microflowers: Superior Li Storage Performance and Insertion/Extraction Mechanism

Na₃V₂(PO₄)₃ (NVP) has excellent electrochemical stability and fast ion diffusion coefficient due to the 3D Na⁺ ion superionic conductor framework, which make it an attractive cathode material for lithium ion batteries (LIBs). However, the electrochemical performance of NVP needs to be further improved for applications in electric vehicles and hybrid electric vehicles. Here, nanoflake‐assembled hierarchical NVP/C microflowers are synthesized using a facile method. The structure of as‐synthesized materials enhances the electrochemical performance by improving the electron conductivity, increasing electrode–electrolyte contact area, and shortening the diffusion distance. The as‐synthesized material exhibits a high capacity (230 mAh g^?1), excellent cycling stability (83.6% of the initial capacity is retained after 5000 cycles), and remarkable rate performance (91 C) in hybrid LIBs. Meanwhile, the hybrid LIBs with the structure of NVP || 1 m LiPF₆/EC (ethylene carbonate) + DMC (dimethyl carbonate) || NVP and Li₄Ti₅O₁₂ || 1 m LiPF₆/EC + DMC || NVP are assembled and display capacities of 79 and 73 mAh g^?1, respectively. The insertion/extraction mechanism of NVP is systematically investigated, based on in situ X‐ray diffraction. The superior electrochemical performance, the design of hybrid LIBs, and the insertion/extraction mechanism investigation will have profound implications for developing safe and stable, high‐energy, and high‐power LIBs.     

Sucrose nonfermenting AMPK-related kinase (SNARK) regulates exercise-stimulated and ischemia-stimulated glucose transport in the heart

The signaling mechanisms mediating myocardial glucose transport are not fully understood. Sucrose nonfermenting AMP-activated protein kinase (AMPK)-related kinase (SNARK) is an AMPK-related protein kinase that is expressed in the heart and has been implicated in contraction-stimulated glucose transport in mouse skeletal muscle. We first determined if SNARK is phosphorylated on Thr²⁰⁸, a site critical for SNARK activity. Mice were treated with exercise, ischemia, submaximal insulin, or maximal insulin. Treadmill exercise slightly, but significantly increased SNARK Thr²⁰⁸ phosphorylation. Ischemia also increased SNARK Thr²⁰⁸ phosphorylation, but there was no effect of submaximal or maximal insulin. HL1 cardiomyocytes were used to overexpress wild-type (WT) SNARK and to knockdown endogenous SNARK. Overexpression of WT SNARK had no effect on ischemia-stimulated glucose transport; however, SNARK knockdown significantly decreased ischemia-stimulated glucose transport. SNARK overexpression or knockdown did not alter insulin-stimulated glucose transport or glycogen concentrations. To study SNARK function in vivo, SNARK heterozygous knockout mice (SNARK^+/−) and WT littermates performed treadmill exercise. Exercise-stimulated glucose transport was decreased by ~50% in hearts from SNARK^+/− mice. In summary, exercise and ischemia increase SNARK Thr²⁰⁸ phosphorylation in the heart and SNARK regulates exercise-stimulated and ischemia-stimulated glucose transport. SNARK is a novel mediator of insulin-independent glucose transport in the heart.     

Integrated regulation of the photosynthetic gene family from Arabidopsis thaliana in transformed tobacco cells

Summary Expression of the three chlorophyll a/b binding protein (cab) genes of Arabidopsis thaliana was studied in transformed tobacco tissues. For each cab gene, approximately 1000 bp of the promoter region plus a portion of the structural gene was inserted into a promoter-expression vector such that a translational fusion between the cab gene and the promoter-less chloramphenicol acetyltransferase (cat) gene was formed. The constructed molecules were introduced into either cultured tobacco cells or tobacco leaves and the promoter activity was monitored as chloramphenicol acetyltransferase activity. The light-grown tissues exhibited 1.5- to 60-fold greater promoter activity than did dark-grown tissues. Expression of the cab promoters was tissue specific: activities were much stronger in green leaves than other tissues. The cab promoters were almost equally active in transformed calli or shoots derived from leaves. However, in cultured tobacco cells, one promoter was two to three times stronger than the other two. The chimeric gene fusion, cab-cat, segregated in the F1 generation as a dominant Mendelian trait.     

玉米根冠脱落细胞中微丝分布的荧光显微观察(简报)

在对玉米等主要农作物进行病理及致病毒素毒理分析时,经常选用根冠脱落细胞作为试材。许多学眷认为这些脱落的细胞处于垂死状态,然而,后来的一些研究肯定了Knudson的脱落根冠细胞具有活力的观点。Copoorali无菌培养玉米根冠脱落细胞,发现它们可以分裂;Vermeer等提出存活于玉米根际周围的根冠脱落细胞是根系统范围的扩展,它们在土壤中的作用应予以特别注意;Hawes不仅认为它们为活细胞,而且以它们做实验材料用于毒理分析;Guinal对玉米根冠脱落细胞的来源、细胞结构及生理特性进行了详细研究,肯定了细胞     

An Improved Method for Rapid Intubation of the Trachea in Mice

Despite some anatomical and physiological differences, mouse models continue to be an essential tool for studying human lung disease. Bleomycin toxicity is a commonly used model to study both acute lung injury and fibrosis, and multiple methods have been developed for administering bleomycin (and other toxic agents) into the lungs. However, many of these approaches, such as transtracheal instillation, have inherent drawbacks, including the need for strong anesthetics and survival surgery. This paper reports a quick, reproducible method of intratracheal intubation that involves mild inhaled anesthesia, visualization of the trachea, and the use of a surrogate spirometer to confirm exposure. As a proof of concept, 8-12 week old C57BL/6 mice were administered either 2.0 U/kg of bleomycin or an equivalent volume of PBS, and both damage and fibrotic endpoints were measured post-exposure. This procedure allows researchers to treat a large cohort of mice in a relatively short period with little expense and minimal post-procedure care.