Differentiation of marine luminous bacteria using commercial identification plates

Differentiation of marine luminous bacteria using Biolog GN plate combined with API 20e or the BBL Crystal ID plate inoculated with cell suspensions in artifical seawater was accomplished by comparison to type species using cluster analysis. Inoculum density affected the results from Biolog GN plates, but had less of an effect on the reactions obtained from API 20e strip or BBL Crystal ID plate. In a few cases, combination of the Biolog GN traits along with either the API 20e or Crystal ID traits was necessary to differentiate some marine luminous bacteria. © 1998 John Wiley & Sons, Ltd.     

A new gas vacuolated heterotrophic rod from freshwaters

Summary A gas vacuolated rod was isolated from the oxygen-depleted zone of a eutrophic lake. It is a non-motile, euryoxic, gram negative heterotroph that grows slowly under all conditions tested. The vesicles of the gas vacuoles of the bacterium revealed by the electron microscope resemble those of other gas vacuolate prokaryotes. The taxonomy of the isolate is discussed but no names have been proposed.ESE Pub. No. 278.     

Genotoxic action of Luna Experience-SC 400 fungicide on rat bone marrow

Abstract

Background: Fungicides describe all chemicals used to control fungi that infect plants. Luna Experience SC-400 is a new line of fungicide that consist of Fluopyram and Tebuconazole.

Objective: In this study, We investigated the genotoxicty and cytotoxicty of Luna Experience-SC 400 using comet assay, micronucleus test and polychromatic erythrocytes number in rat bone marrow. The present study is the first report indicating the effects of genotoxic and cytotoxic of Luna experience SC-400 on rat bone marrow cells.

Material and Methods: We used three different doses (5mg/kg, 10mg/kg, 20mg/kg) of Luna Experience SC 400 at 48 h intervals during 30 days by gavage in rats.Genotoxicity was evaluated using comet assay and micronucleus test and cytotoxicity was measured the PCE/NCE rate in rat bone marrow.

Results: Based on these experimental results, we report that Luna Experience-SC 400 fungicide presents genotoxic and cytotoxic potential on rat bone marrow. There is a significant difference between negative control group and all the doses of Luna Experience-SC 400 (p?<?0.05) for comet assay and micronucleus. Even moderate and high doses of fungicides seem to have reached the values of almost positive control group for Genetic Damage Index (GDI) and Damaged Cell Percentage (DCP). In this study, we also investigated the PCE/NCE rate. Fungicide caused a decrease in the level of significant in the PCE/NCE ratio (p?<?0.05).

Conclusion: Our in vivo study suggests that the gavage exposure to Luna experience SC 400 used in the present investigation may be genotoxic and cytotoxic in rat bone marrow in view of these findings. Because this findings is first report represented in the pesticide biology, it is important to carry out more investigations using various cytogenetic tests under different experimental conditions to definitively resolve the the possible genotoxic and cytotoxic risk associated with new generation pesticides-fungicides.     

Loss of SIRT4 decreases GLT‐1‐dependent glutamate uptake and increases sensitivity to kainic acid

Glutamate transport is a critical process in the brain that maintains low extracellular levels of glutamate to allow for efficient neurotransmission and prevent excitotoxicity. Loss of glutamate transport function is implicated in epilepsy, traumatic brain injury, and amyotrophic lateral sclerosis. It remains unclear whether or not glutamate transport can be modulated in these disease conditions to improve outcome. Here, we show that sirtuin (SIRT)4, a mitochondrial sirtuin, is up‐regulated in response to treatment with the potent excitotoxin kainic acid. Loss of SIRT4 leads to a more severe reaction to kainic acid and decreased glutamate transporter expression and function in the brain. Together, these results indicate a critical and novel stress response role for SIRT4 in promoting proper glutamate transport capacity and protecting against excitotoxicity.

     

Imaging Cleared Intact Biological Systems at a Cellular Level by 3DISCO

Tissue clearing and subsequent imaging of transparent organs is a powerful method to analyze fluorescently labeled cells and molecules in 3D, in intact organs. Unlike traditional histological methods, where the tissue of interest is sectioned for fluorescent imaging, 3D imaging of cleared tissue allows examination of labeled cells and molecules in the entire specimen. To this end, optically opaque tissues should be rendered transparent by matching the refractory indices throughout the tissue. Subsequently, the tissue can be imaged at once using laser-scanning microscopes to obtain a complete high-resolution 3D image of the specimen. A growing list of tissue clearing protocols including 3DISCO, CLARITY, Sca/e, ClearT2, and SeeDB provide new ways for researchers to image their tissue of interest as a whole. Among them, 3DISCO is a highly reproducible and straightforward method, which can clear different types of tissues and can be utilized with various microscopy techniques. This protocol describes this straightforward procedure and presents its various applications. It also discusses the limitations and possible difficulties and how to overcome them.     

Structural determination of Streptococcus pyogenes M1 protein interactions with human immunoglobulin G using integrative structural biology

Streptococcus pyogenes (Group A streptococcus; GAS) is an important human pathogen responsible for mild to severe, life-threatening infections. GAS expresses a wide range of virulence factors, including the M family proteins. The M proteins allow the bacteria to evade parts of the human immune defenses by triggering the formation of a dense coat of plasma proteins surrounding the bacteria, including IgGs. However, the molecular level details of the M1-IgG interaction have remained unclear. Here, we characterized the structure and dynamics of this interaction interface in human plasma on the surface of live bacteria using integrative structural biology, combining cross-linking mass spectrometry and molecular dynamics (MD) simulations. We show that the primary interaction is formed between the S-domain of M1 and the conserved IgG Fc-domain. In addition, we show evidence for a so far uncharacterized interaction between the A-domain and the IgG Fc-domain. Both these interactions mimic the protein G-IgG interface of group C and G streptococcus. These findings underline a conserved scavenging mechanism used by GAS surface proteins that block the IgG-receptor (FcγR) to inhibit phagocytic killing. We additionally show that we can capture Fab-bound IgGs in a complex background and identify XLs between the constant region of the Fab-domain and certain regions of the M1 protein engaged in the Fab-mediated binding. Our results elucidate the M1-IgG interaction network involved in inhibition of phagocytosis and reveal important M1 peptides that can be further investigated as future vaccine targets.     

Hsp70 chaperone ligands control domain association via an allosteric mechanism mediated by the interdomain linker

Hsp70 chaperones assist in protein folding, disaggregation, and membrane translocation by binding to substrate proteins with an ATP-regulated affinity that relies on allosteric coupling between ATP-binding and substrate-binding domains. We have studied single- and two-domain versions of the E. coli Hsp70, DnaK, to explore the mechanism of interdomain communication. We show that the interdomain linker controls ATPase activity by binding to a hydrophobic cleft between subdomains IA and IIA. Furthermore, the domains of DnaK dock only when ATP binds and behave independently when ADP is bound. Major conformational changes in both domains accompany ATP-induced docking: of particular importance, some regions of the substrate-binding domain are stabilized, while those near the substrate-binding site become destabilized. Thus, the energy of ATP binding is used to form a stable interface between the nucleotide- and substrate-binding domains, which results in destabilization of regions of the latter domain and consequent weaker substrate binding.