IL-1beta expressing neutrophil extracellular traps in Legionella pneumophila infection

IntroductionLegionella pneumophila is the causative agent of Legionnaires’ Disease (LD), an atypical pneumonia with potentially fatal outcome. Neutrophils, the first line of defense, infiltrate the lungs during L. pneumophila infection, although the precise immune mechanisms involved remain unclear.MethodsThis study aims to examine in vitro the interaction of neutrophils with L. pneumophila. Neutrophils from healthy individuals were infected with opsonized and non-opsonized bacteria. Phagocytosis was assessed by immunolabeling, and reactive oxygen species (ROS) generation by flow cytometry. The ability of neutrophils to form Neutrophil Extracellular Traps (NETs) in response to L. pneumophila and the impact of these NETs on bacterial proliferation were examined. Immunolabeling and Western blotting were used for specific NET-associated epitope detection.ResultsIt was demonstrated that neutrophils phagocytose opsonized L. pneumophila, while non-opsonized bacteria were not phagocytosed. Opsonized bacteria triggered ROS production, unlike non-opsonized bacteria. Neutrophils released NETs upon L. pneumophila interaction in a ROS-independent manner, but these NETs failed to inhibit bacterial proliferation. Notably, IL-1b was detected on NETs.DiscussionThis study provides evidence that neutrophils react to L. pneumophila through phagocytosis, the production of ROS, and NET release. IL-1b on NETs could play a role in complicated LD cases. These findings contribute to the understanding of neutrophil-mediated immune responses in LD

Quantitative Proteome Profiling of C. burnetii under Tetracycline Stress Conditions

The recommended antibiotic regimen against Coxiella burnetii, the etiological agent of Q fever, is based on a semi-synthetic, second-generation tetracycline, doxycycline. Here, we report on the comparison of the proteomes of a C. burnetii reference strain either cultured under control conditions or under tetracycline stress conditions. Using the MS-driven combined fractional diagonal chromatography proteomics technique, out of the 531 proteins identified, 5 and 19 proteins were found significantly up- and down-regulated respectively, under tetracycline stress. Although the predicted cellular functions of these regulated proteins did not point to known tetracycline resistance mechanisms, our data clearly reveal the plasticity of the proteome of C. burnetii to battle tetracycline stress. Finally, we raise several plausible hypotheses that could further lead to more focused experiments on studying tetracycline resistance in C. burnetii and thus reduced treatment failures of Q fever

Isolation and Characterization of Carbonosomes from <i>Pseudomonas</i> sp. phDV1 Grown Using Phenol as Carbon Source

The Pseudomonas sp. strain phDV1 was found to utilize monocyclic aromatic compounds as a sole carbon source and has a variety of potential applications in the bioremediation and biosynthesis of biodegradable plastics. It was possible to produce polyhydroxybutyrate when cultivated in the presence of monocyclic aromatic compounds as the sole carbon source. This study provides the small-scale optimization for phenol bioremediation and polyhydroxybutyrate production. The bacterium was cultivated in minimal medium supplemented with different concentrations of phenol. The formation and localization of the polyhydroxybutyrate granules (carbonosomes) in the cell were determined after 72 h of cultivation using Nile Red stain in combination with fluorescence microscopy. Analytical HPLC was also used to quantify the PHB content in the cells and to optimize the production. Finally, comparative proteomic analysis of isolated carbonosomes was used to characterize of their protein composition

The ultrastructure of Chlorobaculum tepidum revealed by cryo-electron tomography

Chlorobaculum (Cba) tepidum is a green sulfur bacterium that oxidizes sulfide, elemental sulfur, and thiosulfate for photosynthetic growth. As other anoxygenic green photosynthetic bacteria, Cba tepidum synthesizes bacteriochlorophylls for the assembly of a large light-harvesting antenna structure, the chlorosome. Chlorosomes are sac-like structures that are connected to the reaction centers in the cytoplasmic membrane through the BChl alpha-containing Fenna-Matthews-Olson protein. Most components of the photosynthetic machinery are known on a biophysical level, however, the structural integration of light harvesting with charge separation is still not fully understood. Despite over two decades of research, gaps in our understanding of cellular architecture exist. Here we present an in-depth analysis of the cellular architecture of the thermophilic photosynthetic green sulfur bacterium of Cba tepidum by cryo-electron tomography. We examined whole hydrated cells grown under different electron donor conditions. Our results reveal the distribution of chlorosomes in 3D in an unperturbed cell, connecting elements between chlorosomes and the cytoplasmic membrane and the distribution of reaction centers in the cytoplasmic membrane. (C) 2014 Elsevier B.V. All rights reserved

Membrane proteome of the green sulfur bacterium Chlorobium tepidum (syn. Chlorobaculum tepidum) analyzed by gel-based and gel-free methods

Chlorobium tepidum is a Gram-negative bacterium of the green sulfur phylum (Chlorobia). Chlorobia are obligate anaerobic photolithoautotrophs that are widely distributed in aquatic environments where anoxic layers containing reduced sulfur compounds are exposed to light. The envelope of C. tepidum is a complex organelle composed of the outer membrane, the periplasm-peptidoglycan layer, and the cytoplasmic membrane. In addition to the outer and plasma membranes, C. tepidum contains chlorosomes attached to the cytoplasmic side of the plasma membrane. Each cellular compartment has a unique set of proteins, called sub-proteome. An important aim of proteome analysis is to study the level of the expressed genes and their response to environmental changes. Membrane protein studies are of primary importance to understand how nutrients are transported inside the cell, how toxic molecules are exported, and the mechanisms of photosynthesis and energy metabolism