Utilizing nutrient type compounds as anti-bacterial compounds: arginine and cysteine inhibit Salmonella survival in egg white.

Because of growing levels of antibiotic resistance, new methods to combat bacteria are needed. We hypothesized that because bacteria evolved to survive in specific environments, the addition of compounds, including nutrient type compounds, to an environment, might result in a modification of that environment that will disrupt bacterial growth or in maladaptive bacterial behavior, i.e., gene expression. As a proof of concept, we focused on the egg white environment and the pathogen Salmonella. Despite egg white's antibacterial nature, Salmonella is able to survive and grow in egg white, and this ability of Salmonella leads to infection of chicks and humans. Here, the 20 L-amino-acids were screened for their ability to affect the growth of Salmonella in egg white. L-arginine and L-cysteine were found to reduce growth in egg white in physiologically relevant concentrations. To determine the mechanism behind L-arginine inhibition TnSeq was utilized. TnSeq identified many Salmonella genes required for survival in egg white including genes required for iron import, biotin synthesis, stress responses, cell integrity, and DNA repair. However, a comparison of Salmonella in egg white with and without L-arginine identified only a few differences in the frequency of transposon insertions, including the possible contribution of perturbations in the cell envelope to the inhibition mechanism. Finally, both D-arginine and D-cysteine were found to inhibit Salmonella in egg white. This implied that the effect of arginine and cysteine in egg white is chemical rather than biological, likely on the egg white environment or on the bacterial outer membrane. To conclude, these results show that this approach of addition of compounds, including nutrient type compounds, to an environment can be used to limit bacterial growth. Importantly, these compounds have no inherent anti-bacterial properties, are used as nutrients by animals and bacteria, and only become anti-bacterial in a specific environmental context. Future research screening for the effects of compounds in relevant environments might uncover new ways to reduce pathogen levels in the poultry industry and beyond.

The Effect of Oil-Based Cannabis Extracts on Metabolic Parameters and Microbiota Composition of Mice Fed a Standard and a High-Fat Diet.

The prevalence of obesity and obesity-related pathologies is lower in frequent cannabis users compared to non-users. It is well established that the endocannabinoid system has an important role in the development of obesity. We recently demonstrated that prolonged oral consumption of purified Δ-9 Tetrahydrocannabinol (THC), but not of cannabidiol (CBD), ameliorates diet-induced obesity and improves obesity-related metabolic complications in a high-fat diet mouse model. However, the effect of commercially available medical cannabis oils that contain numerous additional active molecules has not been examined. We tested herein the effects of THC- and CBD-enriched medical cannabis oils on obesity parameters and the gut microbiota composition of C57BL/6 male mice fed with either a high-fat or standard diet. We also assessed the levels of prominent endocannabinoids and endocannabinoid-like lipid mediators in the liver. THC-enriched extract prevented weight gain by a high-fat diet and attenuated diet-induced liver steatosis concomitantly with reduced levels of the lipid mediators palmitoyl ethanolamide (PEA) and docosahexaenoyl ethanolamide (DHEA) in the liver. In contrast, CBD-enriched extract had no effect on weight gain, but, on the contrary, it even exacerbated liver steatosis. An analysis of the gut microbiota revealed that mainly time but not treatment exerted a strong effect on gut microbiota alterations. From our data, we conclude that THC-enriched cannabis oil where THC is the main constituent exerts the optimal anti-obesity effects.

Cyclic-di-GMP regulation promotes survival of a slow-replicating subpopulation of intracellular Salmonella Typhimurium.

Salmonella Typhimurium can invade and survive within macrophages where the bacterium encounters a range of host environmental conditions. Like many bacteria, S. Typhimurium rapidly responds to changing environments by the use of second messengers such as cyclic di-GMP (c-di-GMP). Here, we generate a fluorescent biosensor to measure c-di-GMP concentrations in thousands of individual bacteria during macrophage infection and to define the sensor enzymes important to c-di-GMP regulation. Three sensor phosphodiesterases were identified as critical to maintaining low c-di-GMP concentrations generated after initial phagocytosis by macrophages. Maintenance of low c-di-GMP concentrations by these phosphodiesterases was required to promote survival within macrophages and virulence for mice. Attenuation of S Typhimurium virulence was due to overproduction of c-di-GMP-regulated cellulose, as deletion of the cellulose synthase machinery restored virulence to a strain lacking enzymatic activity of the three phosphodiesterases. We further identified that the cellulose-mediated reduction in survival was constrained to a slow-replicating persister population of S. Typhimurium induced within the macrophage intracellular environment. As utilization of glucose has been shown to be required for S. Typhimurium macrophage survival, one possible hypothesis is that this persister population requires the glucose redirected to the synthesis of cellulose to maintain a slow-replicating, metabolically active state.

Dynamics of the type III secretion system activity of enteropathogenic Escherichia coli.

UNLABELLED: Type III secretion systems (TTSSs) are employed by pathogens to translocate host cells with effector proteins, which are crucial for virulence. The dynamics of effector translocation, behavior of the translocating bacteria, translocation temporal order, and relative amounts of each of the translocated effectors are all poorly characterized. To address these issues, we developed a microscopy-based assay that tracks effector translocation. We used this assay alongside a previously described real-time population-based translocation assay, focusing mainly on enteropathogenic Escherichia coli (EPEC) and partly comparing it to Salmonella. We found that the two pathogens exhibit different translocation behaviors: in EPEC, a subpopulation that formed microcolonies carried out most of the translocation activity, while Salmonella executed protein translocation as planktonic bacteria. We also noted variability in host cell susceptibility, with some cells highly resistant to translocation. We next extended the study to determine the translocation dynamics of twenty EPEC effectors and found that all exhibited distinct levels of translocation efficiency. Further, we mapped the global effects of key TTSS-related components on TTSS activity. Our results provide a comprehensive description of the dynamics of the TTSS activity of EPEC and new insights into the mechanisms that control the dynamics. IMPORTANCE: EPEC and the closely related enterohemorrhagic Escherichia coli (EHEC) represent a global public health problem. New strategies to combat EPEC and EHEC infections are needed, and development of such strategies requires better understanding of their virulence machinery. The TTSS is a critical virulence mechanism employed by these pathogens, and by others, including Salmonella. In this study, we aimed at elucidating new aspects of TTSS function. The results obtained provide a comprehensive description of the dynamics of TTSS activity of EPEC and new insights into the mechanisms that control these changes. This knowledge sets the stage for further analysis of the system and may accelerate the development of new ways to treat EPEC and EHEC infections. Further, the newly described microscopy-based assay can be readily adapted to study the dynamics of TTSS activity in other pathogens.

The nuclear lamina and its functions in the nucleus.

The nuclear lamina is a structure near the inner nuclear membrane and the peripheral chromatin. It is composed of lamins, which are also present in the nuclear interior, and lamin-associated proteins. The increasing number of proteins that interact with lamins and the compound interactions between these proteins and chromatin-associated proteins make the nuclear lamina a highly complex but also a very exciting structure. The nuclear lamina is an essential component of metazoan cells. It is involved in most nuclear activities including DNA replication, RNA transcription, nuclear and chromatin organization, cell cycle regulation, cell development and differentiation, nuclear migration, and apoptosis. Specific mutations in nuclear lamina genes cause a wide range of heritable human diseases. These diseases include Emery-Dreifuss muscular dystrophy, limb girdle muscular dystrophy, dilated cardiomyopathy (DCM) with conduction system disease, familial partial lipodystrophy (FPLD), autosomal recessive axonal neuropathy (Charcot-Marie-Tooth disorder type 2, CMT2), mandibuloacral dysplasia (MAD), Hutchison Gilford Progeria syndrome (HGS), Greenberg Skeletal Dysplasia, and Pelger-Huet anomaly (PHA). Genetic analyses in Caenorhabditis elegans, Drosophila, and mice show new insights into the functions of the nuclear lamina, and recent structural analyses have begun to unravel the molecular structure and assembly of lamins and their associated proteins.