A convenient and robust in vivo reporter system to monitor gene expression in the human pathogen helicobacter pylori

Thirty years of intensive research have significantly contributed to our understanding of Helicobacter pylori biology and pathogenesis. However, the lack of convenient genetic tools, in particular the limited effectiveness of available reporter systems, has notably limited the toolbox for fundamental and applied studies. Here, we report the construction of a bioluminescent H. pylori reporter system based on the Photorhabdus luminescens luxCDABE cassette. The system is constituted of a promoterless lux acceptor strain in which promoters and sequences of interest can be conveniently introduced by double homologous recombination of a suicide transformation vector. We validate the robustness of this new lux reporter system in noninvasive in vivo monitoring of dynamic transcriptional responses of inducible as well as repressible promoters and demonstrate its suitability for the implementation of genetic screens in H. pylori. © 2012, American Society for Microbiology

Comparison of zootechnical performances, welfare condition and quality traits of hybrid striped bass (Morone chrysops x Morone saxatilis) reared in different Italian farms.

Growth performances of sunshine bass (initial m.b.w.=200\ub150 g) were examined in 3 different rearing conditions: concrete square basins (HSB-1); raceways (HSB-2); natural ponds (HSB-3). Fish received the same extruded feed (prot. 48.8%; lip. 17.8%). The trial lasted 16 months in HSB-1 and HSB-2 and 12 months in HSB-3 due to unfavourable welfare status and low growth performances. At harvest (16 months), sunshine bass reached a satisfactory mean weight without significant differences between groups. HSB-2 fish were longer with a smaller circumference, probably due to different hydrodynamics of the raceway. Glucose, cholesterol, triglycerides, protein, lactic dehydrogenase (LDH), aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), creatine kinase (CK), glycogen of plasma or tissue were not affected by rearing conditions as well as the proximate composition and the fatty acid profile of the fillet

Molecular Cloning and Characterization of the Human Diacylglycerol Kinase β (DGKβ) Gene ALTERNATIVE SPLICING GENERATES DGKβ ISOTYPES WITH DIFFERENT PROPERTIES

Diacylglycerol kinases are key modulators of levels of diacylglycerol, a second messenger involved in a variety of cellular responses to extracellular stimuli. A number of diacylglycerol kinases encoded by separate genes are present in mammalian genomes. We have cloned cDNAs encoding several isoforms of the human homologue of the rat diacylglycerol kinase β gene and characterized two such isoforms that differ at their carboxyl terminus through alternative splicing and the usage of different polyadenylation signals. Quantitative analysis of gene expression in a panel of human tissue cDNAs revealed that transcripts corresponding to both isoforms are co-expressed in central nervous system tissues and in the uterus, with one variant being expressed at relatively higher levels. As green fluorescent protein fusions, the two isoforms displayed localization to different subcellular compartments, with one variant being associated with the plasma membrane, while the other isoform was predominantly localized within the cytoplasm. Differences were also observed in their subcellular localization in response to phorbol ester stimulation. Enzymatic assays demonstrated that the two isoforms display comparable diacylglycerol kinase activities. Therefore, the human diacylglycerol kinase β gene can generate several enzyme isoforms, which can display different expression levels and subcellular localization but similar enzymatic activities in vitro

Paradigm Shift in Gastric Cancer Prevention: Harnessing the Potential of Aristolochia olivieri Extract

Gastric cancer, particularly adenocarcinoma, is a significant global health concern. Environmental risk factors, such as Helicobacter pylori infection and diet, play a role in its development. This study aimed to characterize the chemical composition and evaluate the in vitro antibacterial and antitumor activities of an Aristolochia olivieri Colleg. ex Boiss. Leaves’ methanolic extract (AOME). Additionally, morphological changes in gastric cancer cell lines were analyzed. AOME was analyzed using HPLC-MS/MS, and its antibacterial activity against H. pylori was assessed using the broth microdilution method. MIC and MBC values were determined, and positive and negative controls were included in the evaluation. Anticancer effects were assessed through in vitro experiments using AGS, KATO-III, and SNU-1 cancer cell lines. The morphological changes were examined through SEM and TEM analyses. AOME contained several compounds, including caffeic acid, rutin, and hyperoside. The extract displayed significant antimicrobial effects against H. pylori, with consistent MIC and MBC values of 3.70 ± 0.09 mg/mL. AOME reduced cell viability in all gastric cancer cells in a dose- and time-dependent manner. Morphological analyses revealed significant ultrastructural changes in all tumor cell lines, suggesting the occurrence of cellular apoptosis. This study demonstrated that AOME possesses antimicrobial activity against H. pylori and potent antineoplastic properties in gastric cancer cell lines. AOME holds promise as a natural resource for innovative nutraceutical approaches in gastric cancer management. Further research and in vivo studies are warranted to validate its potential clinical applications

Association of FOXO3A locus with extreme longevity in the Southern Italian Centenarian Study

A number of potential candidate genes in a variety of biological pathways have been associated with longevity in model organisms. Many of these genes have human homologs and thus have the potential to provide insights into human longevity. Recently, several studies suggested that FOXO3A functions as a key bridge for various signaling pathways that influence aging and longevity. Interestingly, Willcox and colleagues identified several variants that displayed significant genotype-gender interaction in male human longevity. In particular, a nested case-control study was performed in an ethnic Japanese population in Hawaii, and five candidate longevity genes were chosen based on links to the insulin-insulin-like growth factor-1 (IGF-1) signaling pathway. In the Willcox study, the investigated genetic variations (rs2802292, rs2764264, and rs13217795) within the FOXO3A gene were significantly associated with longevity in male centenarians. We validated the association of FOXO3A polymorphisms with extreme longevity in males from the Southern Italian Centenarian Study. Particularly, rs2802288, a proxy of rs2802292, showed the best allelic association-minor allele frequency (MAF) = 0.49; p = 0.003; odds ratio (OR) = 1.51; 95% confidence interval (CI), 1.15-1.98). Furthermore, we undertook a meta-analysis to explore the significance of rs2802292 association with longevity by combining the association results of the current study and the findings coming from the Willcox et al. investigation. Our data point to a key role of FOXO3A in human longevity and confirm the feasibility of the identification of such genes with centenarian-controls studies. Moreover, we hypothesize the susceptibility to the longevity phenotype may well be the result of complex interactions involving genes and environmental factors but also gender

Changes in microphytobenthos fluorescence over a tidal cycle: implications for sampling designs

Intertidal microphytobenthos (MPB) are important primary producers and provide food for herbivores in soft sediments and on rocky shores. Methods of measuring MPB biomass that do not depend on the time of collection relative to the time of day or tidal conditions are important in any studies that need to compare temporal or spatial variation, effects of abiotic factors or activity of grazers. Pulse amplitude modulated (PAM) fluorometry is often used to estimate biomass of MPB because it is a rapid, non-destructive method, but it is not known how measures of fluorescence are altered by changing conditions during a period of low tide. We investigated this experimentally using in situ changes in minimal fluorescence (F) on a rocky shore and on an estuarine mudflat around Sydney (Australia), during low tides. On rocky shores, the time when samples are taken during low tide had little direct influence on measures of fluorescence as long as the substratum is dry. Wetness from wave-splash, seepage from rock pools, run-off, rainfall, etc., had large consequences for any comparisons. On soft sediments, fluorescence was decreased if the sediment dried out, as happens during low-spring tides on particularly hot and dry days. Surface water affected the response of PAM and therefore measurements used to estimate MPB, emphasising the need for care to ensure that representative sampling is done during low tide

Protecting the environment through insect farming as a means to produce protein for use as livestock, poultry, and aquaculture feed

Securing protein for the approximate 10 billion humans expected to inhabit our planet by 2050 is a major priority for the global community. Evidence has accrued over the past 30 years that strongly supports and justifies the sustainable use of insects as a means to produce protein products as feed for pets, livestock, poultry, and aquacultured species. Researchers and entrepreneurs affiliated with universities and industries, respectively, from 18 nations distributed across North and South America, Europe, Asia, Africa and Australia contributed to the development of this article, which is an indication of the global interest on this topic. A brief overview of insects as feed for the aquaculture industry along with a review of the black soldier fly, Hermetia illucens (Diptera: Stratiomyidae), as a model for such systems is provided

Characterizing Ligand-Gated Ion Channel Receptors with Genetically Encoded Ca++ Sensors

We present a cell based system and experimental approach to characterize agonist and antagonist selectivity for ligand-gated ion channels (LGIC) by developing sensor cells stably expressing a Ca2+ permeable LGIC and a genetically encoded Förster (or fluorescence) resonance energy transfer (FRET)-based calcium sensor. In particular, we describe separate lines with human α7 and human α4β2 nicotinic acetylcholine receptors, mouse 5-HT3A serotonin receptors and a chimera of human α7/mouse 5-HT3A receptors. Complete concentration-response curves for agonists and Schild plots of antagonists were generated from these sensors and the results validate known pharmacology of the receptors tested. Concentration-response relations can be generated from either the initial rate or maximal amplitudes of FRET-signal. Although assaying at a medium throughput level, this pharmacological fluorescence detection technique employs a clonal line for stability and has versatility for screening laboratory generated congeners as agonists or antagonists on multiple subtypes of ligand-gated ion channels. The clonal sensor lines are also compatible with in vivo usage to measure indirectly receptor activation by endogenous neurotransmitters