Functional Metagenomics: A High Throughput Screening Method to Decipher Microbiota-Driven NF-κB Modulation in the Human Gut

Background/Aim: The human intestinal microbiota plays an important role in modulation of mucosal immune responses. To study interactions between intestinal epithelial cells (IECs) and commensal bacteria, a functional metagenomic approach was developed. One interest of metagenomics is to provide access to genomes of uncultured microbes. We aimed at identifying bacterial genes involved in regulation of NF-kappa B signaling in IECs. A high throughput cell-based screening assay allowing rapid detection of NF-kappa B modulation in IECs was established using the reporter-gene strategy to screen metagenomic libraries issued from the human intestinal microbiota. Methods: A plasmid containing the secreted alkaline phosphatase (SEAP) gene under the control of NF-kappa B binding elements was stably transfected in HT-29 cells. The reporter clone HT-29/kb-seap-25 was selected and characterized. Then, a first screening of a metagenomic library from Crohn's disease patients was performed to identify NF-kappa B modulating clones. Furthermore, genes potentially involved in the effect of one stimulatory metagenomic clone were determined by sequence analysis associated to mutagenesis by transposition. Results: The two proinflammatory cytokines, TNF-alpha and IL-1 beta, were able to activate the reporter system, translating the activation of the NF-kappa B signaling pathway and NF-kappa B inhibitors, BAY 11-7082, caffeic acid phenethyl ester and MG132 were efficient. A screening of 2640 metagenomic clones led to the identification of 171 modulating clones. Among them, one stimulatory metagenomic clone, 52B7, was further characterized. Sequence analysis revealed that its metagenomic DNA insert might belong to a new Bacteroides strain and we identified 2 loci encoding an ABC transport system and a putative lipoprotein potentially involved in 52B7 effect on NF-kappa B. Conclusions: We have established a robust high throughput screening assay for metagenomic libraries derived from the human intestinal microbiota to study bacteria-driven NF-kappa B regulation. This opens a strategic path toward the identification of bacterial strains and molecular patterns presenting a potential therapeutic interest

The Influence of the Type of Lime on the Hygric Behaviour and Bio-Receptivity of Hemp Lime Composites Used for Rendering Applications in Sustainable New Construction and Repair Works

The benefits of using sustainable building materials are linked not only to the adoption of manufacturing processes that entail reduced pollution, CO2 emissions and energy consumption, but also to the onset of improved performance in the building. In particular, hemp-lime composite shows low shrinkage and high thermal and acoustic insulating properties. However, this material also shows a great ability to absorb water, an aspect that can turn out to be negative for the long-term durability of the building. For this reason, the hygric properties of hemp-based composites need to be studied to ensure the correct use of this material in construction and repair works. The water absorption, drying and transpirability of hemp composites made with aerial (in the form of dry powder and putty) and hydraulic limes were investigated here and related to the microbial growth induced by the water movements within the material. Results show that hemp-natural hydraulic lime mixes exhibit the highest transpirability and drying rate, the lowest water absorption by immersion and capillary uptake and the least intense microbial attack and chromatic change. A microscopical study of the hemp shives also related their great ability to absorb water to the near-irreversible swelling of their structure under dry-wet conditions.The research leading to these results has received funding from the European Union's Seventh Framework Programme (FP7/2007-2013) under grant agreement n° 326983 (NaturALiMe), and the Spanish project MAT-2012-34473 of the Ministerio de Ciencia y Competitividad. Author MB, owner of the CANNABRIC company, had some role in the design and preparation of mortar samples and in the preparation of this manuscript, but did not have any additional role in data collection and analysis

Approaches in biotechnological applications of natural polymers

Natural polymers, such as gums and mucilage, are biocompatible, cheap, easily available and non-toxic materials of native origin. These polymers are increasingly preferred over synthetic materials for industrial applications due to their intrinsic properties, as well as they are considered alternative sources of raw materials since they present characteristics of sustainability, biodegradability and biosafety. As definition, gums and mucilages are polysaccharides or complex carbohydrates consisting of one or more monosaccharides or their derivatives linked in bewildering variety of linkages and structures. Natural gums are considered polysaccharides naturally occurring in varieties of plant seeds and exudates, tree or shrub exudates, seaweed extracts, fungi, bacteria, and animal sources. Water-soluble gums, also known as hydrocolloids, are considered exudates and are pathological products; therefore, they do not form a part of cell wall. On the other hand, mucilages are part of cell and physiological products. It is important to highlight that gums represent the largest amounts of polymer materials derived from plants. Gums have enormously large and broad applications in both food and non-food industries, being commonly used as thickening, binding, emulsifying, suspending, stabilizing agents and matrices for drug release in pharmaceutical and cosmetic industries. In the food industry, their gelling properties and the ability to mold edible films and coatings are extensively studied. The use of gums depends on the intrinsic properties that they provide, often at costs below those of synthetic polymers. For upgrading the value of gums, they are being processed into various forms, including the most recent nanomaterials, for various biotechnological applications. Thus, the main natural polymers including galactomannans, cellulose, chitin, agar, carrageenan, alginate, cashew gum, pectin and starch, in addition to the current researches about them are reviewed in this article.. }To the Conselho Nacional de Desenvolvimento Cientfíico e Tecnológico (CNPq) for fellowships (LCBBC and MGCC) and the Coordenação de Aperfeiçoamento de Pessoal de Nvíel Superior (CAPES) (PBSA). This study was supported by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UID/BIO/04469/2013 unit, the Project RECI/BBB-EBI/0179/2012 (FCOMP-01-0124-FEDER-027462) and COMPETE 2020 (POCI-01-0145-FEDER-006684) (JAT)

Geology of the construction materials of the church of Santa Maria degli Angeli in Alì Superiore (NE Sicily, Italy)

This study deals with the petrographic analysis of a great variety of construction materials used to build a 16th century church in NE Sicily. X-ray diffraction and optical microscopy techniques were useful to establish the composition and the texture of natural stones (phyllites, schists, marbles and volcanic scoriae) and artificial materials (mortars and bricks). The quarry areas of natural stones are located around the church where they can be correlated to geological formations. Information about mortars and brick production is provide

Direct observation of microcrack development in marble caused by thermal weathering

One of the properties that makes marble such an excellent construction and ornamental material is its low porosity. It is very difficult for water or decay agents to penetrate the internal structure of materials with no or few pores, so enhancing the durability of these materials. However, environmental temperature fluctuations bring about significant physical changes in marbles that result in an increase in porosity, due to the appearance of new microcracks and the expansion of existing ones. These cracks offer new paths into the marble which make it easier for solutions containing pollutants to penetrate the material. Thermal expansion tests were performed on three different types of marble known as White, Tranco, and Yellow Macael (Almeria, Spain), after which an increase in porosity (from 17 to 73% depending on marble type) was observed, mainly due to crack formation. The structural changes occurring during thermal expansion tests were more significant in the case of White Macael samples, a fact that is not only related to its mineralogical composition but also to the morphology of the grains, grain boundaries and crystal size. Our research suggests that thermally weathered White Macael marble could be more susceptible to decay by other contaminant agents than Tranco or Yellow Macael. The use of hot-stage environmental scanning electron microscopy is proposed as a valid tool for observing, both in situ and at high magnification, changes in the fracture system of building stones induced by thermal stress