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BACKGROUND AND AIM: Nitrogen-fixing bacteria or diazotrophs have been isolated for many years using different formulations of N-free semi-solid media. However, the strategies used to isolate them, and the recipes of these media, are scattered through the published literature and in other sources that are more difficult to access and which are not always retrievable. Therefore, the aim of this work was to collate the various methods and recipes, and to provide a comprehensive methodological guide and their use by the scientific community working in the field of biological nitrogen fixation (BNF), particularly with non-leguminous plants. METHODS: Procedures used for bacterial counting and identification either from rhizosphere soil or on the surface of, or within, plant tissues (to access “endophytic” bacteria) are presented in detail, including colony and cell morphologies. More importantly, appropriate recipes available for each N-free semi-solid culture medium that are used to count and isolate various diazotrophs are presented. RESULTS: It is recognized by those working in the field of BNF with non-legumes that the development of the N-free semi-solid medium has allowed a tremendous accumulation of knowledge on the ecology and physiology of their associated diazotrophs. At least 20 nitrogen-fixing species have been isolated and identified based on the enrichment method originally developed by Döbereiner, Day and collaborators in the 70’s. In spite of all the advances in molecular techniques used to detect bacteria, in most cases the initial isolation and identification of these diazotrophs still requires semi-solid media. CONCLUSIONS: The introduction of the N-free semi-solid medium opened new opportunities for those working in the area of BNF with non-legumes not only for elucidating the important role played by their associated microorganisms, but also because some of these bacteria that were isolated using semi-solid media are now being recommended as plant growth-promoting inoculants for sugarcane (Saccharum sp.), maize (Zea mays) and wheat (Triticum aestivum) in Brazil and other countries. Further progress in the field could be made by using a combination of culture-independent molecular community analyses, in situ activity assessments with probe-directed enrichment, and isolation of target strains using modified or standard semi-solid media.

Adipose tissue-derived stem cells (ADSCs) are an available source of mesenchymal stem cells with the appropriate capacity to in vitro survive, propagate, and differentiate into cells from three lineages of ectoderm, mesoderm, and endoderm. The biological features of ADSCs depend on the donor physiology and health status, isolation procedure, culture conditions, and differentiation protocols used. Adipose tissue samples are provided by surgery and lipoaspiration-based methods and subjected to various mechanical and chemical digestion techniques to finally generate a heterogeneous mixture named stromal vascular fraction (SVF). ADSCs are purified through varied cell populations that exist within SVF and cultured under standard conditions to give rise to a highly rich resource of stem cells directly applied in the clinic or differentiated into a wide range of cells. The development and optimization of conventional isolation, expansion, and differentiation methods seem noteworthy to preserve the desirable biological functions of ADSCs in pre-clinical and clinical investigations.

The study was conducted to standardize the parameters of centrifugation method of starch isolation for sweet potato to yield better quality starch. The effects of solid-to-solvent ratio in slurry-making, numbers of blending, slurry filtration and starch washings were studied for obtaining maximum yield and purity of starch with desired color and amylose content. The goal of the study was to develop a suitable process for small scale/on-farm processing of sweet potato for preparation of starch. A widely grown pink-skinned cream fleshed cultivar ( Kanjangad ) was used for the experiments. The studied parameters were slurry making solid-to-solvent ratio (1:2, 1:3 and 1:4), number of blendings (1 and 2 times), number of slurry filtration water washings (2, 3 and 4 times) and starch washings (2, 3 and 4 times). The study showed that the starch yield, purity, color and amylose were influenced by altering the starch isolation procedures. The solid-to-solvent ratio of 1:2 in the starch isolation process yielded the best results. Also, the increase in number of blendings degraded the purity, amylose and color of starch. The optimal process was slurry making at 1:2 solid-to-solvent ratio and once blending, slurry filtration washing for 2 times and starch washing for 2 times. Accordingly, the starch yield, purity, amylose, and L, a and b values were 13.967 ± 1.135%, 80.640 ± 1.139%, 29.220 ± 0.759%, and 57.88 ± 2.982, 0.386 ± 0.204, and 2.68 ± 0.127, respectively. The prepared starch was characterized for chemical composition, water activity, paste clarity, swelling power, water solubility index, pasting property and granule size and morphology. The study suggested that the prepared starch was suitable for food uses.

The ability to form biofilms is important for environmental survival, transmission, and infectivity of Vibrio cholerae, the causative agent of cholera in humans. To form biofilms, V. cholerae produces an extracellular matrix composed of proteins, nucleic acids and a glycoconjugate, termed Vibrio exopolysaccharide (VPS). Here, we present the data on isolation and characterization of the polysaccharide part of the VPS (VPS-PS), which has the following structure: -4)-α-GulpNAcAGly3OAc-(1-4)-β-D-Glcp-(1-4)-α-Glcp-(1-4)-α-D-Galp-(1- where α-D-Glc is partially (,20%) replaced with α-D-GlcNAc. α-GulNAcAGly is an amide between 2-acetamido-2-deoxy-aguluronic acid and glycine. Apparently, the polysaccharide is bound to a yet unidentified component, which gives it high viscosity and completely suppresses any NMR signals belonging to the sugar chains of the VPS. The only reliable method to remove this component at present is a treatment of the whole glycoconjugate with concentrated hydrochloric acid.

subspecies is explored as an anti-cariogenic probiotic. Here, subjecting freshly stimulated saliva samples of 35 healthy volunteers, six epidemiologically unrelated and two related strains were isolated (prevalence around 20%) applying a newly developed three-step procedure. Furthermore, the probiotic strain 7746 (AB-Dentisanium®) was tested under a variety of environmental conditions for its inhibitory effect on six , two , 15 other oral or intestinal streptococci, 15 strains, and six representatives of other species including periodontopathogens. All except one of the strains were inhibited by 7746 colonies or culture supernatant concentrate but only if either the test cell number was low or the producer or its bacteriocin concentration, respectively, was high. OMI 332, OMI 315, OMI 335, OMI 238, and the intestinal OMI 339 were not inhibited, while the other 10 streptococcal strains (especially OMI 334 and intestinal OMI 326) showed a certain degree of inhibition. From the panel of other bacterial species only was slightly inhibited. With the exception of OMI 285 and OMI 291 that possessed a 7746 bacteriocin-like gene cluster, all strains and especially type strain 7747 were strongly inhibited by 7746. In conclusion, probiotic strain 7746 might antagonize the initiation and progression of dental caries by reducing if not too abundant. strains inhibit each other, but strains with similar bacteriocin-related gene clusters, including immunity genes, are able to co-exist due to cross-resistance. In addition, development of resistance and adaptation to 7746-bacteriocins was observed during our study and needs attention. Hence, mechanisms underlying such processes need to be further investigated using omics-approaches. On the manufacturing level, probiotic strains should be continuously tested for function. Further clinical studies investigating inhibition of by AB-Dentisanium® are required that should also monitor the impact on the oral microbiome composition including resident strains.

Two new cyclotetrapeptides, sartoryglabramides A (5) and B (6), and a new analog of fellutanine A (8) were isolated, together with six known compounds including ergosta-4, 6, 8 (14), 22-tetraen-3-one, ergosterol 5, 8-endoperoxide, helvolic acid, aszonalenin (1), (3R)-3-(1H-indol-3-ylmethyl)-3,4-dihydro-1H-1,4-benzodiazepine-2,5-dione (2), takakiamide (3), (11aR)-2,3-dihydro-1H-pyrrolo[2,1-c][1,4]benzodiazepine-5,11(10H,11aH)-dione (4), and fellutanine A (7), from the ethyl acetate extract of the culture of the marine sponge-associated fungus Neosartorya glabra KUFA 0702. The structures of the new compounds were established based on extensive 1D and 2D spectral analysis. X-ray analysis was also used to confirm the relative configuration of the amino acid constituents of sartoryglabramide A (5), and the absolute stereochemistry of the amino acid constituents of sartoryglabramide A (5) and sartoryglabramides B (6) was determined by chiral HPLC analysis of their hydrolysates by co-injection with the D- and L- amino acids standards. Compounds 1-8 were tested for their antibacterial activity against Gram-positive (Escherichia coli ATCC 25922) and Gram-negative (Staphyllococus aureus ATCC 25923) bacteria, as well as for their antifungal activity against filamentous (Aspergillus fumigatus ATCC 46645), dermatophyte (Trichophyton rubrum ATCC FF5) and yeast (Candida albicans ATCC 10231). None of the tested compounds exhibited either antibacterial (MIC > 256 μg/mL) or antifungal activities (MIC > 512 μg/mL). © 2016 by the authors; licensee MDPI.

This work was developed in the Natural Products Research Laboratory of the Department of Chemistry, Instituto de Ciências Biomédicas Abel Salazar (ICBAS), of the University of Porto and partially supported through national funds provided by FCT-Foundation for Science and Technology and European Regional Development Fund (ERDF) and COMPETE, under the projects PEst-C/MAR/LA0015/2013, PTDC/MAR-BIO/4694/2014, as well as by the project INNOVMAR (Innovation and Sustainability in the Management and Exploitation of Marine Resources) (Reference NORTE-01-0145-FEDER-000035, within Research Line NOVELMAR/INSEAFOOD/ECOSERVICES), supported by the North Portugal Regional Operational Programme (NORTE 2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF). We thank Mrs. Júlia Bessa and Sara Cravo for technical support.

Background Infectious gastroenteritis is common in the emergency department (ED). Patients infected with either Norovirus or toxigenic Clostridium difficile require special isolation procedures. The aims were to describe the aetiology of infectious gastroenteritis in the ED, evaluate whether current isolation procedures, based on clinical judgement are sufficient, and to identify information that might be used to identify patients requiring isolation. Methods Prospective, observational, multicentre study. We collected information on symptoms, vital signs, travel history, the recent use of antibiotics, and infectious contacts and tested faecal samples for Norovirus, C. difficile, and enteropathogenic bacteria. Results The study enrolled 227 patients, of whom 163 (71%) delivered a faecal sample for Norovirus analysis (13% positive), 171 (74%) for C. difficile (13% positive), and 173 (76%) for enteropathogenic bacteria (16% positive). In total 71% of the patients were isolated using strict precautions, 29% of the isolated patient and 14% of the patients who were not isolated had had a highly contagious GE. Risk factors for Norovirus included frequent vomiting (OR 5.5), recent admission of another patient with Norovirus (OR 2.6), and a short duration of diarrhoea. Risk factors for C. difficile infections included older age (OR 6.0), longer duration of diarrhoea (OR 5.2), mucus in stool (OR 3.5), and previous antibiotic use (OR 23.4). Conclusion Highly contagious GE occurs in ¼ of the GE patients in the EDs, isolation based on clinical judgement is not very efficient. Several risk factors can predict the presence of Norovirus or toxigenic Clostridium difficile . It is uncertain whether this knowledge can improve isolation practices in ED settings. Trial registration This study was retrospectively registered in the Clinical Trials Data Base ( NCT02685527 ) and prospectively approved by the Regional Committees on Health Research Ethics for Southern Denmark (project ID S20140200) and Ethics Committee at the Medical Association of Schleswig-Holstein [“Ethikkommission bei der Ärztekammer Schleswig-Holstein”, project ID 120/15(I)] and registered with the Danish Data Protection Agency (project ID nr. 2008-58-0035/ 1608).

WHO recommends surveillance for COVID-19 among travelers at Points of Entry (POE) to countries. At 13 selected POE at the Nepal-India border, between March 2021 and July 2021, we describe the screening, testing, diagnosis and isolation practices of COVID-19 amongst travelers. Those who stayed in India or elsewhere for > one day and those who did not have a negative RT-PCR result within the last 72 h of travel were tested for COVID-19 with rapid antigen diagnostic tests. Daily surveillance reports maintained at POE were used for analysis. Of 337,338 travelers screened, 69,886 (21%) were tested and 3907 (6%) were diagnosed with COVID-19. The proportions tested averaged 15% during April-May when screened numbers were high and increased to 35% in July when screened numbers had decreased. The proportions diagnosed positive peaked at 10% in April-May, but decreased to below 1% in June and July. Testing coverage varied from 0–99% in the different POE. Most COVID-19 cases were Nepalese, male, <60 years of age, migrant workers and presented with fever. Of COVID-19 cases, 32% had home-based isolation, 64% underwent community-based isolation and the remainder either went to hospital or returned to India. In conclusion, about one fifth of travelers overall were tested, with coverage varying considerably over time and among different POE. Strengthening surveillance processes at POE is needed.

For the last decades, the fate of lignins in soil was analyzed mainly with cupric oxide (CuO) oxidation, which is traditionally used to quantify soil lignin content and characterize its state of degradation. This method presents limitations due to incomplete depolymerization of the lignin structure. In this study, we used a physicochemical soil lignin isolation procedure, which permits recovery of a milled wall enzymatic lignin (MWEL) fraction. Elemental composition and chemical structure of MWEL isolated from plants and soil were characterized. Its incorporation rate into an agricultural loamy soil was studied using stable isotope analyses of MWEL isolated from soils after 0 to 9 years of maize cultivation after wheat. Comparison of MWEL isolated from maize tissues and soil provided information on evolution of the lignin structure once incorporated into soil. We observed aromatic–aliphatic complex formation, which could lead to its sequestration in soil evidenced by increasing MWEL content after 9 years of maize cultivation. The 13 C natural abundance of isolated MWEL showed faster incorporation of MWEL (17.4 % of renewed lignins after 9 years) compared to total soil organic matter (9 % of total soil organic carbon (SOC) was renewed). This faster incorporation rate of MWEL compared to bulk soil organic matter is in agreement with lignin turnover observed by CuO oxidation. Radiocarbon dating of MWEL suggested a mean age of around 50 years. We conclude that lignin isolation allows (1) access to a different fraction compared to CuO oxidation and (2) a detailed characterization of lignin transformation in soil. We suggest that interaction with aliphatic compounds could be one possible pathway of lignin preservation in soil.