Interactions between Spider Silk and Cells - NIH/3T3 Fibroblasts Seeded on Miniature Weaving Frames

Background: Several materials have been used for tissue engineering purposes, since the ideal matrix depends on the desired tissue. Silk biomaterials have come to focus due to their great mechanical properties. As untreated silkworm silk has been found to be quite immunogenic, an alternative could be spider silk. Not only does it own unique mechanical properties, its biocompatibility has been shown already in vivo. In our study, we used native spider dragline silk which is known as the strongest fibre in nature. Methodology/Principal Findings: Steel frames were originally designed and manufactured and woven with spider silk, harvesting dragline silk directly out of the animal. After sterilization, scaffolds were seeded with fibroblasts to analyse cell proliferation and adhesion. Analysis of cell morphology and actin filament alignment clearly revealed adherence. Proliferation was measured by cell count as well as determination of relative fluorescence each after 1, 2, 3, and 5 days. Cell counts for native spider silk were also compared with those for trypsin-digested spider silk. Spider silk specimens displayed less proliferation than collagen-and fibronectin-coated cover slips, enzymatic treatment reduced adhesion and proliferation rates tendentially though not significantly. Nevertheless, proliferation could be proven with high significance (p<0.01). Conclusion/Significance: Native spider silk does not require any modification to its application as a biomaterial that can rival any artificial material in terms of cell growth promoting properties. We could show adhesion mechanics on intracellular level. Additionally, proliferation kinetics were higher than in enzymatically digested controls, indicating that spider silk does not require modification. Recent findings concerning reduction of cell proliferation after exposure could not be met. As biotechnological production of the hierarchical composition of native spider silk fibres is still a challenge, our study has a pioneer role in researching cellular mechanics on native spider silk fibres.

Assessment of fungal contamination in a Portuguese maternity unit

A descriptive study was developed to monitor air fungal contamination in one Portuguese maternity. Sixty air samples were collected through impaction method. Air sampling was performed in food storage facilities, kitchen, food plating, canteen, pharmacy, sterilization areas, genecology wards, intensive care unit, operating rooms, urgency and also, outside premises, since this was the place regarded as reference. Besides air samples, forty three samples were collected by swabbing the surfaces using a 10 by 10 cm square stencil. Simultaneously, temperature, relative humidity and particles counting (PM10) were registered. Twenty three species of fungi were identified in air, being the two most commonly isolated the genera Penicillium (41,5%) and Cladosporium (28,4%). Regarding yeasts, only Rhodotorula sp. (45,2%), Trichosporon mucoides (51,6%) and Cryptococcus neoformans (3,2%) were found. Thirteen species of fungi were identified in surfaces, being the most frequent the Penicillium genus (91,6%). Concerning yeasts found in surfaces, four species were identified being Rhodotorula sp. (29,1%) the most frequent. There was no coincidence between prevailing genera indoors and outside premises. Moreover, some places presented fungal species different from the ones isolated outside. In the inside environment, Aspergillus species were isolated in air and surfaces. There was no significant relationship (p>0,05) between fungal contamination and the studied environmental variables. Keywords: air, surfaces, fungal contamination, environmental variables, maternity.

Bacterial biofilms, antibiotic resistance and healthcare-associated infections: a dangerous connection

In 2012, were estimated 6.7 million cases of healthcare-associated infections (HAI) either in long-term care facilities or acute-care hospitals from which result 37,000 deaths configuring a serious public health problem. The etiological agents are diverse and often resistant to antimicrobial drugs. One of the mechanisms responsible for the emergence of drug resistance is biofilm assembly. Biofilms are defined as thin layers of microorganisms adhering to the surface of a structure, which may be organic or inorganic, together with the polymers that they secrete. They are dynamic structures which experience different stages of organization with the ageing and are linked to an increase in bacterial resistance to host defense mechanisms, antibiotics, sterilization procedures other than autoclaving, persistence in water distribution systems and other surfaces. The understanding of bacteria organization within the biofilm and the identification of differences between planktonic and sessile forms of bacteria will be a step forward to fight HAIs.

UNDERSTANDING THE RELATIONSHIP BETWEEN THE BROWN MARMORATED STINK BUG, HALYOMORPHA HALYS (STÅL), AND ITS SYMBIONT, PANTOEA CARBEKII, WITH IMPLICATIONS FOR STINK BUG MANAGEMENT

Symbiotic relationships between insects and beneficial microbes are very common in nature, especially within the Hemiptera. The brown marmorated stink bug, Halyomorpha halys Stål, harbors a symbiont, Pantoea carbekii, within the fourth region of the midgut in specialized crypts. In this dissertation, I explored this insect- microbe relationship. I determined that the brown marmorated stink bug is heavily reliant on its symbiont, and that experimental removal of the symbiont from the egg mass surface prior to nymphal acquisition led to lower survival, longer development, lower fecundity, and aberrant nymphal behavior. Additionally, I determined that even when the symbiont is acquired and housed in the midgut crypts, it is susceptible to stressors. Stink bugs reared at a higher temperature showed lower survival, longer development, and a cease in egg mass production, and when bugs were screened for their symbiont, fewer had successfully retained it while under heat stress. Finally, with the knowledge that the stink bug suffers decreases in fitness when its symbiont is missing or stressed, I wanted to determine if targeting the symbiont was a possible management technique for the stink bug. I tested the efficacy of a number of different insecticidal and antimicrobial products to determine whether prevention of symbiont acquisition from the egg mass was possible, and results indicated that transmission of the symbiont from the egg mass to the newly hatched nymph was negatively impacted when certain products were applied (namely surfactants or products containing surfactants). Additionally, direct effects on hatch rate and survival were reported for certain products, namely the insect growth regulator azadirachtin, which suggests that nymphs can pick up residues from the egg mass surface while probing for the symbiont. I conclude that P. carbekii plays a critically important role in the survival of its host, the brown marmorated stink bug, and its presence on the egg mass surface before nymphal hatch makes it targetable as a potential management technique.

Ultra high pressure homogenization (UHPH) inactivation of Bacillus amyloliquefaciens spores in phosphate buffered saline (PBS) and milk

Ultra high pressure homogenization (UHPH) opens up new areas for dynamic high pressure assisted thermal sterilization of liquids. Bacillus amyloliquefaciens spores are resistant to high isostatic pressure and temperature and were suggested as potential surrogate for high pressure thermal sterilization validation. B. amyloliquefaciens spores suspended in PBS buffer (0.01 M, pH 7.0), low fat milk (1.5%, pH 6.7), and whole milk (3.5%, pH 6.7) at initial concentration of similar to 10(6) CFU/mL were subjected to UHPH treatments at 200, 300, and 350 MPa with an inlet temperature at similar to 80 degrees C. Thermal inactivation kinetics of B. amyloliquefaciens spores in PBS and milk were assessed with thin wall glass capillaries and modeled using first-order and Weibull models. The residence time during UHPH treatments was estimated to determine the contribution of temperature to spore inactivation by UHPH. No sublethal injury was detected after UHPH treatments using sodium chloride as selective component in the nutrient agar medium. The inactivation profiles of spores in PBS buffer and milk were compared and fat provided no clear protective effect for spores against treatments. Treatment at 200 MPa with valve temperatures lower than 125 degrees C caused no reduction of spores. A reduction of 3.5 log(10)CFU/mL of B. amyloliquefaciens spores was achieved by treatment at 350 MPa with a valve temperature higher than 150 degrees C. The modeled thermal inactivation and observed inactivation during UHPH treatments suggest that temperature could be the main lethal effect driving inactivation.

Production of bioplastics from hydrolysates of paper industry by Haloferax mediterranei

The biorefinery concept has attracted much attention over the last decade due to increasing concerns about the use of fossil resources. In this context emerged the use of bioplastics, namely polyhydroxyalkanoates (PHA). PHA are biocompatible and biodegradable plastics that can be obtained from renewable raw materials and can constitute an alternative solution to conventional plastics. In this work, hydrolysed cellulose pulp, coming from Eucalyptus globulus wood cooking, was used as substrate to the PHA-storing bacteria Haloferax mediterranei. The hydrolysed pulp is rich in simple sugars, mainly glucose (81.96 g.L-1) and xylose (20.90 g.L-1). Tests were made in defined medium with glucose and xylose and in hydrolysate supplemented with salts and yeast extract. Different concentrations of glucose were tested, namely 10, 15, 20, 30 and 40 g.L-1. The best accumulation results (27.1 % of PHA) were obtained in hydrolysate medium with 10 g.L-1. Using this concentration, assays were performed in fed-batch and sequencing batch reactor conditions in order to determine the best feeding strategy. The strategy that led to the best results was fed-batch assay with 24.7 % of PHA. An assay without sterile conditions was performed, in which was obtained the same growth than in sterilization test. Finally it was performed an assay in a bioreactor and a fast growth (0.14 h-1) with high glucose and xylose consumption rates (0.368 g.L-1.h-1 and 0.0947 g.L-1.h-1, respectively) were obtained. However 1.50 g.L-1 of PHA, corresponding to 16.1 % (92.52 % of 3HB and 3HV of 7.48 %) of % PHA were observed. The polymer was further characterized by DSC with a glass transition temperature of -6.07 °C, a melting temperature of 156.3 °C and a melting enthalpy of 63.07 J.g-1, values that are in accordance with the literature. This work recognizes for the first time the suitability of the pulp paper hydrolysate as a substrate for PHA production by H. mediterranei.

MANICURE/PEDICURE TECHNICIANS: BIOSECURITY KNOWLEDGE AND PRACTICES FOR VIRAL HEPATITIS

Objetivo: Investigar o conhecimento e as práticas de biossegurança para hepatites virais de manicures/pedicures. Métodos: Estudo descritivo, transversal, quantitativo, através de questionário, utilizando instrumento de coleta de dados autoaplicado elaborado pelos pesquisadores, contendo dados da população (sexo, idade, tempo de atuação profissional) e conhecimentos básicos sobre transmissão de hepatite e práticas de biossegurança e higiene. Resultados: Entrevistaram-se 96 manicures/pedicures que atuam no Noroeste do Paraná. A maioria das profissionais já ouviu falar da patologia, mas somente 41,7% (n=40) fizeram o exame para detecção do vírus da hepatite; 38,39% (n=77) relataram como via de transmissão o sangue e 31,8% (n=63), a relação sexual. A reutilização de materiais descartáveis foi relatada por 60,4% (n=58); 55,2% (n=53) realizam esterilização de materiais e 27,1% (n=26) não a realizam. Não ficou evidenciada associação significativa entre tempo de profissão e as variáveis utilizadas: ouviu sobre hepatite (p=0,77025), realização de exames (p=0,035476), reutilização de materiais descartáveis (p=0,42691), lavagem de mãos (p=0,32876), uso de luvas descartáveis (p=0,33752) e esterilização de materiais (p=0,84443). Conclusão: As manicures entrevistadas não conhecem as exigências da Vigilância Sanitária no que concerne à prevenção da transmissão de hepatites.

A New Journal in the Field of Pharmaceutical Technology Is Born

International audience

Hidrógeis de Poli(N-Vinil 2-Pirrolidona) com nano argilas laponite RD sinterizados por irradiação gama

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Développement et validation d’une méthode de dosage des traces de détergents inactivants totaux du prion

International audience

Stability and Local Toxicity Evaluation of a Liposomal Prilocaine Formulation

This study reports a physicochemical stability evaluation of a previously reported liposomal prilocaine (PLC(LUV)) formulation (Cereda el al. J. Pharm. Pharmaceut. Sci. 7:235, 2004) before and after steam sterilization as well as its local toxicity evaluation. Prilocaine (PLC) was encapsulated into extruded unilamellar liposomes (LUVs) composed by egg phosphatidylcholine:cholesterol:alfa-tocopherol (4:3:0.07, mole %). Laser light-scattering analysis (p > 0.05) and thiobarbituric acid reaction (p > 0.05) were used to evaluate the liposomes physical (size) and chemical (oxidation) stability, respectively. The prilocaine chemical stability was followed by (1)H-nuclear magnetic resonance. These tests detected no differences on the physicochemical stability of PLC or PLCLUV, sterilized or not, up to 30 days after preparation (p > 0.05). Finally, the paw edema test and histological analysis of rat oral mucosa were used to assess the possible inflammatory effects of PLC(LUV). PLC(LUV) did not evoke rat paw edema (p > 0.05), and no significant differences were found in histological analysis, when compared to the control groups (p > 0.05). The present work shows that PLC(LUV) is stable for a 30-day period and did not induce significant inflammatory effects both in the paw edema test and in histological analysis, giving supporting evidence for its safely and possible clinical use in dentistry.

Efecto de la extirpación de las espículas del glande del cuy como técnia de esterilización reproductiva y su influencia en agresividad y ganancia de peso en comparación con un método químico alcohol yodado al 2%

La presente investigación evaluó el efecto de la extirpación de las espículas del glande en cobayos como método de esterilización reproductiva y su influencia en agresividad y ganancia de peso en comparación con un método químico (alcohol yodado 2%) y un testigo. Se contemplaron dos etapas de estudio, la primera valoró agresividad y ganancia de peso en 90 cuyes machos de 45 días de edad con peso promedio de 658,3±8,54gr los cuales fueron distribuidos en grupos de 5 animales por jaula; para la segunda etapa se consideró fertilidad y prolificidad en 90 cuyes hembras sexualmente maduras de 3 meses de edad, las cuales fueron empadradas con 18 machos obtenidos al azar de una sub-muestra de la primera etapa, la relación de empadre fue cinco hembras un macho. Los animales en estudio fueron distribuidos en los siguientes tratamientos: T1 animales enteros (n=30), T2 animales extirpados las espículas (n=30) y T3 animales castrados con alcohol yodado 2% (n=30). Los datos obtenidos fueron procesados y analizados en el programa SPSS® versión 22.0 determinándose que la ganancia total de peso fue para T1 836,4±33,89gr, T2 860,5±24,54gr y T3 725,5±30,45 con significancia estadística (P<0,05) para T2 y T1 con relación a T3. En referencia a agresividad medida a través del daño de la canal a nivel de la zona dorso posterior se obtuvieron resultados porcentuales estadísticos no significativos (p>0,05) entre tratamientos. En cuanto a fertilidad se obtuvo: para T1 un valor de 66,7%, T2 86,7% y T3 12% existiendo significancia estadística (p<0,05) entre tratamientos. Para la variable prolificidad se obtuvo valores medios de: T1 2,47±0,34; T2 3,43±0,32 y T3 0,40±0.22, con diferencias significativas (p<0,05) únicamente para el T3 que fue el que menos crías obtuvo. En conclusión la extirpación de las espículas del glande del cuy no esteriliza reproductivamente por el contrario aumenta el porcentaje de fertilidad, no disminuye agresividad, obtiene pesos no distintos al testigo pero alcanza mejores pesos que el método químico.

Dimensional changes of impression materials after 6 months storage period

Poster presented at the 24th Annual Meeting of the Portuguese Dental Association, 12-14 November 2015 Meo Arena, Lisboa, Portugal.