Engineering design of a bioprocess for the production of natural red colourants by submerged fermentation of the thermophilic fungus Penicillium purpurogenum GH2

The development of a new bioprocess requires several steps from initial concept to a practical and feasible application. Industrial applications of fungal pigments will depend on: (i) safety of consumption, (ii) stability of the pigments to the food processing conditions required by the products where they will be incorporated and (iii) high production yields so that production costs are reasonable. Of these requirements the first involves the highest research costs and the practical application of this type of processes may face several hurdles until final regulatory approval as a new food ingredient. Therefore, before going through expensive research to have them accepted as new products, the process potential should be assessed early on, and this brings forward pigment stability studies and process optimisation goals. Only ingredients that are usable in economically feasible conditions should progress to regulatory approval. This thesis covers these two aspects, stability and process optimisation, for a potential new ingredient; natural red colour, produced by microbial fermentation. The main goal was to design, optimise and scale-up the production process of red pigments by Penicillium purpurogenum GH2. The approach followed to reach this objective was first to establish that pigments produced by Penicillium purpurogenum GH2 are sufficiently stable under different processing conditions (thermal and non-thermal) that can be found in food and textile industries. Once defined that pigments were stable enough, the work progressed towards process optimisation, aiming for the highest productivity using submerged fermentation as production culture. Optimum production conditions defined at flask scale were used to scale up the pigment production process to a pilot reactor scale. Finally, the potential applications of the pigments were assessed. Based on this sequence of specific targets, the thesis was structured in six parts, containing a total of nine chapters. Engineering design of a bioprocess for the production of natural red colourants by submerged fermentation of the thermophilic fungus Penicillium purpurogenum GH2.

Attenuation of inflammatory events in human intervertebral disc cells with a tumor necrosis factor antagonist.

STUDY DESIGN: The inflammatory responses of primary human intervertebral disc (IVD) cells to tumor necrosis factor α (TNF-α) and an antagonist were evaluated in vitro. OBJECTIVE: To investigate an ability for soluble TNF receptor type II (sTNFRII) to antagonize TNF-α-induced inflammatory events in primary human IVD cells in vitro. SUMMARY OF BACKGROUND DATA: TNF-α is a known mediator of inflammation and pain associated with radiculopathy and IVD degeneration. sTNFRs and their analogues are of interest for the clinical treatment of these IVD pathologies, although information on the effects of sTNFR on human IVD cells remains unknown. METHODS: IVD cells were isolated from surgical tissues procured from 15 patients and cultured with or without 1.4 nmol/L TNF-α (25 ng/mL). Treatment groups were coincubated with varying doses of sTNFRII (12.5-100 nmol/L). Nitric oxide (NO), prostaglandin E₂ (PGE₂), and interleukin-6 (IL6) levels in media were quantified to characterize the inflammatory phenotype of the IVD cells. RESULTS: Across all patients, TNF-α induced large, statistically significant increases in NO, PGE₂, and IL6 secretion from IVD cells compared with controls (60-, 112-, and 4-fold increases, respectively; P < 0.0001). Coincubation of TNF-α with nanomolar doses of sTNFRII significantly attenuated the secretion of NO and PGE₂ in a dose-dependent manner, whereas IL6 levels were unchanged. Mean IC₅₀ values for NO and PGE₂ were found to be 35.1 and 20.5 nmol/L, respectively. CONCLUSION: Nanomolar concentrations of sTNFRII were able to significantly attenuate the effects of TNF-α on primary human IVD cells in vitro. These results suggest this sTNFR to be a potent TNF antagonist with potential to attenuate inflammation in IVD pathology.

Phosphorus export from a restored wetland ecosystem in response to natural and experimental hydrologic fluctuations

Wetland restoration is a commonly used approach to reduce nutrient loading to freshwater and coastal ecosystems, with many wetland restoration efforts occurring in former agricultural fields. Restored wetlands are expected to be effective at retaining or removing both nitrogen and phosphorus (P), yet restoring wetland hydrology to former agricultural fields can lead to the release of legacy fertilizer P. Here, we examined P cycling and export following rewetting of the Timberlake Restoration Project, a 440 ha restored riverine wetland complex in the coastal plain of North Carolina. We also compared P cycling within the restored wetland to two minimally disturbed nearby wetlands and an adjacent active agricultural field. In the restored wetland we observed increased soluble reactive phosphorus (SRP) concentrations following initial flooding, consistent with our expectations that P bound to iron would be released under reducing conditions. SRP concentrations in spring were 2.5 times higher leaving the restored wetland than a forested wetland and an agricultural field. During two large-scale drawdown and rewetting experiments we decreased the water depth by 1 m in ∼10 ha of inundated wetland for 2 weeks, followed by reflooding. Rewetting following experimental drainage had no effect on SRP concentrations in winter, but SRP concentrations did increase when the experiment was repeated during summer. Our best estimates suggest that this restored wetland could release legacy fertilizer P for up to a decade following hydrologic restoration. The time lag between restoration and biogeochemical recovery should be incorporated into management strategies of restored wetlands. Copyright 2010 by the American Geophysical Union.

Natural streamflow regime alterations: Damming of the Piave river basin (Italy)

A novel approach is proposed to estimate the natural streamflow regime of a river and to assess the extent of the alterations induced by dam operation related to anthropogenic (e.g., agricultural, hydropower) water uses in engineered river basins. The method consists in the comparison between the seasonal probability density function (pdf) of observed streamflows and the purportedly natural streamflow pdf obtained by a recently proposed and validated probabilistic model. The model employs a minimum of landscape and climate parameters and unequivocally separates the effects of anthropogenic regulations from those produced by hydroclimatic fluctuations. The approach is applied to evaluate the extent of the alterations of intra-annual streamflow variability in a highly engineered alpine catchment of north-eastern Italy, the Piave river. Streamflows observed downstream of the regulation devices in the Piave catchment are found to exhibit smaller means/modes, larger coefficients of variation, and more pronounced peaks than the flows that would be observed in the absence of anthropogenic regulation, suggesting that the anthropogenic disturbance leads to remarkable reductions of river flows, with an increase of the streamflow variability and of the frequency of preferential states far from the mean. Some structural limitations of management approaches based on minimum streamflow requirements (widely used to guide water policies) as opposed to criteria based on whole distributions are also discussed. Copyright © 2010 by the American Geophysical Union.

Analysis of the genome and transcriptome of Cryptococcus neoformans var. grubii reveals complex RNA expression and microevolution leading to virulence attenuation.

Cryptococcus neoformans is a pathogenic basidiomycetous yeast responsible for more than 600,000 deaths each year. It occurs as two serotypes (A and D) representing two varieties (i.e. grubii and neoformans, respectively). Here, we sequenced the genome and performed an RNA-Seq-based analysis of the C. neoformans var. grubii transcriptome structure. We determined the chromosomal locations, analyzed the sequence/structural features of the centromeres, and identified origins of replication. The genome was annotated based on automated and manual curation. More than 40,000 introns populating more than 99% of the expressed genes were identified. Although most of these introns are located in the coding DNA sequences (CDS), over 2,000 introns in the untranslated regions (UTRs) were also identified. Poly(A)-containing reads were employed to locate the polyadenylation sites of more than 80% of the genes. Examination of the sequences around these sites revealed a new poly(A)-site-associated motif (AUGHAH). In addition, 1,197 miscRNAs were identified. These miscRNAs can be spliced and/or polyadenylated, but do not appear to have obvious coding capacities. Finally, this genome sequence enabled a comparative analysis of strain H99 variants obtained after laboratory passage. The spectrum of mutations identified provides insights into the genetics underlying the micro-evolution of a laboratory strain, and identifies mutations involved in stress responses, mating efficiency, and virulence.

The potential of natural, photosynthetic pigments to improve the efficiency of dye-sensitized solar cells

Gemstone Team Grenergy

Natural hybridization between genera that diverged from each other approximately 60 million years ago.

A fern from the French Pyrenees-×Cystocarpium roskamianum-is a recently formed intergeneric hybrid between parental lineages that diverged from each other approximately 60 million years ago (mya; 95% highest posterior density: 40.2-76.2 mya). This is an extraordinarily deep hybridization event, roughly akin to an elephant hybridizing with a manatee or a human with a lemur. In the context of other reported deep hybrids, this finding suggests that populations of ferns, and other plants with abiotically mediated fertilization, may evolve reproductive incompatibilities more slowly, perhaps because they lack many of the premating isolation mechanisms that characterize most other groups of organisms. This conclusion implies that major features of Earth's biodiversity-such as the relatively small number of species of ferns compared to those of angiosperms-may be, in part, an indirect by-product of this slower "speciation clock" rather than a direct consequence of adaptive innovations by the more diverse lineages.

Temporal Stability of Molecular Diversity Measures in Natural Populations of Drosophila pseudoobscura and Drosophila persimilis

Many molecular ecological and evolutionary studies sample wild populations at a single point in time, failing to consider that data they collect represents genetic variation from a potentially unrepresentative snapshot in time. Variation across time in genetic parameters may occur quickly in species that produce multiple generations of offspring per year. However, many studies of rapid contemporary microevolution examine phenotypic trait divergence as opposed to molecular evolutionary divergence. Here, we compare genetic diversity in wild caught populations of Drosophila persimilis and D. pseudoobscura collected 16 years apart at the same time of year and same site at four X-linked and two mitochondrial loci to assess genetic stability. We found no major changes in nucleotide diversity in either species, but we observed a drastic shift in Tajima’s D between D. pseudoobscura timepoints at one locus associated with the increased abundance of a set of related haplotypes. Our data also suggests that D. persimilis may have recently accelerated its demographic expansion. While the changes we observed were modest, this study reinforces the importance of considering potential temporal variation in genetic parameters within single populations over short evolutionary timescales.

Conversion of natural forests to managed forest plantations decreases tree resistance to prolonged droughts

© 2015 Published by Elsevier B.V.Throughout the southern US, past forest management practices have replaced large areas of native forests with loblolly pine plantations and have resulted in changes in forest response to extreme weather conditions. However, uncertainty remains about the response of planted versus natural species to drought across the geographical range of these forests. Taking advantage of a cluster of unmanaged stands (85-130year-old hardwoods) and managed plantations (17-20year-old loblolly pine) in coastal and Piedmont areas of North Carolina, tree water use, cavitation resistance, whole-tree hydraulic (Ktree) and stomatal (Gs) conductances were measured in four sites covering representative forests growing in the region. We also used a hydraulic model to predict the resilience of those sites to extreme soil drying. Our objectives were to determine: (1) if Ktree and stomatal regulation in response to atmospheric and soil droughts differ between species and sites; (2) how ecosystem type, through tree water use, resistance to cavitation and rooting profiles, affects the water uptake limit that can be reached under drought; and (3) the influence of stand species composition on critical transpiration that sets a functional water uptake limit under drought conditions. The results show that across sites, water stress affected the coordination between Ktree and Gs. As soil water content dropped below 20% relative extractable water, Ktree declined faster and thus explained the decrease in Gs and in its sensitivity to vapor pressure deficit. Compared to branches, the capability of roots to resist high xylem tension has a great impact on tree-level water use and ultimately had important implications for pine plantations resistance to future summer droughts. Model simulations revealed that the decline in Ktree due to xylem cavitation aggravated the effects of soil drying on tree transpiration. The critical transpiration rate (Ecrit), which corresponds to the maximum rate at which transpiration begins to level off to prevent irreversible hydraulic failure, was higher in managed forest plantations than in their unmanaged counterparts. However, even with this higher Ecrit, the pine plantations operated very close to their critical leaf water potentials (i.e. to their permissible water potentials without total hydraulic failure), suggesting that intensively managed plantations are more drought-sensitive and can withstand less severe drought than natural forests.

Respuestas fenológicas de dos gramíneas C3 y dos C4, del pastizal natural de San Luis (Argentina), a variaciones de agua y temperatura

La temperatura y las precipitaciones son factores claves en la fenología de especies vegetales y su productividad. Los pastizales de la provincia de San Luis son especialmente interesantes porque coexisten gramíneas con vías metabólicas C3 (invernales) y C4 (estivales). Para evaluar cómo se relacionan la producción y la fenología reproductiva de las especies frente a cambios de temperatura y disponibilidad de agua, evalué cuatro gramíneas representativas de la región (dos C3 y dos C4) en dos estudios: 1) observacional a campo, y 2) manipulativo en invernáculo con dos niveles de temperatura y dos niveles de agua. En el estudio observacional comparé diez años de relevamientos fenológicos históricos realizados entre 1976 y 1986, con relevamientos fenológicos propios entre 2008 y 2010, todos llevados a cabo sobre el mismo sitio de estudio. En esta comparación encontré que una de las cuatro especies (Poa ligularis), atrasó significativamente su ciclo reproductivo (38 días la floración y 16 días la diseminación de semillas). La temperatura tuvo menor efecto sobre la producción, pero reflejó un claro control sobre la fenología en las cuatro especies estudiadas. En general, altas temperaturas estivales atrasaron el fin y aumentaron el largo del ciclo reproductivo de todas las especies. En cambio, primaveras más cálidas adelantaron el inicio reproductivo de las C4, pero retrasaron el de las C3. Por otra parte, la mayor disponibilidad de agua incrementó la producción, adelantó el comienzo floral y extendió entre 15 y 30 días el ciclo reproductivo de las cuatro especies. Inviernos lluviosos adelantaron el inicio floral de las especies C3, mientras que años con elevadas precipitaciones estivales retrasaron y extendieron el ciclo reproductivo de las C4. Estos resultados aportan valiosa información sobre las respuestas de la vegetación al clima, y pueden servir de insumo en el diseño de estrategias de manejo sustentable de estos pastizales.

Plan de marketing para la inserción de una bebida natural a base de yacón con piña, en el mercado de Lima, Perú

El consumo de productos naturales en el mundo es cada vez más creciente, y dentro de ello las bebidas funcionales que desde sus orígenes han tenido una performance relativamente satisfactoria. El yacón (Smallanthus sochifolius)es una planta perene, herbácea, que crece en toda la serranía peruana. Su demanda en el mercado exterior ha hecho que cada vez tenga mayor importancia. En el 2014 se exportó USD 2 899.398,00 en forma de polvo, jarabe, deshidratado; sin embargo su consumo en el mercado nacional es bajo, a pesar del gran potencial nutritivo que posee. El mercado de Lima moderna se presenta como un importante atractivo de mercado que podría consumir la bebida. Por ello el objetivo del presente trabajo es Realizar un Plan de Marketing para el mercado de Lima moderna a fin de evaluar la factibilidad de insertar una bebida nueva: yacón con piña, para fomentar el consumo interno de este producto. La metodología utilizada fue de carácter exploratorio, descriptivo cuantitativa y cualitativa mediante información primaria y secundaria que ayudó a su desarrollo. Dentro de la información primaria se utilizó como herramienta la encuesta y la información secundaria a través de informes y estudios que se hicieron para el sector de bebidas no alcohólicas. El sector de bebidas no alcohólicas en el Perú, muestra un importante crecimiento en lo referente a bebidas saludables. En el último año (2014) el crecimiento de este sector fue de 6 por ciento, motivados por la percepción del consumidor en los beneficios saludables de estas bebidas. La zona de Lima moderna acoge al 72,2 por ciento del nivel socio económico A y B, este segmento concentra el más alto promedio de ingresos. En el análisis de Porter, se describe a la empresa como una industria, el cual para este rubro las barreras de ingreso son altas por la inversión en activos físicos y la inversión en publicidad y promoción por ser un producto nuevo. A través del estudio realizado se encontró que existe demanda para este producto, el 31,1 por ciento de los encuestados compraría definitivamente, 25,5 por ciento una vez por semana y el 18,3 por ciento todos los días. La estrategia utilizada en el presente trabajo fue la diferenciación, por las características saludables de la bebida y única en el mercado. La bebida es un producto de introducción en el mercado, el precio fijado es alto porque se orienta a un segmento de mercado selectivo y además fijado en la imagen y calidad del producto. La distribución se realizará a través de los supermercados, minimarkets, hoteles categorizados. Y por último se concluye que la introducción de un producto novedoso, podría marcar estándares en el mercado de bebidas no alcohólicas y permitir la creación de una nueva categoría de productos Premium naturales y funcionales.

Fertilización o reemplazo de especies : revisión bibliográfica de los efectos sobre la productividad del pastizal natural

Los pastizales son los ecosistemas terrestres más extensos que existen en el mundo y se estima que ocupan un área de 52,2 millones de kilómetros cuadrados. Su productividad primaria neta aérea (PPNA) es una propiedad fundamental ya que determina la cantidad de energía que se encuentra disponible para otros niveles tróficos y está controlada por la calidad del suelo y la cantidad el agua. Suele intervenirse el pastizal para lograr diversos objetivos, tales como, aumentar y concentrar o distribuir la PPNA durante el año y aumentar la calidad forrajera de las especies que lo componen. Este trabajo tiene por objetivo responder a dos cuestiones inherentes a la productividad primaria neta de los pastizales: la primera es evaluar el impacto de la fertilización sobre la producción de los pastizales con Nitrógeno (N), Fósforo (P) y ambos (N+P). Y la segunda es evaluar el efecto sobre la producción de biomasa aérea del reemplazo de las comunidades originales del pastizal por una o varias especies forrajeras, las cuales pueden ser exóticas o naturalizadas. Para conseguir estos objetivos se realizó una revisión bibliográfica y un análisis de los datos cuantitativos publicados en diferentes revistas científicas y técnicas con y sin referato. La productividad de los tratamientos de corrección de la fertilidad edáfica con N; N+P y P se diferenciaron estadísticamente (p menor a 0,0001) de la producción del pastizal natural. Al analizar, en promedio, el incremento de biomasa producido se destaca en primer lugar el tratamiento N+P con 1825 kg MS/ha/año, le sigue el tratamiento con N con 768 kg MS/ha/año y en último lugar es el tratamiento P con 531 kg MS/ha/año. En el rango de precipitación entre los 800 y 1.140 mm, similar a los de la región pampeana, la PPNA de las pasturas implantadas se diferenció estadísticamente, (p menor a 0,0001), mediante una prueba t, de la PPNA de los pastizales. La media de producción de los pastizales fue de 3.933 kg MS/ha/año y de las pasturas fue de 5.763 kg MS/ha/año. En la mayoría de los ensayos la productividad de la pastura bajo la misma precipitación fue superior a la del pastizal.

Fermento natural de leche: efecto de su empleo sobre la calidad de quesos de leche de oveja

La fabricación de queso es esencialmente un proceso de deshidratación de la leche cuyo resultado final depende del tiempo de coagulación, tipo de fermento y período de maduración. La combinación de estos factores da lugar a una gran variedad de quesos, los que resultan de las diferentes técnicas empleadas en relación a los aspectos mencionados. El fermento es un cultivo de microorganismos cuyo crecimiento en la leche produce ácido láctico a partir de lactosa, acidificando el medio. Además durante la maduración del queso las enzimas bacterianas del inóculo utilizado intervendrán degradando los diferentes componentes de la leche liberando precursores de sustancias responsables de la textura, sabor y aroma del queso. Los fermentos utilizados pueden ser naturales preparados artesanalmente o comerciales seleccionados elaborados industrialmente. Se denominan naturales a aquellos elaborados a partir de la leche ordeñada y cobran especial interés cuando los volúmenes a procesar son pequeños. Su principal ventaja es que permiten aprovechar la ecología zonal bacteriana en la producción de un queso con características particulares, que puede reconocerse por su origen y que presenta características únicas eirrepetibles. El objetivo de este trabajo es evaluar la aptitud quesera de un fermento natural elaborado a partir de la leche de oveja cruza Frisona x Texel proveniente del tambo ovino de la Faculta de Agronomía de la Universidad de Buenos Aires (FAUBA). Para ello se analizaron sólidos totales y características organolépticas de quesos elaborados con fermento natural y se las contrastó con las de quesos elaborados con fermentos seleccionados comerciales. Los resultados del análisis de sólidos totales en quesos no mostraron diferencias significativas entre tratamientos, sin embargo la evaluación sensorial indicó, entre los atributos más destacables, que los quesos elaborados con fermento de leche tuvieron mayor intensidad de olor a oveja, aroma a manteca y leche cocida, sabor más picante y textura más untuosa que los elaborados con fermento comercial. En conclusión estos resultados sugieren que los atributos descriptos dan cuenta de un producto de características sensoriales distintivas. Esto indica la influencia diferencial que tuvo el fermento natural utilizado sobre las características sensoriales de los quesos a través de las enzimas aportadas por las bacterias durante el proceso de maduración.

Ecosystem services : a travel through natural benefits

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