Spoilage yeasts in Patagonian winemaking: molecular and physiological features of Pichia guilliermondii indigenous isolates Levaduras contaminantes en vinos patagónicos: características moleculares y fisiológicas de los aislamientos indígenas de Picchia guilliermondii
C. A. Lopes
Full Text Available Yeasts belonging to the genus Dekkera/Brettanomyces, especially the species Dekkera bruxellensis, have long been associated with the production of volatile phenols responsible for off-flavour in wines. According to recent reports, the species Pichia guilliermondii could also produce these compounds at the initial stages of fermentation. Based on the abundance of P. guilliermondii in Patagonian winemaking, we decided to study the relevance of indigenous isolates belonging to this species as wine spoilage yeast. Twenty-three indigenous isolates obtained from grape surfaces and red wine musts were analyzed in their capacity to produce volatile phenols on grape must. The relationship between molecular Random Amplified Polymorphic DNA (RAPD and physiological (killer biotype patterns detected in indigenous populations of P. guilliermondii and volatile phenol production was also evaluated. Different production levels of 4-ethylphenol, 4-vinylguaiacol and 4-ethylguaiacol were detected among the isolates; however, the values were always lower than those produced by the D. bruxellensis reference strain in the same conditions. High levels of 4-vinylphenol were detected among P. guilliermondii indigenous isolates. The combined use of RAPD and killer biotype allowed us to identify the isolates producing the highest volatile phenol levels.Las levaduras del género Dekkera/Brettanomyces, sobre todo la especie Dekkera bruxellensis, siempre han sido asociadas con la producción de fenoles volátiles responsables de aromas desagradables en los vinos. Recientemente, se ha demostrado que la especie Pichia guilliermondii también es capaz de producir estos compuestos, particularmente durante las etapas iniciales de la fermentación. Dada la abundancia de P. guilliermondii en las bodegas de la Patagonia, se decidió evaluar la importancia de algunos aislamientos indígenas de esta especie como levaduras alterantes de vinos regionales. Se evaluó la capacidad de
Aug 24, 2011 ... preharvest spraying with P. guilliermondii does not influence the quality attributes of the fruit. In addition, the .... Protein content in enzyme extracts was estimated by using the. Bradford method ..... Use of honey bees and.
Aryuman, Phichayaphorn; Lertsiri, Sittiwat; Visessanguan, Wonnop; Niamsiri, Nuttawee; Bhumiratana, Amaret; Assavanig, Apinya
In this study, 34 yeast isolates were obtained from koji and moromi samples of Thai soy sauce fermentation. However, the most interesting yeast strain was isolated from the enriched 2 month-old (M2) moromi sample and identified as Meyerozyma (Pichia) guilliermondii EM2Y61. This strain is a salt-tolerant yeast that could tolerate up to 20% (w/v) NaCl and produce extracellular and cell-bound glutaminases. Interestingly, its glutaminases were more active in 18% (w/v) NaCl which is a salt concentration in moromi. The extracellular glutaminase's activity was found to be much higher than that of cell-bound glutaminase. The highest specific activity and stability of the extracellular glutaminase were found in 18% (w/v) NaCl at pH4.5 and 37°C. A challenge test by adding partially-purified extracellular glutaminase from M. guilliermondii EM2Y61 into 1 month-old (M1) moromi sample showed an increased conversion of L-glutamine to L-glutamic acid. This is the first report of glutaminase producing M. guilliermondii isolated from the moromi of Thai soy sauce fermentation. The results suggested the potential application of M. guilliermondii EM2Y61 as starter yeast culture to increase l-glutamic acid during soy sauce fermentation. Copyright © 2014 Elsevier B.V. All rights reserved.
Full Text Available Attachment of yeast cells or bacteria on fungal hyphae have been observed in various antagonisms between microorganisms. Physical interactions between yeast Pichia guilliermondii and postharvest fruit pathogen Penicillium expansum in culture were studied in detail using light and transmission electron microscope to give better understanding on their mode of antagonism. Both organisms were co-cultured for 24-hr on potato dextrose agar. Light microscopy observations on the co-culture showed that the yeast cells attached firmly on the fungal hyphae. This attachment was inhibited by several substances such as enzymes degrading protein (protease or trypsin, a respiration inhibitor (sodium azide, an acid (hydrochloric acid or an alkali (sodium hydroxide. Although autoclaved hyphae did not affect the attachment, but boiled enzymes and autoclaved yeast cells totally abolished the attachment. These evidences suggested that the attachment might be an active process mediated by certain protein from live yeast cells. Transmission electron micrographs on the ultrastructure of the co-culture revealed that the hyphae showed abnormalities in their structure and organelles, and a degree of obvious damage. Physical interactions observed in this study could be contributed to the mechanism of antagonism between P. guilliermondii and P. expansum.
Jensen, Susanne Langgård; Umiker, Nicole L.; Arneborg, Nils
Yeast isolates from commercial red wines were characterized with regards to tolerances to molecular SO2, ethanol, and temperature as well as synthesis of 4-ethyl-phenol/4-ethyl-guaiacol in grape juice or wine. Based on rDNA sequencing, nine of the 11 isolates belonged to Dekkera bruxellensis (B1a......, B1b, B2a, E1, F1a, F3, I1a, N2, and P2) while the other two were Candida pararugosa (Q2) and Pichia guilliermondii (Q3). Strains B1b, Q2, and Q3 were much more resistant to molecular SO2 in comparison to the other strains of Dekkera. These strains were inoculated (103-104 cfu/ml) along with lower...... populations of Saccharomyces (red grape juice and red wine incubated at two temperatures, 15 C and 21 C. Although Saccharomyces quickly dominated fermentations in grape juice, B1b and Q2 grew and eventually reached populations >105 cfu/ml. In wine, Q3 never entered logarithmic growth...
Full Text Available In previous studies it was shown that Pichia guilliermondii strain Z1, isolated from healthy Moroccan citrus Valencia-Late oranges, was effective against Penicillium italicum. Here the effectiveness of strain Z1 was assessed against Penicillium digitatum, the causal agent of green mould, under different temperature (5-25°C and relative humidity (RH (45-100% regimes for its reliable and largescale application in packinghouse. All main effects and interactions were significant (P80%. Its applications as a formulated product significantly reduced the incidence of infected fruit (55% and the percentage of infected wounds (47% compared to the only pathogen control treatment. However, disease control with formulated product was significantly less than that obtained with thiabendazole (30% or strain Z1 culturable cells (35%. These results highlight that strain Z1 is an effective biological control agent for control of green mould under varying environmental conditions, and control may be optimized by combining its use with other environmentally-safe post-harvest treatments or improved formulation.
Knoshaug, Eric P; Vidgren, Virve; Magalhães, Frederico; Jarvis, Eric E; Franden, Mary Ann; Zhang, Min; Singh, Arjun
Genes encoding L-arabinose transporters in Kluyveromyces marxianus and Pichia guilliermondii were identified by functional complementation of Saccharomyces cerevisiae whose growth on L-arabinose was dependent on a functioning L-arabinose transporter, or by screening a differential display library, respectively. These transporters also transport D-xylose and were designated KmAXT1 (arabinose-xylose transporter) and PgAXT1, respectively. Transport assays using L-arabinose showed that KmAxt1p has K(m) 263 mM and V(max) 57 nM/mg/min, and PgAxt1p has K(m) 0.13 mM and V(max) 18 nM/mg/min. Glucose, galactose and xylose significantly inhibit L-arabinose transport by both transporters. Transport assays using D-xylose showed that KmAxt1p has K(m) 27 mM and V(max) 3.8 nM/mg/min, and PgAxt1p has K(m) 65 mM and V(max) 8.7 nM/mg/min. Neither transporter is capable of recovering growth on glucose or galactose in a S. cerevisiae strain deleted for hexose and galactose transporters. Transport kinetics of S. cerevisiae Gal2p showed K(m) 371 mM and V(max) 341 nM/mg/min for L-arabinose, and K(m) 25 mM and V(max) 76 nM/mg/min for galactose. Due to the ability of Gal2p and these two newly characterized transporters to transport both L-arabinose and D-xylose, one scenario for the complete usage of biomass-derived pentose sugars would require only the low-affinity, high-throughput transporter Gal2p and one additional high-affinity general pentose transporter, rather than dedicated D-xylose or L-arabinose transporters. Additionally, alignment of these transporters with other characterized pentose transporters provides potential targets for substrate recognition engineering. Copyright © 2015 John Wiley & Sons, Ltd.
Romi, Wahengbam; Keisam, Santosh; Ahmed, Giasuddin; Jeyaram, Kumaraswamy
Meyerozyma guilliermondii (anamorph Candida guilliermondii) and Meyerozyma caribbica (anamorph Candida fermentati) are closely related species of the genetically heterogenous M. guilliermondii complex. Conventional phenotypic methods frequently misidentify the species within this complex and also with other species of the Saccharomycotina CTG clade. Even the long-established sequencing of large subunit (LSU) rRNA gene remains ambiguous. We also faced similar problem during identification of yeast isolates of M. guilliermondii complex from indigenous bamboo shoot fermentation in North East India. There is a need for development of reliable and accurate identification methods for these closely related species because of their increasing importance as emerging infectious yeasts and associated biotechnological attributes. We targeted the highly variable internal transcribed spacer (ITS) region (ITS1-5.8S-ITS2) and identified seven restriction enzymes through in silico analysis for differentiating M. guilliermondii from M. caribbica. Fifty five isolates of M. guilliermondii complex which could not be delineated into species-specific taxonomic ranks by API 20 C AUX and LSU rRNA gene D1/D2 sequencing were subjected to ITS-restriction fragment length polymorphism (ITS-RFLP) analysis. TaqI ITS-RFLP distinctly differentiated the isolates into M. guilliermondii (47 isolates) and M. caribbica (08 isolates) with reproducible species-specific patterns similar to the in silico prediction. The reliability of this method was validated by ITS1-5.8S-ITS2 sequencing, mitochondrial DNA RFLP and electrophoretic karyotyping. We herein described a reliable ITS-RFLP method for distinct differentiation of frequently misidentified M. guilliermondii from M. caribbica. Even though in silico analysis differentiated other closely related species of M. guilliermondii complex from the above two species, it is yet to be confirmed by in vitro analysis using reference strains. This method can be used
... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Candida guilliermondii. 173.160 Section 173.160... CONSUMPTION Enzyme Preparations and Microorganisms § 173.160 Candida guilliermondii. The food additive Candida... the following conditions: (a) The food additive is the enzyme system of the viable organism Candida...
Alessandro Comarú Pasqualotto
Full Text Available Candida guilliermondii is one of the components of human microbiota. This yeast has been infrequently associated with human infections, which may be related to its low pathogenicity. The aim of this study was to provide clinical and epidemiological data for patients infected with C. guilliermondii at Santa Casa Complexo Hospitalar, Brazil. From October 1997 to October 2003, C. guilliermondii was isolated from clinical samples from 11 patients. Three patients were excluded because the isolation of the yeast represented colonisation. Specimens from the eight patients included in the study corresponded to blood (n = 5, ascitis fluid (n = 2, and oesophagus biopsy (n = 1. Three patients (37.5% had major immunosuppressed conditions, including solid organ transplantation, AIDS, and leukaemia. Previous use of antibiotics occurred in 87.5%. Main invasive medical procedures were central venous catheter (50.0%, abdominal surgery (25.0%, and peritoneal dialysis (50.0%. No susceptibility data was obtained. Although risk factors for candidaemia were similar amongst patients infected by with C. guilliermondii or other Candida species, mortality associated with C. guilliermondii was significantly lower.Candida guilliermondii é um dos componentes da microbiota humana e infecções associadas com esta levedura têm sido incomuns, o que pode ser atribuído a sua baixa patogenicidade. O objetivo deste trabalho foi documentar aspectos clínico-epidemiológicos em pacientes que tiveram C. guilliermondii isolada a partir de amostras biológicas. O estudo foi conduzido na Santa Casa Complexo Hospitalar, Brasil. Durante outubro de 1997 e outubro de 2003, C. guilliermondii foi isolada de 11 pacientes, três dos quais foram excluídos por se apresentarem apenas colonizados. Espécimes clínicos corresponderam a sangue (n = 5, líquido de ascite (n = 2 e biópsia de esôfago (n = 1. Três pacientes eram imunodeprimidos, incluindo transplante de órgãos sólidos, SIDA e
Song, Young-Ran; Jeong, Do-Youn; Baik, Sang-Ho
This study deals with understanding the effects of salt reduction on both the physicochemical and microbiological properties of soy sauce fermentation and also the application of indigenous yeast starters to compensate for undesirable changes occurring in salt-reduced processes. Fermentation was tested in situ at a Korean commercial soy sauce processing unit. Salt reduction resulted in higher acidity as well as lower pH and contents of residual sugar and ethanol. Moreover, undesired flavor characteristics, due to a lack of distinctive compounds, was observed. In addition, putrefactive Staphylococcus and Enterococcus spp. were present only during salt-reduced fermentation. To control these adverse effects, a single or mixed culture of two indigenous yeasts, Torulaspora delbrueckii and Pichia guilliermondii, producing high ethanol and 3-methyl-1-butanol, respectively, were tested. Overall, all types of yeast applications inhibited undesirable bacterial growth despite salt reduction. Of the starter cultures tested, the mixed culture resulted in a balance of more complex and richer flavors with an identical flavor profile pattern to that obtained from high salt soy sauce. Hence, this strategy using functional yeast cultures offers a technological option to manufacture salt-reduced soy sauce while preserving its typical sensory characteristics without affecting safety. Copyright © 2015 Elsevier Ltd. All rights reserved.
Greppi, Anna; Krych, Lukasz; Costantini, Antonella
Phytate is known as a strong chelate of minerals causing their reduced uptake by the human intestine. Ninety-three yeast isolates from traditional African fermented food products, belonging to nine species (Pichia kudriavzevii, Saccharomyces cerevisiae, Clavispora lusitaniae, Kluyveromyces...... marxianus, Millerozyma farinosa, Candida glabrata, Wickerhamomyces anomalus, Hanseniaspora guilliermondii and Debaryomyces nepalensis) were screened for phytase production on solid and liquid media. 95% were able to grow in the presence of phytate as sole phosphate source, P. kudriavzevii being the best...
López-Linares, Juan Carlos; Romero, Inmaculada; Cara, Cristobal
This study evaluated the possibility of using rapeseed straw hemicellulosic hydrolysate as a fermentation medium for xylitol production. Two yeast strains, namely Debaryomyces hansenii and Candida guilliermondii, were used for this bioconversion process and their performance to convert xylose...
Qi, Kai; Xia, Xiao-Xia; Zhong, Jian-Jiang
Commercialization of lignocellulosic ethanol fermentation requires its high titer, but the reactive oxygen species (ROS) accumulation during the bioprocess damaged the cells and compromised this goal. To improve the cellular anti-oxidative activity during non-detoxified corncob residue hydrolysate fermentation, seed cells were prepared to possess a higher level of intracellular biotin pool (IBP), which facilitated the biosyntheses of catalase and porphyrin. As a result, the catalase activity increased by 1.3-folds compared to control while the ROS level reduced by 50%. Cell viability in high-IBP cells was 1.7-folds of control and the final ethanol titer increased from 31.2 to 41.8 g L(-1) in batch fermentation. The high-IBP cells were further used for repeated-batch fermentation in the non-detoxified lignocellulosic hydrolysate, and the highest titer and average productivity of ethanol reached 63.7 g L(-1) and 1.2 g L(-1)h(-1). The results were favorable to future industrial application of this lignocellulosic bioethanol process. Copyright © 2015 Elsevier Ltd. All rights reserved.
Anne Caroline Defranceschi Oliveira
Full Text Available Extracellular lipases from the endophytic yeast Candida guilliermondii isolated from castor leaves (Ricinus communis L. were produced using low-cost raw materials such as agro-industrial residues and applying them in the esterification of oleic acid for evaluating their potential use in biodiesel production. After partial purification using ammonium sulfate, the enzyme was characterized and presented higher activity (26.8 ± 1.5 U mL-1 in the presence of 5 mmol L-1 NaCl at 30 ºC and pH 6.5. The production through submerged fermentation was formerly performed in 150 mL erlenmeyer flasks and, once the enzyme production was verified, assays in a 14 L bioreactor were conducted, obtaining 18 ± 1.4 U mL-1. The produced enzyme was applied in the oleic acid esterification under different solvents: hexane, cyclohexane or cyclohexanone and different acid:alcohol molar ratios. Higher ester conversion rate (81% was obtained using hexane and the molar ratio of 1:9 was the best conditions using methanol. The results suggest the potential for development of endophytic yeast in the production of biocatalyst through submerged fermentation using agroindustrial residues as culture medium.
María J. Navarro-Arias
Full Text Available The fungal cell wall contains glycoproteins that interact with the host immune system. In the prominent pathogenic yeast Candida albicans, Pmr1 acts as a Golgi-resident ion pump that provides cofactors to mannosyltransferases, regulating the synthesis of mannans attached to glycoproteins. To gain insight into a putative conservation of such a crucial process within opportunistic yeasts, we were particularly interested in studying the role of the PMR1 homolog in a low-virulent species that rarely causes candidiasis, Candida guilliermondii. We disrupted C. guilliermondii PMR1 and found that loss of Pmr1 affected cell growth and morphology, biofilm formation, susceptibility to cell wall perturbing agents, mannan levels, and the wall composition and organization. Despite there was a significant increment in the amount of β1,3-glucan exposed at the wall surface, this positively influenced only the ability of the mutant to stimulate IL-10 production by human monocytes, suggesting that recognition of both mannan and β1,3-glucan, is required to stimulate strong levels of pro-inflammatory cytokines. Accordingly, our results indicate C. guilliermondii sensing by monocytes was critically dependent on the recognition of N-linked mannans and β1,3-glucan, as reported in other Candida species. In addition, chemical remotion of cell wall O-linked mannans was found to positively influence the recognition of C. guilliermondii by human monocytes, suggesting that O-linked mannans mask other cell wall components from immune cells. This observation contrasts with that reported in C. albicans. Finally, mice infected with C. guilliermondii pmr1 null mutant cells had significantly lower fungal burdens compared to animals challenged with the parental strain. Accordingly, the null mutant showed inability to kill larvae in the Galleria mellonella infection model. This study thus demonstrates that mannans are relevant for the C. guilliermondii-host interaction, with
Frajkin, G.Ya.; Pinyaskina, E.V.; Strakhovskaya, M.G.
Resistance of the Candida guilliermondii cells to ultraviolet radiation (290-320 nm, 400-750 nm) is studied. Presence of previously unknown photoinducible protective mechanism in yeasts, providing for increase in cell stability to mid-ultraviolet radiation, biologically most active in the solar radiation spectrum, is revealed. 9 refs.; 3 figs
Zhang, Qiang; Gong, Jin-Song; Dong, Ting-Ting; Liu, Ting-Ting; Li, Heng; Dou, Wen-Fang; Lu, Zhen-Ming; Shi, Jin-Song; Xu, Zheng-Hong
3-Hydroxypropionic acid (3-HP) is an important platform chemical in organic synthesis. Traditionally, 3-HP was produced by chemical methods and fermentation process. In this work, a novel enzymatic method was developed for green synthesis of 3-HP. A yeast strain harboring nitrile-hydrolyzing enzyme was newly isolated from environmental samples using 3-hydroxypropionitrile (3-HPN) as the sole nitrogen source. It was identified to be Meyerozyma guilliermondii CGMCC12935 by sequencing of the 18S ribosomal DNA and internal transcribed spacer, together with analysis of the morphology characteristics. The catalytic properties of M. guilliermondii CGMCC12935 resting cells were determined, and the optimum activity was achieved at 55 °C and pH 7.5. The enzyme showed broad substrate specificity towards nitriles, especially 3-HPN, aminoacetonitrile and 3-cyanopyridine. The presence of Ag + , Pb 2+ and excess substrate inhibited the enzyme activity, whereas 5% (v/v) ethyl acetate had a positive effect on the enzyme activity. M. guilliermondii CGMCC12935 resting cells by addition of 3% glucose could thoroughly hydrolyze 500 mM 3-HPN into 3-HP within 100 h and the maximal accumulative production of 3-HP reached 216.33 mM, which was over twofolds than the control group with no additional glucose. And this work would lay the foundation for biological production of 3-HP in industry.
... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Pichia pastoris dried yeast. 573.750 Section 573... Food Additive Listing § 573.750 Pichia pastoris dried yeast. (a) Identity. The food additive Pichia pastoris dried yeast may be used in feed formulations of broiler chickens as a source of protein not to...
Being homeless in one’s homeland is a colonial legacy for many Indigenous people in settler societies. The construction of Commonwealth nation-states from colonial settler societies depended on the dispossession of Indigenous peoples from their lands. The legacy of that dispossession and related...... attempts at assimilation that disrupted Indigenous practices, languages, and cultures—including patterns of housing and land use—can be seen today in the disproportionate number of Indigenous people affected by homelessness in both rural and urban settings. Essays in this collection explore the meaning...... and scope of Indigenous homelessness in the Canada, Australia, and New Zealand. They argue that effective policy and support programs aimed at relieving Indigenous homelessness must be rooted in Indigenous conceptions of home, land, and kinship, and cannot ignore the context of systemic inequality...
Thomas W. Jeffries; Jennifer R. Headman Van Vleet
Genome sequencing and subsequent global gene expression studies have advanced our understanding of the lignocellulose-fermenting yeast Pichia stipitis. These studies have provided an insight into its central carbon metabolism, and analysis of its genome has revealed numerous functional gene clusters and tandem repeats. Specialized physiological traits are often the...
Türkşen, Zeynep; Yağcı, Server; Karadağ, Ayşe Serap; Tezer, Ayla; Taner, Omer Faruk; Tekin, Fatih; Arıkan Akdağlı, Sevtap
Opportunistic fungal infections are usually seen in immunocompromised patients. While Candida is the most prevalent agent in such infections, Aspergillus is at the second order. Primary cutaneous aspergillosis is most common in immunocompromised patients but can rarely be seen in healthy hosts as well. We report a case of posttraumatic primary cutaneous aspergillosis and Candida guilliermondii coinfection in a 70-years-old healthy man. The patient had an ulcerous lesion which developed in the site of a trauma on the middle finger of the right hand. Histopathological examination of the biopsy specimens revealed septate hyphae with dichotomous branching small circular blastospores. The cultures of the biopsy specimen yielded yellow-green colored, granular mold colonies and creamy white yeast colonies. Microscopic examination of the lactophenol cotton blue stained mold colonies indicated long conidiophores with vesicles surrounded by uniseriate phialides, compatible with Aspergillus flavus. Yeast colonies were identified as Candida guilliermondii by ID32C (BioMerieux, France) and by their microscopical morphology detected in corn meal-Tween 80 agar incubated at 25°C for 72 hours. The patient was treated properly with surgical debridement and itraconazole therapy. Since the immune system is compressed as a consequence of aging, cutaneous opportunistic fungal infections should be considered in the differential diagnosis of posttraumatic necrotic ulcers and black eschar in aged patients.
Gun Woo Lee
Full Text Available Isolated primary candidal infection of joint is extremely rare, with only a few reported cases. It occurs as a result of accidental implantations of fungus during traumatic procedures, such as surgery, and is usually reported in patients with predisposing factors such as immunosuppression, malignancy, and drug abuse. If left untreated, irreversible deformity and pain with severe osteoarticular destruction occur. Thus, early diagnosis and treatment are important. This paper presents a case of 72-year-old man with primary C. guilliermondii infection of knee joint without predisposing factors and previous traumatic procedures, who was misdiagnosed with advanced degenerative osteoarthritis. Our case is the second case of primary C. guilliermondii arthritis of knee to be reported in the English-language literature and the first to be successfully treated with total knee arthroplasty following IV amphotericin B and oral fluconazole. Primary candidal infection of joint is generally asymptomatic or involves only mild pain and swelling in the affected knee. Thus, although the majority of knee joint infections are of a pyogenic or tuberculous origin, if a patient complains of mild pain and swelling in the knee and has mild signs of infection, the possibility of fungal infection should be considered.
Fraikin, G.Y.; Pospelov, M.E.; Rubin, L.B.
In experiments with the non-photoreactivable yeast Candida guilliermondii, radiations at 313, 334 and 365 nm, having no effect on untreated cell populations, produced an 'enhancing' effect on the lethality of 254 nm-pretreated cells. Wavelengths in the visible region of the spectrum did not exhibit a similar effect. (author)
Full Text Available Abstract Background Sustainable utilization of plant biomass as renewable source for fuels and chemical building blocks requires a complex mixture of diverse enzymes, including hydrolases which comprise the largest class of lignocellulolytic enzymes. These enzymes need to be available in large amounts at a low price to allow sustainable and economic biotechnological processes. Over the past years Pichia pastoris has become an attractive host for the cost-efficient production and engineering of heterologous (eukaryotic proteins due to several advantages. Results In this paper codon optimized genes and synthetic alcohol oxidase 1 promoter variants were used to generate Pichia pastoris strains which individually expressed cellobiohydrolase 1, cellobiohydrolase 2 and beta-mannanase from Trichoderma reesei and xylanase A from Thermomyces lanuginosus. For three of these enzymes we could develop strains capable of secreting gram quantities of enzyme per liter in fed-batch cultivations. Additionally, we compared our achieved yields of secreted enzymes and the corresponding activities to literature data. Conclusion In our experiments we could clearly show the importance of gene optimization and strain characterization for successfully improving secretion levels. We also present a basic guideline how to correctly interpret the interplay of promoter strength and gene dosage for a successful improvement of the secretory production of lignocellulolytic enzymes in Pichia pastoris.
Fraikin, G.Y.; Pospelov, M.E.; Rubin, L.B.
The present communication is concerned with the effects of near-UV radiation (300-380 nm) on yeast Candida guilliermondii. It was found that certain doses of 313 nm irradiation caused inactivation of the yeast which was exhibited in a way different from the lethal action of far-UV radiation. It was also found that the cells inactivated by 313 nm are capable of recovering vitality, if incubated for some time in a non-nutrient medium. The yeast inactivated by far-UV radiation also proved to be capable of recovering, though to a lesser degree. Both 334 nm radiation and non-lethal doses at 313 nm induced the photoprotective effect against far-UV damage. The effect was exhibited if there was a certain time interval (2-4 h) between the exposures to photoprotective light and subsequent far-UV radiation. Within this time interval the extent of photoprotection was dependent on temperature. (author)
Débora Danielle Virgínio da Silva
Full Text Available The effect of glucose on xylose-to-xylitol bioconversion by Candida guilliermondii was examined by adding it to sugarcane bagasse hydrolysate medium to obtain different glucose:xylose ratios (1:25, 1:12, 1:5 and 1:2.5. Under experimental conditions, increasing glucose:xylose ratio improved the assimilation of the xylose present in the hydrolysate by yeast, resulting in biomass increase, and in the formation of xylitol and glycerol/ethanol by-products. Maximum values of xylitol yield (0.59 g g-1 and volumetric productivity (0.53 g l-1.h-1 were obtained with glucose:xylose ratio of 1:5, resulting in the higher conversion efficiency (64.3%.O efeito da glicose na bioconversão de xilose em xilitol por Candida guilliermondii foi avaliado em hidrolisado hemicelulósico de bagaço de cana com diferentes relações glicose:xilose (1:25, 1:12, 1:5 and 1:2,5. Sob as condições experimentais, o aumento da relação glicose:xilose favoreceu a assimilação da xilose presente no hidrolisado, resultando em aumento da biomassa celular e aumento da formação de xilitol e dos sub-produtos glicerol e etanol. Os valores máximos do fator de conversão de xilose em xilitol (0,59 g g-1 e da produtividade volumétrica de xilitol (0,53 g l-1.h-1 foram obtidos com a relação glicose:xilose 1:5, resultando na maior eficiência de conversão (64,3%.
Marcela Villalba Cadavid
Full Text Available En este estudio se empleó cascarilla de arroz como materia prima para la obtención de xilitol, previa hidrólisis durante 60 minutos con ácido sulfúrico al 4% p/v; a 121 ºC y 15 psig,; La xilosa producida se transformó en xilitol mediante su fermentación con Candida guilliermondii. Se estudió el efecto de las variables concentración inicial de xilosa, concentración de inóculo y relación volumen del medio/volumen del matraz, así como sus efectos combinados, sobre la producción de xilitol. Se encontró que las concentraciones iniciales de xilosa e inóculo más adecuadas, entre los valores ensayados, fueron 80 y 5,0 g/l, respectivamente. En estas condiciones, la concentración final de xilitol obtenida fue de 45,2 g/l, con una productividad volumétrica de 0,23 g/l•h y un rendimiento de 0,57 g/g.In this study was used rice husk, previosly hydrolyzed with diluted sulfuric acid at 121 ºC and 15 psig, with a residence time 60 min. The initial concentration of substrate, inoculum, and relationship between media volume/flask volume and their combined effects were studied on the production of xylitol. The initial concentrations of 80 g/l xylose and 5 g/l inoculums led the best xylitol production (45.2 g/l, productivity (0.23 g/l•h and yield (0.57 g/g.
Reclassification of Candida guilliermondii FTI 20037 as Candida tropicalis based on molecular phylogenetic analysis Reclassificação de Candida guilliermondii FTI 20037 como Candida tropicalis baseada na análise filogenética molecular
Luanne Helena Augusto Lima
Full Text Available Yeasts of the genus Candida are of clinical importance and also have many industrial applications, mainly in the food industry. The yeast Candida guilliermondii FTI 20037 has been extensively studied in order to establish a biotechnological process for the production of xylitol. The goal of this study was to verify the taxonomic classification of this strain based on the analysis of rDNA sequences and the xyl1 gene. DNA fragments from these sequences were amplified by PCR and BLAST analysis revealed strong identity with the corresponding sequences from Candida tropicalis. Based on these results, we propose that C. guilliermondii FTI 20037 must be reclassified as C. tropicalis.As leveduras do gênero Candida possuem tanto importância clínica como diversas aplicações industriais, principalmente na indústria de alimentos. A levedura Candida guilliermondii FTI 20037 tem sido exaustivamente estudada pois pretende-se utilizá-la no estabelecimento de um processo biotecnológico para a produção de xilitol. O objetivo deste trabalho foi verificar a classificação taxonômica desta levedura por análise de sequências do rDNA e do gene xyl1. Fragmentos correspondentes a estas regiões foram amplificados por PCR e a análise destas sequências por BLAST revelou alta identidade com sequências correspondentes de Candida tropicalis. Estes resultados nos levam a propor que C. guilliermondii FTI 20037 deva ser reclassificada como C. tropicalis.
Geertz, Armin W.
Dette essay diskuterer en publikation af James L. Cox med titlen From Primitive to Indigenous (2007). Bogen analyserer forskellige forfatteres holdninger til studiet af indfødte kulturers religioner. Cox's analyser tages op i dette essay og de problematiseres i forhold til mit eget arbejde....
Ramírez-Castrillón, Mauricio; Jaramillo-Garcia, Victoria P; Rosa, Priscila D; Landell, Melissa F; Vu, Duong; Fabricio, Mariana F; Ayub, Marco A Z; Robert, Vincent; Henriques, João A P; Valente, Patricia
A high throughput screening (HTS) methodology for evaluation of cellular lipid content based on Nile red fluorescence reads using black background 96-wells test plates and a plate reader equipment allowed the rapid intracellular lipid estimation of strains from a Brazilian phylloplane yeast collection. A new oleaginous yeast, Meyerozyma guilliermondii BI281A, was selected, for which the gravimetric determination of total lipids relative to dry weight was 52.38% for glucose or 34.97% for pure glycerol. The lipid production was optimized obtaining 108 mg/L of neutral lipids using pure glycerol as carbon source, and the strain proved capable of accumulating oil using raw glycerol from a biodiesel refinery. The lipid profile showed monounsaturated fatty acids (MUFA) varying between 56 or 74% in pure or raw glycerol, respectively. M. guilliermondii BI281A bears potential as a new biodiesel feedstock.
Full Text Available A high throughput screening (HTS methodology for evaluation of cellular lipid content based on Nile red fluorescence reads using black background 96-wells test plates and a plate reader equipment allowed the rapid intracellular lipid estimation of strains from a Brazilian phylloplane yeast collection. A new oleaginous yeast, Meyerozyma guilliermondii BI281A, was selected, for which the gravimetric determination of total lipids relative to dry weight was 52.38% for glucose or 34.97% for pure glycerol. The lipid production was optimized obtaining 108 mg/L of neutral lipids using pure glycerol as carbon source, and the strain proved capable of accumulating oil using raw glycerol from a biodiesel refinery. The lipid profile showed monounsaturated fatty acids (MUFA varying between 56 or 74% in pure or raw glycerol, respectively. M. guilliermondii BI281A bears potential as a new biodiesel feedstock.
Solange Inês Mussatto
Full Text Available Xylitol was produced by Candida guilliermondii from charcoal-treated and untreated rice straw hemicellulosic hydrolysate with or without nutrients (ammonium sulphate, calcium chloride, rice bran extract. Both, xylitol yield and volumetric productivity decreased significantly when the nutrients were added to treated and untreated hydrolysates. In the treated hydrolysate, the efficiency of xylose conversion to xylitol was 79% when the nutrients were omitted. The results demonstrated that rice straw hemicellulosic hydrolysate treated with activated charcoal was a cheap source of xylose and other nutrients for xylitol production by C. guilliermondii. The non-necessity of adding nutrients to the hydrolysate media would be very advantageous since the process becomes less costly.Este trabalho avaliou a produção de xilitol pela levedura Candida guilliermondii, a partir de hidrolisado hemicelulósico de palha de arroz não tratado e tratado com carvão ativo, ambos suplementados ou não com nutrientes (sulfato de amônio, cloreto de cálcio e extrato de farelo de arroz. Os resultados mostraram que tanto o rendimento como a produtividade volumétrica em xilitol diminuíram quando os nutrientes foram adicionados em ambos hidrolisados, tratado e não tratado. Em hidrolisado tratado, a eficiência de conversão de xilose em xilitol foi de 79% quando em ausência de nutrientes. Estes resultados mostram que o hidrolisado hemicelulósico de palha de arroz tratado com carvão ativo é uma fonte barata de xilose e outros nutrientes, para a produção de xilitol por Candida guilliermondii. A não necessidade de adicionar nutrientes ao meio a base de hidrolisado é muito vantajosa, uma vez que o processo se torna mais econômico.
Schwan, Rosane F.; Almeida, Euziclei G.; Souza-Dias, Maria Aparecida G.
and peanuts. A fermentation using rice and cassava was conducted, and samples were collected at 4-h intervals for microbial analysis. The yeast population was low at the beginning of the fermentation and reached 6.9 x 10(7) CFU mL(-1) after 48 h. During the fermentation process common yeast species were......The Tapirapé people of the Tapi'itãwa tribe of Brazil produce several fermented foods and beverages, one of which is called 'cauim'. This beverage usually makes up the main staple food for adults and children. Several substrates are used in its production, including cassava, rice, corn, maize...... identified by sequencing of the D1/D2 domain of the large-subunit (26S) rRNA gene. The predominant yeast species found was Candida tropicalis. Candida intermedia, Candida parapsilosis, Pichia guilliermondii, Saccharomyces cerevisiae and Trichosporon asahii were also found in high numbers during...
Lage, Patrícia; Barbosa, Catarina; Mateus, Beatriz; Vasconcelos, Isabel; Mendes-Faia, Arlete; Mendes-Ferreira, Ana
Non-Saccharomyces yeasts include different species which comprise an ecologically and biochemically diverse group capable of altering fermentation dynamics and wine composition and flavour. In this study, single- and mixed-culture of Hanseniaspora guilliermondii and Saccharomyces cerevisiae were used to ferment natural grape-juice, under two nitrogen regimes. In single-culture the strain H. guilliermondii failed to complete total sugar breakdown even though the nitrogen available has not been a limiting factor of its growth or fermentative activity. In mixed-culture, that strain negatively interfered with the growth and fermentative performance of S. cerevisiae, resulting in lower fermentation rate and longer fermentation length, irrespective of the initial nitrogen concentration. The impact of co-inoculation on the volatile compounds profile was more evident in the wines obtained from DAP-supplemented musts, characterised by increased levels of ethyl and acetate esters, associated with fruity and floral character of wines. Moreover, the levels of fatty acids and sulphur compounds which are responsible for unpleasant odours that depreciate wine sensory quality were significantly lower. Accordingly, data obtained suggests that the strain H. guilliermondii has potential to be used as adjunct of S. cerevisiae in wine industry, although possible interactions with S. cerevisiae still need to be elucidated. Copyright © 2013 Elsevier B.V. All rights reserved.
F. da Cunha-Pereira
Full Text Available Abstract This work investigated the ability of a recently isolated strain of Candida guilliermondii to convert hexoses and pentoses obtained from acid-enzymatic soybean hull hydrolysates into ethanol and, in smaller amounts, into xylitol. Operational conditions and media formulation were optimized concerning ethanol production using experimental designs (Plackett-Burman and Central Composite Design. Results showed that C. guilliermondii BL 13 was capable of growing in non-supplemented, non-detoxified biomass hydrolysates, and the best culture conditions were determined to be 28 °C, pH 5.0, and 109 CFU mL-1 of inoculum size. Ethanol productivity reached 1.4 g L-1 h-1, and maximal yields of 0.41 g g-1 were obtained, representing 80.4 % of the expected theoretical yields, whereas small amounts of xylitol were also produced. These results suggest that C. guilliermondii BL13 is a potentially useful yeast strain to be applied in second-generation ethanol production from lignocellulosic biomass based on its natural capacity to metabolize C-5 and C-6 sugars.
Frajkin, G.Ya.; Pospelov, M.E.; Rubin, L.B.
The results of studies on the effect of near uv light on the yeast Candida guilliermondii are presented. It was shown that certain doses of 313 nm light inactivated the yeast. The detailed affect is shown in the loss of the ability of the cells to form microcolonies and outwardly does not differ from inactivation caused by 254 nm uv. It was concluded that the cell destruction caused by the 313 nm light was not due to damage to DNA. Experiments in which yeast cells were inactivated by 313 nm light before plating on agar and held for some time in a non-nutrient medium permitted observation of recovery of their viability. A difference was shown in the level of repair of yeasts irradiated by 313 nm light (up to 100% recovery) and 254 nm light (60% recovery). The nature of the dependence of the photoprotection on the 313 nm light dose was determined. A decrease in photoprotection was noted, starting with 7x10 -7 einstein/cm 2 , with its complete disappearance upon further dose increase. It is suggested that, in this recovery of the yeast, some other, thus far unknown, mechanism participates. Data were obtained on the survival of yeast irradiated with lethal uv doses. Of special importance, in the authors' opinion, is the fact that, for the photoprotection effect to appear, some time is needed between actions of the 313 and 254 nm lights, which suggests a photoinduced formation in the yeast cells of compounds that protect them from lethal injury
Kelly, Christine; Jones, Opal; Barnhart, Christopher; Lajoie, Curtis
Xylitol is a five-carbon sugar alcohol with established commercial use as an alternative sweetener and can be produced from hemicellulose hydrolysate. However, there are difficulties with microbiological growth and xylitol biosynthesis on hydrolysate because of the inhibitors formed from hydrolysis of hemicellulose. This research focused on the effect of furfural, vanillin, and syringaldehyde on growth of Candida guilliermondii and xylitol accumulation from xylose in a semi-synthetic medium in microwell plate and bioreactor cultivations. All three compounds reduced specific growth rate, increased lag time, and reduced xylitol production rate. In general, increasing concentration of inhibitor increased the severity of inhibition, except in the case of 0.5 g vanillin per liter, which resulted in a faster late batch phase growth rate and increased biomass yield. At concentrations of 1 g/1 or higher, furfural was the least inhibitory to growth, followed by syringaldehyde. Vanillin most severely reduced specific growth rate. All three inhibitors reduced xylitol production rate approximately to the same degree.
Greppi, Anna; Krych, Łukasz; Costantini, Antonella; Rantsiou, Kalliopi; Hounhouigan, D Joseph; Arneborg, Nils; Cocolin, Luca; Jespersen, Lene
Phytate is known as a strong chelate of minerals causing their reduced uptake by the human intestine. Ninety-three yeast isolates from traditional African fermented food products, belonging to nine species (Pichia kudriavzevii, Saccharomyces cerevisiae, Clavispora lusitaniae, Kluyveromyces marxianus, Millerozyma farinosa, Candida glabrata, Wickerhamomyces anomalus, Hanseniaspora guilliermondii and Debaryomyces nepalensis) were screened for phytase production on solid and liquid media. 95% were able to grow in the presence of phytate as sole phosphate source, P. kudriavzevii being the best growing species. A phytase coding gene of P. kudriavzevii (PHYPk) was identified and its expression was studied during growth by RT-qPCR. The expression level of PHYPk was significantly higher in phytate-medium, compared to phosphate-medium. In phytate-medium expression was seen in the lag phase. Significant differences in gene expression were detected among the strains as well as between the media. A correlation was found between the PHYPk expression and phytase extracellular activity. Copyright © 2015 Elsevier B.V. All rights reserved.
Young, Travis; Schultz, Peter G.
The invention provides orthogonal translation systems for the production of polypeptides comprising unnatural amino acids in methylotrophic yeast such as Pichia pastoris. Methods for producing polypeptides comprising unnatural amino acids in methylotrophic yeast such as Pichia pastoris are also provided.
Young, Travis [San Diego, CA; Schultz, Peter G [La Jolla, CA
The invention provides orthogonal translation systems for the production of polypeptides comprising unnatural amino acids in methyltrophic yeast such as Pichia pastoris. Methods for producing polypeptides comprising unnatural amino acids in methyltrophic yeast such as Pichia pastoris are also provided.
Saleh, Qasem; Kovács, Renátó; Kardos, Gábor; Gesztelyi, Rudolf; Kardos, Tamás; Bozó, Aliz; Majoros, László
Currently, echinocandins are first-line drugs for treatment of invasive candidiasis. However, data on how serum influences killing activity of echinocandins against uncommon Candida species are limited. Therefore, the killing activity of micafungin in RPMI-1640 and in 50% serum was compared against Candida guilliermondii, Candida lusitaniae, and Candida kefyr. Minimum inhibitory concentration (MIC) ranges in RPMI-1640 were 0.5-1, 0.12-0.25, and 0.06-0.12 mg/L, respectively. In 50% serum, MICs increased 32- to 256-fold. In RPMI-1640 ≥ 0.25, ≥4, and 32 mg/L micafungin was fungicidal against all four C. kefyr (≤4.04 hours), two of three C. lusitaniae (≤16.10 hours), and two of three C. guilliermondii (≤12.30 hours), respectively. In 50% serum, all three species grew at ≤4 mg/L. Micafungin at 16-32 mg/L was fungicidal against all C. kefyr isolates (≤3.03 hours) and at 32 mg/L was fungistatic against one of three C. lusitaniae isolates. Two C. lusitaniae isolates and all three C. guilliermondii grew at all tested concentrations. Adding human serum to susceptibility test media drew attention to loss of fungicidal or fungistatic activity of micafungin in the presence of serum proteins, which is not predicted by MICs in case of C. kefyr and C. lusitaniae in RPMI-1640. Our results strongly suggest that micafungin and probably other echinocandins should be used with caution against rare Candida species.
Hernández-Pérez, Andrés Felipe; de Arruda, Priscila Vaz; Felipe, Maria das Graças de Almeida
Sugarcane straw has become an available lignocellulosic biomass since the progressive introduction of the non-burning harvest in Brazil. Besides keeping this biomass in the field, it can be used as a feedstock in thermochemical or biochemical conversion processes. This makes feasible its incorporation in a biorefinery, whose economic profitability could be supported by integrated production of low-value biofuels and high-value chemicals, e.g., xylitol, which has important industrial and clinical applications. Herein, biotechnological production of xylitol is presented as a possible route for the valorization of sugarcane straw and its incorporation in a biorefinery. Nutritional supplementation of the sugarcane straw hemicellulosic hydrolyzate as a function of initial oxygen availability was studied in batch fermentation of Candida guilliermondii FTI 20037. The nutritional supplementation conditions evaluated were: no supplementation; supplementation with (NH4)2SO4, and full supplementation with (NH4)2SO4, rice bran extract and CaCl2·2H2O. Experiments were performed at pH 5.5, 30°C, 200rpm, for 48h in 125mL Erlenmeyer flasks containing either 25 or 50mL of medium in order to vary initial oxygen availability. Without supplementation, complete consumption of glucose and partial consumption of xylose were observed. In this condition the maximum xylitol yield (0.67gg(-1)) was obtained under reduced initial oxygen availability. Nutritional supplementation increased xylose consumption and xylitol production by up to 200% and 240%, respectively. The maximum xylitol volumetric productivity (0.34gL(-1)h(-1)) was reached at full supplementation and increased initial oxygen availability. The results demonstrated a combined effect of nutritional supplementation and initial oxygen availability on xylitol production from sugarcane straw hemicellulosic hydrolyzate. Copyright © 2016 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.
Bartolo-Aguilar, Yaneth; Dendooven, Luc; Chávez-Cabrera, Cipriano; Flores-Cotera, Luis B; Hidalgo-Lara, María E; Villa-Tanaca, Lourdes; Marsch, Rodolfo
The production of recombinant biopharmaceutical proteins is a multi-billion dollar market. Protein recovery represents a major part of the production costs. Pichia pastoris is one of the microbial systems most used for the production of heterologous proteins. The use of a cold-induced promoter to express lytic enzymes in the yeast after the growth stage could reduce protein recovery costs. This study shows that a cold-shock can be applied to induce lysis of the yeast cells. A strain of P. pastoris was constructed in which the endogenous eng gene encoding a putative endo-β-1,3-glucanase was overexpressed using the cold-shock induced promoter of the cctα gene from Saccharomyces cerevisiae. In the transgenic P. pastoris, the expression of eng increased 3.6-fold after chilling the cells from 30 to 4 °C (cold-shock stage) followed by incubation for 6 h (eng expression stage). The culture was heated to 30 °C for 6 h (ENG synthesis stage) and kept at 37 °C for 24 h (lysis stage). After this procedure the cell morphology changed, spheroplasts were obtained and cellular lysis was observed. Thus, a clone of P. pastoris was obtained, which undergoes autolysis after a cold-shock.
Fraikin, G.Y.; Strakhovskaya, M.G.; Rubin, L.B.
Mechanisms of near-UV (334 nm) induced photoprotection as well as potentiation of far-UV (254 nm) lethality are considered in Candida guilliermondii. Using exogenous precursors of serotonin, it appears that the above two mechanisms involve photoactivated synthesis of serotonin. It has been postulated that the serotonin effect could take place by binding to DNA. (author)
Navarro-Arias, María J; Dementhon, Karine; Defosse, Tatiana A; Foureau, Emilien; Courdavault, Vincent; Clastre, Marc; Le Gal, Solène; Nevez, Gilles; Le Govic, Yohann; Bouchara, Jean-Philippe; Giglioli-Guivarc'h, Nathalie; Noël, Thierry; Mora-Montes, Hector M; Papon, Nicolas
Hybrid histidine kinases (HHKs) progressively emerge as prominent sensing proteins in the fungal kingdom and as ideal targets for future therapeutics. The group X HHK is of major interest, since it was demonstrated to play an important role in stress adaptation, host-pathogen interactions and virulence in some yeast and mold models, and particularly Chk1, that corresponds to the sole group X HHK in Candida albicans. In the present work, we investigated the role of Chk1 in the low-virulence species Candida guilliermondii, in order to gain insight into putative conservation of the role of group X HHK in opportunistic yeasts. We demonstrated that disruption of the corresponding gene CHK1 does not influence growth, stress tolerance, drug susceptibility, protein glycosylation or cell wall composition in C. guilliermondii. In addition, we showed that loss of CHK1 does not affect C. guilliermondii ability to interact with macrophages and to stimulate cytokine production by human peripheral blood mononuclear cells. Finally, the C. guilliermondii chk1 null mutant was found to be as virulent as the wild-type strain in the experimental model Galleria mellonella. Taken together, our results demonstrate that group X HHK function is not conserved in Candida species. Copyright © 2017 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.
Schwarzhans, Jan-Philipp; Luttermann, Tobias; Geier, Martina; Kalinowski, Jörn; Friehs, Karl
The methylotrophic yeast Pichia pastoris is firmly established as a host for the production of recombinant proteins, frequently outperforming other heterologous hosts. Already, a sizeable amount of systems biology knowledge has been acquired for this non-conventional yeast. By applying various omics-technologies, productivity features have been thoroughly analyzed and optimized via genetic engineering. However, challenging clonal variability, limited vector repertoire and insufficient genome annotation have hampered further developments. Yet, in the last few years a reinvigorated effort to establish P. pastoris as a host for both protein and metabolite production is visible. A variety of compounds from terpenoids to polyketides have been synthesized, often exceeding the productivity of other microbial systems. The clonal variability was systematically investigated and strategies formulated to circumvent untargeted events, thereby streamlining the screening procedure. Promoters with novel regulatory properties were discovered or engineered from existing ones. The genetic tractability was increased via the transfer of popular manipulation and assembly techniques, as well as the creation of new ones. A second generation of sequencing projects culminated in the creation of the second best functionally annotated yeast genome. In combination with landmark physiological insights and increased output of omics-data, a good basis for the creation of refined genome-scale metabolic models was created. The first application of model-based metabolic engineering in P. pastoris showcased the potential of this approach. Recent efforts to establish yeast peroxisomes for compartmentalized metabolite synthesis appear to fit ideally with the well-studied high capacity peroxisomal machinery of P. pastoris. Here, these recent developments are collected and reviewed with the aim of supporting the establishment of systems metabolic engineering in P. pastoris. Copyright © 2017. Published
Thomas W. Jeffries; N. Q. Shi; J. Y. Cho; P. Lu; K. Dahn; J. Hendrick; H. K. Sreenath
A useful genetic system has been developed for the transformation of Pichia stipitis. This includes two selectable markers (URA3 and LEU2), integrating and autonomous replication vectors, a pop-out cassette that enables multiple targeted disruptions, and a genomic X-library for rapid cloning. Using this system we have cloned two genes for alcohol dehydrogenase (PsADH1...
The methylotrophic yeast Pichia pastoris was used as a host to generate multicopy clones using in-vitro multimerization and GAP vector approaches. The latter approach relied on the selection of spontaneously occurring multiple integrants based on zeocin resistance. Higher levels of heterologous protein could result using ...
Rodil, Kasper; Winschiers-Theophilus, Heike
to understand indigenous youths’ own conception of storytelling the paper presents empirical data from a study with indigenous Khoisan children in Namibia. This is followed by a discussion of an effort of digitizing indigenous intangible cultural heritage in relation to technologies’ embodied bias...
Porter, Robert; Yellow Bird, Michael
The development of a new Indigenous Nations Studies program at the University of Kansas is described. Success depended on a critical mass of Indigenous and non-Indigenous faculty and students that had a sense of political and social justice and understood the need for institutional change. The biggest challenge was countering the entrenched…
Pan, Heng-Yen; Whittaker, Mei M.; Bouveret, Romaric; Berna, Anne; Bernier, Francois; Whittaker, James W.
High-level secretory expression of wheat (Triticum aestivum) germin/oxalate oxidase was achieved in Pichia pastoris fermentation cultures as an α-mating factor signal peptide fusion, based on the native wheat cDNA coding sequence. The oxalate oxidase activity of the recombinant enzyme is substantially increased (7-fold) by treatment with sodium periodate, followed by ascorbate reduction. Using these methods, approximately 1 g (4 x 10 4 U) of purified, activated enzyme was obtained following eight days of induction of a high density Pichia fermentation culture, demonstrating suitability for large-scale production of oxalate oxidase for biotechnological applications. Characterization of the recombinant protein shows that it is glycosylated, with N-linked glycan attached at Asn47. For potential biomedical applications, a nonglycosylated (S49A) variant was also prepared which retains essentially full enzyme activity, but exhibits altered protein-protein interactions
Yong-Su Jin; Jose Cruz; Thomas W. Jeffries
Xylitol production by Pichia stipitis FPL-YS30, a xyl3-Ã1 mutant that metabolizes xylose using an alternative metabolic pathway, was investigated under aerobic and oxygen-limited culture conditions. Under both culture conditions, FPL-YS30 (xyl3-Ã1) produced a negligible amount of ethanol and converted xylose mainly into xylitol with comparable yields (0.30 and 0.27 g...
The term indigenous, long used to distinguish between those who are "native" and their "others" in specific locales, has also become a term for a geocultural category, presupposing a world collectivity of "indigenous peoples" in contrast to their various "others." Many observers have noted that the stimuli for internationalization of the indigenous category originated principally from particular nation-states-Anglo-American settler colonies and Scandinavia. All, I argue, are relevantly political cultures of liberal democracy and weighty (in different ways) in international institutional affairs. However, international indigeneity has not been supported in any unqualified way by actions taken in the name of several nation-states that were among its main points of origin. In fact, staunch resistance to the international indigenous project has recently come from four of them. In 2007, the only four voting countries to reject the main product of international indigenist activity over the past 30 years, the Declaration on the Rights of Indigenous Peoples, were Australia, the United States, Canada, and New Zealand. In these locations, forms of "indigenous relationship" emerged that launched international indigeneity and that strongly influenced international perceptions of what "indigeneity" is and who "indigenous peoples" may be. Some other countries say the model of indigenous relationship that they see represented by the "establishing" set is inapplicable to themselves (but have nonetheless had to take notice of expanding internationalist indigenism). The apparently paradoxical rejection of the draft declaration by the establishing countries is consistent with the combination of enabling and constraining forces that liberal democratic political cultures offer.
Mewa-Ngongang, Maxwell; du Plessis, Heinrich W; Hutchinson, Ucrecia F; Mekuto, Lukhanyo; Ntwampe, Seteno Ko
Biological antimicrobial compounds from yeast can be used to address the critical need for safer preservatives in food, fruit and beverages. The inhibition of Candida guilliermondii, a common fermented beverage spoilage organism, was achieved using antimicrobial compounds produced by Candida pyralidae KU736785. The antimicrobial production system was modelled and optimised using response surface methodology, with 22.5 ℃ and pH of 5.0 being the optimum conditions. A new concept for quantifying spoilage organism inhibition was developed. The inhibition activity of the antimicrobial compounds was observed to be at a maximum after 17-23 h of fermentation, with C. pyralidae concentration being between 0.40 and 1.25 × 10 9 CFU ml -1 , while its maximum specific growth rate was 0.31-0.54 h -1 . The maximum inhibitory activity was between 0.19 and 1.08 l contaminated solidified media per millilitre of antimicrobial compound used. Furthermore, the antimicrobial compound formation rate was 0.037-0.086 l VZI ml -1 ACU h -1 , respectively. The response surface methodology analysis showed that the model developed sufficiently described the antimicrobial compound formation rate 1.08 l VZI ml -1 ACU, as 1.17 l VZI ml -1 ACU, predicted under the optimum production conditions.
Wang, Haikuan; Zhao, Xinhuai; Lu, Fuping
According to the bias of codon utilization of Pichia methanolica, a fragment encoding bovine lactoferricin has been cloned and expressed in the P. methanolica under the control of the alcohol oxidase promoter, which was followed by the Saccharomyces cerevisiae alpha-factor signal peptide. The alpha-factor signal peptide efficiently directed the secretion of bovine lactoferricin from the recombinant yeast cell. The recombinant bovine lactoferricin appears to be successfully expressed, as it displays antibacterial activity (antibacterial assay). Moreover, the identity of the recombinant product was estimated by Tricine-SDS-PAGE.
Aproveitamento de subproduto industrial de óleos vegetais para produção de riboflavina por Candida guilliermondii DM 644 Utilization industrial waste from vegetal oils riboflavine production by candida guilliermondii DM 644
Maria de Lourdes Andrade Pessoa
Full Text Available A produção e consumo de alimentos industrializados têm aumentado a preocupação com suplementação e enriquecimento de alimentos com vitaminas e sais minerais, visando repor as possíveis perdas durante os processos de fabricação, principalmente das vitaminas hidrossolúveis, mais especificamente da vitamina B2 ou riboflavina. Assim sendo, a proposta deste trabalho foi utilizar como componente principal do meio, para produção da riboflavina, um subproduto do refino de óleos vegetais e o microrganismo Candida guillermondii DM 644. A produção da vitamina B2 foi realizada por fermentação em batelada utilizando Erlenmeyer. As condições empregadas foram agitação orbital, ausência de luz, 30°C, e 24h de incubação. A otimização da produção de riboflavina foi realizada através de Delineamento Fatorial Fracionário, para avaliar os efeitos da concentração de matéria graxa, fonte de nitrogênio, pH, velocidade de agitação, fonte de fósforo e extrato de levedura e as possíveis interações. A concentração máxima de riboflavina foi 19,12mg/mL. Os fatores mais importantes para produção de riboflavina foram a concentração de matéria graxa e a fonte de nitrogênio, enquanto que a fonte de fósforo e o extrato de leveduras não estimularam sua biossíntese. A máxima produção foi obtida com matéria graxa a 10g/L, uréia a 2,5g/L e pH 5,0. A velocidade de agitação (200 e 400rpm não interferiu no processo biotecnológico.The ever growing production and consumption of industrialized foods have increased the concern about supplementation and enrichment of food with vitamins and minerals, attempting the replenishment of the possible losses during their production processes, mainly the hydrosoluble vitamins, more specifically B2 vitamin or riboflavin. A subproduct of vegetal oil refining and the microorganism Candida guilliermondii DM 644 were used as the main substrate. The vitamin B2 was produced by the batch
Njoku, Stephen Ikechukwu; Iversen, Jens Asmus; Uellendahl, Hinrich
liquid hydrolysate to ethanol is essential for economically feasible cellulosic ethanol processes. Fermentation of the separated hemicellulose liquid hydrolysates obtained after the WEx pretreatment was done by Pichia stipitis CBS 6054 (Scheffersomyces stipitis). Results: The fermentation of the WEx...
Full Text Available In this work we analyze the spoiling potential of Meyerozyma guilliermondii in yogurt. The analysis was based on contaminated samples sent to us by an industrial laboratory over two years. All the plain and fruit yogurt packages were heavily contaminated by yeasts, but only the last ones, containing fermentable sugars besides lactose, were spoiled by gas swelling. These strains were unable to grow and ferment lactose (as the type strain; they did grow on lactate plus galactose, fermented glucose and sucrose, and galactose (weakly, but did not compete with lactic acid bacteria for lactose. This enables them to grow in any yogurt, although only those with added jam were spoiled due to the fermentation of the fruit sugars. Fermentation, but not growth, was strongly inhibited at 8 °C. In consequence, in plain yogurt as well as in any yogurt maintained at low temperature, yeast contamination would not be detected by the consumer. The risk could be enhanced because the species has been proposed for biological control of fungal infections in organic agriculture. The combination of the IGS PCR-RFLP (amplification of the intergenic spacer region of rDNA followed by restriction fragment length polymorphism analysis method and mitochondrial DNA-RFLP makes a good tool to trace and control the contamination by M. guilliermondii.
Perna, Michelle Dos Santos Cordeiro; Bastos, Reinaldo Gaspar; Ceccato-Antonini, Sandra Regina
The tolerance of the pentose-fermenting yeast Meyerozyma guilliermondii to the inhibitors released after the biomass hydrolysis, such as acetic acid and furfural, was surveyed. We first verified the effects of acetic acid and cell concentrations and initial pH on the growth of a M. guilliermondii strain in a semi-synthetic medium containing acetic acid as the sole carbon source. Second, the single and combined effects of furfural, acetic acid, and sugars (xylose, arabinose, and glucose) on the sugar uptake, cell growth, and ethanol production were also analysed. Growth inhibition occurred in concentrations higher than 10.5 g l -1 acetic acid and initial pH 3.5. The maximum specific growth rate (µ) was 0.023 h -1 and the saturation constant (ks) was 0.75 g l -1 acetic acid. Initial cell concentration also influenced µ. Acetic acid (initial concentration 5 g l -1 ) was co-consumed with sugars even in the presence of 20 mg l -1 furfural without inhibition to the yeast growth. The yeast grew and fermented sugars in a sugar-based medium with acetic acid and furfural in concentrations much higher than those usually found in hemicellulosic hydrolysates.
Khattab, Sadat Mohammad Rezq; Kodaki, Tsutomu
One osmotolerant strain from among 44 yeast isolates was selected based on its growth abilities in media containing high concentrations of sucrose. This selected strain, named SKENNY, was identified as Meyerozyma guilliermondii by sequencing the internal transcribed spacer regions and partial D1/D2 large-subunit domains of the 26S ribosomal RNA. SK-ENNY was utilized to produce high-fructose glucose syrup (HFGS) from sucrose-containing biomass. Conversion rates to HFGS from 310-610 g/l of pure sucrose and from 75-310 g/l of sugar beet molasses were 73.5-94.1% and 76.2-91.1%, respectively. In the syrups produced, fructose yields were 89.4-100% and 96.5-100% and glucose yields were 57.6-82.5% and 55.3-79.5% of the theoretical values for pure sucrose and molasses sugars, respectively. This is the first report of employing M. guilliermondii for production of HFGS from sucrose-containing biomass.
Donna L. M. Kurtz
Full Text Available Increasing Indigenous health care professional presence in health care aims to reduce health inequities of Indigenous Peoples in Canada. Nurses are the largest health professional group and nurse graduates the main source of recruitment. The quality of graduate transition to practice is evident in the literature; however, little is reported about Indigenous new graduates. We describe using Indigenous methodology and two-eyed seeing (Indigenous and Western perspectives in exploring Indigenous transition experiences. Talking circles provided a safe environment for nurses, nurse educators and students, health managers, and policy makers to discuss Indigenous new graduate case scenarios. The methodology was critical in identifying challenges faced, recommendations for change, and a new collective commitment for cultural safety education, and ethical and respectful relationships within education, practice, and policy.
Received 31 August 1996; accepted 20 March /998. Mitochondrial DNA cleavage patterns from representative animals of the Afrikaner and Nguni sanga cattle breeds, indigenous to Southern Africa, were compared to the mitochondrial DNA cleavage patterns of the Brahman (zebu) and the Jersey. (taurine) cattle breeds.
Zhou, Xiang-Shan; Fan, Wei-Min; Zhang, Yuan-Xing
Chemostat culture was performed to characterize the growth, substrate consumption and the hirudin production, and to disclose their interrelations in the fermentation of recombinant Pichia pastoris. The Andrew substrate-inhibited growth model is more suitable than Monod model to simulate the growth of Pichia pastoris on methanol. Therefore, two stationary states can be obtained in the continuous culture at a certain dilution rate because of the substrate inhibition on cell growth. The stationary state could be obtained if only the dilution rate not more than 0.048 h(-1) in the continuous fermentation. The concentrations of cell, methanol and hirudin were constant after 50 h continuous culture with dilution rate at 0.04 h(-1). However, it could not be obtained when the dilution rate more than 0.048 h(-1) because the other stationary point at S > 0.048 h(-1) is unstable. Therefore, it was found that the cell concentration declined and the methanol concentration increased from 2.9 g/L to 18.1 g/L within 18h at dilution rate 0.06 h(-1). Thus, the fed-batch culture with a constant specific growth rate was carried out to disclose the fermentation behavior at high and constant methanol concentration in aid of a methanol sensor. The theoretical maximum specific growth rate, microm = 0.0464 h(-1), was found under critical methanol concentration, Scrit = 3.1 g/L. The growth of P. pastoris was typically methanol-limited at the methanol concentration S Scrit. The maximum specific Hir65 production rate qp was obtained at 0.2 mg/(g x h) when methanol concentration and mu were 0.5 g/L and 0.02 h(-1), respectively. The specific Hir65 production rate qp increased with the increase of mu and S at mu 0.02 h(-1). The specific methanol consumption rate increased with the increase of S when S 5 g/L. At last, the high Hir65 production rate 0.2 mg/(g x h) was obtained in the fermentation conducted under methanol-limited concentration and mu controlled at 0.5 g/L and 0.02 h(-1
Mohammed N. Baeshen
Full Text Available Increase in the incidence of Insulin Dependent Diabetes Mellitus (IDDM among people from developed and developing countries has created a large global market for insulin. Moreover, exploration of new methods for insulin delivery including oral or inhalation route which require very high doses would further increase the demand of cost-effective recombinant insulin. Various bacterial and yeast strains have been optimized to overproduce important biopharmaceuticals. One of the approaches we have taken is the production of recombinant human insulin along with C-peptide in yeast Pichia pastoris. We procured a cDNA clone of insulin from Origene Inc., USA. Insulin cDNA was PCR amplified and cloned into yeast vector pPICZ-α. Cloned insulin cDNA was confirmed by restriction analysis and DNA sequencing. pPICZ-α-insulin clone was transformed into Pichia pastoris SuperMan5 strain. Several Zeocin resistant clones were obtained and integration of insulin cDNA in Pichia genome was confirmed by PCR using insulin specific primers. Expression of insulin in Pichia clones was confirmed by ELISA, SDS-PAGE, and Western blot analysis. In vivo efficacy studies in streptozotocin induced diabetic mice confirmed the activity of recombinant insulin. In conclusion, a biologically active human proinsulin along with C-peptide was expressed at high level using Pichia pastoris expression system.
A bacterial endo-polygalacturonase (endo-PGase) gene from the plant pathogen Pectobacterium carotovorum was cloned into pGAPZaA vector and constitutively expressed in Pichia pastoris. The recombinant endo-PGase secreted by the Pichia clone showed a 1.7 fold increase when the culture medium included ...
Sarfati, Diana; Garvey, Gail; Robson, Bridget; Moore, Suzanne; Cunningham, Ruth; Withrow, Diana; Griffiths, Kalinda; Caron, Nadine R; Bray, Freddie
It is estimated that there are 370 million indigenous peoples in 90 countries globally. Indigenous peoples generally face substantial disadvantage and poorer health status compared with nonindigenous peoples. Population-level cancer surveillance provides data to set priorities, inform policies, and monitor progress over time. Measuring the cancer burden of vulnerable subpopulations, particularly indigenous peoples, is problematic. There are a number of practical and methodological issues potentially resulting in substantial underestimation of cancer incidence and mortality rates, and biased survival rates, among indigenous peoples. This, in turn, may result in a deprioritization of cancer-related programs and policies among these populations. This commentary describes key issues relating to cancer surveillance among indigenous populations including 1) suboptimal identification of indigenous populations, 2) numerator-denominator bias, 3) problems with data linkage in survival analysis, and 4) statistical analytic considerations. We suggest solutions that can be implemented to strengthen the visibility of indigenous peoples around the world. These include acknowledgment of the central importance of full engagement of indigenous peoples with all data-related processes, encouraging the use of indigenous identifiers in national and regional data sets and mitigation and/or careful assessment of biases inherent in cancer surveillance methods for indigenous peoples. Copyright © 2018 Elsevier Inc. All rights reserved.
Malone, Christine C.; Ciszak, Eva; Karr, Laurel J.
A soluble form of human bone alkaline phosphatase has been expressed in a recombinant strain of the methylotrophic yeast Pichia pastoris. We constructed a plasmid containing cDNA encoding for human bone alkaline phosphatase, with the hydrophobic carboxyl terminal portion deleted. Alkaline phosphatase was secreted into the medium to a level of 32mg/L when cultured in shake flasks, and enzyme activity was 12U/mg, as measured by a spectrophotometric assay. By conversion to a fermentation system, a yield of 880mg/L has been achieved with an enzyme activity of 968U/mg. By gel electrophoresis analysis, it appears that greater than 50% of the total protein in the fermentation media is alkaline phosphatase. Although purification procedures are not yet completely optimized, they are expected to include filtration, ion exchange and affinity chromatography. Our presentation will focus on the purification and crystallization results up to the time of the conference. Structural data should provide additional information on the role of alkaline phosphatase in normal bone mineralization and in certain bone mineralization anomalies.
Full Text Available Abstract Background High cell density cultures of Pichia pastoris grown on methanol tend to develop yellow colored supernatants, attributed to the release of free flavins. The potential of P. pastoris for flavin overproduction is therefore given, but not pronounced when the yeast is grown on glucose. The aim of this study is to characterize the relative regulatory impact of each riboflavin synthesis gene. Deeper insight into pathway control and the potential of deregulation is established by overexpression of the single genes as well as a combined deregulation of up to all six riboflavin synthesis genes. Results Overexpression of the first gene of the riboflavin biosynthetic pathway (RIB1 is already sufficient to obtain yellow colonies and the accumulation of riboflavin in the supernatant of shake flask cultures growing on glucose. Sequential deregulation of all the genes, by exchange of their native promoter with the strong and constitutive glyceraldehyde-3-phosphate dehydrogenase promoter (PGAP increases the riboflavin accumulation significantly. Conclusion The regulation of the pathway is distributed over more than one gene. High cell density cultivations of a P. pastoris strain overexpressing all six RIB genes allow the accumulation of 175 mg/L riboflavin in the supernatant. The basis for rational engineering of riboflavin production in P. pastoris has thus been established.
Krainer, Florian W; Gerstmann, Michaela A; Darnhofer, Barbara; Birner-Gruenberger, Ruth; Glieder, Anton
Horseradish peroxidase (HRP) is a high-demand enzyme for applications in diagnostics, bioremediation, biocatalysis and medicine. Current HRP preparations are isolated from horseradish roots as mixtures of biochemically diverse isoenzymes. Thus, there is a strong need for a recombinant production process enabling a steady supply with enzyme preparations of consistent high quality. However, most current recombinant production systems are limited at titers in the low mg/L range. In this study, we used the well-known yeast Pichia pastoris as host for recombinant HRP production. To enhance recombinant enzyme titers we systematically evaluated engineering approaches on the secretion process, coproduction of helper proteins, and compared expression from the strong methanol-inducible PAOX1 promoter, the strong constitutive PGAP promoter, and a novel bidirectional promoter PHTX1. Ultimately, coproduction of HRP and active Hac1 under PHTX1 control yielded a recombinant HRP titer of 132mg/L after 56h of cultivation in a methanol-independent and easy-to-do bioreactor cultivation process. With regard to the many versatile applications for HRP, the establishment of a microbial host system suitable for efficient recombinant HRP production was highly overdue. The novel HRP production platform in P. pastoris presented in this study sets a new benchmark for this medically relevant enzyme. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.
This article focuses on the impact of colonisation and its associated impact on Indigenous teaching and learning. Western European institutions have dominated Indigenous ways of knowing and in Australia this has led to barriers which restrict the participation of Aboriginal people in education systems. Globally Indigenous people are attempting to bring into the introduced educational systems culturally appropriate teaching and learning practices so that a more holistic approach to education can become the norm rather than the exception. The relationship between Indigenous knowledge and western European concepts of knowledge and knowing need to placed in a framework of mutual interaction so that not only do Indigenous people benefit, but so do non-Indigenous educators and students.
Full Text Available In this study, bovine lactoferrin (bLf, an iron-binding glycoprotein considered an important nutraceutical protein because of its several properties, was expressed in Pichia pastoris KM71-H under AOX1 promoter control, using pJ902 as the recombinant plasmid. Dot blotting analysis revealed the expression of recombinant bovine lactoferrin (rbLf in Pichia pastoris. After Bach fermentation and purification by molecular exclusion, we obtained an expression yield of 3.5 g/L of rbLf. rbLf and predominantly pepsin-digested rbLf (rbLfcin demonstrated antibacterial activity against Escherichia coli (E. coli BL21DE3, Staphylococcus aureus (S. aureus FRI137, and, in a smaller percentage, Pseudomonas aeruginosa (Ps. Aeruginosa ATCC 27833. The successful expression and characterization of functional rbLf expressed in Pichia pastoris opens a prospect for the development of natural antimicrobial agents produced recombinantly.
Ren, Ren; Jiang, Zhengbing; Liu, Meiyun; Tao, Xinyi; Ma, Yushu; Wei, Dongzhi
Two Pichia pastoris cell surface display vectors were constructed. The vectors consisted of the flocculation functional domain of Flo1p with its own secretion signal sequence or the alpha-factor secretion signal sequence, a polyhistidine (6xHis) tag for detection, an enterokinase recognition site, and the insertion sites for target proteins. Adenoregulin (ADR) is a 33-amino-acid antimicrobial peptide isolated from Phyllomedusa bicolor skin. The ADR was expressed and displayed on the Pichia pastoris KM71 cell surface with the system reported. The displayed recombinant ADR fusion protein was detected by fluorescence microscopy and confocal laser scanning microscopy (CLSM). The antimicrobial activity of the recombinant adenoregulin was detected after proteolytic cleavage of the fusion protein on cell surface. The validity of the Pichia pastoris cell surface display vectors was proved by the displayed ADR.
Du, W; Xia, J; Zhang, Y; Liu, M J; Li, H B; Yan, X M; Zhang, J S; Li, N; Zhou, Z Y; Xie, W Z
Myostatin propeptide can inhibit the biological activity of myostatin protein and promote muscle growth. To express myostatin propeptide in vitro with a higher biological activity, we performed codon optimization on the sheep myostatin propeptide gene sequence, and mutated aspartic acid-76 to alanine based on the codon usage bias of Pichia pastoris and the enhanced biological activity of myostatin propeptide mutant. Modified myostatin propeptide gene was cloned into the pPIC9K plasmid to form the recombinant plasmid pPIC9K-Msp. Recombinant plasmid pPIC9K-Msp was transformed into Pichia pastoris GS115 by electrotransformation. Transformed cells were screened, and methanol was used to induce expression. SDS-PAGE and western blotting were used to verify the successful expression of myostatin propeptide with biological activity in Pichia pastoris, providing the basis for characterization of this protein.
Full Text Available During the twentieth century, indigenous peoples have often embraced the category of indigenous while also having to face the ambiguities and limitations of this concept. Indigeneity, whether represented by indigenous people themselves or others, tends to face a “double bind”, as defined by Gregory Bateson, in which “no matter what a person does, he can’t win.” One exit strategy suggested by Bateson is meta-communication—communication about communication—in which new solutions emerge from a questioning of system-internal assumptions. We offer case studies from Ecuador, Peru and Alaska that chart some recent indigenous experiences and strategies for such scenarios.
Liu, Yifei; Chen, Delong; Luo, Yunbo; Huang, Kunlun; Zhang, Wei; Xu, Wentao
A recombinant lactase was expressed in Pichia pastoris, resulting in enzymatic activity of 3600 U/mL in a 5 L fermenter. The lactase product was subjected to a series of toxicological tests to determine its safety for use as an enzyme preparation in the dairy industry. This recombinant lactase had the highest activity of all recombinant strains reported thus far. Acute oral toxicity, mutagenicity, genotoxic, and subchronic toxicity tests performed in rats and mice showed no death in any groups. The lethal dose 50% (LD50) based on the acute oral toxicity study is greater than 30 mL/kg body weight, which is in accordance with the 1500 L milk consumption of a 50 kg human daily. The lactase showed no mutagenic activity in the Ames test or a mouse sperm abnormality test at levels of up to 5 mg/plate and 1250 mg/kg body weight, respectively. It also showed no genetic toxicology in a bone marrow cell micronucleus test at levels of up to 1250 mg/kg body weight. A 90-day subchronic repeated toxicity study via the diet with lactase levels up to 1646 mg/kg (1000-fold greater than the mean human exposure) did not show any treatment-related significant toxicological effects on body weight, food consumption, organ weights, hematological and clinical chemistry, or histopathology compared to the control groups. This toxicological evaluation system is comprehensive and can be used in the safety evaluation of other enzyme preparations. The lactase showed no acute, mutagenic, genetic, or subchronic toxicity under our evaluation system. PMID:25184300
Full Text Available A recombinant lactase was expressed in Pichia pastoris, resulting in enzymatic activity of 3600 U/mL in a 5 L fermenter. The lactase product was subjected to a series of toxicological tests to determine its safety for use as an enzyme preparation in the dairy industry. This recombinant lactase had the highest activity of all recombinant strains reported thus far. Acute oral toxicity, mutagenicity, genotoxic, and subchronic toxicity tests performed in rats and mice showed no death in any groups. The lethal dose 50% (LD50 based on the acute oral toxicity study is greater than 30 mL/kg body weight, which is in accordance with the 1500 L milk consumption of a 50 kg human daily. The lactase showed no mutagenic activity in the Ames test or a mouse sperm abnormality test at levels of up to 5 mg/plate and 1250 mg/kg body weight, respectively. It also showed no genetic toxicology in a bone marrow cell micronucleus test at levels of up to 1250 mg/kg body weight. A 90-day subchronic repeated toxicity study via the diet with lactase levels up to 1646 mg/kg (1000-fold greater than the mean human exposure did not show any treatment-related significant toxicological effects on body weight, food consumption, organ weights, hematological and clinical chemistry, or histopathology compared to the control groups. This toxicological evaluation system is comprehensive and can be used in the safety evaluation of other enzyme preparations. The lactase showed no acute, mutagenic, genetic, or subchronic toxicity under our evaluation system.
Rosa Laura Andrade Melchor
Full Text Available ABSTRACT Coffee rust is a fungal disease that has affected every coffee-producing region in the world. Given that the effectivity of the protectant and systemic fungicides applied routinely to control the spread of the causative agent of the disease (Hemileia vastatrix has gradually diminished, besides are harmful to mammals and ecosystems, the objective of this work was to search for a mixture of harmless natural compounds with the potential to be applied in the field. So, a yeast strain producing a battery of long-chain carboxylic acids (CA with fungicide properties was isolated from soil of coffee crop and identified as Pichia membranifaciens by ITS sequencing. Culture conditions of the yeast were optimized and the CA in the solution were characterized by Gas Chromatography-Mass Spectrometry (GC-MS as ethyl formate (55.5 g L-1, octadecenoic acid (3.5 g L-1, propionic acid (7.2 g L-1, 3-(octadecanoyl-propionic acid (7.2 g L-1 and methyl acetate (8.4 g L-1. Randomized field studies were conducted in three different locations in Chiapas, México. Five treatments were tested including three concentrations of the CA solution (389, 584 and 778 ppm and copper oxychloride (5 000 ppm as conventional control. The initial coffee rust incidence averages varied between sites: Maravillas (3-9%, Santo Domingo (10-16% and Búcaro (16-22%. The treatments of CA solution proved to be effective at slowing down the progress of the rust disease even for the sites where initial incidence was high. Likewise, the CA solution reduced the viability of H. vastatrix spores, as assessed by fluorescence microscopy.
Canilha, Larissa; Carvalho, Walter; Felipe, Maria das Graças de Almeida; Silva, João Batista de Almeida e; Giulietti, Marco
The objective of this study was to evaluate the ethanol production from the sugars contained in the sugarcane bagasse hemicellulosic hydrolysate with the yeast Pichia stipitis DSM 3651. The fermentations were carried out in 250-mL Erlenmeyers with 100 mL of medium incubated at 200 rpm and 30 degrees C for 120 h. The medium was composed by raw (non-detoxified) hydrolysate or by hydrolysates detoxified by pH alteration followed by active charcoal adsorption or by adsorption into ion-exchange resins, all of them supplemented with yeast extract (3 g/L), malt extract (3 g/L), and peptone (5 g/L). The initial concentration of cells was 3 g/L. According to the results, the detoxification procedures removed inhibitory compounds from the hemicellulosic hydrolysate and, thus, improved the bioconversion of the sugars into ethanol. The fermentation using the non-detoxified hydrolysate led to 4.9 g/L ethanol in 120 h, with a yield of 0.20 g/g and a productivity of 0.04 g L(-1) h(-1). The detoxification by pH alteration and active charcoal adsorption led to 6.1 g/L ethanol in 48 h, with a yield of 0.30 g/g and a productivity of 0.13 g L(-1) h(-1). The detoxification by adsorption into ion-exchange resins, in turn, provided 7.5 g/L ethanol in 48 h, with a yield of 0.30 g/g and a productivity of 0.16 g L(-1) h(-1).
Purpose: The purpose of this paper is to draw attention to an indigenous community that lives in the periphery of Taiwan. The Dao on Orchid Island have had to face serious abuse of their human rights in terms of ecological exploitation and environmental injustice. The article highlights the empowerment of the indigenous group through collective…
Hanna, Philippe; Langdon, Esther Jean; Vanclay, Frank
Protests to claim rights are a common practice among Indigenous peoples of the world, especially when their interests conflict with those of nation states and/or multinational corporations regarding the use of their lands and resources. Drawing on a case study of the National Indigenous Mobilization
The thesis (working title: 'Indigenous Education and Heritage Revitalization') focuses on the (possible) roles of tangible and intangible cultural heritage in the education of indigenous peoples in Taiwan, against the background of worldwide discussions and studies of the possibilities to create and
Lilian Alessa; Carlos Andrade; Phil Cash Cash; Christian P. Giardina; Matt Hamabata; Craig Hammer; Kai Henifin; Lee Joachim; Jay T. Johnson; Kekuhi Kealiikanakaoleohaililani; Deanna Kingston; Andrew Kliskey; Renee Pualani Louis; Amanda Lynch; Daryn McKenny; Chels Marshall; Mere Roberts; Taupouri Tangaro; Jyl Wheaton-Abraham; Everett. Wingert
This policy brief explores the use and expands the conversation on the ability of geospatial technologies to represent Indigenous cultural knowledge. Indigenous peoples' use of geospatial technologies has already proven to be a critical step for protecting tribal self-determination. However, the ontological frameworks and techniques of Western geospatial...
Wilde, de W.J.J.O.
A short revision of the indigenous Old World taxa in Passifiora in the form of a key, the enumeration of synonyms, descriptions, and an index accounting for all names proposed for the area. Examined specimens, distributional areas, and some notes are given. In the Old World 20 indigenous species are
Support for indigenous peoples has been increasing over the last few decades. This can be seen internationally, as well as in several domestic contexts. The support for indigenous people has been linked to the increasingly prominent impetus to conserve the Earth’s biodiversity and environment.
Wright, Jason T.
One of the primary open questions of astrobiology is whether there is extant or extinct life elsewhere the solar system. Implicit in much of this work is that we are looking for microbial or, at best, unintelligent life, even though technological artefacts might be much easier to find. Search for Extraterrestrial Intelligence (SETI) work on searches for alien artefacts in the solar system typically presumes that such artefacts would be of extrasolar origin, even though life is known to have existed in the solar system, on Earth, for eons. But if a prior technological, perhaps spacefaring, species ever arose in the solar system, it might have produced artefacts or other technosignatures that have survived to present day, meaning solar system artefact SETI provides a potential path to resolving astrobiology's question. Here, I discuss the origins and possible locations for technosignatures of such a prior indigenous technological species, which might have arisen on ancient Earth or another body, such as a pre-greenhouse Venus or a wet Mars. In the case of Venus, the arrival of its global greenhouse and potential resurfacing might have erased all evidence of its existence on the Venusian surface. In the case of Earth, erosion and, ultimately, plate tectonics may have erased most such evidence if the species lived Gyr ago. Remaining indigenous technosignatures might be expected to be extremely old, limiting the places they might still be found to beneath the surfaces of Mars and the Moon, or in the outer solar system.
Markošová, K.; Weignerová, Lenka; Rosenberg, M.; Křen, Vladimír; Rebroš, M.
Roč. 6, OCT 2015 (2015), s. 1-10 ISSN 1664-302X R&D Projects: GA MŠk(CZ) 7E11010; GA MŠk(CZ) LD15085 Institutional support: RVO:61388971 Keywords : Pichia pastoris * alpha-L-rhamnosidase * recombinant enzyme Subject RIV: CE - Biochemistry Impact factor: 4.165, year: 2015
Jose M. Laplaza; Beatriz Rivas Torres; Yong-Su Jin; Thomas W. Jeffries
Pichia stipitis is widely studied for its capacity to ferment d-xylose to ethanol. Strain improvement has been facilitated by recent completion of the P. stipitis genome. P. stipitis uses CUG to code for serine rather than leucine, as is the case for the universal genetic code thereby limiting the availability of heterologous drug resistance markers for transformation...
Hassan K. Sreenath; Thomas W. Jeffries
The glucose analog 2-deoxyglucose (2-DOG) has been used to obtain mutants derepressed for pentose metabolism. Some researchers have used 2-DOG alone whereas others have used it in the presence of a glucoserepressible carbon source. We examined both methods and screened mutant strains for improved use of xylose in the presence of glucose. Pichia stipitis mutants...
de Jong, Lutea; Grünewald, S; Franke, J.P.; Uges, Donald; Bischoff, Rainer
The human dopamine D2S receptor was expressed in the methylotrophic yeast Pichia pastoris, where the receptor with a molecular mass of approximately 40 kDa exhibited specific and saturable binding properties. The dopamine antagonist [H-3]spiperone showed an average dissociation constant K-d of 0.6
Vester-Christensen, Malene Bech; Abou Hachem, Maher; Næsted, Henrik
biosynthesis by trimming of intermediate branched alpha-glucan structures. Highly active barley LD is obtained by secretory expression during high cell-density fermentation of Pichia pastoris. The LD encoding gene fragment without signal peptide was subcloned in-frame with the Saccharomyces cerevisiae alpha...
The human Iduronate-2-sulfate sulfatase (hIDS-Like) was cloned into the methylotrophic yeast Pichia pastoris under the control of alcohol oxidase promoter (AOX1) and the -mating factor signal peptide (a-factor). Six clones were identified by PCR. Using clone IDS28, the enzyme was secreted into the culture medium, ...
Rita C.L.B. Rodrigues; William R. Kenealy; Thomas W. Jeffries
Corn stover that had been treated with vapor-phase diethyl oxalate released a mixture of mono-and oligosaccharides consisting mainly of xylose and glucose. Following overliming and neutralization, a D-xylulokinase mutant of Pichia stipitis, FPL-YS30 (xyl3 -Ã1), converted the stover hydrolysate into xylitol. This research examined the effects of phosphoric or gluconic...
To construct the Pichia pastoris (P. pastoris) cell surface display system of Bombyx mori acetylcholinesterase (BmAChE), the gene for the anchor protein (AGa1) was obtained from Saccharomyces cerevisiae and was fused with the modified Bombyx mori acetylcholinesterase gene (bmace) and transformed int...
Jun 17, 2009 ... Restriction enzymes,. T4-ligase, Taq-DNA polymerase and culture media were from .... Each experiment was run at least 3 times, with a coef- ficient of variation ..... New York,. Greene Publishing ... Pichia expression kit. Protein ...
Pichia pastoris is a eukaryote and has many of the advantages of higher eukaryotic expression systems, such as protein processing, protein folding, and the availability of posttranslational modifications. It is as easy to manipulate as Escherichia coli or Saccharomyces cerevisiae. However, some serious and unavoidable ...
Molecular characterization and technological properties of wine yeasts isolated during spontaneous fermentation of Vitis vinifera L.cv. Narince grape must grown in ancient wine making area Tokat, Anatolia
Çelik Zeynep Dilan
Full Text Available Narince is a native white grape variety of Vitis vinifera L grown in Tokat and produces rich and balanced wines often with a greenish yellow tint and delicate fruity flavour. Fermentation by indigenous yeasts may produce wines with complex oenological properties that are unique to specific region. In this study yeast population during alcoholic fermentation of Narince was investigated. Yeasts were identified by PCR-RFLP analysis of the 5.8 ITS rRNA region and sequence information for the D1/D2 domains of the 26S gene. Eight different species belonging to nine genera were identified as: Hanseniaspora uvarum, Hansenispora guilliermondii, Pichia kluyveri, Metschnikowiaspp., Pichia occidentalis, Torulaspora delbrueckii, Candida zemplinina, Lachancea thermotolerance and Saccharomyces cerevisiae. Hanseniaspora guilliermondii, Metschnikowiaspp., Pichia occidentalis and Pichia kluyveri were identified only in the early stage of fermentation. Selected yeasts tested for their physiological traits, ethanol, SO2, temperature, pH tolerance, H2S production, killer and enzymatic activity, fermentation rate, flocculation characteristic, foam, volatile acid and volatile compounds production. Among the yeasts, one,Lachancea thermotolerance and four Saccharomyces cerevisiae strain showed remarkable technological properties and results were compared with those obtained by using commercial starter culture.
Full Text Available The paper begins by noting the low level of reference to Indigenous Australians in the Commonwealth Constitution at the start of Federation, and goes on to discuss the limits to what was achieved by the 1967 amendments. The situation represents a marked contrast with the USA and Canada in terms of treaties and constitutional recognition. In Australia, particularly during the period of the ‘Reconciliation’ process in the 1990s, important steps were taken by Indigenous Australians to identify items of ‘unfinished business’ in a ‘Statement of Indigenous Rights’. But there has been limited progress to meet these aspirations. And Australian law still lacks a tradition of recognition of human rights generally, let alone Indigenous rights. International law, too, largely lacked recognition of human rights, generally prior to the adoption in 1945 of the Charter of the United Nations. The brief references in the Charter were subsequently developed in a range of declarations and of treaties. These applied to people generally, with scant reference to Indigenous peoples. But, since the 1970s, there has been growing international recognition of the rights of Indigenous peoples under existing declarations and treaties. Since the 1990s, in particular, the UN system has established specific mechanisms for addressing such issues. On 13 September 2007, the General Assembly finally adopted a Declaration on the Rights of Indigenous Peoples.
Catherine E. Gordon
Full Text Available In this article, the educational attainment of Indigenous peoples of working age (25 to 64 years in Canada is examined. This diverse population has typically had lower educational levels than the general population in Canada. Results indicate that, while on the positive side there are a greater number of highly educated Indigenous peoples, there is also a continuing gap between Indigenous and non-Indigenous peoples. Data also indicate that the proportion with less than high school education declined, which corresponds with a rise of those with a PSE; the reverse was true in 1996. Despite these gains, however, the large and increasing absolute numbers of those without a high school education is alarming. There are intra-Indigenous differences: First Nations with Indian Status and the Inuit are not doing as well as non-Status and Métis peoples. Comparisons between the Indigenous and non-Indigenous populations reveal that the documented gap in post-secondary educational attainment is at best stagnant. Out of the data analysis, and based on the history of educational policy, we comment on the current reform proposed by the Government of Canada, announced in February of 2014, and propose several policy recommendations to move educational attainment forward.
Okonkwo, C C; Azam, M M; Ezeji, T C; Qureshi, N
Studies were performed on the effect of CaCO3 and CaCl2 supplementation to fermentation medium for ethanol production from xylose, glucose, or their mixtures using Scheffersomyces (Pichia) stipitis. Both of these chemicals were found to improve maximum ethanol concentration and ethanol productivity. Use of xylose alone resulted in the production of 20.68 ± 0.44 g L(-1) ethanol with a productivity of 0.17 ± 0.00 g L(-1) h(-1), while xylose plus 3 g L(-1) CaCO3 resulted in the production of 24.68 ± 0.75 g L(-1) ethanol with a productivity of 0.21 ± 0.01 g L(-1) h(-1). Use of xylose plus glucose in combination with 3 g L(-1) CaCO3 resulted in the production of 47.37 ± 0.55 g L(-1) ethanol (aerobic culture), thus resulting in an ethanol productivity of 0.39 ± 0.00 g L(-1) h(-1). These values are 229 % of that achieved in xylose medium. Supplementation of xylose and glucose medium with 0.40 g L(-1) CaCl2 resulted in the production of 44.84 ± 0.28 g L(-1) ethanol with a productivity of 0.37 ± 0.02 g L(-1) h(-1). Use of glucose plus 3 g L(-1) CaCO3 resulted in the production of 57.39 ± 1.41 g L(-1) ethanol under micro-aerophilic conditions. These results indicate that supplementation of cellulosic sugars in the fermentation medium with CaCO3 and CaCl2 would improve economics of ethanol production from agricultural residues.
Jin, Jun; Slepniov, Dmitrij
champions. However, recently growing number of Chinese companies are seeking to create a foundation for growth and development based on innovation. As a result of this, many of them spread their operations to the countries of the traditional industrial ‘triad’ of North America, Europe and Japan to capture...... a foothold in these markets and to tap into the advanced technologies and concepts originating from this developed context. Another category of Chinese companies includes those who seek to move from routine transactional tasks to more innovation-intensive concepts while remaining in China and relying...... on their own in-house resources. The development and implementation of indigenous innovation solutions for these companies is an imperative which has not been adequately addressed in the literature. Therefore, by employing an explorative case of a Chinese company behind an innovative logistics concept...
Li, Peter Ping; Leung, Kwok; Chen, Chao C.
We attempt to provide a definition and a typology of indigenous research on Chinese management as well as outline the general methodological approaches for this type of research. We also present an integrative summary of the four articles included in this special issue and show how they illustrate...... our definition and typology of indigenous research on Chinese management, as well as the various methodological approaches we advocate. Further, we introduce a commentary on the four articles from the perspective of engaged scholarship, and also three additional articles included in this issue....... Finally, we conclude with our suggestions for future indigenous research....
Cortez, Daniela Vieira; Mussatto, Solange I; Roberto, Inês Conceição
Cells of Candida guilliermondii permeabilized with Triton X-100 were able to efficiently produce xylitol from a medium composed only by D-xylose and MgCl 2 ·6H 2 O in potassium phosphate buffer, at 35 °C and pH 6.5. Under these conditions, the results were similar to those obtained when cofactor and co-substrate or nutrients were added to the medium (about 95 % D-xylose was assimilated producing 42 g/L of xylitol, corresponding to 0.80 g/g yield and 2.65 g/L h volumetric productivity). Furthermore, the permeabilized cells kept the D-xylose assimilation in about 90 % and the xylitol production in approx. 40 g/L during three bioconversion cycles of 16 h each. These values are highly relevant when compared to others reported in the literature using enzyme technology and fermentative process, thereby demonstrating the effectiveness of the proposed method. The present study reveals that the use of permeabilized cells is an interesting alternative to obtain high xylitol productivity using low cost medium formulation. This approach may allow the future development of xylitol production from xylose present in lignocellulosic biomass, with additional potential for implementation in biorefinery strategies.
Alves, Lourdes A; Vitolo, Michele; Felipe, Maria das Graças A; de Almeida e Silva, João Batista
The sugarcane bagasse hydrolysate, which is rich in xylose, can be used as culture medium for Candida guilliermondii in xylitol production. However, the hydrolysate obtained from bagasse by acid hydrolysis at 120 degrees C for 20 min has by-products (acetic acid and furfural, among others), which are toxic to the yeast over certain concentrations. So, the hydrolysate must be pretreated before using in fermentation. The pretreatment variables considered were: adsorption time (15,37.5, and 60 min), type of acid used (H2So4 and H3Po4), hydrolysate concentration (original, twofold, and fourfold concentrated), and active charcoal (0.5, 1.75 and 3.0%). The suitability of the pretreatment was followed by measuring the xylose reductase (XR) and xylitol dehydrogenase (XD) activity of yeast grown in each treated hydrolysate. The response surface methodology (2(4) full factorial design with a centered face) indicated that the hydrolysate might be concentrated fourfold and the pH adjusted to 7.0 with CaO, followed by reduction to 5.5 with H3PO4. After that it was treated with active charcoal (3.0%) by 60 min. This pretreated hydrolysate attained the high XR/XD ratio of 4.5.
Gurpilhares, Daniela B; Pessoa, Adalberto; Roberto, Inês C
Remaining cells of Candida guilliermondii cultivated in hemicellulose-based fermentation medium were used as intracellular protein source. Recovery of glucose-6-phosphate dehydrogenase (G6PD) was attained in conventional aqueous two-phase systems (ATPS) was compared with integrated process involving mechanical disruption of cells followed by ATPS. Influences of polyethylene glycol molar mass (M PEG) and tie line lengths (TLL) on purification factor (PF), yields in top (Y T ) and bottom (Y B ) phases and partition coefficient (K) were evaluated. First scheme resulted in 65.9 % enzyme yield and PF of 2.16 in salt-enriched phase with clarified homogenate (M PEG 1500 g mol(-1), TLL 40 %); Y B of 75.2 % and PF B of 2.9 with unclarified homogenate (M PEG 1000 g mol(-1), TLL 35 %). The highest PF value of integrated process was 2.26 in bottom phase (M PEG 1500 g mol(-1), TLL 40 %). In order to optimize this response, a quadratic model was predicted for the response PFB for process integration. Maximum response achieved was PFB = 3.3 (M PEG 1500 g mol(-1), TLL 40 %). Enzyme characterization showed G6P Michaelis-Menten constant (K M ) equal 0.07-0.05, NADP(+) K M 0.02-1.98 and optimum temperature 70 °C, before and after recovery. Overall, our data confirmed feasibility of disruption/extraction integration for single-step purification of intracellular proteins from remaining yeast cells.
Hing, Nerilee; Breen, Helen; Gordon, Ashley; Russell, Alex
The gambling activities of minority groups such as Indigenous peoples are usually culturally complex and poorly understood. To redress the scarcity of information and contribute to a better understanding of gambling by Indigenous people, this paper presents quantitative evidence gathered at three Australian Indigenous festivals, online and in several Indigenous communities. With support from Indigenous communities, the study collected and analyzed surveys from 1,259 self-selected Indigenous adults. Approximately 33 % of respondents gambled on card games while 80 % gambled on commercial gambling forms in the previous year. Gambling participation and involvement are high, particularly on electronic gaming machines (EGMs), the favorite and most regular form of gambling. Men are significantly more likely to participate in gambling and to gamble more frequently on EGMs, horse/dog races, sports betting and instant scratch tickets. This elevated participation and frequency of gambling on continuous forms would appear to heighten gambling risks for Indigenous men. This is particularly the case for younger Indigenous men, who are more likely than their older counterparts to gamble on EGMs, table games and poker. While distinct differences between the gambling behaviors of our Indigenous sample and non-Indigenous Australians are apparent, Australian Indigenous behavior appears similar to that of some Indigenous and First Nations populations in other countries. Although this study represents the largest survey of Indigenous Australian gambling ever conducted in New South Wales and Queensland, further research is needed to extend our knowledge of Indigenous gambling and to limit the risks from gambling for Indigenous peoples.
Burger, Julian; Kapron, Mary
This paper identifies the principal concerns of indigenous peoples with regard to current international treaties on certain psychoactive substances and policies to control and eradicate their production, trafficking, and sale. Indigenous peoples have a specific interest in the issue since their traditional lands have become integrated over time into the large-scale production of coca, opium poppy, and cannabis crops, in response to high demand from the American and European markets, among others. As a consequence, indigenous peoples are persecuted because of their traditional use of these and other plant-based narcotics and hallucinogens. They are also victims of the drug producers who remove them from their lands or forcibly recruit them into the production process. As indigenous peoples are caught in the violent world of illicit drug production, law enforcement often targets them first, resulting in disproportionate rates of criminalization and incarceration.
Indigenous Technological Innovation : Capability and Competitiveness in China's ... IDRC and key partners will showcase critical work on adaptation and ... Call for new OWSD Fellowships for Early Career Women Scientists now open.
This article examines to what extent the recently adopted United Nations Declarations on the Rights of Indigenous Peoples participate to the development of indigenous peoples' international human rights.
Caspeta, Luis; Nielsen, Jens
trends in systems biology of Aspergillus and Pichia species, highlighting the relevance of these developments for systems metabolic engineering of these organisms for the production of hydrolytic enzymes, biofuels and chemicals from biomass. Metabolic engineering is moving from traditional methods...... for the production of hydrolytic enzymes, biofuels and chemicals from biomass. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim....
Zhang, Wenhui; Smith, Leonard A; Plantz, Bradley A; Schlegel, Vicki L; Meagher, Michael M
The methylotrophic yeast Pichia pastoris is an effective system for recombinant protein productions that utilizes methanol as an inducer, and also as carbon and energy source for a Mut(+) (methanol utilization plus) strain. Pichia fermentation is conducted in a fed-batch mode to obtain a high cell density for a high productivity. An accurate methanol control is required in the methanol fed-batch phase (induction phase) in the fermentation. A simple "on-off" control strategy is inadequate for precise control of methanol concentrations in the fermentor. In this paper we employed a PID (proportional, integral and derivative) control system for the methanol concentration control and designed the PID controller settings on the basis of a Pichia growth model. The closed-loop system was built with four components: PID controller, methanol feed pump, fermentation process, and methanol sensor. First, modeling and transfer functions for all components were derived, followed by frequency response analysis, a powerful method for calculating the optimal PID parameters K(c) (controller gain), tau(I) (controller integral time constant), and tau(D) (controller derivative time constant). Bode stability criteria were used to develop the stability diagram for evaluating the designed settings during the entire methanol fed-batch phase. Fermentations were conducted using four Pichia strains, each expressing a different protein, to verify the control performance with optimal PID settings. The results showed that the methanol concentration matched the set point very well with only small overshoot when the set point was switched, which indicated that a very good control performance was achieved. The method developed in this paper is robust and can serve as a framework for the design of other PID feedback control systems in biological processes.
Frank K. Agbogbo; Guillermo Coward-Kelly; Mads Torry-Smith; Kevin Wenger; Thomas W. Jeffries
Xylose was fermented using Pichia stipitis CBS 6054 at different initial cell concentrations. A high initial cell concentration increased the rate of xylose utilization, ethanol formation, and the ethanol yield. The highest ethanol concentration of 41.0 g/L and a yield of 0.38 g/g was obtained using an initial cell concentration of 6.5 g/L. Even though more xylitol was...
Sun, L W; Zhao, Y; Jiang, R; Song, Y; Feng, H; Feng, K; Niu, L P; Qi, C
Pichia secretive expression system is one of powerful eukaryotic expression systems in genetic engineering, which is especially suitable for industrial utilization. Because of the low concentration of the target protein in initial experiment, the methods and conditions for expression of the target protein should be optimized according to the protein yield repetitively. It is necessary to set up a rapid, simple and convenient analysis method for protein expression levels instead of the generally used method such as ultrafiltration, purification, dialysis, lyophilization and so on. In this paper, acetone precipitation method was chosen to concentrate the recombinant protein firstly after comparing with four different protein precipitation methods systematically, and then the protein was analyzed by SDS-Polyacrylamide Gel Electrophoresis. The recombinant protein was determined with the feature of protein band by the Automated Image Capture and 1-D Analysis Software directly. With this method, the optimized expression conditions of basic fibroblast growth factor secreted from pichia were obtained, which is as the same as using traditional methods. Hence, a convenient tool to determine the optimized conditions for the expression of recombinant proteins in Pichia was established.
Acourene, S; Ammouche, A
The present study deals with submerged ethanol, citric acid, and α-amylase fermentation by Saccharomyces cerevisiae SDB, Aspergillus niger ANSS-B5, and Candida guilliermondii CGL-A10, using date wastes as the basal fermentation medium. The physical and chemical parameters influencing the production of these metabolites were optimized. As for the ethanol production, the optimum yield obtained was 136.00 ± 0.66 g/l under optimum conditions of an incubation period of 72 h, inoculum content of 4% (w/v), sugars concentration of 180.0 g/l, and ammonium phosphate concentration of 1.0 g/l. Concerning citric acid production, the cumulative effect of temperature (30°C), sugars concentration of 150.0 g/l, methanol concentration of 3.0%, initial pH of 3.5, ammonium nitrate concentration of 2.5 g/l, and potassium phosphate concentration of 2.5 g/l during the fermentation process of date wastes syrup did increase the citric acid production to 98.42 ± 1.41 g/l. For the production of α-amylase, the obtained result shows that the presence of starch strongly induces the production of α-amylase with a maximum at 5.0 g/l. Among the various nitrogen sources tested, urea at 5.0 g/l gave the maximum biomass and α-amylase estimated at 5.76 ± 0.56 g/l and 2,304.19 ± 31.08 μmol/l/min, respectively after 72 h incubation at 30°C, with an initial pH of 6.0 and potassium phosphate concentration of 6.0 g/l.
Hopkins, Shelley; Sampson, Geoff P; Hendicott, Peter L; Wood, Joanne M
Little is known about the prevalence of refractive error, binocular vision, and other visual conditions in Australian Indigenous children. This is important given the association of these visual conditions with reduced reading performance in the wider population, which may also contribute to the suboptimal reading performance reported in this population. The aim of this study was to develop a visual profile of Queensland Indigenous children. Vision testing was performed on 595 primary schoolchildren in Queensland, Australia. Vision parameters measured included visual acuity, refractive error, color vision, nearpoint of convergence, horizontal heterophoria, fusional vergence range, accommodative facility, AC/A ratio, visual motor integration, and rapid automatized naming. Near heterophoria, nearpoint of convergence, and near fusional vergence range were used to classify convergence insufficiency (CI). Although refractive error (Indigenous, 10%; non-Indigenous, 16%; p = 0.04) and strabismus (Indigenous, 0%; non-Indigenous, 3%; p = 0.03) were significantly less common in Indigenous children, CI was twice as prevalent (Indigenous, 10%; non-Indigenous, 5%; p = 0.04). Reduced visual information processing skills were more common in Indigenous children (reduced visual motor integration [Indigenous, 28%; non-Indigenous, 16%; p < 0.01] and slower rapid automatized naming [Indigenous, 67%; non-Indigenous, 59%; p = 0.04]). The prevalence of visual impairment (reduced visual acuity) and color vision deficiency was similar between groups. Indigenous children have less refractive error and strabismus than their non-Indigenous peers. However, CI and reduced visual information processing skills were more common in this group. Given that vision screenings primarily target visual acuity assessment and strabismus detection, this is an important finding as many Indigenous children with CI and reduced visual information processing may be missed. Emphasis should be placed on identifying
Full Text Available This paper seeks to provide an initial theoretical grounding to assess a practical project: a new software application that attempts to be a beneficial resource in the field of Indigenous representation. As a starting point, we are concerned to provide a theoretical ground for considering the inherited and shifting spaces of Indigenous media representation. To this end, this paper reconsiders the strengths and weaknesses of debates surrounding the ‘Indigenous public sphere’. This is used as grounds for critically understanding the relations that constitute this field. Following this, we consider how a more materialist approach to publics might enable a productive reconceptualization, and in particular how digital media initiatives and shifting news markets may be contributing to change. Finally, drawing on this model, we outline both the ‘Wakul app’ project, and how this framework might inform an assessment of its impact.
Indilinga: African Journal of Indigenous Knowledge Systems ... Yet in Africa, where traditional religions and thought systems of the indigenous people of Africa were formerly ... and the active participation of respondents in the research process.
Indigenous games are an integral component of indigenous knowledge systems. ... and national activities; mathematical concepts associated with the games; possibilities and implications for general classroom ... AJOL African Journals Online.
Faculty of Education, University of South Africa, P.O. Box 392, Unisa, 0003 South Africa ... Indigenous Knowledge also termed Traditional, Endogenous or Classical .... its civilisation, carries both its indigenous and modern knowledge systems.
This chapter describes an innovative program that weaves together adult learning, transformative education, and indigenous epistemology in order to prepare Alaskan rural indigenous social service providers to better serve their communities.
Curriculum enrichment through indigenous Zulu games. ... 1997). The aim of the study was to document and analyze indigenous Zulu games for possible curriculum enrichment of physical ... AJOL African Journals Online. HOW TO USE AJOL.
Cordia millenii, Ficus spp, Markhamia lutea and Albizia spp are the most commonly used indigenous ... activities like construction of roads and expansion of ranches and ... impact of traditional uses of indigenous tress on the sustainability.
Whitinui, Paul; McIvor, Onowa; Robertson, Boni; Morcom, Lindsay; Cashman, Kimo; Arbon, Veronica
There is an Indigenous resurgence in education occurring globally. For more than a century Euro-western approaches have controlled the provision and quality of education to, and for Indigenous peoples. The World Indigenous Research Alliance (WIRA) established in 2012, is a grass-roots movement of Indigenous scholars passionate about making a…
Li, Peter Ping
It has long been recognized that indigenous research should be helpful, if not essential, for an adequate understanding of local phenomena. The indigenous approach is consistent with, but extends beyond, the repeated calls for contextualizing management and organization research. However, the cha......It has long been recognized that indigenous research should be helpful, if not essential, for an adequate understanding of local phenomena. The indigenous approach is consistent with, but extends beyond, the repeated calls for contextualizing management and organization research. However...
Rogers, Wayne P.; Sture, Stein
The utilization of local resources for the construction and operation of a lunar base can significantly reduce the cost of transporting materials and supplies from Earth. The feasibility of processing lunar regolith to form construction materials and structural components is investigated. A preliminary review of potential processing methods such as sintering, hot-pressing, liquification, and cast basalt techniques, was completed. The processing method proposed is a variation on the cast basalt technique. It involves liquification of the regolith at 1200-1300 C, casting the liquid into a form, and controlled cooling. While the process temperature is higher than that for sintering or hot-pressing (1000-1100 C), this method is expected to yield a true engineering material with low variability in properties, high strength, and the potential to form large structural components. A scenario for this processing method was integrated with a design for a representative lunar base structure and potential construction techniques. The lunar shelter design is for a modular, segmented, pressurized, hemispherical dome which could serve as habitation and laboratory space. Based on this design, estimates of requirements for power, processing equipment, and construction equipment were made. This proposed combination of material processing method, structural design, and support requirements will help to establish the feasibility of lunar base construction using indigenous materials. Future work will refine the steps of the processing method. Specific areas where more information is needed are: furnace characteristics in vacuum; heat transfer during liquification; viscosity, pouring and forming behavior of molten regolith; design of high temperature forms; heat transfer during cooling; recrystallization of basalt; and refinement of estimates of elastic moduli, compressive and tensile strength, thermal expansion coefficient, thermal conductivity, and heat capacity. The preliminary
Bunten, Alexis Celeste
Through a comparison of Indigenous-owned cultural tourism businesses in southeastern Alaska and New Zealand as well as secondary data examining Indigenous tourism across the Pacific, this article introduces the concept of "Indigenous capitalism" as a distinct strategy to achieve ethical, culturally appropriate, and successful Indigenous…
Gandrup, Tobias; Jespersgaard Jakobsen, Line
This project is set out to analyse the negotiation of indigeneity. This will be done by unfolding the semiotic practices of two organisations that represents indigenous interests in contemporary Peruvian politics. It examines the rise of the term indigeneity in international politics through the emergence of an international framework and asks to how this has shaped political possibilities for the local indigenous organisations to represent the indigenous interests. The analysis shows that th...
Indigenous species of actinorhizal plants of Casuarinaceae, Elaeagnaceae and Rhamnaceae are found in specific regions of Australia. Most of these plants belong to Casuarinaceae, the dominant actinorhizal family in Australia. Many of them have significant environmental and economical value. The other two families with ...
... as determined by a Steering Committee of experts drawn from government agencies, universities and research institutions all over the country. It is expected to generate a body of evidence that will aid Chinese policymakers to develop and implement effective policies for enhancing indigenous innovations in the west.
An ethnomedicinal study was conducted in the remote Hindukush-Himalayan valleys of Utror and Gabral, during which 36 common folk medicinal recipes of the area were documented. The indigenous methods of medicinal plants collection and their further processing were also explored. It was also observed that huge ...
Maasz, Donovan; Winschiers-Theophilus, Heike; Stanley, Colin
Indigenous Knowledge (IK) preservation and management has been taken up as a serious endeavor by various governments who have realized the value of IK as well as the opportunities given by emerging technologies. Considering the various phases and activities of indigenous knowledge management which...... the indigenous knowledge digitization process, namely, codesign, conceptualization, collection, correction, curation, circulation, and creation of knowledge. We exemplify the application of the model with technologies currently developed under an indigenous knowledge holder’s toolkit promoting the agency...... of digitalizing indigenous knowledge across the phases....
Eom, Gyeong Tae; Lee, Seung Hwan; Song, Bong Keun; Chung, Keun-Wo; Kim, Young-Wun; Song, Jae Kwang
The gene encoding lipase B from Candida antarctica (CalB) was expressed in Pichia pastoris after it was synthesized by the recursive PCR and cloned into the Pichia expression plasmid, pPICZαA. The CalB was successfully secreted in the recombinant P. pastoris strain X-33 with an apparent molecular weight of 34 kDa. For 140 h flask culture, the dry cell weight and the extracellular lipase activity reached at 5.4 g/l and 57.9 U/l toward p-nitrophenyl palmitate, respectively. When we performed the fed-batch fermentation using a methanol feeding strategy for 110 h, the dry cell weight and the extracellular lipase activity were increased to 135.7 g/l and 11,900 U/l; the CalB protein concentration was 1.18 g/l of culture supernatant. The characteristics of CalB recovered from the P. pastoris culture were compared with the commercial form of CalB produced in Aspergillus oryzae. The kinetic constants and specific activity, the effects of activity and stability on temperature and pH, the glycosylation extent, the degree of immobilization on macroporous resin and the yield of esterification reaction between oleic acid and n-butanol were almost identical to each other. Therefore, we successfully proved that the Pichia-based expression system for CalB in this study was industrially promising compared with one of the most efficient production systems. Copyright © 2013 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.
Zhou, Chenyan; Wang, Yongtao; Wu, Minchen; Wang, Wu; Li, Dongfeng
To efficiently produce xylanase for food processing industry, a gene encoding xylanase II (XynII) from Aspergillus usamii has been cloned into the vector pPIC9K and integrated into the genome of Pichia pastoris KM71 by electroporation. By means of minimal dextrose (MD) plates and PCR, the recombinant P. pastoris strains (His+Muts) have been obtained. Activity assay and SDS-PAGE demonstrate that XynII was extracellularly expressed in P. pastoris with the induction of methanol. In shake flask c...
Clark, Lindsay; Zahm, Jacob A.; Ali, Rustam; Kukula, Maciej; Bian, Liangqiao; Patrie, Steven M.; Gardner, Kevin H.; Rosen, Michael K.; Rosenbaum, Daniel M.
13 C Methyl TROSY NMR spectroscopy has emerged as a powerful method for studying the dynamics of large systems such as macromolecular assemblies and membrane proteins. Specific 13 C labeling of aliphatic methyl groups and perdeuteration has been limited primarily to proteins expressed in E. coli, preventing studies of many eukaryotic proteins of physiological and biomedical significance. We demonstrate the feasibility of efficient 13 C isoleucine δ1-methyl labeling in a deuterated background in an established eukaryotic expression host, Pichia pastoris, and show that this method can be used to label the eukaryotic protein actin, which cannot be expressed in bacteria. This approach will enable NMR studies of previously intractable targets
Micheelsen, Pernille Ollendorff; Ostergaard, Peter Rahbek; Lange, Lene
An expression system for high-level expression of the native Hordeum vulgare alpha-amylase/subtilisin inhibitor (BASI) has been developed in Pichia pastoris, using the methanol inducible alcohol oxidase 1 (AOX1) promoter. To optimize expression, two codon-optimized coding regions have been designed...... and expressed alongside the wild-type coding region. To ensure secretion of the native mature protein, a truncated version of the alpha mating factor secretion signal from Saccharomyces cerevisiae was used. In order to be able to compare expression levels from different clones, single insertion transformants...
Kang, Zhen; Huang, Hao; Zhang, Yunfeng; Du, Guocheng; Chen, Jian
Pichia pastoris: (reclassified as Komagataella phaffii), a methylotrophic yeast strain has been widely used for heterologous protein production because of its unique advantages, such as readily achievable high-density fermentation, tractable genetic modifications and typical eukaryotic post-translational modifications. More recently, P. pastoris as a metabolic pathway engineering platform has also gained much attention. In this mini-review, we addressed recent advances of molecular toolboxes, including synthetic promoters, signal peptides, and genome engineering tools that established for P. pastoris. Furthermore, the applications of P. pastoris towards synthetic biology were also discussed and prospected especially in the context of genome-scale metabolic pathway analysis.
Cindy Smithers Graeme
Full Text Available Employing a reflexive and co-constructed narrative analysis, this article explores our experiences as a non-Indigenous doctoral student and a First Nations research assistant working together within the context of a community-based participatory Indigenous geography research project. Our findings revealed that within the research process there were experiences of conflict, and opportunities to reflect upon our identity and create meaningful relationships. While these experiences contributed to an improved research process, at a broader level, we suggest that they also represented our personal stories of reconciliation. In this article, we share these stories, specifically as they relate to reconciliatory processes of re-education and cultural regeneration. We conclude by proposing several policy recommendations to support research as a pathway to reconciliation in Canada.
Full Text Available This paper provides an overview of discourses of the movement for national reconciliation prevailing within the Australian socio-political context since the inception of the Council for Aboriginal Reconciliation in 1991, to the national apology delivered by the Prime Minister Kevin Rudd on 13th February 2008. It provides an framework for the various discourses of reconciliation, by exploring and analysing the accrued meanings to such terms such as ‘genuine’, substantive or ‘true’ reconciliation; the Howard’s Government’s ‘practical reconciliation’ and the Rudd government’s great attempt at ‘symbolic’ reconciliation in the national apology to Indigenous Australians. In the changing political context in Australia today this paper revisits the debates on reconciliation, and endeavours to locate the movement solidly within a human rights framework that includes first nation rights. This requires an examination of the roots of the reconciliation movement including community attitudes to reconciliation and the nature of the peoples’ movement as well as the differing perspectives of policy makers, politicians and of course, Indigenous peoples. It asks crucial questions about the progress of reconciliation and the type of reconciliation mainstream Australians will accept. In truth therefore, was the ‘National Apology’ a grand symbolic gesture by mainstream Australia to maintain the status quo and divert our eyes from the more searching questions of the ‘unfinished business’ of ‘substantive’ reconciliation which encompasses first nations rights for Indigenous peoples.
Marchand, Dawn Marie
Thousands of indigenous people from across North America came to the Enoch Cree Nation for the Alberta Indigenous Games, six days of sport, education, and cultural awakening. The vision of the Alberta Indigenous Games is to recognize the value and potential of Indigenous culture and the young people. Activities include sports, indigenous arts,…
Liu Wenping; Du Jingling
The article explains the significance of Library Science Indigenization, Answer some misunderstanding of Library Science Indigenization,reveals express form of Library Science Indigenization, Discusses criteria of Library Science Indigenization, finally give some suggestions and methods of Library Science Indigenization. (authors)
Full Text Available Indigenous Australians experience a high prevalence of sexual assault, yet a regional sexual assault service found few Indigenous Australians accessed their services. This prompted exploration of how its services might be improved. A resultant systematic search of the literature is reported in this article. Seven electronic databases and seven websites were systematically searched for peer reviewed and gray literature documenting responses to the sexual assault of Indigenous Australians. These publications were then classified by response type and study type. Twenty-three publications met the inclusion criteria. They included studies of legal justice, media, and community-based and mainstream service responses for Indigenous survivors and perpetrators. We located program descriptions, measurement, and descriptive research, but no intervention studies. There is currently insufficient evidence to confidently prescribe what works to effectively respond to Indigenous Australian sexual assault. The study revealed an urgent need for researchers, Indigenous communities, and services to work together to develop the evidence base.
Amaya-Delgado, L; Herrera-López, E J; Arrizon, Javier; Arellano-Plaza, M; Gschaedler, A
Traditionally, industrial tequila production has used spontaneous fermentation or Saccharomyces cerevisiae yeast strains. Despite the potential of non-Saccharomyces strains for alcoholic fermentation, few studies have been performed at industrial level with these yeasts. Therefore, in this work, Agave tequilana juice was fermented at an industrial level using two non-Saccharomyces yeasts (Pichia kluyveri and Kluyveromyces marxianus) with fermentation efficiency higher than 85 %. Pichia kluyveri (GRO3) was more efficient for alcohol and ethyl lactate production than S. cerevisiae (AR5), while Kluyveromyces marxianus (GRO6) produced more isobutanol and ethyl-acetate than S. cerevisiae (AR5). The level of volatile compounds at the end of fermentation was compared with the tequila standard regulation. All volatile compounds were within the allowed range except for methanol, which was higher for S. cerevisiae (AR5) and K. marxianus (GRO6). The variations in methanol may have been caused by the Agave tequilana used for the tests, since this compound is not synthesized by these yeasts.
Wood, Matthew J.; Komives, Elizabeth A.
Heterologous expression in Pichia pastoris has many of the advantages of eukaryotic expression, proper folding and disulfide bond formation, glycosylation, and secretion. Contrary to other eukaryotic systems, protein production from P.pastoris occurs in simple minimal defined media making this system attractive for production of labeled proteins for NMR analysis. P.pastoris is therefore the expression system of choice for NMR of proteins that cannot be refolded from inclusion bodies or that require post-translational modifications for proper folding or function. The yield of expressed proteins from P.pastoris depends critically on growth conditions, and attainment of high cell densities by fermentation has been shown to improve protein yields by 10-100-fold. Unfortunately, the cost of the isotopically enriched fermentation media components, particularly 15NH4OH, is prohibitively high. We report fermentation methods that allow for both 15N- labeling from (15NH4)2SO4 and 13C-labeling from 13C-glucose or 13C-glycerol of proteins produced in Pichia pastoris. Expression of an 83 amino acid fragment of thrombomodulin with two N-linked glycosylation sites shows that fermentation is more cost effective than shake flask growth for isotopic enrichment
Chahardooli, Mahmood; Niazi, Ali; Aram, Farzaneh; Sohrabi, Seyyed Mohsen
Lactoferricin (LFcin) is a strong cationic peptide released from the N-terminus of lactoferrin by gastric pepsin digestion. LFcin has some important properties, including high antimicrobial activity. To date, lactoferricins have been isolated and characterised from various animal species, but not from camel. The aim of this study was to characterise and express recombinant camel lactoferricin (LFcinC) in Pichia pastoris and investigate its antimicrobial activity. After methanol induction, LFcinC was expressed and secreted into a culture broth medium and the results determined by concentrated supernatant culture medium showed high antimicrobial activity against the following microorganisms: Escherichia coli PTCC 1330 (ATCC 8739), Staphylococcus aureus PTCC 1112 (ATCC 6538), Pseudomonas aeruginosa PTCC 1074 (ATCC 9027), Bacillus subtilis PTCC 1023 (ATCC 6633), and Candida albicans PTCC 5027 (ATCC 10231). Thermal stability was clarified with antibacterial activity against Escherichia coli PTCC 1330 (ATCC 8739). Results confirmed that camel lactoferricin had suitable antimicrobial activity and its production by Pichia pastoris can be used for recombinant production. © 2015 Society of Chemical Industry.
Kim, Sun-Il; Wu, Yuanzheng; Kim, Ka-Lyun; Kim, Geun-Joong; Shin, Hyun-Jae
An efficient method for Pichia cell disruption that employs an aminopropyl magnesium phyllosilicate (AMP) clay-assisted glass beads mill is presented. AMP clay is functionalized nanocomposite resembling the talc parent structure Si8Mg6O20(OH)4 that has been proven to permeate the bacterial membrane and cause cell lysis. The recombinant capsid protein of cowpea chlorotic mottle virus (CCMV) expressed in Pichia pastoris GS115 was used as demonstration system for their ability of self-assembly into icosahedral virus-like particles (VLPs). The total protein concentration reached 4.24 mg/ml after 4 min treatment by glass beads mill combined with 0.2 % AMP clay, which was 11.2 % higher compared to glass beads mill only and the time was half shortened. The stability of purified CCMV VLPs illustrated AMP clay had no influence on virus assembly process. Considering the tiny amount added and simple approach of AMP clay, it could be a reliable method for yeast cell disruption.
Mei, Xiang; Du, Renqian; Li, Xi; Huang, Haiyan; Yu, Min; Tang, Qiqun
Adiponectin is an adipokine predominantly synthesized and secreted by adipocytes in the white adipose tissue, and it can lower the blood glucose level and increase free fatty acid oxidation. In the current study, we developed the globular domain of adiponectin (gapM1) to treat type II diabetes. In both flask and fermentor, we cultivated Pichia pastoris expressing recombinant gapM1 and established the purification procedure by using gel filtration and anion exchange chromatography. To evaluate the biological activity of recombinant gapM1, we used rat type II diabetes model fed high-fat diet in combination with low-dose STZ (Streptozocin) induction. We purified 200 mg gapM1 with purity of 96% from 10 liters of supernatant. The recombinant gapM1 significantly lowered blood glucose (34.2%), serum triglyceride (79.6%) and total cholesterol (62.1%) in type II diabetes induced rat. Therefore, the recombinant human gapM1 is successfully expressed in Pichia pastoris and effectively treated type II diabetes in rat models.
Kato, Saemi; Shimizu-Ibuka, Akiko; Mura, Kiyoshi; Takeuchi, Akiko; Tokue, Chiyoko; Arai, Soichi
An alpha-amylase secreted by Pichia burtonii 15-1 isolated from a traditional starter murcha of Nepal, named Pichia burtonii alpha-amylase (PBA), was studied. The gene was cloned and its nucleotide sequence was determined. PBA was deduced to consist of 494 amino acid residues. It shared certain degrees of amino acid sequence identity with other homologous proteins: 60% with Schwanniomyces occidentalis alpha-amylase, 58% with Saccharomycopsis sp. alpha-amylase, and 47% with Taka-amylase A from Aspergillus oryzae. A three-dimensional structural model of PBA generated using the known three-dimensional structure of Taka-amylase A as a template suggested high structural similarity between them. Kinetic analysis revealed that the K(m) values of PBA were lower than those of Taka-amylase A for the oligosaccharides. Although the k(cat) values of PBA were lower than those of Taka-amylase A for the oligosaccharide substrates, the k(cat)/K(m) values of PBA were higher.
Full Text Available Filamentous fungi are the predominant source of lignocellulolytic enzymes used in industry for the transformation of plant biomass into high-value molecules and biofuels. The rapidity with which new fungal genomic and post-genomic data are being produced is vastly outpacing functional studies. This underscores the critical need for developing platforms dedicated to the recombinant expression of enzymes lacking confident functional annotation, a prerequisite to their functional and structural study. In the last decade, the yeast Pichia pastoris has become increasingly popular as a host for the production of fungal biomass-degrading enzymes, and particularly carbohydrate-active enzymes (CAZymes. This study aimed at setting-up a platform to easily and quickly screen the extracellular expression of biomass-degrading enzymes in Pichia pastoris. We first used three fungal glycoside hydrolases that we previously expressed using the protocol devised by Invitrogen to try different modifications of the original protocol. Considering the gain in time and convenience provided by the new protocol, we used it as basis to set-up the facility and produce a suite of fungal CAZymes (glycoside hydrolases, carbohydrate esterases and auxiliary activity enzyme families out of which more than 70% were successfully expressed. The platform tasks range from gene cloning to automated protein purifications and activity tests, and is open to the CAZyme users’ community.
Prabhu, Ashish A; Boro, Bibari; Bharali, Biju; Chakraborty, Shuchishloka; Dasu, V Venkata
Process development involving system metabolic engineering and bioprocess engineering has become one of the major thrust for the development of therapeutic proteins or enzymes. Pichia pastoris has emerged as a prominent host for the production of therapeutic protein or enzymes. Despite of producing high protein titers, various cellular and process level bottlenecks hinders the expression of recombinant proteins in P. pastoris. In the present review, we have summarized the recent developments in the expression of foreign proteins in P. pastoris. Further, we have discussed various cellular engineering strategies which include codon optimization, pathway engineering, signal peptide processing, development of protease deficient strain and glyco-engineered strains for the high yield protein secretion of recombinant protein. Bioprocess development of recombinant proteins in large scale bioreactor including medium optimization, optimum feeding strategy and co-substrate feeding in fed batch as well as continuous cultivation have been described. The recent advances in system and synthetic biology studies including metabolic flux analysis in understanding the phenotypic characteristics of recombinant Pichia and genome editing with CRISPR-CAS system have also been summarized. Copyright© Bentham Science Publishers; For any queries, please email at email@example.com.
Marquard, D; Enders, A; Roth, G; Rinas, U; Scheper, T; Lindner, P
In situ Microscopy (ISM) is an optical non-invasive technique to monitor cells in bioprocesses in real-time. Pichia pastoris is one of the most promising protein expression systems. This yeast combines fast growth on simple media and important eukaryotic features such as glycosylation. In this work, the ISM technology was applied to Pichia pastoris cultivations for online monitoring of the cell concentration during cultivation. Different ISM settings were tested. The acquired images were analyzed with two image processing algorithms. In seven cultivations the cell concentration was monitored by the applied algorithms and offline samples were taken to determine optical density (OD) and dry cell mass (DCM). Cell concentrations up to 74g/L dry cell mass could be analyzed via the ISM. Depending on the algorithm and the ISM settings, an accuracy between 0.3 % and 12 % was achieved. The overall results show that for a robust measurement a combination of the two described algorithms is required. Copyright © 2016 Elsevier B.V. All rights reserved.
Full Text Available Optimising Pichia onychis yeast biomass production was evaluated using different substrates and different physicochemical conditions for liquid fermentation. The Plackett-Burman statistical design was initially applied for screening the most important nutritional variables (three carbon sources and eight nitrogen sources affecting yeast biomass production. Four nutritional sources and two physicochemical variables were subsequently evaluated using a factorial fractionated design as the starting point for optimising the process by applying a central composite rotational design. The results obtained f rom employing a polynomial regression model using the experimental data showed that biomass production was strongly affected by nutritional and physicochemical conditions. The highest yield was obtained in the following conditions: 43,42 g/L carbon source, 0,261 g/L nitrogen organic source, shaking at 110 rpm, 6,0 pH, 48 h total fermentation time during which 8,95 XlO9 cells/mL were obtained, equivalent to 6,30 g/L dry biomass. Key words: Pichia onychis, optimisation, liquid fermentation.
In 2010 and 2011, foreign businesses and governments welcomed measures believed to dramatically reform a highly controversial branch of China’s indigenous innovation policy which provided government procurement preferences to applicants who can meet restrictive indigenous intellectual property (IP) rights requirements. However, this article describes specific examples of (what can be labeled) China’s “indigenous IP policy” that are still very much in force, in particular several programs link...
The Indigenous education agenda in Australia remains focused on mainstream schooling contexts. Although overlooked in Indigenous education discourse, flexi schools appear to be engaging with disproportionately high numbers of Indigenous students and staff. The educative roles of Indigenous peoples in broader Indigenous education discourse are…
Juge, N; Andersen, Jens S.; Tull, D
, and 2-chloro-4-nitrophenyl beta-D-maltoheptaoside as well as in Ca2+ dependency of activity. Pichia pastoris thus produced in high yields recombinant alpha-amylase that is similar with respect to structure and function to the enzyme purified from malt extracts. This greatly facilitates future mutational...
Rita C.L.B. Rodrigues; Chenfeng Lu; Bernice Liu; Thomas W. Jeffries
Spent sulfite pulping liquor (SSL) contains lignin, which is present as lignosulfonate, and hemicelluloses that are present as hydrolyzed carbohydrates. To reduce the biological oxygen demand of SSL associated with dissolved sugars, we studied the capacity of Pichia stipitis FPL-YS30 (xyl3[delta]) to convert these sugars into useful products. FPL-YS30 produces a...
Full Text Available Abstract Abstract: This paper examines the possibilities and limitations of an emergent global discourse of indigeneity to offer an oppositional praxis in the face of the depredations of settler colonialism in post-apartheid South Africa. Self-conscious articulations of indigeneity, we argue, reveal the fraught relationship between increasingly hegemonic and narrow understandings of the indigenous and the carceral logic of apartheid. We examine this by focusing on the meanings and attachments forged through indigenous plants in two realms: the world of indigenous gardening practised by white suburban dwellers and that of subsistence farming undertaken by rural black women. This juxtaposition reveals that in contrast to the pervasive resurrection of colonial time that defines metropolitan indigenous gardening, the social relations of a subsistence cultivator challenge the confines of colonial temporality, revealing a creative mode of dissent structured around dreams, ancestral knowledge, and the commons. Our exploration of struggles around botanical indigeneity suggests that anticolonial modes of indigeneity do not necessarily inhere in recognisable forms and that studies of the indigenous need to proceed beyond those that bear familial resemblance to emergent global understandings.
Indigenous knowledge is multidimensional encompassing the beliefs, practices, arts, spirituality and other forms of traditional and cultural experiences that belong to Indigenous communities globally. In order to protect, preserve and recognize the knowledge of the Indigenous people of Fiji, known as the iTaukei, the University of Fiji has…
Heather A. Knauer
Full Text Available Parenting may be influenced by ethnicity; marginalization; education; and poverty. A critical but unexamined question is how these factors may interact to compromise or support parenting practices in ethnic minority communities. This analysis examined associations between mothers’ stimulating parenting practices and a range of child-level (age; sex; and cognitive and socio-emotional development; household-level (indigenous ethnicity; poverty; and parental education; and community-level (economic marginalization and majority indigenous population variables among 1893 children ages 4–18 months in poor; rural communities in Mexico. We also explored modifiers of associations between living in an indigenous community and parenting. Key findings were that stimulating parenting was negatively associated with living in an indigenous community or family self-identification as indigenous (β = −4.25; SE (Standard Error = 0.98; β = −1.58; SE = 0.83 respectively. However; living in an indigenous community was associated with significantly more stimulating parenting among indigenous families than living in a non-indigenous community (β = 2.96; SE = 1.25. Maternal education was positively associated with stimulating parenting only in indigenous communities; and household crowding was negatively associated with stimulating parenting only in non-indigenous communities. Mothers’ parenting practices were not associated with child sex; father’s residential status; education; or community marginalization. Our findings demonstrate that despite greater community marginalization; living in an indigenous community is protective for stimulating parenting practices of indigenous mothers.
Magson, Natasha R.; Craven, Rhonda G.; Nelson, Genevieve F.; Yeung, Alexander S.; Bodkin-Andrews, Gawaian H.; McInerney, Dennis M.
This research explored gender and cross-cultural similarities and differences in the motivational profiles of Indigenous Papua New Guinean (PNG) and Indigenous and non-Indigenous Australians. Secondary students (N = 1,792) completed self-report motivational measures. Invariance testing demonstrated that the Inventory of School Motivation…
Full Text Available BACKGROUND: Genetic research has potential benefits for improving health, such as identifying molecular characteristics of a disease, understanding disease prevalence and treatment, and developing treatments tailored to patients based on individual genetic characteristics of their disease. Indigenous people are often targeted for genetic research because genes are easier to study in communities that practice endogamy. Therefore, populations perceived to be more homogenous, such as Indigenous peoples, are ideal for genetic studies. While Indigenous communities remain the focal point of many genomic studies, some result in harm and unethical practice. Unfortunately, the harms of poorly formulated and unethical research involving Indigenous people have created barriers to participation that prevent critical and lifesaving research. These harms have led a number of Indigenous communities to develop guidelines for engaging with researchers to assist in safely bridging the gap between genetic research and Indigenous peoples.SPECIFIC AIMS: The specific aims of this study were: (1 to conduct an international review and comparison of Indigenous research guidelines that highlight topics regarding genetics and use of biological samples and identify commonalities and differences among ethical principles of concern to Indigenous peoples; and (2 develop policy recommendations for Indigenous populations interested in creating formal policies around the use of genetic information and protection of biological samples using data from specific aim 1.METHODS: A comparative analysis was performed to identify best research practices and recommendations for Indigenous groups from four countries: Canada, New Zealand, Australia, and the United States. The analysis examined commonalities in political relationships, which support self-determination among these Indigenous communities to control their data. Current international Indigenous guidelines were analyzed to review
The importance of indigenous games: The selected cases of Indigenous games in South Africa. ... do not have enough time to transfer their skills and knowledge of indigenous games to the younger generation. ... AJOL African Journals Online.
Integrating indigenous games and knowledge into Physical Education: Implications for ... The aim of this study was to analyse indigenous Zulu games towards integrating indigenous game skill and knowledge ... AJOL African Journals Online.
M. H. Mahamad Maifiah
Full Text Available Phytases catalyze the hydrolysis of phytate (myo-inositol hexakisphosphate, one of the major storage form of phosphate in plants, with subsequent release of myo-inositol, phosphate and phytate-bound minerals. Non-ruminant animals such as chicken, swine and fish can't use the organic phosphorus and minerals from their diet because there is no phytase activity in their digestive tract. Phytate degrading enzyme is added to the animal feed diet to improve phosphorus availability from the dietary phytate and at the same time this lessen the phosphate pollution level in areas of intensive animal production as the phosphate would not be excreted out to the environment. ASUIA279, a bacterial strain isolated from Malaysian soil has potentially shown good phytase activity. In the present work, the gene encoding for phytase has been amplified from the plasmid DNA of recombinant ASUIA279(5 obtained from a previous study (unpublished data by a polymerase chain reaction (PCR methodology. The amplified phytase gene was extracted, purified then cloned into the pPICZA plasmid and transformed into Pichia pastorisX-33 strain for enzyme production.ABSTRAK: Fitase pemangkinan hidrolisis fitat (myo-inositol hexakisphosphate, merupakan salah satu cara penyimpanan utama fosfat dalam tumbuhan, dengan pelepasan berturut myo-inositol , fosfat dan galian terikat fitat. Haiwan bukan ruminan seperti ayam, khinzir dan ikan tidak dapat memanfaatkan fosforus organik serta galian yang diperolehi daripada makanan kerana tidak mempunyai aktiviti fitase di dalam saluran pencernaan mereka. Enzim pengecilan fitase dicampurkan ke dalam pemakanan haiwan untuk mempertingkatkan keperolehan fosforus dari fitat diet. Pada masa yang sama ia dapat mengurangkan tahap pencemaran fosfat di kawasan yang terdapat penternakan haiwan secara intensif agar fosfat tidak dikumuhkan ke persekitaran. ASUIA279, satu strain bakteria yang diasingkan daripada tanih di Malaysiamenunjukkan aktiviti fitase
The purpose was to discover what aspects of indigenous leadership and cultural resources might be accessed and developed to influence individual behaviour as well as the prevailing community norms, values, sanctions and social controls that are related to sexual behaviour. The indigenous leaders participating in the ...
Indigenous populations are now being encouraged to be involved in the business of gambling as an operator or if not given that status, are actively encouraged to participate in gambling activities. Research both published and unpublished show that different indigenous populations often have a higher prevalence of problem and pathological gambling…
LaFrance, Joan; Nichols, Richard
The American Indian Higher Education Consortium (AIHEC), comprising 34 American Indian tribally controlled colleges and universities, has undertaken a comprehensive effort to develop an "Indigenous Framework for Evaluation" that synthesizes Indigenous ways of knowing and Western evaluation practice. To ground the framework, AIHEC engaged…
Henry, Frances; Dua, Enakshi; Kobayashi, Audrey; James, Carl; Li, Peter; Ramos, Howard; Smith, Malinda S.
This article is based on data from a four-year national study of racialization and Indigeneity at Canadian universities. Its main conclusion is that whether one examines representation in terms of numbers of racialized and Indigenous faculty members and their positioning within the system, their earned income as compared to white faculty, their…
The use of indigenous research ethics has a possibility of contextualising indigenous research. Orthodox research is guided by ethical principles which are meant to protect the institution or researcher and the participants. Despite the existence of the ethical pronouncements, literature has shown that research has proven to ...
This study was aimed at conducting ophthalmic sonographic examination of Nigerian indigenous dogs to provide baseline information on some major ocular parameters. Healthy eyes of eighty (80) indigenous dogs were used for the study. The dogs were adequately restrained physically and the structure of the ocular ...
Dudley, Mary Jo
Describes the effects on indigenous communities of Bolivia's recent Popular Participation Laws, which relocated political and financial decision making to the municipal level; community efforts toward cultural maintenance and nonformal agricultural education; the activism of indigenous university students; and the dual discrimination suffered by…
This paper seeks to understand the nature of knowledge, introduce the concept of indigenous knowledge, provide some idea of the status of Indigenous ... African professionals, scholars, researchers, policy makers and activists attempting to understand or promote IK run the risk of a cool reception, ridicule or even outright ...
Focus and Scope. Welcome to Indilinga: African Journal of Indigenous Knowledge Systems (IAJIKS). The name Indilinga: stands for the "circular orientation" of indigenous African communities which is exhibited in their material culture and behaviour. The journal has been motivated by the need for a dependable expression ...
Objective: To report cases of clinical otosclerosis histologically confirmed among indigenous South African blacks. Design: A retrospective study. Setting: Referral tertiary center, MEDUNSA, Garankuwa Hospital, South Africa. Subjects: All fifteen indigenous South African blacks diagnosed with clinical otosclerosis at ...
Ngeno, K. (2015). Breeding program for indigenous chicken in Kenya. Analysis of diversity in indigenous chicken populations. PhD thesis, Wageningen University, the Netherlands
The objective of this research was to generate knowledge required for the
Appanna, Subhashni Devi
Some Indigenous students are at risk of academic failure and science teachers have a role in salvaging these equally able students. This article firstly elucidates the research entailed in Indigenous science education in Australia and beyond. Secondly, it reviews the cultural and language barriers when learning science, faced by middle and senior…
Haptoglobin Phenotypes and Hypertension in Indigenous Zambians at the University Teaching Hospital, Lusaka, Zambia. MM Phiri, T Kaile, FM Goma. Abstract. Objectives: The aim of the study was to investigate the association between presence of haptoglobin phenotypes and hypertension in indigenous Zambian patients ...
This article examines the global debates about indigenous knowledge and Africa's traditional medicine. It explores whether it is possible to document all the elements of indigenous knowledge about Africa's traditional medicine that is used for the treatment of diverse forms of sickness. Certain types of Africa's traditional ...
Bandias, Susan; Fuller, Don; Larkin, Steven
Important recent objectives of indigenous education policy in Australia have been aimed at redressing indigenous economic and social disadvantage through increasing student retention, progression and completion rates in both compulsory and post-compulsory education. The two sectors of the tertiary education system, vocational education and…
Being victims of racial prejudice, religious intolerance, poverty, disempowerment and language loss it could be expected that indigenous people would be supportive of the Inclusion Movement with its philosophy of valuing and acceptance of all people. This supposition is examined for Maori, the indigenous people of Aotearoa/New Zealand. In…
The conservation of the genetic variability of the indigenous angiosperm community is a sine qua non. A survey of indigenous angiosperm biodiversity of the Olabisi Onabanjo University permanent site was undertaken. Plants collected were dried, poisoned and mounted on herbarium sheets, proper identification and ...
Indigenous knowledge is often dismissed as 'traditional and outdated', and hence irrelevant to modern ecological assessment. This theoretical paper critically examines the arguments advanced to elevate modern western ecological knowledge over indigenous ecological knowledge, as well as the sources and uses of ...
Smith, Linda Tuhiwai
From the vantage point of indigenous peoples, the term "research" is inextricably linked to European imperialism and colonialism. In this book, an indigenous researcher calls for the decolonization of research methods. The first part of the book critically examines the historical and philosophical bases of Western research; Western…
the attention of decision-makers yet it forms part of discussions in such fora as the Convention .... neem tree under the millet heads when they lay them on the ground to dry. .... There is a close competition between the conventional and indigenous practices, .... Figure 3: Most Effective Indigenous Management Practices.
Harrold, Timothy C; Randall, Deborah A; Falster, Michael O; Lujic, Sanja; Jorm, Louisa R
To quantify the independent roles of geography and Indigenous status in explaining disparities in Potentially Preventable Hospital (PPH) admissions between Indigenous and non-Indigenous Australians. Analysis of linked hospital admission data for New South Wales (NSW), Australia, for the period July 1 2003 to June 30 2008. Age-standardised admission rates, and rate ratios adjusted for age, sex and Statistical Local Area (SLA) of residence using multilevel models. PPH diagnoses accounted for 987,604 admissions in NSW over the study period, of which 3.7% were for Indigenous people. The age-standardised PPH admission rate was 76.5 and 27.3 per 1,000 for Indigenous and non-Indigenous people respectively. PPH admission rates in Indigenous people were 2.16 times higher than in non-Indigenous people of the same age group and sex who lived in the same SLA. The largest disparities in PPH admission rates were seen for diabetes complications, chronic obstructive pulmonary disease and rheumatic heart disease. Both rates of PPH admission in Indigenous people, and the disparity in rates between Indigenous than non-Indigenous people, varied significantly by SLA, with greater disparities seen in regional and remote areas than in major cities. Higher rates of PPH admission among Indigenous people are not simply a function of their greater likelihood of living in rural and remote areas. The very considerable geographic variation in the disparity in rates of PPH admission between Indigenous and non-Indigenous people indicates that there is potential to reduce unwarranted variation by characterising outlying areas which contribute the most to this disparity.
Hansen, Annette Skovsted
The re-vitalization of indigenous languages depend on political and legal support and the imple-mentation of language rights depend on knowledge of vocabulary and grammar structures of the individual languages. Throughout the nineteenth century world, compilers of dictionaries adapted indigenous...... languages to match standards defined in nation-building and, thereby, enabled latent possibilities for indigenous populations to re-vitalize their languages in connection with the United Nations Year for Indigenous Peoples in 1993, and the first United Nations Decade for Indigenous Peoples, 1995......–2004. This article focuses on dictionaries of the languages of the Ainu populations in the borderlands between the nation-states Japan and Russia. The main argument is that the Ainu Cultural Promotion Act promulgated in 1997 had a significant impact on the production and purpose of Ainu dictionaries...
Ivers, Rebecca; Clapham, Kathleen; Senserrick, Teresa; Lyford, Marilyn; Stevenson, Mark
Injury prevention in Indigenous communities in Australia is a continuing national challenge, with Indigenous fatality rates due to injury three times higher than the general population. Suicide and transport are the leading causes of injury mortality, and assault, transport and falls the primary causes of injury morbidity. Addressing the complex range of injury problems in disadvantaged Indigenous communities requires considerable work in building or enhancing existing capacity of communities to address local safety issues. Poor data, lack of funding and absence of targeted programs are some of the issues that impede injury prevention activities. Traditional approaches to injury prevention can be used to highlight key areas of need, however adaptations are needed in keeping with Indigenous peoples' holistic approach to health, linked to land and linked to community in order to address the complex spiritual, emotional and social determinants of Indigenous injury.
Babar, S.M.; Hussain, M.; Mahmood, T.
Environmental pollution control is the most important and highly discussed issue at the international level. Our and our's next generation survival highly depends on environment. Environmental security is not less important than territorial security. Living in the Competitive trade, Business and Commerce era. WTO threats of globalization to countries like Pakistan require sharp and immediate actions. SOS(Save our Sole) steps should be taken in Environmental Education in order to reorganizing values and clarifying Concepts to develop the necessary skills and attitude necessary to understand and appreciate the interrelatidness among masses, the Cultures and Ecosystem. Historical backgrounds along with different approaches were discussed particularly reference to Pakistan. In this presentation a new but indigenous idea is flashed to improve the environment education system in poor third world countries including Pakistan. Instead of imported ideas, previous implemented as such, indigenous approach highly Perfumed with Islamic, Ideological and cultural blends will do the right job in right direction if employed with true sense of commitment. (author)
Liachko, Ivan; Youngblood, Rachel A; Tsui, Kyle; Bubb, Kerry L; Queitsch, Christine; Raghuraman, M K; Nislow, Corey; Brewer, Bonita J; Dunham, Maitreya J
The well-studied DNA replication origins of the model budding and fission yeasts are A/T-rich elements. However, unlike their yeast counterparts, both plant and metazoan origins are G/C-rich and are associated with transcription start sites. Here we show that an industrially important methylotrophic budding yeast, Pichia pastoris, simultaneously employs at least two types of replication origins--a G/C-rich type associated with transcription start sites and an A/T-rich type more reminiscent of typical budding and fission yeast origins. We used a suite of massively parallel sequencing tools to map and dissect P. pastoris origins comprehensively, to measure their replication dynamics, and to assay the global positioning of nucleosomes across the genome. Our results suggest that some functional overlap exists between promoter sequences and G/C-rich replication origins in P. pastoris and imply an evolutionary bifurcation of the modes of replication initiation.
Full Text Available The well-studied DNA replication origins of the model budding and fission yeasts are A/T-rich elements. However, unlike their yeast counterparts, both plant and metazoan origins are G/C-rich and are associated with transcription start sites. Here we show that an industrially important methylotrophic budding yeast, Pichia pastoris, simultaneously employs at least two types of replication origins--a G/C-rich type associated with transcription start sites and an A/T-rich type more reminiscent of typical budding and fission yeast origins. We used a suite of massively parallel sequencing tools to map and dissect P. pastoris origins comprehensively, to measure their replication dynamics, and to assay the global positioning of nucleosomes across the genome. Our results suggest that some functional overlap exists between promoter sequences and G/C-rich replication origins in P. pastoris and imply an evolutionary bifurcation of the modes of replication initiation.
Borodina, Irina; Jensen, Bettina M.; Wagner, Tim
Wasp venom allergy is the most common insect venom allergy in Europe. It is manifested by large local reaction or anaphylactic shock occurring after a wasp sting. The allergy can be treated by specific immunotherapy with whole venom extracts. Wasp venom is difficult and costly to obtain...... and is a subject to composition variation, therefore it can be advantageous to substitute it with a cocktail of recombinant allergens. One of the major venom allergens is phospholipase A1, which so far has been expressed in Escherichia coli and in insect cells. Our aim was to produce the protein in secreted form...... in yeast Pichia pastoris, which can give high yields of correctly folded protein on defined minimal medium and secretes relatively few native proteins simplifying purification.Residual amounts of enzymatically active phospholipase A1 could be expressed, but the venom protein had a deleterious effect...
Borodina, Irina; Jensen, Bettina M.; Wagner, Tim
Wasp venom allergy is the most common insect venom allergy in Europe. It is manifested by large local reaction or anaphylactic shock occurring after a wasp sting. The allergy can be treated by specific immunotherapy with whole venom extracts. Wasp venom is difficult and costly to obtain...... and is a subject to composition variation, therefore it can be advantageous to substitute it with a cocktail of recombinant allergens. One of the major venom allergens is phospholipase A1, which so far has been expressed in Escherichia coli and in insect cells. Our aim was to produce the protein in secreted form...... in yeast Pichia pastoris, which can give high yields of correctly folded protein on defined minimal medium and secretes relatively few native proteins simplifying purification. Residual amounts of enzymatically active phospholipase A1 could be expressed, but the venom protein had a deleterious effect...
Borodina, Irina; Jensen, Bettina M; Wagner, Tim
Wasp venom allergy is the most common insect venom allergy in Europe. It is manifested by large local reaction or anaphylactic shock occurring after a wasp sting. The allergy can be treated by specific immunotherapy with whole venom extracts. Wasp venom is difficult and costly to obtain...... and is a subject to composition variation, therefore it can be advantageous to substitute it with a cocktail of recombinant allergens. One of the major venom allergens is phospholipase A1, which so far has been expressed in Escherichia coli and in insect cells. Our aim was to produce the protein in secreted form...... in yeast Pichia pastoris, which can give high yields of correctly folded protein on defined minimal medium and secretes relatively few native proteins simplifying purification.Residual amounts of enzymatically active phospholipase A1 could be expressed, but the venom protein had a deleterious effect...
Petersen, Bent Larsen; Damager, Iben; Faber, Kirsten
Cell). In the present study, Flag-tagged (MDYKDDDD) RGXT2 was expressed in Pichia pastoris as secreted soluble protein, in pea (using the Pea early browning virus as expression vector) as soluble intra-cellular protein and in tobacco as full length membrane bound protein. The amount of expressed...... to participate in plant CW biosynthesis, has been achieved in only a few cases. We have previously reported the characterisation of two highly homologous plant-specific membrane-bound GTs, which when expressed as secreted tagged soluble proteins in the baculo virus system, catalysed the transfer of xylose from...... protein was estimated using anti Flag Ab and corresponding activity monitored. Pros and cons of using the various expression systems are discussed....
Ramon, R; Feliu, J X; Cos, O; Montesinos, J L; Berthet, F X; Valero, F
The Pichia pastoris expression system is widely used for the production of recombinant proteins. A simple and efficient experimental set-up allowing on-line monitoring of the methanol concentration during the fermentation of P. pastoris based on the detection of the methanol vapor concentration in the exhaust air from fermenter by a tin dioxide (SnO2) semiconductor sensor is described. An experimental procedure to allow precise calibration of the system and to reduce methanol sensor's interferences (>95% reduction) are also presented and discussed. Accuracy and measurement error were estimated about 0.05 g x l(-1) and 6%, respectively. The efficient monitoring of methanol will help to advanced control of recombinant protein production and process optimization.
Camattari, Andrea; Weinhandl, Katrin; Gudiminchi, Rama K
The methylotrophic yeast Pichia pastoris is becoming one of the favorite industrial workhorses for protein expression. Due to the widespread use of integration vectors, which generates significant clonal variability, screening methods allowing assaying hundreds of individual clones are of particular importance. Here we describe methods to detect and analyze protein expression, developed in a 96-well format for high-throughput screening of recombinant P. pastoris strains. The chapter covers essentially three common scenarios: (1) an enzymatic assay for proteins expressed in the cell cytoplasm, requiring cell lysis; (2) a whole-cell assay for a fungal cytochrome P450; and (3) a nonenzymatic assay for detection and quantification of tagged protein secreted into the supernatant.
Chen, Yong; Li, Yang; Liu, Peng; Sun, Qun; Liu, Zhu
A weakness of using immobilized metal affinity chromatography (IMAC) to purify recombinant proteins expressed in Pichia pastoris is the co-purification of native proteins that exhibit high affinities for Ni-IMAC. We have determined the elution profiles of P. pastoris proteins and have examined the native proteins that co-purify when eluting with 100 mM imidazole. Four major contaminants were identified: mitochondrial alcohol dehydrogenase isozyme III (mADH), nucleotide excision repair endonuclease, and the hypothetical proteins TPHA_0L01390 and TDEL_0B02190 which are homologous proteins derived from Tetrapisispora phaffii and Torulaspora delbrueckii, respectively. A new P. pastoris expression strain was engineered that eliminated the predominant contaminant, mADH, by gene disruption. The total amount of protein contaminants was reduced by 55 % without effecting cell growth. The present study demonstrates the feasibility of using a proteomic approach to facilitate bioprocess optimization.
Borodina, Irina; Do, Duy Duc; Eriksen, Jens C.
Background Methylotrophic yeast Pichia pastoris is widely used for recombinant protein production, largely due to its ability to secrete correctly folded heterologous proteins to the fermentation medium. Secretion is usually achieved by cloning the recombinant gene after a leader sequence, where...... alpha‐mating factor (MF) prepropeptide from Saccharomyces cerevisiae is most commonly used. Our aim was to test whether signal peptides from P. pastoris native secreted proteins could be used to direct secretion of recombinant proteins. Results Eleven native signal peptides from P. pastoris were tested...... by optimization of expression of three different proteins in P. pastoris. Conclusions Native signal peptides from P. pastoris can be used to direct secretion of recombinant proteins. A novel USER‐based P. pastoris system allows easy cloning of protein‐coding gene with the promoter and leader sequence of choice....
Full Text Available Synthetic dyes are known to be highly toxic to mammalian cells and mutagenic and carcinogenic to humans and, therefore, should be detoxified and removed from industrial effluents. Different approaches for removal and detoxication are extensively sought. Biochemical methods are considered the most economical and effective method of dye decolourization. In this research, the laccase gene from Laccaria bicolor was modified and expressed in Pichia pastoris. The properties of the recombinant laccase and its ability to degrade synthetic dyes were studied. The laccase activity was optimal at pH 2.2 and 50 °C. Its Km value was 0.187 mmol/L for ABTS [2,2'-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid]. The laccase obtained was shown to decolorize the synthetic dyes, malachite green, crystal violet and orange G, with ABTS as a mediator. These results indicated that the laccase obtained may be used to treat industrial effluents containing artificial dyes.
Jeffries, Thomas W.; Grigoriev, Igor; Grimwood, Jane; Laplaza,Jose M.; Aerts, Andrea; Salamov, Asaf; Schmutz, Jeremy; Lindquist, Erika; Dehal, Paramvir; Shapiro, Harris; Jin, Yong-Su; Passoth, Volkmar; Richardson, Paul M.
Xylose is a major constituent of angiosperm lignocellulose,so its fermentation is important for bioconversion to fuels andchemicals. Pichia stipitis is the best-studied native xylose fermentingyeast. Genes from P. stipitis have been used to engineer xylosemetabolism in Saccharomycescerevisiae, and the regulation of the P.stipitis genome offers insights into the mechanisms of xylose metabolismin yeasts. We have sequenced, assembled and finished the genome ofP.stipitis. As such, it is one of only a handful of completely finishedeukaryotic organisms undergoing analysis and manual curation. Thesequence has revealed aspects of genome organization, numerous genes forbiocoversion, preliminary insights into regulation of central metabolicpathways, numerous examples of co-localized genes with related functions,and evidence of how P. stipitis manages to achieve redox balance whilegrowing on xylose under microaerobic conditions.
Bajaj, Bijender Kumar; Raina, Sandeepu; Singh, Satbir
The killer phenomenon of yeast may have technological implications in many areas like beverage fermentation, food technology, biological control in agriculture, and in medicine. In the present study the killer phenomenon in Pichia kudriavzevii (P. kudriavzevii RY55) is being reported for the first time. The P. kudriavzevii RY55 toxin exhibited excellent antibacterial activity against several pathogens of human health significance such as Escherichia coli, Enterococcus faecalis, Klebsiella sp., Staphylococcus aureus, Pseudomonas aeruginosa and Pseudomonas alcaligenes. Killer toxin was purified to homogeneity by using ammonium sulphate precipitation and ion exchange chromatography and characterized for few properties. P. kudriavzevii RY55 killer toxin may be of vast significance in the development of novel antimicrobial chemotherapeutic agents, new bio-based safer candidates for food preservation and biocontrol, and starter cultures for fermentation industries. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
Venskutonytė, Raminta; Elbing, Karin; Lindkvist-Petersson, Karin
Large amounts of pure and homogenous protein are a prerequisite for several biochemical and biophysical analyses, and in particular if aiming at resolving the three-dimensional protein structure. Here we describe the production of the rat glucose transporter 1 (GLUT1), a membrane protein facilitating the transport of glucose in cells. The protein is recombinantly expressed in the yeast Pichia pastoris. It is easily maintained and large-scale protein production in shaker flasks, as commonly performed in academic research laboratories, results in relatively high yields of membrane protein. The purification protocol describes all steps needed to obtain a pure and homogenous GLUT1 protein solution, including cell growth, membrane isolation, and chromatographic purification methods.
Arslan, Yeşim; Eken-Saraçoğlu, Nurdan
In this study, we investigated the use of hazelnut shell as a renewable and low cost lignocellulosic material for bioethanol production for the first time. High lignin content of hazelnut shell is an important obstacle for such a biotransformation. Biomass hydrolysis with acids yields reducing sugar with several inhibitors which limit the fermentability of sugars. The various conditioning methods for biomass and hydrolysate were performed to overcome the toxicity and their effects on the subsequent fermentation of hazelnut shell hydrolysate by Pichia stipitis were evaluated with shaking flasks experiments. Hazelnut shells hydrolysis with 0.7M H(2)SO(4) yielded 49 gl(-1) total reducing sugars and fermentation inhibitors in untreated hydrolysate. First, it was shown that several hydrolysate detoxification methods were solely inefficient in achieving cell growth and ethanol production in the fermentation of hazelnut shell hydrolysates derived from non-delignified biomass. Next, different pretreatments of hazelnut shells were considered for delignification and employed before hydrolysis in conjunction with hydrolysate detoxification to improve alcohol fermentation. Among six delignification methods, the most effective pretreatment regarding to ethanol concentration includes the treatment of shells with 3% (w/v) NaOH at room temperature, which was integrated with sequential hydrolysate detoxification by overliming and then treatment with charcoal twice at 60 degrees C. This treatment brought about a total reduction of 97% in furans and 88.4% in phenolics. Almost all trialed treatments caused significant sugar loss. Under the best assayed conditions, ethanol concentration of 16.79gl(-1) was reached from a hazelnut shell hyrolysate containing initial 50g total reducing sugar l(-1) after partial synthetic xylose supplementation. This value is equal to 91.25% of ethanol concentration that was obtained from synthetic d-xylose under same conditions. The present study
Aramoana, Jaclyn; Alley, Patrick; Koea, Jonathan B
Progress has been made in Australia and New Zealand to increase the numbers of indigenous students (Aboriginal, Torres Strait Islander and Maori) entering primary medical qualification courses. In New Zealand, up to 20 Maori are graduating annually, with similar numbers possible in Australia, creating a potential opportunity to develop an indigenous surgical workforce. A literature review identified factors utilized by medical schools to attract indigenous students into medical careers and the interventions necessary to ensure successful graduation. A further search identified those factors important in encouraging indigenous medical graduates to enter specialist training programmes and achieve faculty appointments. All medical schools have utilized elements of a 'pipeline approach' encompassing contact with students at secondary school level to encourage aspirational goals and assist with suitable subject selection. Bridging courses can ensure students leaving school have appropriate skill sets before entering medical degree courses. Extensive practical help is available during primary medical qualification study. The elements necessary for primary medical qualification success - dedicated and focused study, developing appropriate skill sets, mentoring, support, and an institutional and collegial commitment to success - are also the elements required for postgraduate achievement. The Royal Australasian College of Surgeons (RACS) is primarily involved in training rather than service provision. The increasing numbers of indigenous medical graduates in both Australia and New Zealand represent an opportunity for the College to contribute to improving indigenous health status by implementing specific measures to increase numbers of indigenous surgeons. © 2013 Royal Australasian College of Surgeons.
This paper discusses the individual, family, household and area level characteristics associated with preschool attendance for Indigenous and non-Indigenous Australians (aged three to five years who are not at school). Controlling for these factors explains all of the difference between Indigenous and non-Indigenous attendance rates for…
Gorlewski, Julie; Porfilio, Brad J.
Based upon the life histories of six Indigenous hip hop artists of the Beat Nation artist collective, this essay captures how Indigenous hip hop has the potential to revolutionize environmental education. Hip hop provides Indigenous youth an emancipatory space to raise their opposition to neocolonial controls of Indigenous territories that…
Morozkina, E V; Vavilova, E A; Zatsepin, S S; Klyachko, E V; Yagudin, T A; Chulkin, A M; Dudich, I V; Semenkova, L N; Churilova, I V; Benevolenskii, S V
A system for the production of mutant recombinant human alpha-fetoprotein (rhAFPO) lacking the glycosylation site has been engineered in the yeast Pichia pastoris. A strain of the methylotrophic yeast Pichia pastoris GS 115/pPICZ?A/rhAFP0, which produces unglycosylated rhAFPO and secretes it to the culture medium, has been constructed. Optimization and scale-up of the fermentation technology have resulted in an increase in the rhAFP0 yield to 20 mg/L. A scheme of isolation and purification of biologically active rhAFP0 has been developed. The synthesized protein has the antitumor activity, which is analogous to the activity of natural human embryonic alpha-fetoprotein.
Zhou, Xiang-Shan; Fan, Wei-Min; Zhang, Yuan-Xing
Four different methanol feeding modes were evaluated in the hirudin production in high-density fermentation by Pichia pastoris. It was difficult to avoid methanol excessive in the broth with the feeding strategy only based on DO level. On the other hand, the fluctuation in methanol concentration was observed with methanol feeding strategy by off-line gas chromatography. However, the stable methanol concentration was perfectly achieved by the on-line monitoring with methanol sensor. The supply of energy was improved by feeding glycerol at a limited rate as well as methanol in the induction phase. Therefore, the high cell dry weight (162 g/L) and high hirudin activity (2.4 x 10(4) ATU/mL or 1.7 g/L) was obtained in the fed-batch fermentation of recombinant Pichia pastoris by methanol-glycerol mixed feeding.
Pandya, R. E.; Bennett, B.
Historically, the goal of broadening participation in the geosciences has been expressed and approached from the viewpoint of the majority-dominated geoscience community. The need for more students who are American Indian, Native Hawaiian, or Alaska Native is expressed in terms of the need to diversify the research community, and strategies to engage more students are often posed around the question “what can we do to get more indigenous students interested in coming to our institutions to do geosciences?” This approach can lead to neglecting indigenous ways of knowing, inadvertently prioritizes western values over traditional ones, and doesn’t necessarily honor tribal community’s desire to hold on to their talented youth. Further, while this approach has resulted in some modest success, the overall participation in geoscience by students from indigenous backgrounds remains low. Many successful programs, however, have tried an alternate approach; they begin by approaching the geosciences from the viewpoint of indigenous communities. The questions they ask center around how geosciences can advance the priorities of indigenous communities, and their approaches focus on building capacity for the geosciences within indigenous communities. Most importantly, perhaps, these efforts originate in Tribal communities themselves, and invite the geoscience research community to partner in projects that are rooted in indigenous culture and values. Finally, these programs recognize that scientific expertise is only one among many skills indigenous peoples employ in their relation with their homelands. Climate change, like all things related to the landscape, is intimately connected to the core of indigenous cultures. Thus, emerging concerns about climate change provide a venue for developing new, indigenous-centered, approaches to the persistent problem of broadening participation in the geoscience. This presentation will highlight three indigenous-led efforts in to
Diehl, Eliana Elisabeth; Langdon, Esther Jean; Dias-Scopel, Raquel Paiva
Indigenous community health workers are part of a strategy developed by Brazil in the last two decades to promote a special healthcare model for indigenous peoples. Their role is designed to deal with various aspects of the special health policy, including the link between the heath team and the community and mediation between scientific and indigenous medical knowledge. Despite a significant increase in the number of indigenous community health workers in recent years, an evaluation of their responsibilities and contributions to the success of special care had not been conducted previously. This article, based on a literature review and original research by the authors, analyzes the role of the indigenous community health workers vis-à-vis their training and participation in health teams in different contexts in Brazil. Considering the importance assigned to the role of indigenous community health workers, this analysis reveals various ambiguities and contradictions that hinder both their performance and their potential contribution to the special health services.
Slámová, Kristýna; Bojarová, Pavla; Gerstorferová, Daniela; Fliedrová, Barbora; Hofmeisterová, Jana; Fiala, Martin; Pompach, Petr; Křen, Vladimír
Roč. 82, č. 1 (2012), s. 212-217 ISSN 1046-5928 R&D Projects: GA ČR GP203/09/P024; GA ČR GD305/09/H008; GA ČR(CZ) GAP207/11/0629; GA MŠk(CZ) 7E11011 Keywords : Talaromyces flavus * Pichia pastoris * Yeast expression system Subject RIV: CE - Biochemistry Impact factor: 1.429, year: 2012
Mukherjee, Pranab K.; Chandra, Jyotsna; Retuerto, Mauricio; Sikaroodi, Masoumeh; Brown, Robert E.; Jurevic, Richard; Salata, Robert A.; Lederman, Michael M.; Gillevet, Patrick M.; Ghannoum, Mahmoud A.
Oral microbiota contribute to health and disease, and their disruption may influence the course of oral diseases. Here, we used pyrosequencing to characterize the oral bacteriome and mycobiome of 12 HIV-infected patients and matched 12 uninfected controls. The number of bacterial and fungal genera in individuals ranged between 8–14 and 1–9, among uninfected and HIV-infected participants, respectively. The core oral bacteriome (COB) comprised 14 genera, of which 13 were common between the two groups. In contrast, the core oral mycobiome (COM) differed between HIV-infected and uninfected individuals, with Candida being the predominant fungus in both groups. Among Candida species, C. albicans was the most common (58% in uninfected and 83% in HIV-infected participants). Furthermore, 15 and 12 bacteria-fungi pairs were correlated significantly within uninfected and HIV-infected groups, respectively. Increase in Candida colonization was associated with a concomitant decrease in the abundance of Pichia, suggesting antagonism. We found that Pichia spent medium (PSM) inhibited growth of Candida, Aspergillus and Fusarium. Moreover, Pichia cells and PSM inhibited Candida biofilms (P = .002 and .02, respectively, compared to untreated controls). The mechanism by which Pichia inhibited Candida involved nutrient limitation, and modulation of growth and virulence factors. Finally, in an experimental murine model of oral candidiasis, we demonstrated that mice treated with PSM exhibited significantly lower infection score (P = .011) and fungal burden (P = .04) compared to untreated mice. Moreover, tongues of PSM-treated mice had few hyphae and intact epithelium, while vehicle- and nystatin-treated mice exhibited extensive fungal invasion of tissue with epithelial disruption. These results showed that PSM was efficacious against oral candidiasis in vitro and in vivo. The inhibitory activity of PSM was associated with secretory protein/s. Our findings provide the
Yakovleva, Maria E.; Killyeni, Aniko; Seubert, Oliver; Conghaile, Peter O.; MacAodha, Domhnall; Leech, Donal; Gonaus, Christoph; Popescu, Ionel Catalin; Peterbauer, Clemens K.; Kjellstrom, Sven; Gorton, Lo
The present study focuses on fragmented deglycosylated pyranose dehydrogenase (fdgPDH) from Agaricus meleagris recombinantly expressed in Pichia pastoris. Fragmented deglycosylated PDH is formed from the deglycosylated enzyme (dgPDH) when it spontaneously loses a C-terminal fragment when stored in a buffer solution at 4 °C. The remaining larger fragment has a molecular weight of ∼46 kDa and exhibits higher volumetric activity for glucose oxidation compared with the deglycosylated and glycosyl...
Yunier Serrano-Rivero; Karen Marrero-Domínguez; Rafael Fando-Calzada
La levadura Pichia pastoris ha sido ampliamente empleada como sistema hospedero para la producción de proteínas heterólogas, tales como; fragmentos de anticuerpos, huridina recombinante, eritropoyetina humana recombinante e insulina humana recombinante. Su extendida aplicación se debe a la disponibilidad del promotor de la enzima alcohol oxidasa, el cual es fuerte y regulable por metanol y a su capacidad para alcanzar altas densidades celulares en biorreactores en medios de cultivo baratos. E...
Land is the foundation for the economic sustenance of indigenous peoples and for the continued survival of their cultures. One of the major problems faced by indigenous peoples is the dispossession of their traditional lands and territories. The activities of business interests and economic development projects in indigenous territories – such as forest logging and infrastructure projects - and the environmental implications of such activities, often constitute a great threat to the livelihoo...
Indigenous environmental activists have clearly articulated their views on global climate change policy. The content of these views was explored during the 10-day 2008 World Conservation Congress (WCC) in Barcelona. Data were primarily collected through interviews and participant observation. In addition, policy statements and declarations made by indigenous environmental activists from 2000 to 2009 were analysed to place the perspectives of indigenous leaders and environmental activists in t...
Athmaram, T N; Singh, Anil Kumar; Saraswat, Shweta; Srivastava, Saurabh; Misra, Princi; Kameswara Rao, M; Gopalan, N; Rao, P V L
The present Influenza vaccine manufacturing process has posed a clear impediment to initiation of rapid mass vaccination against spreading pandemic influenza. New vaccine strategies are therefore needed that can accelerate the vaccine production. Pichia offers several advantages for rapid and economical bulk production of recombinant proteins and, hence, can be attractive alternative for producing an effective influenza HA based subunit vaccine. The recombinant Pichia harboring the transgene was subjected to fed-batch fermentation at 10 L scale. A simple fermentation and downstream processing strategy is developed for high-yield secretory expression of the recombinant Hemagglutinin protein of pandemic Swine Origin Influenza A virus using Pichia pastoris via fed-batch fermentation. Expression and purification were optimized and the expressed recombinant Hemagglutinin protein was verified by sodium dodecyl sulfate polyacrylamide gel electrophoresis, Western blot and MALDI-TOF analysis. In this paper, we describe a fed-batch fermentation protocol for the secreted production of Swine Influenza A Hemagglutinin protein in the P. pastoris GS115 strain. We have shown that there is a clear relationship between product yield and specific growth rate. The fed-batch fermentation and downstream processing methods optimized in the present study have immense practical application for high-level production of the recombinant H1N1 HA protein in a cost effective way using P. pastoris.
Ronny Alejandro Leiva
Chilean indigenous institutions recently joined the Convention concerning Indigenous and Tribal Peoples Convention, 1989 (no. 169) of the International Labor Organization (ILO) to its administrative structure. Prior to this, indigenous law 19.253 indigenous organizations were created in order to foster the participation of these peoples. Convention No. 169 and other international instruments on indigenous rights enshrine the right to participation through their own representative institut...
Pringle, Kirsty G; de Meaultsart, Celine Corbisier; Sykes, Shane D; Weatherall, Loretta J; Keogh, Lyniece; Clausen, Don C; Dekker, Gus A; Smith, Roger; Roberts, Claire T; Rae, Kym M; Lumbers, Eugenie R
The intrarenal renin-angiotensin system (iRAS) is implicated in the pathogenesis of hypertension, chronic kidney disease and diabetic nephropathy. Urinary angiotensinogen (uAGT) levels reflect the activity of the iRAS and are altered in women with preeclampsia. Since Indigenous Australians suffer high rates and early onset of renal disease, we hypothesised that Indigenous Australian pregnant women, like non-Indigenous women with pregnancy complications, would have altered uAGT levels. The excretion of RAS proteins was measured in non-Indigenous and Indigenous Australian women with uncomplicated or complicated pregnancies (preeclampsia, diabetes/gestational diabetes, proteinuria/albuminuria, hypertension, small/large for gestational age, preterm birth), and in non-pregnant non-Indigenous women. Non-Indigenous pregnant women with uncomplicated pregnancies, had higher uAGT/creatinine levels than non-Indigenous non-pregnant women (P pregnant women with pregnancy complications, uAGT/creatinine was suppressed in the third trimester (P pregnant women with uncomplicated pregnancies, there was no change in uAGT/creatinine with gestational age and uAGT/creatinine was lower in the 2nd and 3rd trimesters than in non-Indigenous pregnant women with uncomplicated pregnancies (P pregnant women may reflect subclinical renal dysfunction which limits the ability of the kidney to maintain sodium balance and could indicate an increased risk of pregnancy complications and/or future renal disease. Copyright © 2018. Published by Elsevier B.V.
Maharajh, Dheepak M
Full Text Available advantages. Approximately 30% of South African environments favourable for isolating algae have been sampled. Samples were enriched, purified and assessed for lipid content, resulting in a database of indigenous algae. Positive isolates were grown under...
Herbal-based and plant-derived products can be exploited with sustainable comparative and competitive advantage. This review presents some indigenous African plants with chemotherapeutic properties and possible ways of developing them into potent pharmacological agents using biotechnological approaches.
Documenting and disseminating agricultural indigenous knowledge for sustainable food security in Uganda. ... University of Dar es Salaam Library Journal ... Moreover, small-scale farmers should be involved in agricultural extension services ...
knowledge research approaches in a school-based collaborative project of the .... ticipate in informal employment; however, they manage to provide for their families. ... Entitled “Promoting and learning from Cofimvaba community's indigenous.
harvesting and storage of indigenous root crops and animals. .... sample of 351 informants who were selected using random, purposive and snowball sampling ... The farmers in Soroti also spray crops with human and animal urine, dust.
Indigenous and traditional African vegetables (AITVs) are important sources of ... and (iii) marketing: retail markup, price variation by season, year and region, ... size and cost, retailer storage, remainders, processing and less common AITVs.
Indigenous knowledge system should ... Therefore, ignoring these local knowledge practices by the ..... International Journal of Science and Rural Development. ... Ethno-medicine: Its potential in the health care system, Canoe Press.
challenges) in society, sometimes it is marginalized in education because it is seen as non-scientific and non-engaging in formal education. Using the capability approach to human development, this paper investigates the link between indigenous ...
Supporting indigenous women in science, technology, engineering and mathematics careers in Mexico and Central ... ROSSA's latest bulletin puts a focus on women. ... IDRC invites applications for the IDRC Doctoral Research Awards.
Dean SPGS NAU
dance performance presents Ekwe people; situated in Isu local government ... Indigenous dance is not a luxury… it is part of each .... symbols for certain brand products in adverts. ... music, costumes, make-up, set lights and any other effects.
Leaving no one behind: can (indigenous) education ensure effective gender ... in the distribution of socio-economic and political benefits, depict that additional ... of gender equality and equity and explores in different ways the relationships ...
Vogl, Thomas; Gebbie, Leigh; Palfreyman, Robin W; Speight, Robert
Pichia pastoris (syn. Komagataella phaffii ) is one of the most common eukaryotic expression systems for heterologous protein production. Expression cassettes are typically integrated in the genome to obtain stable expression strains. In contrast to Saccharomyces cerevisiae , where short overhangs are sufficient to target highly specific integration, long overhangs are more efficient in P. pastoris and ectopic integration of foreign DNA can occur. Here, we aimed to elucidate the influence of ectopic integration by high-throughput screening of >700 transformants and whole-genome sequencing of 27 transformants. Different vector designs and linearization approaches were used to mimic the most common integration events targeted in P. pastoris Fluorescence of an enhanced green fluorescent protein (eGFP) reporter protein was highly uniform among transformants when the expression cassettes were correctly integrated in the targeted locus. Surprisingly, most nonspecifically integrated transformants showed highly uniform expression that was comparable to specific integration, suggesting that nonspecific integration does not necessarily influence expression. However, a few clones (integrated cassettes showed a greater variation spanning a 25-fold range, surpassing specifically integrated reference strains up to 6-fold. High-expression strains showed a correlation between increased gene copy numbers and high reporter protein fluorescence levels. Our results suggest that for comparing expression levels between strains, the integration locus can be neglected as long as a sufficient numbers of transformed strains are compared. For expression optimization of highly expressible proteins, increasing copy number appears to be the dominant positive influence rather than the integration locus, genomic rearrangements, deletions, or single-nucleotide polymorphisms (SNPs). IMPORTANCE Yeasts are commonly used as biotechnological production hosts for proteins and metabolites. In the yeast
Full Text Available Abstract Background Laccases are blue multi-copper oxidases and catalyze the oxidation of phenolic and non-phenolic compounds. There is considerable interest in using these enzymes for dye degradation as well as for synthesis of aromatic compounds. Laccases are produced at relatively low levels and, sometimes, as isozymes in the native fungi. The investigation of properties of individual enzymes therefore becomes difficult. The goal of this study was to over-produce a previously reported laccase from Cyathus bulleri using the well-established expression system of Pichia pastoris and examine and compare the properties of the recombinant enzyme with that of the native laccase. Results In this study, complete cDNA encoding laccase (Lac from white rot fungus Cyathus bulleri was amplified by RACE-PCR, cloned and expressed in the culture supernatant of Pichia pastoris under the control of the alcohol oxidase (AOX1 promoter. The coding region consisted of 1,542 bp and encodes a protein of 513 amino acids with a signal peptide of 16 amino acids. The deduced amino acid sequence of the matured protein displayed high homology with laccases from Trametes versicolor and Coprinus cinereus. The sequence analysis indicated the presence of Glu 460 and Ser 113 and LEL tripeptide at the position known to influence redox potential of laccases placing this enzyme as a high redox enzyme. Addition of copper sulfate to the production medium enhanced the level of laccase by about 12-fold to a final activity of 7200 U L-1. The recombinant laccase (rLac was purified by ~4-fold to a specific activity of ~85 U mg-1 protein. A detailed study of thermostability, chloride and solvent tolerance of the rLac indicated improvement in the first two properties when compared to the native laccase (nLac. Altered glycosylation pattern, identified by peptide mass finger printing, was proposed to contribute to altered properties of the rLac. Conclusion Laccase of C. bulleri was
Garg, Neha; Bieler, Nora; Kenzom, Tenzin; Chhabra, Meenu; Ansorge-Schumacher, Marion; Mishra, Saroj
Laccases are blue multi-copper oxidases and catalyze the oxidation of phenolic and non-phenolic compounds. There is considerable interest in using these enzymes for dye degradation as well as for synthesis of aromatic compounds. Laccases are produced at relatively low levels and, sometimes, as isozymes in the native fungi. The investigation of properties of individual enzymes therefore becomes difficult. The goal of this study was to over-produce a previously reported laccase from Cyathus bulleri using the well-established expression system of Pichia pastoris and examine and compare the properties of the recombinant enzyme with that of the native laccase. In this study, complete cDNA encoding laccase (Lac) from white rot fungus Cyathus bulleri was amplified by RACE-PCR, cloned and expressed in the culture supernatant of Pichia pastoris under the control of the alcohol oxidase (AOX)1 promoter. The coding region consisted of 1,542 bp and encodes a protein of 513 amino acids with a signal peptide of 16 amino acids. The deduced amino acid sequence of the matured protein displayed high homology with laccases from Trametes versicolor and Coprinus cinereus. The sequence analysis indicated the presence of Glu 460 and Ser 113 and LEL tripeptide at the position known to influence redox potential of laccases placing this enzyme as a high redox enzyme. Addition of copper sulfate to the production medium enhanced the level of laccase by about 12-fold to a final activity of 7200 U L-1. The recombinant laccase (rLac) was purified by ~4-fold to a specific activity of ~85 U mg(-1) protein. A detailed study of thermostability, chloride and solvent tolerance of the rLac indicated improvement in the first two properties when compared to the native laccase (nLac). Altered glycosylation pattern, identified by peptide mass finger printing, was proposed to contribute to altered properties of the rLac. Laccase of C. bulleri was successfully produced extra-cellularly to a high level of 7200
Doyle, Kerrie; Hungerford, Catherine
Indigenous Australians have higher levels of mental illness, self-harm, suicide and substance abuse than non-Indigenous Australians, as well as more frequent contact with the criminal justice system. These indices point to the need for strong leadership to support Close the Gap programmes that have now been implemented across Australia. This article considers leadership as a journey of learning for Australian Indigenous leaders. Through the use of story, it is suggested that a situational leadership approach, incorporating the principles of mindfulness, provides the most appropriate framework for Indigenous leaders who work with Indigenous communities. Flexible approaches are needed to meet the needs of diverse Indigenous populations, and address the complex challenges involved, including lateral violence. Such flexibility will enable Indigenous leaders and communities to work together to achieve improvements in the health outcomes, not only for Indigenous Australians, but also for Indigenous populations worldwide.
Nikolakis, William D.; Grafton, R. Quentin; To, Hang
Drawing upon on the literature on Indigenous values to water, water markets and the empirical findings from a survey of 120 Indigenous and non-Indigenous respondents across northern Australia, the paper makes important qualitative and statistical comparisons between Indigenous and non-Indigenous values to water markets. The study is the first comparison of Indigenous and non-Indigenous values to water markets based on the same survey instrument. Key results from Indigenous respondents include: (1) water markets are held to be an acceptable approach to managing water; (2) markets must be carefully designed to protect customary and ecological values; (3) the allocation of water rights need to encompass equity considerations; and (4) water and land rights should not be separated even if this enhances efficiency, as it runs counter to Indigenous holistic values. Overall, the survey results provide the basis for a proposed adaptive decision loop, which allows decision makers to incorporate stakeholder values in water markets.
Wang, R.; Miao, Y.; Kang, C.
Rehmannia glutinosa (R. glutinosa) is a high demand traditional Chinese medicine, but it suffers serious consecutive monoculture obstacle (CMO). The disability of root swelling is one of the negative impacts caused by the R. glutinosa CMO and is related to allelopathy exudates, such as phenolic acids. It is thought that a microbe agent could improve plant health by eliminating the unfavorable effect of allelopathy exudates. In previous research, we isolated two phenolic acid-degrading microbes from rhizosphere soil surrounding R. glutinosa. These were Bacillus sp. and Pichia pastoris. This study found that Bacillus sp. combined with Pichia pastoris could degrade 97.19% ferulic acid and 98.73% hydroxybenzoic acid over 15 days. R. glutinosa takes a long growth time (7-8 months) under field conditions. We set up a modified tissue culture model to rapidly detect whether Bacillus sp. and Pichia pastoris combination could relieve the CMO. The results showed that our tissue culture model effectively simulated the R. glutinosa growing process in unplanted or second-year monoculture field. Furthermore, the combination of Bacillus sp. and Pichia pastoris can significantly relieve the CMO-induced suppression of root swelling. All these results suggested that: 1) The combination of Bacillus sp. and Pichia pastoris has considerable potential to degrade allelopathy exudates and alleviate the CMO of R. glutinosa; 2) Our tissue culture model could be used to quickly screen effective microbes that could alleviate CMO in plants. (author)
Full Text Available Enhanced rate of fossil fuel extraction is likely to deplete limited natural resources over short period of time. So search for alternative fuel is only the way to overcome this problem of upcoming energy crisis. In this aspect biofuel is a sustainable option. Agricultural lands cannot be compromised for biofuel production due to the requirement of food for the increasing population. Certain species of algae can produce ethanol during anaerobic fermentation and thus serve as a direct source for bioethanol production. The high content of complex carbohydrates entrapped in the cell wall of the microalgae makes it essential to incorporate a pre-treatment stage to release and convert these complex carbohydrates into simple sugars prior to the fermentation process. There have been researches on production of bioethanol from a particular species of algae, but this work was an attempt to produce bioethanol from easily available indigenous algae. Acid hydrolysis was carried out as pre-treatment. Gas Chromatographic analysis showed that 5 days’ fermentation by baker’s yeast had yielded 93% pure bioethanol. The fuel characterization of the bioethanol with respect to gasoline showed comparable and quite satisfactory results for its use as an alternative fuel.DOI: http://dx.doi.org/10.3126/ije.v4i1.12182International Journal of Environment Volume-4, Issue-1, Dec-Feb 2014/15, page: 112-120
Burm, Sarah; Burleigh, Dawn
Identifying as non-Indigenous, we are often left considering our positionality and identity in Indigenous education, how we have come to be invested in this area of research, and what we see as our contribution. In conversation with one another, we realized we choose to share certain stories and not others about our experiences working in…
Araneda, Jacqueline; Amigo, Hugo; Bustos, Patricia
During childhood and adolescence, eating habits become established which are instrumental in determining eating behavior later in life. Various authors have described the acculturation of the Mapuche people toward Western culture. The objective of this study was to analyze the eating characteristics of indigenous and non-indigenous adolescent girls in the Araucania Region of Chile. A cross-sectional design was used with a probabilistic sample of 281 adolescents comprised of 139 indigenous and 142 non-indigenous girls attending 168 elementary schools. A modified food frequency questionnaire was applied, designed to obtain information about eating habits and consumption of Mapuche foods. The eating schedules are similar in both ethnic groups, with dinner being the meal that is least consumed. Total snack consumption per week has a mean of 7 with an interquartile range (IQR) of 5 to 10 without any differences between ethnic groups; of these snacks, only 2 were healthy (IQR = 1 to 3). The indigenous girls had a higher probability of consumption of native foods including mote (boiled wheat) (OR = 2.00; IC = 0.93-4.29), muday (fermented cereal alcohol) (OR = 3.45; IC = 1.90-6.27), and yuyo (field mustard) (OR = 4.40; IC = 2.06-9.39). The study's conclusion is that the the eating habits and behavior of indigenous adolescents are similar to those of non-indigenous girls, though the former still consume more indigenous foods.
Mika, Carl; Stewart, Georgina; Watson, Ka'imi; Silva, Keola; Martin, Brian; Matapo, Jacoba; Galuvao, Akata
In this commentary, various expert authors offer their ideas on indigenous research in the philosophy of education and PESA's role from an indigenous perspective. Georgiana Stewart is the first author to step forward and explain that education is based on knowledge, and so education is centrally concerned with literacy and identity. Stewart goes…
Cláudia Pereira Antunes
Full Text Available The education in the indigenous schools, even though it is build within the legal framework of the State and a modern concept of education, demands new comprehensions from the western society not only in its relation with the indigenous education, but also in its relation with the indigenous people. Because of its condition as a mediator between two different forms of thinking, the indigenous school also represents a fertile ground to think about the western conceptions of education. This article is dedicated to a deeper reflection about some aspects of the relation between the western society, based on rationality and science, and the indian people in the construction of the indigenous schools.
Foulds, Heather J A; Bredin, Shannon S D; Warburton, Darren E R
Limited understanding of Indigenous adults' cardiovascular structure and function exists despite high rates of cardiovascular disease. This investigation characterised cardiovascular structure and function among young Indigenous adults and compared to age- and sex-matched European descendants. Echocardiographic assessments included apical two- and four-chamber images, parasternal short-axis images and Doppler. Analyses included cardiac volumes, dimensions, velocities and strains. Cardiovascular structure and function were similar between Indigenous (n=10, 25 ± 3 years, 4 women) and European-descendant (n=10, 24 ± 4 years, 4 women,) adults, though European descendants demonstrated greater systemic vascular resistance (18.19 ± 3.94 mmHg∙min -1 ∙L -1 vs. 15.36 ± 2.97 mmHg∙min -1 ∙L -1 , p=0.03). Among Indigenous adults, women demonstrated greater arterial elastance (0.80 ± 0.15 mmHg·mL -1 ·m -2 vs. 0.55 ± 0.17 mmHg·mL -1 ·m -2 , p=0.02) and possibly greater systemic vascular resistance (17.51 ± 2.20 mmHg∙min -1 ∙L -1 vs. 13.93 ± 2.61 mmHg∙min -1 ∙L -1 , p=0.07). Indigenous men had greater cardiac size, dimensions and output, though body size differences accounted for cardiac size differences. Similar cardiac rotation and strains were observed across sexes. Arterial elastance and cardiac size were different between Indigenous men and women while cardiovascular structure and function may be similar between Indigenous and European descendants.
Baghban, Roghayyeh; Gargari, Seyed Latif Mousavi; Rajabibazl, Masoumeh; Nazarian, Shahram; Bakherad, Hamid
Botulinum neurotoxins (BoNTs) result in severe and often fatal disease, botulism. Common remedial measures such as equine antitoxin and human botulism immunoglobulin in turn are problematic and time-consuming. Therefore, diagnosis and therapy of BoNTs are vital. The variable domain of heavy-chain antibodies (VHH) has unique features, such as the ability to identify and bind specifically to target epitopes and ease of production in bacteria and yeast. The Pichia pastoris is suitable for expression of recombinant antibody fragments. Disulfide bond formation and correct folds of protein with a high yield are some of the advantages of this eukaryotic host. In this study, we have expressed and purified the camelid VHH against BoNT/E in P. pastoris. The final yield of P. pastoris-expressed antibody was estimated to be 16 mg/l, which is higher than that expressed by Escherichia coli. The nanobody expressed in P. pastoris neutralized 4LD50 of the BoNT/E upon i.p. injection in 25% of mice. The nanobody expressed in E. coli extended the mice's survival to 1.5-fold compared to the control. This experiment indicated that the quality of expressed protein in the yeast is superior to that of the bacterial expression. Favorable protein folding by P. pastoris seems to play a role in its better toxin-binding property. © 2014 International Union of Biochemistry and Molecular Biology, Inc.
Chen, Gen-Hung; Chen, Wei-Ming; Huang, Guo-Ting; Chen, Yu-Wen; Jiang, Shann-Tzong
Four recombinant antimicrobial peptide (rAMP) cDNAs, constructed from two goat lactoferricin-related peptide cDNAs (GLFcin and GLFcin II) with/without (His)(6)-Tag, were cloned into pPICZalphaC and transformed into Pichia pastoris SMD1168H. After methanol induction, these rAMPs were expressed and secreted into broth. They were purified after CM-Sepharose (without His-tg), HisTrap (with His-tg) and Sephadex G-25 chromatographies. The yield of purified rAMP was 0.15 mg/mL of broth. These 4 rAMPs were thermal-stable and with high antibacterial activity against Escherichia coli BCRC 11549, Pseudomonas aeruginosa BCRC 12450, Bacillus cereus BCRC 10603, Staphylococcus aureus BCRC 25923, Propioni bacterium acnes BCRC 10723, and Listera monocytogenes BCRC 14845. The minimum inhibitory concentration (MIC) of rAMPs against these indicators ranged from 4.07 to 16.00 mg/mL.
Tang, Xiang-Shan; Tang, Zhi-Ru; Wang, Sheng-Ping; Feng, Ze-Meng; Zhou, Dong; Li, Tie-Jun; Yin, Yu-Long
Bovine lactoferrampin (LFA) and bovine lactoferricin (LFC) are two antimicrobial peptides located in the N(1) domain of bovine lactoferrin. The bactericidal activity of the fused peptide LFA-LFC is stronger than that of either LFA or LFC. The high cost of peptide production from either native digestion or chemical synthesis limits the clinical application of antimicrobial peptides. The expression of recombinant peptides in yeast may be an effective alternative. In the current study, the expression, purification, and antibacterial activity of LFA-LFC using the Pichia pastoris expression system are reported. The linearized expression vector pPICZaA-LFA-LFC was transformed into P. pastoris KM71 by electroporation, and positive colonies harboring the target genes were screened out and used for fermentation. The recombinant LFA-LFC peptide was purified via two-step column chromatography and identified by tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The results indicate that P. pastoris is a suitable system for secreting LFA-LFC. The fermentation supernate and the purified LFA-LFC show high antimicrobial activities. The current study is the first to report on the expression and purification of LFA-LFC in P. pastoris and may have potential practical applications in microbial peptide production.
Acosta-Rivero, Nelson; Rodriguez, Armando; Musacchio, Alexis; Falcon, Viviana; Suarez, Viana M.; Chavez, Liudmila; Morales-Grillo, Juan; Duenas-Carrera, Santiago
Little is known about the in vivo assembly pathway or structure of the hepatitis C virus nucleocapsid. In this work the intermediates of HCcAg multimerization in Pichia pastoris cells and the nucleic acid binding properties of structured nucleocapsid-like particles (NLPs) were studied. Extensive cross-linking was observed for HCcAg after glutaraldehyde treatment. Data suggest that HCcAg exists in dimeric forms probably representing P21-P21, P21-P23, and P23-P23 dimers. In addition, the presence of HCcAg species that might represent trimers and multimers was observed. After sucrose equilibrium density gradient purification and nuclease digestion, NLPs were shown to contain both RNA and DNA molecules. Finally, the analysis by electron microscopy indicated that native NLPs were resistant to nuclease treatment. These results indicated that HCcAg assembles through dimers, trimers, and multimers' intermediates into capsids in P. pastoris cells. Assembly of NLPs in its natural environment might confer stability to these particles by adopting a compact structure
Ohsawa, Shin; Yurimoto, Hiroya; Sakai, Yasuyoshi
Wsc family proteins are plasma membrane spanning sensor proteins conserved from yeasts to mammalian cells. We studied the functional roles of Wsc family proteins in the methylotrophic yeast Pichia pastoris, and found that PpWsc1 and PpWsc3 function as methanol-sensors during growth on methanol. PpWsc1 responds to a lower range of methanol concentrations than PpWsc3. PpWsc1, but not PpWsc3, also functions during high temperature stress, but PpWsc1 senses methanol as a signal that is distinct from high-temperature stress. We also found that PpRom2, which is known to function downstream of the Wsc family proteins in the cell wall integrity pathway, was also involved in sensing methanol. Based on these results, these PpWsc family proteins were demonstrated to be involved in sensing methanol and transmitting the signal via their cytoplasmic tail to the nucleus via PpRom2, which plays a critical role in regulating expression of a subset of methanol-inducible genes to coordinate well-balanced methanol metabolism. © 2017 John Wiley & Sons Ltd.
Shupe, Alan M; Liu, Shijie
Concentrated dilute acid hydrolysate was obtained from hot water extracts of Acer saccharum (sugar maple) and was fermented to ethanol by Pichia stipitis in a 1.3-L-benchtop bioreactor. The conditions under which the highest ethanol yield was achieved were when the air flow rate was set to 100 cm(3) and the agitation rate was set to 150 rpm resulting in an overall mass transfer coefficient (K(L)a) of 0.108 min(-1). A maximum ethanol concentration of 29.7 g/L was achieved after 120 h of fermentation; however, after 90 h of fermentation, the ethanol concentration was only slightly lower at 29.1 g/L with a yield of 0.39 g ethanol per gram of sugar consumed. Using the same air flow rate and adjusting the agitation rate resulted in lower ethanol yields of 0.25 g/g at 50 rpm and 0.30 g/g at 300 rpm. The time it takes to reach the maximum ethanol concentration was also affected by the agitation rate. The ethanol concentration continued to increase even after 130 h of fermentation when the agitation rate was set at 50 rpm, whereas the maximum ethanol concentration was reached after only 68.5 h at 300 rpm.
Li, Zhemin; Zhao, Genhai; Liu, Hui; Guo, Yugang; Wu, Hefang; Sun, Xiaowen; Wu, Xihua; Zheng, Zhiming
Prenylated quinones, especially menaquinones, have significant physiological activities, but are arduous to synthesize efficiently. Due to the relaxed aromatic substrate specificity and prenylation regiospecificity at the ortho- site of the phenolic hydroxyl group, the aromatic prenyltransferase NovQ from Streptomyces may be useful in menaquinone synthesis from menadione. In this study, NovQ was overexpressed in Pichia pastoris. After fermentation optimization, NovQ production increased by 1617%. Then the different effects of metal ions, detergents and pH on the activity of purified NovQ were investigated to optimize the prenylation reaction. Finally, purified NovQ and cells containing NovQ were used for menadione prenylation in vitro and in vivo, respectively. Menaquinone-1 (MK-1) was detected as the only product in vitro with γ,γ-dimethylallyl pyrophosphate and menadione hydroquinol substrates. MK-3 at a concentration of 90.53 mg/L was detected as the major product of whole cell catalysis with 3-methyl-2-buten-1-ol and menadione hydroquinol substrates. This study realized whole cell catalysis converting menadione to menaquinones.
Lei, Da; Xu, Yang; He, Qinghua; Pang, Yifeng; Chen, Bo; Xiong, Liang; Li, Yanping
Neutral protease I from Aspergillus oryzae 3.042 was expressed in Pichia pastoris and its N-glycosylation properties were analyzed. After purification by nickel-affinity chromatography column, the recombinant neutral protease (rNPI) was confirmed to be N-glycosylated by periodicacid/Schiff's base staining and Endo H digestion. Moreover, the deglycosylated protein's molecular weight decreased to 43.3 kDa from 54.5 kDa analyzed by SDS-PAGE and MALDI-TOF-MS, and the hyperglycosylation extent was 21 %. The N-glycosylation site of rNPI was analyzed by nano LC-MS/MS after digesting by trypsin and Glu-C, and the unique potential site Asn41 of mature peptide was found to be glycosylated. Homology modeling of the 3D structure of rNPI indicated that the attached N-glycans hardly affected neutral protease's activity due to the great distance away from the active site of the enzyme.
Full Text Available Abstract Background Cellobiose dehydrogenase (CDH is an extracellular hemoflavoenzyme produced by lignocellulose-degrading fungi including Pycnoporus cinnabarinus. We investigated the cellulolytic system of P. cinnabarinus, focusing on the involvement of CDH in the deconstruction of lignocellulosic biomass. Results First, P. cinnabarinus growth conditions were optimized for CDH production. Following growth under cellulolytic conditions, the main components secreted were cellulases, xylanases and CDH. To investigate the contribution of P. cinnabarinus secretome in saccharification processes, the Trichoderma reesei enzymatic cocktail was supplemented with the P. cinnabarinus secretome. A significant enhancement of the degradation of wheat straw was observed with (i the production of a large amount of gluconic acid, (ii increased hemicellulose degradation, and (iii increased overall degradation of the lignocellulosic material. P. cinnabarinus CDH was heterologously expressed in Pichia pastoris to obtain large amounts of pure enzyme. In a bioreactor, the recombinant CDH (rCDH expression level reached 7800 U/L. rCDH exhibited values of biochemical parameters similar to those of the natural enzyme, and was able to bind cellulose despite the absence of a carbohydrate-binding module (CBM. Following supplementation of purified rCDH to T. reesei enzymatic cocktail, formation of gluconic acid and increased hemicellulose degradation were observed, thus confirming the previous results observed with P. cinnabarinus secretome. Conclusions We demonstrate that CDH offers an attractive tool for saccharification process enhancement due to gluconic acid production from raw lignocellulosic material.
Sukwong, Pailin; Ra, Chae Hun; Sunwoo, In Yung; Tantratian, Sumate; Jeong, Gwi-Taek; Kim, Sung-Koo
This study employed a statistical method to obtain optimal hyper thermal acid hydrolysis conditions using Gelidium amansii (red seaweed) as a source of biomass. The optimal hyper thermal acid hydrolysis using G. amansii as biomass was determined as 12% (w/v) slurry content, 358.3 mM H 2 SO 4 , and temperature of 142.6 °C for 11 min. After hyper thermal acid hydrolysis, enzymatic saccharification was carried out. The total monosaccharide concentration was 45.1 g/L, 72.2% of the theoretical value of the total fermentable monosaccharides of 62.4 g/L based on 120 g dry weight/L in the G. amansii slurry. To increase ethanol production, 3.8 g/L 5-hydroxymethylfurfural (HMF) in the hydrolysate was removed by treatment with 3.5% (w/v) activated carbon for 2 min and fermented with Pichia stipitis adapted to high galactose concentrations via separate hydrolysis and fermentation. With complete HMF removal and the use of P. stipitis adapted to high galactose concentrations, 22 g/L ethanol was produced (yield 0.50). Fermentation with total HMF removal and yeast adapted to high galactose concentrations increased the fermentation performance and decreased the fermentation time from 96 to 36 h compared to traditional fermentation.
Wu, Xihua; Li, Zhemin; Liu, Hui; Wang, Peng; Wang, Li; Fang, Xue; Sun, Xiaowen; Ni, Wenfeng; Yang, Qiang; Zheng, Zhiming; Zhao, Genhai
The effect of methanol addition on the heterologous expression of isoprenyl transferase NovQ was studied in Pichia pastoris Gpn12, with menadione and isopentenol as precursors to catalyze vitamin K2 (MK-3) synthesis. The expression of NovQ increased by 36% when 2% methanol was added every 24 h. The influence of initial pH, temperature, methanol addition, precursors (menadione, isopentenol) addition, catalytic time and cetyltrimethyl-ammonium bromide (CTAB) addition were explored in the P. pastoris whole-cell catalytic synthesis process of MK-3 in shaking flask. Three significant factors were then studied by response surface method. The optimal catalytic conditions obtained were as follows: catalytic temperature 31.56 ℃, menadione 295.54 mg/L, catalytic time 15.87 h. Consistent with the response surface prediction results, the optimized yield of MK-3 reached 98.47 mg/L in shaking flask, 35% higher than that of the control group. On this basis, the production in a 30-L fermenter reached 189.67 mg/L when the cell catalyst of 220 g/L (dry weight) was used to catalyze the synthesis for 24 h. This method laid the foundation for the large-scale production of MK-3 by P. pastoris Gpn12.
Irani, Zahra Azimzadeh; Kerkhoven, Eduard J; Shojaosadati, Seyed Abbas; Nielsen, Jens
Pichia pastoris is used for commercial production of human therapeutic proteins, and genome-scale models of P. pastoris metabolism have been generated in the past to study the metabolism and associated protein production by this yeast. A major challenge with clinical usage of recombinant proteins produced by P. pastoris is the difference in N-glycosylation of proteins produced by humans and this yeast. However, through metabolic engineering, a P. pastoris strain capable of producing humanized N-glycosylated proteins was constructed. The current genome-scale models of P. pastoris do not address native nor humanized N-glycosylation, and we therefore developed ihGlycopastoris, an extension to the iLC915 model with both native and humanized N-glycosylation for recombinant protein production, but also an estimation of N-glycosylation of P. pastoris native proteins. This new model gives a better prediction of protein yield, demonstrates the effect of the different types of N-glycosylation of protein yield, and can be used to predict potential targets for strain improvement. The model represents a step towards a more complete description of protein production in P. pastoris, which is required for using these models to understand and optimize protein production processes. © 2015 Wiley Periodicals, Inc.
Full Text Available Background: Leishmania major LmSTI1 is a conserved protein among different species of leishmania, and expressed in both amastigote and promastigote forms of L. major life cycle. It has previously been expressed in bacterial systems.Materials and Methods: To express LmSTI1 in the methylotrophic yeast Pichia pastoris (P. pastoris, the shuttle vector pPICZA containing gene lmsti1 was constructed under the control of the AOX1 promoter. The recombinant vector was electro-transformed into P. pastoris, and induced by 0.5% methanol in the buffered medium. The expression of the LmSTI1 protein was visualized in the total soluble protein of P. pastoris by 12% SDS-PAGE, and further confirmed by Western blotting with L.major-infected mouse sera and HRP-conjugated goat anti-mouse IgG as the first and secondary antibodies, respectively.Results: The expression level was 0.2% of total soluble proteins.Conclusion: It might be possible to use this formulation as a whole yeast candidate vaccine against cutaneous leishmanization.
Henry Córdoba Ruiz
Full Text Available Producing human proteins in lower organisms' cells using recombinant technology represents a very promising approach for treating many diseases produced by a particular protein deficiency, including close to 40 lysosomal storage diseases. Although E. coli has been the first host successfully employed in expressing human recombinant proteins, it has some limitations owing to its inability to perform some post-traductional steps such as glycosylation. The yeast Saccharomyces cerevisiae (S. cerevisiae has thusbeen initially considered and used. However, S. cerevisiae glycosylates proteins in a very different way to human cells producing highly antigenic proteins and thus some other non-conventional yeasts such as Pichia pastoris have been used recently. Human protein expression is not assodated with growth in this system; growth may occur at high cell concentrations, increasing heterologous protein productivity and yield. The system employs a very efficient, methanol-induced promoter which may be used as sole carbon and energy source. Post-traductional modifications seem more similar to human cells than those produced by other non-mammalian systems used in producing human glycoproteins; they do not secrete large amounts of endogenous proteins, simplifying expressed protein purification. This review presents some strategies for producing heterologous proteins in high density cultures using P. pastoris as an expression system.
Li, Pengfei; Sun, Hongbing; Chen, Zao; Li, Yin; Zhu, Taicheng
Cellulosic biomass especially agricultural/wood residues can be utilized as feedstock to cost-effectively produce fuels, chemicals and bulk industrial enzymes, which demands xylose utilization from microbial cell factories. While previous works have made significant progress in improving microbial conversion of xylose into fuels and chemicals, no study has reported the engineering of efficient xylose utilizing protein expression systems for the purpose of producing industrial enzymes. In this work, using Pichia pastoris as an example, we demonstrated the successful engineering of xylose metabolizing ability into of protein expression systems. A heterologous XI (xylose isomerase) pathway was introduced into P. pastoris GS115 by overexpressing the Orpinomyces spp. XI or/and the endogenous XK (xylulokinase) gene, and evolutionary engineering strategies were also applied. Results showed that the XI pathway could be functionally expressed in P. pastoris. After 50 generation of sequential batch cultivation, a set of domesticated recombinant P. pastoris strains with different performance metrics on xylose were obtained. One evolved strain showed the highest xylose assimilation ability, whose cell yield on xylose can even be comparable to that on glucose or glycerol. This strain also showed significantly increased β-mannanase production when cultured on xylose medium. Furthermore, transcription analysis of xylose pathway genes suggested that overexpression of XI and XK might be the key factors affecting effective xylose assimilation. To our best knowledge, this study is the first work demonstrating the construction of efficient xylose utilizing P. pastoris strains, thus providing a basis for using cellulosic biomass for bulk industrial enzyme production.
Full Text Available The physiological responses and transcription profiling of Pichia pastoris GS115 to simulated microgravity (SMG were substantially changed compared with normal gravity (NG control. We previously reported that the recombinant P. pastoris grew faster under SMG than NG during methanol induction phase and the efficiencies of recombinant enzyme production and secretion were enhanced under SMG, which was considered as the consequence of changed transcriptional levels of some key genes. In this work, transcriptiome profiling of P. pastoris cultured under SMG and NG conditions at exponential and stationary phases were determined using next-generation sequencing (NGS technologies. Four categories of 141 genes function as methanol utilization, protein chaperone, RNA polymerase and protein transportation or secretion classified according to Gene Ontology (GO were chosen to be analyzed on the basis of NGS results. And 80 significantly changed genes were weighted and estimated by Cluster 3.0. It was found that most genes of methanol metabolism (85% of 20 genes and protein transportation or secretion (82.2% of 45 genes were significantly up-regulated under SMG. Furthermore the quantity and fold change of up-regulated genes in exponential phase of each category were higher than those of stationary phase. The results indicate that the up-regulated genes of methanol metabolism and protein transportation or secretion mainly contribute to enhanced production and secretion of the recombinant protein under SMG.
Full Text Available The purposes of this study was to identify the optimum concentration of the lytic enzyme Glucanex for protoplast isolation and to conduct fusion for the purpose of increasing inulinase production. The study performs the protoplast fusion technique using Pichia manshurica and Rhodosporidium paludigenum. Protoplast fusion consists of a series of stages: protoplast isolation, protoplast fusion, protoplast regeneration, and analysis of hybrid fusion results. Protoplast isolation and fusion success rate are determined by various factors, including age of the culture, media type, and type of lytic enzymes used. Hybrid results were analyzed using a fungicide as a marker and measuring specific growth rate (μ of the hybrid compared with parental growth rates. Results demonstrated that a concentration of 4 mg/mL of Glucanex produces the greatest number of protoplasts, 7.2 x 1010 (cell/mL for P. manshurica and 8.8 x 1010 (cell/mL for Rh. paludigenum. The results of analysis of hybrid fusions indicate that the study has identified a new fusant, called fusant F4. Fusant F4 is capable of producing the highest inulinase, 0.6892 IU, compared with parentals P. manshurica, 0557 IU, and Rh. paludigenum, 0.3263 IU. Fusant F4 has specific growth rate (μ of 0.3360/h and generation time (g of 2.0629 h.
Angela Johana Espejo Mojica
Full Text Available β-hexosaminidases (Hex are dimeric enzymes involved in the lysosomal degradation of glycolipids and glycans. They are formed by α- and/or β-subunits encoded byHEXA and HEXB genes, respectively. Mutations in these genes lead to Tay Sachs or Sandhoff diseases, which are neurodegenerative disorders caused by the accumulation of non-degraded glycolipids. Although tissue-derived Hex have been widely characterized, limited information is available for recombinant β-hexosaminidases. In this study, human lysosomal recombinant Hex (rhHex-A, rhHex-B, and rhHex-S were produced in the methylotrophic yeast Pichia pastoris GS115. The highest specific enzyme activities were 13,124 for rhHexA; 12,779 for rhHex-B; and 14,606 U .mg-1 for rhHex-S. These results were 25- to 50-fold higher than those obtained from normal human leukocytes. Proteins were purified and characterized at different pH and temperature conditions. All proteins were stable at acidic pH, and at 4 °C and 37 °C. At 45 °C rhHex-S was completely inactivated, while rhHex-A and rhHex-B showed high stability. This study demonstrates P. pastoris GS115 potential for polymeric lysosomal enzyme production, and describes the characterization of recombinant β-hexosaminidases produced within the same host.
Full Text Available Abstract Background Pichia stipitis xylose reductase (Ps-XR has been used to design Saccharomyces cerevisiae strains that are able to ferment xylose. One example is the industrial S. cerevisiae xylose-consuming strain TMB3400, which was constructed by expression of P. stipitis xylose reductase and xylitol dehydrogenase and overexpression of endogenous xylulose kinase in the industrial S. cerevisiae strain USM21. Results In this study, we demonstrate that strain TMB3400 not only converts xylose, but also displays higher tolerance to lignocellulosic hydrolysate during anaerobic batch fermentation as well as 3 times higher in vitro HMF and furfural reduction activity than the control strain USM21. Using laboratory strains producing various levels of Ps-XR, we confirm that Ps-XR is able to reduce HMF both in vitro and in vivo. Ps-XR overexpression increases the in vivo HMF conversion rate by approximately 20%, thereby improving yeast tolerance towards HMF. Further purification of Ps-XR shows that HMF is a substrate inhibitor of the enzyme. Conclusion We demonstrate for the first time that xylose reductase is also able to reduce the furaldehyde compounds that are present in undetoxified lignocellulosic hydrolysates. Possible implications of this newly characterized activity of Ps-XR on lignocellulosic hydrolysate fermentation are discussed.
Gonçalves, A M; Pedro, A Q; Maia, C; Sousa, F; Queiroz, J A; Passarinha, L A
During the last few decades, it has become evident that the compatibility of the yeast biochemical environment with the ability to process and translate the RNA transcript, along with its capacity to modify a translated protein, are relevant requirements for selecting this host cell for protein expression in several pharmaceutical and clinical applications. In particular, Pichia pastoris is used as an industrial host for recombinant protein and metabolite production, showing a powerful capacity to meet required biomolecular target production levels in high-throughput assays for functional genomics and drug screening. In addition, there is a great advantage to using P. pastoris for protein secretion, even at high molecular weights, since the recovery and purification steps are simplified owing to relatively low levels of endogenous proteins in the extracellular medium. Clearly, no single microexpression system can provide all of the desired properties for human protein production. Moreover, chemical and physical bioprocess parameters, including culture medium formulation, temperature, pH, agitation, aeration rates, induction, and feeding strategies, can highly influence product yield and quality. In order to benefit from the currently available wide range of biosynthesis strategies using P. pastoris, this mini review focuses on the developments and technological fermentation achievements, providing both a comparative and an overall integration analysis. The main aim is to highlight the relevance and versatility of the P. pastoris biosystem to the design of more cost-effective microfactories to meet the increasing demands for recombinant membrane proteins and clinical antibodies for several therapeutic applications.
Li, Sumei; Hong, Tao; Wang, Kun; Lu, Yinghong; Zhou, Min
Most proteins occur and function in complexes rather than as isolated entities in membranes. In most cases macromolecules with multiple subunits are purified from endogenous sources. In this study, an endogenous membrane-protein complex was obtained from Pichia pastoris, which can be grown at high densities to significantly improve the membrane protein yield. We successfully isolated the membrane-bound Vo complex of V-ATPase from P. pastoris using a fusion FLAG tag attached to the C-terminus of subunit a to generate the vph-tag strain, which was used for dissociation and purification. After FLAG affinity and size exclusion chromatography purification, the production quantity and purity of the membrane-bound Vo complex was 20 μg l -1 and >98%, respectively. The subunits of the endogenous membrane-bound Vo complex observed in P. pastoris were similar to those obtained from S. cerevisiae, as demonstrated by liquid chromatography-tandem mass spectrometry (LC-MS-MS). Therefore, successful dissociation and purification of the membrane-bound Vo complex at a high purity and sufficient quantity was achieved via a rapid and simple procedure that can be used to obtain the endogenous membrane-protein complexes from P. pastoris. Copyright © 2017 Elsevier Inc. All rights reserved.
Gasser, Brigitte; Dragosits, Martin; Mattanovich, Diethard
Biotin plays an essential role as cofactor for biotin-dependent carboxylases involved in essential metabolic pathways. The cultivation of Pichia pastoris, a methylotrophic yeast that is successfully used as host for the production of recombinant proteins, requires addition of high dosage of biotin. As biotin is the only non-salt media component used during P. pastoris fermentation (apart from the carbon source), nonconformities during protein production processes are usually attributed to poor quality of the added biotin. In order to avoid dismissed production runs due to biotin quality issues, we engineered the biotin-requiring yeast P. pastoris to become a biotin-prototrophic yeast. Integration of four genes involved in the biotin biosynthesis from brewing yeast into the P. pastoris genome rendered P. pastoris biotin-prototrophic. The engineered strain has successfully been used as production host for both intracellular and secreted heterologous proteins in fed-batch processes, employing mineral media without vitamins. Another field of application for these truly prototrophic hosts is the production of biochemicals and small metabolites, where defined mineral media leads to easier purification procedures. Copyright © 2010 Elsevier Inc. All rights reserved.
Wriessnegger, Tamara; Augustin, Peter; Engleder, Matthias; Leitner, Erich; Müller, Monika; Kaluzna, Iwona; Schürmann, Martin; Mink, Daniel; Zellnig, Günther; Schwab, Helmut; Pichler, Harald
The sesquiterpenoid (+)-nootkatone is a highly demanded and highly valued aroma compound naturally found in grapefruit, pummelo or Nootka cypress tree. Extraction of (+)-nootkatone from plant material or its production by chemical synthesis suffers from low yields and the use of environmentally harmful methods, respectively. Lately, major attention has been paid to biotechnological approaches, using cell extracts or whole-cell systems for the production of (+)-nootkatone. In our study, the yeast Pichia pastoris initially was applied as whole-cell biocatalyst for the production of (+)-nootkatone from (+)-valencene, the abundant aroma compound of oranges. Therefore, we generated a strain co-expressing the premnaspirodiene oxygenase of Hyoscyamus muticus (HPO) and the Arabidopsis thaliana cytochrome P450 reductase (CPR) that hydroxylated extracellularly added (+)-valencene. Intracellular production of (+)-valencene by co-expression of valencene synthase from Callitropsis nootkatensis resolved the phase-transfer issues of (+)-valencene. Bi-phasic cultivations of P. pastoris resulted in the production of trans-nootkatol, which was oxidized to (+)-nootkatone by an intrinsic P. pastoris activity. Additional overexpression of a P. pastoris alcohol dehydrogenase and truncated hydroxy-methylglutaryl-CoA reductase (tHmg1p) significantly enhanced the (+)-nootkatone yield to 208mg L(-1) cell culture in bioreactor cultivations. Thus, metabolically engineered yeast P. pastoris represents a valuable, whole-cell system for high-level production of (+)-nootkatone from simple carbon sources. Copyright © 2014 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.
de Queiroz Brito Cunha, Carolina Cândida; Gama, Aline Rodrigues; Cintra, Lorena Cardoso; Bataus, Luiz Artur Mendes; Ulhoa, Cirano José
Xylanases (EC 18.104.22.168) are hydrolytic enzymes, which randomly cleave the β-1,4-linked xylose residues from xylan. The synthetic gene xynBS27 from Streptomyces sp. S27 was successfully cloned and expressed in Pichia pastoris. The full-length gene consists of 729 bp and encodes 243 amino acids including 51 residues of a putative signal peptide. This enzyme was purified in two steps and was shown to have a molecular weight of 20 kDa. The purified r-XynBS27 was active against beechwood xylan and oat spelt xylan as expected for GH 11 family. The optimum pH and temperature values for the enzyme were 6.0 and 75 °C, respectively. The Km and Vmax were 12.38 mg/mL and 13.68 μmol min/mg, respectively. The r-XynBS27 showed high xylose tolerance and was inhibited by some metal ions and by SDS. r-XynBS27 was employed as an additive in the bread making process. A decrease in firmness, stiffness and consistency, and improvements in specific volume and reducing sugar content were recorded.
Grillitsch, Karlheinz; Tarazona, Pablo; Klug, Lisa; Wriessnegger, Tamara; Zellnig, Günther; Leitner, Erich; Feussner, Ivo; Daum, Günther
Despite similarities of cellular membranes in all eukaryotes, every compartment displays characteristic and often unique features which are important for the functions of the specific organelles. In the present study, we biochemically characterized the plasma membrane of the methylotrophic yeast Pichia pastoris with emphasis on the lipids which form the matrix of this compartment. Prerequisite for this effort was the design of a standardized and reliable isolation protocol of the plasma membrane at high purity. Analysis of isolated plasma membrane samples from P. pastoris revealed an increase of phosphatidylserine and a decrease of phosphatidylcholine compared to bulk membranes. The amount of saturated fatty acids in the plasma membrane was higher than in total cell extracts. Ergosterol, the final product of the yeast sterol biosynthetic pathway, was found to be enriched in plasma membrane fractions, although markedly lower than in Saccharomyces cerevisiae. A further characteristic feature of the plasma membrane from P. pastoris was the enrichment of inositol phosphorylceramides over neutral sphingolipids, which accumulated in internal membranes. The detailed analysis of the P. pastoris plasma membrane is discussed in the light of cell biological features of this microorganism especially as a microbial cell factory for heterologous protein production. Copyright © 2014 Elsevier B.V. All rights reserved.
Ishikawa, Mai; Shiono, Yoshihito; Koseki, Takuya
An α-l-rhamnosidase-encoding gene from Aspergillus oryzae, which belongs to the glycoside hydrolase family 78, was cloned and expressed in Pichia pastoris. SDS-PAGE of the purified recombinant α-l-rhamnosidase protein revealed smeared bands with apparent molecular mass of 90-130 kDa. After N-deglycosylation, the recombinant enzyme showed a molecular mass of 70 kDa. The enzyme exhibited optimal activity at a pH of 5.0 and a temperature of 70 °C. Specific activity of the enzyme was higher toward hesperidin than toward naringin, which consist of α-1,6 and α-1,2 linkages, respectively. The activity was also higher toward hesperidin than toward rutin, which consist of 7-O- and 3-O-glycosyl linkages of flavonoids, respectively. Kinetic analysis of the enzyme showed that the Michaelis constant (K m ) was lowest toward rutin, moderate toward naringin, and higher toward p-nitrophenyl-α-l-rhamnopyranoside and hesperidin. Its high catalytic efficiency (k cat /K m ) toward rutin was results of its low K m value while its high catalytic efficiency toward hesperidin was results of a considerably high k cat value. Copyright © 2017 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.
Full Text Available Fusarium solani NAN103 lipase was successfully overexpressed in Pichia pastoris using inducible expression system and constitutive expression system under the control of alcohol oxidase 1 promoter (pAOX1 and glyceraldehyde-3-phosphate dehydrogenase promoter (pGAP, respectively. Lipase obtained using the constitutive promoter showed the highest activity of 18.8 U/mg in 3 days of cultivation time. Optimal lipase activity was observed at pH 7.0 and 35 °C using p-nitrophenyl laurate as the substrate. Lipase activity was enhanced by Mn2+, Ba2+, Li+, Ca2+, Ni2+, CHAPS and Triton X-100 but was inhibited by Hg2+, Ag+ and SDS. The addition of 10% v/v of octanol, p-xylene, hexane and isopropanol increased lipase activity. Cultivation of lipase-expressing P. pastoris under pGAP in synthetic wastewater containing 1% w/v palm oil resulted in degradation of 87% of the oil within 72 h. P. pastoris expressing F. solani lipase from constitutive expression system has the potential to be used as an alternative microorganism for lipid degradation.
Joshi, Swati; Satyanarayana, Tulasi
The phytase of the yeast Pichia anomala is a histidine acid phosphatase based on signature sequences and catalytic amino acids identified by site-directed mutagenesis. Among modulators, N-bromosuccinimide and butanedione inhibit phytase, while Ca(2+) and Ni(2+) stimulate slightly. Vanadate exhibits competitive inhibition of phytase, making it bifunctional to act as haloperoxidase. Molecular docking supports vanadate to share its binding site with phytate. The T 1/2, activation energy (E a ), temperature quotient (Q 10), activation energy of thermal inactivation (Ed), and enthalpy (ΔH d (0) ) of the enzyme are 4.0 min (80 °C), 27.72 kJ mol(-1), 2.1, 410.62 kJ mol(-1), and ∼407.8 kJ mol(-1) (65-80 °C), respectively. The free energy of the process (ΔG d (o) ) increases from 49.56 to 71.58 kJ mol(-1) with rise in temperature, while entropy of inactivation (ΔS d (0) ) remains constant at ∼1.36 kJ mol(-1) K(-1). The supplementation of whole wheat dough with rPPHY resulted in 72.5 % reduction in phytic acid content of bread. These characteristics confirm that the phytase has adequate thermostability for its applicability as a food and feed additive.
Hellström, A; Qvirist, L; Svanberg, U; Veide Vilg, J; Andlid, T
Mineral deficiencies cause several health problems in the world, especially for populations consuming cereal-based diets rich in the anti-nutrient phytate. Our aim was to characterize the phytate-degrading capacity of the yeast Pichia kudriavzevii TY13 and its secretion of phytase. The phytase activity in cell-free supernatants from cultures with 100% intact cells was 35-190 mU ml(-1) depending on the media. The Km was 0.28 mmol l(-1) and the specific phytase activity 0.32 U mg(-1) total protein. The phytase activity and secretion of extracellular non-cell-bound phytase was affected by the medium phosphate concentrations. Further, addition of yeast extract had a clearly inducing effect, resulting in over 60% of the cultures total phytase activity as non-cell-bound. Our study reveals that it is possible to achieve high extracellular phytase activity from the yeast P. kudriavzevii TY13 by proper composition of the growth medium. TY13 could be a promising future starter culture for fermented foods with improved mineral bioavailability. Using strains that secrete phytase to the food matrix may significantly improve the phytate degradation by facilitating the enzyme-to-substrate interaction. The secreted non-cell-bound phytase activities by TY13 could further be advantageous in industrial production of phytase. © 2015 The Society for Applied Microbiology.
Azad, Abul Kalam; Sawa, Yoshihiro; Ishikawa, Takahiro; Shibata, Hitoshi
Water channels formed by aquaporins (AQPs) play an important role in the control of water homeostasis in individual cells and in multicellular organisms. Plasma membrane intrinsic proteins (PIPs) constitute a subclass of plant AQPs. TgPIP2;1 and TgPIP2;2 from tulip petals are members of the PIP family. In this study, we overexpressed TgPIP2;1 and TgPIP2;2 in Pichia pastoris and monitored their water channel activity (WCA) either by an in vivo spheroplast-bursting assay performed after hypo-osmotic shock or by growth assay. Osmolarity, pH, and inhibitors of AQPs, protein kinases (PKs), and protein phosphatases (PPs) affect the WCA of heterologous AQPs in this expression system. The WCA of TgPIP2;2-expressing spheroplasts was affected by inhibitors of PKs and PPs, which indicates that the water channel of this homologue is regulated by phosphorylation in P. pastoris. From the results reported herein, we suggest that P. pastoris can be employed as a heterologous expression system to assay the WCA of PIPs and to monitor the AQP-mediated channel gating mechanism, and it can be developed to screen inhibitors/effectors of PIPs. PMID:19251885
Azad, Abul Kalam; Sawa, Yoshihiro; Ishikawa, Takahiro; Shibata, Hitoshi
Water channels formed by aquaporins (AQPs) play an important role in the control of water homeostasis in individual cells and in multicellular organisms. Plasma membrane intrinsic proteins (PIPs) constitute a subclass of plant AQPs. TgPIP2;1 and TgPIP2;2 from tulip petals are members of the PIP family. In this study, we overexpressed TgPIP2;1 and TgPIP2;2 in Pichia pastoris and monitored their water channel activity (WCA) either by an in vivo spheroplast-bursting assay performed after hypo-osmotic shock or by growth assay. Osmolarity, pH, and inhibitors of AQPs, protein kinases (PKs), and protein phosphatases (PPs) affect the WCA of heterologous AQPs in this expression system. The WCA of TgPIP2;2-expressing spheroplasts was affected by inhibitors of PKs and PPs, which indicates that the water channel of this homologue is regulated by phosphorylation in P. pastoris. From the results reported herein, we suggest that P. pastoris can be employed as a heterologous expression system to assay the WCA of PIPs and to monitor the AQP-mediated channel gating mechanism, and it can be developed to screen inhibitors/effectors of PIPs.
Full Text Available Indigenous environmental activists have clearly articulated their views on global climate change policy. The content of these views was explored during the 10-day 2008 World Conservation Congress (WCC in Barcelona. Data were primarily collected through interviews and participant observation. In addition, policy statements and declarations made by indigenous environmental activists from 2000 to 2009 were analysed to place the perspectives of indigenous leaders and environmental activists in the context of their decade-long struggle to gain negotiating power at the United Nations Framework Convention on Climate Change. This study examines the rhetorical strategies indigenous leaders from around the world use to gain political recognition and legitimacy in climate change negotiations. Two core principles, relating to a particular representation of indigenous environmental knowledge are identified as fundamental rhetorical tools. These are a belief that the earth is a living being with rights and the conviction that it is the responsibility of indigenous peoples to protect the earth from over-exploitation. However, reference to indigenous environmental knowledge is not the only rhetorical mechanism used by indigenous leaders in the climate debates. When faced with specific United Nations policies to combat climate change that could have a profound impact on their land rights, some indigenous leaders adopt a more confrontational response. Fearing that new polices would reinforce historical trends of marginalisation, indigenous leaders seeking recognition in climate change debates speak less about their ecological knowledge and responsibility to the earth and more about their shared histories of political and economic marginalisation and land dispossession, experienced first through colonialism and more recently through globalisation.
Full Text Available The School of Public Policy (SPP at the University of Calgary organized a conference to announce the establishment of its Indigenous Policy program and to share knowledge and stories about policy issues critical to Indigenous Peoples in Canada. The conference, titled “Beyond Reconciliation,” was held at the University of Calgary Downtown Campus on Nov. 21, 2016 and was attended by 73 participants. This included Indigenous elders, chiefs and leaders, and members of Indigenous organizations, including a women’s group. Also included were members of universities and academic institutions, including students; industry representatives from the oil and gas, pipeline, forestry, electricity, legal and financial sectors; as well as representatives from government and regulatory agencies. The purpose of the conference was established with the following abstract, which was circulated to speakers and participants: The School of Public Policy is establishing a new Indigenous Policy program in order to produce widely disseminated research and engage in outreach that covers an array of policy areas, such as health, education, self-government, and natural resource development. The program will directly engage Indigenous communities in the search for original, long-term, and evidence-based solutions, as part of an effort to improve our national capacity in problem-solving and policy development. The conference will provide a platform to launch the program, showcasing preliminary research and providing a venue for discussion of policy solutions. The conference included three moderated panel sessions and a keynote speaker.1 The first panel considered business and entrepreneurship in Indigenous communities; the second panel showcased case studies that are examining the experiences of Indigenous communities with natural resource development projects, and particularly their experiences with consultation and engagement. The final panel focused on ways of
Living in a compassionate community is not a new practice in First Nations communities; they have always recognized dying as a social experience. First Nations hold extensive traditional knowledge and have community-based practices to support the personal, familial, and community experiences surrounding end-of-life. However, western health systems were imposed and typically did not support these social and cultural practices at end of life. In fact, the different expectations of western medicine and the community related to end of life care has created stress and misunderstanding for both. One solution is for First Nations communities to develop palliative care programs so that people can receive care at home amongst their family, community and culture. Our research project "Improving End-of-Life Care in First Nations Communities" (EOLFN) was funded by the Canadian Institutes of Health Research [2010-2015] and was conducted in partnership with four First Nations communities in Canada (see www.eolfn.lakeheadu.ca). Results included a community capacity development approach to support Indigenous models of care at end-of-life. The workshop will describe the community capacity development process used to develop palliative care programs in First Nations communities. It will highlight the foundation to this approach, namely, grounding the program in community values and principles, rooted in individual, family, community and culture. Two First Nations communities will share stories about their experiences developing their own palliative care programs, which celebrated cultural capacity in their communities while enhancing medical palliative care services in a way that respected and integrated with their community cultural practices. This workshop shares the experiences of two First Nations communities who developed palliative care programs by building upon community culture, values and principles. The underlying model guiding development is shared.
Afonso, G. B.
We have performed observations of the sky alongside with the Indians of all Brazilian regions that made it possible localize many indigenous constellations. Some of these constellations are the same as the other South American Indians and Australian aborigines constellations. The scientific community does not have much of this information, which may be lost in one or two generations. In this work, we present a planetary-observatory that we have made in the Park of Science Newton Freire-Maia of Paraná State, in order to popularize the astronomical knowledge of the Brazilian Indians. The planetary consists, essentially, of a sphere of six meters in diameter and a projection cylinder of indigenous constellations. In this planetary we can identify a lot of constellations that we have gotten from the Brazilian Indians; for instance, the four seasonal constellations: the Tapir (spring), the Old Man (summer), the Deer (autumn) and the Rhea (winter). A two-meter height wooden staff that is posted vertically on the horizontal ground similar to a Gnomon and stones aligned with the cardinal points and the soltices directions constitutes the observatory. A stone circle of ten meters in diameter surrounds the staff and the aligned stones. During the day we observe the Sun apparent motions and at night the indigenous constellations. Due to the great community interest in our work, we are designing an itinerant indigenous planetary-observatory to be used in other cities mainly by indigenous and primary schools teachers.
Kandasamy, Y; Smith, R; Wright, I M R; Lumbers, E R
Rates of chronic kidney disease (CKD) among Indigenous groups in Australia exceed non-Indigenous rates eight-fold. Using kidney volume as a surrogate for nephron number, we carried out a study to determine if Indigenous neonates have a smaller kidney volume (and thus a reduced nephron number) from birth compared with non-Indigenous neonates. We recruited term and preterm neonates (Indigenous) and 39 term (13 Indigenous) neonates. TKV of Indigenous neonates was significantly lower at 32 weeks [12.0 (2.0) v. 15.4 (5.1) ml; P=0.03] and 38 weeks CA [18.6 (4.0) v. 22.6 (5.9) ml; P=0.04] respectively. Term Indigenous neonates also had smaller kidney volumes compared with non-Indigenous neonates. Despite a smaller kidney volume (and reduced nephron number), Indigenous neonates did not have a significantly lower eGFR. Indigenous neonates achieve similar eGFRs to Non-Indigenous neonates, presumably through a higher single nephron filtration rate. This places Indigenous neonates at a greater risk of long-term kidney damage later in life.
Ashourizadeh, Shayegheh; Schøtt, Thomas; Pişkinsüt Şengüler, Ece
Migrants may become entrepreneurs in their host countries. They may utilize their dual embeddedness in both the home country and the host country, and also use transnational links to gain a competitive advantage in exporting compared to indigenous entrepreneurs. Migrant entrepreneurs’ advantage may......, however, be contingent on attributes such as gender and education, especially among the first generation of migrants, in that being male and educated is more advantageous for migrants than for indigenous entrepreneurs. A representative sample of 50,371 entrepreneurs establishing or operating enterprises...... around the world was surveyed in the Global Entrepreneurship Monitor, which reports on migration and exporting. Hierarchical linear modeling shows that migrant entrepreneurs export more than indigenous entrepreneurs, especially in the first generation, and especially among educated and male migrants...
Storytelling, in its various forms, has often been described as a practice with great emancipatory potential. In turn, Indigenous knowledge shows great promise in guiding a participatory action research (PAR) methodology. Yet these two approaches are rarely discussed in relation to one another, nor, has much been written in terms of how these two approaches may work synergistically toward a decolonizing research approach. In this article, I report on a community-driven knowledge translation activity, the Peoples’ International Health Tribunal, as an exemplar of how narrative and PAR approaches, guided by local Indigenous knowledge, have great potential to build methodologically and ethically robust research processes. Implications for building globally relevant research alliances and scholarship are further discussed, particularly in relation to working with Indigenous communities. PMID:28462305
Indilinga: African Journal of Indigenous Knowledge Systems. ... Resource conservation and utilisation through indigenous knowledge in a tribal community of Orissa, ... \\'The snake will swallow you': supernatural snakes and the creation of the ...
Jun 6, 2016 ... Demand for fresh indigenous vegetables in Nigeria has increased ... greater returns from indigenous vegetables compared to conventional vegetables. ... In Kolli Hills, Tamil Nadu, monocropping of a single, non-edible variety ...
In the 1990s, indigenous knowledge has been fertile ground for research, and ... research and will appeal to both seasoned development professional as well as ... indigenous-knowledge issues with the University of Indonesia, the Institute of ...
Pyper, Evelyn; Henry, David; Yates, Erika A; Mecredy, Graham; Ratnasingham, Sujitha; Slegers, Brian; Walker, Jennifer D
Indigenous data governance principles assert that Indigenous communities have a right to data that identifies their people or communities, and a right to determine the use of that data in ways that support Indigenous health and self-determination. Indigenous-driven use of the databases held at the Institute for Clinical Evaluative Sciences (ICES) has resulted in ongoing partnerships between ICES and diverse Indigenous organizations and communities. To respond to this emerging and complex landscape, ICES has established a team whose goal is to support the infrastructure for responding to community-initiated research priorities. ICES works closely with Indigenous partners to develop unique data governance agreements and supports processes, which ensure that ICES scientists must work with Indigenous organizations when conducting research that involves Indigenous peoples. © 2018 Longwoods Publishing.
Jan 6, 2011 ... CASE STUDY: Chile — Health, environment, and indigenous culture .... For example, the National Corporation for Indigenous Development (CONADI) ... Institute for Agriculture Development (INDAP), and applied research on ...
vernacular mass media and the Kenyan indigenous languages. ... African indigenous languages had, "against all odds", survived as media of communication ..... regulations should, of course, primarily ensure quality and ethical journalism.
Indilinga: African Journal of Indigenous Knowledge Systems. ... halting the spread of HIV and AIDS in South Africa: The case of Soshanguve township in the ... Tourism policies and management practices as perceived by indigenous people in ...
A Physical Education Curriculum Enriched With Indigenous Zulu Games For Improved Social Development ... Therefore, it was necessary to assess these indigenous Zulu games' potential in obtaining overt ... AJOL African Journals Online.
Full Text Available This paper describes the features required of OWL (Web Ontology Language) to realise and enhance Indigenous Knowledge (IK) digital repositories. Several needs for Indigenous Knowledge management systems (IKMSs) are articulated, based on extensive...
Guo, Yu-Qi; Wu, Qing-Ping; Shao, Xiao-Xia; Shen, Ting; Liu, Ya-Li; Xu, Zeng-Guang; Guo, Zhan-Yun
Relaxin family peptides are a group of peptide hormones with divergent biological functions. Mature relaxin family peptides are typically composed of two polypeptide chains with three disulfide linkages, rendering their preparation a challenging task. In the present study, we established an efficient approach for preparation of the chimeric relaxin family peptide R3/I5 through secretory overexpression in Pichia pastoris and in vitro enzymatic maturation. A designed single-chain R3/I5 precursor containing the B-chain of human relaxin-3 and the A-chain of human INSL5 was overexpressed in PichiaPink strain 1 by high-density fermentation in a two-liter fermenter, and approximately 200 mg of purified precursor was obtained from one liter of the fermentation supernatant. We also developed an economical approach for preparation of the uniformly (15)N-labeled R3/I5 precursor by culturing in shaking flasks, and approximately 15 mg of purified (15)N-labeled precursor was obtained from one liter of the culture supernatant. After purification by cation ion-exchange chromatography and reverse-phase high performance liquid chromatography, the R3/I5 precursor was converted to the mature two-chain form by sequential treatment with endoproteinase Lys-C and carboxypeptidase B. The mature R3/I5 peptide had an α-helix-dominated conformation and retained full receptor-binding and receptor activation activities. Thus, Pichia overexpression was an efficient approach for sample preparation and isotopic labeling of the chimeric R3/I5 peptide. This approach could also be extended to the preparation of other relaxin family peptides in future studies.
Tomičić, Ružica; Raspor, Peter
An understanding of adhesion behavior of Candida and Pichia yeast under different environmental conditions is key to the development of effective preventive measures against biofilm-associated infection. Hence in this study we investigated the impact of growth medium and temperature on Candida and Pichia adherence using stainless steel (AISI 304) discs with different degrees of surface roughness (Ra = 25.20-961.9 nm), material typical for the food processing industry as well as medical devices. The adhesion of the yeast strains to stainless steel surfaces grown in Malt Extract broth (MEB) or YPD broth at three temperatures (7 °C, 37 °C, 43 °C for Candida strains and 7 °C, 27 °C, 32 °C for Pichia strains) was assessed by crystal violet staining. The results showed that the nutrient content of medium significantly influenced the quantity of adhered cells by the tested yeasts. Adhesion of C. albicans and C. glabrata on stainless steel surfaces were significantly higher in MEB, whereas for C. parapsilosis and C. krusei it was YPD broth. In the case with P. pijperi and P. membranifaciens, YPD broth was more effective in promoting adhesion than MEB. On the other hand, our data indicated that temperature is a very important factor which considerably affects the adhesion of these yeast. There was also significant difference in cell adhesion on all types of stainless steel surfaces for all tested yeast. Copyright © 2017 Elsevier Ltd. All rights reserved.
Very little has been written on indigenous rights in South-East Asia. This article attempts to address issues concerning indigenous land rights in the region, arguing that there is a clear gap between the existing situation and the relevant standards of the international human rights system. After a short overview of the international human rights framework currently binding South-East Asian states, the article analyses issues of indigenous land ownership and control by indigenous peoples ove...
Dubertret , Fabrice
This document is a draft. It aims at providing a basis for discussion between the different organizations and indigenous land and resources rights experts involved in the wider project of building a world atlas of indigenous territories.; This working paper describes the process of establishing a global categorization of indigenous land and resources rights. From the analysis of a great variability of legislations regarding indigenous territories, common considered topics are identified, such...
Razvi, M.A.N.; Jayasekharan, T.; Thankarajan, K.; Guhagarkar, M.B.; Dixit, M.N.; Bhale, G.L.
Design, fabrication and performance testing of an indigenously built Resonance Ionization Mass Spectrometer (RIMS) is presented in this report. The instrument is totally indigenous, but for the laser components consisting of the excimer laser and tunable dye lasers. Constructional details of atomic beam source and linear time-of-flight mass spectrometer are included. Finally, commissioning and performance testing of the instrument is described. Mass resolving power of 400 and a detection limit of 100 atoms has been achieved using this RIMS set-up. (author)
Dodgson, Joan E; Struthers, Roxanne
Marginalization may affect health care delivery. Ways in which indigenous women experienced marginalization were examined. Data from 57 indigenous women (18 to 65 years) were analyzed for themes. Three themes emerged: historical trauma as lived marginalization, biculturalism experienced as marginalization, and interacting within a complex health care system. Experienced marginalization reflected participants' unique perspective and were congruent with previous research. It is necessary for health care providers to assess the detrimental impact of marginalization on the health status of individuals and/or communities.
Petersen, Bent; Egelund, Jack; Damager, Iben
.011 to 0.013 U (1 U = 1 nmol conversion of substrate * min(-1) * microl medium(-1)) similar to those of RGXT1 and RGXT2 expressed in Baculovirus transfected insect Sf9 cells. In summary, the data presented suggest that Pichia is an attractive host candidate for expression of plant glycosyltransferases.......Two Arabidopsis xylosyltransferases, designated RGXT1 and RGXT2, were recently expressed in Baculovirus transfected insect cells and by use of the free sugar assay shown to catalyse transfer of D-xylose from UDP-alpha-D-xylose to L-fucose and derivatives hereof. We have now examined expression...
Slininger, P J; Dien, B S; Lomont, J M; Bothast, R J; Ladisch, M R; Okos, M R
Scheffersomyces (formerly Pichia) stipitis is a potential biocatalyst for converting lignocelluloses to ethanol because the yeast natively ferments xylose. An unstructured kinetic model based upon a system of linear differential equations has been formulated that describes growth and ethanol production as functions of ethanol, oxygen, and xylose concentrations for both growth and fermentation stages. The model was validated for various growth conditions including batch, cell recycle, batch with in situ ethanol removal and fed-batch. The model provides a summary of basic physiological yeast properties and is an important tool for simulating and optimizing various culture conditions and evaluating various bioreactor designs for ethanol production. © 2014 Wiley Periodicals, Inc.
Wu, Song; Xiao, Yong; Wang, Lu; Zheng, Yue; Chang, Kenlin; Zheng, Zhiyong; Yang, Zhaohui; Varcoe, John R.; Zhao, Feng
Extracellular electron transfer (EET) of microorganisms represents a communicative bridge between the interior and exterior of the cells. Most prior EET studies have focused on Gram-negative bacteria. However, fungi and Gram-positive bacteria, that contain dense cellular walls, have rarely been reported. Herein, two model dense cell wall microorganisms (Bacillus sp. WS-XY1 and the yeast Pichia stipitis) were identified to be electrochemically active. Further analysis indicated that the two microorganisms were able to secrete flavins to mediate their EET. The discovery, that dense cell wall containing microorganisms can undertake mediated EET, adds to the body of knowledge towards building a comprehensive understanding of biogeochemical and bioelectrical processes
Oledzka, G.; Dabrowski, Slawomir; Kur, J.
Aqualysin I is a heat-stable subtilisin-type serine protease which is secreted into the culture medium by Thermus aquaticus YT-1, an extreme thermophile. We report the high-level expression of an aqualysin I protein using its native signal sequence for secretion in the methylotrophic yeast, Pichia...... to that of the native enzyme. We also explored the possibility of secreting the GAP expressed aqualysin I in P. pastoris by in-frame fusion of the Saccharomyces cerevisiae alpha-factor secretion signal. However, the levels of secreted pro-aqualysin I particles were approximately 10 times lower, possibly...
Carter, Jennifer; Hollinsworth, David
Australian universities are increasingly embedding Indigenous content and perspectives within curriculum to promote Indigenous cultural competency. We present teaching challenges in an Indigenous geography course designed to present an engaged, intercultural learning experience. We critically reflect on student evaluations, informal discussions…
What is the relationship between Indigenous peoples and violent reactions to contemporary states? This research explores differing, culturally informed notions of attachment to land or place territory. Mechanistic ties and organic ties to land are linked to a key distinction between Indigenous peoples and non-Indigenous peoples. Utilizing the…
Full Text Available The formal reconciliation process in Australia was conducted between 1991 and 2000 and aimed to reconcile Indigenous and non-Indigenous peoples by 2001. In this paper, I detail the failure of both this reconciliation process and governments, in particular the Howard Government, to recognise Indigenous rights, such as sovereignty, a treaty, self-determination and land rights.
Indigenous counselling has not been given attention in Nigeria's school counselling programme. This counselling gap was created by European colonialism, which succeeded in developing in the minds of the African that anything indigenous is local, unscientific and unorthodox. Indigenous counselling is one of the ...
Hohepa, Margie Kahukura (Ngapuhi)
Educational leadership, it is argued, must play a critical role in improving student outcomes, especially those of minoritized and Indigenous students. In the process of improving education and schooling for Indigenous students, Indigenous educational leadership needs to be considered alongside educational leadership more generally. This article…
Liu, Wan-Cang; Zhu, Ping
Pichia pastoris has been one of the most successful heterologous overexpression systems in generating proteins for large-scale production through high-cell-density fermentation. However, optimizing conditions of the large-scale high-cell-density fermentation for biochemistry and industrialization is usually a laborious and time-consuming process. Furthermore, it is often difficult to produce authentic proteins in large quantities, which is a major obstacle for functional and structural features analysis and industrial application. For these reasons, we have developed a protocol for efficient demonstration-scale high-cell-density fermentation of P. pastoris, which employs a new methanol-feeding strategy-biomass-stat strategy and a strategy of increased air pressure instead of pure oxygen supplement. The protocol included three typical stages of glycerol batch fermentation (initial culture phase), glycerol fed-batch fermentation (biomass accumulation phase), and methanol fed-batch fermentation (induction phase), which allows direct online-monitoring of fermentation conditions, including broth pH, temperature, DO, anti-foam generation, and feeding of glycerol and methanol. Using this protocol, production of the recombinant β-xylosidase of Lentinula edodes origin in 1000-L scale fermentation can be up to ~900 mg/L or 9.4 mg/g cells (dry cell weight, intracellular expression), with the specific production rate and average specific production of 0.1 mg/g/h and 0.081 mg/g/h, respectively. The methodology described in this protocol can be easily transferred to other systems, and eligible to scale up for a large number of proteins used in either the scientific studies or commercial purposes.
Avril, Marion; Hathaway, Marianne J; Cartwright, Megan M; Gose, Severin O; Narum, David L; Smith, Joseph D
VAR2CSA is the main candidate for a vaccine against pregnancy-associated malaria, but vaccine development is complicated by the large size and complex disulfide bonding pattern of the protein. Recent X-ray crystallographic information suggests that domain boundaries of VAR2CSA Duffy binding-like (DBL) domains may be larger than previously predicted and include two additional cysteine residues. This study investigated whether longer constructs would improve VAR2CSA recombinant protein secretion from Pichia pastoris and if domain boundaries were applicable across different VAR2CSA alleles. VAR2CSA sequences were bioinformatically analysed to identify the predicted C11 and C12 cysteine residues at the C-termini of DBL domains and revised N- and C-termimal domain boundaries were predicted in VAR2CSA. Multiple construct boundaries were systematically evaluated for protein secretion in P. pastoris and secreted proteins were tested as immunogens. From a total of 42 different VAR2CSA constructs, 15 proteins (36%) were secreted. Longer construct boundaries, including the predicted C11 and C12 cysteine residues, generally improved expression of poorly or non-secreted domains and permitted expression of all six VAR2CSA DBL domains. However, protein secretion was still highly empiric and affected by subtle differences in domain boundaries and allelic variation between VAR2CSA sequences. Eleven of the secreted proteins were used to immunize rabbits. Antibodies reacted with CSA-binding infected erythrocytes, indicating that P. pastoris recombinant proteins possessed native protein epitopes. These findings strengthen emerging data for a revision of DBL domain boundaries in var-encoded proteins and may facilitate pregnancy malaria vaccine development.
Tredwell, Gregory D.; Edwards-Jones, Bryn; Leak, David J.; Bundy, Jacob G.
Metabolic profiling is increasingly being used to investigate a diverse range of biological questions. Due to the rapid turnover of intracellular metabolites it is important to have reliable, reproducible techniques for sampling and sample treatment. Through the use of non-targeted analytical techniques such as NMR and GC-MS we have performed a comprehensive quantitative investigation of sampling techniques for Pichia pastoris. It was clear that quenching metabolism using solutions based on the standard cold methanol protocol caused some metabolite losses from P. pastoris cells. However, these were at a low level, with the NMR results indicating metabolite increases in the quenching solution below 5% of their intracellular level for 75% of metabolites identified; while the GC-MS results suggest a slightly higher level with increases below 15% of their intracellular values. There were subtle differences between the four quenching solutions investigated but broadly, they all gave similar results. Total culture extraction of cells + broth using high cell density cultures typical of P. pastoris fermentations, was an efficient sampling technique for NMR analysis and provided a gold standard of intracellular metabolite levels; however, salts in the media affected the GC-MS analysis. Furthermore, there was no benefit in including an additional washing step in the quenching process, as the results were essentially identical to those obtained just by a single centrifugation step. We have identified the major high-concentration metabolites found in both the extra- and intracellular locations of P. pastoris cultures by NMR spectroscopy and GC-MS. This has provided us with a baseline metabolome for P. pastoris for future studies. The P. pastoris metabolome is significantly different from that of Saccharomyces cerevisiae, with the most notable difference being the production of high concentrations of arabitol by P. pastoris. PMID:21283710
Saulo Martins Vieira
Full Text Available Snakebite envenomation is a neglected condition that constitutes a public health problem in tropical and subtropical countries, including Brazil. Interestingly, some animals are resistant to snake envenomation due to the presence of inhibitory glycoproteins in their serum that target toxic venom components. DM64 is an acidic glycoprotein isolated from Didelphis aurita (opossum serum that has been characterized as an inhibitor of the myotoxicity induced by bothropic toxins bearing phospholipase A2 (PLA2 structures. This antitoxic protein can serve as an excellent starting template for the design of novel therapeutics against snakebite envenomation, particularly venom-induced local tissue damage. Therefore, the aim of this work was to produce a recombinant DM64 (rDM64 in the methylotrophic yeast Pichia pastoris and to compare its biological properties with those of native DM64. Yeast fermentation in the presence of Pefabloc, a serine protease inhibitor, stimulated cell growth (~1.5-fold, increased the rDM64 production yield approximately 10-fold and significantly reduced the susceptibility of rDM64 to proteolytic degradation. P. pastoris fermentation products were identified by mass spectrometry and Western blotting. The heterologous protein was efficiently purified from the culture medium by affinity chromatography (with immobilized PLA2 myotoxin and/or an ion exchange column. Although both native and recombinant DM64 exhibit different glycosylation patterns, they show very similar electrophoretic mobilities after PNGase F treatment. rDM64 formed a noncovalent complex with myotoxin II (Lys49-PLA2 from Bothrops asper and displayed biological activity that was similar to that of native DM64, inhibiting the cytotoxicity of myotoxin II by 92% at a 1:1 molar ratio.
Hamaker, Kent H; Johnson, Daniel C; Bellucci, Joseph J; Apgar, Kristie R; Soslow, Sherry; Gercke, John C; Menzo, Darrin J; Ton, Christopher
Large-scale fermentation of Pichia pastoris requires a large volume of methanol feed during the induction phase. However, a large volume of methanol feed is difficult to use in the processing suite because of the inconvenience of constant monitoring, manual manipulation steps, and fire and explosion hazards. To optimize and improve safety of the methanol feed process, a novel automated methanol feed system has been designed and implemented for industrial fermentation of P. pastoris. Details of the design of the methanol feed system are described. The main goals of the design were to automate the methanol feed process and to minimize the hazardous risks associated with storing and handling large quantities of methanol in the processing area. The methanol feed system is composed of two main components: a bulk feed (BF) system and up to three portable process feed (PF) systems. The BF system automatically delivers methanol from a central location to the portable PF system. The PF system provides precise flow control of linear, step, or exponential feed of methanol to the fermenter. Pilot-scale fermentations with linear and exponential methanol feeds were conducted using two Mut(+) (methanol utilization plus) strains, one expressing a recombinant therapeutic protein and the other a monoclonal antibody. Results show that the methanol feed system is accurate, safe, and efficient. The feed rates for both linear and exponential feed methods were within ± 5% of the set points, and the total amount of methanol fed was within 1% of the targeted volume. Copyright © 2011 American Institute of Chemical Engineers (AIChE).
Xi, Caixia; Liu, Nan; Liang, Fei; Zhao, Xiao; Long, Juan; Yuan, Fang; Yun, Song; Sun, Yuying; Xi, Yongzhi
Effective treatment of rheumatoid arthritis can be mediated by native chicken type II collagen (nCCII), recombinant peptide containing nCCII tolerogenic epitopes (CTEs), or a therapeutic DNA vaccine encoding the full-length CCOL2A1 cDNA. As recombinant CCII (rCCII) might avoid potential pathogenic virus contamination during nCCII preparation or chromosomal integration and oncogene activation associated with DNA vaccines, here we evaluated the importance of propeptide and telopeptide domains on rCCII triple helix molecular assembly. We constructed pC- and pN-procollagen (without N- or Cpropeptides, respectively) as well as CTEs located in the triple helical domain lacking both propeptides and telopeptides, and expressed these in yeast Pichia pastoris host strain GS115 (his4, Mut + ) simultaneously with recombinant chicken prolyl-4-hydroxylase α and β subunits. Both pC- and pN-procollagen monomers accumulated inside P. pastoris cells, whereas CTE was assembled into homotrimers with stable conformation and secreted into the supernatants, suggesting that the large molecular weight pC-or pN-procollagens were retained within the endoplasmic reticulum whereas the smaller CTEs proceeded through the secretory pathway. Furthermore, resulting recombinant chicken type II collagen pCα1(II) can induced collagen-induced arthritis (CIA) rat model, which seems to be as effective as the current standard nCCII. Notably, protease digestion assays showed that rCCII could assemble in the absence of C- and N-propeptides or telopeptides. These findings provide new insights into the minimal structural requirements for rCCII expression and folding.
Trinh, L B; Phue, J N; Shiloach, Joseph
Pichia pastoris, a methylotrophic yeast, is an efficient producer of recombinant proteins in which the heterologous gene is under the control of the methanol-induced AOX1 promoter. Hence, the accepted production procedure has two phases: In the first phase, the yeast utilizes glycerol and biomass is accumulated; in the second phase, the yeast utilizes methanol which is used both as an inducer for the expression of the recombinant protein and as a carbon source. Since the yeast is sensitive to methanol concentration, the methanol is supplied gradually to the growing culture. Three methanol addition strategies were evaluated for the purpose of optimizing recombinant endostatin production. Two strategies were based on the yeast metabolism; one responding to the methanol consumption using a methanol sensor, and the other responding to the oxygen consumption. In these two strategies, the methanol supply is unlimited. The third strategy was based on a predetermined exponential feeding rate, controling the growth rate at 0.02 h(-1), in this strategy the methanol supply is limited. Throughout the induction phase glycerol, in addition to methanol, was continuously added at a rate of 1 g L h(-1). Total endostatin production was similar in all three strategies, (400 mg was obtained from 3 L initial volume), but the amount of methanol added and the biomass produced were lower in the predetermined rate method. This caused the specific production of endostatin per biomass and per methanol to be 2 times higher in the predetermined rate than in the other two methods, making the growth control strategy not only more efficient but also more convenient for downstream processing. Copyright 2003 Wiley Periodicals, Inc. Biotechnol Bioeng 82: 438-444, 2003.
Full Text Available Wasp venom allergy is the most common insect venom allergy in Europe. It is manifested by large local reaction or anaphylactic shock occurring after a wasp sting. The allergy can be treated by specific immunotherapy with whole venom extracts. Wasp venom is difficult and costly to obtain and is a subject to composition variation, therefore it can be advantageous to substitute it with a cocktail of recombinant allergens. One of the major venom allergens is phospholipase A1, which so far has been expressed in Escherichia coli and in insect cells. Our aim was to produce the protein in secreted form in yeast Pichia pastoris, which can give high yields of correctly folded protein on defined minimal medium and secretes relatively few native proteins simplifying purification.Residual amounts of enzymatically active phospholipase A1 could be expressed, but the venom protein had a deleterious effect on growth of the yeast cells. To overcome the problem we introduced three different point mutations at the critical points of the active site, where serine137, aspartate165 or histidine229 were replaced by alanine (S137A, D165A and H229A. All the three mutated forms could be expressed in P. pastoris. The H229A mutant did not have any detectable phospholipase A1 activity and was secreted at the level of several mg/L in shake flask culture. The protein was purified by nickel-affinity chromatography and its identity was confirmed by MALDI-TOF mass spectrometry. The protein could bind IgE antibodies from wasp venom allergic patients and could inhibit the binding of wasp venom to IgE antibodies specific for phospholipase A1 as shown by Enzyme Allergo-Sorbent Test (EAST. Moreover, the recombinant protein was allergenic in a biological assay as demonstrated by its capability to induce histamine release of wasp venom-sensitive basophils.The recombinant phospholipase A1 presents a good candidate for wasp venom immunotherapy.
Full Text Available Carboxypeptidase M (CPM is an extracellular glycosylphosphatidyl-inositol-anchored membrane glycoprotein, which removes the C-terminal basic residues, lysine and arginine, from peptides and proteins at neutral pH. CPM plays an important role in the control of peptide hormones and growth factor activity on the cell surface. The present study was carried out to clone and express human CPM in the yeast Pichia pastoris in order to evaluate the importance of this enzyme in physiological and pathological processes. The cDNA for the enzyme was amplified from total placental RNA by RT-PCR and cloned in the vector pPIC9, which uses the methanol oxidase promoter and drives the expression of high levels of heterologous proteins in P. pastoris. The cpm gene, after cloning and transfection, was integrated into the yeast genome, which produced the active protein. The recombinant protein was secreted into the medium and the enzymatic activity was measured using the fluorescent substrate dansyl-Ala-Arg. The enzyme was purified by a two-step protocol including gel filtration and ion-exchange chromatography, resulting in a 1753-fold purified active protein (16474 RFU mg protein-1 min-1. This purification protocol permitted us to obtain 410 mg of the purified protein per liter of fermentation medium. SDS-PAGE showed that recombinant CPM migrated as a single band with a molecular mass similar to that of native placental enzyme (62 kDa, suggesting that the expression of a glycosylated protein had occurred. These results demonstrate for the first time the establishment of a method using P. pastoris to express human CPM necessary to the development of specific antibodies and antagonists, and the analysis of the involvement of this peptidase in different physiological and pathological processes
Narum David L
Full Text Available Abstract Background VAR2CSA is the main candidate for a vaccine against pregnancy-associated malaria, but vaccine development is complicated by the large size and complex disulfide bonding pattern of the protein. Recent X-ray crystallographic information suggests that domain boundaries of VAR2CSA Duffy binding-like (DBL domains may be larger than previously predicted and include two additional cysteine residues. This study investigated whether longer constructs would improve VAR2CSA recombinant protein secretion from Pichia pastoris and if domain boundaries were applicable across different VAR2CSA alleles. Methods VAR2CSA sequences were bioinformatically analysed to identify the predicted C11 and C12 cysteine residues at the C-termini of DBL domains and revised N- and C-termimal domain boundaries were predicted in VAR2CSA. Multiple construct boundaries were systematically evaluated for protein secretion in P. pastoris and secreted proteins were tested as immunogens. Results From a total of 42 different VAR2CSA constructs, 15 proteins (36% were secreted. Longer construct boundaries, including the predicted C11 and C12 cysteine residues, generally improved expression of poorly or non-secreted domains and permitted expression of all six VAR2CSA DBL domains. However, protein secretion was still highly empiric and affected by subtle differences in domain boundaries and allelic variation between VAR2CSA sequences. Eleven of the secreted proteins were used to immunize rabbits. Antibodies reacted with CSA-binding infected erythrocytes, indicating that P. pastoris recombinant proteins possessed native protein epitopes. Conclusion These findings strengthen emerging data for a revision of DBL domain boundaries in var-encoded proteins and may facilitate pregnancy malaria vaccine development.
Full Text Available Abstract Background Pichia pastoris is widely used as a production platform for heterologous proteins and model organism for organelle proliferation. Without a published genome sequence available, strain and process development relied mainly on analogies to other, well studied yeasts like Saccharomyces cerevisiae. Results To investigate specific features of growth and protein secretion, we have sequenced the 9.4 Mb genome of the type strain DSMZ 70382 and analyzed the secretome and the sugar transporters. The computationally predicted secretome consists of 88 ORFs. When grown on glucose, only 20 proteins were actually secreted at detectable levels. These data highlight one major feature of P. pastoris, namely the low contamination of heterologous proteins with host cell protein, when applying glucose based expression systems. Putative sugar transporters were identified and compared to those of related yeast species. The genome comprises 2 homologs to S. cerevisiae low affinity transporters and 2 to high affinity transporters of other Crabtree negative yeasts. Contrary to other yeasts, P. pastoris possesses 4 H+/glycerol transporters. Conclusion This work highlights significant advantages of using the P. pastoris system with glucose based expression and fermentation strategies. As only few proteins and no proteases are actually secreted on glucose, it becomes evident that cell lysis is the relevant cause of proteolytic degradation of secreted proteins. The endowment with hexose transporters, dominantly of the high affinity type, limits glucose uptake rates and thus overflow metabolism as observed in S. cerevisiae. The presence of 4 genes for glycerol transporters explains the high specific growth rates on this substrate and underlines the suitability of a glycerol/glucose based fermentation strategy. Furthermore, we present an open access web based genome browser http://www.pichiagenome.org.
Full Text Available Carboxypeptidase M (CPM is an extracellular glycosylphosphatidyl-inositol-anchored membrane glycoprotein, which removes the C-terminal basic residues, lysine and arginine, from peptides and proteins at neutral pH. CPM plays an important role in the control of peptide hormones and growth factor activity on the cell surface. The present study was carried out to clone and express human CPM in the yeast Pichia pastoris in order to evaluate the importance of this enzyme in physiological and pathological processes. The cDNA for the enzyme was amplified from total placental RNA by RT-PCR and cloned in the vector pPIC9, which uses the methanol oxidase promoter and drives the expression of high levels of heterologous proteins in P. pastoris. The cpm gene, after cloning and transfection, was integrated into the yeast genome, which produced the active protein. The recombinant protein was secreted into the medium and the enzymatic activity was measured using the fluorescent substrate dansyl-Ala-Arg. The enzyme was purified by a two-step protocol including gel filtration and ion-exchange chromatography, resulting in a 1753-fold purified active protein (16474 RFU mg protein-1 min-1. This purification protocol permitted us to obtain 410 mg of the purified protein per liter of fermentation medium. SDS-PAGE showed that recombinant CPM migrated as a single band with a molecular mass similar to that of native placental enzyme (62 kDa, suggesting that the expression of a glycosylated protein had occurred. These results demonstrate for the first time the establishment of a method using P. pastoris to express human CPM necessary to the development of specific antibodies and antagonists, and the analysis of the involvement of this peptidase in different physiological and pathological processes
Full Text Available Abstract Background FAD dependent glucose dehydrogenase (GDH currently raises enormous interest in the field of glucose biosensors. Due to its superior properties such as high turnover rate, substrate specificity and oxygen independence, GDH makes its way into glucose biosensing. The recently discovered GDH from the ascomycete Glomerella cingulata is a novel candidate for such an electrochemical application, but also of interest to study the plant-pathogen interaction of a family of wide-spread, crop destroying fungi. Heterologous expression is a necessity to facilitate the production of GDH for biotechnological applications and to study its physiological role in the outbreak of anthracnose caused by Glomerella (anamorph Colletotrichum spp. Results Heterologous expression of active G. cingulata GDH has been achieved in both Escherichia coli and Pichia pastoris, however, the expressed volumetric activity was about 4800-fold higher in P. pastoris. Expression in E. coli resulted mainly in the formation of inclusion bodies and only after co-expression with molecular chaperones enzymatic activity was detected. The fed-batch cultivation of a P. pastoris transformant resulted in an expression of 48,000 U L-1 of GDH activity (57 mg L-1. Recombinant GDH was purified by a two-step purification procedure with a yield of 71%. Comparative characterization of molecular and catalytic properties shows identical features for the GDH expressed in P. pastoris and the wild-type enzyme from its natural fungal source. Conclusions The heterologous expression of active GDH was greatly favoured in the eukaryotic host. The efficient expression in P. pastoris facilitates the production of genetically engineered GDH variants for electrochemical-, physiological- and structural studies.
Sygmund, Christoph; Staudigl, Petra; Klausberger, Miriam; Pinotsis, Nikos; Djinović-Carugo, Kristina; Gorton, Lo; Haltrich, Dietmar; Ludwig, Roland
FAD dependent glucose dehydrogenase (GDH) currently raises enormous interest in the field of glucose biosensors. Due to its superior properties such as high turnover rate, substrate specificity and oxygen independence, GDH makes its way into glucose biosensing. The recently discovered GDH from the ascomycete Glomerella cingulata is a novel candidate for such an electrochemical application, but also of interest to study the plant-pathogen interaction of a family of wide-spread, crop destroying fungi. Heterologous expression is a necessity to facilitate the production of GDH for biotechnological applications and to study its physiological role in the outbreak of anthracnose caused by Glomerella (anamorph Colletotrichum) spp. Heterologous expression of active G. cingulata GDH has been achieved in both Escherichia coli and Pichia pastoris, however, the expressed volumetric activity was about 4800-fold higher in P. pastoris. Expression in E. coli resulted mainly in the formation of inclusion bodies and only after co-expression with molecular chaperones enzymatic activity was detected. The fed-batch cultivation of a P. pastoris transformant resulted in an expression of 48,000 U L⁻¹ of GDH activity (57 mg L⁻¹). Recombinant GDH was purified by a two-step purification procedure with a yield of 71%. Comparative characterization of molecular and catalytic properties shows identical features for the GDH expressed in P. pastoris and the wild-type enzyme from its natural fungal source. The heterologous expression of active GDH was greatly favoured in the eukaryotic host. The efficient expression in P. pastoris facilitates the production of genetically engineered GDH variants for electrochemical-, physiological- and structural studies.
Shibui, Tatsuro; Hara, Hiroyoshi
Pichia pastoris has been used for the production of many recombinant proteins, and many useful mutant strains have been created. However, the efficiency of mutant isolation by gene-targeting is usually low and the procedure is difficult for those inexperienced in yeast genetics. In order to overcome these issues, we developed a new gene-disruption system with a rescue gene using an inducible Cre/mutant-loxP system. With only short homology regions, the gene-disruption cassette of the system replaces its target-gene locus containing a mutation with a compensatory rescue gene. As the cassette contains the AOX1 promoter-driven Cre gene, when targeted strains are grown on media containing methanol, the DNA fragment, i.e., the marker, rescue and Cre genes, between the mutant-loxP sequences in the cassette is excised, leaving only the remaining mutant-loxP sequence in the genome, and consequently a target gene-disrupted mutant can be isolated. The system was initially validated on ADE2 gene disruption, where the disruption can easily be detected by color-change of the colonies. Then, the system was applied for knocking-out URA3 and OCH1 genes, reported to be difficult to accomplish by conventional gene-targeting methods. All three gene-disruption cassettes with their rescue genes replaced their target genes, and the Cre/mutant-loxP system worked well to successfully isolate their knock-out mutants. This study identified a new gene-disruption system that could be used to effectively and strategically knock out genes of interest, especially whose deletion is detrimental to growth, without using special strains, e.g., deficient in nonhomologous end-joining, in P. pastoris. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1201-1208, 2017. © 2017 American Institute of Chemical Engineers.
Yang, Jinlong; Cheng, Yichao; Zhu, Yuanyuan; Zhu, Junjun; Chen, Tingting; Xu, Yong; Yong, Qiang; Yu, Shiyuan
Lignin degradation products are toxic to microorganisms, which is one of the bottlenecks for fuel ethanol production. We studied the effects of phenolic ketones (4-hydroxyacetophenone, 4-hydroxy-3-methoxy-acetophenone and 4-hydroxy-3,5-dimethoxy-acetophenone) derived from lignin degradation on ethanol fermentation of xylose and cellular lipid composition of Pichia stipitis NLP31. Ethanol and the cellular fatty acid of yeast were analyzed by high performance liquid chromatography (HPLC) and gas chromatography/mass spectrometry (GC/MS). Results indicate that phenolic ketones negatively affected ethanol fermentation of yeast and the lower molecular weight phenolic ketone compound was more toxic. When the concentration of 4-hydroxyacetophenone was 1.5 g/L, at fermentation of 24 h, the xylose utilization ratio, ethanol yield and ethanol concentration decreased by 42.47%, 5.30% and 9.76 g/L, respectively, compared to the control. When phenolic ketones were in the medium, the ratio of unsaturated fatty acids to saturated fatty acids (UFA/SFA) of yeast cells was improved. When 1.5 g/L of three aforementioned phenolic ketones was added to the fermentation medium, the UFA/SFA ratio of yeast cells increased to 3.03, 3.06 and 3.61, respectively, compared to 2.58 of the control, which increased cell membrane fluidity and instability. Therefore, phenolic ketones can reduce the yeast growth, increase the UFA/SFA ratio of yeast and lower ethanol productivity. Effectively reduce or remove the content of lignin degradation products is the key to improve lignocellulose biorefinery.
Huang, Peng; Shi, Jinlei; Sun, Qingwen; Dong, Xianping; Zhang, Ning
Lysozymes are known as ubiquitously distributed immune effectors with hydrolytic activity against peptidoglycan, the major bacterial cell wall polymer, to trigger cell lysis. In the present study, the full-length cDNA sequence of a novel sea urchin Strongylocentrotus purpuratus invertebrate-type lysozyme (sp-iLys) was synthesized according to the codon usage bias of Pichia pastoris and was cloned into a constitutive expression plasmid pPIC9K. The resulting plasmid, pPIC9K-sp-iLys, was integrated into the genome of P. pastoris strain GS115. The bioactive recombinant sp-iLys was successfully secreted into the culture broth by positive transformants. The highest lytic activity of 960 U/mL of culture supernatant was reached in fed-batch fermentation. Using chitin affinity chromatography and gel-filtration chromatography, recombinant sp-iLys was produced with a yield of 94.5 mg/L and purity of > 99%. Recombinant sp-iLys reached its peak lytic activity of 8560 U/mg at pH 6.0 and 30 °C and showed antimicrobial activities against Gram-negative bacteria (Vibrio vulnificus, Vibrio parahemolyticus, and Aeromonas hydrophila) and Gram-positive bacteria (Staphylococcus aureus and Bacillus subtilis). In addition, recombinant sp-iLys displayed isopeptidase activity which reached the peak at pH 7.5 and 37 °C with the presence of 0.05 M Na + . In conclusion, this report describes the heterologous expression of recombinant sp-iLys in P. pastoris on a preparative-scale, which possesses lytic activity and isopeptidase activity. This suggests that sp-iLys might play an important role in the innate immunity of S. purpuratus.
Juturu, Veeresh; Wu, Jin Chuan
Pichia pastoris is a well-known platform strain for heterologous protein expression. Over the past five years, different strategies to improve the efficiency of recombinant protein expression by this yeast strain have been developed; these include a patent-free protein expression kit, construction of the P. pastoris CBS7435Ku70 platform strain with its high efficiency in site-specific recombination of plasmid DNA into the genomic DNA, the design of synthetic promoters and their variants by combining different core promoters with multiple putative transcription factors, the generation of mutant GAP promoter variants with various promoter strengths, codon optimization, engineering the α-factor signal sequence by replacing the native glutamic acid at the Kex2 cleavage site with the other 19 natural amino acids and the addition of mammalian signal sequence to the yeast signal sequence, and the co-expression of single chaperones, multiple chaperones or helper proteins that aid in recombinant protein folding. Publically available high-quality genome data from multiple strains of P. pastoris GS115, DSMZ 70382, and CBS7435 and the continuous development of yeast expression kits have successfully promoted the metabolic engineering of this strain to produce carotenoids, xanthophylls, nootkatone, ricinoleic acid, dammarenediol-II, and hyaluronic acid. The cell-surface display of enzymes has obviously increased enzyme stability, and high-level intracellular expression of acyl-CoA and ethanol O-acyltransferase, lipase and d-amino acid oxidase has opened up applications in whole-cell biocatalysis for producing flavor molecules and biodiesel, as well as the deracemization of racemic amino acids. High-level expression of various food-grade enzymes, cellulases, and hemicellulases for applications in the food, feed and biorefinery industries is in its infancy and needs strengthening. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.
Yang, Hongyu; Zhu, Qiang; Zhou, Nandi; Tian, Yaping
Prolyl aminopeptidases are specific exopeptidases that catalyze the hydrolysis of the N-terminus proline residue of peptides and proteins. In the present study, the prolyl aminopeptidase gene (pap) from Aspergillus oryzae JN-412 was optimized through the codon usage of Pichia pastoris. Both the native and optimized pap genes were inserted into the expression vector pPIC9 K and were successfully expressed in P. pastoris. Additionally, the activity of the intracellular enzyme expressed by the recombinant optimized pap gene reached 61.26 U mL(-1), an activity that is 2.1-fold higher than that of the native gene. The recombinant enzyme was purified by one-step elution through Ni-affinity chromatography. The optimal temperature and pH of the purified PAP were 60 °C and 7.5, respectively. Additionally, the recombinant PAP was recovered at a yield greater than 65 % at an extremely broad range of pH values from 6 to 10 after treatment at 50 °C for 6 h. The molecular weight of the recombinant PAP decreased from 50 kDa to 48 kDa after treatment with a deglycosylation enzyme, indicating that the recombinant PAP was completely glycosylated. The glycosylated PAP exhibited high thermo-stability. Half of the activity remained after incubation at 50 °C for 50 h, whereas the remaining activity of PAP expressed in E. coli was only 10 % after incubation at 50 °C for 1 h. PAP could be activated by the appropriate salt concentration and exhibited salt tolerance against NaCl at a concentration up to 5 mol L(-1).
Full Text Available Abstract Background The microorganism Pichia pastoris is a commonly used microbial host for the expression of recombinant proteins in biotechnology and biopharmaceutical industry. To speed up process development, a fast methodology to determine strain characteristic parameters, which are needed to subsequently set up fed batch feeding profiles, is required. Results Here, we show the general applicability of a novel approach to quantify a certain minimal set of bioprocess-relevant parameters, i.e. the adaptation time of the culture to methanol, the specific substrate uptake rate during the adaptation phase and the maximum specific substrate uptake rate, based on fast and easy-to-do batch cultivations with repeated methanol pulses in a batch culture. A detailed analysis of the adaptation of different P. pastoris strains to methanol was conducted and revealed that each strain showed very different characteristics during adaptation, illustrating the need of individual screenings for an optimal parameter definition during this phase. Based on the results obtained in batch cultivations, dynamic feeding profiles based on the specific substrate uptake rate were employed for different P. pastoris strains. In these experiments the maximum specific substrate uptake rate, which had been defined in batch experiments, also represented the upper limit of methanol uptake, underlining the validity of the determined process-relevant parameters and the overall experimental strategy. Conclusion In this study, we show that a fast approach to determine a minimal set of strain characteristic parameters based on easy-to-do batch cultivations with methanol pulses is generally applicable for different P. pastoris strains and that dynamic fed batch strategies can be designed on the specific substrate uptake rate without running the risk of methanol accumulation.
Qvirist, Linnea; Vorontsov, Egor; Veide Vilg, Jenny; Andlid, Thomas
In this work, we present the development and characterization of a strain of Pichia kudriavzevii (TY1322), with highly improved phytate-degrading capacity. The mutant strain TY1322 shows a biomass-specific phytate degradation of 1.26 mmol g -1 h -1 after 8 h of cultivation in a high-phosphate medium, which is about 8 times higher compared with the wild-type strain. Strain TY1322 was able to grow at low pH (pH 2), at high temperature (46°C) and in the presence of ox bile (2% w/v), indicating this strain's ability to survive passage through the gastrointestinal tract. The purified phytase showed two pH optima, at pH 3.5 and 5.5, and one temperature optimum at 55°C. The lower pH optimum of 3.5 matches the reported pH of the pig stomach, meaning that TY1322 and/or its phytase is highly suitable for use in feed production. Furthermore, P. kudriavzevii TY1322 tolerates ethanol up to 6% (v/v) and shows high osmotic stress tolerance. Owing to the phenotypic characteristics and non-genetically modified organisms nature of TY1322, this strain show great potential for future uses in (i) cereal fermentations for increased mineral bioavailability, and (ii) feed production to increase the phosphate bioavailability for monogastric animals to reduce the need for artificial phosphate fortification. © 2016 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.
Armour, Danielle; Warren, Elizabeth; Miller, Jodie
Indigenous teacher assistants (ITAs) are often employed in schools to assist in addressing educational issues relating to Indigenous students. While, this practice has occurred for over 40 years in most Australian states, little has been written about their contribution in assisting Indigenous students to learn. This paper explores the influence…
Jay Maddock; Nicole K. Taniguchi
BACKGROUND: Genetic research has potential benefits for improving health, such as identifying molecular characteristics of a disease, understanding disease prevalence and treatment, and developing treatments tailored to patients based on individual genetic characteristics of their disease. Indigenous people are often targeted for genetic research because genes are easier to study in communities that practice endogamy. Therefore, populations perceived to be more homogenous, such as Indigenous ...
Background Studies throughout North America and Europe have documented adverse perinatal outcomes for racial/ethnic minorities. Nonetheless, the contrast in newborn characteristics between indigenous and non-indigenous populations in Latin America has been poorly characterized. This is due to many challenges, including a lack of vital registration information on ethnicity. The objective of this study was to analyze trends in anthropometric measures at birth in Chilean indigenous (Mapuche) and non-indigenous children over a 5-year period. Methods We examined weight and length at birth using information available through a national data base of all birth records for the years 2000 through 2004 (n = 1,166.513). Newborns were classified ethnically according to the origins of the parents' last names. Result The average birthweight was stable over the 5 year period with variations of less than 20 g in each group, and with mean values trivially higher in indigenous newborns. The proportion weighing less than 2500 g at birth increased modestly from 5.2% to 5.6% in non-indigenous newborns whereas the indigenous births remained constant at 5.2%. In multiple regression analyses, adjusting flexibly for gestational age and maternal characteristics, the occurrence of an indigenous surname added only 14 g to an average infant's birthweight while holding other factors constant. Results for length at birth were similar, and adjusted time trend variation in both outcomes was trivially small after adjustment. Anthropometric indexes at birth in Chile are quite favorable by international standards. Conclusion There is only a trivial degree of ethnic disparity in these values, in contrast to conditions for ethnic minorities in other countries. Moreover, these values remained roughly constant over the 5 years of observation in this study. PMID:20598150
This paper discusses an experience of relying on indigenous knowledge to resolve a communal conflict between two Nigerian local communities. The authors were working in one of the communities when conflict erupted, and had to initiate moves to restore peace and normality. They relied largely on information on the ...
With the advancement of modern scientific methods and technology, most of these indigenous biological resources are being developed into commercial products, largely without benefiting the very communities that have sustainably managed them over many generations. This paper examines current regulatory ...
This research undertook the study of ectoparasites and haemoparasites found on and in the body of indigenous chicken (Gallus domesticus). Six hundred and nineteen ectoparasites were collected from 375 chicken from 28 households in and around Ibadan city between February and November, 1999. Of these, 455 ...
The study was carried out to characterize two indigenous chickens of Nigeria using protein markers; haemolglobin (HB) and carbonic anhydrase (CA). Separation of the two proteins was achieved by cellulose acetate electrophoresis and direct gene counting method was employed to interpret the result. Palentological ...
A study to evaluate four indigenous chicken – namely: Horasi, Kuchi, Naked neck and Frizzled in order to obtain grand-parent and parent stocks was carried out at Tanzania Livestock Research Institute, Mpwapwa district of Dodoma, Tanzania. The perfomance of the ecotypes were compared so as to come out with the best ...
An ethnobotanical survey and germplasm collection of Ricinodendron heudelotii (Bail.) (an indigenous fruit tree) were carried out in six provinces of the humid rainforest zone in southern Cameroon. Fruit samples were collected at 40-50 km intervals along the main road network of the zone, from homegardens, food crop as ...
Construction projects' high uncertainty rates make them unattractive to non-risk takers. Construction companies are therefore necessarily risk takers, albeit, to varying degrees. This study made an inquiry into the risk management (RM) practices of multinational and indigenous construction companies (MCCs and ICCs, ...
A child inherits from his or family those sets of meaning, quali- ties of style, modes of ... of experience and trial-and-error problem solving by groups of people working to .... indigenous capitals of the past and relinquishes all that is de- skilling or ...
Leong, Frederick T. L.; Pearce, Marina
As a result of a relative lack of cross-cultural validity in most current (Western) psychological models, indigenous models of psychology have recently become a popular approach for understanding behaviour in specific cultures. Such models would be valuable to vocational psychology research with culturally diverse populations. Problems facing…
In the 1970s, the Mediterranean mussel Mytilus galloprovincialis invaded the South African coast and spread rapidly to dominate much of the West Coast, indicating either the opportunity to occupy a vacant niche or its superior competitive capability over indigenous species. In Namaqualand on the West Coast it appears to ...
This study investigated the application of indigenous knowledge among livestock farmers in Southern Ijaw Local Government Area of Bayelsa State. A structured questionnaire was administered to one hundred and fifty four respondents in the study area. The data were analyzed using descriptive and inferential statistics.
The study examined indigenous communication, religion and education as determinants of attitudes towards STIs/HIV/AIDS education in Igando Community Lagos State, Nigeria. A sample of 195 people was randomly selected from the population. The study used four hypotheses to test the respondents' attitude to the use of ...
Jan 24, 2011 ... The production of Nigerian indigenous yoghurt (kindirmo) using Lactobacillus bulgaricus and. Streptococcus thermophilus mutants as starter culture was investigated. The results of milk fermentations using L. bulgaricus and S. thermophilus mutant isolates when compared with their wild- type strains ...
Huss, Hans Henrik
The pathogenic bacteria indigenous to the aquatic and general environment are listed. Their distribution in nature, prevalence in seafood and the possibilities for growth of these organisms in various types of products are outlined These data, combined with what is known regarding the epidemiology...
Indigenous people around the world both in developed and developing countries have long been marginalized by governments and /or by other privileged social groups from main stream social, political and economic activities. As a result they suffer indignity because their legitimate human rights are violated by way of ...
The implementation of intercultural health programs, often understood as the integration of indigenous and biomedical models of medicine, is a common challenge in many countries. Currently there is great interest in implementing intercultural health programs in Peru and throughout the Latin American region. This project ...
Full Text Available The claim that in natural resource management (NRM a change from anthropocentric values and ethics to eco-centric ones is necessary to achieve sustainability leads to the search for eco-centric models of relationship with the environment. Indigenous cultures can provide such models; hence, there is the need for multicultural societies to further include their values in NRM. In this article, we investigate the environmental values placed on a freshwater environment of the Wet Tropics by a community of indigenous Australians. We discuss their environmental values as human values, and so as beliefs that guide communities’ understanding of how the natural world should be viewed and treated by humans. This perspective represents a step forward in our understanding of indigenous environmental values, and a way to overcome the paradigm of indigenous values as valued biophysical attributes of the environment or processes happening in landscapes. Our results show that the participant community holds biospheric values. Restoring these values in the NRM of the Wet Tropics could contribute to sustainability and environmental justice in the area.
This three-year research study examined the influence of interactive technologies on the math achievement of Indigenous students in Years 4, 5, 6 and 7 technology-equipped classrooms in a rural elementary school in British Columbia, Canada. Using a mixed-methods approach, the researcher conducted semistructured interviews and collected math…
M.V.C. Falanruw; Francis Ruegorong
On Yap Island, indigenous management of the fallow in shifting agriculture has resulted in the development of site-stable taro patch and tree garden agroforestry systems. These systems are relatively sustainable and supportive of household economies , with some surplus for local market sales. however, a broad range of crops whose harvest is complementary to those...
multifaceted, multi-sectorial and widely a participatory process in which the .... Important colours used to communicate different meanings among Mbano people are .... of two local government areas – Isiala-Mbano and Ehime-Mbano. ..... view. Again, 190 respondents believed that indigenous communication can assist the.
The production of Nigerian indigenous yoghurt (kindirmo) using Lactobacillus bulgaricus and Streptococcus thermophilus mutants as starter culture was investigated. The results of milk fermentations using L. bulgaricus and S. thermophilus mutant isolates when compared with their wildtype strains (control) indicated that the ...
... the people of this country should harness a synergy between indigenous health care systems, scientific research and modern health care methods. This article attempts to address the historical evolution of health care methods in South Africa, its effect on the community as well as challenges facing the health professions.
Full Text Available This article comments on Indigenous Australian art from an intercultural perspective. The painting Bush Tomato Dreaming (1998, by the Anmatyerre artist Lucy Ngwarai Kunoth serves as model case for my argument that art expresses existential social knowledge. In consequence, I will argue that social theory and art theory together provide tools for intercultural understanding and competence.
Studies of cultural tourism and indigenous identity are fraught with questions concerning exploitation, entitlement, ownership and authenticity. Unease with the idea of leveraging a group identity for commercial gain is ever-present. This anthology articulates some of these debates from a multitude
Three indigenous Mozambican cattle breeds, namely the Angone, Landim and Bovino de Tete were characterized using six proteins, 13 autosomal microsatellite loci and one Y-specific microsatellite locus (INRA124). The Mashona breed from Zimbabwe was also studied to elucidate the origin of the Bovino de Tete cattle.
Full Text Available The IDRC-funded project 'Empowering Indigenous Peoples and Knowledge Systems Related to Climate Change and Intellectual Property Rights' is part of the Open and Collaborative Science in Development Network (OCSDNet. The project “examiners processes of open and collaborative science related to indigenous peoples’ knowledge, climate change and intellectual property rights”. Natural Justice, the lead organisation has a strong ethical stance on the agency and control over knowledge being vested with the contributing project participants, communities of the Nama and Griqua peoples of the Western Cape of South Africa. The project focuses on questions of how climate change is affecting these communities, how do they produce and maintain knowledge relating to climate change, how that knowledge is characterised and shared (or not with wider publics, and how legal frameworks promote or hinder the agenda of these indigenous communities and their choices to communicate and collaborate with wider publics. Indigenous Knowledge is an area where ethical issues of informed consent, historical injustice, non-compatible epistemologies and political, legal, and economic issues all collide in ways that challenge western and Anglo-American assumptions about data sharing. The group seeks to strongly model and internally critique their own ethical stance in the process of their research, through for instance, using community contracts and questioning institutional informed consent systems.
Anthias, Penelope Fay
A growing body of literature examines how the rise of “neo-extractivist” states in Latin America is reconfiguring the relationship between resources, nation, territory, and citizenship. However, the implications for indigenous territorial projects remain underexplored. Ethnographic research in th...... challenge resistance narratives and resource-curse theories, revealing how resources act as conduits for deeper postcolonial struggles over territory, sovereignty, and citizenship....
The rapid increase in the number of industries may have increased the levels of trace metals in the soil. Phytoremediation of these polluted soils using indigenous grasses is now considered an alternative method in remediating these polluted soils. The present study investigated and compared the ability of three ...
Manu'atu, Linita; Kepa, Mere
This paper, written from the perspectives of indigenous Maori and Tongan researchers, critiques the Auckland Secondary Schools Principals Association's (ASSPA) perspective that culture disrupts students' schooling. It discusses the relations of schooling to the cultural and political forces inside and outside of school; the relations of indigenous…
Use of FBG sensors for real time health monitoring of various civil engineering structures is well-established in western world since last decade, whereas in the Indian context this technology is still in a nascent stage. In this paper, performance assessment of indigenously developed FBG sensors for the application of health ...
Indilinga: African Journal of Indigenous Knowledge Systems. Journal Home · ABOUT THIS JOURNAL · Advanced Search · Current Issue · Archives · Journal Home > Vol 12, No 2 (2013) >. Log in or Register to get access to full text downloads. Username, Password, Remember me, or Register. DOWNLOAD FULL TEXT ...
Indilinga: African Journal of Indigenous Knowledge Systems. Journal Home · ABOUT THIS JOURNAL · Advanced Search · Current Issue · Archives · Journal Home > Vol 12, No 1 (2013) >. Log in or Register to get access to full text downloads. Username, Password, Remember me, or Register. DOWNLOAD FULL TEXT ...
Rodil, Kasper; Winschiers-Theophilus, Heike; Jensen, Kasper L
The article presents in-situ findings of introducing a tablet prototype, with touch interaction and 3D graphical visualizations, to empower knowledgeable village elders in Namibia to locally re-create a 3D graphical context for previously recorded video clips of indigenous practices and narratives...
*Corresponding author. E-mail: firstname.lastname@example.org. This work is licensed under the Creative Commons Attribution 4.0 International License. ASSESSMENT OF METALS IN ROASTED INDIGENOUS COFFEE VARIETIES OF. ETHIOPIA. Abera Gure1,2, Bhagwan Singh Chandravanshi1* and Taddese Wondimu Godeto1, ...
Jane Felipe Beltrão
Full Text Available Considering the frequent rights violations perpetrated against indigenous peoples, which affect people and territories, compromising their lives and even their right to mourn the dead, it is imperative to understand the care and concerns of the indigenous towards life and death. Thus, we propose to analyze ethnographic narratives about the Apinayé, Ka'apor, Tapirapé, Tembé, Tenetehara, Terena and Asurini, in order to discuss the caring of people, considering the context of funerary rituals. The texts analyzed are able to reveal: (1 the existence (or not of the practice; (2 the specific contexts in which the funeral rites are (or not practiced; and (3 the meanings that the practice gain in ethnically differentiated societies. The narratives of indigenous peoples are included in order to attempt to make the peoples that nowadays find themselves accused by both the media and (reportedly pro-life organizations “be heard”. Therefore, using the classical literature we study the heritage of ritual practices, which besides confering dignity to the dead, indicate that life is the greater good among indigenous peoples.
Full Text Available Abstract The study of inequities in health is a critical component of monitoring government obligations to uphold the rights of Indigenous Peoples. In Aotearoa/New Zealand the indigenous Māori population has a substantially younger age structure than the non-indigenous population making it necessary to account for age differences when comparing population health outcomes. An age-standardised rate is a summary measure of a rate that a population would have if it had a standard age structure. Changing age standards have stimulated interest in the potential impact of population standards on disparities data and consequently on health policy. This paper compares the age structure of the Māori and non-Māori populations with two standard populations commonly used in New Zealand: Segi's world and WHO world populations. The performance of these standards in Māori and non-Māori mortality data was then measured against the use of the Māori population as a standard. It was found that the choice of population standard affects the magnitude of mortality rates, rate ratios and rate differences, the relative ranking of causes of death, and the relative width of confidence intervals. This in turn will affect the monitoring of trends in health outcomes and health policy decision-making. It is concluded that the choice of age standard has political implications and the development and utilisation of an international indigenous population standard should be considered.
Lactic acid bacteria play an important role in milk coagulation and cheese ripening. To select strains showing interesting industrial features, two indigenous lactobacilli (Lactobacillus brevis and Lactobacillus plantarum) were studied for aminopeptidase activity. Cell and cells free extract were tested for leucyl aminopeptidase ...
In this article, I explore the relationship between housing, home and health amongst Indigenous homeless people living in the Canadian North. In particular, I examine the ways in which Indigenous homemaking practices conflict with housing policy, and exacerbate individual pathways to homelessness....... I argue that integral components in northern Indigenous conceptualizations of home and, in turn, health are not only unrecognized in housing policy, but actively discouraged. The potential for homemaking to inform health and housing policy speaks to the relevance of cultural safety not only...... to Indigenous health services, but also to a comprehensive framing of Indigenous health....
Indigenous peoples often have the worst health status in comparison to non-indigenous people in their own nations; urgent action to address the health inequities for indigenous people is required. The role of rehabilitation in addressing health and disability inequities is particularly important due to the health need of indigenous peoples; the unequal distribution of health determinants; and disparities in access to, quality of care through and outcomes following rehabilitation. This article will present a perspective for Māori, the indigenous peoples of New Zealand, on a framework for improving rehabilitation services for Māori and ultimately their health and wellbeing.
Kesler, Dylan C; Walker, Robert S
The headwaters of the Amazon Basin harbor most of the world's last indigenous peoples who have limited contact with encroaching colonists. Knowledge of the geographic distribution of these isolated groups is essential to assist with the development of immediate protections for vulnerable indigenous settlements. We used remote sensing to document the locations of 28 isolated villages within the four Brazilian states of Acre, Amazonas, Roraima, and Rondônia. The sites were confirmed during previous over-flights and by image evidence of thatched-roof houses; they are estimated to host over 1,700 individuals. Locational data were used to train maximum entropy models that identified landscape and anthropogenic features associated with the occurrence of isolated indigenous villages, including elevation, proximity to streams of five different orders, proximity to roads and settlements, proximity to recent deforestation, and vegetation cover type. Isolated villages were identified at mid elevations, within 20 km of the tops of watersheds and at greater distances from existing roads and trails. We further used model results, combined with boundaries of the existing indigenous territory system that is designed to protect indigenous lands, to assess the efficacy of the existing protected area network for isolated peoples. Results indicate that existing indigenous territories encompass all of the villages we identified, and 50% of the areas with high predicted probabilities of isolated village occurrence. Our results are intended to help inform policies that can mitigate against future external threats to isolated peoples.
Dylan C Kesler
Full Text Available The headwaters of the Amazon Basin harbor most of the world's last indigenous peoples who have limited contact with encroaching colonists. Knowledge of the geographic distribution of these isolated groups is essential to assist with the development of immediate protections for vulnerable indigenous settlements. We used remote sensing to document the locations of 28 isolated villages within the four Brazilian states of Acre, Amazonas, Roraima, and Rondônia. The sites were confirmed during previous over-flights and by image evidence of thatched-roof houses; they are estimated to host over 1,700 individuals. Locational data were used to train maximum entropy models that identified landscape and anthropogenic features associated with the occurrence of isolated indigenous villages, including elevation, proximity to streams of five different orders, proximity to roads and settlements, proximity to recent deforestation, and vegetation cover type. Isolated villages were identified at mid elevations, within 20 km of the tops of watersheds and at greater distances from existing roads and trails. We further used model results, combined with boundaries of the existing indigenous territory system that is designed to protect indigenous lands, to assess the efficacy of the existing protected area network for isolated peoples. Results indicate that existing indigenous territories encompass all of the villages we identified, and 50% of the areas with high predicted probabilities of isolated village occurrence. Our results are intended to help inform policies that can mitigate against future external threats to isolated peoples.
Barker, Brittany; Goodman, Ashley; DeBeck, Kora
In Canada, Indigenous youth suicide represents one of several health disparities burdening Indigenous populations, and like many other of these disparities, can be understood as an expression of societal, historical, cultural and familial trauma. As the number of Indigenous youth who take their own lives every year in Canada continues to far exceed national averages, it appears that conventional suicide prevention efforts remain ineffective among this population. A growing body of research argues that conventional interventions, largely rooted in Western individual-level behavioural change frameworks, are culturally discordant with Indigenous paradigms. In response, some Indigenous communities are turning to cultural revitalization as a holistic community-driven response to suicide prevention and treatment. The following commentary explores the emerging evidence base for "culture as treatment" - a novel approach to suicide that emphasizes the significance of interconnectedness in healing, alongside the revitalization of traditional values to reclaim community wellness. In doing so, we seek to contribute to a changing discourse surrounding Indigenous youth suicide by acknowledging culture as strength against this national crisis.
Full Text Available A gene encoding Rhizopus oryzae lipase containing prosequence (ProROL was cloned into the pPICZαA and electrotransformed into the Pichia pastoris X-33 strain. The lipase was functionally expressed and secreted in Pichia pastoris with a molecular weight of 35 kDa. The maximum lipase activity of recombinant lipase (rProROL was 21,000 U/mL, which was obtained in a fed-batch cultivation after 168 h induction with methanol in a 50-L bioreactor. After fermentation, the supernatant was concentrated by ultrafiltration with a 10 kDa cut off membrane and purified with ion exchange chromatography using SP Sepharose Fast Flow chromatography. The optimum pH and temperature of the rProROL were pH 9.0 and 40 °C, respectively. The lipase was stable from pH 4.0 to 9.0 and from 25 to 55 °C. The enzyme activity was enhanced by Ca2+ and inhibited by Hg2+ and Ag+. The lipase showed high activity toward triglyceride-Tripalmitin (C16:0 and triglyceride-Trilaurin (C12:0.
Diego Gil de Los Santos
Full Text Available In a previous paper we showed that the yeast Pichia pastoris X-33 grown in parboiled rice effluent supplemented with glycerol byproduct from the biodiesel industry improved the quality of the effluent. In this paper we show the validation of this yeast (PPE as probiotic for broilers. Its effect on feed efficiency and immunomodulation was compared with the same yeast grown in yeast peptone dextrose medium (PPY, with Saccharomyces boulardii (SBY and with the controls fed unsupplemented feed (CON. One-day-old female chicks were vaccinated against infectious bursal disease (IBD and the titers of anti-IBD antibodies were measured by ELISA. PPE group had the highest mean titres on days 14 and 28 (p<0,05, and at 28 days, 64% of the animals showed seroconvertion. The PPE group also showed the best weight gains at 42 days of age, that, on days 7, 14 and 21 were 19%, 15%, and 8.7% higher, respectively, than the control group. The best feed conversion, 8.2% higher than the control group, was obtained by PPY at 42 days. Histopathological studies did not detect any undesirable effects in the supplemented animals. We concluded that Pichia pastoris X-33 when grown in effluents of the rice parboiling industry supplemented with glycerol byproduct from the biodiesel has probiotic properties for poultry.
Takeuchi, Akiko; Shimizu-Ibuka, Akiko; Nishiyama, Yoshitaka; Mura, Kiyoshi; Okada, Sanae; Tokue, Chiyoko; Arai, Soichi
Among more than 20 yeast strains isolated from the traditional starter "murcha" in Nepal, we characterized a yeast that might be involved in saccharification. This strain, identified as Pichia burtonii, produced an extracellular amylolytic enzyme when cultured in the presence of starch in the medium. Since no amylase secreted by P. burtonii has yet been reported, we purified the enzyme and determined its N-terminal amino acid sequence. Together with the results of a hydrolyzing activity assay toward various substrates, it was found to be an alpha-amylase. The purified enzyme, named Pichia burtonii alpha-amylase (PBA), was a glycoprotein with an apparent molecular mass of 51 kDa. Enzyme activity was optimal at pH 5.0 at 40 degrees C. The enzyme retained 80% of its original activity after incubation under the optimal pH condition at 50 degrees C for 30 min. The activity was inhibited by metal ions such as Cd(2+), Cu(2+), Hg(2+), Al(3+), and Zn(2+).
Chahal, Sabreen; Wei, Peter; Moua, Pachai; Park, Sung Pil James; Kwon, Janet; Patel, Arth; Vu, Anthony T; Catolico, Jason A; Tsai, Yu Fang Tina; Shaheen, Nadia; Chu, Tiffany T; Tam, Vivian; Khan, Zill-E-Huma; Joo, Hyun Henry; Xue, Liang; Lin-Cereghino, Joan; Tsai, Jerry W; Lin-Cereghino, Geoff P
The methylotrophic yeast Pichia pastoris has been used extensively for expressing recombinant proteins because it combines the ease of genetic manipulation, the ability to provide complex posttranslational modifications and the capacity for efficient protein secretion. The most successful and commonly used secretion signal leader in Pichia pastoris has been the alpha mating factor (MATα) prepro secretion signal. However, limitations exist as some proteins cannot be secreted efficiently, leading to strategies to enhance secretion efficiency by modifying the secretion signal leader. Based on a Jpred secondary structure prediction and knob-socket modeling of tertiary structure, numerous deletions and duplications of the MATα prepro leader were engineered to evaluate the correlation between predicted secondary structure and the secretion level of the reporters horseradish peroxidase (HRP) and Candida antarctica lipase B. In addition, circular dichroism analyses were completed for the wild type and several mutant pro-peptides to evaluate actual differences in secondary structure. The results lead to a new model of MATα pro-peptide signal leader, which suggests that the N and C-termini of MATα pro-peptide need to be presented in a specific orientation for proper interaction with the cellular secretion machinery and for efficient protein secretion. Copyright Â© 2016 Elsevier B.V. All rights reserved.
Fiori, Stefano; Scherm, Barbara; Liu, Jia; Farrell, Robert; Mannazzu, Ilaria; Budroni, Marilena; Maserti, Bianca E; Wisniewski, Michael E; Migheli, Quirico
Pichia fermentans (strain DISAABA 726) is an effective biocontrol agent against Monilinia fructicola and Botrytis cinerea when inoculated in artificially wounded apple fruit but is an aggressive pathogen when inoculated on wounded peach fruit, causing severe fruit decay. Pichia fermentans grows as budding yeast on apple tissue and exhibits pseudohyphal growth on peach tissue, suggesting that dimorphism may be associated with pathogenicity. Two complementary suppressive subtractive hybridization (SSH) strategies, that is, rapid subtraction hybridization (RaSH) and PCR-based subtraction, were performed to identify genes differentially expressed by P. fermentans after 24-h growth on apple vs. peach fruit. Gene products that were more highly expressed on peach than on apple tissue, or vice versa, were sequenced and compared with available yeast genome sequence databases. Several of the genes more highly expressed, when P. fermentans was grown on peach, were related to stress response, glycolysis, amino acid metabolism, and alcoholic fermentation but surprisingly not to cell wall degrading enzymes such as pectinases or cellulases. The dual activity of P. fermentans as both a biocontrol agent and a pathogen emphasizes the need for a thorough risk analysis of potential antagonists to avoid unpredictable results that could negatively impact the safe use of postharvest biocontrol strategies. © 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.
Willems, Catherine; Stassijns, Gaetane; Cornelis, Wim; D'Août, Kristiaan
This study investigates biomechanical implications of walking with indigenous "Kolhapuri" footwear compared to barefoot walking among a population of South Indians. Ten healthy adults from South India walked barefoot and indigenously shod at voluntary speed on an artificial substrate. The experiment was repeated outside, on a natural substrate. Data were collected from (1) a heel-mounted 3D-accelerometer recording peak impact at heel contact, (2) an ankle-mounted 3D-goniometer (plantar/dorsiflexion and inversion/eversion), and (3) sEMG electrodes at the m. tibialis anterior and the m. gastrocnemius medialis. Data show that the effect of indigenous footwear on the measured variables, compared to barefoot walking, is relatively small and consistent between substrates (even though subjects walked faster on the natural substrate). Walking barefoot, compared to shod walking yields higher impact accelerations, but the differences are small and only significant for the artificial substrate. The main rotations of the ankle joint are mostly similar between conditions. Only the shod condition shows a faster ankle rotation over the rapid eversion motion on the natural substrate. Maximal dorsiflexion in late stance differs between the footwear conditions on an artificial substrate, with the shod condition involving a less dorsiflexed ankle, and the plantar flexion at toe-off is more extreme when shod. Overall the activity pattern of the external foot muscles is similar. The indigenous footwear studied (Kolhapuri) seems to alter foot biomechanics only in a subtle way. While offering some degree of protection, walking in this type of footwear resembles barefoot gait and this type of indigenous footwear might be considered "minimal". © 2017 The Authors American Journal of Physical Anthropology Published by Wiley Periodicals, Inc.
Li, Haiying; Rasmussen, Morten I; Larsen, Martin R
A mutant Trypanosoma rangeli sialidase, Tr7, expressed in Pichia pastoris, exhibits significant trans-sialidase activity, and has been used for analytical-scale production of sialylated human milk oligosaccharides. Mass spectrometry-based site-specific N-glycoprofiling of Tr7 showed that heteroge...
van Oort, Erica; Lerouge, Patrice; de Heer, Pleuni G.; Séveno, Martial; Coquet, Laurent; Modderman, Piet W.; Faye, Loïc; Aalberse, Rob C.; van Ree, Ronald
Background: Recombinant proteins from Pichia pastoris need to be fully evaluated before used as diagnostic tools. Objective: The objective of this study was to investigate whether glycosylation by P. pastoris interferes with the specificity of diagnostic tests. Methods: An autoantigen involved in
Michalak, Malwina; Thomassen, Lise Vestergaard; Roytio, Henna
was to use potato β-1,4-galactan and the SPPP as substrates for enzymatic production of potentially prebiotic compounds of lower and narrower molecular weight. A novel endo-1,4-β-galactanase from Emericella nidulans (anamorph Aspergillus nidulans), GH family 53, was produced in a recombinant Pichia pastoris...
Klompmaker, Sandra H.; Kilic, Aysun; Baerends, Richard J.; Veenhuis, Marten; van der Klei, Ida J.; Goffeau, André
d-Amino acid oxidase (DAO) is an important flavo-enzyme that catalyzes the oxidative deamination of d-amino acids into the corresponding alpha-keto acid, ammonia and H(2)O(2). We identified two amino acid oxidases in the methylotrophic yeast Pichia pastoris: Dao1p, which preferentially uses
For more than 20 years, Jens Dahl has observed and now analyzed how a relatively independent space, the Indigenous Space, has been constructed within the confines of the United Nations. In the UN, indigenous peoples have achieved more than any other group of people, minorities included. The book...... traces this to the ability of indigenous peoples to create consensus among themselves; the establishment of an indigenous caucus; and the construction of a global indigenousness in a continuously developing process in which contentious relationships and symbols have been constructed, reformulated......, negotiated and re-negotiated internally and with the states. In this process 'indigenous peoples' developed as a category and an evolving concept. Dahl looks into the ability of different indigenous representatives to make an impact on the UN processes and use achievements for purposes at home. Combining...
Luis Fernando ANGOSTO FERRÁNDEZ
Full Text Available This article examines the Venezuelan regional elections of 2008 as a contextual event for the analysis of electoral strategies and results associated with the indigenous representation. Three factors intertwined in the electoral moment are analyzed: 1. the existence of minimum guaranteed representation for indigenous population in legislative organs; 2. the participation of indigenous candidates and electors; 3. the maneuvers of political parties and civil organizations that attempt to channel and/or benefit from such indigenous representation and participation. The description of the electoral context facilitates the identification of factors that, beyond the normative structure of the State, condition the agency of individuals and parties involved in electoral processes. Among those factors are the symbolic value of indigeneity in the current process of national identity re-definition, the interest of political parties in controlling the vote of the indigenous representation and the tendency towards the consolidation of professionalized elites within the indigenous activism.
Melissa J. Stoneham
Full Text Available It is acknowledged that health outcomes for Australian Indigenous peoples are lower than those of non-Indigenous Australians. Research suggests negative media in relation to Indigenous Australians perpetuates racist stereotypes among the wider population and impacts on the health of Indigenous Australians. This study examined the media portrayal of Indigenous Australian public health issues in selected media over a twelve month period and found that, overwhelmingly, the articles were negative in their portrayal of Indigenous health. A total of 74 percent of the coverage of Australian Indigenous related articles were negative, 15 percent were positive, and 11 percent were neutral. The most common negative subject descriptors related to alcohol, child abuse, petrol sniffing, violence, suicide, deaths in custody, and crime.
Lu, Flora; Gray, Clark; Bilsborrow, Richard E; Mena, Carlos F; Erlien, Christine M; Bremner, Jason; Barbieri, Alisson; Walsh, Stephen J
To examine differences in land use and environmental impacts between colonist and indigenous populations in the northern Ecuadorian Amazon, we combined data from household surveys and remotely sensed imagery that was collected from 778 colonist households in 64 colonization sectors, and 499 households from five indigenous groups in 36 communities. Overall, measures of deforestation and forest fragmentation were significantly greater for colonists than indigenous peoples. On average, colonist households had approximately double the area in agriculture and cash crops and 5.5 times the area in pasture as indigenous households. Nevertheless, substantial variation in land-use patterns existed among the five indigenous groups in measures such as cattle ownership and use of hired agricultural labor. These findings support the potential conservation value of indigenous lands while cautioning against uniform policies that homogenize indigenous ethnic groups.
Full Text Available By analyzing the forms and contents of the presentations made by indigenous performers and writers at the I Literary Party of Indigenous Poetics, this article exposes the challenges faced by traditional genre theories in tackling indigenous narratives and analyses how this “crisis” contributes to widening hierarchical and Western biased conceptions. On a stage open to contemporary indigenous expression, as is the literary party, the concepts of performance and storytelling, with the social function of maintaining tradition, continuous learning and transformation, better define this indigenous expression.
Full Text Available Abstract Background It has become evident that host cells react to recombinant protein production with a variety of metabolic and intrinsic stresses such as the unfolded protein response (UPR pathway. Additionally, environmental conditions such as growth temperature may have a strong impact on cell physiology and specific productivity. However, there is little information about the molecular reactions of the host cells on a genomic level, especially in context to recombinant protein secretion. For the first time, we monitored transcriptional regulation of a subset of marker genes in the common production host Pichia pastoris to gain insights into the general physiological status of the cells under protein production conditions, with the main focus on secretion stress related genes. Results Overexpression of the UPR activating transcription factor Hac1p was employed to identify UPR target genes in P. pastoris and the responses were compared to those known for Saccharomyces cerevisiae. Most of the folding/secretion related genes showed similar regulation patterns in both yeasts, whereas genes associated with the general stress response were differentially regulated. Secretion of an antibody Fab fragment led to induction of UPR target genes in P. pastoris, however not to the same magnitude as Hac1p overproduction. Overexpression of S. cerevisiae protein disulfide isomerase (PDI1 enhances Fab secretion rates 1.9 fold, but did not relief UPR stress. Reduction of cultivation temperature from 25°C to 20°C led to a 1.4-fold increase of specific product secretion rate in chemostat cultivations, although the transcriptional levels of the product genes (Fab light and heavy chain were significantly reduced at the lower temperature. A subset of folding related genes appeared to be down-regulated at the reduced temperature, whereas transcription of components of the ER associated degradation and the secretory transport was enhanced. Conclusion Monitoring of
Background The methylotrophic yeast Pichia pastoris has emerged as one of the most promising yeast hosts for the production of heterologous proteins. Mixed feeds of methanol and a multicarbon source instead of methanol as sole carbon source have been shown to improve product productivities and alleviate metabolic burden derived from protein production. Nevertheless, systematic quantitative studies on the relationships between the central metabolism and recombinant protein production in P. pastoris are still rather limited, particularly when growing this yeast on mixed carbon sources, thus hampering future metabolic network engineering strategies for improved protein production. Results The metabolic flux distribution in the central metabolism of P. pastoris growing on a mixed feed of glucose and methanol was analyzed by Metabolic Flux Analysis (MFA) using 13C-NMR-derived constraints. For this purpose, we defined new flux ratios for methanol assimilation pathways in P. pastoris cells growing on glucose:methanol mixtures. By using this experimental approach, the metabolic burden caused by the overexpression and secretion of a Rhizopus oryzae lipase (Rol) in P. pastoris was further analyzed. This protein has been previously shown to trigger the unfolded protein response in P. pastoris. A series of 13C-tracer experiments were performed on aerobic chemostat cultivations with a control and two different Rol producing strains growing at a dilution rate of 0.09 h−1 using a glucose:methanol 80:20 (w/w) mix as carbon source. The MFA performed in this study reveals a significant redistristribution of carbon fluxes in the central carbon metabolism when comparing the two recombinant strains vs the control strain, reflected in increased glycolytic, TCA cycle and NADH regeneration fluxes, as well as higher methanol dissimilation rates. Conclusions Overall, a further 13C-based MFA development to characterise the central metabolism of methylotrophic yeasts when growing on mixed
Βackground The methylotrophic yeast Pichia pastoris has become an important host organism for recombinant protein production and is able to use methanol as a sole carbon source. The methanol utilization pathway describes all the catalytic reactions, which happen during methanol metabolism. Despite the importance of certain key enzymes in this pathway, so far very little is known about possible effects of overexpressing either of these key enzymes on the overall energetic behavior, the productivity and the substrate uptake rate in P. pastoris strains. Results A fast and easy-to-do approach based on batch cultivations with methanol pulses was used to characterize different P. pastoris strains. A strain with MutS phenotype was found to be superior over a strain with Mut+ phenotype in both the volumetric productivity and the efficiency in expressing recombinant horseradish peroxidase C1A. Consequently, either of the enzymes dihydroxyacetone synthase, transketolase or formaldehyde dehydrogenase, which play key roles in the methanol utilization pathway, was co-overexpressed in MutS strains harboring either of the reporter enzymes horseradish peroxidase or Candida antarctica lipase B. Although the co-overexpression of these enzymes did not change the stoichiometric yields of the recombinant MutS strains, significant changes in the specific growth rate, the specific substrate uptake rate and the specific productivity were observed. Co-overexpression of dihydroxyacetone synthase yielded a 2- to 3-fold more efficient conversion of the substrate methanol into product, but also resulted in a reduced volumetric productivity. Co-overexpression of formaldehyde dehydrogenase resulted in a 2-fold more efficient conversion of the substrate into product and at least similar volumetric productivities compared to strains without an engineered methanol utilization pathway, and thus turned out to be a valuable strategy to improve recombinant protein production. Conclusions Co
Gao, Limei; Cai, Menghao; Shen, Wei; Xiao, Siwei; Zhou, Xiangshan; Zhang, Yuanxing
Polyketides are one of the most important classes of secondary metabolites and usually make good drugs. Currently, heterologous production of fungal polyketides for developing a high potential industrial application system with high production capacity and pharmaceutical feasibility was still at its infancy. Pichia pastoris is a highly successful system for the high production of a variety of heterologous proteins. In this work, we aim to develop a P. pastoris based in vivo fungal polyketide production system for first time and evaluate its feasibility for future industrial application. A recombinant P. pastoris GS115-NpgA-ATX with Aspergillus nidulans phosphopantetheinyl transferase (PPtase) gene npgA and Aspergillus terrus 6-methylsalicylic acid (6-MSA) synthase (6-MSAS) gene atX was constructed. A specific compound was isolated and identified as 6-MSA by HPLC, LC-MS and NMR. Transcription of both genes were detected. In 5-L bioreactor, the GS115-NpgA-ATX grew well and produced 6-MSA quickly until reached a high value of 2.2 g/L by methanol induction for 20 hours. Thereafter, the cells turned to death ascribing to high concentration of antimicrobial 6-MSA. The distribution of 6-MSA changed that during early and late induction phase it existed more in supernatant while during intermediate stage it mainly located intracellular. Different from 6-MSA production strain, recombinant M. purpureus pksCT expression strains for citrinin intermediate production, no matter PksCT located in cytoplasm or in peroxisomes, did not produce any specific compound. However, both npgA and pksCT transcripted effectively in cells and western blot analysis proved the expression of PPtase. Then the PPTase was expressed and purified, marked by fluorescent probes, and reacted with purified ACP domain and its mutant ACPm of PksCT. Fluoresence was only observed in ACP but not ACPm, indicating that the PPTase worked well with ACP to make it bioactive holo-ACP. Thus, some other factors may
Raúl Alberto Poutou
Full Text Available El objetivo de este trabajo fue expresar la enzima humana iduronato 2-sulfato sulfatasa en la levadura metilotrófica Pichia pastoris. La deficiencia de esta enzima causa una enfermedad denominada Síndrome de Hunter. Siete clones fueron seleccionados por PCR (10, 28, 92, 94, 144, 149 y 153 al amplificar una banda de 826pb, indicativo de la presencia de la IDSh y de la orientación del fragmento. Diferentes medios de cultivo se emplearon en la fase de crecimiento (YPG, YPGli y BMGliY, la fase de inducción fue llevada a cabo cambiando el medio de crecimiento por BMMY. Una vez crecido en YPGli, los clones 10, 28, 144, 149 y 153 mostraron actividad IDS de 1.53, 2.95, 4.35, 4,07 y 4.15 nmol/h mg a las 24, 48, 72 y 120h de inducción respectivamente. El clon 28 produjo 4.21nmol/h mg al crecer en YPG; sólo los clones 92 y 94 produjeron mejores actividades cuando se crecieron en BMGliY; reportando 1.62 y 1.20nmol/mg h respectivamente. La producción de la IDShr se logró en fermentador de 1l, con medio salino (MBS-sF con el clon IDS28. El crecimiento se realizó en cultivo discontinuo utilizando glicerol (fuente de carbono y energía hasta obtener 12.08g/l de biomasa seca. Para el paso de inducción se utilizó un cultivo alimentado con metanol (<1% (v/v; este último sirvió como fuente de carbono y energía e indujo la expresión de IDShr actuando sobre el promotor nativo AOX1 presente en el constructo pPIC9- IDSh. La actividad específica osciló entre 25.4 y 29.36 nmol/mg h. Se destaca que el valor de referencia de nuestro laboratorio para la IDSh en plasma humano es 12.58 nmol/mg h.
Krainer Florian W
Full Text Available Abstract Βackground The methylotrophic yeast Pichia pastoris has become an important host organism for recombinant protein production and is able to use methanol as a sole carbon source. The methanol utilization pathway describes all the catalytic reactions, which happen during methanol metabolism. Despite the importance of certain key enzymes in this pathway, so far very little is known about possible effects of overexpressing either of these key enzymes on the overall energetic behavior, the productivity and the substrate uptake rate in P. pastoris strains. Results A fast and easy-to-do approach based on batch cultivations with methanol pulses was used to characterize different P. pastoris strains. A strain with MutS phenotype was found to be superior over a strain with Mut+ phenotype in both the volumetric productivity and the efficiency in expressing recombinant horseradish peroxidase C1A. Consequently, either of the enzymes dihydroxyacetone synthase, transketolase or formaldehyde dehydrogenase, which play key roles in the methanol utilization pathway, was co-overexpressed in MutS strains harboring either of the reporter enzymes horseradish peroxidase or Candida antarctica lipase B. Although the co-overexpression of these enzymes did not change the stoichiometric yields of the recombinant MutS strains, significant changes in the specific growth rate, the specific substrate uptake rate and the specific productivity were observed. Co-overexpression of dihydroxyacetone synthase yielded a 2- to 3-fold more efficient conversion of the substrate methanol into product, but also resulted in a reduced volumetric productivity. Co-overexpression of formaldehyde dehydrogenase resulted in a 2-fold more efficient conversion of the substrate into product and at least similar volumetric productivities compared to strains without an engineered methanol utilization pathway, and thus turned out to be a valuable strategy to improve recombinant protein
Kreil David P
Full Text Available Abstract Background DNA Microarrays are regarded as a valuable tool for basic and applied research in microbiology. However, for many industrially important microorganisms the lack of commercially available microarrays still hampers physiological research. Exemplarily, our understanding of protein folding and secretion in the yeast Pichia pastoris is presently widely dependent on conclusions drawn from analogies to Saccharomyces cerevisiae. To close this gap for a yeast species employed for its high capacity to produce heterologous proteins, we developed full genome DNA microarrays for P. pastoris and analyzed the unfolded protein response (UPR in this yeast species, as compared to S. cerevisiae. Results By combining the partially annotated gene list of P. pastoris with de novo gene finding a list of putative open reading frames was generated for which an oligonucleotide probe set was designed using the probe design tool TherMODO (a thermodynamic model-based oligoset design optimizer. To evaluate the performance of the novel array design, microarrays carrying the oligo set were hybridized with samples from treatments with dithiothreitol (DTT or a strain overexpressing the UPR transcription factor HAC1, both compared with a wild type strain in normal medium as untreated control. DTT treatment was compared with literature data for S. cerevisiae, and revealed similarities, but also important differences between the two yeast species. Overexpression of HAC1, the most direct control for UPR genes, resulted in significant new understanding of this important regulatory pathway in P. pastoris, and generally in yeasts. Conclusion The differences observed between P. pastoris and S. cerevisiae underline the importance of DNA microarrays for industrial production strains. P. pastoris reacts to DTT treatment mainly by the regulation of genes related to chemical stimulus, electron transport and respiration, while the overexpression of HAC1 induced many genes
Full Text Available Abstract Background The methylotrophic yeast Pichia pastoris has emerged as one of the most promising yeast hosts for the production of heterologous proteins. Mixed feeds of methanol and a multicarbon source instead of methanol as sole carbon source have been shown to improve product productivities and alleviate metabolic burden derived from protein production. Nevertheless, systematic quantitative studies on the relationships between the central metabolism and recombinant protein production in P. pastoris are still rather limited, particularly when growing this yeast on mixed carbon sources, thus hampering future metabolic network engineering strategies for improved protein production. Results The metabolic flux distribution in the central metabolism of P. pastoris growing on a mixed feed of glucose and methanol was analyzed by Metabolic Flux Analysis (MFA using 13C-NMR-derived constraints. For this purpose, we defined new flux ratios for methanol assimilation pathways in P. pastoris cells growing on glucose:methanol mixtures. By using this experimental approach, the metabolic burden caused by the overexpression and secretion of a Rhizopus oryzae lipase (Rol in P. pastoris was further analyzed. This protein has been previously shown to trigger the unfolded protein response in P. pastoris. A series of 13C-tracer experiments were performed on aerobic chemostat cultivations with a control and two different Rol producing strains growing at a dilution rate of 0.09 h−1 using a glucose:methanol 80:20 (w/w mix as carbon source. The MFA performed in this study reveals a significant redistristribution of carbon fluxes in the central carbon metabolism when comparing the two recombinant strains vs the control strain, reflected in increased glycolytic, TCA cycle and NADH regeneration fluxes, as well as higher methanol dissimilation rates. Conclusions Overall, a further 13C-based MFA development to characterise the central metabolism of methylotrophic
Tollefson, D; Bloss, E; Fanning, A; Redd, J T; Barker, K; McCray, E
The burden of tuberculosis (TB) in the estimated 370 million indigenous peoples worldwide is unknown. To conduct a literature review to summarize the TB burden in indigenous peoples, identify gaps in current knowledge, and provide the foundation for a research agenda prioritizing indigenous health within TB control. A systematic literature review identified articles published between January 1990 and November 2011 quantifying TB disease burden in indigenous populations worldwide. Among the 91 articles from 19 countries included in the review, only 56 were from outside Australia, Canada, New Zealand and the United States. The majority of the studies showed higher TB rates among indigenous groups than non-indigenous groups. Studies from the Amazon generally reported the highest TB prevalence and incidence, but select populations from South-East Asia and Africa were found to have similarly high rates of TB. In North America, the Inuit had the highest reported TB incidence (156/100000), whereas the Metis of Canada and American Indians/Alaska Natives experienced rates of indigenous groups. Where data exist, indigenous peoples were generally found to have higher rates of TB disease than non-indigenous peoples; however, this burden varied greatly. The paucity of published information on TB burden among indigenous peoples highlights the need to implement and improve TB surveillance to better measure and understand global disparities in TB rates.
Full Text Available OBJECTIVE: Assess the epidemiological aspects of tuberculosis in Brazilian indigenous children and actions to control it. METHODS: An epidemiological study was performed with 356 children from 0 to 14 years of age in Rondônia State, Amazon, Brazil, during the period 1997-2006. Cases of TB reported to the Notifiable Diseases Surveillance System were divided into indigenous and non-indigenous categories and analyzed according to sex, age group, place of residence, clinical form, diagnostic tests and treatment outcome. A descriptive analysis of cases and hypothesis test (χ² was carried out to verify if there were differences in the proportions of illness between the groups investigated. RESULTS: A total of 356 TB cases were identified (125 indigenous, 231 non-indigenous of which 51.4% of the cases were in males. In the indigenous group, 60.8% of the cases presented in children aged 0-4 years old. The incidence mean was much higher among indigenous; in 2001, 1,047.9 cases/100,000 inhabitants were reported in children aged < 5 years. Pulmonary TB was reported in more than 80% of the cases, and in both groups over 70% of the cases were cured. Cultures and histopathological exams were performed on only 10% of the patients. There were 3 cases of TB/HIV co-infection in the non-indigenous group and none in the indigenous group. The case detection rate was classified as insufficient or fair in more than 80% of the indigenous population notifications, revealing that most of the diagnoses were performed based on chest x-ray. CONCLUSIONS: The approach used in this study proved useful in demonstrating inequalities in health between indigenous and non-indigenous populations and was superior to the conventional analyses performed by the surveillance services, drawing attention to the need to improve childhood TB diagnosis among the indigenous population.
Ignacio César Cruz Islas
Full Text Available Mexican migration to other countries, primarily United States, is a phenomenon that has been studied from different approaches. It is an important flow of people who, for decades, has left Mexico in search of employment opportunities and higher income. This is due to the weakness of opportunities structure present in Mexico, predominantly in rural areas, as well as budget constraints that prevent households to improve their living conditions. Remittances from other countries, in turn, are an alternative for families to address the lack of employment opportunities and income in their homeland, as well as life-deficit conditions. To see how remittances impact on living conditions of indigenous population, in this paper we analyze living conditions of indigenous households.
Caroline Hermínio Maldonado
Full Text Available The works of authors memoir, researched this, depict the lifestyles Guarani and conflicts involving territorial disputes between the eighteenth and mid-twentieth century. They report experiences and seek their opinion about the relationship between indigenous people and migrants, as the authors themselves, who came to southern Ontario then on business and looking for productive land. From these works it is understood as the Indians managed to hide the regional history and justify their judgments about this population without considering the culture and without social organization. The research is to understand the phenomenon of invisibility to which they are subject, not only the Guarani, but the other indigenous peoples today the state of Mato Grosso do Sul.
Full Text Available This study was conducted in the country of Zambia, Southern Africa, to investigate the occurrence of endo-parasites in indigenous Zambian dogs. Faecal samples were collected from 41 indigenous Zambian dogs from different areas of the Mbabala region in the Southern province of Zambia during the “hot wet” season, although at the time that the samples were collected, the country was experiencing a drought. Faecal samples were analysed using the concentration flotation method with zinc sulphate for the determination of the presence of gastrointestinal parasites. The most prevalent parasites were species from the family Ancylostomatidae (65.0 % infection rate which followed by: Isospora canis (9.8 %, Dipylidium caninum (4.8 %, and Toxascaris leonina (2.4 %. There were in addition, two cases of co-infections with the family Ancylostomatidae and D. caninum, as well as the family Ancylostomatidae and I. canis.
Globalisation is often viewed as a threat to cultural and linguistic diversity and therefore is a central concern of educational practices and policy. The present study challenges this common view by demonstrating that local communities can use global means to support and enhance their specific practices and policies. An historical exploration of education policy in Mexico reveals that there has been a continuing struggle by indigenous peoples to maintain locally relevant modes of teaching. Indigenous peoples have increasingly used technology to maintain their languages and local cultural practices. Such accentuation of the local in a global context is exemplified by the people of Chiapas: They live in subsistence-type communities, yet their recent education movements and appeals to international solidarity (such as in the Zapatista rebellion) have employed computer-aided technologies.
Full Text Available Proponents of the discipline of Native Studies (in its various guises have attempted to produce a methodologically and theoretically distinctive body of scholarship to justify its existence in the field of academia. Critiquing Duane Champagne’s recent article published in a flagship journal for North American Native Studies, I argue that while establishing Native Studies as a discipline has little or nothing to do with securing Native Studies departments on university campuses, a place nonetheless exists for these departments. Marrying Native Studies literature on the importance of producing tribally specific knowledge with Australian-based Whiteness Studies literature focusing on the utility of indigeneity for denaturalising white privilege, I argue that the discipline of Native Studies should justify itself departmentally by teaching about the complex forms of local indigeneity upon which white privilege is reproduced.
This paper considers a novel concept for a Martian descent and ascent vehicle, called NIMF (for nuclear rocket using indigenous Martian fuel), the propulsion for which will be provided by a nuclear thermal reactor which will heat an indigenous Martian propellant gas to form a high-thrust rocket exhaust. The performance of each of the candidate Martian propellants, which include CO2, H2O, CH4, N2, CO, and Ar, is assessed, and the methods of propellant acquisition are examined. Attention is also given to the issues of chemical compatibility between candidate propellants and reactor fuel and cladding materials, and the potential of winged Mars supersonic aircraft driven by this type of engine. It is shown that, by utilizing the nuclear landing craft in combination with a hydrogen-fueled nuclear thermal interplanetary vehicle and a heavy lift booster, it is possible to achieve a manned Mars mission in one launch. 6 refs
Full Text Available Most part of indigenous adults in the Guarani communities of Mato Grosso do Sul is bilingual and has one of the indigenous languages, Guarani Kaiowá or Guarani Ñandeva, as their mother tongue and Portuguese as a second language; only a few elderly and young children still who do not go to school speak only the mother tongue. In this paper, we try to verify which impression the speakers have for each of these languages and the importance they attribute to each one of them. Data analysis showed that the mother tongue is closely related to the expression of their traditional culture; in general, the indigenous claim their languages are being transmitted to new generations, and therefore preserved in an appropriate manner in the two communities. The Portuguese is also considered very important by all informants and the main motivation for its teaching/learning is the need to contact with the non-indigenous population. These results may help us understand issues related to the future of these indigenous languages and Portuguese language in the investigated communities.
Limaye, P.K.; Soni, N.L.; Agrawal, R.G.
Water Lubricated Bearings (WLB) are used in various mechanisms of fuel handling systems of PHWRs and AHWR. Availability and random failures of these bearings was a major factor in refuelling operations. Indigenous development of these bearings was taken up and 7 types of antifriction bearings in various sizes (totaling 37 variants) for PHWR, AHWR and Dhruva applications were successfully developed. This paper deals with various aspects of WLB development. (author)
Shajib, Md. Tariqul Islam; Kawser, Mahbuba; Miah, Md. Nuruddin
In line of the development of a food composition database for Bangladesh, 10 minor indigenous fruits were analysed for their nutrient composition comprising ascorbic acid, carotenoids and mineral values. Nutrient data obtained have been compared with published data reported in different literatur...... values of these minor fruits would make awareness among the people for their mass consumption for healthy life and to grow more minor fruit trees from extinction in order to maintain biodiversity....
Hitzman, D.O.; Stepp, A.K.; Dennis, D.M.; Graumann, L.R.
This research program was directed at improving the knowledge of reservoir ecology and developing practical microbial solutions for improving oil production. The goal was to identify indigenous microbial populations which can produce beneficial metabolic products and develop a methodology to stimulate those select microbes with inorganic nutrient amendments to increase oil recovery. This microbial technology has the capability of producing multiple oil-releasing agents.
Hitzman, D.O.; Bailey, S.A.; Stepp, A.K.
This research program was directed at improving the knowledge of reservoir ecology and developing practical microbial solutions for improving oil production. The goal was to identify indigenous microbial populations which can produce beneficial metabolic products and develop a methodology to stimulate those select microbes with inorganic nutrient amendments to increase oil recovery. This microbial technology has the capability of producing multiple oil releasing agents. The potential of the system will be illustrated and demonstrated by the example of biopolymer production on oil recovery.
Winschiers-Theophilus, Heike; Jensen, Kasper Løvborg; Rodil, Kasper
Digital re-presentation of indigenous knowledge remains an absurdity as long as we fail to deconstruct the prevalent design paradigm and techniques continuously re-framing technology within a western epistemology. This paper discusses key challenges in attempts of co-constructing a digital......’s views are brought to light within the design interactions. A new digital reality is created at the periphery of the situated knowledge through continuous negotiations and joint meaning making....
Lee, Huei; Yen, Chiung-Fen; Aikenhead, Glen S.
This preliminary ethnographic investigation focused on how Indigenous traditional wisdom can be incorporated into school science and what students learned as a result. Participants included community elders and knowledge keepers, as well as 4th grade (10-year-old) students, all of Amis ancestry, an Indigenous tribe in Taiwan. The students' non-Indigenous teacher played a central role in developing a science module `Measuring Time' that combined Amis knowledge and Western science knowledge. The study identified two cultural worldview perspectives on time; for example, the place-based cyclical time held by the Amis, and the universal rectilinear time presupposed by scientists. Students' pre-instructional fragmented concepts from both knowledge systems became more informed and refined through their engagement in `Measuring Time'. Students' increased interest and pride in their Amis culture were noted.
Waniamara de Jesus dos Santos
Full Text Available Adopting the theories postulated by Classical Rhetoric and Argumentation Discourse, this paper comes up with a reflection about the identity built by Daniel Munduruku in his memories ‘Meu vô Apolinário: um mergulho no rio da (minha memória’ The indigenous writer assumes the world of classical rhetorical tradition and expose his ideas in order to change preconceived concepts about the visibility of indigenous people by non-indigenous brazilian society. Munduruku shows a new indigenous appearence resulting from the performance between previosly and shown ethos, working both: his ‘fame’ and a new person built in his book and captured by the reader. Adopting gender epidictic, the construction of the indigenous ethos is done in a apparently harmless way, lying hidden the purpose to recreate new values about the world’s indigenous peoples in Brazil.
Hamilton, Marcus J.; Walker, Robert S.; Kesler, Dylan C.
Lowland South America has long been a battle-ground between European colonization and indigenous survival. Initial waves of European colonization brought disease epidemics, slavery, and violence that had catastrophic impacts on indigenous cultures. In this paper we focus on the demography of 238 surviving populations in Brazil. We use longitudinal censuses from all known indigenous Brazilian societies to quantify three demographic metrics: 1) effects of European contact on indigenous populations; 2) empirical estimates of minimum viable population sizes; and 3) estimates of post-contact population growth rates. We use this information to conduct population viability analysis (PVA). Our results show that all surviving populations suffered extensive mortality during, and shortly after, contact. However, most surviving populations exhibit positive growth rates within the first decade post-contact. Our findings paint a positive demographic outlook for these indigenous populations, though long-term survival remains subject to powerful externalities, including politics, economics, and the pervasive illegal exploitation of indigenous lands.
Full Text Available Australia’s civil service has had some success in attracting substantial numbers of Indigenous employees. But significant numbers also regularly exit the bureaucracy. Retaining Indigenous employees is recognised as an ongoing difficulty for government. This research with former and current Indigenous civil servants outlines factors they identify as contributing to decisions to leave the bureaucracy. A key finding involves their general sense of being underutilised and undervalued— that forms of experience and understanding as Indigenous people go largely unrecognised within government, which in turn constrains their potential to meaningfully contribute to improving government relations with Indigenous Australians or to enhancing the effectiveness of the bureaucracy more broadly. Work as an Indigenous civil servant emerges as a space of contestation with the possibilities and limits of statecraft.
Brad M. Farrant
Full Text Available The current study sought to increase our understanding of the factors involved in the early vocabulary development of Australian Indigenous children. Data from the Longitudinal Study of Indigenous Children were available for 573 Indigenous children (291 boys who spoke English (M=37.0 months, SD=5.4 months, at wave 3. Data were also available for 86 children (51 boys who spoke an Indigenous language (M=37.1 months, SD=6.0 months, at wave 3. As hypothesised, higher levels of parent-child book reading and having more children’s books in the home were associated with better English vocabulary development. Oral storytelling in Indigenous language was a significant predictor of the size of children’s Indigenous vocabulary.
Davis, S.H. (The World Bank, Environment Department, Washington DC (United States)); Wali, A. (University of Maryland, College of Behavioral and Social Sciences, Department of Anthropology, College Park, MD (United States))
Indigenous peoples have received much attention as potential resource managers of threatened tropical forest ecosystems. Using data from Latin America, this article argues that fundamental changes need to take place in the legal recognition and demarcation of indigenous territories in order for this potential to be fulfilled. A comparison is made between different national land-tenure models for forest-dwelling indigenous peoples and a new model proposed by Latin American indigenous organizations. This comparison suggests that not only do indigenous peoples need to be provided with some degree of control over their territories and resources, but there needs to be a new type of partnership among indigenous peoples, the scientific community, national governments and international development agencies for the management of tropical forests. 37 refs, 3 tabs
Full Text Available It has previously been shown that being of aboriginal descent is a risk factor for hamstring injuries in Australian football. The aim of this study was to review the Australian Football League (AFL injury database to determine whether there were any injuries where indigenous players had different relative risks to non-indigenous players. Analysis was conducted using data from the AFL injury database, which included data from 4,492 players over 21 years (1992–2012, covering 162,683 player-matches at AFL level, 91,098 matches at lower levels and 328,181 weeks (possible matches of exposure. Compared to non-indigenous players, indigenous players had a significantly higher risk of hamstring injuries (RR 1.52, 95% CI 1.32–1.73 and calf strains (RR 1.30, 95% CI 1.00–1.69. Conversely, indigenous players had a significantly lower risk of lumbar/thoracic spine injuries (RR 0.61, 95% CI 0.41–0.91, groin strains/osteitis pubis (RR 0.75, 95% CI 0.58–0.96 and Achilles tendon injuries (RR 0.32, 95% CI 0.12–0.86. The results for the above injuries were also significant in terms of games missed. There was no difference between overall risk of injury (RR 1.03, 95% CI 0.96–1.10 or missed games (RR 1.00, 95% CI 0.97–1.04. This suggests that indigenous AFL players have the same overall number of injuries and missed games, but a slightly different injury profile.
Wang, Xiahui; Tsang, Yiu Fai; Li, Yuhao; Ma, Xiubing; Cui, Shouqing; Zhang, Tian-Ao; Hu, Jiajun; Gao, Min-Tian
In this study, it was found that the type of phenolic acids derived from rice straw was the major factor affecting ethanol fermentation by Pichia stipitis. The aim of this study was to investigate the inhibitory effect of phenolic acids on ethanol fermentation with rice straw. Different cellulases produced different ratios of free phenolic acids to soluble conjugated phenolic acids, resulting in different fermentation efficiencies. Free phenolic acids exhibited much higher inhibitory effect than conjugated phenolic acids. The flow cytometry results indicated that the damage to cell membranes was the primary mechanism of inhibition of ethanol fermentation by phenolic acids. The removal of free phenolic acids from the hydrolysates increased ethanol productivity by 2.0-fold, indicating that the free phenolic acids would be the major inhibitors formed during saccharification. The integrated process for ethanol and phenolic acids may constitute a new strategy for the production of low-cost ethanol. Copyright © 2017 Elsevier Ltd. All rights reserved.
Nakasaki, Kiyohiko; Araya, Shogo; Mimoto, Hiroshi
In this study, the yeast strain Pichia kudriavzevii RB1 was used as an inoculum to accelerate organic matter degradation of rabbit food with added organic acids, which was used as a model food waste for composting. The RB1 strain rapidly degraded the organic acids present in the raw compost material, leading to an increase in pH beyond the neutral level, within 2 days. Both mesophilic and thermophilic bacteria proliferated faster in the compost with RB1 inoculation than in that without inoculation. Although the yeast died with the increase in compost temperature, it affected the early stages of composting prior to the thermophilic stage and accelerated the composting process by 2 days by eliminating the initial lag phase seen in the growth of other microorganisms. Moreover, populations of Bacillus thermoamylovorans, Bacillus foraminis, and Bacillus coagulans became dominant during the thermophilic stages of both composting with and without RB1 inoculation. Copyright © 2013 Elsevier Ltd. All rights reserved.
Biswas, Rajib; Uellendahl, Hinrich; Ahring, Birgitte Kiær
Sugarcane bagasse is a potential feedstock for cellulosic ethanol production, rich in both glucan and xylan. This stresses the importance of utilizing both C6 and C5 sugars for conversion into ethanol in order to improve the process economics. During processing of the hydrolysate degradation...... products such as acetate, 5-hydroxymethylfurfural (HMF) and furfural are formed, which are known to inhibit microbial growth at higher concentrations. In the current study, conversion of both glucose and xylose sugars into ethanol in wet exploded bagasse hydrolysates was investigated without detoxification...... using Scheffersomyces (Pichia) stipitis CBS6054, a native xylose utilizing yeast strain. The sugar utilization ratio and ethanol yield (Yp/s) ranged from 88-100% and 0.33-0.41 ± 0.02 g/g, respectively, in all the hydrolysates tested. Hydrolysate after wet explosion at 185°C and 6 bar O2, composed...
Ximena Cuadra Montoya
This article analyses the emergence of transnational activism in the context of collective action organised around socio-environmental conflicts in Chile’s indigenous areas. It details the main events in the process of indigenous mobilisation in the form of three emblematic cases carried out on an international scale, together with their implications for the national political arena. The author explains how, after the indigenous people’s demands were blocked at home, they then mobilised abro...
K Kanagasabapathi; V Sakthivel
Indigenous farming is a production system, based on renewal of ecological processes and strengthening of ecological functions of farm ecosystem to produce safe, healthy and sustainable food. Indigenous agriculture is being adopted by the farmers of Kolli Hills in centre Tamil Nadu for different reasons. However, a lot of constraints prevent the farmers in adopting indigenous farming practices, that include poor yield, poor marketing facilities, higher production cost and urbanization
Meeting, June 8, 2015: met with representatives of Honduran Land Management Program (PATH II), indigenous Miskitu leaders from MASTA, and...Granados of Honduran Land Management Program (PATH II), Norvin Goff (President of MASTA indigenous federation), and Darío Cruz (Vice Rector at UPNFM). ...Government, Indigenous Federations, and NGOs Our cartographic research results on the CA Indígena website are used by Honduran government agencies
Maldonado, Julie; Bennett, Bull; Chief, Karletta; Cochran, Patricia; Cozetto, Karen; Gough, Bob; Hiza-Redsteer, Margaret M.; Lynn, Kathy; Maynard, Nancy; Voggesser, Garrit
The organizers of the 2014 US National Climate Assessment (NCA) made a concerted effort to reach out to and collaborate with Indigenous peoples, resulting in the most comprehensive information to date on climate change impacts to Indigenous peoples in a US national assessment. Yet, there is still much room for improvement in assessment processes to ensure adequate recognition of Indigenous perspectives and Indigenous knowledge systems. This article discusses the process used in creating the Indigenous Peoples, Land, and Resources NCA chapter by a team comprised of tribal members, agencies, academics, and non-governmental organizations, who worked together to solicit, collect, and synthesize traditional knowledges and data from a diverse array of Indigenous communities across the US. It also discusses the synergy and discord between traditional knowledge systems and science and the emergence of cross-cutting issues and vulnerabilities for Indigenous peoples. The challenges of coalescing information about climate change and its impacts on Indigenous communities are outlined along with recommendations on the types of information to include in future assessment outputs. We recommend that future assessments – not only NCA, but other relevant local, regional, national, and international efforts aimed at the translation of climate information and assessments into meaningful actions – should support integration of Indigenous perspectives in a sustained way that builds respectful relationships and effectively engages Indigenous communities. Given the large number of tribes in the US and the current challenges and unique vulnerabilities of Indigenous communities, a special report focusing solely on climate change and Indigenous peoples is warranted.This article is part of a special issue on “The National Climate Assessment: Innovations in Science and Engagement” edited by Katharine Jacobs, Susanne Moser, and James Buizer.
Lavoie, Josée G; Kornelsen, Derek; Boyer, Yvonne; Wylie, Lloy
The settlement of the land now known as Canada meant the erasure - sometimes from ignorance, often purposeful - of Indigenous place-names, and understandings of territory and associated obligations. The Canadian map with its three territories and ten provinces, electoral boundaries and districts, reflects boundaries that continue to fragment Indigenous nations and traditional lands. Each fragment adds institutional requirements and organizational complexities that Indigenous nations must engage with when attempting to realize the benefits taken for granted under the Canadian social contract.
Full Text Available Indigenous farming is a production system, based on renewal of ecological processes and strengthening of ecological functions of farm ecosystem to produce safe, healthy and sustainable food. Indigenous agriculture is being adopted by the farmers of Kolli Hills in centre Tamil Nadu for different reasons. However, a lot of constraints prevent the farmers in adopting indigenous farming practices, that include poor yield, poor marketing facilities, higher production cost and urbanization
Klafke, Gabriel Baracy; Moreira, Gustavo Marçal Schmidt Garcia; Pereira, Juliano Lacava; Oliveira, Patrícia Diaz; Conceição, Fabricio Rochedo; Lund, Rafael Guerra; Grassmann, André Alex; Dellagostin, Odir Antonio; da Silva Pinto, Luciano
Lectins are non-immune proteins that reversibly bind to carbohydrates in a specific manner. Bauhinia variegata lectin I (BVL-I) is a Gal/GalNAc-specific, single-chain lectin isolated from Bauhinia variegata seeds that has been implicated in the inhibition of bacterial adhesion and the healing of damaged skin. Since the source of the native protein (nBVL) is limited, this study aimed to produce recombinant BVL-I in Pichia pastoris (rBVL-Ip). The coding sequence for BVL-I containing preferential codons for P. pastoris was cloned into the pPICZαB plasmid. A single expressing clone was selected and fermented, resulting in the secretion and glycosylation of the protein. Fed-batch fermentation in 7L-scale was performed, and the recombinant lectin was purified from culture supernatant, resulting in a yield of 1.5mg/L culture. Further, rBVL-Ip was compared to nBVL and its recombinant version expressed in Escherichia coli BL21 (DE3) (rBVL-Ie). Although it was expressed as a monomer, rBVL-Ip retained its biological activity since it was able to impair the initial adhesion of Streptococcus mutans and S. sanguinis in an in vitro model of biofilm formation and bacterial adhesion. In summary, rBVL-Ip produced in Pichia pastoris represents a viable alternative to large-scale production, encouraging further biological application studies with this lectin. Copyright © 2016 Elsevier B.V. All rights reserved.
Full Text Available This paper focuses upon a place called the Vertical Village, a building in the center of Rio de Janeiro inhabited exclusively by indigenous peoples from different ethnic groups belonging to different parts of the country. In this paper, we discuss questions related to the experience of being indigenous in a city, the construction of a residential space as a village, and the constitution of indigenous identity in the urban context. Following the paths of three inhabitants of the building, the questions considered emerge from their transiting between cities and villages, frontiers either real or imaginary, prejudices and expectations of indigenous identity.
Full Text Available One of the main issues regarding the implementation of REDD+ in Latin America has been the growing concern that such projects may infringe upon the rights and negatively affect the livelihoods of forest-dependent communities. Various indigenous and civil society organizations are ardently opposed to the initiative. Such is the case in Ecuador, where indigenous opposition to REDD+ represents a considerable obstacle in the creation of a national strategy since more than 60% of the country’s remaining forest cover is on indigenous land or under indigenous occupation. Thus one of the most critical challenges remaining for Ecuador will be the construction of a strong legal, financial, and institutional framework—one that the greater indigenous community might be willing to accept. Closer examination of this topic however, reveals just how difficult this may become. Lack of information, a recent political split between national authorities and the indigenous sector, and the dissimilar organizational capacity levels of indigenous communities make the feasibility of carrying out REDD+ projects on these lands extremely complex. However, the biggest obstacle may be ideological. Many indigenous groups view REDD+, with its possible emphasis on international markets and neoliberal mechanisms, as a continuation of the type of policies that have impeded their quest for sovereignty and self determination. As such, indigenous people are only willing to consider such projects if they clearly see preconditions in place that would safeguard their cultures, territories, and autonomy.
Thomas Merlin C
Full Text Available Abstract Background Indigenous Australians have high rates of diabetes and its complications. This study examines ethnic differences in the management of patients with type 2 diabetes in Australian primary care. Methods Diabetes management and outcomes in Indigenous patients enrolled in the NEFRON study (n = 144 was systematically compared with that in non-Indigenous patients presenting consecutively to the same practitioner (n = 449, and the NEFRON cohort as a whole (n = 3893. Results Indigenous Australians with diabetes had high rates of micro- and macrovascular disease. 60% of Indigenous patients had an abnormal albumin to creatinine ratio compared to 33% of non-Indigenous patients (p 1c ≥ 8.0%, observed in 55% of all Indigenous patients, despite the similar frequency use of oral antidiabetic agents and insulin. Smoking was also more common in Indigenous patients (38%vs 10%, p Conclusion Although seeing the same doctors and receiving the same medications, glycaemic and smoking cessation targets remain unfulfilled in Indigenous patients. This cross-sectional study confirms Aboriginal ethnicity as a powerful risk factor for microvascular and macrovascular disease, which practitioners should use to identify candidates for intensive multifactorial intervention.
Full Text Available For many non-Indigenous Australians the only time they have any engagement with Indigenous peoples, history or issues is through watching sport on television or being at a football match at the MCG. This general myopia and indifference by settler Australians with Indigenous Australians manifests itself in many ways but perhaps most obscenely in the simple fact that Indigenous Australians die nearly 20 years younger than the rest of Australias citizens. Many non-Indigenous Australians do not know this. Sport in many ways has offered Indigenous Australians a platform from which to begin the slow, hard process for social justice and equity to be actualised. This paper will discuss the participation of Indigenous Australians in sport and show how sport has enabled Indigenous Australians to create a space so that they can speak out against the injustices they have experienced and to further improve on relations going into the future. The central contention is that through sport all Australians can begin a process of engaging with Indigenous history as a means to improve race relations between the two groups.
Vanessa Alvarenga Caldeira
Full Text Available The objective of this paper is to present a reflection about the processes of environmental licensing of enterprises located in internal or near areas to indigenous lands in Brazil. The analysis is based on work experiences, bibliographic research and subject tracking with the indigenous communities and the government. The article analyzes the environmental licencing process current in Brazil; the common practice of granting environmental licenses without previous studies of the indigenous component and the Brazilian government and enterpreneurs reluctance in hearing the indigenous peoples in these processes.
Full Text Available In November 2014, the lead researcher’s interest in the conceptual development of digital technology and her cultural connection to Indigenous Knowledge Systems created an opportunity to explore a culturally relevant use of technology with urban Indigenous youth: the Indigenous Robotics Prototype Workshop. The workshop achieved a sense of cultural pride and confidence in Indigenous traditional knowledge while inspiring the youth to continue with their engagement in coding and programming through building robots. Yet, the outcomes from the prototype workshop further revealed a need to investigate how Indigenous Knowledge Systems, and particularly Pattern Thinking, might hint toward a possible paradigm shift for the ethical and advanced design of new technologies. This article examines the implications of such a hypothetical shift in autonomous systems in robotics and artificial intelligence (AI, using the Indigenous Robotics Prototype Workshop as a case study and springboard.
This study creates new knowledge regarding the impact of European colonization on Indigenous (Aboriginal, First Nations, Inuit, Metis) families in Canada. It particularly focuses on the issues in families whose children were forcibly removed by the government to institutions called residential schools. Members of Indigenous families voluntarily attended a family therapy practice which utilized a family systems approach and was uniquely based in the Indigenous worldview. This worldview is spir...
Smirnov, Andrew; Kemp, Robert; Ward, James; Henderson, Suzanna; Williams, Sidney; Dev, Abhilash; Najman, Jake M
Despite over-representation of Indigenous Australians in sentinel studies of injecting drug use, little is known about relevant patterns of drug use and dependence. This study compares drug dependence and possible contributing factors in Indigenous and non-Indigenous Australians who inject drugs. Respondent-driven sampling was used in major cities and 'peer recruitment' in regional towns of Queensland to obtain a community sample of Indigenous (n = 282) and non-Indigenous (n = 267) injectors. Data are cross sectional. Multinomial models were developed for each group to examine types of dependence on injected drugs (no dependence, methamphetamine-dependent only, opioid-dependent only, dependent on methamphetamine and opioids). Around one-fifth of Indigenous and non-Indigenous injectors were dependent on both methamphetamine and opioids in the previous 12 months. Psychological distress was associated with dual dependence on these drugs for Indigenous [adjusted relative risk (ARR) 4.86, 95% confidence interval (CI) 2.08-11.34] and non-Indigenous (ARR 4.14, 95% CI 1.59-10.78) participants. Unemployment (ARR 8.98, 95% CI 2.25-35.82) and repeated (> once) incarceration as an adult (ARR 3.78, 95% CI 1.43-9.97) were associated with dual dependence for Indigenous participants only. Indigenous participants had high rates of alcohol dependence, except for those dependent on opioids only. The drug dependence patterns of Indigenous and non-Indigenous people who inject drugs were similar, including the proportions dependent on both methamphetamine and opioids. However, for Indigenous injectors, there was a stronger association between drug dependence and contextual factors such as unemployment and incarceration. Expansion of treatment options and community-level programs may be required. [Smirnov A, Kemp R, Ward J, Henderson S, Williams S, Dev A, Najman J M. Patterns of drug dependence in a Queensland (Australia) sample of Indigenous and non-Indigenous people who
Noer Fauzi Rachman
Full Text Available Under the New Order authoritarian regime, the state endorsed terra-nullification of the customary territories had been the basis for the stipulation of state forest (hutan negara.After the fall of the General Suharto led regime in 1998 generated a new phase for the struggles of the customary groups in different parts of the archipelago. This article examines the rise of indigeneity and counter-hegemonic indigenous legal maneuvering spearheaded by Aliansi Masyarakat Adat Nusantara (AMAN against ongoing land dispossession in Indonesia since the fall of New Order authoritarian regime which includes the indigenous mobilizations (strategy, organization and tactics in the post-authoritarian country, including the avenue of new types of legal activism when it comes to the creative destruction of global capitalism today. It focuses on two modes of policy advocacy and campaign against land dispossession: (a the production of the Constitutional Court Ruling No. 35/PUU-X/2012, a new legal landmark that establishes the constitutional norm of the citizenship status of Indonesian indigenous peoples (masyarakat hukum adat as rights bearing subjects, and the owners of their customary territory; and (b the National Inquiry on Indigenous Peoples’ Rights held by the Indonesian National Human Rights Commission (Komnas HAM. The discussion describes The Colonialism of ‘State-Izing’ Customary Communities’ Territory, Contemporary Indigeneity Politics in Indonesia, Counter-Hegemonic Indigenous Legal Maneuvering, Judicial Review against The1999 Law No. 41on Forestry, National Inquiry on Indigenous Peoples’ Rights, and Connecting Counter-Hegemonic Indigenous Legal Maneuvering with the Grassroots Struggles which focuses on Mobilizing at Multiple Scales. It is concluded from this article that the efficacy of legal struggles is very much depend on the capacity to connect with the grassroots mobilization by continuously promulgating the resurgence of indigeneity
Shah, Mahsood; Widin, Jacquie
Indigenous student satisfaction with the university learning and teaching experience matters. From a student perspective, retention matters as successful completion of tertiary education improves the life chances of students in relation to employment opportunities, being able to support themselves financially and contributing to the society in…
Kennedy, Chona Pineda
This study investigated the alignment of the teaching and learning practices with a student-centred learning approach in an indigenous educational institution. The findings indicated that when a western concept is applied in the classroom, it is vital for it to be culturally relevant and appropriate to the cultural beliefs and values of the…
Pandey, R.; Bandyopadhyay, M.; Parmar, D.; Yadav, R.; Tyagi, H.; Soni, J.; Shishangiya, H.; Sudhir Kumar, D.; Shah, S.; Bansal, G.; Pandya, K.; Parmar, K.; Vuppugalla, M.; Gahlaut, A.; Chakraborty, A.
TWIN source is two RF driver based negative ion source that has been planned to bridge the gap between single driver based ROBIN source (currently operational) and eight river based DNB source (to be operated under IN-TF test facility). TWIN source experiments have been planned at IPR keeping the objective of long term domestic fusion programme to gain operational experiences on vacuum immersed multi driver RF based negative ion source. High vacuum compatible components of twin source are designed at IPR keeping an aim on indigenous built in attempt. These components of TWIN source are mainly stainless steel and OFC-Cu. Being high heat flux receiving components, one of the major functional requirements is continuous heat removal via water as cooling medium. Hence for the purpose stainless steel parts are provided with externally milled cooling lines and that shall be covered with a layer of OFC-cu which would be on the receiving side of high heat flux. Manufacturability of twin source components requires joining of these dissimilar materials via process like electrode position, electron beam welding and vacuum brazing. Any of these manufacturing processes shall give a vacuum tight joint having proper joint strength at operating temperature and pressure. Taking the indigenous development effort vacuum brazing (in non-nuclear environment) has been opted for joining of dissimilar materials of twin source being one of the most reliable joining techniques and commercially feasible across the suppliers of country. Manufacturing design improvisation for the components has been done to suit the vacuum brazing process requirement and to ease some of the machining without comprising over the functional and operational requirements. This paper illustrates the details on the indigenous development effort, design improvisation to suits manufacturability, vacuum brazing basics and its procedures for twin source components.
Chavez Alba, Rafael; Sanchez Arancibia, Oscar Armando [TRANSIERRA S.A., Santa Cruz (Bolivia)
Transierra is a Bolivian company created in the year 2000 with the goal of transporting natural gas from the fields of San Alberto and San Antonio, in Tarija, to the Rio Grande Gas Compression Plant in Santa Cruz, for export to Brazil. Transierra has implemented a Social Action Plan, which allowed it to execute more than 800 community projects for the benefit of over 40 thousand families living in it's area of influence, with the presence of 146 indigenous communities, generally lagging behind in economic and productive life in the region and country. The Support Program to Guarani Development Plans (PA-PDG) is part of the Social Plan and is part of a long-term agreement signed between Transierra and indigenous organizations. The program has implemented more than one hundred projects for productive development, health, education, cultural revaluation, and strengthening organizational infrastructure, generating huge benefits in improving the living conditions of thousands of families of the Guarani people. This year a unique initiative was created with 4 Indigenous Captains and with the support of the International Finance Corporation (World Bank Group), including Business Plans to promote sustainable economic growth, created productive economic cycles involving improvements to the production and productivity to enter the commercial distribution of local and national markets. These four initiatives have meant a shift in the implementation and is helping to generate new dynamics in production, in addition to capturing significant resources from public and private investment, laying the groundwork for the improvement of the incomes and quality of life of its beneficiaries. (author)
Raymond Eder, María L.; Reynoso, Cristina; Lauret, Santiago C.; Rosa, Alberto L.
Grape must harbors a complex community of yeast species responsible for spontaneous alcoholic fermentation. Although there are detailed studies on the microbiota of Vitis vinifera L. grapes, less is known about the diversity and behavior of yeast communities present on fermenting grape must from other species of Vitis. In this work, we used a culture-dependent method to study the identity and dynamics of the indigenous yeast population present during the spontaneous fermentation of Isabella (Vitis labrusca L.) grape must. Alcoholic fermentation was conducted using standard enological practices, and the associated non-Saccharomyces and S. cerevisiae yeast community was analyzed using selective growth media and 5.8-ITS DNA sequencing. Candida californica, Candida hellenica, Starmerella bacillaris (synonym Candida zemplinina), Hanseniaspora uvarum, and Hanseniaspora vineae were the main non-Saccharomyces species identified on Isabella fermenting must. Issatchenkia hanoiensis, a yeast species rarely found on Vitis vinifera L. grapes, was also recognized on Isabella grape must. Candida azymoides, Candida californica and Pichia cecembensis, identified in this work on Isabella fermenting must, have not previously been found on Vitis vinifera L. grape must. Interestingly, C. azymoides, I. hanoiensis and P. cecembensis have recently been isolated from the surface of Vitis labrusca L. grapes from vineyards in the Azores archipelago, suggesting that specific Vitis-yeast species associations are formed independently of geographic origin. We suggest that C. azymoides, C. californica, and P. cecembensis are yeast species preferentially associated with Vitis labrusca L. grapes. Specific biological interactions between grapevines and yeast species may underlie the assembly of differential Vitis-microbial communities. PMID:28424672
Ambastha, K.P.; Chaudhari, Y.V.; Pal, Suvadip; Tikaria, Amit; Pious, Lizy; Dubey, B.P.; Chadda, V.K.
Scanning electron microscope (SEM) is a precision instrument and plays very important role in scientific studies. Bhabha Atomic Research Centre has taken up the job of development of SEM indigenously. Standard and commercially available components like computer, high voltage power supply, detectors etc. shall be procured from market. Focusing and scanning coils, vacuum chamber, specimen stage, control hardware and software etc. shall be developed at BARC with the help of Indian industry. Procurement, design and fabrication of various parts of SEM are in progress. (author)
Hitzman, D.O.; Stepp, A.K.
This research program was directed at improving the knowledge of reservoir ecology and developing practical microbial solutions for improving oil production. The goal was to identify indigenous microbial populations which can produce beneficial metabolic products and develop a methodology to stimulate those select microbes with inorganic nutrient amendments to increase oil recovery. This microbial technology has the capability of producing multiple oil-releasing agents. The potential of the system will be illustrated and demonstrated by the example of biopolymer production on oil recovery.
Debates on the representation of indigenous cultures in museums have come to the fore in the past thirty years. This paper examines the context for the opening of Waipapa Marae at the University of Auckland in 1988. It outlines a history of Māori meeting houses used for teaching and learning in a specifically Māori context in the New Zealand tertiary sector. The challenge for the university curator with a marae as part of the collection is how to interpret it for the 21st century. Facilitatin...
Cloning, production, and functional expression of the bacteriocin enterocin A, produced by Enterococcus faecium T136, by the yeasts Pichia pastoris, Kluyveromyces lactis, Hansenula polymorpha, and Arxula adeninivorans.
Borrero, Juan; Kunze, Gotthard; Jiménez, Juan J; Böer, Erik; Gútiez, Loreto; Herranz, Carmen; Cintas, Luis M; Hernández, Pablo E
The bacteriocin enterocin A (EntA) produced by Enterococcus faecium T136 has been successfully cloned and produced by the yeasts Pichia pastoris X-33EA, Kluyveromyces lactis GG799EA, Hansenula polymorpha KL8-1EA, and Arxula adeninivorans G1212EA. Moreover, P. pastoris X-33EA and K. lactis GG799EA produced EntA in larger amounts and with higher antimicrobial and specific antimicrobial activities than the EntA produced by E. faecium T136.
Drawing on examples mostly from Iñupiaq and Yup’ik sea-ice expertise in coastal Alaska, this contribution examines how local, indigenous knowledge (LIK) can inform and guide geophysical and biological sea-ice research. Part of the relevance of LIK derives from its linkage to sea-ice use and the services coastal communities derive from the ice cover. As a result, indigenous experts keep track of a broad range of sea-ice variables at a particular location. These observations are embedded into a broader worldview that speaks to both long-term variability or change and to the system of values associated with ice use. The contribution examines eight different contexts in which LIK in study site selection and assessment of a sampling campaign in the context of inter annual variability, the identification of rare or inconspicuous phenomena or events, the contribution by indigenous experts to hazard assessment and emergency response, the record of past and present climate embedded in LIK, and the value of holistic sea-ice knowledge in detecting subtle, intertwined patterns of environmental change. The relevance of local, indigenous sea-ice expertise in helping advance adaptation and responses to climate change as well as its potential role in guiding research questions and hypotheses are also examined. The challenges that may have to be overcome in creating an interface for exchange between indigenous experts and seaice researchers are considered. Promising approaches to overcome these challenges include cross-cultural, interdisciplinary education, and the fostering of Communities of Practice.
Full Text Available Indigenous children and young people are overrepresented in the Australian out-of-home care (OOHC system. To date, specific research has not been undertaken on workers' perspectives regarding the Indigenous-specific and non-Indigenous supports and services available to Indigenous young people exiting the system. This exploratory research involved focus group consultations with workers from seven child and family welfare agencies to examine the current support services available to Indigenous young people who are in or will be leaving out-of-home care in the State of Victoria. Findings suggest that Aboriginal Community Controlled Organisations (ACCOs play a positive role in working with non-Indigenous agencies to assist Indigenous care leavers. Participants identified some key strategies to improve outcomes, such as facilitating stronger relationships between Indigenous and non-Indigenous services, and improving the resourcing of ACCOs.
A survey was conducted in Dhemaji district of Assam, India comprising 15 villages and 300 households. Both purposive and random sampling methods were used to evaluate the socio-economic status of the farmers involved in rearing of indigenous chicken, systems of management of indigenous chicken, their ...
Australia is rapidly losing its Indigenous multicultural and multilingual identity. This vast continent has lost 90 per cent of its Indigenous languages and cultures, without adequate documentation, and risks losing the rest by 2050 if action is not taken. There are formal, accredited linguistics courses designed specifically for Indigenous…
This paper reports on indigenous social impact issues in the Kakadu/Alligators Rivers region of Australia. It briefly outlines the social history of the region, reflects on local, national and international attention being given to the impact of regional development on local indigenous (bininj) people, notes how social impact issues are being addressed and suggests some lessons learnt. (author)
With the adoption of the 2030 Agenda for Sustainable Development, the international community committed to address a great number of challenges. Among those emphasised by the SDGs, some are highly relevant for indigenous groups. Education, poverty, access to justice and climate change are only a few of the issues affecting indigenous people's…
conservation rather than exploitation. On this note, Ajibade .... money carelessly at the problems of local small holder agricultural farmers, it is pertinent to ... establishing the basis for the utility of indigenous knowledge in agricultural .... so that indigenous capabilities play only a marginal role in effecting technical change.
One way is for African bioethicists to begin to apply indigenous African philosophy, thought and values to ethical issues. This project is important (i) to restore dignity; (ii) because a bioethics grounded in indigenous ideas is more likely to be accepted by Africans; and (iii) because such ideas can enrich bioethical discourse.
Shay, Marnee; Heck, Deborah
This article will discuss some of the findings from a qualitative research project that explored the connections between alternative education and Indigenous learners. This study investigated how flexi school leaders reported they were supporting Indigenous young people to remain engaged in education. The results of the survey provide demographic…
This article describes the educational situation of indigenous peoples in Latin America, and in particular their scant participation in adult education activities. It analyses the historical, structural and institutional barriers to their greater involvement in adult education. The article proposes to look at indigenous demands on education as a…
Indigenous peoples and other rural or remote populations often bear the social and environmental cost of extractive industries while obtaining little of the wealth they generate. Recent developments including national and international recognition of Indigenous rights, and the growth of "corporate social responsibility" initiatives among…
Huaman, Elizabeth Alva Sumida
This article attempts to contribute to our expanding definitions of Indigenous education within a globalized world. Additionally, the article critiques notions of progress modeled by powerful nation-states due to their histories based on the intended consequences of marginalizing Indigenous populations for the purposes of material gain. Last,…
This article discusses the role of indigenous knowledge in traditional agriculture and its potential in contributing to food security, poverty and hunger eradication, and increased employment in South Africa. It is noted that indigenous knowledge can inform rural agricultural production, storage, processing, marketing, and food
This article explores the concepts advanced from the Australian Learning and Teaching Council (ALTC)-funded project, "Exploring Problem-Based Learning pedagogy as transformative education in Indigenous Australian Studies". As an Indigenous art historian teaching at a mainstream university in Canada, I am constantly reflecting on how to…
Villegas, Malia, Ed.; Neugebauer, Sabina Rak, Ed.; Venegas, Kerry R., Ed.
This book brings together essays that explore Indigenous ways of knowing and that consider how such knowledge can inform educational practices and institutions. Indigenous Knowledge is resiliently local in character and thus poses a distinct contrast to the international, more impersonal system of knowledge prevalent in Western educational…
Winschiers-Theophilus, Heike; Rodil, Kasper; Zaman, Tariq
In this paper we explore the opportunities of mobile technologies in three of our own development endeavors with rural communities, promoting the preservation of indigenous knowledge. We reflect upon and recognize the fact that the representation of indigenous knowledge will be transformed within...
Quijada, Adrian; Cassadore, Edison; Perry, Gaye Bumsted; Geronimo, Ronald; Lund, Kimberley; Miguel, Phillip; Montes-Helu, Mario; Newberry, Teresa; Robertson, Paul; Thornbrugh, Casey
The U.S.-Mexico border region of the Sonoran Desert is home to 30 Native nations in the United States, and about 15 Indigenous communities in Mexico. Imposed on Indigenous peoples' ancestral lands, the border is an artificial line created in 1848, following the war between the U.S. and Mexico. Tohono O'odham Community College (TOCC) seeks to…
Gallegos, Bernardo P.
Indigenous slavery was a critical aspect of New Mexican life and culture during the Spanish, Mexican, and early American (Territorial) periods. Aside from the labor and military support provided by indigenous slaves for the expansion of the province, the genetic contribution to the population growth was enormous. Ramón Gutiérrez (1991) speculates…
Indigenous people are among the most directly affected by climate change. Yet, there is limited understanding of the health dimensions of climate change and opportunities for adaptation among indigenous populations. Researchers have tended to focus on other vulnerable regions or on populations as a whole.
This may well have contributed to the high failure rate amongst mathematics learners as well as the perceived difficulty of mathematics. It is believed that indigenous knowledge, in general, can be used to promote the teaching of mathematics in multicultural classes. We report on an enquiry into the use of the indigenous ...
Biddle, Nicholas; Cameron, Timothy
This report examines two sets of issues, the first being whether Indigenous Australians obtain a lower return on investment in education and training than other Australians. If they do, then this would partly explain why, in general, Indigenous participation in education and training is relatively low. The second issue is whether Indigenous…
Khobondo, J O; Muasya, T K; Miyumo, S
This review gives insights into genetic and feeding regime development for indigenous chicken genetic resources. We highlight and combine confirming evidence of genetic diversity and variability using morphological and molecular techniques. We further discuss previous past and current genetic...... requirement for indigenous chicken and report nutritive contents of various local feedstuffs under various production systems. Various conservation strategies for sustainable utilization are hereby reviewed...
This article is based on research that focussed on the documentation and the validation of indigenous knowledge on the use of the Dwarf Sage plant in Namibia. The purpose of the study was four-fold: (a) to conduct a systematic recording of the body of indigenous knowledge of the Dwarf Sage; (b) to record and document ...
The study assessed the awareness and indigenous coping mechanism employed by women crop farmers to cope with climate change in Kogi State, Nigeria. Respondents' socioeconomic characteristic, level of awareness about climate change, and indigenous coping strategies to climate change as well as activities of ...
Full Text Available A study has been done with indigenous plants to explore their phytochemcial constituents. About 7 indigenous plants collected from Agra–Mathura Region. The collected plants under gone extraction followed by evaporation. The prepared plant extract goes through phytochemical investigation to explore active constituents which are very significant drug development.
27 mars 2008 ... Integrating Indigenous Knowledge in Climate Risk Management in support of Community Based Adaptation. Traditionally, African farmers have used indigenous knowledge to understand weather and climate patterns and make decisions about crop and irrigation cycles. However, increased variability ...
Partington, Gary; Galloway, Ann
Hearing impairment due to conductive hearing loss can have a devastating effect on children's language development, and consequently educational outcomes, especially for Indigenous students, for whom there may be the additional issue of being educated in their second or third language. With appropriate interventions, however, Indigenous students…
ENSHRINING INDIGENOUS KNOWLEDGE AS A PUBLIC GOOD: INDIGENOUS EDUCATION AND THE MAORI SENSE OF PLACE · EMAIL FULL TEXT EMAIL FULL TEXT DOWNLOAD FULL TEXT DOWNLOAD FULL TEXT. Michael E. Jones, Joshua Hunter, 103-111. http://dx.doi.org/10.4314/indilinga.v3i2.26358 ...
Theodore, Reremoana; Tustin, Karen; Kiro, Cynthia; Gollop, Megan; Taumoepeau, Mele; Taylor, Nicola; Chee, Kaa-Sandra; Hunter, Jackie; Poulton, Richie
Maori, the indigenous population of New Zealand, are gaining university qualifications in greater numbers. This article describes the history of Maori university graduates, their current situation and the implications for indigenous futures. Section one provides a brief overview of historical policies and practices that, similar to those used on…
Indigenous foods contain phytochemicals that are linked to protection against the development of diseases such as cancer, diabetes and hypertension. Some of these indigenous foods have been chemically analysed and contain active compounds such as organic sulphur, hypoglycaemic alkaloids, flavonoids, phytosterin ...
The study investigated the relationship between indigenous information channels and rural communication development in Okuku community in Nigeria. It sought to find out the extent to which indigenous information channels correlate to rural community development. The research design is a correlation survey. A total of ...
Watson, Nicole L
In August 2006, the Aboriginal Land Rights (Northern Territory) Amendment Bill 2006 (Cwlth) was passed into law, introducing, among other things, a system of 99-year leases over Indigenous townships. The leasing scheme will diminish the control that traditional owners previously exercised over their lands. This is at odds with research indicating that control over land is a positive influence on Indigenous health.
Galla, Candace Kaleimamoowahinekapu
Within the last two decades, there has been increased interest in how technology supports Indigenous language revitalization and reclamation efforts. This paper considers the effect technology has on Indigenous language learning and teaching, while conceptualizing how language educators, speakers, learners, and technology users holistically…
Sexing has been a challenging task in Nigerian indigenous chickens due to the monomorphism of chicks which makes it impossible to distinguish the male from the female until eight weeks. . Therefore, this study was carried out to determine the sex of Nigerian indigenous chicks using the common auto-sexing methods.
This paper therefore attempts to examine gender issues in five indigenous communities in Port Harcourt. The communities are: Abuloma, Oroazi, Rumuadaolu, Elekahia and Ogbuna-abali. The key questions explored in the study are: how are urban indigenous women been marginalized in the process of urban growth and ...
All the birds were offered the same grower feed and fresh water ad libitum. Indigenous Venda hens supplemented with ascorbic acid produced chicks with improved (P<0.05) feed intake, feed conversion ratio, growth rate and live weight at 7 weeks of age. However, ascorbic acid supplementation to the diets of indigenous ...
Sumida Huaman, Elizabeth; Abeita, Shawn
Reflecting on our experiences within a program of graduate education in Justice Studies, we offer a discussion of how building and maintaining an iterative teacher-learner stance results in strengthening practices of Indigenous education toward social justice. Through this reflection, we discuss the tenets in Indigenous higher education practices…
This article presents insights from an inquiry into renewable energy development by Indigenous communities across Canada. The focus is on Indigenous leadership in developing renewable energy projects that align with traditional ecological philosophies while also providing increased economic and energy security, sovereignty, and educational…
of Nigerian indigenous language implementation and use in the ... Nigeria, it will become obvious that language is central to ... major indigenous languages. .... personality and thought pattern would be given as credit to the ... (13) The purpose of pre-primary education shall be to: .... Ado-Ekiti and sampled the five of them.
English · Français ... Indigenous peoples today are enmeshed in the expanding modern economy, subject to the ... It explores the complex relationships between indigenous peoples' organizations, civil society, and the environment. ... is pleased to announce that the first call for applications for the new Early Career Women.
Indigenous knowledge is often dismissed as 'traditional and outdated', and hence irrelevant to ... What is more, there has been a consistent bias towards modern .... practices and representations that are developed and maintained by peoples with long histories of ..... “Indigenous Technical Knowledge: Analysis, Implications.
Abstract. This study was an attempt to determine empirically the effect of physics in the development of indigenous technology (production of local talking drum and canoe) in Akwa Ibom State. A total of 70 indigenous technologists (40 canoe makers and 30 talking drum makers) from Ikot Abasi, Essien Udim and Ikot. Ekpene ...
Akena Adyanga, Francis
This study examines African Indigenous Science (AIS) in higher education in Uganda. To achieve this, I use anticolonial theory and Indigenous knowledge discursive frameworks to situate the subjugation of Indigenous science from the education system within a colonial historical context. These theories allow for a critical examination of the intersection of power relations rooted in the politics of knowledge production, validation, and dissemination, and how this process has become a systemic and complex method of subjugating one knowledge system over the other. I also employ qualitative and autoethnographic research methodologies. Using a qualitative research method, I interviewed 10 students and 10 professors from two universities in Uganda. My research was guided by the following key questions: What is African Indigenous Science? What methodology would help us to indigenize science education in Uganda? How can we work with Indigenous knowledge and anticolonial theoretical discursive frameworks to understand and challenge the dominance of Eurocentric knowledge in mainstream education? My research findings revealed that AIS can be defined in multiple ways, in other words, there is no universal definition of AIS. However, there were some common elements that my participants talked about such as: (a) knowledge by Indigenous communities developed over a long period of time through a trial and error approach to respond to the social, economic and political challenges of their society. The science practices are generational and synergistic with other disciplines such as history, spirituality, sociology, anthropology, geography, and trade among others, (b) a cumulative practice of the use, interactions with and of biotic and abiotic organism in everyday life for the continued existence of a community in its' totality. The research findings also indicate that Indigenous science is largely lacking from Uganda's education curriculum because of the influence of colonial and
Full Text Available One of the most common Aboriginalist representations of Indigenous Australian people is, as Indigenous female performer Lou Bennett points out, ‘basically a man, out in the desert, black skin, flat nose with a lap-lap on, standing on one leg, resting against a spear’. Her comment raises many issues. In what ways are discourses of Aboriginalism gendered? How does Aboriginalism affect performance and specifically Aboriginal women performers? In exploring these questions, I examine Aboriginalist representations of Aboriginal women performers by white male scholars and the role of women anthropologists in the production of Aboriginalist discourse about Aboriginal women. Drawing on interviews with Indigenous women performers and musical examples of their songs, I explore the impact of Aboriginalism on non-Indigenous expectations of Indigenous Australian women performing in contemporary music contexts, the strategies performers use to work within and against these constructions and my own relationship to Aboriginalism.
Full Text Available One of the most common Aboriginalist representations of Indigenous Australian people is, as Indigenous female performer Lou Bennett points out, ‘basically a man, out in the desert, black skin, flat nose with a lap-lap on, standing on one leg, resting against a spear’. Her comment raises many issues. In what ways are discourses of Aboriginalism gendered? How does Aboriginalism affect performance and specifically Aboriginal women performers? In exploring these questions, I examine Aboriginalist representations of Aboriginal women performers by white male scholars and the role of women anthropologists in the production of Aboriginalist discourse about Aboriginal women. Drawing on interviews with Indigenous women performers and musical examples of their songs, I explore the impact of Aboriginalism on non-Indigenous expectations of Indigenous Australian women performing in contemporary music contexts, the strategies performers use to work within and against these constructions and my own relationship to Aboriginalism.
Kathleen E. Absolon
Full Text Available This paper begins with a poem and is inclusive of my voice as Anishinaabekwe (Ojibway woman and is authored from my spirit, heart, mind and body. The idea of social inclusion and Indigenous peoples leave more to the imagination and vision than what is the reality and actuality in Canada. This article begins with my location followed with skepticism and hope. Skepticism deals with the exclusion of Indigenous peoples since colonial contact and the subsequent challenges and impacts. Hope begins to affirm the possibilities, strengths and Indigenous knowledge that guides wholistic cultural frameworks and ethics of social inclusion. A wholistic cultural framework is presented; guided by seven sacred teachings and from each element thoughts for consideration are guided by Indigenous values and principles. From each element this paper presents a wholistic and ethical perspective in approaching social inclusion and Indigenous peoples.
Nakata, Martin; Hamacher, Duane W.; Warren, John; Byrne, Alex; Pagnucco, Maurice; Harley, Ross; Venugopal, Srikumar; Thorpe, Kirsten; Neville, Richard; Bolt, Reuben
Indigenous Knowledge is important for Indigenous communities across the globe and for the advancement of our general scientific knowledge. In particular, Indigenous astronomical knowledge integrates many aspects of Indigenous Knowledge, including seasonal calendars, navigation, food economics, law, ceremony, and social structure. Capturing, managing, and disseminating this knowledge in the digital environment poses a number of challenges, which we aim to address using a collaborative project emerging between experts in the higher education, library, archive and industry sectors. Using Microsoft's WorldWide Telescope and Rich Interactive Narratives technologies, we propose to develop software, media design, and archival management solutions to allow Indigenous communities to share their astronomical knowledge with the world on their terms and in a culturally sensitive manner.
Simonds, Vanessa W; Christopher, Suzanne
Indigenous communities have long experienced exploitation by researchers and increasingly require participatory and decolonizing research processes. We present a case study of an intervention research project to exemplify a clash between Western research methodologies and Indigenous methodologies and how we attempted reconciliation. We then provide implications for future research based on lessons learned from Native American community partners who voiced concern over methods of Western deductive qualitative analysis. Decolonizing research requires constant reflective attention and action, and there is an absence of published guidance for this process. Continued exploration is needed for implementing Indigenous methods alone or in conjunction with appropriate Western methods when conducting research in Indigenous communities. Currently, examples of Indigenous methods and theories are not widely available in academic texts or published articles, and are often not perceived as valid.
McWhirter, Rebekah E; Mununggirritj, Djapirri; Marika, Dipililnga; Dickinson, Joanne L; Condon, John R
Indigenous populations, in common with all populations, stand to benefit from the potential of genetic research to lead to improvements in diagnostic and therapeutic tools for a wide range of complex diseases. However, many Indigenous communities, especially ones that are isolated, are not included in genetic research efforts. This situation is largely a consequence of the challenges of ethically conducting genetic research in Indigenous communities and compounded by Indigenous peoples' negative past experiences with genetic issues. To examine ways of addressing these challenges, we review one investigation of a cancer cluster in remote Aboriginal communities in Arnhem Land, Australia. Our experiences demonstrate that genetic research can be both ethically and successfully conducted with Indigenous communities by respecting the authority of the community, involving community members, and including regular community review throughout the research process. Copyright © 2012 Elsevier Ltd. All rights reserved.
Robert S Walker
Full Text Available At least 50 indigenous groups spread across lowland South America remain isolated and have only intermittent and mostly hostile interactions with the outside world. Except in emergency situations, the current policy of governments in Brazil, Colombia, and Peru towards isolated tribes is a "leave them alone" strategy, in which isolated groups are left uncontacted. However, these no-contact policies are based on the assumption that isolated populations are healthy and capable of persisting in the face of mounting external threats, and that they can maintain population viability in the long-term. Here, we test this assumption by tracking the sizes and movements of cleared horticultural areas made by 8 isolated groups over the last 10-14 years. We used deforestation data derived from remote sensing Landsat satellite sensors to identify clearings, and those were then validated and assessed with high-resolution imagery. We found only a single example of a relatively large and growing population (c. 50 cleared ha and 400 people, whereas all of the other 7 groups exhibited much smaller villages and gardens with no sizable growth through time. These results indicated that the smaller groups are critically endangered, and it prompts an urgent re-thinking of policies toward isolated populations, including plans for well-organized contacts that may help save lives and rescue isolated indigenous populations from imminent extinction.
Walker, Robert S; Kesler, Dylan C; Hill, Kim R
At least 50 indigenous groups spread across lowland South America remain isolated and have only intermittent and mostly hostile interactions with the outside world. Except in emergency situations, the current policy of governments in Brazil, Colombia, and Peru towards isolated tribes is a "leave them alone" strategy, in which isolated groups are left uncontacted. However, these no-contact policies are based on the assumption that isolated populations are healthy and capable of persisting in the face of mounting external threats, and that they can maintain population viability in the long-term. Here, we test this assumption by tracking the sizes and movements of cleared horticultural areas made by 8 isolated groups over the last 10-14 years. We used deforestation data derived from remote sensing Landsat satellite sensors to identify clearings, and those were then validated and assessed with high-resolution imagery. We found only a single example of a relatively large and growing population (c. 50 cleared ha and 400 people), whereas all of the other 7 groups exhibited much smaller villages and gardens with no sizable growth through time. These results indicated that the smaller groups are critically endangered, and it prompts an urgent re-thinking of policies toward isolated populations, including plans for well-organized contacts that may help save lives and rescue isolated indigenous populations from imminent extinction.
Prue, Robert; Voss, Richard W
This essay frames an invitation to pastoral counselors and pastoral theologians to examine connections and perhaps interactions between themselves and traditional shamanic healers who use ayahuasca in their healing ceremonies. Indigenous people in South America have used ayahuasca for centuries, and the ritual has become common among the mestizo populations in urban areas of the Amazon, particularly as a curing ritual for drug addiction (Dobkin de Rios, 1970; Moir, 1998). Like peyote in the United States (Calabrese, 1997) ayahuasca use amongst the indigenous people of the Amazon is a form of cultural psychiatry. A review of the literature reveals very little commentary or discussion of shamanic practice in Pastoral Counseling (Pastoral Theology). The scant literature identifies an antithetical relationship at best. The current authors wonder about the possibility of to including shamanic practices in the context of pastoral counseling? This essay seeks to provide some basic information about the ritual use of ayahuasca and to offer a rationale for pastoral counselors to engage in a dialogue about its utility.
Gupta, U.C.; Jayaraj, R.N.; Meena, R.; Sastry, V.S.; Radhakrishna, C.; Rao, S.M.; Sinha, K.K.
Nuclear Fuel Complex (NFC), established in early 70's for production of fuel for PHWRs and BWRs in India, has made several improvements in different areas of fuel manufacturing. Starting with wire-wrap type of fuel bundles, NFC had switched over to split spacer type fuel bundle production in mid 80's. On the upstream side slurry extraction was introduced to prepare the pure uranyl nitrate solution directly from the MDU cake. Applying a thin layer of graphite to the inside of the tube was another modification. The Complex has developed cost effective and innovative techniques for these processes, especially for resistance welding of appendages on the fuel elements which has been a unique feature of the Indian PHWR fuel assemblies. Initially, the fuel fabrication plants were set-up with imported process equipment for most of the pelletisation and assembly operations. Gradually with design and development of indigenous equipment both for production and quality control, NFC has demonstrated total self reliance in fuel production by getting these special purpose machines manufactured indigenously. With the expertise gained in different areas of process development and equipment manufacturing, today NFC is in a position to offer know-how and process equipment at very attractive prices. The paper discusses some of the new processes that are developed/introduced in this field and describes different features of a few PLC based automatic equipment developed. Salient features of innovative techniques being adopted in the area Of UO 2 powder production are also briefly indicated. (author)
Suárez-Torres, José; Suárez-López, José Ricardo; López-Paredes, Dolores; Morocho, Hilario; Cachiguango-Cachiguango, Luis Enrique; Dellai, William
The article aims to systematize and disseminate the main contributions of indigenous ancestral wisdom in the agroecological production of food, especially in Latin America. For this purpose, it is necessary to ask whether such knowledge can be accepted by academia research groups and international forums as a valid alternative that could contribute to overcome the world's nutritional problems. Although no new findings are being made, the validity of ancestral knowledge and agroecology is recognized by scientific research, and by international forums organized by agencies of the United Nations. These recommend that governments should implement them in their policies of development, and in the allocation of funds to support these initiatives. Agroecology and ancestral knowledge are being adopted by a growing number of organizations, indigenous peoples and social groups in various parts of the world, as development alternatives that respond to local needs and worldviews. Its productive potential is progressively being recognized at an international level as a model that contributes to improve the condition of people regarding nutritional food.
Aswani, Shankar; Lauer, Matthew
When sudden catastrophic events occur, it becomes critical for coastal communities to detect and respond to environmental transformations because failure to do so may undermine overall ecosystem resilience and threaten people's livelihoods. We therefore asked how capable of detecting rapid ecological change following massive environmental disruptions local, indigenous people are. We assessed the direction and periodicity of experimental learning of people in the Western Solomon Islands after a tsunami in 2007. We compared the results of marine science surveys with local ecological knowledge of the benthos across 3 affected villages and 3 periods before and after the tsunami. We sought to determine how people recognize biophysical changes in the environment before and after catastrophic events such as earthquakes and tsunamis and whether people have the ability to detect ecological changes over short time scales or need longer time scales to recognize changes. Indigenous people were able to detect changes in the benthos over time. Detection levels differed between marine science surveys and local ecological knowledge sources over time, but overall patterns of statistically significant detection of change were evident for various habitats. Our findings have implications for marine conservation, coastal management policies, and disaster-relief efforts because when people are able to detect ecological changes, this, in turn, affects how they exploit and manage their marine resources. © 2014 Society for Conservation Biology.
Rodrigo Casquero Cunha
Full Text Available The cattle tick Rhipicephalus (Boophilus microplus is responsible for great economic losses. It is mainly controlled chemically, with limitations regarding development of resistance to the chemicals. Vaccines may help control this parasite, thereby reducing tick pesticide use. In this light, we performed subcloning of the gene of the protein Bm86-GC, the homologue protein that currently forms the basis of vaccines (GavacTM and TickGardPLUS that have been developed against cattle ticks. The subcloning was done in the pPIC9 expression vector, for transformation in the yeast Pichia pastoris. This protein was characterized by expression of the recombinant Mut+ strain, which expressed greater quantities of protein. The expressed protein (rBm86-CG was recognized in the Western-blot assay using anti-Gavac, anti-TickGard, anti-larval extract and anti-rBm86-CG polyclonal sera. The serum produced in cattle vaccinated with the antigen CG rBm86 presented high antibody titers and recognized the native protein. The rBm86-GC has potential relevance as an immunogen for vaccine formulation against cattle ticks.O carrapato-do-boi Rhipicephalus (Boophilus microplus é responsável por grandes perdas econômicas. Seu controle é principalmente químico e apresenta limitações quanto ao desenvolvimento de resistência aos princípios ativos. As vacinas podem auxiliar no controle deste parasita diminuindo as aplicações de carrapaticidas. Considerando isso, foi realizada a subclonagem do gene da proteína Bm86-CG, proteína homologa a que atualmente é a base das vacinas desenvolvidas (GavacTM e TickGardPLUS contra o carrapato-do-boi, no vetor de expressão pPIC9, para ser transformado em levedura, Pichia pastoris. Esta proteína foi caracterizada pela expressão da cepa recombinante Mut+ que expressou maior quantidade de proteína. A proteína expressa, rBm86-CG, foi reconhecida no ensaio de Western-blot pelos soros policlonais anti-Gavac, anti-TickGard, anti
Woods John A
Full Text Available Abstract Background Cardiovascular diseases contribute substantially to the poor health and reduced life expectancy of Indigenous Australians. Heart failure is a common, disabling, progressive and costly complication of these disorders. The epidemiology of heart failure and the adequacy of relevant health service provision in Indigenous Australians are not well delineated. Methods A systematic search of the electronic databases PubMed, Embase, Web of Science, Cinahl Plus, Informit and Google Scholar was undertaken in April 2012 for peer-reviewed journal articles relevant to the topic of heart failure in Indigenous Australians. Additionally, a website search was done to identify other pertinent publications, particularly government reports. Results There was a paucity of relevant peer-reviewed research, and government reports dominated the results. Ten journal articles, 1 published conference abstract and 10 reports were eligible for inclusion. Indigenous Australians reportedly have higher morbidity and mortality from heart failure than their non-Indigenous counterparts (age-standardised prevalence ratio 1.7; age-standardised hospital separation ratio ≥3; crude per capita hospital expenditure ratio 1.58; age-adjusted mortality ratio >2. Despite the evident disproportionate burden of heart failure in Indigenous Australians, the accuracy of estimation from administrative data is limited by poor indigenous identification, inadequate case ascertainment and exclusion of younger subjects from mortality statistics. A recent journal article specifically documented a high prevalence of heart failure in Central Australian Aboriginal adults (5.3%, noting frequent undiagnosed disease. One study examined barriers to health service provision for Indigenous Australians in the context of heart failure. Conclusions Despite the shortcomings of available published data, it is clear that Indigenous Australians have an excess burden of heart failure. Emerging data
Ehrich, John; Wolgemuth, Jennifer R.; Helmer, Janet; Oteng, Georges; Lea, Tess; Bartlett, Claire; Smith, Heather; Emmett, Sue
As part of an evaluation of a web-based early literacy intervention, ABRACADABRA, a small exploratory study was conducted over one term in three primary schools in the Northern Territory. Of particular concern was the relationship between attendance and the acquisition of early literacy skills of Indigenous and non-Indigenous children. Using the…
Hornberger, Nancy H.
Drawing from long-term ethnographic research in the Andes, this paper examines one Quechua-speaking Indigenous bilingual educator's trajectory as she traversed (and traverses) from rural highland communities of southern Peru through development as teacher, teacher educator, researcher, and advocate for Indigenous identity and language…