Application of a stir-tank bioreactor for perfusion culture and continuous harvest of Glycyrrhiza inflata suspension cells

We developed a perfusion stir-tank bioreactor that allowed continuous cultivation and harvest of plant cells and challenged for Glycyrrhiza inflata cell suspensions. Continuous separation of cell cultures from the medium occurred using a gravitational sedimentation column. We then conducted cell retention studies of this bioreactor by detecting the cell loss in the discarded medium. We determined that complete cell retention had been achieved when the perfusion rate (perfusion rate is the ratio ofmedium feeding volume per day to bioreactor working volume) was lower than 100% d-1 and the cell suspension recirculation speed was more than 0.5 ml s-1. Growth kinetic measurements showed that the maximum cell concentration reached 25 ± 0.5 g l-1 DW (dry weight). The maximum growth rate occurred on Day 18. Under the given perfusion medium feed rate, the bioreactor operated smoothly, maintaining a relatively stable concentration of 20 g l-1 through continuous cell harvest. The bioreactorwe used could be an efficient cell culture system and demonstrates industrial potential

A Minimal Threshold of c-di-GMP Is Essential for Fruiting Body Formation and Sporulation in Myxococcus xanthus

Generally, the second messenger bis-(3’-5’)-cyclic dimeric GMP (c-di-GMP) regulates the switch between motile and sessile lifestyles in bacteria. Here, we show that c-di-GMP is an essential regulator of multicellular development in the social bacterium Myxococcus xanthus. In response to starvation, M. xanthus initiates a developmental program that culminates in formation of spore-filled fruiting bodies. We show that c-di-GMP accumulates at elevated levels during development and that this increase is essential for completion of development whereas excess c-di-GMP does not interfere with development. MXAN3735 (renamed DmxB) is identified as a diguanylate cyclase that only functions during development and is responsible for this increased c-di-GMP accumulation. DmxB synthesis is induced in response to starvation, thereby restricting DmxB activity to development. DmxB is essential for development and functions downstream of the Dif chemosensory system to stimulate exopolysaccharide accumulation by inducing transcription of a subset of the genes encoding proteins involved in exopolysaccharide synthesis. The developmental defects in the dmxB mutant are non-cell autonomous and rescued by co-development with a strain proficient in exopolysaccharide synthesis, suggesting reduced exopolysaccharide accumulation as the causative defect in this mutant. The NtrC-like transcriptional regulator EpsI/Nla24, which is required for exopolysaccharide accumulation, is identified as a c-diGMP receptor, and thus a putative target for DmxB generated c-di-GMP. Because DmxB can be—at least partially—functionally replaced by a heterologous diguanylate cyclase, these results altogether suggest a model in which a minimum threshold level of c-di-GMP is essential for the successful completion of multicellular development in M. xanthus

Persistence Parameter: a Reliable Measurement for Behavioral Responses of Medaka (Oryzias latipes) to Environmental Stress

Online monitoring systems provided a significant evidence for feasibility of the stepwise behavioral response model in detecting the effects of organophosphorus pesticides on movements of medaka (Oryzias latipes), being able to determine the state of indicator organisms, "no effect," "stimulation," "acclimation," "adjustment (readjustment)," and "toxic effect." Though the stepwise behavioral response model postulated that an organism displays a time-dependent sequence of compensatory stepwise behavioral response during exposure to pollutants above their respective thresholds of resistance, it was still a conceptual model based on tendency only in analysis. In this study, the phenomenon of bacterial persistence was used to interpret the relationship between the stepwise behavioral response model and the environmental stress caused by both exposure time and different treatments. Quantitative measurements of the stepwise behavioral response model led to a simple mathematical description of the threshold switch, which evaluated the effects of environmental stress on behavioral responses to decide the tendency. The adjustment ability correlated to "persisters (p)" is very important for test individuals to overcome the "threshold" from the outside environmental stress. The computational modeling results suggested that "persister (p)," as described in the general equations of bacterial persistence model in changing environments, illustrated behavior acclimation and adjustment (or readjustment) clearly. Consequently, the persistence parameter, p, was critical in addressing for medaka to be adapted to fluctuating environments under different environmental stress

Electrospray deposition and direct patterning of polylactic acid nanofibrous microcapsules for tissue engineering

Electrospun nanofibers composed of biodegradable polymers are attractive candidates for cell culture scaffolds in tissue engineering. Their fine-meshed structures, resembling natural extracellular matrices, effectively interact with cell surfaces and promote cell proliferation. The application of electrospinning, however, is limited to two-dimensional (2D) or single tube-like scaffolds, and the fabrication of arbitrary three-dimensional (3D) scaffolds from electrospun nanofibers is still very difficult due to the fibers’ continuous and entangled form. To address this issue, in this paper, we describe the use of phase-separation-assisted electrospray and electrostatic focusing to perform continuous direct 3D patterning of nanofibrous microcapsules of biodegradable polylactic acid (PLA). These microcapsules exhibit fiber-particle duality because they are composed of nanofibers suitable for cell attachment while also being easy to handle as particles for direct 3D patterning. By varying the flow rate of the polymer solution and the humidity of the electrospray atmosphere during electrospraying, the diameter of the microcapsule and its surface porosity can be controlled. The utility of the direct-patterning process is demonstrated by fabricating high-aspect-ratio microscaffolds and subsequent cell cultures. The nanofibrous and hollow structure of the microcapsules combined with the direct 3D patterning process offers a new approach for fabricating tailor-made scaffolds for regenerative medicine

Medicinal plants used by the Yi ethnic group: a case study in central Yunnan

<p>Abstract</p> <p>Background</p> <p>This paper is based on ethnomedicinal investigation conducted from 1999–2002 in Chuxiong, central Yunnan Province, Southwest China. The Yi medicine has made a great contribution to the ethnomedicinal field in China. Neither case studies nor integrated inventories have previously been conducted to investigate the traditional Yi plants. This paper aims to argue the status and features of medicinal plants used in traditional Yi societies through a case study.</p> <p>Methods</p> <p>The approaches of ethnobotany, anthropology, and participatory rural appraisal were used in the field surveys. Twenty-two informants in four counties were interviewed during eight field trips. Medicinal plant specimens were identified according to taxonomic methods.</p> <p>Results</p> <p>One hundred sixteen medicinal plant species were found to be useful by the local people in the treatment of various diseases or disorders, especially those relating to trauma, gastrointestinal disorders and the common cold. Among these 116 species, 25 species (21.55%) were found to have new curative effects and 40 species (34.48%) were recorded for their new preparation methods; 55 different species were used in treating wounds and fractures, and 47 were used to treat gastrointestinal disorders. Traditional Yi herbal medicines are characterized by their numerous quantities of herbaceous plants and their common preparation with alcohol.</p> <p>Conclusion</p> <p>Totally 116 species in 58 families of medicinal plants traditionally used by the Yi people were inventoried and documented. The characteristics of medicinal plants were analyzed. Some new findings (such as new curative effects and new preparation methods) were recorded These newly gathered ethnobotanical and medicinal data are precious sources for the future development of new drugs, and for further phytochemical, pharmacological and clinical studies.</p

Transgenic Expression of Entire Hepatitis B Virus in Mice Induces Hepatocarcinogenesis Independent of Chronic Liver Injury

Hepatocellular carcinoma (HCC), the third leading cause of cancer deaths worldwide, is most commonly caused by chronic hepatitis B virus (HBV) infection. However, whether HBV plays any direct role in carcinogenesis, other than indirectly causing chronic liver injury by inciting the host immune response, remains unclear. We have established two independent transgenic mouse lines expressing the complete genome of a mutant HBV (“preS2 mutant”) that is found at much higher frequencies in people with HCC than those without. The transgenic mice show evidence of stress in the endoplasmic reticulum (ER) and overexpression of cyclin D1 in hepatocytes. These mice do not show any evidence of chronic liver injury, but by 2 years of age a majority of the male mice develop hepatocellular neoplasms, including HCC. Unexpectedly, we also found a significant increase in hepatocarcinogenesis independent of necroinflammation in a transgenic line expressing the entire wildtype HBV. As in the mutant HBV mice, HCC was found only in aged—2-year-old—mice of the wildtype HBV line. The karyotype in all the three transgenic lines appears normal and none of the integration sites of the HBV transgene in the mice is near an oncogene or tumor suppressor gene. The significant increase of HCC incidence in all the three transgenic lines—expressing either mutant or wildtype HBV—therefore argues strongly that in absence of chronic necroinflammation, HBV can contribute directly to the development of HCC

A stable genetic polymorphism underpinning microbial syntrophy

Syntrophies are metabolic cooperations, whereby two organisms co-metabolize a substrate in an interdependent manner. Many of the observed natural syntrophic interactions are mandatory in the absence of strong electron acceptors, such that one species in the syntrophy has to assume the role of electron sink for the other. While this presents an ecological setting for syntrophy to be beneficial, the potential genetic drivers of syntrophy remain unknown to date. Here, we show that the syntrophic sulfate-reducing species Desulfovibrio vulgaris displays a stable genetic polymorphism, where only a specific genotype is able to engage in syntrophy with the hydrogenotrophic methanogen Methanococcus maripaludis. This 'syntrophic' genotype is characterized by two genetic alterations, one of which is an in-frame deletion in the gene encoding for the ion-translocating subunit cooK of the membrane-bound COO hydrogenase. We show that this genotype presents a specific physiology, in which reshaping of energy conservation in the lactate oxidation pathway enables it to produce sufficient intermediate hydrogen for sustained M. maripaludis growth and thus, syntrophy. To our knowledge, these findings provide for the first time a genetic basis for syntrophy in nature and bring us closer to the rational engineering of syntrophy in synthetic microbial communities

A Bayesian Model for Detection of Highorder Interactions Among Genetic Variants in Genome-Wide Association Studies

Background: A central question for disease studies and crop improvements is how genetics variants drive phenotypes. Genome Wide Association Study (GWAS) provides a powerful tool for characterizing the genotypephenotype relationships in complex traits and diseases. Epistasis (gene-gene interaction), including high-order interaction among more than two genes, often plays important roles in complex traits and diseases, but current GWAS analysis usually just focuses on additive effects of single nucleotide polymorphisms (SNPs). The lack of effective computational modelling of high-order functional interactions often leads to significant under-utilization of GWAS data. Results: We have developed a novel Bayesian computational method with a Markov Chain Monte Carlo (MCMC) search, and implemented the method as a Bayesian High-order Interaction Toolkit (BHIT) for detecting epistatic interactions among SNPs. BHIT first builds a Bayesian model on both continuous data and discrete data, which is capable of detecting high-order interactions in SNPs related to case—control or quantitative phenotypes. We also developed a pipeline that enables users to apply BHIT on different species in different use cases. Conclusions: Using both simulation data and soybean nutritional seed composition studies on oil content and protein content, BHIT effectively detected some high-order interactions associated with phenotypes, and it outperformed a number of other available tools. BHIT is freely available for academic users at http://digbio.missouri.edu/BHIT/

Introgression of Chromosome 3Ns from Psathyrostachys huashanica into Wheat Specifying Resistance to Stripe Rust

Wheat stripe rust is a destructive disease in the cool and humid wheat-growing areas of the world. Finding diverse sources of stripe rust resistance is critical for increasing genetic diversity of resistance for wheat breeding programs. Stripe rust resistance was identified in the alien species Psathyrostachys huashanica, and a wheat- P. huashanica amphiploid line (PHW-SA) with stripe rust resistance was reported previously. In this study, a P. huashanica 3Ns monosomic addition line (PW11) with superior resistance to stripe rust was developed, which was derived from the cross between PHW-SA and wheat J-11. We evaluated the alien introgressions PW11-2, PW11-5 and PW11-8 which were derived from line PW11 for reaction to new Pst race CYR32, and used molecular and cytogenetic tools to characterize these lines. The introgressions were remarkably resistant to CYR32, suggesting that the resistance to stripe rust of the introgressions thus was controlled by gene(s) located on P. huashanica chromosome 3Ns. All derived lines were cytologically stable in term of meiotic chromosome behavior. Two 3Ns chromosomes of P. huashanica were detected in the disomic addition line PW11-2. Chromosomes 1B of substitution line PW11-5 had been replaced by a pair of P. huashanica 3Ns chromosomes. In PW11-8, a small terminal segment from P. huashanica chromosome arm 3NsS was translocated to the terminal region of wheat chromosomes 3BL. Thus, this translocated chromosome is designated T3BL-3NsS. These conclusions were further confirmed by SSR analyses. Two 3Ns-specific markers Xgwm181 and Xgwm161 will be useful to rapidly identify and trace the translocated fragments. These introgressions, which had significant characteristics of resistance to stripe rust, could be utilized as novel germplasms for wheat breeding