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78 result(s) for "Axenic Culture - methods"
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Ochrobactrum sp. MPV1 from a dump of roasted pyrites can be exploited as bacterial catalyst for the biogenesis of selenium and tellurium nanoparticles
Background Bacteria have developed different mechanisms for the transformation of metalloid oxyanions to non-toxic chemical forms. A number of bacterial isolates so far obtained in axenic culture has shown the ability to bioreduce selenite and tellurite to the elemental state in different conditions along with the formation of nanoparticles—both inside and outside the cells—characterized by a variety of morphological features. This reductive process can be considered of major importance for two reasons: firstly, toxic and soluble (i.e. bioavailable) compounds such as selenite and tellurite are converted to a less toxic chemical forms (i.e. zero valent state); secondly, chalcogen nanoparticles have attracted great interest due to their photoelectric and semiconducting properties. In addition, their exploitation as antimicrobial agents is currently becoming an area of intensive research in medical sciences. Results In the present study, the bacterial strain Ochrobactrum sp. MPV1, isolated from a dump of roasted arsenopyrites as residues of a formerly sulfuric acid production near Scarlino (Tuscany, Italy) was analyzed for its capability of efficaciously bioreducing the chalcogen oxyanions selenite (SeO 3 2− ) and tellurite (TeO 3 2− ) to their respective elemental forms (Se 0 and Te 0 ) in aerobic conditions, with generation of Se- and Te-nanoparticles (Se- and TeNPs). The isolate could bioconvert 2 mM SeO 3 2− and 0.5 mM TeO 3 2− to the corresponding Se 0 and Te 0 in 48 and 120 h, respectively. The intracellular accumulation of nanomaterials was demonstrated through electron microscopy. Moreover, several analyses were performed to shed light on the mechanisms involved in SeO 3 2− and TeO 3 2− bioreduction to their elemental states. Results obtained suggested that these oxyanions are bioconverted through two different mechanisms in Ochrobactrum sp. MPV1. Glutathione (GSH) seemed to play a key role in SeO 3 2− bioreduction, while TeO 3 2− bioconversion could be ascribed to the catalytic activity of intracellular NADH-dependent oxidoreductases. The organic coating surrounding biogenic Se- and TeNPs was also characterized through Fourier-transform infrared spectroscopy. This analysis revealed interesting differences among the NPs produced by Ochrobactrum sp. MPV1 and suggested a possible different role of phospholipids and proteins in both biosynthesis and stabilization of such chalcogen-NPs. Conclusions In conclusion, Ochrobactrum sp. MPV1 has demonstrated to be an ideal candidate for the bioconversion of toxic oxyanions such as selenite and tellurite to their respective elemental forms, producing intracellular Se- and TeNPs possibly exploitable in biomedical and industrial applications.
The Effect of pH on Antibiotic Efficacy against Coxiella burnetii in Axenic Media
Coxiella burnetii , the etiologic agent of Q fever, replicates in an intracellular phagolysosome with pH between 4 and 5. The impact of this low pH environment on antimicrobial treatment is not well understood. An in vitro system for testing antibiotic susceptibility of C. burnetii in axenic media was set up to evaluate the impact of pH on C. burnetii growth and survival in the presence and absence of antimicrobial agents. The data show that C. burnetii does not grow in axenic media at pH 6.0 or higher, but the organisms remain viable. At pH of 4.75, 5.25, and 5.75 moxifloxacin, doxycycline, and rifampin are effective at preventing growth of C. burnetii in axenic media, with moxifloxacin and doxycycline being bacteriostatic and rifampin having bactericidal activity. The efficacy of doxycycline and moxifloxacin improved at higher pH, whereas rifampin activity was pH independent. Hydroxychloroquine is thought to inhibit growth of C. burnetii in vivo by raising the pH of typically acidic intracellular compartments. It had no direct bactericidal or bacteriostatic activity on C. burnetii in axenic media, suggesting that raising pH of acidic intracellular compartments is its primary mechanism of action in vivo . The data suggest that doxycycline and hydroxychloroquine are primarily independent bacteriostatic agents.
Establishment of Anthoceros agrestis as a model species for studying the biology of hornworts
Background Plants colonized terrestrial environments approximately 480 million years ago and have contributed significantly to the diversification of life on Earth. Phylogenetic analyses position a subset of charophyte algae as the sister group to land plants, and distinguish two land plant groups that diverged around 450 million years ago – the bryophytes and the vascular plants. Relationships between liverworts, mosses hornworts and vascular plants have proven difficult to resolve, and as such it is not clear which bryophyte lineage is the sister group to all other land plants and which is the sister to vascular plants. The lack of comparative molecular studies in representatives of all three lineages exacerbates this uncertainty. Such comparisons can be made between mosses and liverworts because representative model organisms are well established in these two bryophyte lineages. To date, however, a model hornwort species has not been available. Results Here we report the establishment of Anthoceros agrestis as a model hornwort species for laboratory experiments. Axenic culture conditions for maintenance and vegetative propagation have been determined, and treatments for the induction of sexual reproduction and sporophyte development have been established. In addition, protocols have been developed for the extraction of DNA and RNA that is of a quality suitable for molecular analyses. Analysis of haploid-derived genome sequence data of two A. agrestis isolates revealed single nucleotide polymorphisms at multiple loci, and thus these two strains are suitable starting material for classical genetic and mapping experiments. Conclusions Methods and resources have been developed to enable A. agrestis to be used as a model species for developmental, molecular, genomic, and genetic studies. This advance provides an unprecedented opportunity to investigate the biology of hornworts.
Testing the Purity of Limnospira fusiformis Cultures After Axenicity Treatments
Contaminations are challenging for monocultures, as they impact the culture conditions and thus influence the growth of the target organism and the overall biomass composition. In phycology, axenic cultures comprising a single living species are commonly strived for both basic research and industrial applications, because contaminants reduce significance for analytic purposes and interfere with the safety and quality of commercial products. We aimed to establish axenic cultures of Limnospira fusiformis, known as the food additive “Spirulina”. Axenicity is strived because it ensures that pathogens or harmful microorganisms are absent and that the harvested biomass is consistent in terms of quality and composition. For the axenic treatment, we applied sterile filtration, ultrasonication, pH treatment, repeated centrifugation, and administration of antibiotics. For testing axenicity, we considered the most common verification method plate tests with Lysogeny Broth (LB) medium, which indicated axenicity after treatments were performed. In addition, we included plate tests with Reasoner’s 2A (R2A) agar and modified Zarrouk+ medium, the latter comparable to the biochemical properties of L. fusiformis’ cultivation medium. In contrast to LB plates, the other media, particularly Zarrouk+, indicated bacterial contamination. We conclude that LB-agar plates are inappropriate for contamination screening of extremophiles. Contamination was also verified by cultivation-independent methods like flow cytometry and 16S rRNA genome amplicon sequencing. We detected taxa of the phyla Proteobacteria, Bacteriodota, Firmicutes and to a lesser extent Verrucomicrobiota. Contaminants are robust taxa, as they survived aggressive treatments. Sequencing data suggest that some of them are promising candidates for in-depth studies to commercially exploit them.
COX Inhibitory and Cytotoxic Naphthoketal-Bearing Polyketides from Sparticola junci
Axenic fermentation on solid rice of the saprobic fungus Sparticola junci afforded two new highly oxidized naphthalenoid polyketide derivatives, sparticatechol A (1) and sparticolin H (2) along with sparticolin A (3). The structures of 1 and 2 were elucidated on the basis of their NMR and HR-ESIMS spectroscopic data. Assignment of absolute configurations was performed using electronic circular dichroism (ECD) experiments and Time-Dependent Density Functional Theory (TDDFT) calculations. Compounds 1–3 were evaluated for COX inhibitory, antiproliferative, cytotoxic and antimicrobial activities. Compounds 1 and 2 exhibited strong inhibitory activities against COX-1 and COX-2. Molecular docking analysis of 1 conferred favorable binding against COX-2. Sparticolin H (2) and A (3) showed a moderate antiproliferative effect against myelogenous leukemia K-562 cells and weak cytotoxicity against HeLa and mouse fibroblast cells.
RNA-seq data of Ganoderma boninense at axenic culture condition and under in planta pathogen-oil palm (Elaeis guineensis Jacq.) interaction
Objective Basal stem rot disease causes severe economic losses to oil palm production in South-east Asia and little is known on the pathogenicity of the pathogen, the basidiomyceteous Ganoderma boninense . Our data presented here aims to identify both the house-keeping and pathogenicity genes of G. boninense using Illumina sequencing reads. Description The hemibiotroph G. boninense establishes via root contact during early stage of colonization and subsequently kills the host tissue as the disease progresses. Information on the pathogenicity factors/genes that causes BSR remain poorly understood. In addition, the molecular expressions corresponding to G. boninense growth and pathogenicity are not reported. Here, six transcriptome datasets of G. boninense from two contrasting conditions (three biological replicates per condition) are presented. The first datasets, collected from a 7-day-old axenic condition provide an insight onto genes responsible for sustenance, growth and development of G. boninense while datasets of the infecting G. boninense collected from oil palm- G. boninense pathosystem ( in planta condition) at 1 month post-inoculation offer a comprehensive avenue to understand G. boninense pathogenesis and infection especially in regard to molecular mechanisms and pathways. Raw sequences deposited in Sequence Read Archive (SRA) are available at NCBI SRA portal with PRJNA514399, bioproject ID.
Unified field studies of the algae testbed public-private partnership as the benchmark for algae agronomics
National scale agronomic projections are an important input for assessing potential benefits of algae cultivation on the future of innovative agriculture. The Algae Testbed Public-Private Partnership was established with the goal of investigating open pond algae cultivation across different geographic, climatic, seasonal, and operational conditions while setting the benchmark for quality data collection, analysis, and dissemination. Identical algae cultivation systems and data analysis methodologies were established at testbed sites across the continental United States and Hawaii. Within this framework, the Unified Field Studies were designed for algae cultivation during all 4 seasons across the testbed network. With increasingly diverse algae research and development, and field deployment strategies, the challenges associated with data collection, quality, and dissemination increase dramatically. The dataset presented here is the complete, curated, climatic, cultivation, harvest, and biomass composition data for each season at each site. These data enable others to do in-depth cultivation, harvest, techno-economic, life cycle, resource, and predictive growth modelling analysis, as well as development of crop protection strategies throughout the algae cultivation industry.
Selection of strains for shiitake production in axenic substrate
Shiitake mushroom consumption is increasing in Brazil. In addition to the implementation of new production methods, it is also important to increase productivity, quality and reduce production costs. In this study, six commercial Lentinula edodes strains were characterized for genetic diversity (rep-PCR analysis) and mushroom production (yield, number and weight of individual mushrooms) using different substrates and cultural conditions. All strains showed genetic differences by repetitive element palindromic based-polymerase chain reaction (rep-PCR). The richest substrate resulted in the greatest production under both environmental conditions. Strains LE4 and LE6 produced the majority of their mushrooms earlier than the other strains. The highest number of mushrooms was observed in the LE6 strain while the highest weights of individual mushrooms were observed in the LE4 strain. Controlled environmental conditions resulted in superior production for all strains, except for LE4, which had empirically greater yield in the semi-controlled environmental condition.
Charcoal filter paper improves the viability of cryopreserved filamentous ectomycorrhizal and saprotrophic Basidiomycota and Ascomycota
We assessed viability of 18 strains of filamentous ectomycorrhizal and saprotrophic basid-iomycetes and ascomycetes after cryopreservation with a novel technique based on charcoal filter paper strips (CFS). The results indicate that axenic fungal cultures grown on CFS recovered from freezing within a few days, even though none survived cryopreservation by the conventional straw method. Fungal growth on CFS was more vigorous, with morphological differentiations such as rhizomorphs and an increased amount of aerial mycelia compared to the unamended culture media. Accordingly CFS allows the cryopreservation of a wide range of rare and important ectomycorrhizal and saprotrophic fungi, which hitherto were difficult to revive from liquid nitrogen storage with the conventional and widely applied straw technique.
Axenic Cultures of Nitrosomonas europaea and Nitrobacter winogradskyi in Autotrophic Conditions: a New Protocol for Kinetic Studies
As a part of a natural biological N-cycle, nitrification is one of the steps included in the conception of artificial ecosystems designed for extraterrestrial life support systems (LSS) such as Micro-Ecological Life Support System Alternative (MELiSSA) project, which is the LSS project of the European Space Agency. Nitrification in aerobic environments is carried out by two groups of bacteria in a two-step process. The ammonia-oxidizing bacteria ( Nitrosomonas europaea ) realize the oxidation of ammonia to nitrite, and the nitrite-oxidizing bacteria ( Nitrobacter winogradskyi ), the oxidation of nitrite to nitrate. In both cases, the bacteria achieve these oxidations to obtain an energy and reductant source for their growth and maintenance. Furthermore, both groups also use CO 2 predominantly as their carbon source. They are typically found together in ecosystems, and consequently, nitrite accumulation is rare. Due to the necessity of modeling accurately conversion yields and transformation rates to achieve a complete modeling of MELiSSA, the present study focuses on the experimental determination of nitrogen to biomass conversion yields. Kinetic and mass balance studies for axenic cultures of Nitrosomonas europaea and Nitrobacter winogradskyi in autotrophic conditions are performed. The follow-up of these cultures is done using flow cytometry for assessing biomass concentrations and ionic chromatography for ammonium, nitrite, and nitrate concentrations. A linear correlation is observed between cell count and optical density (OD) measurement (within a 10 % accuracy) validating OD measurements for an on-line estimation of biomass quantity even at very low biomass concentrations. The conversion between cell count and biomass concentration has been determined: 7.1 × 10 12 cells g dry matter (DM) −1 for Nitrobacter and 6.3 × 10 12 cells g DM −1 for Nitrosomonas . Nitrogen substrates and products are assessed redundantly showing excellent agreement for mass balance purposes and conversion yields determination. Although the dominant phenomena are the oxidation of NH 4 + into nitrite (0.95 mol mol N −1 for Nitrosomonas europaea within an accuracy of 3 %) and nitrite into nitrate (0.975 mol mol N −1 for Nitrobacter winogradskyi within an accuracy of 2 %), the Nitrosomonas europaea conversion yield is estimated to be 0.42 g DM mol N −1 , and Nitrobacter winogradskyi conversion yield is estimated to be 0.27 g DM mol N −1 . The growth rates of both strains appear to be dominated by the oxygen transfer into the experimental setups.