Feasibility of composted green waste amended by vermiculite and earthworm casts as the growth media for three common ornamental plants.

This study demonstrated the effects of adding specific proportions of vermiculite (VMT: 0%, 10%, and 20%) and earthworm casts (EWCs: 0%, 10%, and 20%) on the physico-chemical properties of composted green waste (CGW), and the impacts of amended CGW as growth media on the growth of three common ornamental plants (Dahlia pinnata Cav. [dahlia], Centaurea cyanus L. [cornflower], and Consolida ajacis [L.] Schur [delphinium]). Compared with Treatment T1 (CK), the addition of 10% VMT and 20% EWCs greatly (p < 0.05) increased the total porosity, aeration porosity, water-holding porosity, total nitrogen, available phosphorus, available potassium, and organic matter of CGW by 9%, 35%, 4%, 18%, 27%, 13%, and 33%, respectively. In addition, this pattern increased (p < 0.05) the total fresh biomass, total chlorophyll content, and root length of dahlias by 9%, 19%, and 27%, respectively; those of cornflowers by 17%, 30%, and 29%, respectively (p < 0.05); and those of delphiniums by 23%, 14%, and 63%, respectively. Therefore, the amended CGW supplemented with 10% VMT and 20% EWCs was an ideal growth medium for the three plants.

Lactococcus cell envelope proteases enable lactococcal growth in minimal growth media supplemented with high molecular weight proteins of plant and animal origin.

Lactic acid bacteria (LAB) have evolved into fastidious microorganisms that require amino acids from environmental sources. Some LAB have cell envelope proteases (CEPs) that drive the proteolysis of high molecular weight proteins like casein in milk. CEP activity is typically studied using casein as the predominant substrate, even though CEPs can hydrolyze other protein sources. Plant protein hydrolysis by LAB has rarely been connected to the activity of specific CEPs. This study aims to show the activity of individual CEPs using LAB growth in a minimal growth medium supplemented with high molecular weight casein or potato proteins. Using Lactococcus cremoris MG1363 as isogenic background to express CEPs, we demonstrate that CEP activity is directly related to growth in the protein-supplemented minimal growth media. Proteolysis is analyzed based on the amino acid release, allowing a comparison of CEP activities and analysis of amino acid utilization by L. cremoris MG1363. This approach provides a basis to analyze CEP activity on plant-based protein substrates as casein alternatives and to compare activity of CEP homologs.

Cultivation of monoxenous trypanosomatids: A minireview.

Trypanosomatids are obligatory parasites, some of which are responsible for important human and animal diseases, but the vast majority of trypanosomatids are restricted to invertebrate hosts. Isolation and in vitro cultivation of trypanosomatids from insect hosts enable their description, characterization, and subsequently genetic and genomic studies. However, exact nutritional requirements are still unknown for most trypanosomatids and thus very few defined media are available. This mini review provides information about the role of different ingredients, recommendations and advice on essential supplements and important physicochemical parameters of culture media with the aim of facilitating first attempts to cultivate insect-infesting trypanosomatids, with a focus on monoxenous trypanosomatids.

A novel medium for long-term primary culture of hemocytes of Metapenaeus ensis.

The development of a suitable shrimp cell medium is essential for achieving a long-term cell culture and finite cell line of shrimps routinely. In this study, we have successfully developed an optimal shrimp cell medium that can be used for long-term in vitro culture and continuous subculture of the hemolymph cells (or hemocytes) of greasyback shrimp Metapenaeus ensis, designated as MeH cells, by shrimp serum-based and supplements-based optimization of the basic and growth medium. In this article, we have focused on the details for the preparation of the optimal shrimp cell medium by diluting and mixing of various stock solutions as well as the methods for isolation and primary culture of MeH cells.•A novel shrimp cell growth medium is developed for long-term shrimp hemocytes culture.•The preparation method of shrimp cell growth medium is successfully established.•Obvious cell activity and proliferation potential of isolated shrimp cells can be maintained beyond 30 days.

Shiga-Toxin-Producing Strains of Escherichia coli O104:H4 and a Strain of O157:H7, Which Can Cause Human Hemolytic Uremic Syndrome, Differ in Biofilm Formation in the Presence of CO2 and in Their Ability to Grow in a Novel Cell Culture Medium.

One pathogen that commonly causes gastrointestinal illnesses from the consumption of contaminated food is Escherichia coli O157:H7. In 2011 in Germany, however, there was a prominent outbreak of bloody diarrhea with a high incidence of hemolytic uremic syndrome (HUS) caused by an atypical, more virulent E. coli O104:H4 strain. To facilitate the identification of this lesser-known, atypical E. coli O104:H4 strain, we wanted to identify phenotypic differences between it and a strain of O157:H7 in different media and culture conditions. We found that E. coli O104:H4 strains produced considerably more biofilm than the strain of O157:H7 at 37 °C (p = 0.0470-0.0182) Biofilm production was significantly enhanced by the presence of 5% CO2 (p = 0.0348-0.0320). In our study on the innate immune response to the E. coli strains, we used HEK293 cells that express Toll-like receptors (TLRs) 2 or 4. We found that E. coli O104:H4 strains had the ability to grow in a novel HEK293 cell culture medium, while the E. coli O157:H7 strain could not. Thus, we uncovered previously unknown phenotypic properties of E. coli O104:H4 to further differentiate this pathogen from E. coli O157:H7.

Plant Growth-Promoting Rhizobacteria (PGPR) with Microbial Growth Broth Improve Biomass and Secondary Metabolite Accumulation of Cannabis sativa L.

Plant growth-promoting rhizobacteria (PGPR) are a sustainable crop production input; some show positive effects under laboratory conditions but poorly colonize host field-grown plants. Inoculating with PGPR in microbial growth medium (e.g., King's B) could overcome this. We evaluated cannabis plant (cv. CBD Kush) growth promotion by inoculating three PGPR (Bacillus sp., Mucilaginibacter sp., and Pseudomonas sp.) in King's B at vegetative and flower stages. At the vegetative stage, Mucilaginibacter sp. inoculation increased flower dry weight (24%), total CBD (11.1%), and THC (11.6%); Pseudomonas sp. increased stem (28%) dry matter, total CBD (7.2%), and THC (5.9%); and Bacillus sp. increased total THC by 4.8%. Inoculation with Mucilaginibacter sp. and Pseudomonas sp. at the flowering stage led to 23 and 18% increases in total terpene accumulation, respectively. Overall, vegetative inoculation with PGPR enhanced cannabis yield attributes and chemical profiles. Further research into PGPR inoculation onto cannabis and the subsequent level of colonization could provide key insights regarding PGPR-host interactions.

Jammed microgel growth medium prepared by flash-solidification of agarose for 3D cell culture and 3D bioprinting.

Although cells cultured in three-dimensional (3D) platforms are proven to be beneficial for studying cellular behavior in settings similar to their physiological state, due to the ease, convenience, and accessibility, traditional 2D culturing approaches are widely adopted. Jammed microgels are a promising class of biomaterials extensively suited for 3D cell culture, tissue bioengineering, and 3D bioprinting. However, existing protocols for fabricating such microgels either involve complex synthesis steps, long preparation times, or polyelectrolyte hydrogel formulations that sequester ionic elements from the cell growth media. Hence, there is an unmet need for a broadly biocompatible, high-throughput, and easily accessible manufacturing process. We address these demands by introducing a rapid, high-throughput, and remarkably straightforward method to synthesize jammed microgels composed of flash-solidified agarose granules directly prepared in a culture medium of choice. Our jammed growth media are optically transparent, porous, yield stress materials with tunable stiffness and self-healing properties, which makes them ideal for 3D cell culture as well as 3D bioprinting. The charge-neutral and inert nature of agarose make them suitable for culturing various cell types and species, the specific growth media for which do not alter the chemistry of the manufacturing process. Unlike several existing 3D platforms, these microgels are readily compatible with standard techniques such as absorbance-based growth assays, antibiotic selection, RNA extraction, and live cell encapsulation. In effect, we present a versatile, highly accessible, inexpensive, and easily adoptable biomaterial for 3D cell culture and 3D bioprinting. We envision their widespread application not just in routine laboratory settings but also in designing multicellular tissue mimics and dynamic co-culture models of physiological niches.

Developmental performance of Eristalis tenax larvae (Diptera: Syrphidae): Influence of growth media and yeast addition during captive rearing.

In holometabolous insects, life-history characteristics can vary in response to diet. The main aim of this contribution was to examine which diet best promotes larval development and survival of the aquatic saprophagous hoverfly Eristalis tenax L. (Diptera: Syrphidae). This study was motivated by a need to optimize techniques for rearing these insects in captivity. We studied how adding yeast to several rearing media based on animal droppings or decaying plant material affected development and survival in captive larvae, and whether these effects could be optimized depending on the amount of yeast and the rearing medium. In addition, premature exit of larvae was examined in two medium volumes to investigate differences in pupation. Larvae in yeast supplemented rabbit growth medium pupated and emerged faster than those in horse and antelope growth media. A high number of adult females emerged when compared to males, and both seemed to have a shorter developmental period in yeast supplemented growth media. Pupal survival was significantly greater in a mixture of droppings and plant organic matter, and a high medium volume of 140 ml (p < 0.05). Between 10% and 17% of larvae prematurely exited the aquatic medium in high (140) and low (70 ml) medium volumes, respectively. These results provide additional information that may be crucial for the successful mass rearing of E. tenax in captivity, and suggest that apart from the addition of yeast, growth medium quality and volume may be limiting factors for the production of large colonies of adults.

A closer look into the microbiome of microalgal cultures.

Although bacteria are commonly co-occurring in microalgal cultivation and production systems, little is known about their community structure and how it might be affected by specific microalgal groups or growth conditions. A better understanding about the underlying factors that determine the growth of specific bacterial populations is not only important for optimizing microalgal production processes, but also in the context of product quality when the algal biomass is to be used for future food or feed. We analyzed the bacterial community composition associated with nine microalgal strains in stock culture, maintained in two different growth media, to explore how specific taxonomic microalgal groups, microalgal origin, or the growth medium affect the bacterial community composition. Furthermore, we monitored the bacterial community composition for three Phaeodactylum strains during batch cultivation in bubble columns to examine if the bacterial composition alters during cultivation. Our results reveal that different microalgal genera, kept at the same cultivation conditions over many years, displayed separate and unique bacterial communities, and that different strains of the same genus had very similar bacterial community compositions, despite originating from different habitats. However, when maintained in a different growth medium, the bacterial composition changed for some. During batch cultivation, the bacterial community structure remained relatively stable for each Phaeodactylum strain. This indicates that microalgae seem to impact the development of the associated bacterial communities and that different microalgal genera could create distinct conditions that select for dominance of specific bacteria. However, other factors such as the composition of growth medium also affect the formation of the bacterial community structure.

Tie2-Cre-Induced Inactivation of Non-Nuclear Estrogen Receptor-α Signaling Abrogates Estrogen Protection Against Vascular Injury.

Using the Cre-loxP system, we generated the first mouse model in which estrogen receptor-α non-nuclear signaling was inactivated in endothelial cells. Estrogen protection against mechanical vascular injury was impaired in this model. This result indicates the pivotal role of endothelial estrogen receptor-α non-nuclear signaling in the vasculoprotective effects of estrogen.

Survival on land: A dark-grown seedling searching for path.

To initiate its development into a plant, a small dark-grown seedling (prior to its emergence from the ground) must penetrate through the growth media. The path that the seedling takes during this journey has yet to be explained. As such, we conducted non-destructive tests using CT scans to observe the growth of dark-grown seedlings in soil over time; we also developed a model to simulate the dynamics of an emerging seedling, and to examine effects of various growth medium conditions, including Lunar soil. It was previously postulated that, with gravitropism in a terrestrial growth medium, a dark-grown seedling would grow directly upright. However, our CT scan results showed that dark-grown soybean seedlings departed from the vertical path in soil, as far as a lateral distance of approximately 10 mm. The phenomenon of the non-straight path was also demonstrated by the model results. Through simulations, we found that an emerging seedling naturally weaves through the particles of growth medium, in search for the path of least resistance. As a result, the seedling ends up travelling a longer distance. Compared with a seedling that was artificially forced to take a straight path in a growth media, the seedling taking the natural path encountered significantly lower resistances (20% lower) from the growth medium, while travelled 12% longer distance during the emergence process. A seedling encountered a much higher impedance in Lunar soil. Our results suggest that taking the path of least resistance, in addition to shaping and orientating itself for mechanical advantage, are strategies evolved by plant species that have contributed to its vast success. An understanding of plant behavior and survival strategies on Earth lay the foundation for future research in agriculture in novel environments, including on celestial bodies.

In Situ Investigation of Pseudomonas aeruginosa Biofilm Development: Interplay between Flow, Growth Medium, and Mechanical Properties of Substrate.

To better understand the impact of biomaterial mechanical properties and growth medium on bacterial adhesion and biofilm formation under flow, we investigated the biofilm formation ability of Pseudomonas aeruginosa in different media on polydimethylsiloxane (PDMS) of different stiffness in real time using a microfluidic platform. P. aeruginosa colonization was recorded with optical microscopy and automated image analysis. The bacterial intracellular level of cyclic diguanylate (c-di-GMP), which regulates biofilm formation, was monitored using the transcription of the putative adhesin gene (cdrA) as a proxy. Contrary to the previous supposition, we revealed that PDMS material stiffness within the tested range has negligible impact on biofilm development and biofilm structures, whereas culture media not only influence the kinetics of biofilm development but also affect the biofilm morphology and structure dramatically. Interestingly, magnesium rather than previously reported calcium was identified here to play a decisive role in the formation of dense P. aeruginosa aggregates and high levels of c-di-GMP. These results demonstrate that although short-term adhesion assays bring valuable insight into bacterial and material interactions, long-term evaluations are essential to better predict overall biofilm outcome. The microfluidic system developed here presents a valuable application potential for studying biofilm development in situ. .

Quick bioassay test from tillers for detecting ALS herbicide resistance of weedy rice and barnyardgrass

Resistance to acetolactate synthase (ALS) inhibitors have increased recently in South Brazil where the major weeds of flooded rice (barnyardgrass and weedy rice) have evolved resistance to imazapyr+imazapic. The aim of this research was to evaluate a growth medium for tissue regeneration of tillers in barnyardgrass, as well as an agar-based bioassays test (also from tillers) to detect susceptible and resistant biotypes of weedy rice and barnyardgrass to imazapyr+imazapic in vitro. Greenhouse experiments were conducted to detect ALS-resistant (R) and susceptible (S) weedy rice and barnyardgrass biotypes, and bioassays were carried out to evaluate an adequate growth medium for barnyardgrass tiller regeneration and determine the concentration of herbicide to distinguish R and S plants. The culture medium that provided a suitable barnyardgrass growth was MS 50% with the addition of benzylamino-purine. The tissue regeneration in vitro with the growth medium containing imazapyr+imazapic allowed to discriminate between R and S barnyardgrass and weedy rice plants. The concentration required for satisfactory control of susceptible barnyardgrass and weedy rice explants grown in vitro was 0.9 µM and 1.3 µM of imazapyr+imazapic herbicide, respectively. The bioassay in vitro using tiller regeneration provides an opportunity to predict effectively imazapyr+imazapic resistance in barnyardgrass and weedy rice.

Preparative protein concentration from biofluids by epitachophoresis.

With increasing demands on protein analyses in complex biological matrices, the insistence on developing new sample preparation techniques is rising. Recently, we introduced a new displacement electrophoresis technique (epitachophoresis) and instrumentation for preparative concentration and cleaning of DNA samples. This work describes the possibility of applying this device to protein samples. We have developed a method for the epitachophoretic concentration of proteins in a cationic mode and tested it by concentrating and collecting the protein zones from complex biological matrices (urine and growth medium). Under optimized conditions, we have obtained recoveries up to 99%. Furthermore, the applicability of the developed method was proven by concentrating and collecting the cytochrome c zone from a HeLa cell line growth medium, where the protein cytochrome c was released during cell apoptosis.

The challenges of growing orchids from seeds for conservation: An assessment of asymbiotic techniques.

Lewis Knudson first successfully germinated orchid seeds asymbiotically on artificial medium in 1922. While many orchid species have since been grown asymbiotically, the tremendous variation in how species respond to artificial medium and growth conditions ex situ has also become apparent in the past century. In this study, we reviewed published journal articles on asymbiotic orchid seed germination to provide a summary of techniques used and to evaluate if these differ between terrestrial and epiphytic species, to identify areas where additional research is needed, and to evaluate whether asymbiotic germination could be used more often in ex situ conservation. We found articles reporting successful asymbiotic germination of 270 species and 20 cultivars across Orchidaceae. Researchers often used different techniques with epiphytic versus terrestrial species, but species-specific responses to growth media and conditions were common, indicating that individualized protocols will be necessary for most species. The widespread success in generating seedlings on artificial media suggests that asymbiotic techniques should be another tool for the conservation of rare orchid species. Further advances are needed in understanding how to introduce mycorrhizae to axenically grown orchids and to maximize the viability of seedlings reintroduced into natural habitats to fully utilize these methods for conservation.

Sustainable utilization of unavoidable food waste into nutritional media for the isolation of bacterial culture for the removal of heavy metals.

This study aims to reuse food waste (FW) as growth media for bacterial cultures for bioremediation of heavy metal. The best natural medium was selected based on the carbon, nitrogen, and other elements. The batch culture of Comamonas terrae showed growth stability for 16 days in the pig bone medium. C. terrae showed the best growth at pH of 7.4, temperature of 35 °C, and medium concentration of 10 g/L. The C. terrae showed heavy metal (HM) removal efficiencies of Cd (52 %) Cr (63 %) Pb (62 %) and Zn (55 %). In addition, the Fourier transform infrared spectroscopy results revealed the bioadsorption of HM in C. terrae. The study suggests the C. terrae can efficiently remove HM and C. terrae may be used for bioremediation of HM. Therefore, pig bone waste is a cost-effective medium and a good solution for the valorization and reuse of FW in line with the circular economy.

Supply Chain Disruptions During COVID-19 Pandemic Uncover Differences in Keratinocyte Culture Media.

Various culture media are used to propagate keratinocytes (KCs) in vitro. The COVID-19 pandemic resulted in supply chain shortages necessitating substitutions to standard laboratory protocols, which resulted in many laboratories having to use culture media different from those they typically use. We screened available media on the KC line N/TERT2G and found that biological responses varied considerably across three culture media: KC serum-free media, KC growth medium 2, and defined media. We observed qualitative and quantitative differences in proliferation; KCs cultured in defined media had significantly lower proliferative capacity. KC differentiation was assessed by western blot for CLDN1, occludin, cytokeratin-10, and loricrin. Elevated expression of differentiation markers was observed in cells cultured in either KC growth medium 2 or defined media compared with those in cells cultured in KC serum-free media. KC barrier function was measured by transepithelial electrical resistance. KCs cultured in KC growth medium 2 and defined media developed significantly higher transepithelial electrical resistance than those cultured in KC serum-free media, and when treated with IL-4 and IL-13 or IL-17A, we observed variable responses. H&E staining on day 5 -post-differentiation showed greater epithelial thickness in KCs cultured in defined media and KC growth medium 2 than in those cultured in KC serum-free media. These findings show that the choice of culture media impacts the biological response of KCs in a manner that persists through differentiation in the same media.

Production of lipids and proteins from marine diatoms under changing pH and silica.

Diatom algae are increasingly explored as an alternative sustainable source for functional biomolecules likes fucoxanthin, and eicosapentaenoic acid. But biomolecule quantity and quantity are influenced by growth conditions. So, effect of differential silica concentration (0-120 mg L-1) and medium pH (5.5-9.5) on growth and cellular biochemical composition of commercially important marine diatom species were studied. Growth rate of Thalassiosira sp., Skeletonema sp., and Chaetoceros sp., was higher with 30 mg L-1 Si at a pH of 7.5-8.5. Highest carbohydrate (153.71 mg g-1) and protein (17.34 mg g-1) content was found in Skeletonema sp. Silica concentration positively influenced chlorophyll and carotenoid content in a dose dependent manner. A medium pH of 8.5 and Si concentration between 60 and 120 mg L-1 was ideal for lipid production. The optimum concentration of Si and pH for maximum biomolecule production have been reported with further scope of utilizing these conditions in commercial scale systems.

Biochemical and Physiological Changes during Early Adventitious Root Formation in Chrysanthemum indicum Linné Cuttings.

Chrysanthemum indicum is an important ornamental and medicinal plant that is often difficult to propagate commercially because of its poor germination and low seed viability. This plant is mostly propagated by cutting, but the rooting is slow and non-uniform. The present investigation evaluated the regeneration capacity of stem cutting by examining the influence of auxins, growth medium, temperature, and explant type on adventitious root formation in C. indicum. The auxin-treated cuttings were planted in different growth substrates under greenhouse conditions. Among the different auxins tested, indole-3-butyric acid (IBA) more effectively induced roots. The cutting position of stock plants influenced rooting capacity. Cutting the stock plants from the apical region enhanced root number and length in the explants. Among the different explant types, apical stem cuts with 2000 ppm IBA produced a significantly higher number of adventitious roots when grown in vermiculite and perlite (V + P) at a ratio of 1:1 at 25 °C. High-performance liquid chromatography (HPLC) analysis revealed that protocatechuic acid, gentisic acid, chlorogenic acid, biochanin A, salicylic acid, caffeic acid, glycitein, and luteolin were the most dominant phenolic compounds present in C. indicum. These results indicate that IBA treatment promoted the synthesis and accumulation of phenolic compounds in C. indicum stem cuttings at the time of root formation. The present results demonstrate that applying auxins is essential for early root initiation and higher rooting success and thus may be beneficial for vegetative C. indicum propagation.

Accelerating gametophytic growth in the model hornwort Anthoceros agrestis.

Premise: Hornworts belong to a unique lineage of bryophytes with critical traits for elucidating the evolution of land plants; however, the development of functional genetic tools for hornworts has been hampered by their relatively slow gametophytic growth. Methods: To identify the external factors that influence the development of hornwort gametophytes and potentially augment their growth, we evaluated the contributions of several culture medium components on the axenic gametophytic growth of Anthoceros agrestis, a model hornwort. A streamlined growth assay utilizing semiautomated image analysis was developed to rapidly quantify and compare tissue development spanning four weeks of culture on solidified medium. Results: The addition of sucrose, ammonium nitrate, activated charcoal, pH buffering, and growth regulators (2,4-dichlorophenoxyacetic acid, 6-benzylaminopurine, and thidiazuron) affected gametophyte tissue survival, growth patterns, and the rate of thalli growth. Subsequently, an optimized medium composition and growth regimen for accelerating A. agrestis gametophytic growth were formulated, which at four weeks of culture increased the tissue wet weight by 2.1- to 8.5-fold compared with other previously utilized hornwort growth media. Discussion: Our protocol for generating vigorous starting material and accelerated tissue regeneration is pertinent for advancing gene function characterization and genome editing in hornworts.