Petri Dish Cultures Research Articles

Gel microspheres enhance the stemness of ADSCs by regulating cell-ECM interaction

The gel microsphere culture system (GMCS) showed various advantages for mesenchymal stem cell (MSC) expansion and delivery, such as high specific surface area, small and regular shape, extensive adjustability, and biomimetic properties. Although various technologies and materials have been developed to promote the development of gel microspheres, the differences in the biological status of MSCs between the GMCS and the traditional Petri dish culture system (PDCS) are still unknown, hindering gel microspheres from becoming a culture system as widely used as petri dishes. In the previous study, an excellent “all-in-one” GMCS has been established for the expansion of human adipose-derived MSCs (hADSCs), which showed convenient cell culture operation. Here, we performed transcriptome and proteome sequencing on hADSCs cultured on the “all-in-one” GMCS and the PDCS. We found that hADSCs cultured in the GMCS kept in an undifferentiation status with a high stemness index, whose transcriptome profile is closer to the adipose progenitor cells (APCs) in vivo than those cultured in the PDCS. Further, the high stemness status of hADSCs in the GMCS was maintained through regulating cell-ECM interaction. For application, bilayer scaffolds were constructed by osteo- and chondro-differentiation of hADSCs cultured in the GMCS and the PDCS. The effect of osteochondral regeneration of the bilayer scaffolds in the GMCS group was better than that in the PDCS group. This study revealed the high stemness and excellent functionality of MSCs cultured in the GMCS, which promoted the application of gel microspheres in cell culture and tissue regeneration.

Diffuse Associations Between Drosophila suzukii and Filamentous Fungal Microbes May Alter Caneberry Disease Dynamics.

Interactions between microorganisms and frugivorous insects can modulate fruit rot disease epidemiology. Insect feeding and/or oviposition wounds may create opportunities for fungal infection. Passive and active dispersal of fungal inoculums by adult insects also increases disease incidence. In fall-bearing raspberries and blackberries, such vectoring interactions could increase crop damage from the invasive pestiferous vinegar fly Drosophila suzukii (spotted-wing drosophila). Periods of peak D. suzukii activity are known to overlap with several species of primary fruit rot pathogen, particularly Botrytis cinerea and Cladosporium cladosporioides, and previous work indicates that larvae co-occur with and feed on various filamentous fungi at low rates. To further our understanding of the epidemiological consequences that may emerge from these associations, we surveyed the filamentous fungal community associated with adult D. suzukii, isolating and molecularly identifying fungi externally and internally (indicating feeding) from field-collected adults over 3 years. We isolated and identified 37 unique genera of fungi in total, including known raspberry pathogens. Most fungi were detected infrequently, and flies acquired and carried fungi externally at higher richness, frequency, and density relative to internally. In a worst-case scenario laboratory vectoring assay, D. suzukii adults were able to transfer B. cinerea and C. cladosporioides to sterile media at 0, 24, 48, and 72 h after exposure to sporulating cultures in Petri dishes. These results collectively suggest an adventitious vectoring association between D. suzukii and fruit rot fungi that has the potential to alter caneberry disease dynamics.

Performance valuation of onion (Allium cepa L.) genotypes under different levels of salinity for the development of cultivars suitable for saline regions.

Abiotic stress, especially salt stress, is one of the major barriers to crop production worldwide. Crops like onion that belong to the glycophytic group are more sensitive to salinity stress. A huge study regarding the influence of salinity stress on the growth and development of crops has already been done and is still ongoing. One of the major targets of the research is to develop genotypes that have enhanced performance under stress environments. The world needs more of these types of genotypes to combat the ever-growing salt-stressed soils. Therefore, a number of germplasm were studied during the 2019-2020 and 2020-2021 seasons under different salt concentrations to identify tolerant genotypes as well as to study the plants' responses at different growth stages against elevated salinity levels. A 2-year study was conducted where germination potential was evaluated in the first year and carried out in petri dish culture of seeds, followed by plastic pot culture for plant establishment and bulb development evaluation during the second year. Four different saline water solutions having different salt concentrations (0, 8, 10, and 12 dS m-1) were applied to the petri dishes and pots as the source of water for plants in both seasons. Results indicated that a significant reduction in plants' performance occurs under higher salinity levels. Salt concentration had an adverse impact on germination, leaf development and growth, the height of plants, bulb size and shape, and the bulb weight of onion. All the growth phases of onion are sensitive to elevated concentrations. Variable performances were observed in the genotypes under stress conditions, and a few genotypes (Ac Bog 409, Ac Bog 414, Ac Bog 424, Ac Bog 430, Ac Bog 417, Ac Bog 419, Ac Bog 420, Ac Bog 422, and Ac Bog 425) having some sort of tolerance to salt stress were identified, which might be recommended for mass production. Tolerance indices could successfully be applied in selecting the salt-tolerant genotypes. Thus, the present findings and the identified genotypes could be further utilized in salt stress improvement research on onion.

Dispersion characteristics of bioaerosols during treatment of rural solid waste in northwest China

In rural China, the release of bioaerosols containing pathogens from solid waste dumps poses a potential health risk to the local population. Here, we sampled bioaerosols from rural solid waste-treatment in four provinces of northwest China to investigate their emission and dispersion characteristics in order to provide a scientific basis for control and risk reduction of bioaerosols released from rural sanitation facilities. The airborne bioaerosol concentrations and particle size distributions were calculated using an Anderson six-stage airborne microbial sampler and counting with its internal Petri dish culture. High-throughput sequencing was used to characterize the microbial composition at different sampling sites and to explore possible influencing factors, while the health risk associated with exposure was estimated based on average daily dose-rate. The highest concentration point values of bacteria and fungi in bioaerosols near the solid waste were 63,617 ± 15,007 and 8044 ± 893 CFU/m³, respectively. Furthermore, the highest concentration point values of Enterobacteriaceae was 502 ± 35 CFU/m³. Most bioaerosols were coarse particles larger than 3.3 μm. Potentially pathogenic genera of winter-indicator species detected in the air were primarily Delftia, Rhodococcus and Aspergillus. The composition of solid waste and environmental conditions are important factors in determining the characteristics of bioaerosols. Local residents are exposed to bioaerosols mainly through inhalation. Children are at a particularly high risk of exposure through both inhalation and skin contact. The results of this study show that bioaerosols in the vicinity of rural solid waste dumps pose a health risk to the surrounding population. More suitable risk assessment criteria for rural areas should be established, and corresponding control and protection measures should be taken from three aspects: generation source and transmission pathway, as well as the recipient.

A microfluidic platform integrating dynamic cell culture and dielectrophoretic manipulation for in situ assessment of endothelial cell mechanics.

The function of vascular endothelial cells (ECs) within the complex vascular microenvironment is typically modulated by biochemical cues, cell-cell interactions, and fluid shear stress. These regulatory factors play a crucial role in determining cell mechanical properties, such as elastic and shear moduli, which are important parameters for assessing cell status. However, most studies on the measurement of cell mechanical properties have been conducted in vitro, which is labor-intensive and time-consuming. Notably, many physiological factors are lacking in Petri dish culture compared with in vivo conditions, leading to inaccurate results and poor clinical relevance. Herein, we developed a multi-layer microfluidic chip that integrates dynamic cell culture, manipulation and dielectrophoretic in situ measurement of mechanical properties. Furthermore, we numerically and experimentally simulated the vascular microenvironment to investigate the effects of flow rate and tumor necrosis factor-alpha (TNF-α) on the Young's modulus of human umbilical vein endothelial cells (HUVECs). Results showed that greater fluid shear stress results in increased Young's modulus of HUVECs, suggesting the importance of hemodynamics in modulating the biomechanics of ECs. In contrast, TNF-α, an inflammation inducer, dramatically decreased HUVEC stiffness, demonstrating an adverse impact on the vascular endothelium. Blebbistatin, a cytoskeleton disruptor, significantly reduced the Young's modulus of HUVECs. In summary, the proposed vascular-mimetic dynamic culture and monitoring approach enables the physiological development of ECs in organ-on-a-chip microsystems for accurately and efficiently studying hemodynamics and pharmacological mechanisms underlying cardiovascular diseases.

Morphological and cultural characterization of two Fusarium isolates causing wheat fusarium head blight in Algeria

In Algeria, fusarium head blight is a serious fungal disease of wheat caused by Fusarium genus. The study of the epidemiological cycle of the disease in the field is always hampered by a major problem which is the identification of the parasitic complex of Fusarium species. These species can only be identified after Petri dish culture and observation of a set of morphological criteria such as color, cultural aspect of colonies, growth rate of mycelium and the form of isolate macroconidia. To this end, the objective of this present study is to identify two Fusarium isolates based on a set of morphological and cultural criteria mentioned above. The results obtained for all the morphological characters studied show phenotypic variability between the two isolates. Based on these results, we were able to characterize these two isolates as F. graminearum and F. culmorum. Indeed, morphological and cultural characterization, although important, remains insufficient. It should therefore be further investigated by molecular characterization, in order to highlight any differences between the two isolates studied.

A Novel Fluidic Platform for Semi-Automated Cell Culture into Multiwell-like Bioreactors.

In this work, we developed and characterized a novel fluidic platform that enables long-term in vitro cell culture in a semi-automated fashion. The system is constituted by a control unit provided with a piezoelectric pump, miniaturized valves, and a microfluidic network for management and fine control of reagents’ flow, connected to a disposable polymeric culture unit resembling the traditional multiwell-like design. As a proof of principle, Human Umbilical Vein Endothelial Cells (HUVEC) and Human Mesenchymal Stem Cells (hMSC) were seeded and cultured into the cell culture unit. The proliferation rate of HUVEC and the osteogenic differentiation of hMSC were assessed and compared to standard culture in Petri dishes. The results obtained demonstrated that our approach is suitable to perform semi-automated cell culture protocols, minimizing the contribution of human operators and allowing the standardization and reproducibility of the procedures. We believe that the proposed system constitutes a promising solution for the realization of user-friendly automated control systems that will favor the standardization of cell culture processes for cell factories, drug testing, and biomedical research.

Boron-mediated aluminium stress tolerance under aluminium toxicity at germination and early seedling stages of wheat

Aluminium stress is one of the major problems of wheat production that significantly reduces the growth and development. Al toxicity can be recovered by exogenous application of different growth inducing nutrient elements. Thus, this study was conducted to evaluate the ameliaration effect of B under Al toxicity. Therefore, a petri dish and hydroponic culture experiment of wheat was conducted at Crop Physiology Laboratory, Department of Crop Botany, Bangladesh Agricultural University, Mymensingh during the period from January 2016 to February 2017 to investigate the effect of boron on amelioration of aluminium toxicity in germination and seedling stage. Both the experiments were designed in completely randomized design (CRD) with three replications. The experiments were comprised of four levels of boron and aluminium concentrations viz., 0 µM B + 0 µM Al (control), 0 µM Al + 40 µM B, 200 µM Al + 40 µM B and 0 µM B + 200 µM Al and five wheat varieties viz; BARI Gom-23, BARI Gom-24, BARI Gom-28, BARI Gom-27 and BARI Gom-30. Results indicated that germination percentage, radicle and plumule length, root and shoot length, leaf length, fresh and dry mass plant-1 were greater in 0 µM Al + 40 µM B treated plants than 0 µM B + 200 µM Al induced conditions. It indicates that wheat seedlings are susceptible to aluminium and boron can ameliorate aluminium toxicity. However, among the studied varieties, the reduction of dry mass under aluminium stress was minimum in BARI Gom-28 followed by BARI Gom-23 indicating BARI Gom-28 was more tolerant to aluminium stress than the other varieties. On the contrary, the varieties, BARI Gom-27 and BARI Gom-24 were more susceptible to aluminium stress. So it indicates that aluminium stress severely affects the growth and developments especially in the sensitive varieties and tolerant varieties have the self-ability to grow and develop even under aluminium stress condition.
 Progressive Agriculture 32 (2): 127-139, 2021

Biocompatible micro tweezers for 3D hydrogel organoid array mechanical characterization.

This study presents novel biocompatible Polydimethylsiloxane (PDMS)-based micromechanical tweezers (μTweezers) capable of the stiffness characterization and manipulation of hydrogel-based organoids. The system showed great potential for complementing established mechanical characterization methods such as Atomic Force Microscopy (AFM), parallel plate compression (PPC), and nanoindentation, while significantly reducing the volume of valuable hydrogels used for testing. We achieved a volume reduction of ~0.22 μl/sample using the μTweezers vs. ~157 μl/sample using the PPC, while targeting high-throughput measurement of widely adopted micro-mesoscale (a few hundred μm-1500 μm) 3D cell cultures. The μTweezers applied and measured nano-millinewton forces through cantilever’ deflection with high linearity and tunability for different applications; the assembly is compatible with typical inverted optical microscopes and fit on standard tissue culture Petri dishes, allowing mechanical compression characterization of arrayed 3D hydrogel-based organoids in a high throughput manner. The average achievable output per group was 40 tests per hour, where 20 organoids and 20 reference images in one 35 mm petri dish were tested, illustrating efficient productivity to match the increasing demand on 3D organoids’ applications. The changes in stiffness of collagen I hydrogel organoids in four conditions were measured, with ovarian cancer cells (SKOV3) or without (control). The Young’s modulus of the control group (Control—day 0, E = 407± 146, n = 4) measured by PPC was used as a reference modulus, where the relative elastic compressive modulus of the other groups based on the stiffness measurements was also calculated (control-day 0, E = 407 Pa), (SKOV3-day 0, E = 318 Pa), (control-day 5, E = 528 Pa), and (SKOV3-day 5, E = 376 Pa). The SKOV3-embedded hydrogel-based organoids had more shrinkage and lowered moduli on day 0 and day 5 than controls, consistently, while SKOV3 embedded organoids increased in stiffness in a similar trend to the collagen I control from day 0 to day 5. The proposed method can contribute to the biomedical, biochemical, and regenerative engineering fields, where bulk mechanical characterization is of interest. The μTweezers will also provide attractive design and application concepts to soft membrane-micro 3D robotics, sensors, and actuators.

Xanthoepocin, a photolabile antibiotic of Penicillium ochrochloron CBS\xa0123823 with high activity against multiresistant gram-positive bacteria

BackgroundWith the steady increase of antibiotic resistance, several strategies have been proposed in the scientific community to overcome the crisis. One of many successful strategies is the re-evaluation of known compounds, which have been early discarded out of the pipeline, with state-of-the-art know-how. Xanthoepocin, a polyketide widespread among the genus Penicillium with an interesting bioactivity spectrum against gram-positive bacteria, is such a discarded antibiotic. The purpose of this work was to (i) isolate larger quantities of this metabolite and chemically re-evaluate it with modern technology, (ii) to explore which factors lead to xanthoepocin biosynthesis in P. ochrochloron, and (iii) to test if it is beside its known activity against methicillin-resistant Staphylococcus aureus (MRSA), also active against linezolid and vancomycin-resistant Enterococcus faecium (LVRE)—a very problematic resistant bacterium which is currently on the rise.ResultsIn this work, we developed several new protocols to isolate, extract, and quantify xanthoepocin out of bioreactor batch and petri dish-grown mycelium of P. ochrochloron. The (photo)chemical re-evaluation with state-of-the-art techniques revealed that xanthoepocin is a photolabile molecule, which produces singlet oxygen under blue light irradiation. The intracellular xanthoepocin content, which was highest under ammonium-limited conditions, varied considerably with the applied irradiation conditions in petri dish and bioreactor batch cultures. Using light-protecting measures, we achieved MIC values against gram-positive bacteria, including methicillin-resistant Staphylococcus aureus (MRSA), which were up to 5 times lower than previously published. In addition, xanthoepocin was highly active against a clinical isolate of linezolid and vancomycin-resistant Enterococcus faecium (LVRE).ConclusionsThis interdisciplinary work underlines that the re-evaluation of known compounds with state-of-the-art techniques is an important strategy in the combat against multiresistant bacteria and that light is a crucial factor on many levels that needs to receive more attention. With appropriate light protecting measures in the susceptibility tests, xanthoepocin proved to be a powerful antibiotic against MRSA and LVRE. Exploring the light response of other polyketides may be pivotal for re-introducing previously discarded metabolites into the antibiotic pipeline and to identify photosensitizers which might be used for (antimicrobial) photodynamic therapies.

Synthesis and herbicidal activity of pyrimidyl‐1,2,4‐triazole derivatives containing aryl sulfonyl moiety

AbstractTo discover novel heterocyclic compounds with high herbicidal activity, a series of new pyrimidyl‐1,2,4‐triazole derivatives (Ι‐1 to Ι‐41) was designed and synthesized by incorporating the activating 1,2,4‐triazole, pyrimidine, and phenyl sulfonyl structural templets. Their structures were clearly identified by proton nuclear magnetic resonance (1H NMR), carbon‐13 nuclear magnetic resonance (13C NMR), high‐resolution mass spectroscopy (HRMS), and X‐ray single‐crystal diffraction (compound Ι‐3). Their herbicidal activities against monocotyledons (Echinochloa crusgalli, Sorghum bicolor) and dicotyledons (Raphanus sativus, Brassica campestris, Cucumis sativus, and Medicago sativa) were evaluated using a Petri dish culture method. The results indicated that most of title compounds exhibited significant herbicidal potencies against the plant growth at 100 mg·L−1, while some of them, such as Ι‐2 and Ι‐4, showed remarkable inhibitory effects on weed control at 10 mg·L−1 and 100 mg·L−1. On the whole, compound Ι‐3, which was found to possess an even broader spectrum of inhibitory activities against the roots and stalks of Echinochloa crusgalli, Raphanus sativus, Brassica campestris, Cucumis sativus, and Medicago sativa at 10 and 100 mg·L−1 than that of the commercial herbicide Flumetsulam, could be employed as a new lead structure for further optimization to find more potent herbicides.

Lytic Exoenzymes of Soil Strains of Bacillus Representatives and Manifestations of their Biological Activity

Bacteria of the genus Bacillus make up a significant (8–12%) part of the soil microbiome. Manifestation of their biological activity, in particular, the antagonistic and lytic activity against other microorganisms directly depends on their exometabolites. According to the literature, such properties of soil bacteria of the genus Bacillus thus can be related to their various lytic exoenzymes. Aim. To evaluate the role of lytic exoenzymes of the studied soil bacteria strains of the genus Bacillus in the manifestation of their biological (antagonistic, lysing) activity. Methods. The antagonistic activity of bacteria strains of the genus Bacillus against phytopathogenic micromycetes was determined by the method of double culture in Petri dishes on potato-glucose agar. For qualitative analysis of the presence of extracellular enzymes, strains of bacteria of the genus Bacillus were plated on Petri dishes with solid mineral-salt medium and a suitable substrate inducer. The ratio of the diameter of substrate hydrolysis zone to the diameter of the colony was taken as the relative enzymatic activity of the culture. Bacteriolytic activity of the studied strains was determined by the change in optical density of living cells of phytopathogenic bacteria suspension at 540 nm. Results. Six strains of bacteria of the genus Bacillus were selected by the results of preliminary screening, with at least five types of lytic activity, namely proteolytic, chitinase, amylolytic, cellulase, and xylanase of different levels (low, average, high). Analysis of the antagonistic activity of the selected strains of bacteria of the genus Bacillus to the main groups of phytopathogenic bacteria (six test cultures) singled out the strain Bacillus sp. 41 for a careful study of the nature and spectrum of its antagonism. Analysis of the level of antagonistic activity of the selected Bacillus strains against the phytopathogenic micromycetes showed that the minimum decrease of antagonism (the decrease of growth inhibition zones) during the observation period (at the 3rd and 7th days) was in Bacillus sp. 41 strain. Therefore, only this strain showed a stable and relatively wide range of antagonistic activity against phytopathogens of bacterial and fungal etiology. The nature of this antagonism is probably complex and conditioned by the participation of various biochemical mechanisms, in particular, the synthesis of a complex of lytic exoenzymes. To assess the lysing activity of Bacillus strains, three strains with the highest proteolytic and cellulolytic activity of exoenzymes were taken from the six previously chosen. Only Bacillus sp.1913 strain showed high (70%) lytic activity against gram-negative polyphagous phytopathogen Pseudomonas syringae pv. syringae UCM B-1027T. Such activity of the strain did not manifest against the rest of the phytopathogenic test cultures. The high lytic activity of Bacillus sp. 1913 strain may be associated with high activity of exogenous proteases and cellulases of the lytic complex, which is quite consistent with the literature data on the lytic activity of bacteria of the genus Bacillus. Conclusions. The spectrum and activity of lytic exoenzymes of strains of the studied soil bacteria of the genus Bacillus indicate the indirect participation of these enzymes in the manifestation of biological activity (antagonistic and lytic).

Upgrading Hepatic Differentiation and Functions on 3D Printed Silk-Decellularized Liver Hybrid Scaffolds.

We developed hybrid liver-specific three-dimensional (3D) printed scaffolds using a solubilized native decellularized liver (DCL) matrix and silk fibroin (SF) and investigated their ability to support functional cultures of hepatic cells. Rat livers were decellularized by perfusing detergents via the portal vein, solubilized using pepsin to form DCL, and characterized. SF blended with gelatin (8% w/v) was optimized with varying percentages of DCL to obtain silk gelatin-DCL bioink (SG-DCL). Different compositions of SG-DCL were studied by rheology for optimum versatility and print fidelity. 3D printed six-layered scaffolds were fabricated using a sophisticated direct-write 3D bioprinter. Huh7 cells were cultured on the 3D printed scaffolds for 3 weeks. 3D printed SG scaffolds without DCL along with 2D films (SG and SG-DCL) and 2D culture on tissue culture Petri dish control were used for comparative studies. The DCL matrix showed the absence of cells in histology and SEM. The combined SG-DCL ink at all of the studied DCL percentages (1-10%) revealed shear-thinning behavior in the printable range. The storage modulus value for the SG-DCL ink at all DCL percentages was higher than the loss modulus. In comparison to 2D controls, hepatic cells cultured on 3D SG-DCL revealed increased proliferation until 2 weeks and an upregulated expression of hepatocyte markers, including asialoglycoprotein receptor 1 (ASGR1). The Wnt pathway gene β-catenin was upregulated by more than 4-fold in 3D SG-DCL on day 3, while it showed a decline on day 7 as compared to 3D SG and also 2D controls. The expression of the epithelial cell adhesion molecule (EpCAM) was however lower in both 2D SG-DCL (2-fold) and 3D SG-DCL (2.5-fold) as compared to that in 2D controls. Immunofluorescence studies validated the protein expression of ASGR1 in 3D SG-DCL. Albumin (ALB) was not identified on SG scaffolds but prominently expressed in 3D SG-DCL constructs. In comparison to 2D SG, both ALB (1.8-fold) and urea (5-fold) were enhanced in cells cultured on 3D SG-DCL on day 7 of culture. Hence, the SG-DCL 3D printed scaffolds provide a conducive microenvironment for elevating differentiation and functions of hepatic cells possibly through an involvement of the Wnt/β-catenin signaling pathway.

In Vitro Development of a New Sponge-Based Delivery System for Intracanal Antimicrobial Administration in Endodontic Treatment.

This study aimed to evaluate the in vitro performance of collagen-based sponges as a drug delivery system for intracanal antimicrobial administration. Four groups of loaded collagen-based sponges (A, 0.3% w/v amoxicillin trihydrate: potassium clavulanate (4:1); B, 0.03% w/v chlorhexidine gluconate [CHX]; C, 0.3% w/v amoxicillin trihydrate: potassium clavulanate (4:1) and 0.03% w/v CHX; D, 1% w/v amoxicillin trihydrate: potassium clavulanate (4:1) and 0.03% w/v CHX) were designed. Release kinetics were tested in vitro on cultures in Petri dishes, and the effect on bacterial biofilms was studied ex vivo on 114 extracted human single-rooted teeth. Biofilm formation was tested by scanning electron microscopy (SEM). Collagen sponges containing amoxicillin and chlorhexidine showed a time-sustained antimicrobial effect in vitro and were also able to destroy mature biofilms ex vivo. This datum was validated by means of SEM-based study of E. faecalis and S. aureus biofilms.

Volatile 1-octanol of tea (Camellia sinensis L.) fuels cell division and indole-3-acetic acid production in phylloplane isolate Pseudomonas sp. NEEL19

Tea leaves possess numerous volatile organic compounds (VOC) that contribute to tea’s characteristic aroma. Some components of tea VOC were known to exhibit antimicrobial activity; however, their impact on bacteria remains elusive. Here, we showed that the VOC of fresh aqueous tea leaf extract, recovered through hydrodistillation, promoted cell division and tryptophan-dependent indole-3-acetic acid (IAA) production in Pseudomonas sp. NEEL19, a solvent-tolerant isolate of the tea phylloplane. 1-octanol was identified as one of the responsible volatiles stimulating cell division, metabolic change, swimming motility, putative pili/nanowire formation and IAA production, through gas chromatography-mass spectrometry, microscopy and partition petri dish culture analyses. The bacterial metabolic responses including IAA production increased under 1-octanol vapor in a dose-dependent manner, whereas direct-contact in liquid culture failed to elicit such response. Thus, volatile 1-octanol emitting from tea leaves is a potential modulator of cell division, colonization and phytohormone production in NEEL19, possibly influencing the tea aroma.

Toward improved in vitro models of human cancer

Cancer is a leading cause of death across the world and continues to increase in incidence. Despite years of research, multiple tumors (e.g., glioblastoma, pancreatic cancer) still have limited treatment options in the clinic. Additionally, the attrition rate and cost of drug development have continued to increase. This trend is partly explained by the poor predictive power of traditional in vitro tools and animal models. Moreover, multiple studies have highlighted that cell culture in traditional Petri dishes commonly fail to predict drug sensitivity. Conversely, animal models present differences in tumor biology compared with human pathologies, explaining why promising therapies tested in animal models often fail when tested in humans. The surging complexity of patient management with the advent of cancer vaccines, immunotherapy, and precision medicine demands more robust and patient-specific tools to better inform our understanding and treatment of human cancer. Advances in stem cell biology, microfluidics, and cell culture have led to the development of sophisticated bioengineered microscale organotypic models (BMOMs) that could fill this gap. In this Perspective, we discuss the advantages and limitations of patient-specific BMOMs to improve our understanding of cancer and how these tools can help to confer insight into predicting patient response to therapy.

Antagonistic activity of dominant strains of bacteria of spring barley rhizosphere against phytopathogenic micromycetes

The purpose of the work. The aim of the study was to conduct research on the manifestation of the dominant representatives of the bacterial biome of the rhizosphere of spring barley antagonistic activity against phytopathogenic micromycetes. Research methods. The standard diffusion method of double culture in Petri dishes was used to study the antagonistic properties of the dominant strains of rhizosphere bacteria. The level of antagonistic activity of microorganisms was assessed by the indicator (%) of inhibition of growth and development of the mycelium of micromycetes Fusarium sporotrichioides Sherb. 23.2, Alternaria alternata (Fr.) Keissl. 3.45, Nigrospora oryzae (Berk. & Broome) Petch. 18.77. Results and scope of their application. Studies have shown that Bacillus methylotrophicus 10 had an inhibitory effect on Fusarium sporotrichioides Sherb. 23.2 ‒ 77,4%, Alternaria alternata (Fr.) Keissl. 3.45 – 66,6% and Nigrospora oryzae (Berk. & Broome) Petch. 18.77 – 86,7%, while the bacteria Phyllobacterium ifriqiyense 1 showed activity against phytopathogens by 45,1%, 63,1% and 65,0%, respectively. Conclusions. Thus, both strains of rhizosphere dominant bacteria have high antagonistic activity against phytopathogenic micromycetes, which gives grounds for their further study.

酚酸物质对红松种子萌发及苗木生长和生理特性的影响

通过室内模拟试验，阐明阔叶红松林中已测得含量较高的3种酚酸物质（苯甲酸、丁香酸和香草酸）对红松种子萌发及苗木生长的影响，为探索阔叶红松林内化感作用机理及解决红松更新障碍问题提供科学依据。采用培养皿培养法及室内盆栽培养法，以红松种子和3年生红松苗为试验对象，设置不同浓度（2、20、200 mg/L）苯甲酸、丁香酸、香草酸处理液，以蒸馏水为对照（CK），进行红松种子萌发试验及红松苗木生长试验，研究3种酚酸物质对红松种子发芽、苗木生长、光合色素、抗氧化酶活性、膜脂过氧化作用及渗透调节物质的影响。结果表明，（1）不同浓度3种酚酸均抑制红松种子萌发，但酚酸浓度变化仅对红松种子发芽率影响差异显著。（2）3种酚酸对红松苗木生长及物质积累抑制作用显著。浓度变化对红松苗株高及地径影响不显著，对生物量、根干重和茎干重影响显著。（3）针叶叶绿素a、叶绿素b、类胡萝卜素含量变化对酚酸处理反应一致，20 mg/L的3种酚酸均显著抑制光合色素产生，而200 mg/L丁香酸溶液及2 mg/L香草酸溶液均显著促进叶绿素a和类胡萝卜素积累。（4）酚酸处理使红松苗针叶中POD、CAT活性降低，SOD活性增加。针叶中MDA含量显著增加，200 mg/L丁香酸溶液处理组针叶MDA含量高于CK处理组70.51%。（5）不同浓度苯甲酸溶液促进可溶性糖增加，抑制可溶性蛋白增加；不同浓度丁香酸溶液促进可溶性蛋白增加，而不同浓度香草酸溶液抑制可溶性蛋白增加，二者对可溶性糖含量影响受浓度变化影响显著。苯甲酸、丁香酸、香草酸影响红松种子萌发，通过对红松苗光合色素、抗氧化酶活性及渗透调节物质的影响导致其生长受抑制、生物量减少，产生膜脂过氧化伤害。因此，解决阔叶红松林内红松更新障碍问题时，凋落物及土壤中酚酸物质的化感作用不容忽视。;Through the relevant indoor simulation experiments, this research aims to clarify the effects of three phenolic acids (benzoic acid, syringic acid, and vanillic acid) with high content in broad-leaved Pinus koraiensis forest on the seed germination and seedling growth of Pinus koraiensis, so as to provide scientific basis for exploring the allelopathy mechanism of broad-leaved Pinus koraiensis forest and solving the problem of regeneration obstacle of Pinus koraiensis. The experiment adopts petri dish culture method and the indoor potted culture method. The Pinus koraiensis seeds and triennial Pinus koraiensis seedlings as experimental object were treated with different concentrations (2, 20, and 200 mg/L) of benzoic acid, syringic acid, vanillic acid solution, and distilled water was used as control. The germination experiment of Pinus koraiensis seed and the Pinus koraiensis seedling growth test were carried out to study the effects of three phenolic acids on germination, seedling growth, photosynthetic pigment, antioxidant enzyme activity, membrane lipid peroxidation, and osmoregulation substances of Pinus koraiensis seeds. The results show that: (1) all the three phenolic acids with different concentrations had the effects of inhibiting germination of Pinus koraiensis seeds, but the change of phenolic acid concentration only had a significant difference in the germination rate of Pinus koraiensis seeds. (2) The three phenolic acids had significant inhibitory effects on the growth and substance accumulation of Pinus koraiensis seedlings. Phenolic acid concentration had no significant effect on plant height and root length, but had significant effect on biomass, root weight and stem weight. (3) The changes in the contents of chlorophyll a, chlorophyll b, and carotenoids in needles had same reaction to the phenolic acid treatment. All the three phenolic acids of 20 mg/L had significant inhibition on the production of photosynthetic pigments, while the syringic acid solution of 200 mg/L and the vanillic acid solution of 2 mg/L had significant effects on promoting the accumulation of chlorophyll a and carotenoids. (4) Phenolic acid treatment reduced the activity of peroxidase and catalase and increased the activity of superoxide dismutase. The content of malondialdehyde in needles was significantly increased, while the content of MDA in needles treated with 200 mg/L syringic acid group was 70.51% higher than that in control group. (5) Benzoic acid solutions of different concentrations could promote the increase of soluble sugar and inhibit the increase of soluble protein. Different concentrations of syringic acid could promote the increase of soluble protein, while different concentrations of vanillic acid could inhibit the increase of soluble protein. The concentration changes of both had a significant influence on the change of soluble sugar content. Benzoic acid, syringic acid, and vanillic acid affected the germination of Pinus koraiensis seeds, which led to growth inhibition, biomass reduction and membrane lipid peroxidation of Pinus koraiensis seedlings by affecting photosynthetic pigment, antioxidant enzyme activity and osmotic regulation substances. Therefore, allelopathy of phenolic acids in litter and soil should not be ignored when solving the problem of Pinus koraiensis regeneration in broad-leaved Pinus koraiensis forests.

Design, Fabrication, and Validation of a Petri Dish-Compatible PDMS Bioreactor for the Tensile Stimulation and Characterization of Microtissues.

In this paper, we report on a novel biocompatible micromechanical bioreactor (actuator and sensor) designed for the in situ manipulation and characterization of live microtissues. The purpose of this study was to develop and validate an application-targeted sterile bioreactor that is accessible, inexpensive, adjustable, and easily fabricated. Our method relies on a simple polydimethylsiloxane (PDMS) molding technique for fabrication and is compatible with commonly-used laboratory equipment and materials. Our unique design includes a flexible thin membrane that allows for the transfer of an external actuation into the PDMS beam-based actuator and sensor placed inside a conventional 35 mm cell culture Petri dish. Through computational analysis followed by experimental testing, we demonstrated its functionality, accuracy, sensitivity, and tunable operating range. Through time-course testing, the actuator delivered strains of over 20% to biodegradable electrospun poly (D, L-lactide-co-glycolide) (PLGA) 85:15 non-aligned nanofibers (~91 µm thick). At the same time, the sensor was able to characterize time-course changes in Young’s modulus (down to 10–150 kPa), induced by an application of isopropyl alcohol (IPA). Furthermore, the actuator delivered strains of up to 4% to PDMS monolayers (~30 µm thick), simultaneously characterizing their elastic modulus up to ~2.2 MPa. The platform repeatedly applied dynamic (0.23 Hz) tensile stimuli to live Human Dermal Fibroblast (HDF) cells for 12 hours (h) and recorded the cellular reorientation towards two angle regimes, with averages of −58.85° and +56.02°. The device biocompatibility with live cells was demonstrated for one week, with no signs of cytotoxicity. We can conclude that our PDMS bioreactor is advantageous for low-cost tissue/cell culture micromanipulation studies involving mechanical actuation and characterization. Our device eliminates the need for an expensive experimental setup for cell micromanipulation, increasing the ease of live-cell manipulation studies by providing an affordable way of conducting high-throughput experiments without the need to open the Petri dish, reducing manual handling, cross-contamination, supplies, and costs. The device design, material, and methods allow the user to define the operational range based on their targeted samples/application.

The Petri Dish-N2B27 Culture Condition Maintains RPE Phenotype by Inhibiting Cell Proliferation and mTOR Activation.

Objective To develop a method for the rapid isolation of rat RPE cells with high yield and maintain its epithelial state in modified culture system. Methods The eyeballs were incubated with dispase. The retina was isolated with RPE attached and cut into several pieces. Following a brief incubation in growth medium, large RPE sheets can be harvested rapidly. RPE cells were divided into four groups and cultured for several weeks, that is, (1) in cell culture dishes with 10% FBS containing medium (CC dish-FBS), (2) in petri dishes with 10% FBS containing medium (Petri dish-FBS), (3) in cell culture dishes with N2 and B27 containing medium (CC dish-N2B27), and (4) in petri dishes with N2 and B27 containing medium (Petri dish-N2B27). Morphological and biological characteristics were investigated using light microscopy, Q-PCR, and western blot. Results The retina would curl inwardly during the growth medium incubation period, releasing RPE sheets in the medium. Compared with low density group (5,000 cells/cm2), RPE cells plated at high density (15,000 cells/cm2) can maintain RPE morphology for a more extended period. Meanwhile, plating RPE cells at low density significantly reduced the expression of RPE cell type-specific genes (RPE65, CRALBP, and bestrophin) and increased the expression of EMT-related genes (N-cadherin, fibronectin, and α-SMA), in comparison with the samples from the high density group. The petri dish culture condition reduced cell adhesion and thus inhibited RPE cell proliferation. As compared with other culture conditions, RPE cells in the petri dish-N2B27 condition could maintain RPE phenotype with increased expression of RPE-specific genes and decreased expression of EMT-related genes. The AKT/mTOR pathway was also decreased in petri dish-N2B27 condition. Conclusion The current study provided an alternative method for easy isolation of RPE cells with high yield and maintenance of its epithelial morphology in the petri dish-N2B27 condition.