Growth Medium Research Articles

Cyclization increases bactericidal activity of arginine-rich cationic cell-penetrating peptide for Neisseria gonorrhoeae.

We previously reported that a linear cationic 12-amino acid cell-penetrating peptide (CPP) was bactericidal for Neisseria gonorrhoeae. In this study, our objectives were to determine the effect of cyclization of the linear CPP on its antibacterial activity for N. gonorrhoeae and cytotoxicity for human cells. We compared the bactericidal effect of 4-hour treatment with the linear CPP to that of CPPs cyclized by a thioether or a disulfide bond on human challenge and multi-drug resistant (MDR) strains of N. gonorrhoeae grown in cell culture media with 10% fetal bovine serum (FBS). The effect of lipooligosaccharide (LOS) sialylation on bactericidal activity was analyzed. We determined the ability of the CPPs to treat human cells infected in vitro with N. gonorrhoeae, to reduce the inflammatory response of human monocytic cells to gonococci, to kill strains of three commensal Neisseria species, and to inhibit gonococcal biofilms. The cyclized CPPs killed 100% of gonococci from all strains at 100 µM and >90% at 20 µM and were more potent than the linear form. The thioether-linked but not the disulfide-linked CPP was less cytotoxic for human cervical cells compared to the linear CPP. LOS sialylation had minimal effect on bactericidal activity. In treating infected human cells, the thioether-linked CPP at 20 µM killed >60% of extra- and intracellular bacteria and reduced TNF-α expression by THP-1 cells. The potency of the CPPs for the pathogenic and the commensal Neisseria was similar. The thioether-linked CPP partially eradicated gonococcal biofilms. Future studies will focus on determining efficacy in the female mouse model of gonorrhea.IMPORTANCENeisseria gonorrhoeae remains a major cause of sexually transmitted infections with 82 million cases worldwide in 2020, and 710,151 confirmed cases in the US in 2021, up 25% from 2017. N. gonorrhoeae can infect multiple tissues including the urethra, cervix, rectum, pharynx, and conjunctiva. The most serious sequelae are suffered by infected women as gonococci ascend to the upper reproductive tract and cause pelvic inflammatory disease, chronic pelvic pain, and infertility in 10%-20% of women. Control of gonococcal infection is widely recognized as increasingly challenging due to the lack of any vaccine. N. gonorrhoeae has quickly developed resistance to all but one class of antibiotics and the emergence of multidrug-resistant strains could result in untreatable infections. As such, gonorrhea is classified by the Center for Disease Control (CDC) as an urgent public health threat. The research presented herein on new therapeutics for gonorrhea has identified a cyclic cell-penetrating peptide (CPP) as a potent molecule targeting N. gonorrhoeae.

Methods of karyoplast cell cycle synchronization for increasing the efficiency of somatic cloning of farm animals

This article is aimed to the analysis of methods for synchronizing the cell cycle of the somatic cells (karyoplasts) upon somatic cloning of farm animals. To obtain cloned animals, the oocytes at the metaphase of the second meiotic division without prior activation are applied as recipient cells in most experiments. In most experiments to obtain cloned offspring, oocytes are used as recipient cells at metaphase of the second meiotic division without prior activation, which determines the choice of somatic cells as karyoplasts at the G0/G1 stage, the most optimal for subsequent nuclear reprogramming by oocyte cytoplasmic factors. The most commonly used methods for arresting cells in this phase are serum starvation and/or contact inhibition, which allows up to 90% of cells to be stopped at the G0/G1 stage. However, despite the effectiveness of these methods, they have a number of significant limitations, and therefore the addition of components to the culture medium of somatic cells that prevent the progression of cells through the cell cycle stages is becoming widespread. The advantage of using chemical inhibitors is the ability of a number of them to have a protective effect on somatic cells, preventing the induction of apoptotic changes. The efficacy of butyrolactone-I, mimosine and aphidicolin application is controversial but a number of studies attests the possible using of these drugs to synchronize the karyoplasts. Thus, there is a wide range of methods for effective synchronization of the cell cycle of karyoplasts, and when choosing the optimal method, one should pay attention to the type of cells, the species of animals from which they were obtained, the permissible duration of cultivation under given conditions in order to minimize the negative impact of conditions on karyoplast viability.

Micropropagation and genetic transformation of Byblis liniflora

AbstractByblis, a small genus of carnivorous plants predominantly found in Australia, is characterized by its passive trapping mechanism and unique floral features. The chemical composition of Byblis, including identified phenylethanoid glycosides, particularly acteoside, highlights its pharmacological potential with various biological activities. In vitro culture techniques have been established for propagation, with micropropagation protocols developed for different Byblis species. However, information on genetic transformation, vital for trait modification and enhanced pharmacological interest, remains limited. This study focuses on optimizing micropropagation, adventitious regeneration, and genetic transformation methods for Byblis liniflora. Adventitious regeneration rates were highest in medium with reduced Murashige and Skoog salts (MS/10) and sucrose (3 gL−1) concentrations. Zeatin supplementation (1 mgL−1) further improved regeneration rates and bud development with 100% of regenerated root explants and 8.8 shoots per explant. Liquid MB3 medium supplemented with indole-3-acetic acid (IAA) 5 mgL−1 facilitated efficient rooting and acclimatization. The establishment of an efficient Rhizobium-mediated genetic transformation method yielded transgenic plants expressing green fluorescent protein (GFP). Molecular analysis confirmed transgene integration, marking the first successful genetic transformation in the Byblis genus. These advancements pave the way for exploring gene function and enhancing pharmacological properties, thereby broadening our understanding and utilization of carnivorous plants like Byblis.

Composite Liquid Media Influence on the Optical and Bactericidal Properties of Silver Nanoparticles Synthesized by Pulsed Laser Ablation in Liquids

Composite Liquid Media Influence on the Optical and Bactericidal Properties of Silver Nanoparticles Synthesized by Pulsed Laser Ablation in Liquids

Influence of the Culture Conditions on Camellia sinensis Cell Cultures

Since the last century, it has been shown that dedifferentiated cells of Camellia sinensis can produce catechins and other secondary metabolites under in vitro conditions, with potential applications in the cosmetic, pharmaceutical and food industries. In this work, cell suspension cultures of a C. sinensis cell line (LSC-5Y) were established in a liquid medium in order to optimize the biomass productivity, catechin monomer (GC, EGC, C, EC, CG, and ECG) and alkaloid (TB and CAF) productivity. The following factors were evaluated: concentration of growth regulators (BA and IBA), inoculum size, age of the cell line, light exposure, and effect of biotic elicitors (MeJA and extracts of Ciborinia camelliae). GC, EGC, and ECG increased approximately 1.80-fold when the auxin IBA concentration was increased from 0.1 to 2.0 mg/L. In addition, better productivity of EGC, C, EC, and CAF was achieved by using inoculum densities between 50 and 100 g/L. Although lower inoculum densities (25 g/L) showed a higher growth rate (0.20 d−1), the use of inoculum densities higher than 25 g/L favors a 2–4-fold increase in total catechin (TC) productivity, with maximum productivity being reached after 21 days of culture. However, the cell line showed instability in TC productivity: in the short term (in three successive subcultures), the coefficient of variation was 32.80%, and catechin production capacity was 2.5 years with maximum productivity at 0.5 years. Finally, it was observed that ethanol, used as an elicitor solvent, has a strong elicitor effect capable of increasing the accumulation of catechins up to 5.24 times compared to the treatment without an elicitor.

Three-phase-lag analysis of transversely isotropic double porous thermoelastic waves with liquid medium

Three-phase-lag analysis of transversely isotropic double porous thermoelastic waves with liquid medium

Fouling resistance of novel amphiphilic polyampholyte membranes with varying hydrophobicity towards common foulants and cell culture media

Self-assembly is a spontaneous and scalable manufacturing method to create much-needed fouling resistant membranes with narrow pore size distributions. However, there is a limited number of self-assembled membrane chemistries that can access pore sizes below 2 nm. Novel thin film composite membranes with a selective layer manufactured from a random Polyampholyte Amphiphilic Copolymer (r-PAC) have an effective pore size of 1.5 nm, relevant to various nanofiltration and water treatment applications. This is the first study that reports the membrane performance of novel r-PAC layers, prepared with varying degrees of ionic content, in filtering several classes of foulants (oil, organics, proteins, and complex mixtures). The net neutral charge of the r-PACs used enabled steady salt rejections independent of the ionic strength of the feed. The best performing r-PAC membrane composition showed no irreversible flux decline in all fouling tests and was further tested with a realistic feed, a complex cell culture media mixture utilized in cellular agriculture. This membrane exhibited an impressive 0 % irreversible flux decline after filtering the cell culture media for 20 h while a comparable commercial membrane exhibited 90 % irreversible flux decline. The chemical stability and excellent fouling resistance behavior of these membrane materials to a broad range of feeds implies these materials can enable the filtration of feeds with extreme fouling potential in biomanufacturing, wastewater treatment, and resource recovery applications.

Repetitive shock waves generated by a single long pulse underwater arc discharge

The electrohydraulic effect induced by underwater arc discharge is an efficient way to generate controllable, high-intensity shock waves. However, the development process of underwater arc discharge involves the complex coupling of plasma arc, gas bubble, and liquid medium, of which the evolution mechanism is not well understood. In this paper, the underwater arc discharge process at a millisecond pulse (>50 ms) was investigated by high-speed shadow imaging and colorimetric temperature measurement, and a simulation model of bubble pulsation was proposed to quantitatively estimate the state variation and energy transfer of the gas bubble. The results indicate that the whole arc discharge process can be categorized into three successive stages: short-period oscillation, long-period oscillation, and quasi-steady state. The vapor inside the bubble can reach a supercritical state (827 K and 140 MPa) at the minimum bubble radius. The simulation shows that the light radiation absorption and the heat conduct loss through metal electrodes are the two dominant factors influencing the pulsation of the bubble, and further analysis indicates that the dynamic evolution of the arc determines the bubble pulsation mode. Our findings demonstrate why and how repetitive electrohydraulic shock waves can be generated by a single long pulse underwater arc discharge, providing a low-cost way of shock wave generator based on an AC/DC high-voltage power source.

TARGETING ALTERNATIVE SPLICING OF FIBRONECTIN IN HUMAN RENAL PROXIMAL TUBULE EPITHELIAL CELLS WITH ANTISENSE OLIGONUCLEOTIDES TO REDUCE EDA+ FIBRONECTIN PRODUCTION AND BLOCK AN AUTOCRINE LOOP THAT DRIVES RENAL FIBROSIS

TGFβ1 is a powerful regulator of fibrosis; secreted in a latent form, it becomes active after release from the latent complex. During tissue fibrosis, the EDA+ isoform of cellular fibronectin is overexpressed. In pulmonary fibrosis it has been proposed that the fibronectin splice variant including the EDA domain (FN EDA+) activates latent TGFβ. Our work investigates the potential of blocking the ‘splicing in’ of EDA with antisense oligonucleotides to inhibit TGFβ1-induced EDA+ fibronectin and to prevent the cascade of events initiated by TGFβ1 in human renal proximal tubule cells (PTEC).Human primary PTEC were treated with TGFβ1 for 48 h, medium removed and the cells transfected with RNase H-independent antisense oligonucleotides (ASO) designed to block EDA exon inclusion (ASO5). The efficacy of ASO to block EDA exon inclusion was assessed by EDA+ fibronectin RNA and protein expression; the expression of TGFβ, αSMA (α smooth muscle actin), MMP2 (matrix metalloproteinse-2), MMP9 (matrix metalloproteinse-9), Collagen I, K Cadherin and connexin 43 was analysed.Targeting antisense oligonucleotides designed to block EDA exon inclusion in fibronectin pre mRNA were effective in reducing the amount of TGFβ1 -induced cellular EDA+ fibronectin RNA and secreted EDA+ fibronectin protein (assessed by western immunoblotting and immunocytochemistry) in human proximal tubule cells in an in vitro cell culture model. The effect was selective for EDA+ exon with no effect of EDB+ fibronectin RNA and total fibronectin mRNA.Exogenous TGFβ1 induced endogenous TGFβ, αSMA, MMP2, MMP9 and Col I mRNA. TGFβ1 treatment for 48h reduced the expression of K-Cadherin expression and increased the expression of connexin-43. These TGFβ1-induced pro-fibrotic changes were attenuated by ASO5 treatment. 48 h after the removal of exogenous TGFβ, further increase in αSMA, MMP2, MMP9 was observed; ASO5 significantly inhibited this subsequent increase. ASO5 treatment also significantly inhibited the ability of the cell culture medium harvested at the end of the experiment (96h) to stimulate SMAD3 reporter cells. The role of endogenous TGFβ1 was confirmed by the use of a TGFβ receptor inhibitor.Our results demonstrate a critical role of FN EDA+ in a cycle of TGFβ driven pro-fibrotic responses in human PTEC and blocking its production with ASO technology offers a potential therapy to interrupt this vicious circle and hence limit the progression of renal fibrosis.

Forming Single-Cell-Derived Colon Cancer Organoid Arrays on a Microfluidic Chip for High Throughput Tumor Heterogeneity Analysis.

Single-cell-derived tumor organoids (STOs) possess a distinct genetic background, making them valuable tools for demonstrating tumor heterogeneity. In order to fulfill the high throughput demands of STO assays, we have developed a microfluidic chip containing 30 000 microwells, which is dedicated to a single cell culture approach for selective expansion and differential induction of cancer stem cells. The microwells are coated with a hydrophilic copolymer to eliminate cell adhesion, and the cell culture is supported by poly(ethylene glycol) (PEG) to establish a nonadhesive culture environment. By utilizing an input cell density of 7 × 103·mL-1, it is possible to construct a 4000 single cell culture system through stochastic cell occupation. We demonstrate that the addition of 15% PEG10000 in the cell culture medium effectively prevents cell loss while facilitating tumor stem cell expansion. As were demonstrated by HCT116, HT29, and SW480 colon cancer cells, the microfluidic approach achieved a STO formation rate of ∼20%, resulting in over 800 STOs generated from a single culture. Comprehensive analysis through histomorphology, immunohistochemistry, drug response evaluation, assessment of cell invasion, and biomarker detection reveals the heterogeneity among individual STOs. Specifically, the smaller STOs exhibited higher invasion and drug resistance capabilities compared with the larger ones. The developed microfluidic approach effectively facilitates STO formation and offers promising prospects for investigating tumor heterogeneity, as well as conducting personalized therapy-focused drug screening.

Scalable isolation of surface-engineered extracellular vesicles and separation of free proteins via tangential flow filtration and size exclusion chromatography (TFF-SEC)

BackgroundExtracellular vesicles (EVs) represent small lipid bilayer structures pivotal in mediating intercellular communication via biomolecular transfer. Their inherent characteristics, including packaging, non-immunogenicity, and biofluid stability, position EVs as promising drug delivery vectors. However, developing clinical quality EVs requires multifaceted technological advancement.MethodsIn this study, a method is introduced for engineering extracellular vesicles (eEVs) from cultured cells and their subsequent isolation using lab-scale tangential flow filtration (TFF). This is the first study to evaluate DNA loading efficacy into EVs isolated by TFF, marking a significant milestone in the field of targeted drug delivery. Initially, cells are transfected with EV-display constructs to facilitate the secretion of eEVs bearing the desired coding molecules. Following brief centrifugation, the cell culture media undergoes filtration using hollow fiber filters. TFF, by applying a constant flow, effectively segregates molecules based on designated molecular weight cut-off (MWCO), enriching particles between 50 and 650 nm.ResultsCompared to conventional methods like ultracentrifugation, TFF demonstrates higher efficiency in removing undesired molecules/aggregates while exerting less stress on EVs. Characterization of eEVs through various assays confirms TFF’s superiority in isolating pure EV populations. Additionally, the necessity of size-exclusion chromatography (SEC) after tangential flow filtration (TFF) becomes evident for effectively removing unbound protein contaminants.ConclusionIn conclusion, TFF-SEC emerges as a scalable and superior approach for eEV isolation, promising significant advancements in clinical applications.

Selective hydrolysis of traditional Chinese medicine residue into reducing sugars catalysed by sulfonated carbon catalyst and application of hydrolysate

The conversion of waste into high-value products has long been a focus point of research in green and sustainable chemistry. In this study, a novel value-adding strategy was devised to not only facilitate the resource utilization of biomass waste like Chinese medicine residue (CMR) and pomelo peel but also contribute to the advancement of microalgae cultivation. Sulfonated biochar PP-700-S was synthesized from pomelo peel waste, and its morphology and structural characteristics were analyzed. An effective method for the hydrolysis of CMR using PP-700-S as a catalyst in the aqueous phase was then explored, with a thorough investigation of the process and key influencing factors. Under the reaction conditions of 130 °C, 24 h, 20 mL H2O, CMR: PP-700-S = 1: 1, a 51.87 % yield of reducing sugars was directly obtained from the CMR, with PP-700-S exhibiting the selectivity of glucose production. Subsequently, the hydrolysate (HL-CMR) was utilized as a growth medium for microalgae, revealing that the presence of HL-CMR enhanced the nutrient balance in the medium, creating a more conducive environment for microalgae growth. Overall, this study presents a sustainable and efficient pathway for the utilization of lignocellulosic waste, facilitating the conversion of waste into higher-value products and contributing to the development of a green and sustainable science.

A new model of endotracheal tube biofilm identifies combinations of matrix-degrading enzymes and antimicrobials able to eradicate biofilms of pathogens that cause ventilator-associated pneumonia.

Ventilator-associated pneumonia is defined as pneumonia that develops in a patient who has been on mechanical ventilation for more than 48 hours through an endotracheal tube. It is caused by biofilm formation on the indwelling tube, which introduces pathogenic microbes such as Pseudomonas aeruginosa, Klebsiella pneumoniae and Candida albicans into the patient's lower airways. Currently, there is a lack of accurate in vitro models of ventilator-associated pneumonia development. This greatly limits our understanding of how the in-host environment alters pathogen physiology and the efficacy of ventilator-associated pneumonia prevention or treatment strategies. Here, we showcase a reproducible model that simulates the biofilm formation of these pathogens in a host-mimicking environment and demonstrate that the biofilm matrix produced differs from that observed in standard laboratory growth medium. In our model, pathogens are grown on endotracheal tube segments in the presence of a novel synthetic ventilated airway mucus medium that simulates the in-host environment. Matrix-degrading enzymes and cryo-scanning electron microscopy were employed to characterize the system in terms of biofilm matrix composition and structure, as compared to standard laboratory growth medium. As seen in patients, the biofilms of ventilator-associated pneumonia pathogens in our model either required very high concentrations of antimicrobials for eradication or could not be eradicated. However, combining matrix-degrading enzymes with antimicrobials greatly improved the biofilm eradication of all pathogens. Our in vitro endotracheal tube model informs on fundamental microbiology in the ventilator-associated pneumonia context and has broad applicability as a screening platform for antibiofilm measures including the use of matrix-degrading enzymes as antimicrobial adjuvants.

Thermal Energy Generated During Ultrasonication Dominates Pinto Bean Hydration.

Ultrasound-assisted hydration (UAH) stands out as a potent strategy, offering multiple advantages over conventional thermal hydration methods. It is an energy efficient, cost and time effective, easily scalable and relatively greener approach. Acoustic cavitation is the major working mechanism in aqueous phase sonication. Cavitation creates immense localised heat and pressure, which ultimately ends up heating the liquid media. To the best of our knowledge, no study dealing with the synergistic relationship between the physical effects of ultrasound and the consequent temperature rise during UAH is available in open literature. This work investigates this complex interplay with aim to identify the dominant working mechanism leading to the hydration of the pinto beans (Phaseolus vulgaris), a variety of common beans. Three different hydration protocols were devised to discern these effects, normal hydration (N) at 25°C, temperature-controlled UAH (TC) between 18-20 °C using an ice bath, and temperature-uncontrolled UAH (TuC). Additionally, the post sonication residual effects of ultrasound treatment were examined. Our findings reveal that sonication induces changes in both TC and TuC treated beans, but more pronounced effects were observed in the absence of temperature control. During TC, acoustic streaming emerged as the predominant sonic action mechanism. Conversely, during TuC, temperature itself emerged as the dominant force, leading to irreversible alterations in the bean's physical and chemical structures.

Tolerance of Young Blackberries to a Selection of Preemergence Herbicides and Rates

Limited preemergence herbicides are registered for new blackberry (Rubus subgenus Rubus) plantings. This greenhouse experiment was designed to investigate the effects of a broad selection of preemergence herbicides at multiple rates on blackberry transplants. Screening was initiated Aug 2021 and repeated Mar 2022 in Fayetteville, AR, USA, in a greenhouse at the Milo J. Shult Agricultural Research and Extension Center. ‘Ouachita’ blackberry plugs were transplanted into utility pots that contained field soil and growth media treated with preemergence herbicides. After transplanting, plant height was measured from the substrate to the highest apical meristem of 25 representative plants. Initial blackberry plant heights were 13.5 and 9.2 cm in 2021 and 2022, respectively. Twenty-five treatments were evaluated, consisting of 12 preemergence herbicides at 1× and 2× field rates, and one untreated control. Herbicide treatments included diuron, flumioxazin, halosulfuron, indaziflam, mesotrione, napropamide, oryzalin, pendimethalin, rimsulfuron, S-metolachlor, simazine, and sulfentrazone applied to substrate in containers at their respective 1× or 2× field rates. Data were collected on plant height, blackberry injury ratings, internode length, leaf chlorophyll content, and destructive harvest, including leaf count, leaf dry biomass, and aboveground dry biomass. Specific leaf areas and leaf area-to-dry matter ratios were calculated. When observed, plant injury tended to increase from 7 days after treatment (DAT) until 42 DAT. Greater injury levels were observed in response to treatment with mesotrione at the 1× (78%) and 2× rates (90%), halosulfuron at the 1× rate (58%), halosulfuron at the 2× rate (68%), and diuron at the 2× rate (73%). Injury from diuron was rate dependent, with the 1× rate causing relatively low injury (19%). At both the 1× and 2× rates, flumioxazin, indaziflam, napropamide, S-metolachlor, and pendimethalin treatments exhibited similar responses to the untreated control.

Engineering a GPCR-based yeast biosensor for a highly sensitive melatonin detection from fermented beverages

Melatonin is a multifunctional molecule with diverse biological roles that holds great value as a health-promoting bioactive molecule in any food product and yeast’s ability to produce it has been extensively demonstrated in the last decade. However, its quantification presents costly analytical challenges due to the usual low concentrations found as the result of yeast metabolism. This study addresses these analytical challenges by optimizing a yeast biosensor based on G protein-coupled receptors (GPCR) for melatonin detection and quantitation. Strategic genetic modifications were employed to significantly enhance its sensitivity and fluorescent signal output, making it suitable for detection of yeast-produced melatonin. The optimized biosensor demonstrated significantly improved sensitivity and fluorescence, enabling the screening of 101 yeast strains and the detection of melatonin in various wine samples. This biosensor’s efficacy in quantifying melatonin in yeast growth media underscores its utility in exploring melatonin production dynamics and potential applications in functional food development. This study provides a new analytical approach that allows a rapid and cost-effective melatonin analysis to reach deeper insights into the bioactivity of melatonin in fermented products and its implications for human health. These findings highlight the broader potential of biosensor technology in streamlining analytical processes in fermentation science.

Tolerance of Young Blackberries to a Selection of Preemergence Herbicides and Rates

Limited preemergence herbicides are registered for new blackberry (Rubus subgenus Rubus) plantings. This greenhouse experiment was designed to investigate the effects of a broad selection of preemergence herbicides at multiple rates on blackberry transplants. Screening was initiated Aug 2021 and repeated Mar 2022 in Fayetteville, AR, USA, in a greenhouse at the Milo J. Shult Agricultural Research and Extension Center. ‘Ouachita’ blackberry plugs were transplanted into utility pots that contained field soil and growth media treated with preemergence herbicides. After transplanting, plant height was measured from the substrate to the highest apical meristem of 25 representative plants. Initial blackberry plant heights were 13.5 and 9.2 cm in 2021 and 2022, respectively. Twenty-five treatments were evaluated, consisting of 12 preemergence herbicides at 1× and 2× field rates, and one untreated control. Herbicide treatments included diuron, flumioxazin, halosulfuron, indaziflam, mesotrione, napropamide, oryzalin, pendimethalin, rimsulfuron, S-metolachlor, simazine, and sulfentrazone applied to substrate in containers at their respective 1× or 2× field rates. Data were collected on plant height, blackberry injury ratings, internode length, leaf chlorophyll content, and destructive harvest, including leaf count, leaf dry biomass, and aboveground dry biomass. Specific leaf areas and leaf area-to-dry matter ratios were calculated. When observed, plant injury tended to increase from 7 days after treatment (DAT) until 42 DAT. Greater injury levels were observed in response to treatment with mesotrione at the 1× (78%) and 2× rates (90%), halosulfuron at the 1× rate (58%), halosulfuron at the 2× rate (68%), and diuron at the 2× rate (73%). Injury from diuron was rate dependent, with the 1× rate causing relatively low injury (19%). At both the 1× and 2× rates, flumioxazin, indaziflam, napropamide, S-metolachlor, and pendimethalin treatments exhibited similar responses to the untreated control.

In Vitro Biocompatibility Assessment of Bioengineered PLA-Hydrogel Core–Shell Scaffolds with Mesenchymal Stromal Cells for Bone Regeneration

Human mesenchymal stromal cells (hMSCs), whether used alone or together with three-dimensional scaffolds, are the best-studied postnatal stem cells in regenerative medicine. In this study, innovative composite scaffolds consisting of a core–shell architecture were seeded with bone-marrow-derived hMSCs (BM-hMSCs) and tested for their biocompatibility and remarkable capacity to promote and support bone regeneration and mineralization. The scaffolds were prepared by grafting three different amounts of gelatin–chitosan (CH) hydrogel into a 3D-printed polylactic acid (PLA) core (PLA-CH), and the mechanical and degradation properties were analyzed. The BM-hMSCs were cultured in the scaffolds with the presence of growth medium (GM) or osteogenic medium (OM) with differentiation stimuli in combination with fetal bovine serum (FBS) or human platelet lysate (hPL). The primary objective was to determine the viability, proliferation, morphology, and spreading capacity of BM-hMSCs within the scaffolds, thereby confirming their biocompatibility. Secondly, the BM-hMSCs were shown to differentiate into osteoblasts and to facilitate scaffold mineralization. This was evinced by a positive Von Kossa result, the modulation of differentiation markers (osteocalcin and osteopontin), an expression of a marker of extracellular matrix remodeling (bone morphogenetic protein-2), and collagen I. The results of the energy-dispersive X-ray analysis (EDS) clearly demonstrate the presence of calcium and phosphorus in the samples that were incubated in OM, in the presence of FBS and hPL, but not in GM. The chemical distribution maps of calcium and phosphorus indicate that these elements are co-localized in the same areas of the sections, demonstrating the formation of hydroxyapatite. In conclusion, our findings show that the combination of BM-hMSCs and PLA-CH, regardless of the amount of hydrogel content, in the presence of differentiation stimuli, can provide a construct with enhanced osteogenicity for clinically relevant bone regeneration.

Gaseous ozone inactivation of Bacillus atrophaeus spores on ceramic and porcelain tiles

In this study, we investigated the ozone inactivation efficiency of Bacillus atrophaeus spores attached to various tile surfaces. Eight different types of tiles were employed, considering factors such as porosity (ceramic, porcelain), color (white, black), and glossiness (matte, glossy). Inactivation was performed by exposing the spore-loaded tiles to ozone gas for a specified duration. The inactivation efficiencies of ozone gas on different tile surfaces were compared by analyzing the colony-forming units of desorbed Bacillus atrophaeus cultured in a growth medium. Results revealed a reduction in colony counts with increasing ozone exposure time, indicating a proportional enhancement in inactivation effectiveness on ozone exposure time. After exposure to ozone gas for 30 min or longer, more than 90% of spores on each tile were inactivated. Regarding porosity, ceramic tiles exhibited slightly superior inactivation effects compared to porcelain tiles. Additionally, in terms of glossiness, glossy tiles demonstrated better inactivation effects than matte tiles. However, no significant differences were observed in inactivation effects based on the color of the tiles.

Antarctic Spore-Forming Microorganisms from Deception Island Inhibit the Growth of Various Bacterial Strains

Antarctic microbes have evolved and adapted unique strategies to survive in the harsh polar environment. Apart from the ability to adapt to the low nutrient soil content and extremely dry and cold polar environment, a particular strategy used by Antarctic bacteria is the production of antimicrobial compounds that can eliminate rivals in the same niche, giving them a competitive edge over other microbes. In contrast, it is unclear whether spore-forming microbes possess similar antimicrobial properties as one of their survival strategies, especially those from the Antarctic volcanic Deception island in the West Antarctic. Hence, this study aims to isolate and characterize the spore-forming microbes in Deception Island, Antarctica, as well as to identify the ones that are equipped with the ability to inhibit other microorganisms. Microbes were isolated using various growth media and were segregated into clusters based on their random amplified polymorphic DNA (RAPD) fingerprints. A total of 90 strains were isolated and clustered into 30 groups at a similarity of 60%. Representative strains from each cluster were assayed for antimicrobial activities against 13 Gram-positive and Gram-negative test bacteria comprising human pathogens. Twenty-five strains exhibited the ability to inhibit at least one test bacterium. The four strains, A60, Im31, Im32 and Im33 that showed the strongest inhibitory activities were subjected to 16S or 18S rDNA sequencing and analysis to identify them. They were identified as Pseudogymnoascus, Bacillus, Leohumicola, and Talaromyces spp. The ability of the aforementioned microbes to thrive in harsh environments and compete with fierce competitors for scarce nutrients is probably due to their ability to produce antimicrobial compounds that target and kill their rivals.